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1. Ibrahim  A, Liesack  W, Pike  S, Stackebrandt  E,     ( 1992 )

The polymerase chain reaction: an epidemiological tool to differentiate between two clusters of pathogenic Yersinia enterocolitica strains.

FEMS microbiology letters 76 (1��2��)
PMID : 1427005  :   DOI  :   10.1016/0378-1097(92)90364-t    
Abstract >>
A primer set designed to amplify the enterotoxin (yst) gene of pathogenic Yersinia enterocolitica strains generated two different electrophoretic profiles of the target sequence when a collection of strains of worldwide origin was screened. Serovars O:1,3; O:2a,3; O:3; O:5,27 and O:9, known as European strains, produced a 200-bp fragment that matched the size of the target sequence. However, serovars O:4,32; O:8; O:13a,13b; O:20 and O:21, known as American strains, generated two fragments of 1.4 and 1.6 kb. The amplified products of one American strain were sequenced and the presence of the yst gene was confirmed in both fragments. Thus, the potential of the polymerase chain reaction to be used as an epidemiological tool in differentiation between the two clusters of pathogenic strains of Y. enterocolitica could be demonstrated.
KeywordMeSH Terms
2. Zhang  ZY, Clemens  JC, Schubert  HL, Stuckey  JA, Fischer  MW, Hume  DM, Saper  MA, Dixon  JE,     ( 1992 )

Expression, purification, and physicochemical characterization of a recombinant Yersinia protein tyrosine phosphatase.

The Journal of biological chemistry 267 (33)
PMID : 1429715  :  
Abstract >>
The Yersinia protein tyrosine phosphatase (PTPase) Yop51, a C235R point mutation (Yop51*), and a protein lacking the first 162 amino acids at the NH2 terminus (Yop51*delta 162) have been overexpressed in Escherichia coli and purified to homogeneity through the use of CM Sephadex C25 cation exchange chromatography followed by Sephadex G-100 gel filtration. Greater than 50 mg of homogeneous Yop51* and Yop51*delta 162 can be obtained from a single liter of bacterial culture, whereas the same procedure yields only 5 mg of pure Yop51. Large, diffraction-quality crystals have been obtained for Yop51*delta 162. Size exclusion chromatography, sedimentation equilibrium, and enzyme concentration dependence experiments have established that the Yersinia PTPases exist and function as monomers in solution. Yop51 and Yop51* display identical UV, CD, and fluorescence spectra and have identical kinetic and structural stability properties. These full-length Yersinia PTPases have 31% alpha-helix, an emission maximum of 342 nm, a turn-over number of 1200 s-1 at pH 5.0, 30 degrees C, and an unfolding delta G value of 6 kcal/mol at 25 degrees C. Yop51*delta 162 has very similar kinetic and fluorescence characteristics to the full-length molecules, whereas its CD and UV spectra show noticeable differences due to the elimination of 162 NH2-terminal residues. The Yersinia PTPases are by far the most active PTPases known, and their kinetic parameters are extremely sensitive to the ionic strength of reaction medium.
KeywordMeSH Terms
3. Stojiljkovic  I, Hantke  K,     ( 1992 )

Hemin uptake system of Yersinia enterocolitica: similarities with other TonB-dependent systems in gram-negative bacteria.

The EMBO journal 11 (12)
PMID : 1425573  :   PMC  :   PMC557009    
Abstract >>
The hemin receptor HemR of Yersinia enterocolitica was identified as a 78 kDa iron regulated outer membrane protein. Cells devoid of the HemR receptor as well as cells mutated in the tonB gene were unable to take up hemin as an iron source. The hemin uptake operon from Y. enterocolitica was cloned in Escherichia coli K12 and was shown to encode four proteins: HemP (6.5 kDa), HemR (78 kDa), HemS (42 kDa) and HemT (27 kDa). When expressed in E.coli hemA aroB, a plasmid carrying genes for HemP and HemR allowed growth on hemin as a porphyrin source. Presence of genes for HemP, HemR and HemS was necessary to allow E.coli hemA aroB cells to use hemin as an iron source. The nucleotide sequence of the hemR gene and its promoter region was determined and the amino acid sequence of the HemR receptor deduced. HemR has a signal peptide of 28 amino acids and a typical TonB box at its amino-terminus. Upstream of the first gene in the operon (hemP), a well conserved Fur box was found which is in accordance with the iron-regulated expression of HemR.
KeywordMeSH Terms
Escherichia coli Proteins
Receptors, Cell Surface
4. Foultier  B, Cornelis  GR,     ( 2003 )

DNA sequence and analysis of the pYVa127/90 virulence plasmid of Yersinia enterocolitica strain A127/90.

Research in microbiology 154 (8)
PMID : 14527656  :   DOI  :   10.1016/S0923-2508(03)00147-5    
Abstract >>
The complete nucleotide sequence and organization of the Yersinia enterocolitica strain A127/90 serotype O:8 biotype 1B virulence plasmid (pYVa127/90) were determined. In contrast to other biotype 1B strains, usually isolated in the USA, strain A127/90 was isolated in Japan. The organization of the genes carried by the 66591-bp pYVa127/90 plasmid is globally similar to that of 67720-bp pYVe8081, the virulence plasmid from 8081, an American isolate of serotype O:8 biotype 1B. Differences are noticeable in the amino acid sequences of the SycH chaperone, the YopM cytotoxin and the plague-protective antigen LcrV. Interestingly, pYVa127/90 contains the entire coding sequence of the YlpA lipoprotein, which is missing from pYVe8081. In addition, we identified five potential new genes. Several vestigial insertion sequence elements appeared to be different between the two plasmids. These observations suggest that both pYV plasmids are from a common ancestor but may have evolved independently following the conquest of two distant ecological niches.
KeywordMeSH Terms
Plasmids
5. Dube  PH, Handley  SA, Revell  PA, Miller  VL,     ( 2003 )

The rovA mutant of Yersinia enterocolitica displays differential degrees of virulence depending on the route of infection.

Infection and immunity 71 (6)
PMID : 12761136  :   DOI  :   10.1128/iai.71.6.3512-3520.2003     PMC  :   PMC155726    
Abstract >>
Yersinia enterocolitica is an invasive enteric pathogen that causes significant inflammatory disease. Recently, we identified and characterized a global regulator of virulence (rovA). When mice are infected orally with the rovA mutant they are attenuated by 50% lethal dose (LD(50)) analysis and have altered kinetics of infection. Most significantly, mice orally infected with the rovA mutant have greatly reduced inflammation in the Peyer's patches compared to those infected with wild-type Y. enterocolitica. However, we present data here indicating that when the rovA mutant bacteria are delivered intraperitoneally (i.p.), they are significantly more virulent than when delivered orally. The i.p. LD(50) for the rovA mutant is only 10-fold higher than that of the wild-type Y. enterocolitica, and there are significant inflammatory responses to the rovA mutant that are evident in the liver and spleen. Altogether, these data suggest that the RovA regulon may be required for the early events of the infection that occur in the Peyer's patches. Furthermore, these data suggest that the RovA regulon may be dispensable for Y. enterocolitica systemic disease and inflammatory responses if the Peyer's patches are bypassed.
KeywordMeSH Terms
6. Sánchez-Céspedes  J, Navia  MM, Martínez  R, Orden  B, Millán  R, Ruiz  J, Vila  J,     ( 2003 )

Clonal dissemination of Yersinia enterocolitica strains with various susceptibilities to nalidixic acid.

Journal of clinical microbiology 41 (4)
PMID : 12682183  :   DOI  :   10.1128/jcm.41.4.1769-1771.2003     PMC  :   PMC153901    
Abstract >>
Ten epidemiologically related Yersinia enterocolitica clinical isolates were studied. Six isolates were nalidixic acid resistant (MIC > 512 microg/ml), with mutations in the quinolone resistance-determining region (QRDR) of the gyrA gene, suggesting clonal dissemination of a nalidixic acid-susceptible Y. enterocolitica strain which has acquired different mutations generating resistance to nalidixic acid.
KeywordMeSH Terms
Mutation
7. Iwobi  A, Heesemann  J, Garcia  E, Igwe  E, Noelting  C, Rakin  A,     ( 2003 )

Novel virulence-associated type II secretion system unique to high-pathogenicity Yersinia enterocolitica.

Infection and immunity 71 (4)
PMID : 12654803  :   DOI  :   10.1128/iai.71.4.1872-1879.2003     PMC  :   PMC152056    
Abstract >>
Yersinia enterocolitica strains comprise an important group of bacterial enteropathogens that cause a broad range of gastrointestinal syndromes. Three groups are distinguishable within this bacterial species, namely, the nonpathogenic group (biotype 1A strains), the low-pathogenicity, non-mouse-lethal group (biotypes 2 to 5), and the high-pathogenicity, mouse-lethal group (biotype 1B). To date, the presence of the high-pathogenicity island (HPI), a chromosomal locus that encodes the yersiniabactin system (involved in iron uptake), defines essentially the difference between low-pathogenicity and high-pathogenicity Y. enterocolitica strains, with the low-pathogenicity strains lacking the HPI. Using the powerful tool of representational difference analysis between the nonpathogenic 1A strain, NF-O, and its high-pathogenicity 1B counterpart, WA-314, we have identified a novel type II secretion gene cluster (yts1C-S) occurring exclusively in the high-pathogenicity group. The encoded secreton, designated Yts1 (for Yersinia type II secretion 1) was shown to be important for virulence in mice. A close examination of the almost completed genome sequence of another high-pathogenicity representative, Y. enterocolitica 8081, revealed a second putative type II secretion cluster uniformly distributed among all Y. enterocolitica isolates. This putative species-specific cluster (designated yts2) differed significantly from yts1, while resembling more closely the putative type II cluster present on the genome of Y. pestis. The Yts1 secreton thus appears to have been additionally acquired by the high-pathogenicity assemblage for a virulence-associated function.
KeywordMeSH Terms
Multigene Family
8. Antonenko  V, Pawlow  V, Heesemann  J, Rakin  A,     ( 2003 )

Characterization of a novel unique restriction-modification system from Yersinia enterocolitica O:8 1B.

FEMS microbiology letters 219 (2)
PMID : 12620628  :   DOI  :   10.1016/S0378-1097(03)00047-8    
Abstract >>
Genetic manipulations with enteropathogenic Yersinia enterocolitica O:8 are complicated by the presence of an efficient PstI-like YenI restriction-modification (R-M) system. We have characterized the YenI R-M system in Y. enterocolitica O:8, biotype 1B. A 5039 bp DNA fragment of the pSAK2 recombinant plasmid carrying the yenI locus was used to determine the nucleotide sequence. DNA sequence analysis identified a single 2481 bp open reading frame (ORF) that encodes an 826 amino acid large polypeptide having an apparent molecular mass of 93 kDa. The N-terminal part of the YenI ORF has 45 and 40% identity to PstI and BsuI methyltransferases (MTases), respectively; while the C-terminal part depicts 55 and 45% identity to endonucleases (ENases) of both isoschyzomeric enzymes. The yenI gene was cloned into pT7-5 plasmid and has been shown to encode a single polypeptide of expected molecular mass. A specific recognition sequence, typical to the type II R-M systems and single peptide organization, typical to type IV R-M systems, make YenI unique among known restriction-modification systems. We have constructed a truncated recombinant variant of YenI enzyme, which conserved only MTase activity, and that can be applied to YenI methylation of the DNA to be transformed into Y. enterocolitica O:8 biotype 1B strains.
KeywordMeSH Terms
9. Roggenkamp  A, Ackermann  N, Jacobi  CA, Truelzsch  K, Hoffmann  H, Heesemann  J,     ( 2003 )

Molecular analysis of transport and oligomerization of the Yersinia enterocolitica adhesin YadA.

Journal of bacteriology 185 (13)
PMID : 12813066  :   DOI  :   10.1128/jb.185.13.3735-3744.2003     PMC  :   PMC161578    
Abstract >>
The Yersinia adhesin YadA is the prototype of a novel class of bacterial adhesins which form oligomeric lollipop-like structures and are anchored in the outer membrane by the C terminus. For YadA, six different regions (R) or domains (D) are predicted from the amino acid sequence: the N-terminal leader sequence, head-D, neck-D, stalk-D, linking-R, and a C-terminal transmembrane region consisting of four beta-strands. To identify structural and functional features of these domains, we performed in-frame deletion mutagenesis and constructed N-terminally tagged YadA variants. Diverse YadA variants were analyzed for outer membrane localization, surface exposure, oligomerization adhesion properties, and ability to protect against complement-mediated lysis. We demonstrated that (i) the C-terminal region (amino acids [aa] 353 to 422) is sufficient for outer membrane insertion and formation of trimers in the outer membrane; (ii) the head, neck, and stalk domains (aa 26 to 330) are surface exposed, forming a passenger domain; and (iii) the linking region (aa 331 to 369) is responsible for outer membrane translocation of the passenger domain. Thus, YadA meets all the criteria of an autotransporter. The same may be true for all other members of the YadA family, forming a subfamily of surface-attached oligomeric autotransporters. Moreover, in-frame truncation mutagenesis suggested that the head and neck domains together form the YadA-binding module which is located on the top of the stalk. However, the YadA-binding module did not confer serum resistance. Mutants lacking the head and neck domain were resistant to complement-mediated lysis. In-frame truncation of the stalk domain did not result in significant attenuation of the mutant in an orogastric mouse infection model.
KeywordMeSH Terms
10. Seoane  A, Sánchez  E, García-Lobo  JM,     ( 2003 )

Tandem amplification of a 28-kilobase region from the Yersinia enterocolitica chromosome containing the blaA gene.

Antimicrobial agents and chemotherapy 47 (2)
PMID : 12543678  :   DOI  :   10.1128/aac.47.2.682-688.2003     PMC  :   PMC151765    
Abstract >>
Most Yersinia enterocolitica strains are resistant to beta-lactam antibiotics due to the production of one or two chromosomally encoded beta-lactamases. Strain Y56 is a Y. enterocolitica O:3 serotype natural isolate that is resistant to moderate amounts of penicillins and that produces a single class A beta-lactamase. To select mutants with increased levels of resistance to beta-lactam antibiotics, strain Y56 was grown on plates containing increasing amounts of ampicillin, and variants resistant to up to 500 micro g of ampicillin per ml were obtained. Chromosomal DNA from hyperresistant isolates was analyzed by Southern hybridization with a blaA-specific probe to detect gene rearrangements. The use of pulsed-field gel electrophoresis revealed that the increase in the resistance level correlated with the amplification in tandem of a DNA fragment of about 28 kb containing the blaA gene. The phenotype of these isolates was not stable, and they recovered the basal low resistance level when the ampicillin used for selection was withdrawn from the growth medium. This loss of resistance was followed by the recovery of the original chromosomal structure. To understand this amplification process, the 28-kb amplification unit was cloned, and the ends were sequenced. The analysis of these sequences did not reveal the presence of either repeats or transposable elements to explain this process. However, we found short sequences similar to some DNA gyrase target sequences that have been described. In addition, we observed that the frequency of appearance of ampicillin-hyperresistant isolates by amplification of the blaA locus was lowered in the presence of the gyrase inhibitor novobiocin. These findings suggest that the DNA gyrase could be involved in this amplification event.
KeywordMeSH Terms
Bacterial Proteins
11. Foultier  B, Troisfontaines  P, Vertommen  D, Marenne  MN, Rider  M, Parsot  C, Cornelis  GR,     ( 2003 )

Identification of substrates and chaperone from the Yersinia enterocolitica 1B Ysa type III secretion system.

Infection and immunity 71 (1)
PMID : 12496172  :   DOI  :   10.1128/iai.71.1.242-253.2003     PMC  :   PMC143280    
Abstract >>
All pathogenic Yersinia enterocolitica strains carry the pYV plasmid encoding the Ysc-Yop type III secretion (TTS) system, which operates at 37 degrees C. In addition, biovar 1B Y. enterocolitica strains possess a second, chromosomally encoded, TTS system called Ysa, which operates, at least in vitro, under low-temperature and high-salt (LTHS) conditions. Six open reading frames, sycB, yspB, yspC, yspD, yspA, and acpY, neighbor the ysa genes encoding the Ysa TTS apparatus. Here we show that YspA, YspB, YspC, and YspD are secreted by the Ysa TTS system under LTHS conditions. SycB is a chaperone for YspB and YspC and stabilizes YspB. YspB, YspC, and SycB share some similarity with TTS substrates and the chaperone encoded by the Mxi-Spa locus of Shigella flexneri and SPI-1 of Salmonella enterica. In addition, Ysa also secretes the pYV-encoded YopE under LTHS conditions, indicating that YopE is a potential effector of both Y. enterocolitica TTS systems. YspC could also be secreted by S. flexneri, but no functional complementation of ipaC was observed, which indicates that despite their similarity the Ysa and the Mxi-Spa systems are not interchangeable. When expressed from the yopE promoter, YspB and YspC could also be secreted via the Ysc injectisome. However, they could not form detectable pores in eukaryotic target cells and could not substitute for YopB and YopD for translocation of Yop effectors.
KeywordMeSH Terms
12. Izquierdo  L, Abitiu  N, Coderch  N, Hita  B, Merino  S, Gavin  R, Tomás  JM, Regué  M,     ( 2002 )

The inner-core lipopolysaccharide biosynthetic waaE gene: function and genetic distribution among some Enterobacteriaceae.

Microbiology (Reading, England) 148 (Pt 11)
PMID : 12427940  :   DOI  :   10.1099/00221287-148-11-3485    
Abstract >>
To determine the function of the waaE gene in the biosynthesis of the inner-core LPS of Klebsiella pneumoniae, a waaE non-polar mutant has been constructed. Data obtained from the comparative chemical analysis of LPS samples obtained from the wild-type, the mutant strain and the complemented mutant demonstrated that the waaE gene is involved in substitution of alpha-L-glycero-D-manno-heptopyranose I (L,D-HeppI) at the O-4 position by a beta-D-glucopyranose (beta-D-Glcp) residue. In addition, DNA amplification and nucleotide sequence determination studies revealed that waaE homologues located between the waaA and coaD genes are present in clinical isolates of Enterobacteriaceae containing the structure beta-D-Glcp-(1-->4)-alpha-L,D-HeppI (K. pneumoniae, Proteus mirabilis and Yersinia enterocolitica), as well as in strains of Serratia marcescens and Enterobacter aerogenes of unknown LPS-core structures. Complementation studies using non-polar waaE mutants prove that all the waaE homologues perform the same function. Furthermore, K. pneumoniae, Ser. marcescens and P. mirabilis non-polar waaE mutants showed reduced adhesion and pathogenicity. In addition, the Ser. marcescens and P. murabilis waaE mutants showed reduced swarming motility and ability to form biofilms in vitro. All these characteristics were rescued by reintroduction of the waaE gene independently of its origin. An easy DNA amplification method to detect this gene was established, which also helps in finding the potential presence of this structural feature [beta-D-Glcp-(1-->4)-alpha-L,D-HeppI] in the inner-core LPS of Enterobacteriaceae members with unknown LPS-core structures.
KeywordMeSH Terms
Glucosyltransferases
13. Foultier  B, Troisfontaines  P, Müller  S, Opperdoes  FR, Cornelis  GR,     ( 2002 )

Characterization of the ysa pathogenicity locus in the chromosome of Yersinia enterocolitica and phylogeny analysis of type III secretion systems.

Journal of molecular evolution 55 (1)
PMID : 12165841  :   DOI  :   10.1007/s00239-001-0089-7    
Abstract >>
Several Gram negative bacteria use a complex system called "type III secretion system" (TTSS) to engage their host. The archetype of TTSS is the plasmid-encoded "Yop virulon" shared by the three species of pathogenic Yersinia (Y. pestis, Y. pseudotuberculosis, and Y. enterocolitica). A second TTSS, called Ysa (for Yersinia secretion apparatus) was recently described in Y. enterocolitica 8081, a strain from serotype O:8. In this study, we describe the ysa locus from A127/90, another strain of serotype O:8, and we extend the sequence to several new genes encoding Ysp proteins which are the substrates of this secretion system, and a putative chaperone SycB. According to the deduced protein sequences, the ysa system from A127/90 is identical to that of 8081. It is different from the chromosome-encoded TTSS of Y. pestis but is instead closely related to the Mxi-Spa TTSS of Shigella and to the SPI-1 encoded TTSS of Salmonella enterica. We further demonstrated that the ysa locus is only present in biotype IB strains of Y. enterocolitica. Including this new Ysa system, a phylogenetic analysis of the 26 known TTSSs was carried out, based on the sequence analysis of three conserved proteins. All the TTSSs fall into five different clusters. The phylogenetic tree of these TTSSs is completely different from the evolutionary tree based on 16S RNA, indicating that TTSSs have been distributed by horizontal transfer.
KeywordMeSH Terms
Chromosomes, Bacterial
Phylogeny
14. Fukushima  M, Kakinuma  K, Kawaguchi  R,     ( 2002 )

Phylogenetic analysis of Salmonella, Shigella, and Escherichia coli strains on the basis of the gyrB gene sequence.

Journal of clinical microbiology 40 (8)
PMID : 12149329  :   DOI  :   10.1128/jcm.40.8.2779-2785.2002     PMC  :   PMC120687    
Abstract >>
Phylogenetic analysis of about 200 strains of Salmonella, Shigella, and Escherichia coli was carried out using the nucleotide sequence of the gene for DNA gyrase B (gyrB), which was determined by directly sequencing PCR fragments. The results establish a new phylogenetic tree for the classification of Salmonella, Shigella, and Escherichia coli in which Salmonella forms a cluster separate from but closely related to Shigella and E. coli. In comparison with 16S rRNA analysis, the gyrB sequences indicated a greater evolutionary divergence for the bacteria. Thus, in screening for the presence of bacteria, the gyrB gene might be a useful tool for differentiating between closely related species of bacteria such as Shigella spp. and E. coli. At present, 16S rRNA sequence analysis is an accurate and rapid method for identifying most unknown bacteria to the genus level because the highly conserved 16S rRNA region is easy to amplify; however, analysis of the more variable gyrB sequence region can identify unknown bacteria to the species level. In summary, we have shown that gyrB sequence analysis is a useful alternative to 16S rRNA analysis for constructing the phylogenetic relationships of bacteria, in particular for the classification of closely related bacterial species.
KeywordMeSH Terms
Phylogeny
15. Petersen  S, Young  GM,     ( 2002 )

Essential role for cyclic AMP and its receptor protein in Yersinia enterocolitica virulence.

Infection and immunity 70 (7)
PMID : 12065508  :   DOI  :   10.1128/iai.70.7.3665-3672.2002     PMC  :   PMC128101    
Abstract >>
Insertion mutations were isolated in cya and crp of Yersinia enterocolitica, which encode adenylate cyclase and the cyclic AMP (cAMP) receptor protein (CRP). The cya and crp mutants were affected for the production of proteins exported by the Ysc, Ysa, and flagellar type III secretion systems (TTSS). Protein production by each TTSS was restored when the respective mutation was complemented by a plasmid-encoded copy of the wild-type gene. Both cya and crp mutants exhibited reduced virulence for orally infected BALB/c mice in a 50% lethal dose analysis. Examination of bacterial survival in host tissues showed that cya and crp mutants colonized Peyer's patches and, to a lesser extent, mesenteric lymph nodes. However, the mutants did not appear to disseminate to the liver and spleen of infected mice. An initial examination of the effectiveness of Y. enterocolitica cya and crp mutants to stimulate protective immunity against subsequent challenge with virulent bacteria in mice was promising. The results indicate that the cAMP-CRP regulatory system is required for Y. enterocolitica virulence.
KeywordMeSH Terms
16. Nieto  JM, Madrid  C, Miquelay  E, Parra  JL, Rodríguez  S, Juárez  A,     ( 2002 )

Evidence for direct protein-protein interaction between members of the enterobacterial Hha/YmoA and H-NS families of proteins.

Journal of bacteriology 184 (3)
PMID : 11790731  :   DOI  :   10.1128/jb.184.3.629-635.2002     PMC  :   PMC139531    
Abstract >>
Escherichia coli nucleoid-associated H-NS protein interacts with the Hha protein, a member of a new family of global modulators that also includes the YmoA protein from Yersinia enterocolitica. This interaction has been found to be involved in the regulation of the expression of the toxin alpha-hemolysin. In this study, we further characterize the interaction between H-NS and Hha. We show that the presence of DNA in preparations of copurified His-Hha and H-NS is not directly implicated in the interaction between the proteins. The precise molecular mass of the H-NS protein retained by Hha, obtained by mass spectrometry analysis, does not show any posttranslational modification other than removal of the N-terminal Met residue. We constructed an H-NS-His recombinant protein and found that, as expected, it interacts with Hha. We used a Ni(2+)-nitrilotriacetic acid agarose method for affinity chromatography copurification of proteins to identify the H-NS protein of Y. enterocolitica. We constructed a six-His-YmoA recombinant protein derived from YmoA, the homologue of Hha in Y. enterocolitica, and found that it interacts with Y. enterocolitica H-NS. We also cloned and sequenced the hns gene of this microorganism. In the course of these experiments we found that His-YmoA can also retain H-NS from E. coli. We also found that the hns gene of Y. enterocolitica can complement an hns mutation of E. coli. Finally, we describe for the first time systematic characterization of missense mutant alleles of hha and truncated Hha' proteins, and we report a striking and previously unnoticed similarity of the Hha family of proteins to the oligomerization domain of the H-NS proteins.
KeywordMeSH Terms
Escherichia coli Proteins
17. Nummelin  H, El Tahir  Y, Ollikka  P, Skurnik  M, Goldman  A,     ( 2002 )

Expression, purification and crystallization of a collagen-binding fragment of Yersinia adhesin YadA.

Acta crystallographica. Section D, Biological crystallography 58 (Pt 6 Pt 2)
PMID : 12037311  :   DOI  :   10.1107/s0907444902005231    
Abstract >>
A recombinant form of a collagen-binding fragment of Yersinia enterocolitica serotype O:3 adhesin YadA with an N-terminal polyhistidine affinity tag has been produced in Escherichia coli, purified and crystallized using the sitting-drop vapour-diffusion technique. Crystals belong to the trigonal space group R3, with unit-cell parameters a = b = 67.05, c = 221.95 A, and diffract to 1.55 A resolution on a synchrotron-radiation source.
KeywordMeSH Terms
18. Sorg  I, Goehring  UM, Aktories  K, Schmidt  G,     ( 2001 )

Recombinant Yersinia YopT leads to uncoupling of RhoA-effector interaction.

Infection and immunity 69 (12)
PMID : 11705930  :   DOI  :   10.1128/IAI.69.12.7535-7543.2001     PMC  :   PMC98844    
Abstract >>
Yersinia enterocolitica, Yersinia pseudotuberculosis, and Yersinia pestis deliver different Yop (Yersinia outer proteins) effector proteins into mammalian cells by a type III secretion mechanism. Recently, it was shown that Yersinia producing YopT leads to disruption of the actin cytoskeleton of HeLa cells (M. Iriarte and G. R. Cornelis, Mol. Microbiol. 29:915-929, 1998). To analyze the molecular mechanism of YopT, we cloned and expressed YopT as a glutathione S-transferase fusion protein. Recombinant YopT caused rounding up of embryonic bovine lung cells and redistribution of the actin cytoskeleton rapidly after microinjection. The Escherichia coli cytotoxic necrotizing factor (CNF1), which constitutively activates Rho proteins, was not able to inhibit or revert YopT-induced cell rounding. YopT caused release of RhoA from embryonic bovine lung membranes and released recombinant isoprenylated RhoA from artificial PE or PE/PIP2 vesicles. Incubation of lysate or cytosol with YopT caused inhibition of the RhoA-rhotekin interaction but led to increased RhoA-RhoGDI interaction. It is suggested that inhibition of the interaction between RhoA and effectors is the underlying mechanism of the YopT action on the cytoskeleton.
KeywordMeSH Terms
Escherichia coli Proteins
Intracellular Signaling Peptides and Proteins
19. Jacobi  CA, Gregor  S, Rakin  A, Heesemann  J,     ( 2001 )

Expression analysis of the yersiniabactin receptor gene fyuA and the heme receptor hemR of Yersinia enterocolitica in vitro and in vivo using the reporter genes for green fluorescent protein and luciferase.

Infection and immunity 69 (12)
PMID : 11705959  :   DOI  :   10.1128/IAI.69.12.7772-7782.2001     PMC  :   PMC98873    
Abstract >>
The enteropathogenic Yersinia enterocolitica strains have several systems for scavenging iron from their environment. We have studied the expression of the fyuA gene, which encodes the outer membrane receptor for the siderophore yersiniabactin (Ybt), and the hemR gene, which encodes the receptor for heme, using the reporter genes gfp (encoding green fluorescent protein) and luc (encoding firefly luciferase). To study gene expression in vitro as well as in vivo, we have constructed several translational reporter gene fusions to monitor simultaneously expression of fyuA and hemR or expression of one gene by a gfp-luc tandem reporter. Results of the in vitro expression analysis (liquid media) indicated that fyuA and hemR are strongly derepressed under iron starvation conditions, resulting in strong fluorescence and/or luminescence at 27 degrees C. In the in vivo BALB/C mouse infection model, tissue-specific expression of fyuA and hemR reporter fusions was observed. Surprisingly, fyuA and hemR reporter constructs were weakly expressed by yersiniae located in the liver and intestinal lumen, whereas strong expression was found for yersiniae in the peritoneal cavity and moderate expression was found in the spleen. Strikingly, yersiniae carrying fyuA or hemR reporter fusions exhibited threefold-stronger signals when grown in the peritoneal cavity of mice than those growing under iron derepression in vitro. This hyperexpression suggests that besides Fur derepression, additional activators may be involved in the enhanced expression of fyuA and hemR under peritoneal growth conditions. Differential expression of the fyuA and hemR reporter fusions could not be observed, suggesting similar regulation of fyuA and hemR in the mouse infection model.
KeywordMeSH Terms
Bacterial Proteins
Phenols
Thiazoles
20. Cheng  LW, Kay  O, Schneewind  O,     ( 2001 )

Regulated secretion of YopN by the type III machinery of Yersinia enterocolitica.

Journal of bacteriology 183 (18)
PMID : 11514512  :   DOI  :   10.1128/jb.183.18.5293-5301.2001     PMC  :   PMC95411    
Abstract >>
During infection, Yersinia enterocolitica exports Yop proteins via a type III secretion pathway. Secretion is activated when the environmental concentration of calcium ions is below 100 microM (low-calcium response). Yersiniae lacking yopN (lcrE), yscB, sycN, or tyeA do not inactivate the type III pathway even when the concentration of calcium is above 100 microM (calcium-blind phenotype). Purified YscB and SycN proteins form cytoplasmic complexes that bind a region including amino acids 16 to 100 of YopN, whereas TyeA binds YopN residues 101 to 294. Translational fusion of yopN gene sequences to the 5' end of the npt reporter generates hybrid proteins that are transported by the type III pathway. The signal necessary and sufficient for the type III secretion of hybrid proteins is located within the first 15 codons of yopN. Expression of plasmid-borne yopN, but not of yopN(1-294)-npt, complements the calcium-blind phenotype of yopN mutants. Surprisingly, yopN mutants respond to environmental changes in calcium concentration and secrete YopN(1-294)-Npt in the absence but not in the presence of calcium. tyeA is required for the low-calcium regulation of YopN(1-294)-Npt secretion, whereas sycN and yscB mutants fail to secrete YopN(1-294)-Npt in the presence of calcium. Experiments with yopN-npt fusions identified two other signals that regulate the secretion of YopN. yopN codons 16 to 100 prevent the entry of YopN into the type III pathway, a negative regulatory effect that is overcome by expression of yscB and sycN. The portion of YopN encoded by codons 101 to 294 prevents transport of the polypeptide across the bacterial double membrane envelope in the presence of functional tyeA. These data support a model whereby YopN transport may serve as a regulatory mechanism for the activity of the type III pathway. YscB/SycN binding facilitates the initiation of YopN into the type III pathway, whereas TyeA binding prevents transport of the polypeptide across the bacterial envelope. Changes in the environmental calcium concentration relieve the TyeA-mediated regulation, triggering YopN transport and activating the type III pathway.
KeywordMeSH Terms
Gene Expression Regulation, Bacterial
Membrane Proteins
21. Nelson  KM, Young  GM, Miller  VL,     ( 2001 )

Identification of a locus involved in systemic dissemination of Yersinia enterocolitica.

Infection and immunity 69 (10)
PMID : 11553561  :   DOI  :   10.1128/IAI.69.10.6201-6208.2001     PMC  :   PMC98752    
Abstract >>
A putative LysR-type transcriptional activator, Hre20, was identified previously in an in vivo expression technology screen designed to identify factors which are expressed early during infection by Yersinia enterocolitica (G. M. Young and V. L. Miller, Mol. Microbiol. 25:319-328, 1997). An insertion in hre20, now designated rscR, resulted in increased splenic dissemination of bacteria during infection in a BALB/c mouse model. A nonpolar mutation was generated in rscR, and examination of this strain in the BALB/c mouse model demonstrated that the mutation in rscR was responsible for the increased dissemination to the spleen that was seen in the original experiments. RscR is homologous to the LysR family of transcriptional regulators; thus, a screen was undertaken to identify genes regulated by RscR. A strain containing an insertion in the chromosomal rscR gene and carrying rscR on a plasmid under the control of the inducible araBAD promoter was mutagenized with an mTn5Km-2 transposon containing a promoterless lacZY. Eighteen insertions were identified which appeared to respond to levels of RscR, and these were classified into four allelic groups based on Southern blot hybridization analysis. Representative members were sequenced from three allelic groups. Sequencing revealed insertions in an ORF with no known homologues, a homologue of OmpF of Serratia marcescens, and a locus (designated rscBAC) with similarity to the hmwABC locus of Haemophilus influenzae. The hmwABC locus promotes adherence of H. influenzae to host cells (S. J. Barenkamp and J. W. St. Geme III, Infect. Immun. 62:3320-3328, 1994; J. W. St. Geme III, S. Falkow, and S. J. Barenkamp, Proc. Natl. Acad. Sci. USA 90:2875-2879, 1993). A strain containing a deletion mutant of rscA, the hmwA homologue, exhibits increased splenic dissemination of bacteria during infection in a BALB/c mouse model, similar to the rscR mutant. This suggests that the phenotype of an rscR mutant is due to the loss of RscA.
KeywordMeSH Terms
Genes, Bacterial
22. Snellings  NJ, Popek  M, Lindler  LE,     ( 2001 )

Complete DNA sequence of Yersinia enterocolitica serotype 0:8 low-calcium-response plasmid reveals a new virulence plasmid-associated replicon.

Infection and immunity 69 (7)
PMID : 11402007  :   DOI  :   10.1128/IAI.69.7.4627-4638.2001     PMC  :   PMC98540    
Abstract >>
The complete nucleotide sequence and organization of the Yersinia enterocolitica serotype 0:8 low-calcium-response (LCR) plasmid, pYVe8081, were determined. The 67,720-bp plasmid encoded all the genes known to be part of the LCR stimulon except for ylpA. Eight of 13 intact open reading frames of unknown function identified in pYVe8081 had homologues in Yersinia pestis plasmid pCD1 or in Y. enterocolitica serotype 0:9 plasmid pYVe227. A region of approximately 17 kbp showed no DNA identity to pCD1 or pYVe227 and contained six potential new genes, a possible new replicon, and two intact insertion sequence (IS) elements. One intact IS element, ISYen1, was a new IS belonging to the IS256 family. Several vestigial IS elements appeared different from the IS distribution seen in the other LCR plasmids. The RepA proteins encoded by Y. enterocolitica serotype 0:8 pYVeWA and pYVe8081 were identical. The putative pYVe8081 replicon showed significant homology to the IncL/M replicon of pMU407.1 but was only distantly related to the replicons of pCD1 and pYVe227. In contrast, the putative partitioning genes of pYVe8081 showed 97% DNA identity to the spy/sopABC loci of pCD1 and pYVe227. Sequence analysis suggests that Yersinia LCR plasmids are from a common ancestor but that Y. enterocolitica serotype 0:8 plasmid replicons may have evolved independently via cointegrate formation following a transposition event. The change in replicon structure is predicted to change the incompatibility properties of Y. enterocolitica serotype 0:8 plasmids from those of Y. enterocolitica serotype 0:9 and Y. pestis LCR plasmids.
KeywordMeSH Terms
DNA, Bacterial
Plasmids
Replicon
23. Heusipp  G, Young  GM, Miller  VL,     ( 2001 )

HreP, an in vivo-expressed protease of Yersinia enterocolitica, is a new member of the family of subtilisin/kexin-like proteases.

Journal of bacteriology 183 (12)
PMID : 11371518  :   DOI  :   10.1128/JB.183.12.3556-3563.2001     PMC  :   PMC95231    
Abstract >>
The role of proteases in pathogenesis is well established for several microorganisms but has not been described for Yersinia enterocolitica. Previously, we identified a gene, hreP, which showed significant similarity to proteases in a screen for chromosomal genes of Y. enterocolitica that were exclusively expressed during an infection of mice. We cloned this gene by chromosome capture and subsequently determined its nucleotide sequence. Like inv, the gene encoding the invasin protein of Y. enterocolitica, hreP is located in a cluster of flagellum biosynthesis and chemotaxis genes. The genomic organization of this chromosomal region is different in Escherichia coli, Salmonella, and Yersinia pestis than in Y. enterocolitica. Analysis of the distribution of hreP between different Yersinia isolates and the relatively low G+C content of the gene suggests acquisition by horizontal gene transfer. Sequence analysis also revealed that HreP belongs to a family of eukaryotic subtilisin/kexin-like proteases. Together with the calcium-dependent protease PrcA of Anabaena variabilis, HreP forms a new subfamily of bacterial subtilisin/kexin-like proteases which might have originated from a common eukaryotic ancestor. Like other proteases of this family, HreP is expressed with an N-terminal prosequence. Expression of an HreP-His(6) tag fusion protein in E. coli revealed that HreP undergoes autocatalytic processing at a consensus cleavage site of subtilisin/kexin-like proteases, thereby releasing the proprotein.
KeywordMeSH Terms
Bacterial Proteins
Genome, Bacterial
Proprotein Convertases
Saccharomyces cerevisiae Proteins
24. Bertin  P, Hommais  F, Krin  E, Soutourina  O, Tendeng  C, Derzelle  S, Danchin  A,     ( 2001 )

H-NS and H-NS-like proteins in Gram-negative bacteria and their multiple role in the regulation of bacterial metabolism.

Biochimie 83 (2)
PMID : 11278074  :   DOI  :   10.1016/s0300-9084(01)01247-0    
Abstract >>
In Escherichia coli, the H-NS protein plays an important role in the structure and the functioning of bacterial chromosome. A homologous protein has also been identified in several enteric bacteria and in closely related organisms such as Haemophilus influenzae. To get information on their structure and their function, we identified H-NS-like proteins in various microorganisms by different procedures. In silico analysis of their amino acid sequence and/or in vivo experiments provide evidence that more than 20 proteins belong to the same class of regulatory proteins. Moreover, large scale technologies demonstrate that, at least in E. coli, the loss of motility in hns mutants results from a lack of flagellin biosynthesis, due to the in vivo repression of flagellar gene expression. In contrast, several genes involved in adaptation to low pH are strongly induced in a H-NS deficient strain, resulting in an increased resistance to acidic stress. Finally, expression profiling and phenotypic analysis suggest that, unlike H-NS, its paralogous protein StpA does not play any role in these processes.
KeywordMeSH Terms
25. Darwin  AJ, Miller  VL,     ( 2001 )

The psp locus of Yersinia enterocolitica is required for virulence and for growth in vitro when the Ysc type III secretion system is produced.

Molecular microbiology 39 (2)
PMID : 11136463  :   DOI  :   10.1046/j.1365-2958.2001.02235.x    
Abstract >>
The phage shock protein locus (pspFpspABCDE) of Escherichia coli has proved to be something of an enigma since its discovery. The physiological functions of the psp locus, including those of the predicted effector protein PspA, are unknown. In a previous genetic screen, we determined that a Yersinia enterocolitica pspC mutant was severely attenuated for virulence. In this study, the psp locus of Y. enterocolitica was characterized further. The pspC gene of Y. enterocolitica was found to be important for normal growth when the Ysc type III secretion system was expressed in the laboratory. This growth defect was specifically caused by production of the secretin protein, YscC. Expression of the psp genes was induced when the type III secretion system was functional or when only the yscC gene was expressed. This induction of psp gene expression required a functional pspC gene. Most significantly, evidence suggests that the expression of at least one gene that is not part of the psp locus is regulated by Psp proteins. This unidentified gene (or genes) may also be important for growth when the type III secretion system is expressed. These conclusions are supported by the effects of various psp mutations on virulence. This is the first indication that Psp proteins might be involved in the regulation of genes besides the psp locus itself.
KeywordMeSH Terms
Gene Expression Regulation, Bacterial
26. Carlson  S, Pederson  KJ, Haller  JC,     ( 2000 )

A chromosomally encoded type III secretion pathway in Yersinia enterocolitica is important in virulence.

Molecular microbiology 36 (6)
PMID : 10931293  :   DOI  :   10.1046/j.1365-2958.2000.01964.x    
Abstract >>
Numerous Gram-negative bacteria use a type III, or contact dependent, secretion system to deliver proteins into the cytosol of host cells. All of these systems identified to date have been shown to have a role in pathogenesis. We have identified 13 genes on the Yersinia enterocolitica chromosome that encode a type III secretion apparatus plus two associated putative regulatory genes. In order to determine the function of this chromosomally-encoded secretion apparatus, we created an in frame deletion of a gene that has homology to the hypothesized inner membrane pore, ysaV. The ysaV mutant strain failed to secrete eight proteins, called Ysps, normally secreted by the parental strain when grown at 28 degrees C in Luria-Bertani (LB) broth supplemented with 0.4 M NaCl. Disruption of the ysaV gene had no effect on motility or phospholipase activity, suggesting this chromosomally encoded type III secretion pathway is distinct from the flagella secretion pathway of Y. enterocolitica. Deletion of the ysaV gene in a virulence plasmid positive strain had no effect on in vitro secretion of Yops by the plasmid-encoded type III secretion apparatus. Secretion of the Ysps was unaffected by the presence or absence of the virulence plasmid, suggesting the chromosomally encoded and plasmid-encoded type III secretion pathways act independently. Y. enterocolitica thus has three type III secretion pathways that appear to act independently. The ysaV mutant strain was somewhat attenuated in virulence compared with the wild type in the mouse oral model of infection (an approximately 0.9 log difference in LD50). The ysaV mutant strain was nearly as virulent as the wild type when inoculated intraperitoneally in the mouse model. A ysaV probe hybridized to sequences in other Yersinia spp. and homologues were found in the incomplete Y. pestis genome sequence, indicating a possible role for this system throughout the genus.
KeywordMeSH Terms
Chromosomes, Bacterial
Genes, Bacterial
27. Kuusela  P, Tahir  YE,     ( 2000 )

Functional mapping of the Yersinia enterocolitica adhesin YadA. Identification Of eight NSVAIG - S motifs in the amino-terminal half of the protein involved in collagen binding.

Molecular microbiology 37 (1)
PMID : 10931316  :   DOI  :   10.1046/j.1365-2958.2000.01992.x    
Abstract >>
The virulence plasmid-encoded YadA of Yersinia enterocolitica serotype O:3 is a 430-amino-acid outer membrane protein, synthesized with a 25-amino-acid signal peptide. YadA forms homotrimeric surface structures that function as adhesin between bacteria and collagen as well as other host proteins. The structure-function relationships of YadA were studied, and the collagen-binding determinants of YadA were located to its amino-terminal half. Collagen did not bind to any of the overlapping 16-mer YadA peptides, indicating that the collagen binding site of YadA is conformational. Epitope mapping of YadA identified 12 linear antigenic epitopes altogether. Seven epitopes were uniquely recognized by an anti-YadA antiserum able to inhibit collagen binding. Four of these epitopes shared a motif NSVAIG-S that is repeated eight times within the N-terminal half of YadA. Site-directed mutagenesis showed that these motifs are absolutely required for YadA-mediated collagen binding, revealing a novel type of collagen-binding mechanism.
KeywordMeSH Terms
28. Hoiczyk  E, Roggenkamp  A, Reichenbecher  M, Lupas  A, Heesemann  J,     ( 2000 )

Structure and sequence analysis of Yersinia YadA and Moraxella UspAs reveal a novel class of adhesins.

The EMBO journal 19 (22)
PMID : 11080146  :   DOI  :   10.1093/emboj/19.22.5989     PMC  :   PMC305836    
Abstract >>
The non-fimbrial adhesins, YadA of enteropathogenic Yersinia species, and UspA1 and UspA2 of Moraxella catarrhalis, are established pathogenicity factors. In electron micrographs, both surface proteins appear as distinct 'lollipop'-shaped structures forming a novel type of surface projection on the outer membranes. These structures, amino acid sequence analysis of these molecules and yadA gene manipulation suggest a tripartite organization: an N-terminal oval head domain is followed by a putative coiled-coil rod and terminated by a C-terminal membrane anchor domain. In YadA, the head domain is involved in autoagglutination and binding to host cells and collagen. Analysis of the coiled-coil segment of YadA revealed unusual pentadecad repeats with a periodicity of 3.75, which differs significantly from the 3.5 periodicity found in the Moraxella UspAs and other canonical coiled coils. These findings predict that the surface projections are formed by oligomers containing right- (Yersinia) or left-handed (Moraxella) coiled coils. Strikingly, sequence comparison revealed that related proteins are found in many proteobacteria, both human pathogenic and environmental species, suggesting a common role in adaptation to specific ecological niches.
KeywordMeSH Terms
29. Anderson  MS, Eveland  SS, Price  NP,     ( 2000 )

Conserved cytoplasmic motifs that distinguish sub-groups of the polyprenol phosphate:N-acetylhexosamine-1-phosphate transferase family.

FEMS microbiology letters 191 (2)
PMID : 11024259  :   DOI  :   10.1111/j.1574-6968.2000.tb09335.x    
Abstract >>
WecA, MraY and WbcO are conserved members of the polyprenol phosphate:N-acetylhexosamine-1-phosphate transferase family involved in the assembly of bacterial cell walls, and catalyze reactions involving a membrane-associated polyprenol phosphate acceptor substrate and a cytoplasmically located UDP-D-amino sugar donor. MraY, WbcO and WecA purportedly utilize different UDP-sugars, although the molecular basis of this specificity is largely unknown. However, domain variations involved in specificity are predicted to occur on the cytoplasmic side of the membrane, adjacent to conserved domains involved in the mechanistic activity, and with access to the cytoplasmically located sugar nucleotides. Conserved C-terminal domains have been identified that satisfy these criteria. Topological analyses indicate that they form the highly basic, fifth cytoplasmic loop between transmembrane regions IX and X. Four diverse loops are apparent, for MraY, WecA, WbcO and RgpG, that uniquely characterize these sub-groups of the transferase family, and a correlation is evident with the known or implied UDP-sugar specificity.
KeywordMeSH Terms
Escherichia coli Proteins
Transferases
30. Badger  JL, Young  BM, Darwin  AJ, Miller  VL,     ( 2000 )

Yersinia enterocolitica ClpB affects levels of invasin and motility.

Journal of bacteriology 182 (19)
PMID : 10986262  :   DOI  :   10.1128/jb.182.19.5563-5571.2000     PMC  :   PMC111002    
Abstract >>
Expression of the Yersinia enterocolitica inv gene is dependent on growth phase and temperature. inv is maximally expressed at 23 degrees C in late-exponential- to early-stationary-phase cultures. We previously reported the isolation of a Y. enterocolitica mutant (JB1A8v) that shows a decrease in invasin levels yet is hypermotile when grown at 23 degrees C. JB1A8v has a transposon insertion within uvrC. Described here is the isolation and characterization of a clone that suppresses these mutant phenotypes of the uvrC mutant JB1A8v. This suppressing clone encodes ClpB (a Clp ATPase homologue). The Y. enterocolitica ClpB homologue is 30 to 40% identical to the ClpB proteins from various bacteria but is 80% identical to one of the two ClpB homologues of Yersinia pestis. A clpB::TnMax2 insertion mutant (JB69Qv) was constructed and determined to be deficient in invasin production and nonmotile when grown at 23 degrees C. Analysis of inv and fleB (flagellin gene) transcript levels in JB69Qv suggested that ClpB has both transcriptional and posttranscriptional effects. In contrast, a clpB null mutant, BY1v, had no effect on invasin levels or motility. A model accounting for these observations is presented.
KeywordMeSH Terms
Adhesins, Bacterial
Escherichia coli Proteins
31. Voigt  I, Strauch  E,     ( 2000 )

Use of a plasmid of a yersinia enterocolitica biogroup 1A strain for the construction of cloning vectors.

Journal of biotechnology 79 (1)
PMID : 10817342  :  
Abstract >>
A plasmid with a size of 2,682 base pairs isolated from the Yersinia enterocolitica biogroup 1A strain # 29807 was characterized in respect to its suitability as a basic replicon for cloning vectors. The copy number of the plasmid was determined to be approximately 14 copies per cell and it was shown to be compatible with vectors with an origin of replication derived from ColE1 and p115A. The replication region of the plasmid encodes a primer RNAI and countertranscript RNAII. Two vectors, pIV1 and pIV2, containing a kanamycin resistance gene and the lacZalpha fragment with the multiple cloning site of pBluescriptSK + were constructed. A mobilizable derivative was successfully introduced into different bacteria belonging to the family Enterobacteriacea. To prove the applicability of the novel vectors for cloning purposes, a 13 kb hemolysin operon of Escherichia coli was inserted into pIV1, and the resulting recombinant plasmid was stably maintained and expressed in E. coli and Y. enterocolitica.
KeywordMeSH Terms
Cloning, Molecular
Genetic Vectors
32. Seoane  A,     ( 2000 )

Identification of a streptogramin A acetyltransferase gene in the chromosome of Yersinia enterocolitica.

Antimicrobial agents and chemotherapy 44 (4)
PMID : 10722489  :   DOI  :   10.1128/aac.44.4.905-909.2000     PMC  :   PMC89790    
Abstract >>
Streptogramins are polypeptide antibiotics inhibiting protein synthesis by the prokaryotic ribosome. Gram-positive organisms are susceptible to streptogramins, while most gram-negative bacteria are intrinsically resistant. We have found a genomic fragment from a Yersinia enterocolitica isolate with an open reading frame coding for a polypeptide similar to the virginiamycin acetyltransferases found in various plasmids from gram-positive bacteria. The susceptible Escherichia coli strain DB10 was transformed to resistance to the type A streptogramins and to mixed (A + B) streptogramins upon introduction of a plasmid containing that gene. In addition, we showed streptogramin acetylating activity in vitro dependent on the presence of the Y. enterocolitica sat gene. Southern blot hybridization experiments showed that the sat gene was present in all the Y. enterocolitica isolates examined. These data together show that the gene in the Y. enterocolitica chromosome encoded an active streptogramin acetyltransferase. The deduced sequence of the Y. enterocolitica Sat protein was close to those of sat gene products found in gram-positive bacteria and cyanobacteria, suggesting a common evolutionary origin.
KeywordMeSH Terms
Bacterial Proteins
33. Revell  PA,     ( 2000 )

A chromosomally encoded regulator is required for expression of the Yersinia enterocolitica inv gene and for virulence.

Molecular microbiology 35 (3)
PMID : 10672189  :   DOI  :   10.1046/j.1365-2958.2000.01740.x    
Abstract >>
The primary invasion factor of Yersinia enterocolitica, invasin, is encoded by inv. inv expression is regulated in response to pH, growth phase and temperature. In vitro, inv is maximally expressed at 26 degrees C, pH 8.0, or 37 degrees C, pH 5.5, in early stationary phase. At 37 degrees C, pH 8.0, inv is weakly expressed. To identify which gene(s) are required for inv regulation, we screened for transposon insertions that decreased expression of an inv-'phoA chromosomal reporter at 26 degrees C. Of 30 000 mutants screened, two were identified that had negligible inv expression in all conditions tested. Both of these independent mutants had an insertion into the same gene, designated rovA (regulator of virulence). RovA has 77% amino acid identity to the Salmonella typhimurium transcriptional regulator SlyA. Complementation with the wild-type rovA allele restores wild-type inv expression as monitored by Western blot analysis, tissue culture invasion assay and alkaline phosphatase assay. There is also a significant decrease in invasin levels in bacteria recovered from mice infected with the rovA mutant; therefore, RovA regulates inv expression in vivo as well as in vitro. In the mouse infection model, an inv mutant has a wild-type LD50, even though the kinetics of infection is changed. In contrast, the rovA mutant has altered kinetics, as well as a 70-fold increase in the LD50 compared with wild type. Furthermore, because the rovA mutant is attenuated in the mouse model, this suggests that RovA regulates other virulence factors in addition to inv. Analysis of other proposed virulence factors such as Ail, YadA and the Yop proteins shows no regulatory role for RovA. The more severe animal phenotype combined with the lack of impact on known virulence genes aside from inv suggests RovA regulates potentially novel virulence genes of Y. enterocolitica during infection.
KeywordMeSH Terms
Adhesins, Bacterial
Transcription Factors
34. Zurth  K, Morelli  G, Torrea  G, Achtman  M,     ( 1999 )

Yersinia pestis, the cause of plague, is a recently emerged clone of Yersinia pseudotuberculosis.

Proceedings of the National Academy of Sciences of the United States of America 96 (24)
PMID : 10570195  :   DOI  :   10.1073/pnas.96.24.14043     PMC  :   PMC24187    
Abstract >>
Plague, one of the most devastating diseases of human history, is caused by Yersinia pestis. In this study, we analyzed the population genetic structure of Y. pestis and the two other pathogenic Yersinia species, Y. pseudotuberculosis and Y. enterocolitica. Fragments of five housekeeping genes and a gene involved in the synthesis of lipopolysaccharide were sequenced from 36 strains representing the global diversity of Y. pestis and from 12-13 strains from each of the other species. No sequence diversity was found in any Y. pestis gene, and these alleles were identical or nearly identical to alleles from Y. pseudotuberculosis. Thus, Y. pestis is a clone that evolved from Y. pseudotuberculosis 1,500-20,000 years ago, shortly before the first known pandemics of human plague. Three biovars (Antiqua, Medievalis, and Orientalis) have been distinguished by microbiologists within the Y. pestis clone. These biovars form distinct branches of a phylogenetic tree based on restriction fragment length polymorphisms of the locations of the IS100 insertion element. These data are consistent with previous inferences that Antiqua caused a plague pandemic in the sixth century, Medievalis caused the Black Death and subsequent epidemics during the second pandemic wave, and Orientalis caused the current plague pandemic.
KeywordMeSH Terms
35. Francis  KP, Rapposch  S, Neuhaus  K,     ( 1999 )

Pathogenic Yersinia species carry a novel, cold-inducible major cold shock protein tandem gene duplication producing both bicistronic and monocistronic mRNA.

Journal of bacteriology 181 (20)
PMID : 10515936  :   PMC  :   PMC103781    
Abstract >>
Inverse PCR was used to amplify major cold shock protein (MCSP) gene families from a diverse range of bacteria, including the psychrotolerant Yersinia enterocolitica, which was found to have two almost identical MCSP coding regions (cspA1 and cspA2) located approximately 300 bp apart. This tandem gene duplication was also found in Y. pestis, Y. pseudotuberculosis, and Y. ruckeri but not in other bacteria. Analysis of the transcriptional regulation of this MCSP gene in Y. enterocolitica, performed by using both reverse transcriptase-PCR and Northern blot assays, showed there to be two cold-inducible mRNA templates arising from this locus: a monocistronic template of approximately 450 bp (cspA1) and a bicistronic template of approximately 900 bp (cspA1/A2). The former may be due to a secondary structure between cspA1 and cspA2 causing either 3' degradation protection of cspA1 or, more probably, partial termination after cspA1. Primer extension experiments identified a putative transcriptional start site (+1) which is flanked by a cold-box motif and promoter elements (-10 and -35) similar to those found in Escherichia coli cold-inducible MCSP genes. At 30 degrees C, the level of both mRNA molecules was negligible; however, upon a temperature downshift to 10 degrees C, transcription of the bicistronic mRNA was both substantial (300-fold increase) and immediate, with transcription of the monocistronic mRNA being approximately 10-fold less (30-fold increase) and significantly slower. The ratio of bicistronic to monocistronic mRNA changed with time after cold shock and was higher when cells were shocked to a lower temperature. High-resolution, two-dimensional protein gel electrophoresis showed that synthesis of the corresponding proteins, both CspA1 and CspA2, was apparent after only 10 min of cold shock from 30 degrees C to 10 degrees C. The data demonstrate an extraordinary capacity of the psychrotolerant Y. enterocolitica to produce major cold shock proteins upon cold shock.
KeywordMeSH Terms
Cold Temperature
Gene Duplication
Genes, Bacterial
36. Andor  A, Jacobi  CA, Aepfelbacher  M, Zumbihl  R,     ( 1999 )

The cytotoxin YopT of Yersinia enterocolitica induces modification and cellular redistribution of the small GTP-binding protein RhoA.

The Journal of biological chemistry 274 (41)
PMID : 10506187  :   DOI  :   10.1074/jbc.274.41.29289    
Abstract >>
Pathogenic Yersinia enterocolitica produces two virulence plasmid-encoded cytotoxins, YopE and YopT, that are translocated into target cells where they disrupt the actin cytoskeleton. Here we show that infection of cells with wild type Y. enterocolitica and a yopE mutant, but not with a yopT mutant, induces an increase in the electrophoretic mobility of the small GTPase RhoA. As tested by isoelectric focusing, YopT-dependent modification resulted in an acidic shift of RhoA. Furthermore, RhoA modification induced by YopT was accompanied by redistribution of membrane-bound RhoA toward the cytosol. Finally, a yopE mutant of Y. enterocolitica expressing the cytotoxic activity of YopT specifically disrupted RhoA-controlled actin stress fibers. These findings provide evidence for inactivation of RhoA by the translocated Y. enterocolitica cytotoxin YopT and suggest a novel inhibitory modification of RhoA by a bacterial virulence factor.
KeywordMeSH Terms
37. Fischer  D, Schubert  S,     ( 1999 )

Ferric enterochelin transport in Yersinia enterocolitica: molecular and evolutionary aspects.

Journal of bacteriology 181 (20)
PMID : 10515929  :   PMC  :   PMC103774    
Abstract >>
Yersinia enterocolitica is well equipped for siderophore piracy, encompassing the utilization of siderophores such as ferrioxamine, ferrichrome, and ferrienterochelin. In this study, we report on the molecular and functional characterization of the Yersinia fep-fes gene cluster orthologous to the Escherichia coli ferrienterochelin transport genes (fepA, fepDGC, and fepB) and the esterase gene fes. In vitro transcription-translation analysis identified polypeptides of 30 and 35 kDa encoded by fepC and fes, respectively. A frameshift mutation within the fepA gene led to expression of a truncated polypeptide of 40 kDa. The fepD, fepG, and fes genes of Y. enterocolitica were shown to complement corresponding E. coli mutants. Insertional mutagenesis of fepD or fes genes abrogates enterochelin-supported growth of Y. enterocolitica on iron-chelated media. In contrast to E. coli, the fep-fes gene cluster in Y. enterocolitica consists solely of genes required for uptake and utilization of enterochelin (fep) and not of enterochelin synthesis genes such as entF. By Southern hybridization, fepDGC and fes sequences could be detected in Y. enterocolitica biotypes IB, IA, and II but not in biotype IV strains, Yersinia pestis, and Yersinia pseudotuberculosis strains. According to sequence alignment data and the coherent structure of the Yersinia fep-fes gene cluster, we suggest early genetic divergence of ferrienterochelin uptake determinants among species of the family Enterobacteriaceae.
KeywordMeSH Terms
Escherichia coli Proteins
Evolution, Molecular
Genes, Bacterial
Multigene Family
38. Shen  P, Samsel  L, Liu  R, Allen  T,     ( 1999 )

Phylogenetic analysis of L4-mediated autogenous control of the S10 ribosomal protein operon.

Journal of bacteriology 181 (19)
PMID : 10498727  :   PMC  :   PMC103642    
Abstract >>
We investigated the regulation of the S10 ribosomal protein (r-protein) operon among members of the gamma subdivision of the proteobacteria, which includes Escherichia coli. In E. coli, this 11-gene operon is autogenously controlled by r-protein L4. This regulation requires specific determinants within the untranslated leader of the mRNA. Secondary structure analysis of the S10 leaders of five enterobacteria (Salmonella typhimurium, Citrobacter freundii, Yersinia enterocolitica, Serratia marcescens, and Morganella morganii) and two nonenteric members of the gamma subdivision (Haemophilus influenzae and Vibrio cholerae) shows that these foreign leaders share significant structural homology with the E. coli leader, particularly in the region which is critical for L4-mediated autogenous control in E. coli. Moreover, these heterologous leaders produce a regulatory response to L4 oversynthesis in E. coli. Our results suggest that an E. coli-like L4-mediated regulatory mechanism may operate in all of these species. However, the mechanism is not universally conserved among the gamma subdivision members, since at least one, Pseudomonas aeruginosa, does not contain the required S10 leader features, and its leader cannot provide the signals for regulation by L4 in E. coli. We speculate that L4-mediated autogenous control developed during the evolution of the gamma branch of proteobacteria.
KeywordMeSH Terms
Gene Expression Regulation, Bacterial
Operon
Regulatory Sequences, Nucleic Acid
39. Noelting  C, Rakin  A,     ( 1999 )

Common and specific characteristics of the high-pathogenicity island of Yersinia enterocolitica.

Infection and immunity 67 (10)
PMID : 10496905  :   PMC  :   PMC96880    
Abstract >>
Yersinia pestis, Y. pseudotuberculosis O:1, and Y. enterocolitica biogroup 1B strains carry a high-pathogenicity island (HPI), which mediates biosynthesis and uptake of the siderophore yersiniabactin and a mouse-lethal phenotype. The HPI of Y. pestis and Y. pseudotuberculosis (Yps HPI) are highly conserved in sequence and organization, while the HPI of Y. enterocolitica (Yen HPI) differs significantly. The 43,393-bp Yen HPI sequence of Y. enterocolitica WA-C, serotype O:8, was completed and compared to that of the Yps HPI of Y. pseudotuberculosis PB1, serotype O:1A. A common GC-rich region (G+C content, 57.5 mol%) of 30.5 kb is conserved between yersinia strains. This region carries genes for yersiniabactin biosynthesis, regulation, and uptake and thus can be considered the functional "core" of the HPI. In contrast, the second part of the HPI is AT rich and completely different in two evolutionary lineages of the HPI, being 12.8 kb in the Yen HPI and 5.6 kb in the Yps HPI. The variable part acquired one IS100 element in the Yps HPI and accumulated four insertion elements, IS1328, IS1329, IS1400, and IS1222, in the Yen HPI. The insertion of a 125-bp ERIC sequence modifies the structure of the promoter of the ybtA yersiniabactin regulator in the Yen HPI. In contrast to the precise excision of the Yps HPI in Y. pseudotuberculosis, the Yen HPI suffers imprecise deletions. The Yen HPI is stably integrated in one of the three asn tRNA copies in Y. enterocolitica biogroup 1B (serotypes O:8, O:13, O:20, and O:21), probably due to inactivation of the putative integrase. The 17-bp duplications of the 3' end of the asnT RNA are present in both Yersinia spp. The HPI attachment site is unoccupied in nonpathogenic Y. enterocolitica NF-O, biogroup 1A, serotype O:5. The HPI of Yersinia is a composite and widely spread genomic element with a highly conserved yersiniabactin functional "core" and a divergently evolved variable part.
KeywordMeSH Terms
40. Buchrieser  C, Prentice  M, Guiyoule  A, Bach  S,     ( 1999 )

The high-pathogenicity island of Yersinia enterocolitica Ye8081 undergoes low-frequency deletion but not precise excision, suggesting recent stabilization in the genome.

Infection and immunity 67 (10)
PMID : 10496882  :   PMC  :   PMC96857    
Abstract >>
Highly pathogenic strains of Yersinia pestis, Y. pseudotuberculosis, and Y. enterocolitica are characterized by the possession of a pathogenicity island designated the high-pathogenicity island (HPI). This 35- to 45-kb island carries an iron uptake system named the yersiniabactin locus. While the HPIs of Y. pestis and Y. pseudotuberculosis are subject to high-frequency spontaneous deletion from the chromosome, we were initially unable to obtain HPI-deleted Y. enterocolitica 1B isolates. In the present study, using a positive selection strategy, we identified three HPI-deleted mutants of Y. enterocolitica strain Ye8081. In these three independent clones, the chromosomal deletion was not limited to the HPI but encompassed a larger DNA fragment of approximately 140 kb. Loss of this fragment, which occurred at a frequency of approximately 5 x 10(-7), resulted in the disappearance of several phenotypic traits, such as growth in a minimal medium, hydrolysis of o-nitrophenyl-beta-D-thiogalactopyranoside, Tween esterase activity, and motility, and in a decreased virulence for mice. However, no precise excision of the Ye8081 HPI was observed. To gain more insight into the molecular basis for this phenomenon, the putative machinery of HPI excision in Y. enterocolitica was analyzed and compared to that in Y. pseudotuberculosis. We show that the probable reasons for failure of precise excision of the HPI of Y. enterocolitica Ye8081 are (i) the interruption of the P4-like integrase gene located close to its right-hand boundary by a premature stop codon and (ii) lack of conservation of 17-bp att-like sequences at both extremities of the HPI. These mutations may represent a process of HPI stabilization in the species Y. enterocolitica.
KeywordMeSH Terms
Genome, Bacterial
41. Odaert  M, Trieu-Cuot  P, Simonet  M,     ( 1999 )

Characterization of IS1541-like elements in Yersinia enterocolitica and Yersinia pseudotuberculosis.

FEMS microbiology letters 176 (1)
PMID : 10418150  :   DOI  :   10.1111/j.1574-6968.1999.tb13666.x    
Abstract >>
We characterized Yersinia enterocolitica and Yersinia pseudotuberculosis insertion sequences related to insertion sequence 1541, recently identified in Yersinia pestis. For each of the two species, two insertion sequence copies were cloned and sequenced. Genetic elements from Y. pseudotuberculosis were almost identical to insertion sequence 1541, whereas these from Y. enterocolitica were less related. Phylogenetic analysis of the putative transposases encoded by insertion sequences from the three pathogenic members of the genus Yersinia showed that they clustered with those encoded by Escherichia coli and Salmonella enterica elements belonging to the insertion sequence 200/insertion sequence 605 group. Insertion sequences originating from Y. pestis and Y. pseudotuberculosis constitute a monophyletic lineage distinct from that of Y. enterocolitica.
KeywordMeSH Terms
42. Revear  L, Hotchkiss  A, Savary  B,     ( 1999 )

Genetic and biochemical characterization of an exopolygalacturonase and a pectate lyase from Yersinia enterocolitica.

Canadian journal of microbiology 45 (5)
PMID : 10446714  :   DOI  :   10.1139/w99-034    
Abstract >>
Yersinia enterocolitica, an invasive foodborne human pathogen, degrades polypectate by producing two depolymerizing enzymes, pectate lyase (PL) and polygalacturonase (PG). The gene encoding the PG activity, designated pehY, was located in a 3-kb genomic fragment of Y. enterocolitica ATCC 49397. The complete nucleotide sequence of this 3-kb fragment was determined and an open reading frame consisting of 1803 bp was predicted to encode a PG protein with an estimated M(r) of 66 kDa and pI of 6.3. The amino acid sequence of prePG showed 59 and 43% identity to that of the exopolygalacturonase (exoPG) of Erwinia chrysanthemi and Ralstonia solanacearum, respectively. The Y. enterocolitica PG overproduced in Escherichia coli was purified to near homogeneity using perfusion cation exchange chromatography. Analysis of the PG depolymerization products by high performance anion-exchange chromatography and pulsed amperometric detection (HPAEC-PAD) revealed the exolytic nature of this enzyme. The Y. enterocolitica PL overproduced in E. coli was also partially purified and the M(r) and pI were estimated to be 55 kDa and 5.2, respectively. HPAEC-PAD analysis of the PL depolymerization products indicated the endolytic nature of this enzyme. Southern hybridization analyses revealed that pehY and pel genes of Y. enterocolitica are possibly encoded in the chromosome rather than in the plasmid. Purified exopolygalacturonase (over 10 activity units) was unable to macerate plant tissues.
KeywordMeSH Terms
43. Forst  S,     ( 1999 )

Identification of a conserved N-terminal sequence involved in transmembrane signal transduction in EnvZ.

Journal of bacteriology 181 (17)
PMID : 10464234  :   PMC  :   PMC94069    
Abstract >>
To determine whether N-terminal sequences are involved in the transmembrane signaling mechanism of EnvZ, the nucleotide sequences of envZ genes from several enteric bacteria were determined. Comparative analysis revealed that the amino acid sequence between Pro41 and Glu53 was highly conserved. To further analyze the role of the conserved sequence, envZ of Escherichia coli was subjected to random PCR mutagenesis and mutant alleles that produced a high-osmolarity phenotype, in which ompF was repressed, were isolated. The mutations identified clustered within, as well as adjacent to, the Pro41-to-Glu53 sequence. These findings suggest that the conserved Pro41-to-Glu53 sequence is involved in the signal transduction mechanism of EnvZ.
KeywordMeSH Terms
Conserved Sequence
Escherichia coli Proteins
Multienzyme Complexes
Signal Transduction
44. Young  GM, Miller  VL,     ( 1999 )

The Yersinia enterocolitica motility master regulatory operon, flhDC, is required for flagellin production, swimming motility, and swarming motility.

Journal of bacteriology 181 (9)
PMID : 10217774  :   PMC  :   PMC93725    
Abstract >>
The ability to move over and colonize surface substrata has been linked to the formation of biofilms and to the virulence of some bacterial pathogens. Results from this study show that the gastrointestinal pathogen Yersinia enterocolitica can migrate over and colonize surfaces by swarming motility, a form of cooperative multicellular behavior. Immunoblot analysis and electron microscopy indicated that swarming motility is dependent on the same flagellum organelle that is required for swimming motility, which occurs in fluid environments. Furthermore, motility genes such as flgEF, flgMN, flhBA, and fliA, known to be required for the production of flagella, are essential for swarming motility. To begin to investigate how environmental signals are processed and integrated by Y. enterocolitica to stimulate the production of flagella and regulate these two forms of cell migration, the motility master regulatory operon, flhDC, was cloned. Mutations within flhDC completely abolished swimming motility, swarming motility, and flagellin production. DNA sequence analysis revealed that this locus is similar to motility master regulatory operons of other gram-negative bacteria. Genetic complementation and functional analysis of flhDC indicated that it is required for the production of flagella. When flhDC was expressed from an inducible ptac promoter, flagellin production was shown to be dependent on levels of flhDC expression. Phenotypically, induction of the ptac-flhDC fusion also corresponded to increased levels of both swimming and swarming motility.
KeywordMeSH Terms
Operon
45. Young  JM, Park  DC,     ( 2007 )

Relationships of plant pathogenic enterobacteria based on partial atpD, carA, and recA as individual and concatenated nucleotide and peptide sequences.

Systematic and applied microbiology 30 (5)
PMID : 17451899  :   DOI  :   10.1016/j.syapm.2007.03.002    
Abstract >>
Relationships of the genera in the Enterobacteriaceae containing plant pathogenic species: Brenneria, Dickeya, Enterobacter, Erwinia, Pantoea, Pectobacterium, and Samsonia, were investigated by comparison of their nucleotide and peptide sequences of atpD, carA, recA, and the concatenated sequences. Erwinia spp. and Pantoea spp., with Pectobacterium cypripedii, formed a group distinct from other pathogenic taxa. Pectobacterium, Brenneria, Dickeya, and Samsonia formed a contiguous clade. Samsonia was usually concurrent with Pectobacterium. Most Brenneria were also close to Pectobacterium, suggesting that these three taxa might be better represented as a single genus. Brenneria quercina was not closely associated with other members of this genus and may represent a separate genus. The sequences representing Dickeya were distinct, further supporting the generic status of the taxon. Plant pathogenic Enterobacter spp. display such sequence variability that few definite conclusions as to their specific placement could be made. These data highlight the difficulty of drawing reliable and robust taxonomic conclusions based on comparative analysis of sequence data without some independent criterion to calibrate a scale for diversity.
KeywordMeSH Terms
46. Gulati  PS, Virdi  JS,     ( 2007 )

The rrn locus and gyrB genotyping confirm the existence of two clonal groups in strains of Yersinia enterocolitica subspecies palearctica biovar 1A.

Research in microbiology 158 (3)
PMID : 17349780  :   DOI  :   10.1016/j.resmic.2006.11.011    
Abstract >>
Eighty-one strains of Yersinia enterocolitica biovar 1A representing several serotypes isolated from India, France, Germany and the USA were analyzed using ribotyping, 16S-23S rDNA intergenic spacer length polymorphism analysis (PCR-ribotyping) and gyrB restriction fragment length polymorphism. Ribotyping with BglI, NciI and EcoRV distinguished 81 strains into 4, 3 and 2 ribotypes respectively. BglI-NciI combination gave the highest Simpson's diversity index (DI=0.43). Strains with identical ribotypes were further differentiated by PCR-ribotyping. The combination of BglI-NciI ribotyping with PCR ribotyping increased DI to 0.72. This suggested that the combination of the two may be used for molecular epidemiological studies of Y. enterocolitica biovar 1A. This approach clearly resolved the strains into two clonal groups, each comprising strains isolated from humans, swine, pork and wastewater. PCR-RFLP of the gyrB gene using three enzymes (AluI, MspI and HinfI) distinguished strains into seven types and confirmed the existence of two clonal groups. Thus, assessment of heterogeneity based on chromosomal restriction analysis (ribotyping), rRNA spacer length polymorphism (PCR-ribotyping) and gyrB gene analysis were in concordance and provided unequivocal evidence for the presence of two groups amongst strains of Y. enterocolitica biovar 1A despite their diverse geographic origins. These data also grouped clinical and non-clinical strains of serotype O:6,30-6,31 into discrete subgroups.
KeywordMeSH Terms
47. Pham  HN, Ohkusu  K, Mishima  N, Noda  M, Monir Shah  M, Sun  X, Hayashi  M, Ezaki  T,     ( 2007 )

Phylogeny and species identification of the family Enterobacteriaceae based on dnaJ sequences.

Diagnostic microbiology and infectious disease 58 (2)
PMID : 17368802  :   DOI  :   10.1016/j.diagmicrobio.2006.12.019    
Abstract >>
Phylogenetic relations within the family Enterobacteriaceae were analyzed using partial dnaJ sequences of 165 strains belonging to 93 species from 27 enterobacterial genera. The dnaJ phylogeny was in relative agreement with that constructed by 16S rDNA sequences, but more monophyletic groups were obtained from the dnaJ tree than from the 16S rDNA tree. The degree of divergence of the dnaJ gene was approximately 6 times greater than that of 16S rDNA. Also, the dnaJ gene showed the most discriminatory power in comparison with tuf and atpD genes, facilitating clear differentiation of any 2 enterobacterial species by dnaJ sequence analysis. The application of dnaJ sequences to the identification was confirmed by assigning 72 clinical isolates to the correct enterobacterial species. Our data indicate that analysis of the dnaJ gene sequences can be used as a powerful marker for phylogenetic study and identification at the species level of the family Enterobacteriaceae.
KeywordMeSH Terms
HSP40 Heat-Shock Proteins
Phylogeny
48. Abbott  DW, Boraston  AB,     ( 2007 )

The structural basis for exopolygalacturonase activity in a family 28 glycoside hydrolase.

Journal of molecular biology 368 (5)
PMID : 17397864  :   DOI  :   10.1016/j.jmb.2007.02.083    
Abstract >>
Family 28 glycoside hydrolases (polygalacturonases) are found in organisms across the plant, fungal and bacterial kingdoms, where they are central to diverse biological functions such as fruit ripening, biomass recycling and plant pathogenesis. The structures of several polygalacturonases have been reported; however, all of these enzymes utilize an endo-mode of digestion, which generates a spectrum of oligosaccharide products with varying degrees of polymerization. The structure of a complementary exo-acting polygalacturonase and an accompanying explanation of the molecular determinants for its specialized activity have been noticeably lacking. We present the structure of an exopolygalacturonase from Yersinia enterocolitica, YeGH28 in a native form (solved to 2.19 A resolution) and a digalacturonic acid product complex (solved to 2.10 A resolution). The activity of YeGH28 is due to inserted stretches of amino acid residues that transform the active site from the open-ended channel observed in the endopolygalacturonases to a closed pocket that restricts the enzyme to the exclusive attack of the non-reducing end of oligogalacturonide substrates. In addition, YeGH28 possesses a fused FN3 domain with unknown function, the first such structure described in pectin active enzymes.
KeywordMeSH Terms
Protein Structure, Tertiary
49. Bäumler  AJ, Hantke  K,     ( 1992 )

A lipoprotein of Yersinia enterocolitica facilitates ferrioxamine uptake in Escherichia coli.

Journal of bacteriology 174 (3)
PMID : 1732192  :   DOI  :   10.1128/jb.174.3.1029-1035.1992     PMC  :   PMC206184    
Abstract >>
A cloned fragment of Yersinia enterocolitica DNA complemented the defect in ferrioxamine B uptake of an Escherichia coli fhuE mutant lacking the outer membrane high-affinity transport protein FhuE. Subcloning revealed that a 13.7-kDa outer membrane protein was required for complementation. The amino acid sequence deduced from the nucleotide sequence showed extensive homology to PCPHi, an outer membrane lipoprotein of Haemophilus influenzae. We therefore termed this protein PCPYe. Plasmid-encoded pcpY mediated a low-affinity uptake of ferrioxamine B which may be caused by changes in the permeability of the outer membrane due to an overexpression of this outer membrane protein. A transposon insertion mutant in the plasmid-encoded pcpY gene was transferred into the chromosome of Y. enterocolitica. The resulting mutation had no effect on the high-affinity uptake of ferrioxamine B in Yersinia cells. Using the antibiotic ferrimycin we were able to isolate a Y. enterocolitica mutant lacking the high-affinity outer membrane receptor for ferrioxamine uptake, termed FoxA.
KeywordMeSH Terms
Escherichia coli Proteins
Receptors, Cell Surface
50. Bhagat  N, Virdi  JS,     ( 2007 )

Distribution of virulence-associated genes in Yersinia enterocolitica biovar 1A correlates with clonal groups and not the source of isolation.

FEMS microbiology letters 266 (2)
PMID : 17233728  :   DOI  :   10.1111/j.1574-6968.2006.00524.x    
Abstract >>
Yersinia enterocolitica, an important food- and water-borne enteric pathogen, is represented by six biovars viz. 1A, 1B and 2-5. Some biovar 1A strains, despite lacking virulence plasmid (pYV) and chromosomal virulence genes, have been reported to cause symptoms similar to that produced by isolates belonging to known pathogenic biovars. Virulence-associated genes viz. ail, virF, inv, myfA, ystA, ystB, ystC, tccC, hreP, fepA, fepD, fes, ymoA and sat were studied in 81 clinical and nonclinical strains of Y. enterocolitica biovar 1A by PCR amplification. All strains lacked ail, virF, ystA and ystC genes. The distribution of other genes with respect to clonal groups revealed that four genes viz. ystB, hreP, myfA and sat were associated exclusively with strains belonging to clonal group A. The clonal groups A and B were differentiated previously based on rep (REP-/ERIC) - PCR genomic fingerprinting. The distribution of virulence-associated genes, however, did not differ significantly between clinical and nonclinical strains. In strains of Y. enterocolitica biovar 1A, clonal groups seem to reflect virulence potential better than the source (clinical vs. nonclinical) of isolation.
KeywordMeSH Terms
51. Kosti?  T, Weilharter  A, Rubino  S, Delogu  G, Uzzau  S, Rudi  K, Sessitsch  A, Bodrossy  L,     ( 2007 )

A microbial diagnostic microarray technique for the sensitive detection and identification of pathogenic bacteria in a background of nonpathogens.

Analytical biochemistry 360 (2)
PMID : 17123456  :   DOI  :   10.1016/j.ab.2006.09.026    
Abstract >>
A major challenge in microbial diagnostics is the parallel detection and identification of low-bundance pathogens within a complex microbial community. In addition, a high specificity providing robust, reliable identification at least at the species level is required. A microbial diagnostic microarray approach, using single nucleotide extension labeling with gyrB as the marker gene, was developed. We present a novel concept applying competitive oligonucleotide probes to improve the specificity of the assay. Our approach enabled the sensitive and specific detection of a broad range of pathogenic bacteria. The approach was tested with a set of 35 oligonucleotide probes targeting Escherichia coli, Shigella spp., Salmonella spp., Aeromonas hydrophila, Vibrio cholerae, Mycobacterium avium, Mycobacterium tuberculosis, Helicobacter pylori, Proteus mirabilis, Yersinia enterocolitica, and Campylobacter jejuni. The introduction of competitive oligonucleotides in the labeling reaction successfully suppressed cross-reaction by closely related sequences, significantly improving the performance of the assay. Environmental applicability was tested with environmental and veterinary samples harboring complex microbial communities. Detection sensitivity in the range of 0.1% has been demonstrated, far below the 5% detection limit of traditional microbial diagnostic microarrays.
KeywordMeSH Terms
52. Mildiner-Earley  S, Miller  VL,     ( 2006 )

Characterization of a novel porin involved in systemic Yersinia enterocolitica infection.

Infection and immunity 74 (7)
PMID : 16790812  :   DOI  :   10.1128/IAI.00154-06     PMC  :   PMC1489722    
Abstract >>
Yersinia enterocolitica is an enteric pathogen capable of causing systemic disease in a murine model. We have identified a novel protein, systemic factor protein A (SfpA), conserved in other pathogenic bacteria that is involved in systemic disease. Analysis of bacterial colonization revealed that a DeltasfpA strain is defective in mesenteric lymph node colonization. Bioinformatics and functional studies suggest that SfpA is a porin.
KeywordMeSH Terms
53. Axler-Diperte  GL, Miller  VL, Darwin  AJ,     ( 2006 )

YtxR, a conserved LysR-like regulator that induces expression of genes encoding a putative ADP-ribosyltransferase toxin homologue in Yersinia enterocolitica.

Journal of bacteriology 188 (23)
PMID : 16997967  :   DOI  :   10.1128/JB.01159-06     PMC  :   PMC1698212    
Abstract >>
Yersinia enterocolitica causes human gastroenteritis, and many isolates have been classified as either "American" or "non-American" strains based on their geographic prevalence and virulence properties. In this study we describe identification of a transcriptional regulator that controls expression of the Y. enterocolitica ytxAB genes. The ytxAB genes have the potential to encode an ADP-ribosylating toxin with similarity to pertussis toxin. However, a ytxAB null mutation did not affect virulence in mice. Nevertheless, the ytxAB genes are conserved in many Y. enterocolitica strains. Interestingly, American and non-American strains have different ytxAB alleles encoding proteins that are only 50 to 60% identical. To obtain further insight into the ytxAB locus, we investigated whether it is regulated as part of a known or novel regulon. Transposon mutagenesis identified a LysR-like regulator, which we designated YtxR. Expression of ytxR from a nonnative promoter increased Phi(ytxA-lacZ) operon fusion expression up to 35-fold. YtxR also activated expression of its own promoter. DNase I footprinting showed that a His(6)-YtxR fusion protein directly interacted with the ytxA and ytxR control regions at similar distances upstream of their probable transcription initiation sites, identified by primer extension. Deletion analysis demonstrated that removal of the regions protected by His(6)-YtxR in vitro eliminated YtxR-dependent induction in vivo. The ytxAB locus is not present in most Yersinia species. In contrast, ytxR is conserved in multiple Yersinia species, as well as in the closely related organisms Photorhabdus luminescens and Photorhabdus asymbiotica. These observations suggest that YtxR may play a conserved role involving regulation of other genes besides ytxAB.
KeywordMeSH Terms
Genes, Bacterial
54. Delmas  J, Breysse  F, Devulder  G, Flandrois  JP, Chomarat  M,     ( 2006 )

Rapid identification of Enterobacteriaceae by sequencing DNA gyrase subunit B encoding gene.

Diagnostic microbiology and infectious disease 55 (4)
PMID : 16626902  :   DOI  :   10.1016/j.diagmicrobio.2006.02.003    
Abstract >>
Real-time polymerase chain reaction and sequencing were used to characterize a 506-bp-long DNA fragment internal to the gyrB gene (gyrBint). The sequences obtained from 32 Enterobacteriaceae-type strains and those available in the Genbank nucleotide sequence database (n = 24) were used as a database to identify 240 clinical enterobacteria isolates. Sequence analysis of the gyrBint fragment of 240 strains showed that gyrBint constitutes a discriminative target sequence to differentiate between Enterobacteriaceae species. Comparison of these identifications with those obtained by phenotypic methods (Vitek 1 system and/or Rapid ID 32E; bioM?rieux, Marcy l'Etoile, France) revealed discrepancies essentially with genera Citrobacter and Enterobacter. Most of the strains identified as Enterobacter cloacae by phenotypic methods were identified as Enterobacter hormaechei strains by gyrBint sequencing. The direct sequencing of gyrBint would be useful as a complementary tool in the identification of clinical Enterobacteriaceae isolates.
KeywordMeSH Terms
55. Miller  VL, Bliska  JB, Falkow  S,     ( 1990 )

Nucleotide sequence of the Yersinia enterocolitica ail gene and characterization of the Ail protein product.

Journal of bacteriology 172 (2)
PMID : 1688838  :   DOI  :   10.1128/jb.172.2.1062-1069.1990     PMC  :   PMC208537    
Abstract >>
The ability to enter (invade) eucaryotic cells is a property common to many pathogenic bacteria. Yersinia enterocolitica is a facultative intracellular pathogen whose primary site of multiplication is the reticuloendothelial system. In an effort to understand how Y. enterocolitica crosses the intestinal epithelial cell layer, we previously reported the cloning of two loci from Y. enterocolitica that individually conferred an invasive phenotype to the normally noninvasive Escherichia coli HB101. One of these loci, ail, is encoded by a region of DNA that is less than 650 base pairs. We have identified the ail gene product in maxicells as a 17-kilodalton membrane-associated protein. The Ail protein has been purified, and its N-terminal sequence has been determined. The nucleotide sequence of the ail gene revealed a single unique open reading frame of 178 amino acids. Comparison of amino acid sequences deduced from the gene and obtained by analysis of the purified protein identified the first 23 amino acids as a signal sequence. The site(s) at which transcription initiates on the ail gene was identified by primer extension analysis and shown to be identical in E. coli and Y. enterocolitica. Two small open reading frames downstream of ail were found and shown to exhibit considerable identity to the proposed IS3 transposase.
KeywordMeSH Terms
Genes, Bacterial
56. Sharma  S, Mittal  S, Mallik  S, Virdi  JS,     ( 2006 )

Molecular characterization of beta-lactamase genes blaA and blaB of Yersinia enterocolitica biovar 1A.

FEMS microbiology letters 257 (2)
PMID : 16553870  :   DOI  :   10.1111/j.1574-6968.2006.00191.x    
Abstract >>
The beta-lactamase genes blaA and blaB were detected by PCR amplification in strains of Yersinia enterocolitica biovar 1A isolated from India, Germany, France and the USA. Both genes were detected in all strains. Polymerase chain reaction-restriction fragment length polymorphism revealed genetic heterogeneity in blaA but not in blaB. Cluster analysis of blaA restriction profiles grouped the strains into three groups. The blaA gene of Y. enterocolitica biovar 1A showed a high degree of sequence homology to that of Y. enterocolitica 8081 (biovar 1B) and Y. enterocolitica Y-56 (biovar 4), whereas homology was low with class A beta-lactamase genes of other members of the family Enterobacteriaceae. The pI 8.7 of enzyme Bla-A of Y. enterocolitica biovar 1A was similar to that of biovars 2, 3 and 4. The enzyme Bla-B focused at 6.8 and 7.1, indicating that biovar 1A strains produced a 'B-like' enzyme. This is the first study to have investigated the genetic heterogeneity of the beta-lactamase genes of Y. enterocolitica.
KeywordMeSH Terms
57. Bäumler  AJ, Hantke  K,     ( 1992 )

Ferrioxamine uptake in Yersinia enterocolitica: characterization of the receptor protein FoxA.

Molecular microbiology 6 (10)
PMID : 1640832  :   DOI  :   10.1111/j.1365-2958.1992.tb00852.x    
Abstract >>
The gene for the high-affinity outer membrane ferrioxamine receptor FoxA of Yersinia enterocolitica was cloned in Escherichia coli K-12. A foxA mutant of Yersinia could be complemented by the cloned DNA fragment. The FoxA encoding region was sequenced and an open reading frame encoding 710 amino acids, including a signal sequence of 26 amino acids, was deduced. The mature FoxA protein consisted of 684 amino acids and had a molecular mass of 75,768 Da. FoxA shared 33% amino acid sequence homology with FhuA, the ferrichrome receptor of Escherichia coli. Based on the homologies with FhuA and other TonB-dependent receptors a topological model of FoxA is presented.
KeywordMeSH Terms
Escherichia coli Proteins
Genes, Bacterial
Receptors, Cell Surface
58. Kerbarh  O, Chirgadze  DY, Blundell  TL, Abell  C,     ( 2006 )

Crystal structures of Yersinia enterocolitica salicylate synthase and its complex with the reaction products salicylate and pyruvate.

Journal of molecular biology 357 (2)
PMID : 16434053  :   DOI  :   10.1016/j.jmb.2005.12.078    
Abstract >>
The salicylate synthase, Irp9, from Yersinia enterocolitica is involved in the biosynthesis of the siderophore yersiniabactin. It is a bifunctional enzyme that forms salicylate and pyruvate from chorismate and water via the intermediate isochorismate. Here we report the first crystal structure of Irp9 and also of its complex with the reaction products salicylate and pyruvate at 1.85 A and 2.1 A resolution, respectively. Like other members of the chorismate-utilizing enzyme family, e.g. the TrpE subunit of anthranilate synthase and the PabB subunit of 4-amino-4-deoxychorismate synthase, Irp9 has a complex alpha/beta fold. The crystal structure of Irp9 contains one molecule each of phosphate and acetate derived from the crystallization buffer. The Irp9-products complex structure was obtained by soaking chorismate into Irp9, demonstrating that the enzyme is still catalytically active in the crystal. Both structures contain Mg(2+) in the active site. There is no evidence of the allosteric tryptophan binding site found in TrpE and PabB. Mutagenesis of Glu240, His321 and Tyr372 provided some insight into the mechanism of the two transformations catalyzed by Irp9. Knowledge of the structure of Irp9 will guide the search for potent inhibitors of salicylate formation, and hence of bacterial iron uptake, which is directly related to the virulence of Yersinia.
KeywordMeSH Terms
Protein Structure, Quaternary
59. Kotetishvili  M, Kreger  A, Wauters  G, Morris  JG, Sulakvelidze  A, Stine  OC,     ( 2005 )

Multilocus sequence typing for studying genetic relationships among Yersinia species.

Journal of clinical microbiology 43 (6)
PMID : 15956383  :   DOI  :   10.1128/JCM.43.6.2674-2684.2005     PMC  :   PMC1151872    
Abstract >>
The intra- and interspecies genetic relationships of 58 strains representing all currently known species of the genus Yersinia were examined by multilocus sequence typing (MLST), using sequence data from 16S RNA, glnA, gyrB, recA, and Y-HSP60 loci. Yersinia aldovae, Y. bercovieri, Y. intermedia, Y. pestis, Y. pseudotuberculosis, Y. rohdei, and Y. ruckeri were genetically more homogeneous than were Y. enterocolitica, Y. frederiksenii, Y. kristensenii, and Y. mollaretii. The MLST data concerning the genetic relatedness within and among various species of Yersinia support the idea that Y. pestis and Y. pseudotuberculosis are two lineages within the same species rather than two distinct species. Y. ruckeri is the genetically most distant species within the genus. There was evidence of O-antigen switching and genetic recombination within and among various species of Yersinia. The genetic relatedness data obtained by MLST of the four housekeeping genes and 16S RNA agreed in most, but not all, instances. MLST was better suited for determining genetic relatedness among yersiniae than was 16S RNA analysis. Some strains of Y. frederiksenii and Y. kristensenii are genetically less related to other strains within those species, compared to strains of all other species within the genus. The taxonomic standing of these strains should be further examined because they may represent currently unrecognized Yersinia species.
KeywordMeSH Terms
Sequence Analysis, DNA
60. Sing  A, Reithmeier-Rost  D, Granfors  K, Hill  J, Roggenkamp  A, Heesemann  J,     ( 2005 )

A hypervariable N-terminal region of Yersinia LcrV determines Toll-like receptor 2-mediated IL-10 induction and mouse virulence.

Proceedings of the National Academy of Sciences of the United States of America 102 (44)
PMID : 16239347  :   DOI  :   10.1073/pnas.0504728102     PMC  :   PMC1276055    
Abstract >>
The virulence antigen LcrV of Yersinia enterocolitica O:8 induces IL-10 in macrophages via Toll-like receptor 2 (TLR2). The TLR2-active region of LcrV is localized within its N-terminal amino acids (aa) 31-57. Sequencing of codons 25-92 of the lcrV gene from 59 strains of the three pathogenic Yersinia species revealed a hypervariable hotspot within aa 40-61. According to these sequence differences, seven LcrV groups were identified, with Y. pestis and Y. pseudotuberculosis represented in group I and the other six distributed within Y. enterocolitica. By testing LcrV sequence-derived synthetic oligopeptides of all seven LcrV groups in CD14/TLR2-transfected human embryonic kidney 293 cells, we found the highest TLR2 activity with a peptide derived from group IV comprising exclusively Y. enterocolitica O:8 strains. These findings were verified in murine peritoneal macrophages by using recombinant LcrV truncates representing aa 1-130 from different Yersinia spp. By systematically replacing charged aa residues by glutamine in synthetic oligopeptides, we show that the K42Q substitution leads to abrogation of TLR2 activity in both in vitro cell systems. This K42Q substitution was introduced in the lcrV gene from Y. enterocolitica O:8 WA-C(pYV), resulting in WA-C(pYVLcrV(K42Q)), which turned out to be less virulent for C57BL/6 mice than the parental strain. This difference in virulence was not observed in TLR2(-/-) or IL-10(-/-) mice, proving that LcrV contributes to virulence by TLR2-mediated IL-10 induction. LcrV is a defined bacterial virulence factor shown to target the TLR system for evasion of the host's immune response.
KeywordMeSH Terms
61. Tennant  SM, Skinner  NA, Joe  A, Robins-Browne  RM,     ( 2005 )

Homologues of insecticidal toxin complex genes in Yersinia enterocolitica biotype 1A and their contribution to virulence.

Infection and immunity 73 (10)
PMID : 16177365  :   DOI  :   10.1128/IAI.73.10.6860-6867.2005     PMC  :   PMC1230942    
Abstract >>
Yersinia enterocolitica is an enteric pathogen that consists of six biotypes: 1A, 1B, 2, 3, 4, and 5. Strains of the latter five biotypes can carry a virulence plasmid, known as pYV, and several well-characterized chromosomally encoded virulence determinants. Y. enterocolitica strains of biotype 1A lack the virulence-associated markers of pYV-bearing strains and were once considered to be avirulent. There is growing epidemiological, clinical, and experimental evidence, however, to suggest that some biotype 1A strains are virulent and can cause gastrointestinal disease. To identify potential virulence genes of pathogenic strains of Y. enterocolitica biotype 1A, we used genomic subtractive hybridization to determine genetic differences between two biotype 1A strains: an environmental isolate, Y. enterocolitica IP2222, and a clinical isolate, Y. enterocolitica T83. Among the Y. enterocolitica T83-specific genes we identified were three, tcbA, tcaC, and tccC, that showed homology to the insecticidal toxin complex (TC) genes first discovered in Photorhabdus luminescens. The Y. enterocolitica T83 TC gene homologues were expressed by Y. enterocolitica T83 and were significantly more prevalent among clinical biotype 1A strains than other Yersinia isolates. Inactivation of the TC genes in Y. enterocolitica T83 resulted in mutants which were attenuated in the ability to colonize the gastrointestinal tracts of perorally infected mice. These results indicate that products of the TC gene complex contribute to the virulence of some strains of Y. enterocolitica biotype 1A, possibly by facilitating their persistence in vivo.
KeywordMeSH Terms
Genes, Bacterial
62. Golubov  A, Gierczynski  R, Heesemann  J, Rakin  A,     ( 2005 )

A novel insertion sequence element, IS Yen2, as an epidemiological marker for weakly pathogenic bioserotypes of Yersinia enterocolitica.

International journal of medical microbiology : IJMM 295 (4)
PMID : 16128396  :   DOI  :   10.1016/j.ijmm.2005.05.007    
Abstract >>
The structure and distribution of IS Yen2, a new Yersinia enterocolitica insertion sequence element, were investigated. ISYen2 is related to IS elements of the IS21 family and is present in two isoforms in Y. enterocolitica serotype O:3. Analysis of all copies of both isoforms, IS Yen2A and IS Yen2B, by PCR and sequencing indicated that they are not flanked by direct repeats which are typical of the IS21 family of repetitive elements. IS Yen2 is present in multiple copies in Y. enterocolitica O:3 and O:9 and in a single copy in Y. enterocolitica O:1 and O:2 serotypes. The probe for IS Yen2 efficiently detected all weakly pathogenic Y. enterocolitica bioserotypes investigated, whereas it did not hybridize with other strains. This indicates that IS Yen2 can serve as an additional tool for Y. enterocolitica differentiation and epidemiological studies. Distribution of the different groups of IS elements in two Y. enterocolitica pathotypes is in favor of the parallel evolution of American and European Y. enterocolitica strains.
KeywordMeSH Terms
Mutagenesis, Insertional
63. Cocolin  L, Comi  G,     ( 2005 )

Use of a culture-independent molecular method to study the ecology of Yersinia spp. in food.

International journal of food microbiology 105 (1)
PMID : 16085330  :   DOI  :   10.1016/j.ijfoodmicro.2005.05.006    
Abstract >>
A culture-independent method for the direct detection in food of Yersinia spp. was developed in this study. It is based on the amplification of a 359 bp PCR product from the RNA polymerase beta-subunit gene (rpoB) and subsequent analysis by denaturing gradient gel electrophoresis (DGGE). Direct detection of Yersinia spp. by PCR-DGGE was carried out in ready-to-eat vegetables and the results compared with the results of the traditional, culture-dependent method. The DGGE profiles were determined to be species-specific. As a matter of fact, Yersinia enterocolitica, Yersinia intermedia, Yersinia frederiskenii and Yersinia kristensenii showed differential migrations in the gels. Moreover, Y. enterocolitica serotypes O:3, O:5 and O:9 were distinguishable, as well. Only for a limited number of traditionally isolated strains, the biochemical and molecular identification agree. In particular, an overestimation of Y. enterocolitica, as determined biochemically, was observed. Finally when the protocol was applied to 27 food samples, a good correlation was obtained when the results of traditional and direct methods were analyzed. The molecular method was able to identify Y. enterocolitica, not detected by plating analysis. However, for 4 samples, that, by plating analysis, were determined to contain Yersinia spp., no PCR product could be obtained after enrichment, probably due to low numbers of target cells, thereby not allowing the possibility to perform DGGE analysis. The protocol described here represents a reliable tool for the detection of Yersinia spp. in food, which can be used to obtain the needed results faster than with traditional culturing methods.
KeywordMeSH Terms
64. Demarta  A, De Respinis  S, Dolina  M, Peduzzi  R,     ( 2004 )

Molecular typing of Yersinia frederiksenii strains by means of 16s rDNA and gyrB genes sequence analyses.

FEMS microbiology letters 238 (2)
PMID : 15358429  :   DOI  :   10.1016/j.femsle.2004.08.005    
Abstract >>
The phenospecies Yersinia frederiksenii is known to comprise three genospecies, indistinguishable on the basis of phenotypic characteristics. In previous works, 13 strains, identified biochemically as Y. frederiksenii, were characterized using Multilocus enzyme electrophoresis and Ribotyping. In order to elucidate the phylogenetic position of these strains we performed their molecular typing by means of 16S rDNA and gyrB sequences analyses. Results demonstrated that gyrB is a better phylogenetic marker than 16S rDNA. The classification achieved by gyrB sequences analyses was in agreement with results obtained with more laborious methods. Moreover, a good phylogenetic identification could be reached also with partial gyrB sequences of only 350 bp.
KeywordMeSH Terms
Bacterial Typing Techniques
65. Skurnik  M, Toivanen  P,     ( 1992 )

LcrF is the temperature-regulated activator of the yadA gene of Yersinia enterocolitica and Yersinia pseudotuberculosis.

Journal of bacteriology 174 (6)
PMID : 1548243  :   DOI  :   10.1128/jb.174.6.2047-2051.1992     PMC  :   PMC205814    
Abstract >>
The virulence plasmid of human pathogenic Yersinia species, pYV, encodes secreted proteins, Yop proteins, and an outer membrane protein, YadA. YadA has been associated with binding to a variety of substrates and with interference with host defense. YadA is regulated by temperature and is expressed only at 37 degrees C. Unlike the yop regulon, the yadA gene is not under Ca2+ regulation. Here, we show that LcrF (VirF), the temperature-regulated activator of the yop regulon, also acts as an activator for yadA.
KeywordMeSH Terms
Gene Expression Regulation, Bacterial
66. Phan  J, Tropea  JE, Waugh  DS,     ( 2004 )

Structure of the Yersinia pestis type III secretion chaperone SycH in complex with a stable fragment of YscM2.

Acta crystallographica. Section D, Biological crystallography 60 (Pt 9)
PMID : 15333930  :   DOI  :   10.1107/S0907444904017597    
Abstract >>
Pathogenic Yersinia species use a type III secretion system to inject cytotoxic effector proteins directly into the cytosol of mammalian cells, where they neutralize the innate immune response by interfering with the signal-transduction pathways that control phagocytosis and inflammation. To be exported efficiently, some effectors must transiently associate with cognate cytoplasmic secretion chaperones. SycH is the chaperone for YopH, a potent eukaryotic-like protein tyrosine phosphatase that is essential for virulence. SycH also binds two negative regulators of type III secretion, YscM1 and YscM2, both of which share significant sequence homology with the chaperone-binding domain of YopH. Here, the structure of a complex between SycH and a stable fragment of YscM2 that was designed on the basis of limited proteolysis experiments is presented. The overall fold of SycH is very similar to the structures of other homodimeric secretion chaperones that have been determined to date. YscM2 wraps around SycH in an extended fashion, with some secondary but no tertiary structure, assuming a conformation distinct from the globular fold that it is predicted to adopt in the absence of SycH.
KeywordMeSH Terms
67. Seoane  A, Francia  MV, García Lobo  JM,     ( 1992 )

Nucleotide sequence of the ampC-ampR region from the chromosome of Yersinia enterocolitica.

Antimicrobial agents and chemotherapy 36 (5)
PMID : 1510392  :   DOI  :   10.1128/aac.36.5.1049     PMC  :   PMC188833    
Abstract >>
The nucleotide sequence of a 3.1-kb region from the chromosome of the Yersinia enterocolitica O:5b strain IP97 containing the gene for an inducible chromosomal cephalosporinase has been determined. The cephalosporinase gene was homologous to other enterobacterial chromosomal cephalosporinase genes, and it was accompanied by a gene homologous to the regulatory ampR gene. The arrangement of genes in the Y. enterocolitica ampCR unit was identical to that in the Enterobacter cloacae and Citrobacter freundii ampCR units.
KeywordMeSH Terms
Base Sequence
Chromosomes, Bacterial
Genes, Bacterial
68. Nummelin  H, Merckel  MC, Leo  JC, Lankinen  H, Skurnik  M, Goldman  A,     ( 2004 )

The Yersinia adhesin YadA collagen-binding domain structure is a novel left-handed parallel beta-roll.

The EMBO journal 23 (4)
PMID : 14765110  :   DOI  :   10.1038/sj.emboj.7600100     PMC  :   PMC381008    
Abstract >>
The crystal structure of the recombinant collagen-binding domain of Yersinia adhesin YadA from Yersinia enterocolitica serotype O:3 was solved at 1.55 A resolution. The trimeric structure is composed of head and neck regions, and the collagen binding head region is a novel nine-coiled left-handed parallel beta-roll. Before the beta-roll, the polypeptide loops from one monomer to the rest, and after the beta-roll the neck region does the same, making the transition from the globular head region to the narrower stalk domain. This creates an intrinsically stable 'lock nut' structure. The trimeric form of YadA is required for collagen binding, and mutagenesis of its surface residues allowed identification of a putative collagen-binding surface. Furthermore, a new structure-sequence motif for YadA beta-roll was used to identify putative YadA-head-like domains in a variety of human and plant pathogens. Such domains may therefore be a common bacterial strategy for avoiding host response.
KeywordMeSH Terms
69. de la Cruz  F, Carmona  M, Juaréz  A,     ( 1992 )

The Hha protein from Escherichia coli is highly homologous to the YmoA protein from Yersinia enterocolitica.

Molecular microbiology 6 (22)
PMID : 1484495  :   DOI  :   10.1111/j.1365-2958.1992.tb02214.x    
Abstract >>
N/A
KeywordMeSH Terms
DNA-Binding Proteins
Escherichia coli Proteins
70. Wren  BW, Colby  SM, Cubberley  RR, Pallen  MJ,     ( 1992 )

Degenerate PCR primers for the amplification of fragments from genes encoding response regulators from a range of pathogenic bacteria.

FEMS microbiology letters 78 (2��3��)
PMID : 1490612  :   DOI  :   10.1016/0378-1097(92)90042-m    
Abstract >>
Many bacterial responses to environmental stimuli are mediated by response regulators which coordinately regulate genes involved in particular adaptive responses. Degenerate oligonucleotide primers were used to amplify by the polymerase chain reaction (PCR), fragments from genes encoding eleven novel response regulators. Sequence and phylogenetic analysis revealed that phoB, phoP and creB gene fragments had been amplified from Yersinia enterocolitica and Yersinia pseudotuberculosis, and that a creB sequence had been amplified from Campylobacter jejuni. Four amplified fragments from C. jejuni, Listeria monocytogenes, Mycobacterium tuberculosis and Escherichia coli clearly came from response regulator genes, but were not closely related to any of the known genes. Mutagenesis of the newly identified genes should allow us to determine their function and the genes under their control.
KeywordMeSH Terms
Genes, Bacterial
Polymerase Chain Reaction
71. Ellison  DW, Young  B, Nelson  K, Miller  VL,     ( 2003 )

YmoA negatively regulates expression of invasin from Yersinia enterocolitica.

Journal of bacteriology 185 (24)
PMID : 14645275  :   DOI  :   10.1128/jb.185.24.7153-7159.2003     PMC  :   PMC296258    
Abstract >>
inv encodes invasin, which is the primary invasion factor of Yersinia enterocolitica. inv expression in vitro is regulated in response to temperature, pH, and growth phase. In vitro, inv is maximally expressed at 26 degrees C and repressed at 37 degrees C at neutral pH but, when the pH of the media is adjusted to 5.5, levels of inv expression at 37 degrees C are comparable to those at 26 degrees C. A previous genetic screen for regulators of inv identified RovA, which was found to be required for activation of inv in vitro under all conditions tested as well as in vivo. Here we describe a screen that has identified a negative regulator of inv expression, ymoA. The ymoBA locus was identified by transposon mutagenesis as a repressor of inv expression in vitro at 37 degrees C at neutral pH. This mutant shows increased inv expression at 37 degrees C. The mutant can be fully complemented for inv expression by a plasmid expressing ymoA. These results indicate that YmoA plays a role in the negative regulation of inv.
KeywordMeSH Terms
72. Kraushaar  B, Dieckmann  R, Wittwer  M, Knabner  D, Konietzny  A, Mäde  D, Strauch  E,     ( 2011 )

Characterization of a Yersinia enterocolitica biotype 1A strain harbouring an ail gene.

Journal of applied microbiology 111 (4)
PMID : 21794036  :   DOI  :   10.1111/j.1365-2672.2011.05112.x    
Abstract >>
The chromosomal ail gene (attachment and invasion locus) is commonly used as target gene for the detection of pathogenic Y. enterocolitica strains in food testing. The ail PCR does not detect strains of biotype 1A (BT1A), which are regarded as non-pathogenic because BT1A strains lack the virulence plasmid and chromosomally encoded virulence genes. In some recent reports, however, BT1A strains were discovered that harboured the ail gene. We isolated an ail-positive strain and characterized this strain with phenotypic and genotypic methods to study its possible relation to pathogenic Y. enterocolitica strains. The ail region of the BT1A strain was sequenced and compared with the corresponding region of nonpathogenic BT1A strains and pathogenic strains. Pulsed field gel electrophoresis (PFGE) analysis was applied revealing no similarity of the PFGE pattern of this strain to the patterns of pathogenic strains. Virulence-gene-based PCR analyses showed the strain to be positive for ystB, but negative for virulence genes ystA, virF and yadA. Whole-cell MALDI-TOF MS combined with a shrinkage discriminant analysis approach was applied and clearly classified the ail-positive biotype 1A strain within the cluster of BT1A strains. PCR detection of ail sequences in food matrices should be followed by the isolation of the responsible strain and its characterization using phenotypic or genotypic methods. The ail gene may be present in Y. enterocolitica BT1A strains, which are commonly considered as nonpathogenic. Efficient methods such as PCR typing of other virulence genes or rapid MALDI-TOF MS-based bacterial profiling allow a more comprehensive assessment of the pathogenicity potential of Yersinia strains.
KeywordMeSH Terms
73. Fu  D, Li  Z, Huang  H, Yuan  T, Shi  P, Luo  H, Meng  K, Yang  P, Yao  B,     ( 2011 )

Catalytic efficiency of HAP phytases is determined by a key residue in close proximity to the active site.

Applied microbiology and biotechnology 90 (4)
PMID : 21380516  :   DOI  :   10.1007/s00253-011-3171-0    
Abstract >>
The maximum activity of Yersinia enterocolitica phytase (YeAPPA) occurs at pH 5.0 and 45 �XC, and notably, its specific activity (3.28 �� 0.24 U mg(-1)) is 800-fold less than that of its Yersinia kristeensenii homolog (YkAPPA; 88% amino acid sequence identity). Sequence alignment and molecular modeling show that the arginine at position 79 (Arg79) in YeAPPA corresponding to Gly in YkAPPA as well as other histidine acid phosphatase (HAP) phytases is the only non-conserved residue near the catalytic site. To characterize the effects of the corresponding residue on the specific activities of HAP phytases, Escherichia coli EcAPPA, a well-characterized phytase with a known crystal structure, was selected for mutagenesis-its Gly73 was replaced with Arg, Asp, Glu, Ser, Thr, Leu, or Tyr. The results show that the specific activities of all of the corresponding EcAPPA mutants (17-2,400 U mg(-1)) were less than that of the wild-type phytase (3,524 U mg(-1)), and the activity levels were approximately proportional to the molecular volumes of the substituted residues' side chains. Site-directed replacement of Arg79 in YeAPPA (corresponding to Gly73 of EcAPPA) with Ser, Leu, and Gly largely increased the specific activity, which further verified the key role of the residue at position 79 for determining phytase activity. Thus, a new determinant that influences the catalytic efficiency of HAP phytases has been identified.
KeywordMeSH Terms
74. Michiels  T, Wattiau  P, Brasseur  R, Ruysschaert  JM, Cornelis  G,     ( 1990 )

Secretion of Yop proteins by Yersiniae.

Infection and immunity 58 (9)
PMID : 2129533  :   PMC  :   PMC313576    
Abstract >>
Upon incubation at 37 degrees C in the absence of Ca2+ ions, pathogenic strains of the genus Yersinia cease growing and produce large amounts of a series of plasmid-encoded proteins involved in pathogenicity. These proteins, called Yops (for Yersinia outer membrane proteins), are detected in both the outer membrane fraction and the culture supernatant. We present here the nucleotide sequence of genes yop20 and yop25 from Yersinia enterocolitica O:9. Protein Yop25 is very similar to YopE, the corresponding protein from Yersinia pestis, Y. pseudotuberculosis, and Y. enterocolitica O:8 (A. Forsberg and H. Wolf-Watz, J. Bacteriol. 172:1547-1555, 1990). This is the first report of a yop20 sequence of yersiniae. We present evidences that Yops are not membrane proteins. Their detection in the membrane fraction results either from copurification of large aggregates of extracellular Yops with the membrane fraction or from the adsorption of released proteins to the cell surface. In contrast with Yops, protein P1 has characteristics of a true membrane protein. The release of Yops by Y. enterocolitica occurs by a novel secretion mechanism that does not involve the cleavage of a typical signal sequence or the recognition of a carboxy-terminal domain.
KeywordMeSH Terms
75. Mallik  S, Virdi  JS,     ( 2010 )

Genetic relationships between clinical and non-clinical strains of Yersinia enterocolitica biovar 1A as revealed by multilocus enzyme electrophoresis and multilocus restriction typing.

BMC microbiology 10 (N/A)
PMID : 20509911  :   DOI  :   10.1186/1471-2180-10-158     PMC  :   PMC2889952    
Abstract >>
Genetic relationships among 81 strains of Y. enterocolitica biovar 1A isolated from clinical and non-clinical sources were discerned by multilocus enzyme electrophoresis (MLEE) and multilocus restriction typing (MLRT) using six loci each. Such studies may reveal associations between the genotypes of the strains and their sources of isolation. All loci were polymorphic and generated 62 electrophoretic types (ETs) and 12 restriction types (RTs). The mean genetic diversity (H) of the strains by MLEE and MLRT was 0.566 and 0.441 respectively. MLEE (DI = 0.98) was more discriminatory and clustered Y. enterocolitica biovar 1A strains into four groups, while MLRT (DI = 0.77) identified two distinct groups. BURST (Based Upon Related Sequence Types) analysis of the MLRT data suggested aquatic serotype O:6,30-6,31 isolates to be the ancestral strains from which, clinical O:6,30-6,31 strains might have originated by host adaptation and genetic change. MLEE revealed greater genetic diversity among strains of Y. enterocolitica biovar 1A and clustered strains in four groups, while MLRT grouped the strains into two groups. BURST analysis of MLRT data nevertheless provided newer insights into the probable evolution of clinical strains from aquatic strains.
KeywordMeSH Terms
Bacterial Typing Techniques
Polymorphism, Restriction Fragment Length
76. Souza  RA, Pitondo-Silva  A, Falcão  DP, Falcão  JP,     ( 2010 )

Evaluation of four molecular typing methodologies as tools for determining taxonomy relations and for identifying species among Yersinia isolates.

Journal of microbiological methods 82 (2)
PMID : 20493215  :   DOI  :   10.1016/j.mimet.2010.05.005    
Abstract >>
In the last few decades, molecular typing has become an important tool in taxonomic, phylogenetic and identification studies of numerous groups of bacteria, including the yersiniae. In this study, Enterobacterial Repetitive Intergenic Consensus PCR (ERIC-PCR), Pulsed-Field Gel Electrophoresis (PFGE), 16S rRNA gene sequencing and Multilocus Sequence Analysis (MLSA) were performed to determine the ability of these techniques to be used in taxonomy and identification of Yersinia strains. A total of 60 Yersinia strains were genotyped by ERIC-PCR and PFGE. Moreover, an in silico analysis was carried out for 16S rRNA gene sequencing and MLSA, using 68 and 49 Yersinia strains, respectively. A phylogenetic tree constructed from the ERIC-PCR, 16S rRNA gene sequencing and MLSA data grouped most of the Yersinia species into distinct species-specific clusters. In the PFGE assay these clusters were not observed. On this basis, ERIC-PCR, 16S rRNA gene sequencing and MLSA seem to be valuable techniques for use in taxonomic and identification studies of the genus Yersinia, whereas PFGE does not. Furthermore, ERIC-PCR has the advantage of being a cheaper, easier and faster assay than 16S rRNA gene sequencing or MLSA, and for these reasons can be considerate an alternative tool in taxonomic studies of yersiniae.
KeywordMeSH Terms
77. Huang  Y, Wang  X, Cui  Z, Yang  Y, Xiao  Y, Tang  L, Kan  B, Xu  J, Jing  H,     ( 2010 )

Possible use of ail and foxA polymorphisms for detecting pathogenic Yersinia enterocolitica.

BMC microbiology 10 (N/A)
PMID : 20691098  :   DOI  :   10.1186/1471-2180-10-211     PMC  :   PMC2924855    
Abstract >>
Yersinia enterocolitica is an enteric pathogen that invades the intestinal mucosa and proliferates within the lymphoid follicles (Peyer's patches). The attachment invasion locus (ail) mediates invasion by Y. enterocolitica and confers an invasive phenotype upon non-invasive E. coli; ail is the primary virulence factor of Y. enterocolitica. The ferrioxamine receptor (foxA) located on the Y. enterocolitica chromosome, together with its transport protein, transports a siderophore specific for ferric ion. Currently, ail is the primary target gene for nucleic acid detection of pathogenic Y. enterocolitica. The genes ail and foxA in 271 pathogenic and 27 non-pathogenic Y. enterocolitica strains isolated from China and 10 reference strains were sequenced, aligned, compared to the ail and foxA sequences of Yersinia enterocolitica subsp. enterocolitica 8081 (Genbank: NC_008800), and analyzed for sequence polymorphism. The ail from the 282 strains showed 3 sequence patterns: 277 strains of serotypes O:3, O:9 and O:5, 27 with identical nucleic acid sequences formed pattern A1; 4 strains of serotype 1B/O:8 with identical nucleic acid sequences formed pattern A2; and one Chinese isolate 2/O:9 formed pattern A3. In the primary coding region of the foxA ORF (Genebank: X60447 nt 433-1866; nt 28 to 1,461 in the ORF), the sequences formed 3 groups and were further divided into 8 sequence patterns. The ail and foxA loci of pathogenic Y. enterocolitica have been analyzed. The ail sequence was highly conserved among the same serotype strains from different sources; and foxA was highly conserved among the pathogenic strains, although there was some sequence diversity. Fewer strains were used from outside China, which is a limitation of the study.
KeywordMeSH Terms
Polymorphism, Genetic
78. Bhagat  N, Virdi  JS,     ( 2009 )

Molecular and biochemical characterization of urease and survival of Yersinia enterocolitica biovar 1A in acidic pH in vitro.

BMC microbiology 9 (N/A)
PMID : 20017936  :   DOI  :   10.1186/1471-2180-9-262     PMC  :   PMC2806259    
Abstract >>
Yersinia enterocolitica, an important food- and water-borne enteric pathogen is represented by six biovars viz. 1A, 1B, 2, 3, 4 and 5. Despite the lack of recognized virulence determinants, some biovar 1A strains have been reported to produce disease symptoms resembling that produced by known pathogenic biovars (1B, 2-5). It is therefore imperative to identify determinants that might contribute to the pathogenicity of Y. enterocolitica biovar 1A strains. Y. enterocolitica invariably produces urease and the role of this enzyme in the virulence of biovar 1B and biovar 4 strains has been reported recently. The objective of this work was to study genetic organization of the urease (ure) gene complex of Y. enterocolitica biovar 1A, biochemical characterization of the urease, and the survival of these strains under acidic conditions in vitro. The ure gene complex (ureABCEFGD) of Y. enterocolitica biovar 1A included three structural and four accessory genes, which were contiguous and was flanked by a urea transport (yut) gene on the 3' side. Differences were identified in ure gene complex of biovar 1A strain compared to biovar 1B and 4 strains. This included a smaller ureB gene and larger intergenic regions between the structural genes. The crude urease preparation exhibited optimal pH and temperature of 5.5 and 65 degrees C respectively, and Michaelis-Menten kinetics with a Km of 1.7 +/- 0.4 mM urea and Vmax of 7.29 +/- 0.42 micromol of ammonia released/min/mg protein. The urease activity was dependent on growth temperature and growth phase of Y. enterocolitica biovar 1A, and the presence of nickel in the medium. The molecular mass of the enzyme was > 545 kDa and an isoelectric point of 5.2. The number of viable Y. enterocolitica biovar 1A decreased significantly when incubated at pH 2.5 for 2 h. However, no such decrease was observed at this pH in the presence of urea. The ure gene cluster of biovar 1A strains though similar to biovar 1B and 4 strains, exhibited important differences. The study also showed the ability of biovar 1A strains of Y. enterocolitica to survive at highly acidic pH in vitro in the presence of urea.
KeywordMeSH Terms
Multigene Family
79. Hirvas  L, Koski  P, Vaara  M,     ( 1991 )

The ompH gene of Yersinia enterocolitica: cloning, sequencing, expression, and comparison with known enterobacterial ompH sequences.

Journal of bacteriology 173 (3)
PMID : 1991717  :   DOI  :   10.1128/jb.173.3.1223-1229.1991     PMC  :   PMC207246    
Abstract >>
We have recently described a previously uncharacterized outer membrane protein of Salmonella typhimurium and Escherichia coli and cloned and sequenced the corresponding gene, the ompH gene, of S. typhimurium (P. Koski, M. Rhen, J. Kantele, and M. Vaara, J. Biol. Chem. 264:18973-18980, 1989). We report here the cloning, sequencing, and expression of the corresponding gene of Yersinia enterocolitica. It is significantly homologous to the ompH genes of E. coli and S. typhimurium (homology percentages, 65 and 64%, respectively), has a promoter region strongly homologous to the E. coli 17-bp class consensus promoter, and encodes a protein consisting of 165 amino acids (22 of which form the signal sequence). The plasmid-borne Y. enterocolitica ompH was found to be expressed both in the E. coli host and in minicells. The isolated outer membrane of Y. enterocolitica was shown to contain OmpH. The homology of the Y. enterocolitica OmpH protein is 66% with E. coli OmpH and 64% with S. typhimurium OmpH. All OmpH proteins have almost identical hydrophobic profiles, charge distributions, and predicted secondary structures. Because yersiniae are considered rather distant relatives of E. coli and S. typhimurium in the Enterobacteriaceae family, these results might indicate that most or all strains of the family Enterobacteriaceae have OmpH proteins remarkably homologous to those now sequenced.
KeywordMeSH Terms
Gene Expression Regulation, Bacterial
80. Cornelis  GR, Sluiters  C, Delor  I, Geib  D, Kaniga  K, Lambert de Rouvroit  C, Sory  MP, Vanooteghem  JC, Michiels  T,     ( 1991 )

ymoA, a Yersinia enterocolitica chromosomal gene modulating the expression of virulence functions.

Molecular microbiology 5 (5)
PMID : 1956283  :   DOI  :   10.1111/j.1365-2958.1991.tb01875.x    
Abstract >>
The virulence functions of Yersinia enterocolitica include the pYV-encoded Yop proteins and YadA adhesin as well as the chromosome-encoded enterotoxin, Yst. The yop and yadA genes form a temperature-activated regulon controlled by the transcriptional activator VirF. Gene virF, also localized on pYV, is itself thermoinduced in the absence of other pYV genes. The enterotoxin yst gene is silent in some collection strains including strain W22703. This paper describes two Tn5-Tc1 chromosomal insertion mutants of W22703 transcribing virF, and hence the yop and yadA genes, at low temperature. These mutants also resumed their production of Yst, with its typical temperature dependence. Both mutations were insertions in the same gene called ymoA for 'Yersinia modulator'. The cloned ymoA gene fully complemented the two mutations. Several properties of the mutants suggest that ymoA encodes a histone-like protein. According to the nucleic acid sequence, the product of ymoA is an 8064 Da protein rich in aspartic acid (9%), glutamic acid (9%) and lysine (10.5%), but the predicted amino acid sequence shows no similarity with any described histone-like protein. This work supports recent reports which propose a role for DNA topology and bacterial chromatin structure in thermoregulation of virulence functions.
KeywordMeSH Terms
Genes, Bacterial
81. Kuhnert  P, Korczak  BM, Stephan  R, Joosten  H, Iversen  C,     ( 2009 )

Phylogeny and prediction of genetic similarity of Cronobacter and related taxa by multilocus sequence analysis (MLSA).

International journal of food microbiology 136 (2)
PMID : 19321218  :   DOI  :   10.1016/j.ijfoodmicro.2009.02.022    
Abstract >>
Multilocus sequence analysis (MLSA) based on recN, rpoA and thdF genes was done on more than 30 species of the family Enterobacteriaceae with a focus on Cronobacter and the related genus Enterobacter. The sequences provide valuable data for phylogenetic, taxonomic and diagnostic purposes. Phylogenetic analysis showed that the genus Cronobacter forms a homogenous cluster related to recently described species of Enterobacter, but distant to other species of this genus. Combining sequence information on all three genes is highly representative for the species' %GC-content used as taxonomic marker. Sequence similarity of the three genes and even of recN alone can be used to extrapolate genetic similarities between species of Enterobacteriaceae. Finally, the rpoA gene sequence, which is the easiest one to determine, provides a powerful diagnostic tool to identify and differentiate species of this family. The comparative analysis gives important insights into the phylogeny and genetic relatedness of the family Enterobacteriaceae and will serve as a basis for further studies and clarifications on the taxonomy of this large and heterogeneous family.
KeywordMeSH Terms
Phylogeny
82. Annamalai  T, Venkitanarayanan  K,     ( 2009 )

Role of proP and proU in betaine uptake by Yersinia enterocolitica under cold and osmotic stress conditions.

Applied and environmental microbiology 75 (6)
PMID : 19114512  :   DOI  :   10.1128/AEM.01644-08     PMC  :   PMC2655450    
Abstract >>
Yersinia enterocolitica is a food-borne pathogen with the ability to grow at cold temperatures and tolerate high osmolarity. The bacterium tolerates osmotic stress by intracellular accumulation of osmolytes, such as betaine. The proP gene and proU operon of Y. enterocolitica were sequenced, and single (ProP(-) ProU(+) and ProP(+) ProU(-)) and double (ProP(-) ProU(-)) mutants were generated. Upon exposure to osmotic or chill stress, the single and double mutants demonstrated a reduction in betaine uptake compared to that in the wild type, suggesting that proP and proU play a role in betaine uptake during osmotic and chill stress responses of Y. enterocolitica.
KeywordMeSH Terms
Cold Temperature
Osmotic Pressure
83. Wiesand  U, Sorg  I, Amstutz  M, Wagner  S, van den Heuvel  J, Lührs  T, Cornelis  GR, Heinz  DW,     ( 2009 )

Structure of the type III secretion recognition protein YscU from Yersinia enterocolitica.

Journal of molecular biology 385 (3)
PMID : 18976663  :   DOI  :   10.1016/j.jmb.2008.10.034    
Abstract >>
The inner-membrane protein YscU has an important role during the assembly of the Yersinia enterocolitica type III secretion injectisome. Its cytoplasmic domain (YscU(C)) recognizes translocators as individual substrates in the export hierarchy. Activation of YscU entails autocleavage at a conserved NPTH motif. Modification of this motif markedly changes the properties of YscU, including translocator export cessation and production of longer injectisome needles. We determined the crystal structures of the uncleaved variants N263A and N263D of YscU(C) at 2.05 A and 1.55 A resolution, respectively. The globular domain is found to consist of a central, mixed beta-sheet surrounded by alpha-helices. The NPTH motif forms a type II beta-turn connecting two beta-strands. NMR analysis of cleaved and uncleaved YscU(C) indicates that the global structure of the protein is retained in cleaved YscU(C). The structure of YscU(C) variant N263D reveals that wild type YscU(C) is poised for cleavage due to an optimal reaction geometry for nucleophilic attack of the scissile bond by the side chain of Asn263. In vivo analysis of N263Q and H266A/R314A YscU variants showed a phenotype that combines the absence of translocator secretion with normal needle-length control. Comparing the structure of YscU to those of related proteins reveals that the linker domain between the N-terminal transmembrane domain and the autocleavage domain can switch from an extended to a largely alpha-helical conformation, allowing for optimal positioning of the autocleavage domain during injectisome assembly.
KeywordMeSH Terms
84. Michiels  T, Vanooteghem  JC, Lambert de Rouvroit  C, China  B, Gustin  A, Boudry  P, Cornelis  GR,     ( 1991 )

Analysis of virC, an operon involved in the secretion of Yop proteins by Yersinia enterocolitica.

Journal of bacteriology 173 (16)
PMID : 1860816  :   DOI  :   10.1128/jb.173.16.4994-5009.1991     PMC  :   PMC208188    
Abstract >>
Upon incubation at 37 degrees C in the absence of Ca2+ ions, pathogenic yersiniae release large amounts of pYV plasmid-encoded proteins called Yops that are involved in pathogenesis. Yersinia enterocolitica also expresses an outer membrane protein that is considered an adhesin and called YadA (previously called P1 or YopA). The production of Yops is coordinately regulated by a 20-kb region of the plasmid referred to as the Ca2+ dependence region and containing at least four loci called virA, virB, virC, and virF. The virF gene encodes a key transcriptional activator of yop genes. We have shown here that virF is also required for transcription of yadA and that virB is necessary for full transcription of the yop and yadA genes. In contrast, mutations in genes virA and virC had only a weak influence on the transcription of yop and yadA genes. These mutations did not affect the production of YadA but they completely inhibited the translocation of Yops from the intracellular compartment to the extracellular milieu. We inferred from these data that virA and virC are involved in the specific transport of Yops. We analyzed the 8.5-kb virC region and showed that it is most probably a single operon containing 13 open reading frames called yscA to yscM (for Yop secretion). Protein YscC has a putative signal sequence and shares significant homology with outer membrane proteins involved in the secretion of pullulanase by Klebsiella pneumoniae (PulD) or in the assembly of filamentous bacteriophages (gene IV product). At least the putative products of yscD, yscJ, and yscL were shown to be required for the export of Yops. YscJ turned out to be YlpB, a lipoprotein that we had detected previously. The yscM gene shares homology with yopH, the adjacent gene on the pYV plasmid. Its product does not appear to be necessary for the production of Yops. Transcription of the virC operon was subjected to the same regulation as the yop genes.
KeywordMeSH Terms
85. Seoane  A, García Lobo  JM,     ( 1991 )

Nucleotide sequence of a new class A beta-lactamase gene from the chromosome of Yersinia enterocolitica: implications for the evolution of class A beta-lactamases.

Molecular & general genetics : MGG 228 (1��2��)
PMID : 1886608  :   DOI  :   10.1007/bf00282468    
Abstract >>
The nucleotide sequence has been determined of a 1400 bp fragment from the chromosome of Yersinia enterocolitica containing the gene for beta-lactamase I. An ORF of 882 bp was identified, which could code for a polypeptide of 294 amino acids, closely related to other beta-lactamases of molecular class A. Amino acids 1-30 could constitute a signal peptide. The mature protein would be 264 amino acids long with a calculated pI of 6.2. Alignment of the amino acid sequence of the class A beta-lactamases suggested the existence of two subgroups in the same class, and this is discussed in the context of the evolution of the enzymes.
KeywordMeSH Terms
Genes, Bacterial
86. Isabel  S, Leblanc  E, Boissinot  M, Boudreau  DK, Grondin  M, Picard  FJ, Martel  EA, Parham  NJ, Chain  PS, Bader  DE, Mulvey  MR, Bryden  L, Roy  PH, Ouellette  M, Bergeron  MG,     ( 2008 )

Divergence among genes encoding the elongation factor Tu of Yersinia Species.

Journal of bacteriology 190 (22)
PMID : 18790860  :   DOI  :   10.1128/JB.01067-08     PMC  :   PMC2576667    
Abstract >>
Elongation factor Tu (EF-Tu), encoded by tuf genes, carries aminoacyl-tRNA to the ribosome during protein synthesis. Duplicated tuf genes (tufA and tufB), which are commonly found in enterobacterial species, usually coevolve via gene conversion and are very similar to one another. However, sequence analysis of tuf genes in our laboratory has revealed highly divergent copies in 72 strains spanning the genus Yersinia (representing 12 Yersinia species). The levels of intragenomic divergence between tufA and tufB sequences ranged from 8.3 to 16.2% for the genus Yersinia, which is significantly greater than the 0.0 to 3.6% divergence observed for other enterobacterial genera. We further explored tuf gene evolution in Yersinia and other Enterobacteriaceae by performing directed sequencing and phylogenetic analyses. Phylogenetic trees constructed using concatenated tufA and tufB sequences revealed a monophyletic genus Yersinia in the family Enterobacteriaceae. Moreover, Yersinia strains form clades within the genus that mostly correlate with their phenotypic and genetic classifications. These genetic analyses revealed an unusual divergence between Yersinia tufA and tufB sequences, a feature unique among sequenced Enterobacteriaceae and indicative of a genus-wide loss of gene conversion. Furthermore, they provided valuable phylogenetic information for possible reclassification and identification of Yersinia species.
KeywordMeSH Terms
Genetic Variation
87. Weitzel  AC, Larsen  RA,     ( 2008 )

Differential complementation of DeltatolA Escherichia coli by a Yersinia enterocolitica TolA homologue.

FEMS microbiology letters 282 (1)
PMID : 18355278  :   DOI  :   10.1111/j.1574-6968.2008.01115.x    
Abstract >>
The Tol system is an interactive set of envelope proteins that includes both outer and cytoplasmic membrane proteins. Central to this system is TolA, which spans the periplasmic space to communicate with residents of each membrane. To identify motifs involved in the protein/protein interactions through which TolA acts, the ability of a phylogenetically distinct TolA protein from Yersinia enterocolitica to function in the Tol system of Escherichia coli was examined. Although at least 59 codons shorter and only c. 67% identical to its E. coli homologue, the Y. enterocolitica gene encoded a protein that supported the physiological function of the Tol system in E. coli, and conferred sensitivity to the TolA-dependent colicins A, K, and E1, but interestingly, not to colicin N. The correlation of conferred phenotype with sequence similarities and differences provides a first step in defining essential structural motifs and their specific contributions to function.
KeywordMeSH Terms
88. Merhej  V, Adékambi  T, Pagnier  I, Raoult  D, Drancourt  M,     ( 2008 )

Yersinia massiliensis sp. nov., isolated from fresh water.

International journal of systematic and evolutionary microbiology 58 (Pt 4)
PMID : 18398169  :   DOI  :   10.1099/ijs.0.65219-0    
Abstract >>
Two bacterial organisms, 50640T and 823, were isolated from fresh water in Marseilles, France, and were further identified as members of the genus Yersinia on the basis of their phenotypic characteristics and 16S rRNA gene sequencing. Their unique phenotypic profile differed from that of closely related species of Yersinia bercovieri and Yersinia mollaretii by exhibiting positive indole and inositol tests, and from that of Yersinia frederiksenii by lacking the ability to ferment l-rhamnose. A polyphasic approach, including almost complete 16S rRNA gene sequencing (1461 bp) and partial sequencing of hsp60 (683 bp), gyrB (662 bp), sodA (624 bp) and rpoB (1049 bp) showed that isolates 50640T and 823 exhibited 98.5, 93.5, 90.4, 92.4 and 96.6 % similarity with Y. mollaretii, 98.7, 93.0, 90.1, 89.1 and 96.2 % with Y. bercovieri, and 98.4, 93.2, 89.8, 88.9 and 95.2 % with Y. frederiksenii, respectively. Both isolates exhibited an identical 16S rRNA gene sequence and differed by one to five point mutations in housekeeping gene sequences. Phylogenetic reconstructions based on the combination of these four housekeeping genes indicated that the two isolates formed a unique branch supported by a bootstrap value of 93 %. Their unique phenotypic traits, 16S rRNA gene sequence, together with housekeeping gene sequences exhibiting <97 % similarity with closely related species, and phylogenetic analyses suggested that the two isolates represent a so far undescribed Yersinia species. The name Yersinia massiliensis sp. nov. is proposed for this new taxon (type strain 50640T=CIP 109351T=CCUG 53443T; isolate 823=CIP 109352=CCUG 53444).
KeywordMeSH Terms
89. Hammerl  JA, Klein  I, Lanka  E, Appel  B, Hertwig  S,     ( 2008 )

Genetic and functional properties of the self-transmissible Yersinia enterocolitica plasmid pYE854, which mobilizes the virulence plasmid pYV.

Journal of bacteriology 190 (3)
PMID : 18055592  :   DOI  :   10.1128/JB.01467-07     PMC  :   PMC2223581    
Abstract >>
Yersinia strains frequently harbor plasmids, of which the virulence plasmid pYV, indigenous in pathogenic strains, has been thoroughly characterized during the last decades. Yet, it has been unknown whether the nonconjugative pYV can be transferred by helper plasmids naturally occurring in this genus. We have isolated the conjugative plasmids pYE854 (95.5 kb) and pYE966 (70 kb) from a nonpathogenic and a pathogenic Yersinia enterocolitica strain, respectively, and demonstrate that both plasmids are able to mobilize pYV. The complete sequence of pYE854 has been determined. The transfer proteins and oriT of the plasmid reveal similarities to the F factor. However, the pYE854 replicon does not belong to the IncF group and is more closely related to a plasmid of gram-positive bacteria. Plasmid pYE966 is very similar to pYE854 but lacks two DNA regions of the larger plasmid that are dispensable for conjugation.
KeywordMeSH Terms
Conjugation, Genetic
Gene Transfer, Horizontal
90. Büttner  CR, Sorg  I, Cornelis  GR, Heinz  DW, Niemann  HH,     ( 2008 )

Structure of the Yersinia enterocolitica type III secretion translocator chaperone SycD.

Journal of molecular biology 375 (4)
PMID : 18054956  :   DOI  :   10.1016/j.jmb.2007.11.009    
Abstract >>
Many Gram-negative bacteria use a type III secretion (T3S) system to directly inject effector molecules into eucaryotic cells in order to establish a symbiotic or pathogenic relationship with their host. The translocation of many T3S proteins requires specialized chaperones from the bacterial cytosol. SycD belongs to a class of T3S chaperones that assists the secretion of pore-forming translocators and, specifically chaperones the translocators YopB and YopD from enteropathogenic Yersinia enterocolitica. In addition, SycD is involved in the regulation of virulence factor biosynthesis and secretion. In this study, we present two crystal structures of Y. enterocolitica SycD at 1.95 and 2.6 A resolution, the first experimental structures of a T3S class II chaperone specific for translocators. The fold of SycD is entirely alpha-helical and reveals three tetratricopeptide repeat-like motifs that had been predicted from amino acid sequence. In both structures, SycD forms dimers utilizing residues from the first tetratricopeptide repeat motif. Using site-directed mutagenesis and size exclusion chromatography, we verified that SycD forms head-to-head homodimers in solution. Although in both structures, dimerization largely depends on the same residues, the two assemblies represent alternative dimers that exhibit different monomer orientations and overall shape. In these two distinct head-to-head dimers, both the concave and the convex surface of each monomer are accessible for interactions with the SycD binding partners YopB and YopD. A SycD variant carrying two point mutations in the dimerization interface is properly folded but defective in dimerization. Expression of this stable SycD monomer in Yersinia does not rescue the phenotype of a sycD null mutant, suggesting a physiological relevance of the dimerization interface.
KeywordMeSH Terms
91. Peracchi  A, Veiga-da-Cunha  M, Kuhara  T, Ellens  KW, Paczia  N, Stroobant  V, Seliga  AK, Marlaire  S, Jaisson  S, Bommer  GT, Sun  J, Huebner  K, Linster  CL, Cooper  AJL, Van Schaftingen  E,     ( 2017 )

Nit1 is a metabolite repair enzyme that hydrolyzes deaminated glutathione.

Proceedings of the National Academy of Sciences of the United States of America 114 (16)
PMID : 28373563  :   DOI  :   10.1073/pnas.1613736114     PMC  :   PMC5402446    
Abstract >>
The mammalian gene Nit1 (nitrilase-like protein 1) encodes a protein that is highly conserved in eukaryotes and is thought to act as a tumor suppressor. Despite being ?35% sequence identical to �s-amidase (Nit2), the Nit1 protein does not hydrolyze efficiently �\-ketoglutaramate (a known physiological substrate of Nit2), and its actual enzymatic function has so far remained a puzzle. In the present study, we demonstrate that both the mammalian Nit1 and its yeast ortholog are amidases highly active toward deaminated glutathione (dGSH; i.e., a form of glutathione in which the free amino group has been replaced by a carbonyl group). We further show that Nit1-KO mutants of both human and yeast cells accumulate dGSH and the same compound is excreted in large amounts in the urine of Nit1-KO mice. Finally, we show that several mammalian aminotransferases (transaminases), both cytosolic and mitochondrial, can form dGSH via a common (if slow) side-reaction and provide indirect evidence that transaminases are mainly responsible for dGSH formation in cultured mammalian cells. Altogether, these findings delineate a typical instance of metabolite repair, whereby the promiscuous activity of some abundant enzymes of primary metabolism leads to the formation of a useless and potentially harmful compound, which needs a suitable "repair enzyme" to be destroyed or reconverted into a useful metabolite. The need for a dGSH repair reaction does not appear to be limited to eukaryotes: We demonstrate that Nit1 homologs acting as excellent dGSH amidases also occur in Escherichia coli and other glutathione-producing bacteria.
KeywordMeSH Terms
amidase
aminotransferases
deaminated glutathione
metabolite repair
92. Marimon  O, Teixeira  JM, Cordeiro  TN, Soo  VW, Wood  TL, Mayzel  M, Amata  I, García  J, Morera  A, Gay  M, Vilaseca  M, Orekhov  VY, Wood  TK, Pons  M,     ( 2016 )

An oxygen-sensitive toxin-antitoxin system.

Nature communications 7 (N/A)
PMID : 27929062  :   DOI  :   10.1038/ncomms13634     PMC  :   PMC5155140    
Abstract >>
The Hha and TomB proteins from Escherichia coli form an oxygen-dependent toxin-antitoxin (TA) system. Here we show that YmoB, the Yersinia orthologue of TomB, and its single cysteine variant [C117S]YmoB can replace TomB as antitoxins in E. coli. In contrast to other TA systems, [C117S]YmoB transiently interacts with Hha (rather than forming a stable complex) and enhances the spontaneous oxidation of the Hha conserved cysteine residue to a -SOxH-containing species (sulfenic, sulfinic or sulfonic acid), which destabilizes the toxin. The nuclear magnetic resonance structure of [C117S]YmoB and the homology model of TomB show that the two proteins form a four-helix bundle with a conserved buried cysteine connected to the exterior by a channel with a diameter comparable to that of an oxygen molecule. The Hha interaction site is located on the opposite side of the helix bundle.
KeywordMeSH Terms
93. Skurnik  M, Wolf-Watz  H,     ( 1989 )

Analysis of the yopA gene encoding the Yop1 virulence determinants of Yersinia spp.

Molecular microbiology 3 (4)
PMID : 2761389  :   DOI  :   10.1111/j.1365-2958.1989.tb00198.x    
Abstract >>
The Yop proteins of Yersinia are important virulence determinants. The Yop1 protein sequences of Yersinia pestis, Yersinia pseudotuberculosis, and two Yersinia enterocolitica serotypes, 0:3 and 0:8, deduced from the nucleotide sequences of the corresponding yopA genes, were compared. Most differences were found in the hydrophilic domains of the proteins, whereas the hydrophobic domains were conserved. The amino acid sequences revealed a signal sequence 25 amino acids long. No cysteine residues were present, even though Yop1 forms a polymeric structure. The transcription startpoint of yopA was determined by primer extension. The coding region and transcription startpoint were separated by a leader sequence 270 nucleotides long. The yopA promoter sequence of Y.pestis is identical to the corresponding sequence of Y. pseudotuberculosis and transcription studies revealed that this promoter is active in Y.pestis. Thus, the inability of Y.pestis to express the Yop1 protein is due to a single base pair deletion in the coding region of the yopA gene of Y.pestis.
KeywordMeSH Terms
Adhesins, Bacterial
Genes
Genes, Bacterial
94. Meneely  KM, Ronnebaum  TA, Riley  AP, Prisinzano  TE, Lamb  AL,     ( 2016 )

Holo Structure and Steady State Kinetics of the Thiazolinyl Imine Reductases for Siderophore Biosynthesis.

Biochemistry 55 (38)
PMID : 27601130  :   DOI  :   10.1021/acs.biochem.6b00735     PMC  :   PMC5046821    
Abstract >>
Thiazolinyl imine reductases catalyze the NADPH-dependent reduction of a thiazoline to a thiazolidine, a required step in the formation of the siderophores yersiniabactin (Yersinia spp.) and pyochelin (Pseudomonas aeruginosa). These stand-alone nonribosomal peptide tailoring domains are structural homologues of sugar oxidoreductases. Two closed structures of the thiazolinyl imine reductase from Yersinia enterocolitica (Irp3) are presented here: an NADP(+)-bound structure to 1.45 ? resolution and a holo structure to 1.28 ? resolution with NADP(+) and a substrate analogue bound. Michaelis-Menten kinetics were measured using the same substrate analogue and the homologue from P. aeruginosa, PchG. The data presented here support the hypothesis that tyrosine 128 is the likely general acid residue for catalysis and also highlight the phosphopantetheine tunnel for tethering of the substrate to the nonribosomal peptide synthetase module during assembly line biosynthesis of the siderophore.
KeywordMeSH Terms
95. Platt-Samoraj  A, Syczy?o  K, Bancerz-Kisiel  A, Szczerba-Turek  A, Giżejewska  A, Szweda  W,     ( 2015 )

Yersinia enterocolitica strains isolated from beavers (Castor fiber).

Polish journal of veterinary sciences 18 (2)
PMID : 26172198  :   DOI  :   10.1515/pjvs-2015-0058    
Abstract >>
Pseudocloacal swabs and palatine tonsils from beavers have been examined for the Yersinia enterocolitica presence. Thirty-six samples from 24 beavers were collected and subjected to bacteriological examinations including sero- and biotypisation. Amplicons confirmed by PCR as Y. enterocolitica were sequenced. Positive samples originated from 4 out of the 24 beavers (16.7 %) and all the strains belonged to biotype 1A. The study suggested that Y. enterocolitica could be isolated from beavers, which may therefore be treated as a reservoir, a significant factor of water contamination and a vector of the Y. enterocolitica.
KeywordMeSH Terms
96. Lee  WL, Grimes  JM, Robinson  RC,     ( 2015 )

Yersinia effector YopO uses actin as bait to phosphorylate proteins that regulate actin polymerization.

Nature structural & molecular biology 22 (3)
PMID : 25664724  :   DOI  :   10.1038/nsmb.2964     PMC  :   PMC4745138    
Abstract >>
Pathogenic Yersinia species evade host immune systems through the injection of Yersinia outer proteins (Yops) into phagocytic cells. One Yop, YopO, also known as YpkA, induces actin-filament disruption, impairing phagocytosis. Here we describe the X-ray structure of Yersinia enterocolitica YopO in complex with actin, which reveals that YopO binds to an actin monomer in a manner that blocks polymerization yet allows the bound actin to interact with host actin-regulating proteins. SILAC-MS and biochemical analyses confirm that actin-polymerization regulators such as VASP, EVL, WASP, gelsolin and the formin diaphanous 1 are directly sequestered and phosphorylated by YopO through formation of ternary complexes with actin. This leads to a model in which YopO at the membrane sequesters actin from polymerization while using the bound actin as bait to recruit, phosphorylate and misregulate host actin-regulating proteins to disrupt phagocytosis.
KeywordMeSH Terms
Models, Molecular
97. Cordeiro  TN, García  J, Bernadó  P, Millet  O, Pons  M,     ( 2015 )

A Three-protein Charge Zipper Stabilizes a Complex Modulating Bacterial Gene Silencing.

The Journal of biological chemistry 290 (35)
PMID : 26085102  :   DOI  :   10.1074/jbc.M114.630400     PMC  :   PMC4571853    
Abstract >>
The Hha/YmoA nucleoid-associated proteins help selectively silence horizontally acquired genetic material, including pathogenicity and antibiotic resistance genes and their maintenance in the absence of selective pressure. Members of the Hha family contribute to gene silencing by binding to the N-terminal dimerization domain of H-NS and modifying its selectivity. Hha-like proteins and the H-NS N-terminal domain are unusually rich in charged residues, and their interaction is mostly electrostatic-driven but, nonetheless, highly selective. The NMR-based structural model of the complex between Hha/YmoA and the H-NS N-terminal dimerization domain reveals that the origin of the selectivity is the formation of a three-protein charge zipper with interdigitated complementary charged residues from Hha and the two units of the H-NS dimer. The free form of YmoA shows collective microsecond-millisecond dynamics that can by measured by NMR relaxation dispersion experiments and shows a linear dependence with the salt concentration. The number of residues sensing the collective dynamics and the population of the minor form increased in the presence of H-NS. Additionally, a single residue mutation in YmoA (D43N) abolished H-NS binding and the dynamics of the apo-form, suggesting the dynamics and binding are functionally related.
KeywordMeSH Terms
charge zipper complexes
electrostatics
gene silencing
horizontal gene transfer
nuclear magnetic resonance (NMR)
nucleoid-associated proteins
protein dynamic
protein-protein interaction
Gene Expression Regulation, Bacterial
Gene Transfer, Horizontal
98. Cornelis  G, Sluiters  C, de Rouvroit  CL, Michiels  T,     ( 1989 )

Homology between virF, the transcriptional activator of the Yersinia virulence regulon, and AraC, the Escherichia coli arabinose operon regulator.

Journal of bacteriology 171 (1)
PMID : 2644192  :   DOI  :   10.1128/jb.171.1.254-262.1989     PMC  :   PMC209580    
Abstract >>
Virulent yersiniae (Yersinia pestis, Y. pseudotuberculosis, and Y. enterocolitica) restrict their growth at 37 degrees C in rich medium deprived of calcium. This property, called calcium dependency, correlates with the secretion of Yersinia outer membrane proteins (Yops) and with pathogenicity. It is mediated by a 70-kilobase plasmid called pYV. The structural genes of the Yops (yop genes), as well as genes involved in the control of their expression (vir genes), have been localized on pYV. In this communication we show that virF encodes a transcriptional activator controlling the yop regulon. This activator is a 30,879-dalton protein related to AraC, the regulator of the Escherichia coli and Salmonella typhimurium arabinose operons. We also show in this paper that transcription of virF is thermodependent and presumably autoregulated. virF is thus responsible for the effect of temperature on the production of the Yops. Finally, we show that virF activates transcription of the yop genes independently of the presence of calcium ions. The role of calcium therefore remains unaccounted for.
KeywordMeSH Terms
Bacterial Proteins
Genes, Regulator
Operon
99. Emödy  L, Heesemann  J, Wolf-Watz  H, Skurnik  M, Kapperud  G, O'Toole  P, Wadström  T,     ( 1989 )

Binding to collagen by Yersinia enterocolitica and Yersinia pseudotuberculosis: evidence for yopA-mediated and chromosomally encoded mechanisms.

Journal of bacteriology 171 (12)
PMID : 2592347  :   DOI  :   10.1128/jb.171.12.6674-6679.1989     PMC  :   PMC210562    
Abstract >>
Binding of Yersinia enterocolitica and Yersinia pseudotuberculosis strains to type I, II, and IV collagens has been studied. Wild-type strains which harbored the 40- to 50-megadalton virulence plasmid specifically bound all three types of collagen. Curing of the virulence plasmid or Tn5 insertion in the yopA gene encoding the temperature-inducible outer membrane protein YOP1 abolished the binding of all three collagen types to Y. enterocolitica and type I and II collagens to Y. pseudotuberculosis. Full binding capacity was restored by introduction of the yopA gene into nonbinding Yersinia strains. Binding of type I, II, and IV collagens was expressed in Escherichia coli constructs harboring the yopA gene of either Y. enterocolitica or Y. pseudotuberculosis. The interaction of bacterial cells with type I collagen could be blocked by nonradiolabeled native collagens or denatured collagen but not with other serum and connective-tissue proteins. Unlabeled collagen could not displace bound radiolabeled collagen. The binding was inhibited by YOP1-specific polyclonal antibodies, in contrast to normal rabbit serum. The interaction was rapid and was quite resistant to heat treatment, to proteolytic enzymes, to various pHs in both acidic and alkaline ranges, and to the chaotropic agent urea. We propose that this newly identified interaction may be involved both in the first steps of the pathogenesis and in the complications of Yersinia infections affecting connective tissue.
KeywordMeSH Terms
100. Breitsprecher  D, Gherardi  E, Bleymüller  WM, Niemann  HH,     ( 2014 )

Crystal structure of an engineered YopM-InlB hybrid protein.

BMC structural biology 14 (N/A)
PMID : 24669959  :   DOI  :   10.1186/1472-6807-14-12     PMC  :   PMC3986869    
Abstract >>
The multi-domain protein InlB (internalin B) from Listeria monocytogenes is an agonist of the human receptor tyrosine kinase MET. Only the internalin domain directly interacts with MET. The internalin domain consists of seven central leucine-rich repeats (LRRs) flanked by an N-terminal helical cap domain and a C-terminal immunoglobulin-like structure. A potential function of the N-terminal cap in receptor binding could so far not be demonstrated by deleting the cap, since the cap is also implicated in nucleating folding of the LRR domain. We generated an InlB variant (YopM-InlB) in which the InlB cap domain was replaced by the unrelated N-terminal capping structure of the LRR protein YopM from Yersinia enterocolitica. The crystal structure of the engineered protein shows that it folds properly. Because the first LRR is structurally closely linked to the cap domain, we exchanged LRR1 along with the cap domain. This resulted in unexpected structural changes extending to LRR2 and LRR3, which are deeply involved in MET binding. As a consequence, the binding of YopM-InlB to MET was substantially weaker than that of wild type InlB. The engineered protein was about one order of magnitude less active in colony scatter assays than wild type InlB. We obtained a well-behaved InlB variant with an altered N-terminal capping structure through protein design. The reduced affinity for MET precludes a straightforward interpretation of the results from cell-based assays. Still, the engineered hybrid protein induced cell scatter, suggesting that the cap is required for folding and stability of InlB but is not essential for interactions that assemble the signalling-active receptor complex. The cap swap approach described here is clearly applicable to other L. monocytogenes internalins and other LRR proteins such as YopM and may yield useful structure/function correlates within this protein family.
KeywordMeSH Terms
Protein Engineering
101. Liang  J, Bi  Z, Shi  G, Xiao  Y, Qiu  H, Kou  Z, Hu  B, Jing  H, Wang  X,     ( 2014 )

Two novel ail-positive biotype 1A strains of Yersinia enterocolitica isolated in China with unequal adhesion and invasion properties.

Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases 27 (N/A)
PMID : 25038297  :   DOI  :   10.1016/j.meegid.2014.07.009    
Abstract >>
Yersinia enterocolitica is an enteric pathogen having six biotypes: 1A, 1B, 2, 3, 4, and 5. Different bioserotypes have been associated with varying pathogenicity, and the strains of biotype 1A lack the virulence-associated pYV-bearing genes and were once considered to be avirulent. However, there is growing epidemiological, clinical, and experimental evidence to suggest some biotype 1A isolates are virulent and can cause gastrointestinal disease. Here, we describe two biotype 1A strains discovered from 3807 isolates that carry the ail (attachment and invasion locus) gene. The two strains showed unique PFGE patterns compared to all other isolates in the Chinese Y. enterocolitica isolate PFGE database. Strain SDWL-003 isolated from a sheep shared ail sequence identical to A1 pattern, and the foxA (ferrioxamine receptor) sequence was identical to the pathogenic F5 pattern, besides, the PFGE patterns of SDWL-003 was also cluster to pathogenic branch; however it does not attach to or invade Hep-2 cells. The ail sequence of strain 2006RAT isolated from a Microtus fortis showed several mutations compared to other published genomes, and therefore formed an entirely new pathogenic pattern. Though it clustered to non-pathogenic block with foxA sequence polymorphism analysis or PFGE assay, the strain 2006RAT showed adhesion properties. The data here bring new insights into the molecular genetics of Y. enterocolitica biotype 1A, show some isolates of 1A biotype gaining potential pathogenicity using the function of the virulence gene - ail, and indicate the lateral gene transfer of ail virulence genes proceeded between pathogenic and nonpathogenic Y. enterocolitica.
KeywordMeSH Terms
Adhesion
Biotype 1A
Invasion
Yersinia enterocolitica
ail gene
Genes, Bacterial
102. Souza  RA, Imori  PF, Falcão  JP,     ( 2013 )

Multilocus sequence analysis and 16S rRNA gene sequencing reveal that Yersinia frederiksenii genospecies 2 is Yersinia massiliensis.

International journal of systematic and evolutionary microbiology 63 (Pt 8)
PMID : 23908151  :   DOI  :   10.1099/ijs.0.047175-0    
Abstract >>
Since Yersinia frederiksenii was first described in 1980, it has been recognized genotypically as a heterogeneous species, comprising three phenotypically indistinguishable genospecies. In this study, the sequence of the 16S rRNA gene and the concatenated sequences of six housekeeping genes (glnA, gyrB, hsp60, recA, rpoB and sodA) of all the currently known species of the genus Yersinia were used to determine the phylogenetic position of Y. frederiksenii genospecies 2 in the genus Yersinia. The phylogenetic analyses grouped the Y. frederiksenii genospecies 2 strains in a monophyletic group together with representative strains of Yersinia massiliensis. Moreover, the Y. frederiksenii genospecies 2 strains were also grouped apart from the other species of the genus Yersinia and far from the other two genospecies of Y. frederiksenii. All of the observations made in this study support the conclusion that Y. frederiksenii genospecies 2 should be reclassified as Y. massiliensis.
KeywordMeSH Terms
Phylogeny
103. Dhar  MS, Gupta  V, Virdi  JS,     ( 2013 )

Detection, distribution and characterization of novel superoxide dismutases from Yersinia enterocolitica Biovar 1A.

PloS one 8 (5)
PMID : 23704955  :   DOI  :   10.1371/journal.pone.0063919     PMC  :   PMC3660340    
Abstract >>
Superoxide dismutases (SODs) cause dismutation of superoxide radicals to hydrogen peroxide and oxygen. Besides protecting the cells against oxidative damage by endogenously generated oxygen radicals, SODs play an important role in intraphagocytic survival of pathogenic bacteria. The complete genome sequences of Yersinia enterocolitica strains show presence of three different sod genes. However, not much is known about the types of SODs present in Y. enterocolitica, their characteristics and role in virulence and intraphagocytic survival of this organism. This study reports detection and distribution of the three superoxide dismutase (sodA, sodB and sodC) genes in 59 strains of Y. enterocolitica and related species. The majority (94%) of the strains carried all three genes and constitutive expression of sodA and sodB was detected in 88% of the strains. Expression of sodC was not observed in any of the strains. The sodA, sodB and sodC genes of Y. enterocolitica were cloned in pET28a (+) vector. Recombinant SodA (82 kDa) and SodB (21 kDa) were expressed as homotetramer and monomer respectively, and showed activity over a broad range of pH (3.0-8.0) and temperature (4-70�XC). SodA and SodB showed optimal activity at 4�XC under acidic pH of 6.0 and 4.0 respectively. The secondary structures of recombinant SodA and SodB were studied using circular dichroism. Production of YeSodC was not observed even after cloning and expression in E. coli BL21(DE3) cells. A SodA(-) SodB(-) Escherichia coli strain which was unable to grow in medium supplemented with paraquat showed normal growth after complementation with Y. enterocolitica SodA or SodB. This is the first report on the distribution and characterization of superoxide dismutases from Y. enterocolitica. The low pH optima of both SodA and SodB encoded by Y. enterocolitica seem to implicate their role in acidic environments such as the intraphagocytic vesicles.
KeywordMeSH Terms
104. Young  VB, Miller  VL, Falkow  S, Schoolnik  GK,     ( 1990 )

Sequence, localization and function of the invasin protein of Yersinia enterocolitica.

Molecular microbiology 4 (7)
PMID : 2233250  :   DOI  :   10.1111/j.1365-2958.1990.tb00686.x    
Abstract >>
The inv locus of Yersinia enterocolitica is sufficient to convert a non-invasive Escherichia coli K12 strain into a microorganism that is able to penetrate cultured mammalian cells. The nucleotide sequence of inv reveals an open reading frame corresponding to an 835-amino-acid protein that is homologous to the invasin protein from Yersinia pseudotuberculosis. A polyclonal antiserum elicited by a synthetic peptide corresponding to the C-terminal 88 amino acids of this open reading frame detected a unique 100 kD protein in cell lysates of Y. enterocolitica strain 8081 c and in an E. coli strain harbouring the cloned inv gene. This protein localized to the outer membranes of both microorganisms and was cleaved by low concentrations of extracellular trypsin. HEp-2 cells were shown to attach to surfaces coated with bacterial outer membranes containing invasin and this attachment was destroyed by treatment of the membranes with trypsin. Thus it appears that the invasin protein from Y. enterocolitica is able to mediate both attachment to and entry of cultured epithelial cells.
KeywordMeSH Terms
Adhesins, Bacterial
105. Viitanen  AM, Toivanen  P, Skurnik  M,     ( 1990 )

The lcrE gene is part of an operon in the lcr region of Yersinia enterocolitica O:3.

Journal of bacteriology 172 (6)
PMID : 2160939  :   DOI  :   10.1128/jb.172.6.3152-3162.1990     PMC  :   PMC209120    
Abstract >>
The low-calcium response (lcr) region of the virulence plasmid of Yersinia pestis, Yersinia pseudotuberculosis, and Yersinia enterocolitica has been associated with calcium-dependent growth of bacteria. Mutations in the previously identified lcrE locus within the lcr region lack the repressor control of production of the lcr specific proteins, Yersinia outer membrane proteins (Yops) and V and W antigens. We sequenced a 3.3-kilobase-pair BamHI-ClaI fragment of the lcrE locus of pYVO3, the virulence plasmid of Y. enterocolitica O:3. The sequence of lcrE locus revealed six tightly packed open reading frames (ORFs), one of which was identified as the structural gene, lcrE, of the 32.9-kilodalton outer membrane protein LcrE (formerly known as Yop4b or YopN). Detection of large (greater than 2.3-kilobase-pair) transcripts strongly supports the conclusion that the lcrE gene and ORF1 to -5 function as an operon. Transcription of the lcrE-containing operon and the adjacent lcrB locus was found to be divergent, and the corresponding transcripts overlapped about 1,200 nucleotides. This extremely long overlap of the 5' ends of the transcripts produced from face-to-face promoters is a new finding; the longest overlap thus far found has been a few hundred nucleotides. Temperature was found to play the major role in regulation of transcription of the lcrE-containing operon of pYVO3, whereas Ca2+ concentration seemed to affect it only moderately.
KeywordMeSH Terms
Genes, Bacterial
Operon
106. Delor  I, Kaeckenbeeck  A, Wauters  G, Cornelis  GR,     ( 1990 )

Nucleotide sequence of yst, the Yersinia enterocolitica gene encoding the heat-stable enterotoxin, and prevalence of the gene among pathogenic and nonpathogenic yersiniae.

Infection and immunity 58 (9)
PMID : 2201642  :   PMC  :   PMC313599    
Abstract >>
The gene encoding the heat-stable enterotoxin (yst) was cloned from the chromosome of Yersinia enterocolitica W1024 (serotype O:9), and the nucleotide sequence was determined. The yst gene encodes a 71-amino-acid polypeptide. The C-terminal 30 amino acids of the predicted protein exactly correspond to the amino acid sequence of the toxin extracted from culture supernatants (T. Takao, N. Tominaga, and Y. Shimonishi, Biochem. Biophys. Res. Commun. 125:845-851, 1984). The N-terminal 18 amino acids have the properties of a signal sequence. The central 22 residues are removed during or after the secretion process. This organization in three domains (Pre, Pro, and mature Yst) resembles that of the enterotoxin STa of Escherichia coli. The degree of conservation between the E. coli and Y. enterocolitica toxins is much lower in the Pre and the Pro domains than in the mature proteins. The mature toxin of Y. enterocolitica is much larger than that of E. coli, but the active domain appears to be highly conserved. The yst gene of Y. enterocolitica introduced in E. coli K-12 directed the secretion of an active toxin. The cloned yst gene was used as an epidemiological probe among a collection of 174 strains representative of all Yersinia species except Yersinia pestis and numerous Y. enterocolitica subgroups. In Y. enterocolitica, there was a clear-cut difference between pathogenic and nonpathogenic strains: 89 of 89 pathogenic and none of 51 nonpathogenic strains contained yst-homologous DNA, suggesting that Yst is involved in pathogenesis. Among the other Yersinia species, only four strains of Yersinia kristensenii had DNA homologous to yst.
KeywordMeSH Terms
Genes, Bacterial
107. Guan  KL, Dixon  JE,     ( 1990 )

Protein tyrosine phosphatase activity of an essential virulence determinant in Yersinia.

Science (New York, N.Y.) 249 (4968)
PMID : 2166336  :   DOI  :   10.1126/science.2166336    
Abstract >>
Yersinia is the genus of bacteria that is the causative agent in plague or the black death, and on several occasions this organism has killed a significant portion of the world's population. An essential virulence determinant of Yersinia was shown to be a protein tyrosine phosphatase. The recombinant 50-kilodalton Yersinia phosphatase had a specificity for removal of phosphate from Tyr-containing as opposed to Ser/Thr-containing phosphopeptides and proteins. Site-directed mutagenesis was used to show that the Yersinia phosphatase possesses an essential Cys residue required for catalysis. Amino acids surrounding an essential Cys residue are highly conserved, as are other amino acids in the Yersinia and mammalian protein tyrosine phosphatases, suggesting that they use a common catalytic mechanism.
KeywordMeSH Terms
108. Stenkova  AM, Isaeva  MP, Shubin  FN, Rasskazov  VA, Rakin  AV,     ( 2011 )

Trends of the major porin gene (ompF) evolution: insight from the genus Yersinia.

PloS one 6 (5)
PMID : 21655186  :   DOI  :   10.1371/journal.pone.0020546     PMC  :   PMC3105102    
Abstract >>
OmpF is one of the major general porins of Enterobacteriaceae that belongs to the first line of bacterial defense and interactions with the biotic as well as abiotic environments. Porins are surface exposed and their structures strongly reflect the history of multiple interactions with the environmental challenges. Unfortunately, little is known on diversity of porin genes of Enterobacteriaceae and the genus Yersinia especially. We analyzed the sequences of the ompF gene from 73 Yersinia strains covering 14 known species. The phylogenetic analysis placed most of the Yersinia strains in the same line assigned by 16S rDNA-gyrB tree. Very high congruence in the tree topologies was observed for Y. enterocolitica, Y. kristensenii, Y. ruckeri, indicating that intragenic recombination in these species had no effect on the ompF gene. A significant level of intra- and interspecies recombination was found for Y. aleksiciae, Y. intermedia and Y. mollaretii. Our analysis shows that the ompF gene of Yersinia has evolved with nonrandom mutational rate under purifying selection. However, several surface loops in the OmpF porin contain positively selected sites, which very likely reflect adaptive diversification Yersinia to their ecological niches. To our knowledge, this is a first investigation of diversity of the porin gene covering the whole genus of the family Enterobacteriaceae. This study demonstrates that recombination and positive selection both contribute to evolution of ompF, but the relative contribution of these evolutionary forces are different among Yersinia species.
KeywordMeSH Terms
109.     ( 1997 )

Flagellar flhA, flhB and flhE genes, organized in an operon, cluster upstream from the inv locus in Yersinia enterocolitica.

Microbiology (Reading, England) 143 (Pt 11) (N/A)
PMID : 9387224  :   DOI  :   10.1099/00221287-143-11-3461    
Abstract >>
The inv gene of Yersinia enterocolitica codes for invasin, a member of the invasin/intimin-like protein family, which mediates the internalization of the bacterium into cultured epithelial cells. The putative inclusion of inv into a pathogenicity island was tested by investigating its flanking sequences. Indeed, the enteropathogenic Escherichia coli (EPEC) intimin, a member of the same family of proteins, is encoded by eaeA, a gene which belongs to a pathogenicity island. An ORF located upstream from inv was of particular interest since it appeared homologous both to the flagellar flhA gene and to sepA, an EPEC gene lying inside the same pathogenicity island as eaeA. A mutant in this ORF was non-motile and non-flagellated while its invasion phenotype remained unaffected. These data indicated that the ORF corresponded to the flhA gene of Y. enterocolitica. Subsequently, the flhB and flhE genes, located respectively upstream and downstream from flhA, were identified. The three flh genes appear to be transcribed from a single operon called flhB, according to the nomenclature used for Salmonella typhimurium. Intergenic sequence between flhE and inv includes a grey hole, with no recognizable function. Downstream from inv, we have detected the flagellar flgM operon as already reported. Finally, the incongruous localization of inv amidst the flagellar cluster is discussed; while transposition could explain this phenomenon, no trace of such an event was detected.
KeywordMeSH Terms
Adhesins, Bacterial
Carrier Proteins
Escherichia coli Proteins
110.     ( 1997 )

Yersinia enterocolitica induces apoptosis in macrophages by a process requiring functional type III secretion and translocation mechanisms and involving YopP, presumably acting as an effector protein.

Proceedings of the National Academy of Sciences of the United States of America 94 (23)
PMID : 9356502  :   DOI  :   10.1073/pnas.94.23.12638     PMC  :   PMC25065    
Abstract >>
Yersiniae, causative agents of plague and gastrointestinal diseases, secrete and translocate Yop effector proteins into the cytosol of macrophages, leading to disruption of host defense mechanisms. It is shown in this report that Yersinia enterocolitica induces apoptosis in macrophages and that this effect depends on YopP. Functional secretion and translocation mechanisms are required for YopP to act, strongly suggesting that this protein exerts its effect intracellularly, after translocation into the macrophages. YopP shows a high level of sequence similarity with AvrRxv, an avirulence protein from Xanthomonas campestris, a plant pathogen that induces programmed cell death in plant cells. This indicates possible similarities between the strategies used by pathogenic bacteria to elicit programmed cell death in both plant and animal hosts.
KeywordMeSH Terms
Apoptosis
Yersinia enterocolitica
111.     ( 1997 )

The novel heat-stable enterotoxin subtype gene (ystB) of Yersinia enterocolitica: nucleotide sequence and distribution of the yst genes.

Microbial pathogenesis 23 (4)
PMID : 9344780  :  
Abstract >>
The gene (ystB) encoding the novel subtype of the heat-stable enterotoxin (Y-STb) was cloned from the chromosome of a clinical isolate of Yersinia enterocolitica 84-50 (serotype O:5, biotype 1A) and the nucleotide sequence was determined. The ystB contained 216 base pairs that encoded a protein of 71 amino acid residues. The C-terminal 30 residues of the precursor protein exactly corresponded to the amino acid sequence of the Y-STb toxin, purified from the culture supernatant of the wild strain. Homology search revealed that there are 76.9% nucleotide sequence similarity between ystB and the Yersinia kristensenii ST gene, and 73.5% with the Y. enterocolitica prototype sequence of yst (ystA). When tested with the PCR generated ystB specific probe, 36 of 304 Y. enterocolitica strains from 18 countries hybridized with the probe. All the ystB probe positive strains belonged to biotype 1A and mostly to the so-called non-pathogenic serotype O:5, O:6, O:7,8 O:7,13 and O:10, while ystA was predominantly found among the pathogenic serotypes (78.5%). Out of 36 ystB gene positive strains, 18 were clinical origin from six countries, which were also positive in the suckling mice assay suggesting that ystB may play an important role in the pathogenesis, and the so-called non-pathogenic serotypes could be virulent for human.
KeywordMeSH Terms
Genes, Bacterial
112.     ( 1997 )

Identification of novel chromosomal loci affecting Yersinia enterocolitica pathogenesis.

Molecular microbiology 25 (2)
PMID : 9282744  :   DOI  :   10.1046/j.1365-2958.1997.4661829.x    
Abstract >>
Pathogenic species of the genus Yersinia have a marked tropism for lymphoid tissue during the early stages of infection. Bacterial survival at this site determines whether the disease is localized or progresses systemically, leading to a high rate of mortality. Several plasmid-encoded virulence genes are known to be required for survival and pathogenesis, but the contribution of chromosomal genes has been largely unexplored. This study represents the first intensive effort to characterize and determine the function of Yersinia chromosomal genes expressed in lymphoid tissue after intragastric infection. Strains harbouring cat fusions expressed in the host were isolated from Peyer's patch tissue of mice intragastrically infected and treated with chloramphenicol (Cm); genes identified in this manner were designated hre for host responsive element. The hre::cat strains that were Cm resistant in vivo (in mouse tissue) and Cm sensitive in vitro (on laboratory media at 26 degrees C) were identified and shown to consist of 61 different allelic groups. The hre::cat fusions from 48 of the allelic groups were cloned and characterized by DNA sequence analysis. The results identified genes necessary for iron acquisition, protection from environmental stresses, biosynthesis of cell envelope components and other diverse metabolic activities. However, the DNA sequence of many clones had no homology to other known genes. Insertion mutations were constructed for four hre genes and the resulting Y. enterocolitica mutants were tested in the mouse model for effects on pathogenesis. All of the mutant strains were affected for virulence when assayed for survival in host tissues and LD50 analysis.
KeywordMeSH Terms
Gene Expression Regulation, Bacterial
Genes, Bacterial
113.     ( 1996 )

The X-ray crystal structures of Yersinia tyrosine phosphatase with bound tungstate and nitrate. Mechanistic implications.

The Journal of biological chemistry 271 (31)
PMID : 8702535  :   DOI  :   10.1074/jbc.271.31.18780    
Abstract >>
X-ray crystal structures of the Yersinia tyrosine phosphatase (PTPase) in complex with tungstate and nitrate have been solved to 2. 4-A resolution. Tetrahedral tungstate, WO42-, is a competitive inhibitor of the enzyme and is isosteric with the substrate and product of the catalyzed reaction. Planar nitrate, NO3-, is isosteric with the PO3 moiety of a phosphotransfer transition state. The crystal structures of the Yersinia PTPase with and without ligands, together with biochemical data, permit modeling of key steps along the reaction pathway. These energy-minimized models are consistent with a general acid-catalyzed, in-line displacement of the phosphate moiety to Cys403 on the enzyme, followed by attack by a nucleophilic water molecule to release orthophosphate. This nucleophilic water molecule is identified in the crystal structure of the nitrate complex. The active site structure of the PTPase is compared to alkaline phosphatase, which employs a similar phosphomonoester hydrolysis mechanism. Both enzymes must stabilize charges at the nucleophile, the PO3 moiety of the transition state, and the leaving group. Both an associative (bond formation preceding bond cleavage) and a dissociative (bond cleavage preceding bond formation) mechanism were modeled, but a dissociative-like mechanism is favored for steric and chemical reasons. Since nearly all of the 47 invariant or highly conserved residues of the PTPase domain are clustered at the active site, we suggest that the mechanism postulated for the Yersinia enzyme is applicable to all the PTPases.
KeywordMeSH Terms
114.     ( 1996 )

Characterization of a large chromosomal "high-pathogenicity island" in biotype 1B Yersinia enterocolitica.

Journal of bacteriology 178 (23)
PMID : 8955291  :   DOI  :   10.1128/jb.178.23.6743-6751.1996     PMC  :   PMC178570    
Abstract >>
Pathogenic Yersinia spp. can be subdivided into highly pathogenic (high-pathogenicity) and low-pathogenicity strains. Several genes specific for the high-pathogenicity strains are clustered on a chromosomal fragment designated a "high-pathogenicity island" (HPI). In the present work, the HPI of biotype 1B strain Ye 8081 of Y. enterocolitica was characterized. We demonstrate important differences from the HPI of Y. pestis. The HPI of Y. enterocolitica is smaller (45 kb) and is not flanked by insertion sequences. A copy of the gene coding for the tRNA-Asn is present at one extremity of the HPI and may, as in uropathogenic Escherichia coli, participate in the excision of the island. In addition to the genes encoding the yersiniabactin-pesticin receptor and the high-molecular-weight protein 2, four repeated sequences are present on the HPI of Y. enterocolitica. At least two of them are insertion elements: previously described IS1328 and newly characterized IS1400. Comparison of the HPI of strain Ye 8081 with that of other Y. enterocolitica strains of biotype 1B indicates that most of the island is conserved, apart from 15 kb at the left-hand end which is variable, especially in the region where three repeated sequences are clustered.
KeywordMeSH Terms
Bacterial Proteins
Chromosomes, Bacterial
Genes, Bacterial
115.     ( 1996 )

Identification and characterization of the Yersinia enterocolitica gsrA gene, which protectively responds to intracellular stress induced by macrophage phagocytosis and to extracellular environmental stress.

Infection and immunity 64 (8)
PMID : 8757824  :   PMC  :   PMC174178    
Abstract >>
Yersinia enterocolitica is able to resist the microbicidal mechanisms of macrophages and to grow within phagocytic cells. Some bacteria including Y. enterocolitica have been shown to respond to the hostile environment in macrophages by producing a set of stress proteins which are also induced by environmental stresses. To understand the role of stress proteins in intracellular survival of bacteria, we identified and cloned a Y. enterocolitica gene, called gsrA (global stress requirement). The gsrA gene was identified because its insertional inactivation by a transposon resulted in the inability of the organism to grow at an elevated temperature and to survive within macrophages after phagocytosis. The gsrA gene was sequenced and shown to encode a basic, 49,500-Da protein. The GsrA protein shows significant amino acid sequence homology to the HtrA stress protein which was originally identified in Escherichia coli. Furthermore, the genetically defined Y. enterocolitica gsrA mutant was constructed and characterized. The insertional mutation of gsrA resulted in inhibition of growth at temperatures above 39 degrees C and greatly increased susceptibility to oxidative and osmotic stresses. The mutant additionally lost the ability to survive and replicate within macrophages. These results, taken together, indicate that the gsrA gene is an essential component of the protection mechanism employed by Y. enterocolitica, allowing it to respond to the intracellular stress in macrophages as well as extracellular environmental stress.
KeywordMeSH Terms
Bacterial Proteins
Gene Expression Regulation, Bacterial
Genes, Bacterial
116.     ( 1994 )

The evolutionarily conserved ribosomal protein L23 and the cationic urease beta-subunit of Yersinia enterocolitica O:3 belong to the immunodominant antigens in Yersinia-triggered reactive arthritis: implications for autoimmunity.

Molecular medicine (Cambridge, Mass.) 1 (1)
PMID : 8790600  :   PMC  :   PMC2229931    
Abstract >>
Reactive arthritis (ReA) is a T cell mediated inflammatory process. The immune response is primarily directed against a triggering organism, although autoimmunity has been invoked in long-lasting, antibiotic-resistant disease. Although a variety of different species are known to trigger Reactive arthritis, the clinical manifestations are strikingly similar as well as closely associated to the HLA-B27 (70%). Various antigenic fractions and single antigens of Yersinia enterocolitica were prepared, and their immunological activity was assessed by proliferation of synovial fluid mononuclear cells from 10 Reactive arthritis patients. The gene encoding one hitherto unknown antigen has been sequenced. Nonapeptides deduced from sequences of the target antigens were tested in an assembly assay. Two immunodominant proteins of Yersinia enterocolitica were found, one being the urease beta-subunit and the other the 50 S ribosomal protein L23. The latter has been sequenced and belongs to the evolutionarily conserved ribosomal proteins with homology to procaryotes and eucaryotes. One nonapeptide derived from the urease beta-subunit was identified as a possible epitope for HLA-B27-restricted cytotoxic T cells by its high affinity. This epitope is also highly conserved. Sharing of conserved immunodominant proteins between different disease triggering microorganisms could provide an explanation of the shared clinical picture in Reactive arthritis. Moreover, autoimmunity in Reactive arthritis might be mediated by antigen mimicry between evolutionarily conserved epitopes of ribosomal proteins and their host analogs.
KeywordMeSH Terms
Escherichia coli Proteins
117.     ( 1996 )

The gene cluster directing O-antigen biosynthesis in Yersinia enterocolitica serotype 0:8: identification of the genes for mannose and galactose biosynthesis and the gene for the O-antigen polymerase.

Microbiology (Reading, England) 142 (Pt 2) (N/A)
PMID : 8932701  :   DOI  :   10.1099/13500872-142-2-277    
Abstract >>
The rfb gene cluster of Yersinia enterocolitica serotype O:8 (YeO8) strain 8081-c was cloned by cosmid cloning. Restriction mapping, deletion analysis and transposon mutagenesis showed that about 19 kb of the cloned DNA is essential for the synthesis and expression of the YeO8 O-side-chain in Escherichia coli. Deletion analysis generated a derivative that expressed semi-rough LPS, a phenotype typical of an rfc mutant lacking the O-antigen polymerase. The deletions and transcomplementation experiments allowed localization of the rfc gene to the 3'-end of the rfb gene cluster. The deduced YeO8 Rfc did not share significant amino acid sequence similarity with any other protein, but its amino acid composition and hydrophobicity profile are similar to those of identified Rfc proteins. In addition, the codon usage of the rfc gene is similar to other rfc genes. Nucleotide sequence analysis identified three other genes upstream of rfc. Two of the gene products showed 60-70% identity to the RfbM and RfbK proteins that are biosynthetic enzymes for the GDPmannose pathway of enterobacteria. The third gene product was about 50-80% identical to the bacterial GalE protein, UDPglucose 4-epimerase, which catalyses the epimerization of UDPglucose to UDPgalactose. Since mannose and galactose are both present in the YeO8 O-antigen repeat unit, the above three genes are likely to belong to the rfb gene cluster. A gene similar to the gsk gene downstream of rfc, and genes similar to adk and hemH upstream of the rfb gene cluster, were recognized. Thus the rfb gene cluster of YeO8 is located between the adk-hemH and gsk loci, and the order is adk-hemH-rfb-rfc-gsk in the chromosome. Also in other Yersinia spp., the locus downstream of the hemH gene is occupied by gene clusters associated with LPS biosynthesis.
KeywordMeSH Terms
Genes, Bacterial
Multigene Family
118.     ( 1997 )

Nucleotide sequence of a gene encoding the novel Yersinia enterocolitica heat-stable enterotoxin that includes a pro-region-like sequence in its mature toxin molecule.

Microbial pathogenesis 22 (2)
PMID : 9049998  :  
Abstract >>
A novel ST (Y-STc) produced by a pathogenic strain of Yersinia enterocolitica was recently purified and the mature toxin was found to include the pro-region-like sequence (Yoshino et al., FEBS Lett 362: 319-322, 1995). In the present study, we report the nucleotide sequence of the ystC gene. We also present the amino acid sequence of a ystC gene product secreted by a recombinant strain of Escherichia coli. From the nucleotide sequence of ystC gene, the Y-STc precursor protein was deduced to consist of 72 amino acid residues and to be comprised of two regions (pre- and mature toxin regions), while the known ST precursor proteins are comprised of three regions (pre-, pro-, and mature toxin regions). Therefore, we concluded that a long polypeptide corresponding to a pro-toxin of other Sts is secreted as the Y-STc mature toxin. This implies that the proteolytic processing of a pro-form appearing in the periplasm is not essential for the secretion of Yersinia-ST to the extracellular through the outer membrane. The recombinant Y-STc released by E. coli was found to be shorter and composed of C-terminal 24 amino acid residues of the native Y-STc. On the basis of these results, we propose that the cleavage site between the pro- and mature toxin regions of ST depends on both the amino acid sequence of the pro-toxin and on the bacterial protease system responsible for the maturation.
KeywordMeSH Terms
119.     ( 1996 )

Bacterial polysaccharide synthesis and gene nomenclature.

Trends in microbiology 4 (12)
PMID : 9004408  :  
Abstract >>
Gene nomenclature for bacterial surface polysaccharides is complicated by the large number of structures and genes. We propose a scheme applicable to all species that distinguishes different classes of genes, provides a single name for all genes of a given function and greatly facilitates comparative studies.
KeywordMeSH Terms
Terminology as Topic
120.     ( 1995 )

yst gene expression in Yersinia enterocolitica is positively regulated by a chromosomal region that is highly homologous to Escherichia coli host factor 1 gene (hfq).

Molecular microbiology 18 (5)
PMID : 8825090  :   DOI  :   10.1111/j.1365-2958.1995.18050859.x    
Abstract >>
Yersinia enterocolitica produces heat-stable enterotoxin (Y-ST) as one of its virulence factors. The yst gene, however, frequently and spontaneously becomes inactive (silent) during storage, which is accompanied by concurrent changes in some biological properties such as colony morphology, growth rate, carbon fermentation and ornithine decarboxylase activity. Northern blot analysis revealed that the level of mRNA for yst was repressed. To investigate the regulatory region, we transformed a yst-silent strain with a chromosomal gene library of Y-ST producing an isogenic counterpart. Out of 3604 clones, one clone resumed the Y-ST production and concurrently other biological properties. An open reading frame in this clone was designated as yrp, yersinia regulator for pleiotropic phenotype. Deduced from the nucleotide sequence, Yrp was a small protein of I01 amino acids with no similarity with any regulatory factor described, but showed high homology with an Escherichia coli host factor 1 required for Q beta-replicase, and with an Azorhizobium caulinodans NrfA required for the expression of nifA. The yrp gene mutation caused decreased negative supercoiling of plasmids, as did hfq. The yrp gene could similarly complement Y-ST production in two other silent strains of Y. enterocolitica. In all three silent strains examined, we found various mutations in the yrp region.
KeywordMeSH Terms
Chromosomes, Bacterial
121.     ( 1997 )

Passive immunity to infection with Yersinia spp. mediated by anti-recombinant V antigen is dependent on polymorphism of V antigen.

Infection and immunity 65 (2)
PMID : 9009295  :   PMC  :   PMC174615    
Abstract >>
The V antigen is a 37-kDa secreted polypeptide encoded on the 70-kb virulence plasmid of pathogenic Yersinia spp. Besides having regulatory functions, it is known to be a virulence factor and a protective antigen. DNA sequencing of the most common serotypes of human pathogenic Yersinia enterocolitica and Y. pseudotuberculosis revealed that two evolutionary distinct types of V antigen exist in Yersinia spp. One type is represented by Y. enterocolitica serotype 08 strains WA, WA-314, and NCTC 10938 (designated LcrV-YenO8); the other type comprises Y. pestis, Y. pseudotuberculosis, and Y. enterocolitica serotypes O3, O9, and O5,27 (LcrV-Yps). A hypervariable region between amino acids 225 and 232 represents the main difference between the two types. By raising monospecific antisera against both types of V antigen (anti-rVO8 and anti-rVO3), we were able to demonstrate that, in general, passive immunization of mice against a challenge with yersiniae was possible with both anti-Y. enterocolitica V antigen sera. However, anti-V antigen serum was protective only if the immunizing V antigen was the same type as the V antigen produced by the infective strain. The failure of the American V antigen type represented by Y. enterocolitica serotype O8 to protect against Yersinia spp. carrying the other V antigen type (LcrV-Yps) could be an explanation for the presence of plague foci in American countries.
KeywordMeSH Terms
Immunization, Passive
122.     ( 1996 )

Identification of the galE gene and a galE homolog and characterization of their roles in the biosynthesis of lipopolysaccharide in a serotype O:8 strain of Yersinia enterocolitica.

Journal of bacteriology 178 (20)
PMID : 8830687  :   DOI  :   10.1128/jb.178.20.5916-5924.1996     PMC  :   PMC178447    
Abstract >>
A clone that complements mutations in Yersinia enterocolitica lipopolysaccharide (LPS) core biosynthesis was isolated, and the DNA sequence of the clone was determined. Three complete open reading frames and one partial open reading frame were located on the cloned DNA fragment. The first, partial, open reading frame had homology to the rfbK gene. The remaining reading frames had homology to galE, rol, and gsk. Analysis of the galE homolog indicates that although it can complement an Escherichia coli galE mutant, its primary function in Y. enterocolitica is not in the production of UDP galactose but, instead, some other nucleotide sugar required for LPS biosynthesis. This gene has been renamed lse, for LPS sugar epimerase. The rol homolog has been demonstrated to have a role in Y. enterocolitica serotype 0:8 O-polysaccharide antigen chain length determination. An additional galE homolog has been identified in Y. enterocolitica by homology to the E. coli gene. The product of this gene has UDP galactose 4-epimerase activity in both E. coli and Y. enterocolitica. This gene is linked to the other genes of the galactose utilization pathway, similar to what is seen in other members of the family Enterobacteriaceae. Although Y. enterocolitica 0:8 strains are reported to have galactose as a constituent of LPS, a strain containing a mutation in this galE gene does not exhibit any LPS defects.
KeywordMeSH Terms
Genes, Bacterial
123.     ( 1993 )

YopB and YopD constitute a novel class of Yersinia Yop proteins.

Infection and immunity 61 (1)
PMID : 8418066  :   PMC  :   PMC302689    
Abstract >>
Virulent Yersinia species harbor a common plasmid that encodes essential virulence determinants (Yersinia outer proteins [Yops]), which are regulated by the extracellular stimuli Ca2+ and temperature. The V-antigen-encoding operon has been shown to be involved in the Ca(2+)-regulated negative pathway. The genetic organization of the V-antigen operon and the sequence of the lcrGVH genes were recently presented. The V-antigen operon was shown to be a polycistronic operon having the gene order lcrGVH-yopBD (T. Bergman, S. H?kansson, A. Forsberg, L. Norlander, A. Macellaro, A. B?ckman, I. B?lin, and H. Wolf-Watz, J. Bacteriol. 173:1607-1616, 1991; S. B. Price, K. Y. Leung, S. S. Barve, and S. C. Straley, J. Bacteriol. 171:5646-5653, 1989). We present here the sequence of the distal part of the V-antigen operons of Yersinia pseudotuberculosis and Yersinia enterocolitica. The sequence information encompasses the yopB and yopD genes and a downstream region in both species. We conclude that the V-antigen operon ends with the yopD gene. This conclusion is strengthened by the observation of an insertion-like element downstream of the yopD gene. The translational start codons of YopB and YopD have been identified by N-terminal amino acid sequencing. By computer analysis, the yopB and yopD gene products were found to be possible transmembrane proteins, and YopD was shown to contain an amphipathic alpha-helix in its carboxy terminus. These findings contrast with the general globular pattern observed for other Yops. Homology between Yersinia LcrH and Shigella flexneri IppI and between Yersinia YopB and S. flexneri IpaB was found, suggesting conservation of this locus between these two genera. YopB was also found to have a moderate level of homology, especially within the hydrophobic regions, to members of the RTX protein family of alpha-hemolysins and leukotoxins, indicating that YopB might exhibit a similar function.
KeywordMeSH Terms
124.     ( 1997 )

Virulence and arsenic resistance in Yersiniae.

Journal of bacteriology 179 (3)
PMID : 9006011  :   DOI  :   10.1128/jb.179.3.612-619.1997     PMC  :   PMC178738    
Abstract >>
The genus Yersinia contains three pathogenic species: Yersinia pestis, Y. pseudotuberculosis, and Y. enterocolitica. Only a few biotypes and serotypes of Y. enterocolitica are pathogenic, and these form two distinct groups: some are of low virulence, and they are encountered worldwide; others, mainly encountered in North America, are markedly more virulent. All pathogenic yersiniae possess a 70-kb virulence plasmid called pYV which encodes secreted antihost proteins called Yops as well as a type III secretion machinery that is required for Yop secretion. Genes encoding Yop synthesis and secretion are tightly clustered in three quadrants of the pYV plasmid. We show here that in the low-virulence strains of Y. enterocolitica, the fourth quadrant of the plasmid contains a new class II transposon, Tn2502. This transposon encodes a defective transposase, but transposition can be complemented in trans by Tn2501, another class II transposon. Tn2502 was not detected in the pYV plasmids of the more virulent American strains of Y. enterocolitica, of Y. pseudotuberculosis, and of Y. pestis. Tn2502 confers arsenite and arsenate resistance. This resistance involves four genes; three are homologous to the arsRBC genes present on the Escherichia coli chromosome, but no homolog of the fourth one, arsH, has been found. The systematic presence of such a resistance operon on a virulence plasmid is unusual and could be related to the recent spread of low-virulence Y. enterocolitica strains. The presence of this ars operon also constitutes the first significant difference between the pYV plasmids from different Yersinia species.
KeywordMeSH Terms
Bacterial Proteins
Escherichia coli Proteins
Genes, Bacterial
125.     ( 1993 )

Identification of the Yersinia enterocolitica urease beta subunit as a target antigen for human synovial T lymphocytes in reactive arthritis.

Infection and immunity 61 (10)
PMID : 8406844  :   PMC  :   PMC281190    
Abstract >>
The local T-cell response to bacterial antigens is involved in the pathogenesis of reactive arthritis (ReA). Here, we have identified a 19-kDa antigen of Yersinia enterocolitica O:9 recognized by Yersinia-specific synovial fluid CD4+ T cells in two patients with Yersinia-induced ReA. N-terminal amino acid sequencing of this protein revealed that it was identical to the 19-kDa urease beta subunit of Y. enterocolitica O:9. This protein has previously been shown to be arthritogenic in preimmunized rats after intra-articular injection. Analysis of the T-cell response to this protein showed that it contains several T-cell epitopes, one of which cross-reacts with other enterobacteria not able to induce ReA. This indicates that the arthritogenicity of the 19-kDa antigen is not a property of the 19-kDa protein alone but is dependent on its expression in bacteria able to induce ReA.
KeywordMeSH Terms
126.     ( 1994 )

The novel hexapeptide motif found in the acyltransferases LpxA and LpxD of lipid A biosynthesis is conserved in various bacteria.

FEBS letters 337 (3)
PMID : 8293817  :   DOI  :   10.1016/0014-5793(94)80211-4    
Abstract >>
Two bacterial acyltransferases (LpxA of Escherichia coli, LpxD of E. coli and Salmonella typhimurium) have previously been shown to consist of a very unusual tandem-repeat structure with tens of repeating hexapeptides (24 hexapeptides in LpxA, 26 in LpxD). By sequencing LpxD of Yersinia enterocolitica (a distant relative of E. coli and S. typhimurium within Enterobacteriaceae) as well as LpxA of S. typhimurium and Y. enterocolitica, and by analyzing the existing data on these enzymes of Ricketsia rickettsii, it was now shown that the hexapeptide repeat pattern is a very conservative property of these enzymes. Even though the overall homology (allowing equivalent amino acids) between the four proteins was only 59% in LpxA and 58% in LpxD, the homology in the first residue of each hexapeptide was 87% in LpxA and 100% in LpxD. Secondary structure prediction by PredictProtein server suggested a very strong beta strand dominance in all the hexad regions. Accordingly, LpxA and LpxD of various bacterial origins can now be regarded as structurally very unusual enzymes, largely consisting of hexad repeats.
KeywordMeSH Terms
127.     ( 1993 )

The TonB-dependent ferrichrome receptor FcuA of Yersinia enterocolitica: evidence against a strict co-evolution of receptor structure and substrate specificity.

Molecular microbiology 7 (3)
PMID : 8384682  :   DOI  :   10.1111/j.1365-2958.1993.tb01130.x    
Abstract >>
A Yersinia enterocolitica receptor mutant was isolated which is impaired in ferrichrome uptake. The receptor-encoding gene fcuA was cloned in Escherichia coli K-12. A fcuA mutant of Y. enterocolitica could be complemented by the cloned DNA fragment. The FcuA-encoding region was sequenced and an open reading frame encoding 758 amino acids including a signal sequence of 36 amino acids was found. FcuA shared 34.6% amino acid sequence homology with FatA, the anguibactin receptor of Vibrio anguillarum, but only 20.6% homology with FhuA, the ferrichrome receptor of E. coli. Since the structure of anguibactin differs strongly from that of ferrichrome there seems to be no co-evolution of receptor structure and substrate specificity. The ferrichrome receptors FcuA from Y. enterocolitica and FhuA from E. coli had slightly different substrate specificities. In contrast to FhuA from E. coli, FcuA from Y. enterocolitica was more stereoselective and failed to transport enantio ferrichrome. Three additional ferrichrome receptors were cloned from Pantoea agglomerans (formerly Erwinia herbicola), Salmonella paratyphi B and Salmonella typhimurium. Their substrate specificity was similar but not identical.
KeywordMeSH Terms
Escherichia coli Proteins
128.     ( 1995 )

A novel locus of Yersinia enterocolitica serotype O:3 involved in lipopolysaccharide outer core biosynthesis.

Molecular microbiology 17 (3)
PMID : 8559076  :   DOI  :   10.1111/j.1365-2958.1995.mmi_17030575.x    
Abstract >>
Yersinia enterocolitica serotype O:3 strain 6471/76-c (YeO3-c) was sensitive to bacteriophage phi R1-37 when grown at 37 degrees C but not when grown at 22 degrees C because of steric hindrance by abundant lipopolysaccharide (LPS) O-side chain (O-antigen) expressed at 22 degrees C. The transposon library of YeO3-c was grown at 37 degrees C and screened for phage phi R1-37-resistant transposon insertion mutants. Three types of mutant were isolated: (i) phage receptor mutants expressing O-antigen (LPS-smooth), (ii) phage receptor mutants not expressing O-antigen (LPS-rough), and (iii) LPS-smooth mutants with the phage receptor constitutively sterically blocked. Mutant type (i) was characterized in detail; the transposon insertion inactivates an operon, named the trs operon. The main findings based on this mutant are: (i) the frs operon is involved in the biosynthesis of the LPS outer core in YeO3-c; the nucleotide sequence of the trs operon revealed eight novel genes showing similarly to known polysaccharide biosynthetic genes of various Gram-negative bacteria as well as to capsule biosynthesis genes of Staphylococcus aureus; (ii) the biosynthesis of the core of YeO3-c involves at least two genetic loci; (iii) the trs operon is required for the biosynthesis of the bacteriophage phi R1-37 receptor structures; (iv) the homopolymeric O-antigen of YeO3-c is ligated to the inner core in Y. enterocolitica O:3; (v) the trs operon is located between the adk-hemH and galE-gsk gene pairs in the Y. enterocolitica chromosome; and (vi) the phage phi R1-37 receptor is present in many but not in all Y. enterocolitica serotypes. The results also allow us to speculate that the trs operon is a relic of the ancestral rfb region of Y. enterocolitica O:3 carrying genes indispensable for the completion of the core polysaccharide biosynthesis.
KeywordMeSH Terms
Genes, Bacterial
129.     ( 1996 )

Construction and characterization of a Yersinia enterocolitica O:8 high-temperature requirement (htrA) isogenic mutant.

Infection and immunity 64 (6)
PMID : 8675311  :   PMC  :   PMC174040    
Abstract >>
The high-temperature requirement (HtrA) family of stress response proteins are induced by different environmental stress conditions in a variety of bacteria and have been shown to contribute to the pathogenicity of some of these species. In this study, the htrA gene from Yersinia enterocolitica O:8 was amplified, cloned, and sequenced. Analysis of the deduced amino acid sequence predicted that the putative HtrA homolog contains a serine protease active site and a catalytic triad characteristic of trypsin-like serine proteases, structural features characteristic of previously described HtrA proteins. In order to evaluate the biological functions of Y. enterocolitica HtrA, an isogenic mutant was constructed by a reverse-genetics PCR-based approach. Characterization of the mutant provided evidence supporting a stress response function for the Y. enterocolitica htrA gene product. In contrast to the parent strain, the mutant showed increased sensitivity to killing by H2O2, O2- and temperature stress (50 degrees C). The mutant was avirulent in the murine yersiniosis injection model and offered partial protection to mice challenged with the parent strain. Further studies with the Y. enterocolitica htrA mutant should increase our knowledge of the host-pathogen interactions which occur during Yersinia infections.
KeywordMeSH Terms
Heat-Shock Proteins
Periplasmic Proteins
130.     ( 1993 )

High-molecular-weight protein 2 of Yersinia enterocolitica is homologous to AngR of Vibrio anguillarum and belongs to a family of proteins involved in nonribosomal peptide synthesis.

Journal of bacteriology 175 (17)
PMID : 8366034  :   DOI  :   10.1128/jb.175.17.5488-5504.1993     PMC  :   PMC206605    
Abstract >>
The iron-regulated irp2 gene is specific for the highly pathogenic Yersinia species and encodes high-molecular-weight protein 2 (HMWP2). Despite the established correlation between the presence of HMWP2 and virulence, the role of this protein is still unknown. To gain insight into the function of HMWP2, the entire coding sequence and the promoter of irp2 were sequenced. Two putative -35 and -10 promoter sequences were identified upstream of a large open reading frame, and two potential Fur-binding sites were found overlapping the second -35 box. The large open reading frame is composed of 6,126 nucleotides and may encode a protein of 2,035 amino acids (ca. 228 kDa) with a pI of 5.81. A signal sequence was not present at the N terminus of the protein. Despite the existence of 30 cysteine residues, carboxymethylation prevented the formation of most if not all disulfide bonds that otherwise occurred when the cells were sonicated. The protein was composed of three main domains: a central region of ca. 850 residues, bordered on each side by a repeat of 550 residues. A high degree of identity (44.5%) was found between HMWP2 and the protein AngR of Vibrio anguillarum. The central part of HMWP2 (after removal of a loop of 337 residues) also displayed significant homology with proteins belonging to the superfamily of adenylate-forming enzymes and, like them, possessed a putative ATP-binding motif that is also present in AngR. In addition, HMWP2 shared with the group of antibiotic and enterochelin synthetases a potential amino acid-binding site. Six consensus sequences defining the superfamily and four defining the family of synthetases were derived from the multiple alignment of the 30 sequences of proteins or repeated domains. A phylogenetic tree that was constructed showed that HMWP2 and AngR are in a family composed of Lys2, EntF, and the tyrocidine, gramicidin, and Beta-lactam synthetases. This finding suggests that HMWP2 may participate in the nonribosomal synthesis of small biologically active peptides.
KeywordMeSH Terms
Transcription Factors
131.     ( 1993 )

The TonB protein of Yersinia enterocolitica and its interactions with TonB-box proteins.

Molecular & general genetics : MGG 237 (1��2��)
PMID : 8384290  :   DOI  :   10.1007/bf00282796    
Abstract >>
The tonB gene is required for energy-dependent transport processes across the outer membrane of gram-negative bacteria. Using the antibiotics albomycin and ferrimycin, a tonB mutant of Yersinia enterocolitica was isolated. Comparison of the tonB mutant with the parent strain revealed that in Y. enterocolitica the uptake of ferrioxamine, ferrichrome, pesticin and heme is TonB-dependent. The tonB gene from Y. enterocolitica was sequenced and found to be similar to those of other Enterobacteria. The Y. enterocolitica tonB gene complemented a Y. enterocolitica tonB mutant. In contrast, some TonB functions of an Escherichia coli tonB mutant were not restored by the tonB gene of Y. enterocolitica. The observed differences in the ability to complement E. coli TonB functions correlated with the degree to which the TonB boxes of the receptors and colicins differed from the TonB box consensus sequence. Furthermore, the N-terminal membrane anchor of the TonB proteins and the TolA protein are likely to form an alpha-helix with an identical sequence motif (SHLS) located at one face of the alpha-helix, suggesting this region to be involved in the functional cross-talk between the TonB-ExbBD- and TolABQR-dependent transport systems across the outer membrane.
KeywordMeSH Terms
Escherichia coli Proteins
132.     ( N/A )

Cloning and nucleotide sequence analysis of immunodominant heat-shock protein of Yersinia enterocolitica.

Research in microbiology 144 (9)
PMID : 8190995  :  
Abstract >>
Yersinia enterocolitica, a highly antigenic 60-kDa protein, designated cross-reacting protein antigen (CRPA), is a member of the chaperonin-60 family of molecular chaperones. The gene encoding CRPA was cloned, expressed and sequenced. A partial library from Y. enterocolitica 0:3 genomic DNA was constructed in vector pUC19 and was screened by the immunoreactivity to monoclonal antibody 1A4, which has specificity for a species-specific epitope on the CRPA molecule. The crpA gene region consists of a putative two-cistron operon encoding proteins of 549 and 97 amino acids. The operon structure was led by a consensus heat-shock promoter sequence. Homology searches using the derived amino acid sequence have revealed that CRPA is 88.2% identical to GroEL of Escherichia coli. CRPB, another protein encoded by the operon, shows extensive sequence homology, 91.8% identical to GroES of E. coli which is a member of chaperonin-10.
KeywordMeSH Terms
133.     ( 1994 )

The pesticin receptor of Yersinia enterocolitica: a novel virulence factor with dual function.

Molecular microbiology 13 (2)
PMID : 7984105  :   DOI  :   10.1111/j.1365-2958.1994.tb00420.x    
Abstract >>
The iron-repressible outer membrane protein FyuA of Yersinia enterocolitica operates as a receptor with dual function: (i) as a receptor for the Y. pestis bacteriocin pesticin, and (ii) as a receptor for yersiniabactin, a siderophore that is produced by mouse-virulent Y. enterocolitica strains of biogroup IB. Cloning of the FyuA-encoding gene was achieved by mobilization of a genomic cosmid library of the pesticin-sensitive and mouse-virulent Y. enterocolitica O:8 strain WA into the pesticin-resistant WA fyuA mutant and subsequent in vivo selection of transconjugants for the ability to survive and multiply in mice (phenotype mouse virulence). The reisolated transconjugants which survived in mice for 3 d harboured a unique cosmid and phenotypically were pesticin sensitive. From this cosmid a 2650 bp SalI-PstI fragment conferring pesticin sensitivity was subcloned. Sequencing of this DNA fragment revealed a single open reading frame of 2022 bp, which encodes a deduced polypeptide of 673 amino acids with a predicted molecular mass of 73,677 Da. Cleavage of a putative signal sequence composed of 22 amino acids should lead to a mature protein of 651 amino acids with a molecular mass of 71,368 Da. The open reading frame is preceded by a sequence which shares homology with the postulated consensus Fur iron-repressor protein-binding site. FyuA shows homology to other iron-regulated TonB-dependent outer membrane proteins with receptor functions (e.g. BtuB, CirA, FepA, IutA, FhuA, FoxA, FcuA). On the basis of multiple alignment of amino acid sequences of FyuA and other TonB-dependent receptors, a phylogenetic tree was constructed, demonstrating that FyuA probably belongs to the citrate subfamily or represents a new subfamily of TonB-dependent receptors. Moreover, by complementation of the WA fyuA mutant by the cloned fyuA gene, yersiniabactin uptake and mouse virulence were restored. These studies demonstrate that the cloned pesticin/yersiniabactin receptor FyuA of Y. enterocolitica has the typical features of iron-regulated TonB-dependent outer membrane receptors for siderophores and bacteriocins and is required for mouse virulence.
KeywordMeSH Terms
Bacterial Proteins
134.     ( 1994 )

YscU, a Yersinia enterocolitica inner membrane protein involved in Yop secretion.

Journal of bacteriology 176 (15)
PMID : 8045883  :   DOI  :   10.1128/jb.176.15.4534-4542.1994     PMC  :   PMC196272    
Abstract >>
Pathogenic yersiniae secrete antihost Yop proteins by a recently discovered secretion pathway which is also encountered in several animal and plant pathogens. The components of the export machinery are encoded by the virA (lcrA), virB (lcrB), and virC (lcrC) loci of the 70-kb pYV plasmid. In the present paper we describe yscU, the last gene of the virB locus. We determined the DNA sequence and mutated the gene on the pYV plasmid. After inactivation of yscU, the mutant strain was unable to secrete Yop proteins. The topology of YscU was investigated by the analysis of YscU-PhoA translational fusions generated by TnphoA transposition. This showed that the 40.3-kDa yscU product contains four transmembrane segments anchoring a large cytoplasmic carboxyl-terminal domain to the inner membrane. YscU is related to Spa40 from Shigella flexneri, to SpaS from Salmonella typhimurium, to FlhB from Bacillus subtilis, and to HrpN from Pseudomonas solanacearum.
KeywordMeSH Terms
Membrane Proteins
Virulence Factors
135.     ( 1994 )

YscN, the putative energizer of the Yersinia Yop secretion machinery.

Journal of bacteriology 176 (6)
PMID : 8132449  :   DOI  :   10.1128/jb.176.6.1561-1569.1994     PMC  :   PMC205240    
Abstract >>
Pathogenic yersiniae secrete a set of 11 antihost proteins called Yops. Yop secretion appears as the archetype of the type III secretion pathway. Several components of this machinery are encoded by the virA (lcrA) and virC (lcrC) loci of the 70-kb pYV plasmid. In this paper, we describe yscN, another gene involved in this pathway. It is the first gene of the virB locus. It encodes a 47.8-kDa protein similar to the catalytic subunits of F0F1 and related ATPases, as well as to products of other genes presumed to be involved in a type III secretion pathway. YscN contains the two consensus nucleotide-binding motifs (boxes A and B) described by Walker et al. (J. E. Walker, M. Saraste, M. J. Runswick, and N. J. Gay, EMBO J. 1:945-951, 1982). We engineered a pYV mutant encoding a modified YscN protein lacking box A. This mutant, impaired in Yop secretion, can be complemented in trans by a cloned yscN gene. We conclude that YscN is a component of the Yop secretion machinery using ATP. We hypothesize that it is either the energizer of this machinery or a part of it.
KeywordMeSH Terms
Adenosine Triphosphatases
136.     ( 1993 )

The Myf fibrillae of Yersinia enterocolitica.

Molecular microbiology 9 (3)
PMID : 8105362  :   DOI  :   10.1111/j.1365-2958.1993.tb01712.x    
Abstract >>
The Myf antigen produced by Yersinia enterocolitica appeared as a proteic polymer composed of 21 kDa subunits. By transposon mutagenesis we isolated Myf-defective mutants. Those allowed us to clone and sequence a 4.4 kb chromosomal locus involved in Myf production. This region was found to contain three genes that we called myfA, myfB and myfC. Genes myfB and myfC encode an assembly machine related to those involved in the synthesis of many fimbriae: MyfB, the putative chaperone, possesses the consensus residues of the PapD family and myfC encodes a putative outer-membrane protein. MyfA, the major subunit, was found to be 44% identical to the pH 6 antigen of Y. pestis. Myf is thus the Y. enterocolitica counterpart of this antigen, but it is by far not so well conserved as the other virulence determinants such as the Yops, suggesting that Myf and pH 6 antigen do not necessarily play the same role in Y. enterocolitica and Y. pestis. The study of the prevalence of myfA in various species of Yersinia revealed that, like the yst enterotoxin gene, its presence is restricted to the pathogenic serotypes of Y. enterocolitica. By immunogold labelling, Myf appeared as a layer of extracellular material extending locally 2 microns from the bacterial surface, indicative of a fibrillar structure.
KeywordMeSH Terms
Fimbriae, Bacterial
137.     ( 1993 )

The putative arthritogenic cationic 19-kilodalton antigen of Yersinia enterocolitica is a urease beta-subunit.

Infection and immunity 61 (6)
PMID : 8500886  :   PMC  :   PMC280875    
Abstract >>
The gene coding for a putative 19-kDa arthritogenic antigen of Yersinia enterocolitica O:3 (A. K. H. Mertz et al., J. Clin. Invest. 87:632-642, 1991) was cloned and sequenced after amplification of part of the gene by the polymerase chain reaction using degenerate primers, inferred from the amino acid sequence. The deduced amino acid sequence of the antigen showed similarity to small subunits of ureases from several different organisms, including the jack bean urease. Screening of a genomic library of Y. enterocolitica O:3 with a 19-kDa-antigen-specific DNA probe allowed recombinant clones containing the entire urease operon to be obtained. These clones expressed urease activity in Escherichia coli.
KeywordMeSH Terms
138.     ( 1994 )

Crystal structure of Yersinia protein tyrosine phosphatase at 2.5 A and the complex with tungstate.

Nature 370 (6490)
PMID : 8052312  :   DOI  :   10.1038/370571a0    
Abstract >>
Protein tyrosine phosphatases (PTPases) and kinases coregulate the critical levels of phosphorylation necessary for intracellular signalling, cell growth and differentiation. Yersinia, the causative bacteria of the bubonic plague and other enteric diseases, secrete an active PTPase, Yop51, that enters and suppresses host immune cells. Though the catalytic domain is only approximately 20% identical to human PTP1B, the Yersinia PTPase contains all of the invariant residues present in eukaryotic PTPases, including the nucleophilic Cys 403 which forms a phosphocysteine intermediate during catalysis. We present here structures of the unliganded (2.5 A resolution) and tungstate-bound (2.6 A) crystal forms which reveal that Cys 403 is positioned at the centre of a distinctive phosphate-binding loop. This loop is at the hub of several hydrogen-bond arrays that not only stabilize a bound oxyanion, but may activate Cys 403 as a reactive thiolate. Binding of tungstate triggers a conformational change that traps the oxyanion and swings Asp 356, an important catalytic residue, by approximately 6 A into the active site. The same anion-binding loop in PTPases is also found in the enzyme rhodanese.
KeywordMeSH Terms
139.     ( 1994 )

Characterisation of the urease-encoding gene complex of Yersinia enterocolitica.

Gene 145 (1)
PMID : 8045421  :   DOI  :   10.1016/0378-1119(94)90318-2    
Abstract >>
A cosmid gene library of chromosomal DNA from Yersinia enterocolitica A2635 (serogroup O:8) was constructed in Escherichia coli. Subcloning of a urease-positive (Ure+) clone revealed a region of 6.6 kb that was sufficient for expression of Ure activity in E. coli. Sequencing of this fragment disclosed seven ORFs transcribed in the same direction. On the basis of homology to known Ure, these were designated ureA, ureB, ureC, ureE, ureF, ureG and ureD, which are predicted to encode polypeptides of 11.1, 17.9, 61.0, 29.5, 25.0, 24.1 and 36.4 kDa, respectively. The polypeptides encoded by the ure gene complex of Y. enterocolitica are significantly divergent from those encoded by the ure operons of other Enterobacteriaceae, which appear to be closely related to each other. This suggests that the ure genes were acquired by Y. enterocolitica from an unrelated organism or alternatively, that they diverged from those of other Enterobacteriaceae some considerable time ago.
KeywordMeSH Terms
140.     ( 1993 )

SycE, a chaperone-like protein of Yersinia enterocolitica involved in Ohe secretion of YopE.

Molecular microbiology 8 (1)
PMID : 8497188  :   DOI  :   10.1111/j.1365-2958.1993.tb01209.x    
Abstract >>
Pathogenic yersiniae secrete a set of 11 anti-host proteins called Yops. The yop genes, scattered around the pYV plasmid, constitute a thermoinduced regulon controlled by the product of virF gene. The secretion of the Yops also requires the presence of the products of the other vir genes and operons, namely virA, virB and virC. The large virC operon and presumably some genes of the virA region encode a new secretion system. Mutations in any of these vir genes impair the production of all the Yops. In contrast, mutations in the yerA locus, located close to yopE, specifically abolish the expression of the cytotoxin YopE. We describe here the counterpart of yerA in Yersinia enterocolitica W22703. We demonstrate that the gene product of yerA regulates the production of YopE at a post-transcriptional level. It specifically binds the YopE protein. We consider that it acts as a specific chaperone and we call it SycE (for specific YopE chaperone). We hypothesize that SycE is a link between translation and the specific Yop export machinery. It is the first representative of a new family of pYV-encoded proteins.
KeywordMeSH Terms
141. Mikulskis  AV, Delor  I, Thi  VH, Cornelis  GR,     ( 1994 )

Regulation of the Yersinia enterocolitica enterotoxin Yst gene. Influence of growth phase, temperature, osmolarity, pH and bacterial host factors.

Molecular microbiology 14 (5)
PMID : 7715452  :   DOI  :   10.1111/j.1365-2958.1994.tb01326.x    
Abstract >>
The chromosome of Yersinia enterocolitica encodes an enterotoxin called Yst. We analysed transcription of chromosomal yst'--luxAB and plasmid-borne yst'--lacZ operon fusions and we observed that regulation of yst expression occurs at transcriptional level. In a wild-type strain, yst was transcribed from at least two major promoters. yst transcription reached a maximum at the entry to the stationary phase and significantly varied in different Y. enterocolitica strains. In some strains, it gradually decreased during the course of our work, suggesting the existence of a mechanism switching the expression of yst to a silent state. Changes in the status of bacterial host factors rather than modifications in the yst gene are responsible for this silencing. Negative regulator YmoA participates in yst silencing and temperature regulation of yst. YmoA was also required for proper growth-phase regulation of yst, although it is not the only factor involved in this regulation. Physico-chemical parameters of the environment play an important role in yst transcription. In usual culture media (e.g. tryptic soy broth), the enterotoxin gene was transcribed only at temperatures below 30 degrees C, which argued against the role of Yst in a prolonged diarrhoea at body temperatures. However, yst transcription could be induced at 37 degrees C by increasing osmolarity and pH to the values normally present in the ileum lumen. This finding reconciles the observations concerning yst expression in a host environment and in bacterial cultures, thus supporting the idea that enterotoxin Yst is a virulence factor of Y. enterocolitica.
KeywordMeSH Terms
Gene Expression Regulation, Bacterial
Genes, Bacterial
142. Iriarte  M, Cornelis  GR,     ( 1995 )

MyfF, an element of the network regulating the synthesis of fibrillae in Yersinia enterocolitica.

Journal of bacteriology 177 (3)
PMID : 7836309  :   DOI  :   10.1128/jb.177.3.738-744.1995     PMC  :   PMC176651    
Abstract >>
The Yersinia enterocolitica surface antigen Myf is a fibrillar structure that resembles CS3 fimbriae. Gene myfA encodes the 21-kDa major subunit of the antigen, while genes myfB and myfC are required for the transport and assembly of pilin subunits at the bacterial cell surface. Here we show that the expression of Myf is regulated at the transcriptional level by temperature and pH. Gene myfA is transcribed at 37 degrees C and in acidic medium. The transcription start is preceded by a putative -10 box for the vegetative RNA polymerase as well as by sequences resembling the consensus sequence recognized by sigma 28. Thus, myfA could be transcribed either from a classical sigma 70 promoter or from a sigma 28 promoter. Transcription of myfA requires at least two genes, myfF and myfE, situated immediately upstream from myfA. The myfF product does not show similarity to any known regulatory protein. It is an 18.5-kDa protein with no typical helix-turn-helix motif and a unique hydrophobic domain in the NH2-terminal part. T7 expression, osmotic shock, fractionation experiments, and TnphoA fusion analyses carried out in Escherichia coli suggest that MyfF is associated with the inner membrane by means of its hydrophobic domain whereas the hydrophilic part protrudes in the periplasm. These features strikingly evoke ToxS, a protein involved in regulation of Tcp pilus production in Vibrio cholerae. MyfE resembles PsaE, a protein involved in regulation of pH6 antigen in Yersinia pestis. Genes myfF and myfE are presumably part of a whole regulatory network. MyfF could be an element of the signal transducing system.
KeywordMeSH Terms
Fimbriae, Bacterial
Transcription, Genetic
143. Yoshino  K, Takao  T, Huang  X, Murata  H, Nakao  H, Takeda  T, Shimonishi  Y,     ( 1995 )

Characterization of a highly toxic, large molecular size heat-stable enterotoxin produced by a clinical isolate of Yersinia enterocolitica.

FEBS letters 362 (3)
PMID : 7729521  :   DOI  :   10.1016/0014-5793(95)00267-d    
Abstract >>
A novel heat-stable enterotoxin (ST) designated as Y-STc was purified to homogeneity from the culture supernatant of a pathogenic strain of Yersinia enterocolitica serotype O3 and its amino acid sequence was determined. The mature Y-STc was found to consist of 53 amino acid residues, which includes the putative pro-sequence. The molecular weight of Y-STc was 5638 and constituted the largest molecular size in the family of currently known STs. The minimum effective dose of purified Y-STc in the suckling mouse assay was 0.6 ng (0.0 pmol), indicating that, despite the long sequence, Y-STc is the most toxic in the ST family.
KeywordMeSH Terms
Bacterial Toxins
Enterotoxins
144. Osbourn  SE, Turner  AK, Grinsted  J,     ( 1995 )

Nucleotide sequence within Tn3926 confirms this as a Tn21-like transposable element and provides evidence for the origin of the mer operon carried by plasmid pKLH2.

Plasmid 33 (1)
PMID : 7753910  :   DOI  :   10.1006/plas.1995.1008    
Abstract >>
The DNA sequence of the resolvase gene, resolution sites, and the region between the transposition functions and the end of the mercury resistance operon of the bacterial transposon, Tn3926, is presented. The sequence of Tn3926 upstream of the resolution sites is homologous to that bordering the 11.2-kb insert of Tn21, supporting the idea that this insert transposed into a progenitor of Tn3926. This region of Tn3926 also shows 97.0% identity to the mercury-resistant determinant of the plasmid, pKLH2, suggesting that this plasmid once harbored a close relative of Tn3926. It is proposed that the mercury resistance operon of Tn3926 will have a structure very similar to that found on pKLH2.
KeywordMeSH Terms
DNA Transposable Elements
Operon
Plasmids
Transposon Resolvases
145.     ( 1993 )

Hydrophobic domains affect the collagen-binding specificity and surface polymerization as well as the virulence potential of the YadA protein of Yersinia enterocolitica.

Molecular microbiology 10 (5)
PMID : 7934875  :   DOI  :   10.1111/j.1365-2958.1993.tb00971.x    
Abstract >>
The YadA surface protein of enteropathogenic Yersinia species contains two highly hydrophobic regions: one close to the amino terminal, and the other at the carboxy-terminal end of the YadA polypeptide. To study the role of these hydrophobic regions, we constructed 66 bp deletion mutants of the yadA genes of Yersinia enterocolitica serotype O:3 strain 6471/76 (YeO3) and of O:8 strain 8081 (YeO8). The mutant proteins, YadAYeO3-delta 83-104 and YadAYeO8-delta 8O-101, lacked 22 amino acids from the amino-terminal hydrophobic region, formed fibrillae and were expressed on the cell surface. Bacteria expressing the mutated protein lost their auto-agglutination potential as well as their collagen-binding property. Binding to fibronectin and laminin was affected differently in the YeO3 and the YeO8 constructs. The deletion did not influence YadA-mediated complement inhibition. Loss of the collagen-binding property was associated with loss of virulence in mice. We also constructed a number of YadAYeO3 deletion mutants lacking the hydrophobic carboxy-terminal end of the protein. Deletions ranging from 19 to 79 amino acids from the carboxy terminus affected polymerization of the YadA subunits, and also resulted in the loss of the YadA expression on the cell surface. This suggests that the carboxy terminus of YadA is involved in transport of the protein to the bacterial outer surface.
KeywordMeSH Terms
Adhesins, Bacterial
146. Iriarte  M, Stainier  I, Cornelis  GR,     ( 1995 )

The rpoS gene from Yersinia enterocolitica and its influence on expression of virulence factors.

Infection and immunity 63 (5)
PMID : 7729893  :   PMC  :   PMC173233    
Abstract >>
The chromosome of Yersinia enterocolitica encodes a heat-stable enterotoxin called Yst and a surface antigen called Myf, which closely resembles enterotoxin-associated fimbriae. Both factors could act in conjunction to produce diarrhea. Production of the enterotoxin is regulated by temperature, osmolarity, and pH and occurs only when bacteria reach the stationary phase. Myf production is regulated by temperature and pH and, as we show in this work, also occurs after the exponential growth phase. In an attempt to understand the late-phase expression of yst and myf, we cloned, sequenced, and mutagenized the gene encoding RpoS, an alternative sigma factor of the RNA polymerase involved in expression of stationary-phase genes in other enterobacteria. An intact rpoS gene was necessary for full expression of yst in the stationary phase but not for the expression of myf and of pYV-encoded virulence determinants.
KeywordMeSH Terms
Antigens, Bacterial
Bacterial Outer Membrane Proteins
Gene Expression Regulation, Bacterial
147. Iriarte  M, Stainier  I, Mikulskis  AV, Cornelis  GR,     ( 1995 )

The fliA gene encoding sigma 28 in Yersinia enterocolitica.

Journal of bacteriology 177 (9)
PMID : 7730257  :   DOI  :   10.1128/jb.177.9.2299-2304.1995     PMC  :   PMC176884    
Abstract >>
Yersinia enterocolitica is an enterobacterium responsible for gastrointestinal syndromes. Its pathogenicity depends on the presence of the 70-kb pYV plasmid, which directs Yop secretion. The Yop secretion machinery, consisting of the YscA-U and LcrD proteins, presents some structural similarity with the flagellum assembly machinery characterized in other bacteria. Flagellum assembly requires sigma 28, an alternative sigma factor. The region upstream of the lcrD gene resembles promoters recognized by sigma 28, suggesting that the similarity between Yop secretion and flagellum assembly could extend to their regulation. The chromosome of Y. enterocolitica also contains pathogenicity determinants such as myfA, which encodes the Myf antigen subunit. The promoter region of myfA also resembles promoters recognized by sigma 28. In an attempt to clarify the role of sigma 28 in the expression of lcrD, myfA, and flagellar genes, we cloned, sequenced, and mutagenized the fliA gene encoding the sigma 28 homolog in Y. enterocolitica. As is the case in other bacteria, fliA was required for motility. However, it was involved neither in fibrilla synthesis nor in Yop secretion. The fliA mutant allowed us to monitor the role of motility in pathogenesis. At least in the mouse model, motility seemed not to be required for Y. enterocolitica pathogenesis.
KeywordMeSH Terms
Antigens, Bacterial
148. Rakin  A, Heesemann  J,     ( 1995 )

Virulence-associated fyuA/irp2 gene cluster of Yersinia enterocolitica biotype 1B carries a novel insertion sequence IS1328.

FEMS microbiology letters 129 (2��3��)
PMID : 7607411  :   DOI  :   10.1111/j.1574-6968.1995.tb07594.x    
Abstract >>
The fyuA/irp2 gene cluster, which is part of the Yersinia pestis pigmentation (pgm) locus encoding genes involved in iron uptake and virulence, is present in all pesticin-sensitive bacteria. In Y. enterocolitica biotype 1B strains (serotypes O8, O20, O21), the fyuA/irp2 gene cluster carries an insertion of a novel repetitive sequence, IS1328. It was also found in the genome of Y. enterocolitica O5 (biotype 1A) and O13 (biotype 1B), but not in pesticin-sensitive Y. pseudotuberculosis O1 and Escherichia coli Phi. The 1353-bp repetitive element has 12-bp perfect inverted terminal repeats. A single open reading frame is capable of encoding a 334-amino acid polypeptide. IS1328 DNA has high homology with the DNA sequences located downstream of the aggR gene from the enteroaggregative E. coli (EAggEC), to the region of the R751 plasmid flanking Tn501, to the sequence following the merR gene of S. marcescens pDU1358 plasmid, and to the sequences of K. pneumoniae plasmid pCFF04. The putative polypeptide has 36.4% identity with the transposase encoded by the Coxiella burnetii IS1111a insertion sequence. The IS1328 insertion sequence could be responsible for the deletions of the fyuA/irp2 gene cluster observed in Y. enterocolitica O8 and could represent a member of a new group of widely distributed repetitive elements.
KeywordMeSH Terms
149.     ( 1994 )

Individual chaperones required for Yop secretion by Yersinia.

Proceedings of the National Academy of Sciences of the United States of America 91 (22)
PMID : 7937981  :   DOI  :   10.1073/pnas.91.22.10493     PMC  :   PMC45047    
Abstract >>
Pathogenic yersiniae secrete anti-host proteins called Yops, by a recently discovered Sec-independent pathway. The Yops do not have a classical signal peptide at their N terminus and they are not processed during membrane translocation. The secretion domain is nevertheless contained in their N-terminal part but these domains do not resemble each other in the different Yops. We have previously shown that YopE secretion requires SycE, a 15-kDa acidic protein acting as a specific cytosolic chaperone. Here we show that the gene downstream from yopH encodes a 16-kDa acidic protein that binds to hybrid proteins made of the N-terminal part of YopH and either the bacterial alkaline phosphatase or the cholera toxin B subunit. Loss of this protein by mutagenesis led to accumulation of YopH in the cytoplasm and to a severe and selective reduction of YopH secretion. This protein thus behaves like the counterpart of SycE and we called it SycH. We also engineered a mutation in lcrH, the gene upstream from yopB and yopD, known to encode a 19-kDa acidic protein. Although this mutation was nonpolar, the mutant no longer secreted YopB and YopD. The product of lcrH could be immunoprecipitated together with cytoplasmic YopD. lcrH therefore seems to encode a YopD-specific chaperone, which we called SycD. Determination of the dependence of YopB on SycD requires further investigation. SycE, SycH, and SycD appear to be members of a new family of cytosolic chaperones required for Yop secretion.
KeywordMeSH Terms
Protein Tyrosine Phosphatases
150. Rakin  A, Urbitsch  P, Heesemann  J,     ( 1995 )

Evidence for two evolutionary lineages of highly pathogenic Yersinia species.

Journal of bacteriology 177 (9)
PMID : 7730256  :   DOI  :   10.1128/jb.177.9.2292-2298.1995     PMC  :   PMC176883    
Abstract >>
Sensitivity to Yersinia pestis bacteriocin pesticin correlates with the existence of two groups of human pathogenic yersiniae, mouse lethal and mouse nonlethal. The presence of the outer membrane pesticin receptor (FyuA) in mouse-lethal yersiniae is a prerequisite for pesticin sensitivity. Genes that code for FyuA (fyuA) were identified and sequenced from pesticin-sensitive bacteria, including Y. enterocolitica biotype 1B (serotypes O8; O13, O20, and O21), Y. pseudotuberculosis serotype O1, Y. pestis, two known pesticin-sensitive Escherichia coli isolates (E. coli Phi and E. coli CA42), and two newly discovered pesticin-sensitive isolates, E. coli K49 and K235. A 2,318-bp fyuA sequence was shown to be highly conserved in all pesticin-sensitive bacteria, including E. coli strains (DNA sequence homology was 98.5 to 99.9%). The same degree of DNA homology (97.8 to 100%) was established for the sequenced 276-bp fragment of the irp2 gene that encodes high-molecular-weight protein 2, which is also thought to be involved in the expression of virulence by Yersinia species. Highly conserved irp2 was also found in all pesticin-sensitive E. coli strains. On the basis of the fyuA and irp2 sequence homologies, two evolutionary groups of highly pathogenic Yersinia species can be established. One group includes Y. enterocolitica biotype 1B strains, while the second includes Y. pestis, Y. pseudotuberculosis serotype O1, and irp2-positive Y. pseudotuberculosis serotype O3 strains. E. coli Phi, CA42, K49, and K235 belong to the second group. The possible proximity of these two iron-regulated genes (fyuA and irp2), as well as their high levels of sequence conservation and similar G+C contents (56.2 and 59.8 mol%), leads to the assumption that these two genes may represent part of an unstable pathogenicity island that has been acquired by pesticin-sensitive bacteria as a result of a horizontal transfer.
KeywordMeSH Terms
Bacterial Proteins
Biological Evolution
151.     ( 1994 )

Transport of haemin across the cytoplasmic membrane through a haemin-specific periplasmic binding-protein-dependent transport system in Yersinia enterocolitica.

Molecular microbiology 13 (4)
PMID : 7997183  :   DOI  :   10.1111/j.1365-2958.1994.tb00465.x    
Abstract >>
The Yersinia enterocolitica O:8 periplasmic binding-protein-dependent transport (PBT) system for haemin was cloned and characterized. It consisted of four proteins: the periplasmic haemin-binding protein HemT, the haemin permease protein HemU, the ATP-binding hydrophilic protein HemV and the putative haemin-degrading protein HemS. Y. enterocolitica strains mutated in hemU or hemV genes were unable to use haemin as an iron source whereas those mutated in the hemT gene were able to use haemin as an iron source. As Escherichia coli strains expressing only the haemin outer membrane receptor protein HemR from Y. enterocolitica were capable of using haemin as an iron source the existence of an E. coli K-12 haemin-specific PBT system is postulated. The first gene in the Y. enterocolitica haemin-specific PBT system encoded a protein, HemS, which is probably involved in the degradation of haemin in the cytoplasm. The presence of the hemS gene was necessary to prevent haemin toxicity in E. coli strains that accumulate large amounts of haemin in the cytoplasm. We propose a model of haemin utilization in Y. enterocolitica in which HemT, HemU and HemV proteins transport haemin into the cytoplasm where it is degraded by HemS thereby liberating the iron.
KeywordMeSH Terms
Adenosine Triphosphatases
ATP-Binding Cassette Transporters
Escherichia coli Proteins
Oxidoreductases
Periplasmic Binding Proteins
152. De Koning-Ward  TF, Robins-Browne  RM,     ( 1995 )

Contribution of urease to acid tolerance in Yersinia enterocolitica.

Infection and immunity 63 (10)
PMID : 7558281  :   PMC  :   PMC173532    
Abstract >>
The stomach serves as a barrier to enteric infection because of the antibacterial effect of the hydrochloric acid in gastric juice. In this study, we tested the ability of the enteric pathogen Yersinia enterocolitica to tolerate a pH range of 2.0 to 6.0 and found that under the conditions of a normal human fasting stomach (pH < 3 and a gastric emptying time of 2 h), Y. enterocolitica is highly acid resistant, showing approximately 85% survival. The resistance of Y. enterocolitica to acid in vitro depended on the bacterial growth phase and the concentration of urea in the medium, being maximal during stationary phase in the presence of at least 0.3 mM urea. Urease-negative mutants of Y. enterocolitica were constructed by disrupting the urease gene complex of a virulent strain of serogroup O9. Compared with the wild type, these mutants showed an approximately 1,000-fold decrease in the ability to tolerate acid in vitro (< 0.08% survival) and a 10-fold reduction in viability after passage through the stomachs of mice. Complementation of the disrupted urease genes in trans restored the ability of urease-negative mutants to tolerate low pH in vitro and gastric acidity to approximately wild-type levels. These findings indicate that urease is responsible for acid resistance in Y. enterocolitica and suggest that urease contributes to the virulence of Y. enterocolitica by enhancing the likelihood of bacterial survival during passage through the stomach.
KeywordMeSH Terms
153. Throup  JP, Camara  M, Briggs  GS, Winson  MK, Chhabra  SR, Bycroft  BW, Williams  P, Stewart  GS,     ( 1995 )

Characterisation of the yenI/yenR locus from Yersinia enterocolitica mediating the synthesis of two N-acylhomoserine lactone signal molecules.

Molecular microbiology 17 (2)
PMID : 7494483  :   DOI  :   10.1111/j.1365-2958.1995.mmi_17020345.x    
Abstract >>
Yersinia enterocolitica produces compounds capable of transcriptionally activating the Photobacterium fischeri bioluminescence (lux) operon. Using high-performance liquid chromatography, high resolution tandem mass spectrometry in conjunction with chemical synthesis, two signal molecules were identified and shown to be N-hexanoyl-L-homoserine lactone (HHL) and N-(3-oxohexanoyl)-L-homoserine lactone (OHHL). A gene (yenI) was isolated from Y. enterocolitica and demonstrated to direct the synthesis of both HHL and OHHL. DNA sequence analysis revealed an open reading frame (ORF) of 642 bp encoding a protein (YenI) of 24.6 kDa with approximately 20% identity to the LuxI family of proteins. Northern blot analysis of yenI expression indicated yenI is transcribed as a single gene and 5' transcript mapping of yenI identified a transcriptional start site 89 bp upstream of the ORF. DNA sequence analysis of the region downstream of yenI located a second ORF, termed yenR, with significant homology to the LuxR family of transcriptional activators. An insertion mutation of yenI abolishes HHL and OHHL production, indicating its central role in N-acylhomoserine lactone synthesis in Y. enterocolitica. Transcriptional analysis using a chromosomal yenI::luxAB fusion has demonstrated that yenI is not subject to autoinduction but is expressed constitutively. Whilst production of the Yop proteins in the wild type and in yenI mutants is indistinguishable, two-dimensional SDS-PAGE analysis of total cell proteins indicated that a number of proteins lack the yenI mutant.
KeywordMeSH Terms
Repressor Proteins
154. Takao  T, Tominaga  N, Yoshimura  S, Shimonishi  Y, Hara  S, Inoue  T, Miyama  A,     ( 1985 )

Isolation, primary structure and synthesis of heat-stable enterotoxin produced by Yersinia enterocolitica.

European journal of biochemistry 152 (1)
PMID : 4043080  :   DOI  :   10.1111/j.1432-1033.1985.tb09183.x    
Abstract >>
Five heat-stable enterotoxins were isolated from the culture supernatant of Yersinia enterocolitica and purified to homogeneity by DEAE-Sephacel and high-performance liquid chromatographies. They caused acute fluid accumulation in the intestine of suckling mice. The amino acid sequence of one of the enterotoxins was determined to be Ser-Ser-Asp-Trp-Asp-Cys-Cys-Asp-Val-Cys-Cys-Asn-Pro-Ala-Cys-Ala-Gly-Cys, by Edman degradation of its pyridylethylated derivative and a combination of fast atom bombardment mass spectrometry and carboxypeptidase B digestion. This structure was unambiguously confirmed by chemical synthesis. The other enterotoxins had longer or shorter amino acid sequences at their N termini, but the same sequence at their C termini. The six half-cystine residues formed intramolecular disulfide linkages, as shown by measurement of the molecular masses of the enterotoxins by fast atom bombardment mass spectrometry. The sequence of 13 amino acid residues at the C terminus showed similarity to those of heat-stable enterotoxins isolated from enterotoxigenic Escherichia coli [Aimoto, S. et al. (1982) Eur. J. Biochem. 129, 257-263; Takao, T. et al. (1983) FEBS Lett. 152, 1-5] suggesting that these similar sequences are related to the common biological and immunological properties of enterotoxins produced by Y. enterocolitica and enterotoxigenic E. coli.
KeywordMeSH Terms
Bacterial Toxins
Enterotoxins
Yersinia enterocolitica
155. Lång  H, Ferenci  T,     ( 1995 )

Sequence alignment and structural modelling of the LamB glycoporin family.

Biochemical and biophysical research communications 208 (3)
PMID : 7702622  :   DOI  :   10.1006/bbrc.1995.1423    
Abstract >>
lamB gene segments were obtained from Yersinia enterocolitica and Vibrio parahaemolyticus by the PCR and the DNA sequence determined. The deduced polypeptide sequences showed high similarity to six other LamB-related proteins and all contained typical signature sequences present in all members of the family but not other proteins. The aligned amino acid sequences permitted derivation of a model of LamB folding across the bacterial outer membrane using an approach successfully applied in the identification of structural features in other porins (Ferenci,T. (1994) Mol. Microbiol. 14:188-189). The alignment-based model differs from previous LamB structure predictions and is also more complex than that found for OmpF-related porins; more than 16 conserved stretches of amino acid sequence potentially corresponded to membrane-spanning segments.
KeywordMeSH Terms
Protein Folding
Protein Structure, Secondary
156. Yamato  M, Takahashi  Y, Tomotake  H, Ota  F, Hirota  K, Yamaguchi  K,     ( 1994 )

Monoclonal antibodies to spirosin of Yersinia enterocolitica and analysis of the localization of spirosome by use of them.

Microbiology and immunology 38 (3)
PMID : 7521508  :   DOI  :   10.1111/j.1348-0421.1994.tb01762.x    
Abstract >>
Two hybridomas producing monoclonal antibodies (MAbs) were prepared by fusing myeloma cells (Sp2/0-Ag14) with mouse spleen cells immunized with purified spirosin from Yersinia enterocolitica SYT-11-72 (YE72). The antibodies produced by them were designated MAbs-S5 and S27. They were IgG2a and IgG1, respectively, both with kappa light chains. MAbs-S5 and S27 reacted specifically with spirosin from YE72. On Western blotting after limited proteolysis with Staphylococcus aureus V8 protease, YE72 spirosin revealed peptide fragments of 35 and 37 kDa reacting markedly with MAb-S5, which suggested the presence of an antigenic determinant on these fragments. By cellular fractionation of YE72 and subsequent EIA and Western blot analysis, spirosome was shown to be present in the cytoplasm of YE72.
KeywordMeSH Terms
157. Pepe  JC, Badger  JL, Miller  VL,     ( 1994 )

Growth phase and low pH affect the thermal regulation of the Yersinia enterocolitica inv gene.

Molecular microbiology 11 (1)
PMID : 7511772  :   DOI  :   10.1111/j.1365-2958.1994.tb00295.x    
Abstract >>
The inv gene encodes the protein invasin, which is the primary invasion factor for Yersinia enterocolitica in vitro and in vivo. Previous studies of Yersinia species have shown that inv expression and entry into mammalian cells are temperature regulated. Invasin production is reduced at the host temperature of 37 degrees C as compared to production at ambient temperature; consequently, this study was initiated to determine whether other host environmental signals might induce inv expression at 37 degrees C. An inv::phoA translational fusion was recombined on to the Y. enterocolitica chromosome by allelic exchange to monitor inv expression. Molecular characterization of expression of the wild-type inv gene and the inv::phoA fusion showed that invasin is not produced until early stationary phase in bacteria grown at 23 degrees C. Y. enterocolitica grown at 37 degrees C and pH 5.5 showed levels of inv expression comparable to those observed in bacteria grown at 23 degrees C. An increase in Na+ ions caused a slight increase in expression at 37 degrees C. However, expression at 37 degrees C was unaffected by anaerobiosis, growth medium, calcium levels, or iron levels. Additionally, Y. enterocolitica expressed invasin in Peyer's patches two days after being introduced intragastrically into BALB/c mice. These results suggest that invasin expression in Y. enterocolitica may remain elevated early during interaction with the intestinal epithelium, a site at which invasin was shown to be necessary.
KeywordMeSH Terms
Adhesins, Bacterial
158. Allaoui  A, Scheen  R, Lambert de Rouvroit  C, Cornelis  GR,     ( 1995 )

VirG, a Yersinia enterocolitica lipoprotein involved in Ca2+ dependency, is related to exsB of Pseudomonas aeruginosa.

Journal of bacteriology 177 (15)
PMID : 7635810  :   DOI  :   10.1128/jb.177.15.4230-4237.1995     PMC  :   PMC177167    
Abstract >>
Pathogenic yersiniae require Ca2+ for growth at 37 degrees C. They harbor closely related plasmids of about 70 kb that are essential for virulence. At 37 degrees C and in the absence of Ca2+ ions, these plasmids cause a decrease in growth rate and the release of large amounts of proteins called Yops. Here we describe the virG gene of Yersinia enterocolitica; virG is located just upstream of the virF gene, which encodes the transcriptional activator of some plasmid virulence factors. Analysis of the VirG amino acid sequence suggested that virG encodes a lipoprotein, which was confirmed by [3H]palmitate labeling of VirG-PhoA fusion proteins. A nonpolar virG mutant was constructed and found to be Ca2+ independent for growth at 37 degrees C but to still secrete Yops. This phenotype was complemented by the introduction of a plasmid harboring an intact virG gene. VirG was found to be homologous to ExsB, a protein encoded by a Pseudomonas aeruginosa gene located in the locus controlling exoenzyme S synthesis. Interestingly, the exsA gene, located just downstream of exsB, is also homologous to virF.
KeywordMeSH Terms
159. Zhang  L, al-Hendy  A, Toivanen  P, Skurnik  M,     ( 1993 )

Genetic organization and sequence of the rfb gene cluster of Yersinia enterocolitica serotype O:3: similarities to the dTDP-L-rhamnose biosynthesis pathway of Salmonella and to the bacterial polysaccharide transport systems.

Molecular microbiology 9 (2)
PMID : 7692217  :   DOI  :   10.1111/j.1365-2958.1993.tb01692.x    
Abstract >>
The Yersinia enterocolitica O:3 lipopolysaccharide O-antigen is a homopolymer of 6-deoxy-L-altrose. The cloned rfb region was sequenced, and 10 open reading frames were identified. Transposon mutagenesis, deletion analysis and transcomplementation experiments showed that eight of the genes, organized into two operons, rfbABC and rfbDEFGH, are essential for O-antigen synthesis. Functional tandem promoters were identified upstream of both operons. Of the deduced polypeptides RfbA, RfbF and RfbG were similar to Salmonella proteins involved in the dTDP-L-rhamnose biosynthesis. Rhamnose and 6-deoxy-L-altrose are C3-epimers suggesting that analogous pathways function in their biosynthesis. RfbD and RfbE were similar to capsular polysaccharide export proteins, e.g. KpsM and KpsT of Escherichia coli. This and transposon mutagenesis showed that RfbD and RfbE function as O-antigen exporters.
KeywordMeSH Terms
Genes, Bacterial
160. Badger  JL, Miller  VL,     ( 1995 )

Role of RpoS in survival of Yersinia enterocolitica to a variety of environmental stresses.

Journal of bacteriology 177 (18)
PMID : 7665530  :   DOI  :   10.1128/jb.177.18.5370-5373.1995     PMC  :   PMC177337    
Abstract >>
rpoS, a gene that encodes an alternative sigma factor (also known as katF), is critical for the ability of Yersinia enterocolitica grown at 37 degrees C, but not at 26 degrees C, to survive diverse environmental insults such as high temperature, hydrogen peroxide, osmolarity, and low pH. However, a Y. enterocolitica rpoS mutant was not affected in expression of inv or ail, invasion of tissue culture cells, or virulence in mice.
KeywordMeSH Terms
Adhesins, Bacterial
161. Michiels  T, Cornelis  G,     ( 1988 )

Nucleotide sequence and transcription analysis of yop51 from Yersinia enterocolitica W22703.

Microbial pathogenesis 5 (6)
PMID : 3244311  :  
Abstract >>
Virulent strains of Yersinia enterocolitica, pseudotuberculosis and pestis secrete large amounts of plasmid-encoded proteins involved in virulence and called Yops. A 2 kb fragment of the pYVe227 plasmid from Yersinia enterocolitica 0:9 encoding Yop51 was sequenced. Gene yop51 was found to encode a 50,882 Da protein consisting of 468 amino acids. This protein shows 99% identity with Yop2b, its counterpart from Y. pseudotuberculosis YPIII (pIB1), confirming that the virulence machinery is highly conserved among Yersinia spp. The homology stops abruptly 240 bp upstream and 175 bp downstream of the structural yop51 gene suggesting that all the sequences involved in the regulation of yop51 are located within the conserved region and confirming that the homology between the plasmids of Yersinia enterocolitica and Yersinia pseudotuberculosis is made up of boxes of high homology. Gene yop51 is only transcribed at 37 degrees C from a VirF-regulated promoter. This promoter was tentatively identified by determining the messenger transcriptional startpoint. The putative yop51 promoter resembles E. coli promoters despite the fact that it is not active in that species in the absence of VirF. A transcription terminator was found at the end of the gene while a second terminator was detected within the structural gene leading to premature termination of some of the messenger molecules.
KeywordMeSH Terms
Bacterial Outer Membrane Proteins
Genes
Genes, Bacterial
Protein Tyrosine Phosphatases
Transcription, Genetic
162. Richaud  F, Richaud  C, Ratet  P, Patte  JC,     ( 1986 )

Chromosomal location and nucleotide sequence of the Escherichia coli dapA gene.

Journal of bacteriology 166 (1)
PMID : 3514578  :   DOI  :   10.1128/jb.166.1.297-300.1986     PMC  :   PMC214591    
Abstract >>
In Escherichia coli, the first enzyme of the diaminopimelate and lysine pathway is dihydrodipicolinate synthetase, which is feedback-inhibited by lysine and encoded by the dapA gene. The location of the dapA gene on the bacterial chromosome has been determined accurately with respect to the neighboring purC and dapE genes. The complete nucleotide sequence and the transcriptional start of the dapA gene were determined. The results show that dapA consists of a single cistron encoding a 292-amino acid polypeptide of 31,372 daltons.
KeywordMeSH Terms
Chromosome Mapping
Genes, Bacterial
163. Bräuning  B, Bertosin  E, Praetorius  F, Ihling  C, Schatt  A, Adler  A, Richter  K, Sinz  A, Dietz  H, Groll  M,     ( 2018 )

Structure and mechanism of the two-component �\-helical pore-forming toxin YaxAB.

Nature communications 9 (1)
PMID : 29728606  :   DOI  :   10.1038/s41467-018-04139-2     PMC  :   PMC5935710    
Abstract >>
Pore-forming toxins (PFT) are virulence factors that transform from soluble to membrane-bound states. The Yersinia YaxAB system represents a family of binary �\-PFTs with orthologues in human, insect, and plant pathogens, with unknown structures. YaxAB was shown to be cytotoxic and likely involved in pathogenesis, though the molecular basis for its two-component lytic mechanism remains elusive. Here, we present crystal structures of YaxA and YaxB, together with a cryo-electron microscopy map of the YaxAB complex. Our structures reveal a pore predominantly composed of decamers of YaxA-YaxB heterodimers. Both subunits bear membrane-active moieties, but only YaxA is capable of binding to membranes by itself. YaxB can subsequently be recruited to membrane-associated YaxA and induced to present its lytic transmembrane helices. Pore formation can progress by further oligomerization of YaxA-YaxB dimers. Our results allow for a comparison between pore assemblies belonging to the wider ClyA-like family of �\-PFTs, highlighting diverse pore architectures.
KeywordMeSH Terms
Protein Conformation, alpha-Helical
Protein Multimerization
164.     ( 2012 )

Identification and distribution of putative virulence genes in clinical strains of Yersinia enterocolitica biovar 1A by suppression subtractive hybridization.

Journal of applied microbiology 113 (5)
PMID : 22897337  :   DOI  :   10.1111/j.1365-2672.2012.05427.x    
Abstract >>
To detect putative virulence genes in clinical strains of Yersinia enterocolitica biovar 1A by suppression subtractive hybridization between two closely related strains of clinical and nonclinical origin having the same serotype (O:6,30-6,31). Suppression Subtractive Hybridization (SSH) was used to identify genomic differences between clinical (serotype O:6,30-6,31, from diarrhoeic human stools) and nonclinical (serotype O:6,30-6,31, from wastewater) strains of Y. enterocolitica biovar 1A. Following genomic subtraction and DNA sequencing, nine DNA sequences that were present only in clinical biovar 1A strains were identified. The sequences identified using SSH showed similarity to conserved hypothetical proteins, proteins related to iron acquisition and haemin storage, type 1 secretion proteins, flagellar hook proteins, exported protein and ABC transport system. All these sequences showed high similarity with Y. enterocolitica 8081 (biovar 1B). The distribution of these genes was further analysed using PCR in 26 clinical strains of Y. enterocolitica biovar 1A. The results revealed that the distribution of these genes was not uniform. Genes related to iron acquisition and storage, and flagellar proteins might be responsible for virulence of some of the clinical strains of Y. enterocolitica biovar 1A. Genes identified in this study might be useful in understanding the pathogenic potential of clinical strains of Y. enterocolitica biovar 1A.
KeywordMeSH Terms
165.     ( 2012 )

Clinical isolates of Yersinia enterocolitica biotype 1A represent two phylogenetic lineages with differing pathogenicity-related properties.

BMC microbiology 12 (N/A)
PMID : 22985268  :   DOI  :   10.1186/1471-2180-12-208     PMC  :   PMC3512526    
Abstract >>
Y. enterocolitica biotype (BT) 1A strains are often isolated from human clinical samples but their contribution to disease has remained a controversial topic. Variation and the population structure among the clinical Y. enterocolitica BT 1A isolates have been poorly characterized. We used multi-locus sequence typing (MLST), 16S rRNA gene sequencing, PCR for ystA and ystB, lipopolysaccharide analysis, phage typing, human serum complement killing assay and analysis of the symptoms of the patients to characterize 298 clinical Y. enterocolitica BT 1A isolates in order to evaluate their relatedness and pathogenic potential. A subset of 71 BT 1A strains, selected based on their varying LPS patterns, were subjected to detailed genetic analyses. The MLST on seven house-keeping genes (adk, argA, aroA, glnA, gyrB, thrA, trpE) conducted on 43 of the strains discriminated them into 39 MLST-types. By Bayesian analysis of the population structure (BAPS) the strains clustered conclusively into two distinct lineages, i.e. Genetic groups 1 and 2. The strains of Genetic group 1 were more closely related (97% similarity) to the pathogenic bio/serotype 4/O:3 strains than Genetic group 2 strains (95% similarity). Further comparison of the 16S rRNA genes of the BT 1A strains indicated that altogether 17 of the 71 strains belong to Genetic group 2. On the 16S rRNA analysis, these 17 strains were only 98% similar to the previously identified subspecies of Y. enterocolitica. The strains of Genetic group 2 were uniform in their pathogenecity-related properties: they lacked the ystB gene, belonged to the same LPS subtype or were of rough type, were all resistant to the five tested yersiniophages, were largely resistant to serum complement and did not ferment fucose. The 54 strains in Genetic group 1 showed much more variation in these properties. The most commonly detected LPS types were similar to the LPS types of reference strains with serotypes O:6,30 and O:6,31 (37%), O:7,8 (19%) and O:5 (15%). The results of the present study strengthen the assertion that strains classified as Y. enterocolitica BT 1A represent more than one subspecies. Especially the BT 1A strains in our Genetic group 2 commonly showed resistance to human serum complement killing, which may indicate pathogenic potential for these strains. However, their virulence mechanisms remain unknown.
KeywordMeSH Terms
Genetic Variation
166.     ( 1990 )

Genetic analysis of the yopE region of Yersinia spp.: identification of a novel conserved locus, yerA, regulating yopE expression.

Journal of bacteriology 172 (3)
PMID : 2307658  :   DOI  :   10.1128/jb.172.3.1547-1555.1990     PMC  :   PMC208631    
Abstract >>
The yopE gene of Yersinia pseudotuberculosis was recently sequenced, and YopE was identified as an indispensable virulence determinant when tested in a mouse model (A. Forsberg and H. Wolf-Watz, Mol. Microbiol. 2:121-133, 1988). In the study described here, the DNA sequences of the yopE genes of Yersinia pestis EV76 and Yersinia enterocolitica 8081 were determined and compared with that of the Y. pseudotuberculosis gene. Only two codons were found to differ, both leading to amino acid replacements, when the gene from Y. pestis was compared. These two replacements were also present in the gene from Y. enterocolitica; in addition, 18 other codons were found to differ. Thirteen of these substitutions led to amino acid replacements. Downstream of the yopE gene, the plasmid partition locus par was found to be conserved in all three species. In Y. enterocolitica 8081, the sequence homology was interrupted by a putative insertion sequence element inserted between the yopE gene and the par region at a position only 5 base pairs downstream of the yopE stop codon. Upstream of the yopE gene, 620 base pairs were conserved in the three species. This region contained a 130-amino-acid-long open reading frame reading in the opposite direction to the yopE gene and expressed a 14-kilodalton protein in minicells. An insertion mutation in this region constructed in Y. pseudotuberculosis expressed significantly lower amounts of YopE protein in vitro than did the corresponding wild type. The expression level could be restored by transcomplementation. This new locus was designated yerA, for yopE-regulating gene A. The yerA mutant was avirulent when mice were challenged by oral infection.
KeywordMeSH Terms
Gene Expression Regulation, Bacterial
Genes, Bacterial
Genes, Regulator
167.     ( 1998 )

The psychrotrophic bacterium Yersinia enterocolitica requires expression of pnp, the gene for polynucleotide phosphorylase, for growth at low temperature (5 degrees C).

Molecular microbiology 28 (3)
PMID : 9632258  :   DOI  :   10.1046/j.1365-2958.1998.00816.x    
Abstract >>
The psychrotrophic bacterium Yersinia enterocolitica is characterized by temperature-dependent adaptations. To investigate Y. enterocolitica genes involved in cold adaptation, a mutant restricted in its ability to grow at 5 degrees C was isolated from a transposon mutant library. The transposon insertion site in this psychrotrophy-defective (PD) mutant mapped 16 bp upstream of an open reading frame whose predicted amino acid sequence showed 93% similarity with the Escherichia coli exoribonuclease polynucleotide phosphorylase (PNPase), encoded by pnp. Expression of this gene was blocked in the PD mutant. However, the introduction of a second copy of pnp, including 0.33 kbp sequences upstream of its coding region, into the chromosome of the PD mutant restored pnp expression as well as the ability to grow at 5 degrees C. Furthermore, the expression of pnp appeared to be temperature dependent: in the parental Y. enterocolitica strain, the levels of both pnp mRNA and PNPase were 1.6-fold higher at 5 degrees C compared with 30 degrees C. A similarly enhanced level of PNPase at 5 degrees C was observed in the merodiploid recombinant strain, which indicates that the 0.33 kbp region upstream of pnp harboured a cold-inducible promoter. A putative cold shock promoter motif (ATTGG) was observed in this region.
KeywordMeSH Terms
168.     ( 1998 )

Characterization of plasmid regions of foodborne Yersinia enterocolitica biogroup 1A strains hybridizing to the Yersinia enterocolitica virulence plasmid.

Systematic and applied microbiology 21 (2)
PMID : 9704108  :   DOI  :   10.1016/S0723-2020(98)80024-6    
Abstract >>
The aim of our study was to find out if plasmids of foodborne Yersinia enterocolitica biogroup 1A strains harbour genes related to the virulence genes located on the virulence plasmid pYV of Yersinia enterocolitica. The foodborne strains were isolated from pork, as pigs are considered as an important reservoir for enteropathogenic Y. enterocolitica 0:3 and 0:9 strains. The plasmids of the foodborne strains were characterized by restriction enzyme analysis and hybridized to the virulence plasmid pYV of pathogenic Y. enterocolitica strains (0:3 biogroup 4; 0:9 biogroup 2). In several cases the plasmids of the foodborne strains showed homologies to parts of the pYV plasmid. Analysis of the hybridizing regions revealed that genes involved in replication, sequences of transposable elements and an endonuclease gene caused the observed hybridization to the virulence plasmid. In cause of the study also a remnant of a Tn3-like transposon was shown to be present adjacent to the yadA gene on the pYV plasmid. Although there is evidence that at least some strains of Y. enterocolitica biogroup 1A might possess pathogenic properties none of the well known plasmid encoded virulence genes were present on the plasmids of the investigated foodborne biogroup 1A strains.
KeywordMeSH Terms
Food Microbiology
169.     ( 1999 )

Identification of SycN, YscX, and YscY, three new elements of the Yersinia yop virulon.

Journal of bacteriology 181 (2)
PMID : 9882687  :   PMC  :   PMC93427    
Abstract >>
The Yop virulon allows Yersinia spp. to resist the immune response of the host by injecting harmful proteins into host cells. We identified three new elements of the Yop virulon: SycN, required for normal secretion of YopN, and YscX and YscY, two new components of the secretion machinery.
KeywordMeSH Terms
Membrane Proteins
170.     ( 1998 )

Construction and characterisation of a Yersinia enterocolitica O:8 ompR mutant.

FEMS microbiology letters 165 (1)
PMID : 9711851  :   DOI  :   10.1111/j.1574-6968.1998.tb13139.x    
Abstract >>
The ompR-envZ two-component regulatory system has been shown to contribute to virulence in a number of enteric bacterial pathogens. A Yersinia enterocolitica O:8 ompR homologue was amplified, cloned and sequenced, showing 99.2% homology to the Escherichia coli OmpR. An isogenic ompR mutant was constructed by reverse genetics-based methodology. The mutant was shown to have increased sensitivity to high osmolarity, high temperature and low pH stresses in vitro. In the murine yersiniosis model, the mutant was attenuated and offered partial protection against wild-type challenge.
KeywordMeSH Terms
Bacterial Proteins
171.     ( 1998 )

YopT, a new Yersinia Yop effector protein, affects the cytoskeleton of host cells.

Molecular microbiology 29 (3)
PMID : 9723929  :   DOI  :   10.1046/j.1365-2958.1998.00992.x    
Abstract >>
Extracellular Yersinia disarm the immune system of their host by injecting effector Yop proteins into the cytosol of target cells. Five effectors have been described: YopE, YopH, YpkA/YopO, YopP and YopM. Delivery of these effectors by Yersinia adhering at the cell surface requires other Yops (translocators) including YopB. Effector and translocator Yops are secreted by the type III Ysc secretion apparatus, and some Yops also need a specific cytosolic chaperone, called Syc. In this paper, we describe a new Yop, which we have called YopT (35.5kDa). Its secretion required an intact Ysc apparatus and SycT (15.0kDa, pl4.4), a new chaperone resembling SycE. Infection of macrophages with a Yersinia, producing a hybrid YopT-adenylate cyclase, led to the accumulation of intracellular cAMP, indicating that YopT is delivered into the cytosol of eukaryotic cells. Infection of HeLa cells with a mutant strain devoid of the five known Yop effectors (deltaHOPEM strain) but producing YopT resulted in the alteration of the cell cytoskeleton and the disruption of the actin filament structure. This cytotoxic effect was caused by YopT and dependent on YopB. YopT is thus a new effector Yop and a new bacterial toxin affecting the cytoskeleton of eukaryotic cells.
KeywordMeSH Terms
172.     ( 1998 )

Heparin interferes with translocation of Yop proteins into HeLa cells and binds to LcrG, a regulatory component of the Yersinia Yop apparatus.

Molecular microbiology 27 (2)
PMID : 9484897  :   DOI  :   10.1046/j.1365-2958.1998.00691.x    
Abstract >>
Yersiniae are equipped with the Yop virulon, an apparatus that allows extracellular bacteria to deliver toxic Yop proteins inside the host cell cytosol in order to sabotage the communication networks of the host cell or even to cause cell death. LcrG is a component of the Yop virulon involved in the regulation of secretion of the Yops. In this paper, we show that LcrG can bind HeLa cells, and we analyse the role of proteoglycans in this phenomenon. Treatment of the HeLa cells with heparinase I, but not chondroitinase ABC, led to inhibition of binding. Competition assays indicated that heparin and dextran sulphate strongly inhibited binding, but that other glycosaminoglycans did not. This demonstrated that binding of HeLa cells to purified LcrG is caused by heparan sulphate proteoglycans. LcrG could bind directly to heparin-agarose beads and, in agreement with these results, analysis of the protein sequence of Yersinia enterocolitica LcrG revealed heparin-binding motifs. In vitro production and secretion by Y. enterocolitica of the Yops was unaffected by the addition of heparin. However, the addition of exogenous heparin decreased the level of YopE-Cya translocation into HeLa cells. A similar decrease was seen with dextran sulphate, whereas the other glycosaminoglycans tested had no significant effect. Translocation was also decreased by treatment of HeLa cells with heparinitase, but not with chondroitinase. Thus, heparan sulphate proteoglycans have an important role to play in translocation. The interaction between LcrG and heparan sulphate anchored at the surface of HeLa cells could be a signal triggering deployment of the Yop translocation machinery. This is the first report of a eukaryotic receptor interacting with the type III secretion and associated translocation machinery of Yersinia or of other bacteria.
KeywordMeSH Terms
173.     ( 1997 )

YscM1 and YscM2, two Yersinia enterocolitica proteins causing downregulation of yop transcription.

Molecular microbiology 26 (4)
PMID : 9427412  :   DOI  :   10.1046/j.1365-2958.1997.6281995.x    
Abstract >>
Synthesis of the Yop proteins by yersiniae is downregulated when secretion is prevented by closure or destruction of the contact (type III) secretion channel. In Yersinia pseudotuberculosis, a mutation in the IcrQ gene, encoding a secreted protein, reduces this feedback inhibition mechanism. Surprisingly, mutation in the yscM gene, the IcrQ homologue in Y. enterocolitica, does not lead to the same deregulated phenotype. In this paper, we addressed the question of this discrepancy. We found a new gene on the Y. enterocolitica pYV plasmid that encodes a protein with 57% identity to YscM (now called YscM1). Overexpression of this gene, called yscM2, like overexpression of IcrQ and yscM1, blocked Yop secretion. A double yscM1, yscM2 mutant had the same phenotype as that of the IcrQ mutant. The discrepancy can thus be explained by the existence of two functionally equivalent copies of yscM in Y. enterocolitica. Overexpression of yscM1 drastically reduced the expression of a yopH-cat reporter gene when tested in a pYV+ background. However, no effect could be observed in the absence of a pYV plasmid, indicating that YscM1 does not act directly as a transcriptional repressor or as an anti-VirF factor. We have also ruled out that YscM acts by obstructing the secretion channel.
KeywordMeSH Terms
Down-Regulation
Transcription, Genetic
Virulence Factors
174.     ( 1997 )

Detection and speciation of bacteria through PCR using universal major cold-shock protein primer oligomers.

Journal of industrial microbiology & biotechnology 19 (4)
PMID : 9439003  :  
Abstract >>
The detection of bacteria using PCR is a well-established diagnostic technique. However, conventional PCR requires the use of DNA primer oligomers that are specific to the target organism and, as a consequence, a sample can only be tested for the presence of that specific target. A significant advantage would be to probe a sample for the presence of any bacteria, followed by identification. To achieve this it is necessary to identify a DNA sequence common to all bacteria. Here we demonstrate that such a sequence may be that encoding the major cold-shock proteins. Using two universal PCR primer oligomers from conserved regions of these gene homologues, we have amplified a 200 base-pair DNA sequence from more than 30 diverse Gram-positive and Gram-negative bacteria, including representatives from the genera Aeromonas, Bacillus, Citrobacter, Enterobacter, Enterococcus, Escherichia, Klebsiella, Lactobacillus, Lactococcus, Listeria, Pediococcus, Photobacterium, Proteus, Salmonella, Shigella, Staphylococcus, Streptococcus, and Yersinia. Sequence analysis of the amplified products confirmed a high level of DNA homology. Significantly, however, there are sufficient nucleotide variations to allow the unique allocation of each amplified sequence to its parental bacterium.
KeywordMeSH Terms
DNA Primers
175.     ( 1998 )

Phylogenetic relationships of Salmonella based on DNA sequence comparison of atpD encoding the beta subunit of ATP synthase.

FEMS microbiology letters 161 (1)
PMID : 9561735  :   DOI  :   10.1111/j.1574-6968.1998.tb12933.x    
Abstract >>
DNA sequences covering 57% of atpD encoding the beta subunit of ATP synthase were determined for 16 strains of Salmonella enterica, two strains of S. bongori, and one strain each of Citrobacter freundii and Yersinia enterocolitica, and comparison was made with the published Escherichia coli and Enterobacter aerogenes sequences. The phylogenetic tree based on maximum-likelihood analysis showed separation of the subspecies of S. enterica except for two serotypes of subspecies II which were unsupported by a common node. The two serotypes of S. bongori were separated from S. enterica and related to the serotypes of subspecies II. A tight relationship was found between S. enterica subspecies IIIa consisting of monophasic serotypes and subspecies IIIb consisting of diphasic serotypes. This is in conflict with results obtained for most other housekeeping genes and the 23S rRNA gene separating mono- from diphasic subspecies.
KeywordMeSH Terms
176.     ( 1997 )

Contribution of the Mn-cofactored superoxide dismutase (SodA) to the virulence of Yersinia enterocolitica serotype O8.

Infection and immunity 65 (11)
PMID : 9353054  :   PMC  :   PMC175675    
Abstract >>
Enteric pathogens harbor a set of enzymes (e.g., superoxide dismutases [SOD]) for detoxification of endogenous and exogenous reactive oxygen species which are encountered during infection. To analyze the role of the Mn-cofactored SOD (SodA) in the pathogenicity of yersiniae, we cloned the sodA gene of Yersinia enterocolitica serotype O8 by complementation of an Escherichia coli sodA sodB mutant and subsequently constructed an isogenic mutant by allelic exchange. Sequence analysis revealed an open reading frame that enabled the deduction of a sequence of 207 amino acids with 85% identity to SodA of E. coli. In a mouse infection model, the sodA null mutant was strongly attenuated in comparison to its parental strain. After intravenous infection, the survival and multiplication of the mutant in the spleen and liver were markedly reduced. In contrast, inactivation of sodA had only minor effects on survival and multiplication in the gut and Peyer's patches, as could be demonstrated in the orogastric infection model. The reduction in virulence was accompanied by a low but significant increase of susceptibility of the soda mutant to bacterial killing by polymorphonuclear leukocytes (PMN) and an alteration of the intracellular chemiluminescence response of PMN. These results suggest that the resistance of Y. enterocolitica to exogenous oxygen radicals produced by phagocytes involves the Mn-cofactored SOD. The important role of sodA for the pathogenicity of Y. enterocolitica could also be due to detoxification of endogenous, metabolically produced oxygen radicals which are encountered by extracellular enteric pathogens during the invasion of the host.
KeywordMeSH Terms
177.     ( 1998 )

The yersiniabactin biosynthetic gene cluster of Yersinia enterocolitica: organization and siderophore-dependent regulation.

Journal of bacteriology 180 (3)
PMID : 9457855  :   PMC  :   PMC106919    
Abstract >>
The ability to synthesize and uptake the Yersinia siderophore yersiniabactin is a hallmark of the highly pathogenic, mouse-lethal species Yersinia pestis, Y. pseudotuberculosis, and Y. enterocolitica 1B. We have identified four genes, irp1, irp3, irp4, and irp5, on a 13-kb chromosomal DNA fragment of Y. enterocolitica 08, WA-314. These genes constitute the yersiniabactin biosynthetic gene cluster together with the previously defined irp2. The irp1 gene consists of 9,486 bp capable of encoding a 3,161-amino-acid high-molecular-weight protein 1 (HMWP1) polypeptide with a predicted mass of 384.6 kDa. The first 3,000 bp of irp1 show similarity to the corresponding regions of the polyketide synthase genes of Bacillus subtilis and Streptomyces antibioticus. The remaining part of irp1 is most similar to irp2, encoding HMWP2, which might be the reason for immunological cross-reactivity of the two polypeptides. Irp4 was found to have 41.7% similarity to thioesterase-like protein of the anguibactin biosynthetic genes of Vibrio anguillarum. Irp5 shows 41% similarity to EntE, the 2,3-dihydroxybenzoic acid-activating enzyme utilized in enterobactin synthesis of Escherichia coli. Irp4 and Irp5 are nearly identical to YbtT and YbtE, recently identified in Y. pestis. irp3 has no similarity to any known gene. Inactivation of either irp1 or irp2 abrogates yersiniabactin synthesis. Mutations in irp1 or fyuA (encoding yersiniabactin/pesticin receptor) result in downregulation of irp2 that can be upregulated by the addition of yersiniabactin. A FyuA-green fluorescent protein translational fusion was downregulated in an irp1 mutant. Upregulation was achieved by addition of yersiniabactin but not desferal, pesticin, or pyochelin, which indicates high specificity of the FyuA receptor and autoregulation of genes involved in synthesis and uptake of yersiniabactin.
KeywordMeSH Terms
Genes, Bacterial
Operon
178.     ( 1998 )

TyeA, a protein involved in control of Yop release and in translocation of Yersinia Yop effectors.

The EMBO journal 17 (7)
PMID : 9524114  :   DOI  :   10.1093/emboj/17.7.1907     PMC  :   PMC1170537    
Abstract >>
Extracellular Yersinia spp. disarm the immune system by injecting the effector Yersinia outer proteins (Yops) into the target cell. Yop secretion is triggered by contact with eukaryotic cells or by Ca2+ chelation. Two proteins, YopN and LcrG, are known to be involved in Yop-secretion control. Here we describe TyeA, a third protein involved in the control of Yop release. Like YopN, TyeA is localized at the bacterial surface. A tyeA knock-out mutant secreted Yops in the presence of Ca2+ and in the absence of eukaryotic cells. Unlike a yopN null mutant, the tyeA mutant was defective for translocation of YopE and YopH, but not YopM, YopO and YopP, into eukaryotic cells. This is the first observation suggesting that Yop effectors can be divided into two sets for delivery into eukaryotic cells. TyeA was found to interact with the translocator YopD and with residues 242-293 of YopN. In contrast with a yopN null mutant, a yopNDelta248-272 mutant was also unable to translocate YopE and YopH. Our results suggest that TyeA forms part of the translocation-control apparatus together with YopD and YopN, and that the interaction of these proteins is required for selective translocation of Yops inside eukaryotic cells.
KeywordMeSH Terms
Membrane Proteins
179.     ( 1997 )

The ClpP protein, a subunit of the Clp protease, modulates ail gene expression in Yersinia enterocolitica.

Molecular microbiology 26 (1)
PMID : 9383193  :   DOI  :   10.1046/j.1365-2958.1997.5551916.x    
Abstract >>
Yersinia enterocolitica is a gastrointestinal pathogen of humans and animals. Ail is a 17kDa cell-surface protein that confers on Y. enterocolitica resistance to serum killing and the ability to attach to and invade cells in vitro. The ail gene of Y. enterocolitica is regulated by temperature and growth phase. In stationary phase, ail transcript is only detected when bacteria are grown at the host temperature of 37 degrees C. Our laboratory previously described a group of mini-Tn10 mutants, which expressed ail in stationary phase at 28 degrees C. In one of these mutants, DP5102::mini-Tn10 3-2, the mini-Tn 10 inserted into a gene encoding a protein with 90.3% identity to the ClpP protease subunit from Escherichia coli. Expression of ail in stationary phase at 28 degrees C was also derepressed in a directed Y. enterocolitica clpP mutant. Analysis of ail transcripts in the wild-type and clpP mutant strains indicated that there is a single start site of transcription of ail and that the effect of the clpP mutation was on the initiation of transcription at this site. Similar to E. coli, a clpX homologue was identified downstream of clpP. The Y. enterocolitica clpP gene complemented the clpP mutant phenotype, repressing the expression of both ail transcript levels and cell surface-expressed Ail protein. Thus, ClpP has a role in the modulation of ail transcription in Y. enterocolitica.
KeywordMeSH Terms
Gene Expression Regulation, Bacterial

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