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1. Haas  H, Herfurth  E, Stöffler  G, Redl  B,     ( 1992 )

Purification, characterization and partial amino acid sequences of a xylanase produced by Penicillium chrysogenum.

Biochimica et biophysica acta 1117 (3)
PMID : 1420277  :   DOI  :   10.1016/0304-4165(92)90025-p    
Abstract >>
An extracellular xylanase (1,4-beta-D-xylan xylanohydrolase, EC 3.2.1.8, endo 1,4-beta-xylanase) was found to be the major protein in the culture filtrate of Penicillium chrysogenum when grown on 1% xylan. In contrast to other microorganism no xylanase multiplicity was found in P. chrysogenum under the conditions used. This enzyme was purified to homogeneity by high performance anion-exchange and size-exclusion chromatography. It had an M(r) of 35,000 as estimated by SDS-PAGE and was shown to be active as a monomer. No glycosylation of the protein could be detected neither by a sensitive glycostain nor by enzymatic deglycosylation studies. The enzyme hydrolyzed oat spelt and birchwood xylan randomly, yielding xylose and xylobiose as major end products. It had no cellulase, CMCase, beta-xylosidase or arabinogalactanase activity but acted on p-nitrophenylcellobioside. The pH and temperature optima for its activity were pH 6.0 and 40 degrees C, respectively. Eight peptides obtained after endoproteinase LysC digestion of xylanase have been sequenced, six of them showed considerable amino acid similarity to glucanases and high M(r)/acidic xylanases from different bacteria, yeasts and fungi.
KeywordMeSH Terms
2. Shen  HD, Chou  H, Tam  MF, Chang  CY, Lai  HY, Wang  SR,     ( 2003 )

Molecular and immunological characterization of Pen ch 18, the vacuolar serine protease major allergen of Penicillium chrysogenum.

Allergy 58 (10)
PMID : 14510716  :   DOI  :   10.1034/j.1398-9995.2003.00107.x    
Abstract >>
We have suggested previously that the 32 and 34 kDa major allergens of Penicillium chrysogenum (also known as P. notatum) are the vacuolar (Pen ch 18) and the alkaline (Pen ch 13) serine proteases, respectively, of P. chrysogenum. The purpose of this study is to characterize the 32 kDa allergen of P. chrysogenum and its immunoglobulin E (IgE)cross-reactivity with Pen ch 13 allergen. The full-length cDNA of Pen ch 18 was isolated by reverse transcriptase-polymerase chain reaction and the 5'-rapid amplification cDNA end reaction. Recombinant Pen ch 18 was expressed as his-tagged proteins in Escherichia coli. Its reactivity with IgE and monoclonal antibodies against fungal serine protease allergens was analyzed by immunoblotting. The IgE cross-reactivity between Pen ch 18 and Pen ch 13 was analyzed by immunoblot inhibition. Overlapping recombinant fragments and synthetic peptides were used to map the B cell epitopes on Pen ch 18. In this study, we isolated a 1857 bp cDNA fragment containing an open reading frame of 494 amino acids that encodes the preproenzyme of Pen ch 18. Similar to other vacuolar serine proteases, this precursor appears to undergo N- and possibly C-terminal cleavage upon maturation. The his-tagged recombinant Pen ch 18 containing the putative sequence of the mature protein reacted with IgE antibodies in serum samples from asthmatic patients. In addition, IgE-binding to the 32 kDa major allergen of P. chrysogenum was inhibited when a positive serum sample was absorbed with recombinant Pen ch 18 before immunoblotting. Both inhibition and almost no inhibition of IgE-binding to the 32 kDa major allergen of Pen ch 18 were detected when eight positive serum samples were preabsorbed individually with purified Pen ch 13 before immunoblotting. The major IgE binding region was located in a fragment (PN1) encompassing the N-terminal 102 amino acid residues of the recombinant Pen ch 18. A dominant linear IgE epitope was further mapped within residues 73-95 (peptide PN1-e) of the N-terminally processed allergen. Monoclonal antibody FUM20 that reacts with Pen ch 18 but not with Pen ch 13 binds a synthetic peptide with sequence encompassing the N-terminal 23 residues of the recombinant Pen ch 18. Monoclonal antibody PCM39 that reacts with both Pen ch 13 and Pen ch 18 recognizes a peptide containing residues 132-154 of the allergen. Our results confirm that the Pen ch 18 allergen is a vacuolar serine protease of P. chrysogenum that matures through N- and possibly C-terminal processing. The finding that there are cross-reactive and allergen-specific IgE epitopes for Pen ch 18 and Pen ch 13 suggests that both major allergens should be included in clinically diagnostic P. chrysogenum extracts.
KeywordMeSH Terms
3. Sakamoto  T, Kawasaki  H,     ( 2003 )

Purification and properties of two type-B alpha-L-arabinofuranosidases produced by Penicillium chrysogenum.

Biochimica et biophysica acta 1621 (2)
PMID : 12726996  :   DOI  :   10.1016/s0304-4165(03)00058-8    
Abstract >>
Two distinct extracellular alpha-L-arabinofuranosidases (AFases; EC 3.2.1.55) were purified from the culture filtrate of Penicillium chrysogenum 31B. The molecular masses of the enzymes were estimated to be 79 kDa (AFQ1) and 52 kDa (AFS1) by SDS-PAGE. Both enzymes had their highest activities at 50 degrees C and were stable up to 50 degrees C. Enzyme activities of AFQ1 and AFS1 were highest at pH 4.0 to 6.5 and pH 3.3 to 5.0, respectively. Addition of 10 mg/ml arabinose to the reaction mixture decreased the AFS1 activity but hardly affected AFQ1. Both enzymes displayed broad substrate specificities; they released arabinose from branched arabinan, debranched arabinan, arabinoxylan, arabinogalactan, and arabino-oligosaccharides. AFS1 also showed low activity towards p-nitrophenyl-beta-D-xylopyranoside. An exo-arabinanase, which catalyzes the release of arabinobiose from linear arabinan at the nonreducing terminus, acted synergistically with both enzymes to produce L-arabinose from branched arabinan.
KeywordMeSH Terms
4. Chou  H, Chang  CY, Tsai  JJ, Tang  RB, Lee  SS, Wang  SR, Peng  HJ, Shen  HD,     ( 2003 )

The prevalence of IgE antibody reactivity against the alkaline serine protease major allergen of Penicillium chrysogenum increases with the age of asthmatic patients.

Annals of allergy, asthma & immunology : official publication of the American College of Allergy, Asthma, & Immunology 90 (2)
PMID : 12602675  :   DOI  :   10.1016/S1081-1206(10)62150-3    
Abstract >>
Penicillium species are prevalent airborne fungi. However, the prevalence of allergic sensitization to Penicillium antigens and the true impact of these ubiquitous fungi on atopic respiratory disorders remain to be determined. The purpose of this study was to analyze the prevalence of immunoglobulin (Ig)E and IgG antibodies against Penicillium chrysogenum (Pen ch 13), the alkaline serine protease major allergen of P. chrysogenum, in asthmatic patients of different age groups. Pen ch 13 was purified from a culture medium of P. chrysogenum. The reactivity of IgE and IgG antibodies to Pen ch 13 in the serum samples of 212 asthmatic patients was analyzed by immunoblotting methods. Sixty-nine (33%) of the 212 sera analyzed showed IgE and/or IgG immunoblot reactivity to Pen ch 13. Significant differences in the prevalence of IgE and/or IgG antibody reactivity to Pen ch 13 were found among eight different age groups of 212 asthmatic patients. The frequency of IgE-binding reactivity to Pen ch 13 increased significantly with the age of the patients. It was 7% for the group less than 10 years old and 42% for the group older than 70 years old. In addition, a significant difference between the prevalence of IgE (7%) and IgG (33%) antibodies against Pen ch 13 in the group aged 10 or less was also found. Our study demonstrates that IgE and IgG antibodies specific for Pen ch 13 were detected in approximately one-third of the 212 asthmatic patients analyzed. Our results suggest that allergic sensitization to Pen ch 13, and possibly to other airborne Penicillium species, is more common in older asthmatic patients.
KeywordMeSH Terms
5. MacRae  IJ, Segel  IH, Fisher  AJ,     ( 2002 )

Allosteric inhibition via R-state destabilization in ATP sulfurylase from Penicillium chrysogenum.

Nature structural biology 9 (12)
PMID : 12426581  :   DOI  :   10.1038/nsb868    
Abstract >>
The structure of the cooperative hexameric enzyme ATP sulfurylase from Penicillium chrysogenum bound to its allosteric inhibitor, 3'-phosphoadenosine-5'-phosphosulfate (PAPS), was determined to 2.6 A resolution. This structure represents the low substrate-affinity T-state conformation of the enzyme. Comparison with the high substrate-affinity R-state structure reveals that a large rotational rearrangement of domains occurs as a result of the R-to-T transition. The rearrangement is accompanied by the 17 A movement of a 10-residue loop out of the active site region, resulting in an open, product release-like structure of the catalytic domain. Binding of PAPS is proposed to induce the allosteric transition by destabilizing an R-state-specific salt linkage between Asp 111 in an N-terminal domain of one subunit and Arg 515 in the allosteric domain of a trans-triad subunit. Disrupting this salt linkage by site-directed mutagenesis induces cooperative inhibition behavior in the absence of an allosteric effector, confirming the role of these two residues.
KeywordMeSH Terms
Models, Molecular
6. Lansdon  EB, Segel  IH, Fisher  AJ,     ( 2002 )

Ligand-induced structural changes in adenosine 5'-phosphosulfate kinase from Penicillium chrysogenum.

Biochemistry 41 (46)
PMID : 12427029  :   DOI  :   10.1021/bi026556b    
Abstract >>
Adenosine 5'-phosphosulfate (APS) kinase catalyzes the second reaction in the two-step, ATP-dependent conversion of inorganic sulfate to 3'-phosphoadenosine 5'-phosphosulfate (PAPS). PAPS serves as the sulfuryl donor for the biosynthesis of all sulfate esters and also as a precursor of reduced sulfur biomolecules in many organisms. Previously, we determined the crystal structure of ligand-free APS kinase from the filamentous fungus, Penicillium chrysogenum [MacRae et al. (2000) Biochemistry 39, 1613-1621]. That structure contained a protease-susceptible disordered region ("mobile lid"; residues 145-170). Addition of MgADP and APS, which together promote the formation of a nonproductive "dead-end" ternary complex, protected the lid from trypsin. This report presents the 1.43 A resolution crystal structure of APS kinase with both ADP and APS bound at the active site and the 2.0 A resolution structure of the enzyme with ADP alone bound. The mobile lid is ordered in both complexes and is shown to provide part of the binding site for APS. That site is formed primarily by the highly conserved Arg 66, Arg 80, and Phe 75 from the protein core and Phe 165 from the mobile lid. The two Phe residues straddle the adenine ring of bound APS. Arg 148, a completely conserved residue, is the only residue in the mobile lid that interacts directly with bound ADP. Ser 34, located in the apex of the P-loop, hydrogen-bonds to the 3'-OH of APS, the phosphoryl transfer target. The structure of the binary E.ADP complex revealed further changes in the active site and N-terminal helix that occur upon the binding/release of (P)APS.
KeywordMeSH Terms
7. Yu  CJ, Chen  YM, Su  SN, Forouhar  F, Lee  SH, Chow  LP,     ( 2002 )

Molecular and immunological characterization and IgE epitope mapping of Pen n 18, a major allergen of Penicillium notatum.

The Biochemical journal 363 (Pt 3)
PMID : 11964171  :   DOI  :   10.1042/bj3630707     PMC  :   PMC1222523    
Abstract >>
The mould genus, Penicillium, is a significant source of environmental aero-allergens. A major allergen from Penicillium notatum, Pen n 18, was identified by two-dimensional immunoblotting using monoclonal antibody G11A10, raised against the vacuolar serine protease of Penicillium citrinum, followed by matrix-assisted laser-desorption ionization-time-of-flight MS analysis of the peptide digest. Pen n 18 was then cloned and the amino acid sequence deduced from the cDNA sequence. The cDNA encoded a 494 amino acid protein, considerably larger than mature Pen n 18, the differences being due to the N- and C-terminal prosequences. The deduced amino acid sequence showed extensive similarity with those of vacuolar serine proteases from various fungi. The Pen n 18 coding sequence was expressed in Escherichia coli as a His-tagged fusion protein and purified by Ni(2+)-chelate affinity chromatography. On immunoblots, the purified recombinant protein specifically bound IgE from mould-allergic patients, and cross-inhibition assays demonstrated the presence of common IgE-binding epitopes on Pen n 18 and a major allergen of P. citrinum, Pen c 18. When mapping of the allergenic epitopes was performed, at least nine different linear IgE-binding epitopes, located throughout the Pen n 18 protein, were identified. Of these, peptide C12, located in the N-terminal region of the molecule, was recognized by serum from 75% of the patients tested and therefore appears to be an immunodominant IgE-binding epitope.
KeywordMeSH Terms
Epitope Mapping
Immunoglobulin E
8. Holzmann  K, Schreiner  E, Schwab  H,     ( 2002 )

A Penicillium chrysogenum gene (aox) identified by specific induction upon shifting pH encodes for a protein which shows high homology to fungal alcohol oxidases.

Current genetics 40 (5)
PMID : 11935224  :   DOI  :   10.1007/s002940100251    
Abstract >>
Parallel cultures of Penicillium chrysogenum were grown in controlled bioreactors under conditions of penicillin production and one was shifted from the initial pH 6.0 to pH 8.0. RNA was isolated from both cultures and used for a differential hybridization experiment to identify genes specifically induced upon this pH shift. About 2,000 plaques of a cDNA library constructed from pH 8.0 material were screened with a pH 8.0 to pH 6.0 subtracted probe. Two independent clones of which the RNA was highly abundant in pH 8-shifted material and essentially not present in pH 6 material were isolated. Both clones were found to belong to one specific gene, which could be identified by sequence homology as an alcohol oxidase. The identified aox gene encoded for a peptide of 666 amino acids, interrupted by nine introns; and it showed high homology (>65% identity) to alcohol oxidases from Cladosporium fulvum and the methanol-utilizing yeasts Candida boidinii, Hansenula polymorpha (now Pichia angusta) and P. pastoris. The transcription start was identified by primer extension analysis. Southern analysis revealed that related genes are widely distributed among fungal species.
KeywordMeSH Terms
Genes, Fungal
9. Naranjo  L, Martin de Valmaseda  E, Bañuelos  O, Lopez  P, Riaño  J, Casqueiro  J, Martin  JF,     ( 2001 )

Conversion of pipecolic acid into lysine in Penicillium chrysogenum requires pipecolate oxidase and saccharopine reductase: characterization of the lys7 gene encoding saccharopine reductase.

Journal of bacteriology 183 (24)
PMID : 11717275  :   DOI  :   10.1128/JB.183.24.7165-7172.2001     PMC  :   PMC95565    
Abstract >>
Pipecolic acid is a component of several secondary metabolites in plants and fungi. This compound is useful as a precursor of nonribosomal peptides with novel pharmacological activities. In Penicillium chrysogenum pipecolic acid is converted into lysine and complements the lysine requirement of three different lysine auxotrophs with mutations in the lys1, lys2, or lys3 genes allowing a slow growth of these auxotrophs. We have isolated two P. chrysogenum mutants, named 7.2 and 10.25, that are unable to convert pipecolic acid into lysine. These mutants lacked, respectively, the pipecolate oxidase that converts pipecolic acid into piperideine-6-carboxylic acid and the saccharopine reductase that catalyzes the transformation of piperideine-6-carboxylic acid into saccharopine. The 10.25 mutant was unable to grow in Czapek medium supplemented with alpha-aminoadipic acid. A DNA fragment complementing the 10.25 mutation has been cloned; sequence analysis of the cloned gene (named lys7) revealed that it encoded a protein with high similarity to the saccharopine reductase from Neurospora crassa, Magnaporthe grisea, Saccharomyces cerevisiae, and Schizosaccharomyces pombe. Complementation of the 10.25 mutant with the cloned gene restored saccharopine reductase activity, confirming that lys7 encodes a functional saccharopine reductase. Our data suggest that in P. chrysogenum the conversion of pipecolic acid into lysine proceeds through the transformation of pipecolic acid into piperideine-6-carboxylic acid, saccharopine, and lysine by the consecutive action of pipecolate oxidase, saccharopine reductase, and saccharopine dehydrogenase.
KeywordMeSH Terms
10. Chou  H, Lai  HY, Tam  MF, Chou  MY, Wang  SR, Han  SH, Shen  HD,     ( 2002 )

cDNA cloning, biological and immunological characterization of the alkaline serine protease major allergen from Penicillium chrysogenum.

International archives of allergy and immunology 127 (1)
PMID : 11893850  :   DOI  :   10.1159/000048165    
Abstract >>
Penicillium chrysogenum (Penicillium notatum) is a prevalent airborne Penicillium species. A 34-kD major IgE-reacting component from P. chrysogenum has been identified as an alkaline serine protease (Pen ch 13, also known as Pen n 13 before) by immunoblot and N-terminal amino acid sequence analysis. In the present study, Pen ch 13 was further characterized in terms of cDNA cloning, protein purification, enzymatic activity, histamine release and IgE cross-reactivity with alkaline serine protease allergens from two other prevalent fungal species--P. citrinum (Pen c 13) and Aspergillus flavus (Asp fl 13). A 1,478-bp cDNA (Pen ch 13) that encodes a 398-amino-acid alkaline serine protease from P. chrysogenum was isolated. This fungal protease has pre- and pro-enzyme sequences. The previously determined N-terminal amino acid sequence of the P. chrysogenum 34-kD major allergen is identical to that of residues 116-125 of the cDNA. Starting from Ala116, the deduced amino acid sequence (283 residues) of the mature alkaline serine protease has a calculated molecular mass of 28.105 kD with two cysteines and two putative N-glycosylation sites. It has 83 and 49% sequence identity with the alkaline serine proteases from P. citrinum and A. fumigatus, respectively. The recombinant Pen ch 13 was recovered from inclusion bodies and isolated under denaturing condition. This recombinant protein reacted with IgE antibodies in serum from an asthmatic patient and with monoclonal antibodies (PCM8, PCM10, PCM39) that reacted with the 34-kD component from P. chrysogenum. The N-terminal amino acid sequence of the purified native Pen ch 13 is identical to that determined previously for the 34-kD major allergen in crude P. chrysogenum extracts. The purified native Pen ch 13 has proteolytic activity with casein as the substrate at pH 8.0. This enzymatic activity was inhibited by phenylmethylsulfonyl fluoride or diethylpyrocarbonate. Pen ch 13 was also able to degrade gelatin and collagen but not elastin. Basophils from 5 asthmatic patients released histamine (12-73%) when exposed to the purified Pen ch 13. In ELISA (enzyme-linked immunosorbent assay) experiments, IgE for Pen ch 13 was able to compete with purified Pen ch 13, Pen c 13 or Asp fl 13 in a dose-related manner. These results demonstrated that the 34-kD major allergen of P. chrysogenum is an alkaline serine protease. These results also indicated that atopic patients primarily sensitized by either of these prevalent fungal species may develop allergic symptoms by exposure to other environmental fungi due to cross-reacting IgE antibodies against this protease.
KeywordMeSH Terms
Antigens, Fungal
Serine Endopeptidases
11. MacRae  IJ, Segel  IH, Fisher  AJ,     ( 2001 )

Crystal structure of ATP sulfurylase from Penicillium chrysogenum: insights into the allosteric regulation of sulfate assimilation.

Biochemistry 40 (23)
PMID : 11389593  :   DOI  :   10.1021/bi010367w    
Abstract >>
ATP sulfurylase from Penicillium chrysogenum is an allosterically regulated enzyme composed of six identical 63.7 kDa subunits (573 residues). The C-terminal allosteric domain of each subunit is homologous to APS kinase. In the presence of APS, the enzyme crystallized in the orthorhombic space group (I222) with unit cell parameters of a = 135.7 A, b = 162.1 A, and c = 273.0 A. The X-ray structure at 2.8 A resolution established that the hexameric enzyme is a dimer of triads in the shape of an oblate ellipsoid 140 A diameter x 70 A. Each subunit is divided into a discreet N-terminal domain, a central catalytic domain, and a C-terminal allosteric domain. Two molecules of APS bound per subunit clearly identify the catalytic and allosteric domains. The sequence 197QXRN200 is largely responsible for anchoring the phosphosulfate group of APS at the active site of the catalytic domain. The specificity of the catalytic site for adenine nucleotides is established by specific hydrogen bonds to the protein main chain. APS was bound to the allosteric site through sequence-specific interactions with amino acid side chains that are conserved in true APS kinase. Within a given triad, the allosteric domain of one subunit interacts with the catalytic domain of another. There are also allosteric-allosteric, allosteric-N-terminal, and catalytic-catalytic domain interactions across the triad interface. The overall interactions-each subunit with four others-provide stability to the hexamer as well as a way to propagate a concerted allosteric transition. The structure presented here is believed to be the R state. A solvent channel, 15-70 A wide exists along the 3-fold axis, but substrates have access to the catalytic site only from the external medium. On the other hand, a surface "trench" links each catalytic site in one triad with an allosteric site in the other triad. This trench may be a vestigial feature of a bifunctional ("PAPS synthetase") ancestor of fungal ATP sulfurylase.
KeywordMeSH Terms
12. Díez  B, Marcos  AT, Rodríguez  M, de la Fuente  JL, Barredo  JL,     ( 2001 )

Structural and phylogenetic analysis of the gamma-actin encoding gene from the penicillin-producing fungus Penicillium chrysogenum.

Current microbiology 42 (2)
PMID : 11136133  :  
Abstract >>
The nucleotide sequence of a 2994-bp genomic fragment, including the gamma-actin encoding gene from Penicillium chrysogenum, has been determined, showing an open reading frame (ORF) of 1756 bp interrupted by five introns with fungal consensus splice-site junctions. The 5' untranslated region contains a consensus TATA box, five CAAT motifs, and two large pyrimidine stretches. The predicted protein (375 amino acids) revealed high identity to gamma-actins from fungi (>90%), and gene phylogenies support the grouping of P. chrysogenum actin close to those from the majority of the filamentous fungi. The actA gene is present as a single copy in the genome of P. chrysogenum, and its expression is constitutive during penicillin fermentation, showing a single 1.4-kb transcript.
KeywordMeSH Terms
Genes, Fungal
13. van de Kamp  M, Schuurs  TA, Vos  A, van der Lende  TR, Konings  WN, Driessen  AJ,     ( 2000 )

Sulfur regulation of the sulfate transporter genes sutA and sutB in Penicillium chrysogenum.

Applied and environmental microbiology 66 (10)
PMID : 11010912  :   DOI  :   10.1128/aem.66.10.4536-4538.2000     PMC  :   PMC92338    
Abstract >>
Penicillium chrysogenum uses sulfate as a source of sulfur for the biosynthesis of penicillin. Sulfate uptake and the mRNA levels of the sulfate transporter-encoding sutB and sutA genes are all reduced by high sulfate concentrations and are elevated by sulfate starvation. In a high-penicillin-yielding strain, sutB is effectively transcribed even in the presence of excess sulfate. This deregulation may facilitate the efficient incorporation of sulfur into cysteine and penicillin.
KeywordMeSH Terms
Anion Transport Proteins
Cation Transport Proteins
Fungal Proteins
14. Kiel  JA, Hilbrands  RE, Bovenberg  RA, Veenhuis  M,     ( 2000 )

Isolation of Penicillium chrysogenum PEX1 and PEX6 encoding AAA proteins involved in peroxisome biogenesis.

Applied microbiology and biotechnology 54 (2)
PMID : 10968639  :  
Abstract >>
In Penicillium chrysogenum, key enzymes involved in the production of penicillin reside in peroxisomes. As a first step to understand the role of these organelles in penicillin biosynthesis, we set out to isolate the genes involved in peroxisome biogenesis. Here we report the cloning and characterization of P. chrysogenum PEX1 and PEX6, which encode proteins of the AAA family of ATPases. The second AAA module, which is essential for the function of Pex1p and Pex6p in peroxisome biogenesis, is highly conserved in both PcPexlp and PcPex6p. PcPEX1 and PcPEX6 contain three and two introns, respectively.
KeywordMeSH Terms
15. Schmitt  EK,     ( 2000 )

The fungal CPCR1 protein, which binds specifically to beta-lactam biosynthesis genes, is related to human regulatory factor X transcription factors.

The Journal of biological chemistry 275 (13)
PMID : 10734077  :   DOI  :   10.1074/jbc.275.13.9348    
Abstract >>
Here we report the isolation and characterization of a novel transcription factor from the cephalosporin C-producing fungus Acremonium chrysogenum. We have identified a protein binding site in the promoter of the beta-lactam biosynthesis gene pcbC, located 418 nucleotides upstream of the translational start. Using the yeast one-hybrid system, we succeeded in isolating a cDNA clone encoding a polypeptide, which binds specifically to the pcbC promoter. The polypeptid shows significant sequence homology to human transcription factors of the regulatory factor X (RFX) family and was designated CPCR1. A high degree of CPCR1 binding specificity was observed in in vivo and in vitro experiments using mutated versions of the DNA binding site. The A. chrysogenum RFX protein CPCR1 recognizes an imperfect palindrome, which resembles binding sites of human RFX transcription factors. One- and two-hybrid experiments with truncated versions of CPCR1 showed that the protein forms a DNA binding homodimer. Nondenaturing electrophoresis revealed that the CPCR1 protein exists in vitro solely in a multimeric, probably dimeric, state. Finally, we isolated a homologue of the cpcR1 gene from the penicillin-producing fungus Penicillium chrysogenum and determined about 60% identical amino acid residues in the DNA binding domain of both fungal RFX proteins, which show an overall amino acid sequence identity of 29%.
KeywordMeSH Terms
16. Segel  IH, MacRae  IJ,     ( 2000 )

Crystal structure of adenosine 5'-phosphosulfate kinase from Penicillium chrysogenum.

Biochemistry 39 (7)
PMID : 10677210  :   DOI  :   10.1021/bi9924157    
Abstract >>
Adenosine 5'-phosphosulfate (APS) kinase catalyzes the second reaction in the two-step conversion of inorganic sulfate to 3'-phosphoadenosine 5'-phosphosulfate (PAPS). This report presents the 2.0 A resolution crystal structure of ligand-free APS kinase from the filamentous fungus, Penicillium chrysogenum. The enzyme crystallized as a homodimer with each subunit folded into a classic kinase motif consisting of a twisted, parallel beta-sheet sandwiched between two alpha-helical bundles. The Walker A motif, (32)GLSASGKS(39), formed the predicted P-loop structure. Superposition of the APS kinase active site region onto several other P-loop-containing proteins revealed that the conserved aspartate residue that usually interacts with the Mg(2+) coordination sphere of MgATP is absent in APS kinase. However, upon MgATP binding, a different aspartate, Asp 61, could shift and bind to the Mg(2+). The sequence (156)KAREGVIKEFT(166), which has been suggested to be a (P)APS motif, is located in a highly protease-susceptible loop that is disordered in both subunits of the free enzyme. MgATP or MgADP protects against proteolysis; APS alone has no effect but augments the protection provided by MgADP. The results suggest that the loop lacks a fixed structure until MgATP or MgADP is bound. The subsequent conformational change together with the potential change promoted by the interaction of MgATP with Asp 61 may define the APS binding site. This model is consistent with the obligatory ordered substrate binding sequence (MgATP or MgADP before APS) as established from steady state kinetics and equilibrium binding studies.
KeywordMeSH Terms
17. Chiou  SH, Hsiao  MC, Chow  LP,     ( 2000 )

Characterization of Pen n 13, a major allergen from the mold Penicillium notatum.

Biochemical and biophysical research communications 269 (1)
PMID : 10694469  :   DOI  :   10.1006/bbrc.2000.2253    
Abstract >>
Penicillium notatum is a well-known indoor aeroallergen and is frequently included in skin test panels for allergic diagnosis. On two-dimensional immunoblotting using patients' sera containing IgE and monoclonal antibody D7B8 specific for Pen c 1 of P. citrinum, two allergens with a molecular mass of 33 kDa but different isoelectric points were identified. A novel cDNA coding for Pen n 13 was cloned and sequenced. The nucleotide sequence codes for a protein 397 amino acids including a putative signal peptide of 25 amino acids and a propeptide of 90 amino acids. The allergen is an alkaline serine protease that shares more than 39% identical residues with other kinds of mold allergens. The coding cDNA of Pen n 13 was cloned into vector pQE-30 and expressed in E. coli M15 as a His-tag fusion protein and purified to homogeneity. The fusion protein reacted with monoclonal antibodies of Pen c 1 and with IgE from Penicillium-allergic patients. Furthermore, it also cross-reacted strongly with IgE specific for the natural Pen c 1, indicating that similar IgE binding epitopes may exist in the allergens of P. notatum and P. citrinum. Antigenicity index plots indicated that there are several similar epitope regions of high antigenic indices in Pen c 1 and Pen n 13, corroborating that mold allergens belonging to the alkaline serine protease family possess similar protein structure and strong antigenic cross-reactivity.
KeywordMeSH Terms
18. Vos  A, van der Lende  TR, Schuurs  TA, Pizzinini  E, van de Kamp  M,     ( 1999 )

Sulfate transport in Penicillium chrysogenum: cloning and characterization of the sutA and sutB genes.

Journal of bacteriology 181 (23)
PMID : 10572125  :   PMC  :   PMC103684    
Abstract >>
In industrial fermentations, Penicillium chrysogenum uses sulfate as the source of sulfur for the biosynthesis of penicillin. By a PCR-based approach, two genes, sutA and sutB, whose encoded products belong to the SulP superfamily of sulfate permeases were isolated. Transformation of a sulfate uptake-negative sB3 mutant of Aspergillus nidulans with the sutB gene completely restored sulfate uptake activity. The sutA gene did not complement the A. nidulans sB3 mutation, even when expressed under control of the sutB promoter. Expression of both sutA and sutB in P. chrysogenum is induced by growth under sulfur starvation conditions. However, sutA is expressed to a much lower level than is sutB. Disruption of sutB resulted in a loss of sulfate uptake ability. Overall, the results show that SutB is the major sulfate permease involved in sulfate uptake by P. chrysogenum.
KeywordMeSH Terms
Anion Transport Proteins
19. Rodríguez  M, Bernasconi  E, Mellado  E, Díez  B,     ( 1999 )

The NADP-dependent glutamate dehydrogenase gene from Penicillium chrysogenum and the construction of expression vectors for filamentous fungi.

Applied microbiology and biotechnology 52 (2)
PMID : 10499259  :  
Abstract >>
The gdhA gene encoding the NADP-dependent glutamate dehydrogenase activity from Penicillium chrysogenum has been isolated and characterized for its use in gene expression. The nucleotide sequence of a 2816-bp genomic fragment was determined, showing an open reading frame of 1600 bp interrupted by two introns, of 160 bp and 57 bp respectively, with fungal consensus splice-site junctions. The predicted amino acid sequence revealed a high degree of identity to glutamate dehydrogenase enzymes, especially to those from the fungi Aspergillus nidulans (82%) and Neurospora crassa (78%). The gdhA gene was found to be present in a single copy in the genome of several P. chrysogenum strains with different penicillin productivity. The use of the gdhA promoter for homologous and heterologous gene expression in fungi and Escherichia coli was analyzed. Heterologous gene expression was ascertained by the construction of gene fusions with the lacZ gene from E. coli and the bleomycin-resistance determinant (bleR) from Streptoalloteichus hindustanus. Homologous gene expression was shown through the use of the penicillin-biosynthetic genes pchC and penDE from P. chrysogenum and the cephalosporin biosynthetic genes cefEF and cefG from Acremonium chrysogenum.
KeywordMeSH Terms
Genes, Fungal
Genetic Vectors
20. Han  SH, Huang  MH, Tzean  SS, Shen  HD, Wang  SR, Tam  MF,     ( 1999 )

Characterization of allergens from Penicillium oxalicum and P. notatum by immunoblotting and N-terminal amino acid sequence analysis.

Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology 29 (5)
PMID : 10231324  :   DOI  :   10.1046/j.1365-2222.1999.00509.x    
Abstract >>
Penicillium species are important causative agents of extrinsic bronchial asthma. However, little is known about the allergens of these ubiquitous fungal species. Objective The object was to analyse the composition, the allergenic cross-reactivity and the N-terminal sequences of allergens from two prevalent airborne Penicillium species, P. oxalicum and P. notatum. The allergenic composition and the immunoglobulin (Ig)E cross-reactivity were analysed by immunoblot and immunoblot inhibition, respectively, using sera from asthmatic patients. The N-terminal amino acid sequences of major allergens were determined by Edman degradation. Allergens identified were also characterized by immunoblotting using monoclonal antibody (MoAb) PCM39 against the alkaline serine proteinase major allergen of P. citrinum. Among the 70 asthmatic sera tested, 18 (26%) and 17 (24%) had IgE immunoblot reactivity towards components of P. oxalicum and P. notatum, respectively. Major allergens (> 80% frequency of IgE-binding) from both species are the 34 and 30 kDa proteins of P. oxalicum and the 34 and 32 kDa proteins of P. notatum. IgE cross-reactivity among these major allergens and the 33 kDa major allergen of P. citrinum can be detected by immunoblot inhibition studies. The N-terminal amino acid sequences of the 34 kDa allergen of P. oxalicum and of the 32 and the 28 kDa allergens of P. notatum share homology with sequences of the vacuolar serine proteinase from Aspergillus fumigatus. The N-terminal amino acid sequence of the 34 kDa allergen of P. notatum shows sequence homology with that of alkaline serine proteinase from P. citrinum. Results obtained from immunoblotting showed that MoAb PCM39 reacted with the 34, 30 and 16 kDa IgE-binding components of P. oxalicum, and with the 34, 32 and 28 kDa IgE-binding components of P. notatum. Results obtained suggest that the 34 kDa major allergen of P. oxalicum may be a vacuolar serine proteinase. The 34 and the 32 kDa major allergens of P. notatum may be the alkaline and the vacuolar serine proteinases of P. notatum, respectively. The 30 and 16 kDa IgE-binding components of P. oxalicum and the 28 kDa IgE-binding component of P. notatum may be breakdown products of the 34 and the 32 kDa major vacuolar serine proteinase allergens of P. oxalicum and P. notatum, respectively.
KeywordMeSH Terms
21. García-Rico  RO, Martín  JF, Fierro  F,     ( 2007 )

The pga1 gene of Penicillium chrysogenum NRRL 1951 encodes a heterotrimeric G protein alpha subunit that controls growth and development.

Research in microbiology 158 (5)
PMID : 17467244  :   DOI  :   10.1016/j.resmic.2007.03.001    
Abstract >>
The pga1 gene of Penicillium chrysogenum NRRL 1951 has been cloned and shown to participate in the developmental program of this fungus. It encodes a protein showing a high degree of identity to group I alpha subunits of fungal heterotrimeric G proteins, presenting in its sequence all the distinctive characteristics of this group. Northern analysis revealed that pga1 is highly expressed in a constitutive manner in submerged cultures, while its expression changes during development on solid media cultures; it is higher during vegetative growth and decreases significantly at the time of conidiogenesis. Attenuation of pga1 gene expression by antisense RNA, and mutations of pga1 resulting in a constitutively activated (pga1G42R allele) or constitutively inactivated (pga1G203R allele) Pga1 alpha subunit were used to study the function of Pga1 in P. chrysogenum. The phenotype of transformants expressing the antisense construction and the mutant alleles showed substantial morphological differences in colony diameter and conidiation, indicating that Pga1 controls apical extension and negatively regulates conidiogenesis on solid medium, but has no effect on submerged cultures. Pga1 is also functional in Penicillium roqueforti, controlling the same processes.
KeywordMeSH Terms
22. Masuda  N, Kitamura  N, Saito  K,     ( 1991 )

Primary structure of protein moiety of Penicillium notatum phospholipase B deduced from the cDNA.

European journal of biochemistry 202 (3)
PMID : 1722456  :   DOI  :   10.1111/j.1432-1033.1991.tb16433.x    
Abstract >>
Phospholipase B has not yet been well defined. The most important points about this enzyme are its relationships with lysophospholipase and phospholipase A1. As reported [Saito, K., Sugatani, J. & Okumura, T. (1991) Methods Enzymol. 197, 446-456], Penicillium notatum phospholipase B is a glycoprotein with a molecular mass of 95 kDa and intrinsic lysophospholipase and phospholipase B activities; however, by endogenous proteolytic modification, its phospholipase B activity is lost almost completely, whereas its lysophospholipase activity remains unchanged. A cDNA library of P. notatum was screened by hybridization with two synthetic oligodeoxyribonucleotide probes, which corresponds to two different pentapeptides of the enzyme. A hybridization-positive clone, pPLB18, was isolated and its nucleotide sequence was determined. The deduced amino acid sequence was quite different from that found previously. Therefore, we rescreened the cDNA library with a Sau3AI fragment derived from pPLB18 and isolated a new clone, pPLB15. Comparison of the nucleotide sequences of pPLB15 and pPLB18 revealed that pPLB18 contained an insertion sequence of 53 bp. Consequently, the reading frame was open downstream for 603 amino acid residues. From the assigned sequence, it was deduced that the limited proteolysis occurred between Leu175 and Asp176; eight cysteine residues and 16 potential N-glycosylation sites were also found. No amino acid sequence similarity was found with other proteins, including other phospholipases.
KeywordMeSH Terms
23. Hou  Y, Wang  T, Long  H, Zhu  H,     ( 2007 )

Cloning, sequencing and expression analysis of the first cellulase gene encoding cellobiohydrolase 1 from a cold-adaptive Penicillium chrysogenum FS010.

Acta biochimica et biophysica Sinica 39 (2)
PMID : 17277884  :   DOI  :   10.1111/j.1745-7270.2007.00260.x    
Abstract >>
A cellobiohydrolase 1 gene (cbh1) was cloned from Penicillium chrysogenum FS010 by a modified thermal asymmetric interlaced polymerase chain reaction (TAIL-PCR). DNA sequencing shows that cbh1 has an open reading frame of 1590 bp, encoding a putative protein of 529 amino acid residues. The deduced amino acid sequence revealed that CBHI has a modular structure with a predicted molecular mass of 56 kDa and consists of a fungal type carbohydrate binding module separated from a catalytic domain by a threonine rich linker region. The putative gene product is homologous to fungal cellobiohydrolases in Family 7 of the glycosyl hydrolases. A novel cbh1 promoter (1.3 kb) was also cloned and sequenced, which contains seven putative binding sites (5'-SYGGRG-3') for the carbon catabolite repressor CRE1. Effect of various carbon sources to the cbh1 transcription of P. chrysogenum was examined by Northern analysis, suggesting that the expression of cbh1 is regulated at transcriptional level. The cbh1 gene in cold-adaptive fungus P. chysogenum was expressed as an active enzyme in Saccharomyces cerevisiae H158. The recombinant CBHI accumulated intracellularly and could not be secreted into the medium.
KeywordMeSH Terms
24. Meijer  WH, van der Klei  IJ, Veenhuis  M, Kiel  JA,     ( N/A )

ATG genes involved in non-selective autophagy are conserved from yeast to man, but the selective Cvt and pexophagy pathways also require organism-specific genes.

Autophagy 3 (2)
PMID : 17204848  :   DOI  :   10.4161/auto.3595    
Abstract >>
ATG genes encode proteins that are required for macroautophagy, the Cvt pathway and/or pexophagy. Using the published Atg protein sequences, we have screened protein and DNA databases to identify putative functional homologs (orthologs) in 21 fungal species (yeast and filamentous fungi) of which the genome sequences were available. For comparison with Atg proteins in higher eukaryotes, also an analysis of Arabidopsis thaliana and Homo sapiens databases was included. This analysis demonstrated that Atg proteins required for non-selective macroautophagy are conserved from yeast to man, stressing the importance of this process in cell survival and viability. The A. thaliana and human genomes encode multiple proteins highly similar to specific fungal Atg proteins (paralogs), possibly representing cell type-specific isoforms. The Atg proteins specifically involved in the Cvt pathway and/or pexophagy showed poor conservation, and were generally not present in A. thaliana and man. Furthermore, Atg19, the receptor of Cvt cargo, was only detected in Saccharomyces cerevisiae. Nevertheless, Atg11, a protein that links receptor-bound cargo (peroxisomes, the Cvt complex) to the autophagic machinery was identified in all yeast species and filamentous fungi under study. This suggests that in fungi an organism-specific form of selective autophagy may occur, for which specialized Atg proteins have evolved.
KeywordMeSH Terms
Conserved Sequence
Genes, Fungal
25. Kiel  JA, Veenhuis  M, van der Klei  IJ,     ( 2006 )

PEX genes in fungal genomes: common, rare or redundant.

Traffic (Copenhagen, Denmark) 7 (10)
PMID : 16978390  :   DOI  :   10.1111/j.1600-0854.2006.00479.x    
Abstract >>
PEX genes encode proteins, termed peroxins, that are required for the biogenesis and proliferation of microbodies (peroxisomes). We have screened the available protein and DNA databases to identify putative peroxin orthologs in 17 fungal species (yeast and filamentous fungi) and in humans. This analysis demonstrated that most peroxins are present in all fungi under study. Only Pex16p is absent in most yeast species, with the exception of Yarrowia lipolytica, but this peroxin is present in all filamentous fungi. Furthermore, we found that the Y. lipolytica PEX9 gene, a putative orphan gene, might encode a Pex26p ortholog. In addition, in the genomes of Saccharomyces cerevisiae and Candida glabrata, several PEX genes appear to have been duplicated, exemplified by the presence of paralogs of the peroxins Pex5p and Pex21p, which were absent in other organisms. In all organisms, we observed multiple paralogs of the peroxins involved in organelle proliferation. These proteins belong to two groups of peroxins that we propose to designate the Pex11p and Pex23p families. This redundancy may complicate future studies on peroxisome biogenesis and proliferation in fungal species.
KeywordMeSH Terms
Genome, Fungal
26. Benito  MJ, Connerton  IF, Córdoba  JJ,     ( 2006 )

Genetic characterization and expression of the novel fungal protease, EPg222 active in dry-cured meat products.

Applied microbiology and biotechnology 73 (2)
PMID : 16941178  :   DOI  :   10.1007/s00253-006-0498-z    
Abstract >>
EPg222 protease is a novel extracellular enzyme produced by Penicillium chrysogenum (Pg222) isolated from dry-cured hams that has the potential for use over a broad range of applications in industries that produce dry-cured meat products. The gene encoding EPg222 protease has been identified. Peptide sequences of EPg222 were obtained by de novo sequencing of tryptic peptides using mass spectrometry. The corresponding gene was amplified by PCR using degenerated primers based on a combination of conserved serine protease-encoding sequences and reverse translation of the peptide sequences. EPg222 is encoded as a gene of 1,361 bp interrupted by two introns. The deduced amino acid sequence indicated that the enzyme is synthesized as a preproenzyme with a putative signal sequence of 19 amino acids (aa), a prosequence of 96 aa and a mature protein of 283 aa. A cDNA encoding EPg222 has been cloned and expressed as a functionally active enzyme in Pichia pastoris. The recombinant enzyme exhibits similar activities to the native enzyme against a wide range of protein substrates including muscle myofibrillar protein. The mature sequence contains conserved aa residues characteristic of those forming the catalytic triad of serine proteases (Asp42, His76 and Ser228) but notably the food enzyme exhibits specific aa substitutions in the immunoglobulin-E recognition regions that have been identified in protein homologues that are allergenic.
KeywordMeSH Terms
27. Fierro  F, García-Estrada  C, Castillo  NI, Rodríguez  R, Velasco-Conde  T, Martín  JF,     ( 2006 )

Transcriptional and bioinformatic analysis of the 56.8 kb DNA region amplified in tandem repeats containing the penicillin gene cluster in Penicillium chrysogenum.

Fungal genetics and biology : FG & B 43 (9)
PMID : 16713314  :   DOI  :   10.1016/j.fgb.2006.03.001    
Abstract >>
High penicillin-producing strains of Penicillium chrysogenum contain 6-14 copies of the three clustered structural biosynthetic genes, pcbAB, pcbC, and penDE [Barredo, J.L., D?ez, B., Alvarez, E., Mart?n, J.F., 1989. Large amplification of a 35-kb DNA fragment carrying two penicillin biosynthetic genes in high penicillin producing strains of Penicillium chrysogenum. Curr. Genet. 16, 453-459; Smith, D.J., Bull, J.H., Edwards, J., Turner, G., 1989. Amplification of the isopenicillin N synthetase gene in a strain of Penicillium chrysogenum producing high levels of penicillin. Mol. Gen. Genet. 216, 492-497.] . The cluster is located in a 56.8 kb DNA region bounded by a conserved TGTAAA/T hexanucleotide that undergoes amplification in tandem repeats [Fierro, F., Barredo, J.L., D?ez, B., Guti?rrez, S., Fern?ndez, F.J., Mart?n, J.F., 1995. The penicillin gene cluster is amplified in tandem repeats linked by conserved hexanucleotide sequences. Proc. Natl. Acad. Sci. USA 92, 6200-6204; Newbert, R.W., Barton, B., Greaves, P., Harper, J., Turner, G., 1997. Analysis of a commercially improved Penicillium chrysogenum strain series: involvement of recombinogenic regions in amplification and deletion of the penicillin biosynthesis gene cluster. J. Ind. Microbiol. Biotechnol. 19, 18-27]. Transcriptional analysis of this amplified region (AR) revealed the presence of at least eight transcripts expressed in penicillin producing conditions. Three of them correspond to the known penicillin biosynthetic genes, pcbAB, pcbC, and penDE. To locate genes related to penicillin precursor formation, or penicillin transport and regulation we have sequenced and analyzed the 56.8 kb amplified region of P. chrysogenum AS-P-78, finding a total of 16 open reading frames. Two of these ORFs have orthologues of known function in the databases. Other ORFs showed similarities to specific domains occurring in different proteins and superfamilies which allowed to infer their probable function. ORF11 encodes a D-amino acid oxidase that might be responsible for the conversion of D-amino acids in the tripeptide L-alpha-aminoadipyl-L-cysteinyl-D-valine or other beta-lactam intermediates to deaminated by-products. ORF12 encodes a predicted protein with similarity to saccharopine dehydrogenases that seems to be related to biosynthesis of the penicillin precursor alpha-aminoadipic acid. A deletion mutant, P. chrysogenum npe10 lacking the entire AR including ORF12, shows a partial requirement of L-lysine for growth. ORF13 encodes a putative protein containing a Zn(II)2-Cys6 fungal-type DNA-binding domain, probably a transcriptional regulator. Although some of the ORFs in the AR may play roles in increasing penicillin production, none of the 13 ORFs other than pcbAB, pcbC, and penDE seem to be strictly indispensable for penicillin biosynthesis. The genes located in the P. chrysogenum AR have been compared with those found in the Aspergillus nidulans 50 kb DNA region adjacent to the penicillin gene cluster, showing no conservation between these two fungi.
KeywordMeSH Terms
Multigene Family
Transcription, Genetic
28. Frisvad  JC, Larsen  TO, Dalsgaard  PW, Seifert  KA, Louis-Seize  G, Lyhne  EK, Jarvis  BB, Fettinger  JC, Overy  DP,     ( 2006 )

Four psychrotolerant species with high chemical diversity consistently producing cycloaspeptide A, Penicillium jamesonlandense sp. nov., Penicillium ribium sp. nov., Penicillium soppii and Penicillium lanosum.

International journal of systematic and evolutionary microbiology 56 (Pt 6)
PMID : 16738124  :   DOI  :   10.1099/ijs.0.64160-0    
Abstract >>
Penicillium jamesonlandense is a novel species from Greenland that grows exceptionally slowly at 25 degrees C and has an optimum temperature for growth of 17-18 degrees C. The novel species is more psychrotolerant than any other Penicillium species described to date. Isolates of this novel species produce a range of secondary metabolites with a high chemical diversity, represented by kojic acid, penicillic acid, griseofulvin, pseurotin, chrysogine, tryptoquivalins and cycloaspeptide. Penicillium ribium, another novel psychrotolerant species from the Rocky Mountains, Wyoming, USA, produces asperfuran, kojic acid and cycloaspeptide. Originally reported from an unidentified Aspergillus species isolated from Nepal, cycloaspeptide A is reported here for the first time from the two novel Penicillium species and two known psychrotolerant species with high chemical diversity, Penicillium soppii and Penicillium lanosum. All species, except P. ribium, produce a combination of cycloaspeptide and griseofulvin. However, P. ribium (3/5 strains) produced the precursor to griseofulvin, norlichexanthone. The type strain of Penicillium jamesonlandense sp. nov. is DAOM 234087(T) (=IBT 21984(T) = IBT 24411(T) = CBS 102888(T)) and the type strain of Penicillium ribium sp. nov. is DAOM 234091(T) (=IBT 16537(T) = IBT 24431(T)).
KeywordMeSH Terms
29. Wang  L, Zhuang  WY,     ( 2007 )

Phylogenetic analyses of penicillia based on partial calmodulin gene sequences.

Bio Systems 88 (1��2��)
PMID : 16860929  :   DOI  :   10.1016/j.biosystems.2006.04.008    
Abstract >>
Partial sequences (about 600 nucleotides) of the calmodulin gene were used for the phylogenetic studies on Eupenicillium, Talaromyces and Penicillium. This region is from the 3rd base of the codon for the 9th amino acid Gln to the 3rd base of the codon for the 122th amino acid Val, flanking parts of the 2nd and 5th exons with complete sequences of two exons and three introns. Seventy-six isolates of 56 taxa of penicillia were involved. The nucleotide sequences with and without introns were analyzed respectively using the neighbor-joining (NJ) and maximum parsimony (MP) methods. The cluster analysis on relative synonymous codon usage (RSCU) of each sequence was also carried out. The fact that species of penicillia belong to the two subfamilies of the Trichocomaceae proposed by Malloch based on traditional methods is supported by our molecular data, whereas, the development of asci and patterns of penicilli show little phylogenetic information. Nine groups in the lineage of Eupenicillium and two in that of Talaromyces were recognized in our studies. In addition to the teleomorph-holomorph-anamorph evolutionary model of penicillia suggested by LoBuglio et al., and Pitt, we proposed that a mutation bias of holomorphs/anamorphs with or without selection is another evolutionary path of these organisms.
KeywordMeSH Terms
Genes, Fungal
30. Elleuche  S, Nolting  N, Pöggeler  S,     ( 2006 )

Protein splicing of PRP8 mini-inteins from species of the genus Penicillium.

Applied microbiology and biotechnology 72 (5)
PMID : 16544141  :   DOI  :   10.1007/s00253-006-0350-5    
Abstract >>
Inteins are protein-intervening sequences found inside the coding region of different host proteins and are translated in-frame with them. They can self-excise through protein splicing, which ligates the host protein flanks with a peptide bond. In this study, four different species of the genus Penicillium were investigated for the presence of inteins inside the conserved splicing-factor protein PRP8. We identified 157 to 162 amino acid in-frame insertions in the PRP8 protein of Penicillium chrysogenum, Penicillium expansum, and Penicillium vulpinum (formerly Penicillium claviforme). The Penicillium PRP8 inteins are mini-inteins without a conserved endonuclease domain. We demonstrated that the PRP8 mini-inteins of P. chrysogenum, P. expansum, and P. vulpinum undergo autocatalytic protein splicing when heterologously expressed in E. coli, in a model host protein, and in a divided GFP model system. They are, thus, among the smallest known nuclear-encoded, active splicing protein elements. The GFP assay should be valuable as a screening system for protein splicing inhibitors as potential antimycotic agents and as tools for studying the mechanism of protein splicing of fungal mini-inteins.
KeywordMeSH Terms
31. Hou  YH, Wang  TH, Long  H, Zhu  HY,     ( 2006 )

Novel cold-adaptive Penicillium strain FS010 secreting thermo-labile xylanase isolated from Yellow Sea.

Acta biochimica et biophysica Sinica 38 (2)
PMID : 16474906  :   DOI  :   10.1111/j.1745-7270.2006.00135.x    
Abstract >>
A novel cold-adaptive xylanolytic Penicillium strain FS010 was isolated from Yellow sea sediments. The marine fungus grew well from 4 to 20 degrees; a lower (0 degrees) or higher (37 degrees) temperature limits its growth. The strain was identified as Penicillium chrysogenum. Compared with mesophilic P. chrysogenum, the cold-adaptive fungus secreted the cold-active xylanase (XYL) showing high hydrolytic activities at low temperature (2-15 degrees) and high sensitivity to high temperature (>50 degrees). The XYL gene was isolated from the cold-adaptive P. chrysogenum FS010 and designated as xyl. The deduced amino acid sequence of the protein encoded by xyl showed high homology with the sequence of glycoside hydrolase family 10. The gene was subcloned into an expression vector pGEX-4T-1 and the encoded protein was overexpressed as a fusion protein with glutathione-S-transferase in Escherichia coli BL21. The expression product was purified and subjected to enzymatic characterization. The optimal temperature and pH for recombinant XYL was 25 degrees and 5.5, respectively. Recombinant XYL showed nearly 80% of its maximal activity at 4 degrees and was active in the pH range 3.0-9.5.
KeywordMeSH Terms
Adaptation, Physiological
Cold Temperature
32. Tai  HY, Tam  MF, Chou  H, Peng  HJ, Su  SN, Perng  DW, Shen  HD,     ( 2006 )

Pen ch 13 allergen induces secretion of mediators and degradation of occludin protein of human lung epithelial cells.

Allergy 61 (3)
PMID : 16436150  :   DOI  :   10.1111/j.1398-9995.2005.00958.x    
Abstract >>
Alkaline serine proteases from six prevalent airborne Penicillium and Aspergillus species have been identified as a group of major allergens (group 13). After entering human airways, the allergens are in initial contacts with respiratory epithelial cells. The purpose of this study is to investigate interactions between the Pen ch 13 allergen from P. chrysogenum and human lung epithelial cells. A549 cells, 16HBE14o- cells and primary cultures of human bronchial epithelial cells (HBEpC) were exposed to purified Pen ch 13 and mediators released into culture supernatants were assayed with enzyme-linked immunosorbent assay (ELISA) kits. Cleavage of occludin in 16HBE14o- cells was analysed by immunofluorescent staining of whole cells and immunoblot analysis of whole cell extracts. Fragments generated by incubating Pen ch 13 and a synthetic peptide carrying the occludin sequence were analysed by mass spectrometry. Pen ch 13 induced productions of prostaglandin-E2 (PGE2), interleukin (IL)-8 and transforming growth factor (TGF)-beta1 by A549 cells, 16HBE14o- cells and primary cultures of HBEpC. The protease activity of Pen ch 13 is needed for the induction of PGE2 IL-8, TGF-beta1 and cyclo-oxygenase (COX)-2 expression. A tight junction protein occludin of 16HBE14o- cells can be cleaved by Pen ch 13 at Gln202 and Gln211 which are within the second extracellular domain of the protein. Pen ch 13 may contribute to asthma by damaging the barrier formed by the airway epithelium and stimulating the release of mediators that orchestrate local immune responses and inflammatory process from HBEpC.
KeywordMeSH Terms
Allergens
Antigens, Fungal
33. Castillo  NI, Fierro  F, Gutiérrez  S, Martín  JF,     ( 2006 )

Genome-wide analysis of differentially expressed genes from Penicillium chrysogenum grown with a repressing or a non-repressing carbon source.

Current genetics 49 (2)
PMID : 16362424  :   DOI  :   10.1007/s00294-005-0029-y    
Abstract >>
Penicillium chrysogenum is an economically important ascomycete used as industrial producer of penicillin. However, with the exception of penicillin biosynthesis genes, little attention has been paid to the genetics of other aspects of the metabolism of this fungus. In this article we describe the first attempt of systematic analysis of expressed genes in P. chrysogenum, using a suppression subtractive hybridization approach to clone and identify sequences of genes differentially expressed in media with glucose or lactose as carbon source (penicillin-repressing or non-repressing conditions). A total of 167 clones were analysed, 95 from the glucose condition and 72 from the lactose condition. Genes differentially expressed in the glucose condition encode mainly proteins involved in the mitochondrial electron transport chain and primary metabolism. Genes expressed differentially in lactose-containing medium include genes for secondary metabolism (pcbC, isopenicillin N synthase), different hydrolases and a gene encoding a putative hexose transporter or sensor. The results provided information on how the metabolism of this fungus adapts to different carbon sources. The expression patterns of some of the genes support the hypothesis that glucose induces higher rates of respiration in P. chrysogenum while repressing secondary metabolism.
KeywordMeSH Terms
Gene Expression Profiling
Genome, Fungal
34. Naranjo  L, Lamas-Maceiras  M, Ullán  RV, Campoy  S, Teijeira  F, Casqueiro  J, Martín  JF,     ( 2005 )

Characterization of the oat1 gene of Penicillium chrysogenum encoding an omega-aminotransferase: induction by L-lysine, L-ornithine and L-arginine and repression by ammonium.

Molecular genetics and genomics : MGG 274 (3)
PMID : 16163487  :   DOI  :   10.1007/s00438-005-0019-2    
Abstract >>
The Penicillium chrysogenum oat1 gene, which encodes a class III omega-aminotransferase, was cloned and characterized. This enzyme converts lysine into 2-aminoadipic semialdehyde, and plays an important role in the biosynthesis of 2-aminoadipic acid, a precursor of penicillin and other beta-lactam antibiotics. The enzyme is related to ornithine-5-aminotransferases and to the lysine-6-aminotransferases encoded by the lat genes found in bacterial cephamycin gene clusters. Expression of oat1 is induced by lysine, ornithine and arginine, and repressed by ammonium ions. AreA-binding GATA and GATT sequences involved in regulation by ammonium, and an 8-bp direct repeat associated with arginine induction in Emericella (Aspergillus nidulans and Saccharomyces cerevisiae, were found in the oat1 promoter region. Deletion of the oat1 gene resulted in the loss of omega-aminotransferase activity. The null mutants were unable to grow on ornithine or arginine as sole nitrogen sources and showed reduced growth on lysine. Complementation of the null mutant with the oat1 gene restored normal levels of omega-aminotransferase activity and the ability to grow on ornithine, arginine and lysine. The role of the oat1 gene in the biosynthesis of 2-aminoadipic acid is discussed.
KeywordMeSH Terms
35. Damveld  RA, vanKuyk  PA, Arentshorst  M, Klis  FM, van den Hondel  CA, Ram  AF,     ( 2005 )

Expression of agsA, one of five 1,3-alpha-D-glucan synthase-encoding genes in Aspergillus niger, is induced in response to cell wall stress.

Fungal genetics and biology : FG & B 42 (2)
PMID : 15670714  :   DOI  :   10.1016/j.fgb.2004.11.006    
Abstract >>
1,3-alpha-D-Glucan is an important component of the cell wall of filamentous fungi. We have identified a family of five 1,3-alpha-D-glucan synthase-encoding genes in Aspergillus niger. The agsA gene was sequenced and the predicted protein sequence indicated that the overall domain structure of 1,3-alpha-D-glucan synthases is conserved in fungi. Using RT-PCR and Northern blot analysis, we found that expression of the agsA gene and to a lesser extent also of agsE were induced in the presence of the cell wall stress-inducing compounds such as Calcofluor White (CFW), SDS, and caspofungin. Loss of agsA function did not result in an apparent phenotype under normal growth conditions but rendered the cells more sensitive to CFW. The induction of 1,3-alpha-D-glucan synthase-encoding genes in response to cell wall stress was not limited to A. niger, but was also observed in Penicillium chrysogenum. We propose that this response to cell wall stress commonly occurs in filamentous fungi.
KeywordMeSH Terms
36. Kiel  JA, van der Klei  IJ, van den Berg  MA, Bovenberg  RA, Veenhuis  M,     ( 2005 )

Overproduction of a single protein, Pc-Pex11p, results in 2-fold enhanced penicillin production by Penicillium chrysogenum.

Fungal genetics and biology : FG & B 42 (2)
PMID : 15670713  :   DOI  :   10.1016/j.fgb.2004.10.010    
Abstract >>
Current industrial production of beta-lactam antibiotics, using the filamentous fungus Penicillium chrysogenum, is the result of many years of strain improvement by classical mutagenesis. More efficient production strains showed significant increases in the number and volume fraction of microbodies in their cells, organelles that harbor key enzymes involved in the biosynthesis of beta-lactam antibiotics. We have isolated the P. chrysogenum cDNA encoding Pc-Pex11p, a peroxin that is involved in microbody abundance. We demonstrate that overproduction of Pc-Pex11p in P. chrysogenum results in massive proliferation of tubular-shaped microbodies and a 2- to 2.5-fold increase in the level of penicillin in the culture medium. Notably, Pc-Pex11p-overproduction did not affect the levels of the enzymes of the penicillin biosynthetic pathway. Our results suggest that the stimulating effect of enhanced organelle numbers may reflect an increase in the fluxes of penicillin and/or its precursors across the now much enlarged microbody membrane.
KeywordMeSH Terms
37. Lai  HY, Tam  MF, Chou  H, Lee  SS, Tai  HY, Shen  HD,     ( 2004 )

Molecular and structural analysis of immunoglobulin E-binding epitopes of Pen ch 13, an alkaline serine protease major allergen from Penicillium chrysogenum.

Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology 34 (12)
PMID : 15663570  :   DOI  :   10.1111/j.1365-2222.2004.02115.x    
Abstract >>
Through proteomic and genomic approaches we have previously identified and characterized an alkaline serine protease that is a major allergen (88% frequency of IgE binding) of Penicillium chrysogenum (Pen ch 13). The aim of the present study is to identify the linear IgE-binding epitopes of Pen ch 13. IgE-binding regions were identified by dot-blot immunoassay using 11 phage-displayed peptide fragments spanning the whole molecule of Pen ch 13. The minimal epitope requirements for IgE binding were further defined with overlapping peptides synthesized on derivatized cellulose membranes using SPOTs technology. The critical residues on the immunodominant epitopes were mapped through site-directed mutagenesis. The locations of the IgE epitopes identified were correlated with a three-dimensional structure of Pen ch 13. IgE antibodies in 35 serum samples reacted with at least one of the 11 peptide fragments of Pen ch 13. Peptide f-2n (residues 31-61) showed a high-intensity and the highest frequency (77%) of IgE binding. The frequencies of IgE binding to peptide f-4 (residues 93-133), f-1 (residues 1-37) and f-7 (residues 168-206) were 51%, 34% and 31%, respectively. SPOTs assay narrowed down the region of IgE binding of f-2n to residues 48-55 (GHADFGGR). Three, two and one epitope(s) that are four to nine amino acids in length, within f-4, f-1 and f-7, respectively, were found. Site-directed mutagenesis of Pen ch 13 revealed that substitution of His49 and/or Phe52 on Pen ch 13 with methionine resulted in proteins with drastic loss of IgE binding in seven sera tested. Proteins with amino acid replacements at residues 15-18 (RISS), or at residues 112 (I) and 116 (D) have lower IgE-binding reactivity in one of the two patient's sera tested. Substituting residues 117 (W), 119 (V) and 120 (K) also block most of the IgE binding in one of the two patient's sera tested. In addition, replacing residues 203 (V) and 204 (D) along with a deletion at residue 206 (Y) diminished the IgE binding in two serum samples tested. A model was constructed based on the structure of P. cyclopium subtilisin protease that has >90% (256 out of 283 amino acids) sequence identity with Pen ch 13. The major epitope (GHADFGGR) on Pen ch 13 formed a loop-like structure and was located at the surface of the allergen. Several linear IgE-reactive epitopes and their critical core amino acid residues were identified for the Pen ch 13 allergen. The major linear IgE-binding epitope, 48GHADFGGR55, formed a loop-like structure at the surface of the allergen. Substitution of His49 and/or Phe52 with methionine significantly reduced IgE-binding to Pen ch 13. Mapping of these results on a 3D model of the allergen provides valuable information about the molecular basis of allergenicity for Pen ch 13 and for designing specific immunotherapeutics.
KeywordMeSH Terms
38. Díez  B, Rodríguez-Sáiz  M, de la Fuente  JL, Moreno  MA, Barredo  JL,     ( 2005 )

The nagA gene of Penicillium chrysogenum encoding beta-N-acetylglucosaminidase.

FEMS microbiology letters 242 (2)
PMID : 15621446  :   DOI  :   10.1016/j.femsle.2004.11.017    
Abstract >>
We purified the beta-N-acetylglucosaminidase from the filamentous fungus Penicillium chrysogenum and its N-terminal sequence was determined, showing the presence of a mixture of two proteins (P1 and P2). A genomic DNA fragment was cloned by using degenerated oligonucleotides from the Nt sequences. The nucleotide sequence showed the presence of an ORF (nagA gene) lacking introns, with a length of 1791 bp, and coding for a protein of 66.5 kDa showing similarity to acetylglucosaminidases. The NagA deduced protein includes P1 and P2 as incomplete forms of the mature protein, and contains putative features for protein maturation: an 18-amino acid signal peptide, a KEX2 processing site, and four glycosylation motifs. The sequence just after the signal peptide corresponds to P2 and that after the KEX2 site to P1. The nagA transcript has a size of about 2.1 kb and is present until the end of the fermentation process for penicillin production. NagA is one of the most largely represented proteins in P. chrysogenum, increasing along the fermentation process. The suitability of the nagA promoter (PnagA) for gene expression in fungi was demonstrated by expressing the bleomycin resistance gene (ble(R)) from Streptoalloteichus hindustanus in P. chrysogenum.
KeywordMeSH Terms
39. Haas  H, Redl  B, Friedlin  E, Stöffler  G,     ( 1992 )

Isolation and analysis of the Penicillium chrysogenum phoA gene encoding a secreted phosphate-repressible acid phosphatase.

Gene 113 (1)
PMID : 1563629  :   DOI  :   10.1016/0378-1119(92)90680-n    
Abstract >>
We have isolated the genomic sequence encoding a secreted phosphate-repressible acid phosphatase (PHOA) from Penicillium chrysogenum using synthetic oligodeoxyribonucleotide probes. Nucleotide sequence data revealed that this gene consists of two exons of 192 and 1047 bp separated by an intron of 52 bp in length. A sequence encoding a putative signal peptide, resembling known signal sequences of fungi, was identified at the 5'-end of the coding sequence. Northern blot analysis of total cellular RNA indicated that the phoA gene codes for a 1.6-kb transcript. The expression of this gene is regulated at the transcriptional level and is markedly affected by the inorganic phosphate concentration of the growth medium.
KeywordMeSH Terms
Genes, Fungal
40. Trip  H, Evers  ME, Driessen  AJ,     ( 2004 )

PcMtr, an aromatic and neutral aliphatic amino acid permease of Penicillium chrysogenum.

Biochimica et biophysica acta 1667 (2)
PMID : 15581852  :   DOI  :   10.1016/j.bbamem.2004.09.014    
Abstract >>
The gene encoding an aromatic and neutral aliphatic amino acid permease of Penicillium chrysogenum was cloned, functionally expressed and characterized in Saccharomyces cerevisiae M4276. The permease, designated PcMtr, is structurally and functionally homologous to Mtr of Neurospora crassa, and unrelated to the Amino Acid Permease (AAP) family which includes most amino acid permeases in fungi. Database searches of completed fungal genome sequences reveal that Mtr type permeases are not widely distributed among fungi, suggesting a specialized function.
KeywordMeSH Terms
41. Ram  AF, Arentshorst  M, Damveld  RA, vanKuyk  PA, Klis  FM, van den Hondel  CA,     ( 2004 )

The cell wall stress response in Aspergillus niger involves increased expression of the glutamine : fructose-6-phosphate amidotransferase-encoding gene (gfaA) and increased deposition of chitin in the cell wall.

Microbiology (Reading, England) 150 (Pt 10)
PMID : 15470111  :   DOI  :   10.1099/mic.0.27249-0    
Abstract >>
Perturbation of cell wall synthesis in Saccharomyces cerevisiae, either by mutations in cell wall synthesis-related genes or by adding compounds that interfere with normal cell wall assembly, triggers a compensatory response to ensure cell wall integrity. This response includes an increase in chitin levels in the cell wall. Here it is shown that Aspergillus niger also responds to cell wall stress by increasing chitin levels. The increased chitin level in the cell wall was accompanied by increased transcription of gfaA, encoding the glutamine : fructose-6-phosphate amidotransferase enzyme, which is responsible for the first and a rate-limiting step in chitin synthesis. Cloning and disruption of the gfaA gene in A. niger showed that it was an essential gene, but that addition of glucosamine to the growth medium could rescue the deletion strain. When the plant-pathogenic fungus Fusarium oxysporum and food spoilage fungus Penicillium chrysogenum were subjected to cell wall stress, the transcript level of their gfa gene increased as well. These observations suggest that cell wall stress in fungi may generally lead to activation of the chitin biosynthetic pathway.
KeywordMeSH Terms
42. Kiel  JA, van den Berg  M, Bovenberg  RA, van der Klei  IJ, Veenhuis  M,     ( 2004 )

Penicillium chrysogenum Pex5p mediates differential sorting of PTS1 proteins to microbodies of the methylotrophic yeast Hansenula polymorpha.

Fungal genetics and biology : FG & B 41 (7)
PMID : 15275666  :   DOI  :   10.1016/j.fgb.2004.02.006    
Abstract >>
We have isolated the Penicillium chrysogenum pex5 gene encoding the receptor for microbody matrix proteins containing a type 1 peroxisomal targeting signal (PTS1). Pc-pex5 contains 2 introns and encodes a protein of approximately 75 kDa. P. chrysogenum pex5 disruptants appear to be highly unstable, show poor growth, and are unable to sporulate asexually. Furthermore, pex5 cells mislocalize a fluorescent PTS1 reporter protein to the cytosol. Pc-pex5 was expressed in a PEX5 null mutant of the yeast Hansenula polymorpha. Detailed analysis demonstrated that the PTS1 proteins dihydroxyacetone synthase and catalase were almost fully imported into microbodies. Surprisingly, alcohol oxidase, which also depends on Pex5p for import into microbodies, remained mainly in the cytosol. Thus, P. chrysogenum Pex5p has a different specificity of cargo recognition than its H. polymorpha counterpart. This was also suggested by the observation that Pc-Pex5p sorted a reporter protein fused to various functional PTS1 signals with different efficiencies.
KeywordMeSH Terms
43. Wang  L, Zhou  HB, Frisvad  JC, Samson  RA,     ( 2004 )

Penicillium persicinum, a new griseofulvin, chrysogine and roquefortine C producing species from Qinghai Province, China.

Antonie van Leeuwenhoek 86 (2)
PMID : 15280651  :   DOI  :   10.1023/B:ANTO.0000036140.86059.51    
Abstract >>
A unique Penicillium isolate from Chinese soil with terverticillate penicilli and ellipsoidal to cylindrical smooth-walled conidia, produces, in addition to the common metabolite ergosterol, copious amounts of an unknown peach-red pigment and the following secondary metabolites: griseofulvin, dechlorogriseofulvin, lichexanthone, roquefortine C, roquefortine D, chrysogine, 2-pyrovoylaminobenzamide, 2-acetyl-quinazolin-4(3H)-one. This isolate, CBS 111235, is described as Penicillium persicinum sp. nov., which belongs to subgenus Penicillium section Chrysogena but is morphologically similar to P. italicum. On the basis of the production of secondary metabolites it resembles P. griseofulvum and P. coprophilum. Sequence data using part of the beta-tubulin gene showed that it is phylogenetically related to P. chrysogenum and P. aethiopicum in section Chrysogena with which it shares both secondary metabolites and ability to grow at 37 degrees C.
KeywordMeSH Terms
44. Trip  H, Evers  ME, Kiel  JA, Driessen  AJ,     ( 2004 )

Uptake of the beta-lactam precursor alpha-aminoadipic acid in Penicillium chrysogenum is mediated by the acidic and the general amino acid permease.

Applied and environmental microbiology 70 (8)
PMID : 15294814  :   DOI  :   10.1128/AEM.70.8.4775-4783.2004     PMC  :   PMC492385    
Abstract >>
External addition of the beta-lactam precursor alpha-aminoadipic acid to the filamentous fungus Penicillium chrysogenum leads to an increased intracellular alpha-aminoadipic acid concentration and an increase in penicillin production. The exact route for alpha-aminoadipic acid uptake is not known, although the general amino acid and acidic amino acid permeases have been implicated in this process. Their corresponding genes, PcGAP1 and PcDIP5, of P. chrysogenum were cloned and functionally expressed in a mutant of Saccharomyces cerevisiae (M4276) in which the acidic amino acid and general amino acid permease genes (DIP5 and GAP1, respectively) are disrupted. Transport assays show that both PcGap1 and PcDip5 mediated the uptake of alpha-aminoadipic acid, although PcGap1 showed a higher affinity for alpha-aminoadipic acid than PcDip5 (K(m) values, 230 and 800 microM, respectively). Leucine strongly inhibits alpha-aminoadipic acid transport via PcGap1 but not via PcDip5. This difference was exploited to estimate the relative contribution of each transport system to the alpha-aminoadipic acid flux in beta-lactam-producing P. chrysogenum. The transport measurements demonstrate that both PcGap1 and PcDip5 contribute to the alpha-aminoadipic acid flux.
KeywordMeSH Terms
Gene Expression Regulation, Fungal
45. Sakamoto  T, Ihara  H, Shibano  A, Kasai  N, Inui  H, Kawasaki  H,     ( 2004 )

Molecular characterization of a Penicillium chrysogenum exo-1,5-alpha-L-arabinanase that is structurally distinct from other arabinan-degrading enzymes.

FEBS letters 560 (1��3��)
PMID : 14988022  :   DOI  :   10.1016/S0014-5793(04)00106-1    
Abstract >>
The nucleotide sequence of the abnx cDNA gene, which encodes an exo-arabinanase (Abnx) of Penicillium chrysogenum 31B, was determined. Abnx was found to be structurally distinct from known arabinan-degrading enzymes based on its amino acid sequence and a hydrophobic cluster analysis. The protein in the protein database with the highest similarity to Abnx was the Neurospora crassa conserved hypothetical protein. The abnx cDNA gene product expressed in Escherichia coli catalyzed the release of arabinobiose from alpha-1,5-L-arabinan. The activity of the recombinant Abnx towards a series of arabino-oligosaccharides, as expressed by k(cat)/K(m) value, was greatest with arabinohexaose.
KeywordMeSH Terms
46. Sugimoto  M, Saiki  Y, Zhang  D, Kawai  F,     ( 2004 )

Cloning and characterization of preferentially expressed genes in an aluminum-tolerant mutant derived from Penicillium chrysogenum IFO4626.

FEMS microbiology letters 230 (1)
PMID : 14734176  :   DOI  :   10.1016/S0378-1097(03)00886-3    
Abstract >>
cDNAs expressed preferentially in an Al-tolerant microorganism were isolated by subtraction hybridization with cDNAs of Al-sensitive Penicillium chrysogenum IFO4626 as driver cDNA and cDNAs of the Al-tolerant mutant derived from the wild cells by UV irradiation as tester cDNA. Northern blot analysis revealed that mRNA levels of six genes were increased significantly in the Al-tolerant mutant after exposure to Al stress when compared with the wild cells. Two genes accumulated in both the presence and absence of Al stress and four genes were induced by Al stress in the Al-tolerant mutant. cDNA fragments were amplified by rapid amplification of cDNA ends and sequenced to obtain full-length cDNAs of the six genes. Two genes were novel or predicted ones and the others showed significant homology to known genes, ADP/ATP translocase, enolase, cysteine synthase, and glucoamylase, which are induced by environmental stresses in prokaryotic and eukaryotic cells. These enzyme activities increased in the Al-tolerant mutant when compared to those in the wild cells, showing that not only the levels of gene expression but also the levels of enzyme activities increased in the Al-tolerant mutant.
KeywordMeSH Terms
Cloning, Molecular
Gene Expression Profiling
Mutation
47. Sogabe  Y, Kitatani  T, Yamaguchi  A, Kinoshita  T, Adachi  H, Takano  K, Inoue  T, Mori  Y, Matsumura  H, Sakamoto  T, Tada  T,     ( 2011 )

High-resolution structure of exo-arabinanase from Penicillium chrysogenum.

Acta crystallographica. Section D, Biological crystallography 67 (Pt 5)
PMID : 21543843  :   DOI  :   10.1107/S0907444911006299    
Abstract >>
Arabinanase Abnx from Penicillium chrysogenum 31B, which belongs to the GH93 family, releases arabinobiose from the nonreducing terminus of �\-1,5-L-arabinan, which is distributed in the primary cell walls of higher plants. Crystal structures of Abnx and of its complex with arabinobiose were determined at the high resolutions of 1.14 ? to an R(work) of 10.7% (R(free) = 12.8%) and 1.04 ? to an R(work) of 10.4% (R(free) = 12.5%). Abnx has a six-bladed �]-propeller fold with a typical ring-closure mode called `Velcro', in which the last four-stranded �]-sheet is completed by the incorporation of a strand from the N-terminus. Catalytic residues which act as a nucleophile and an acid/base were proposed from the structures and confirmed by site-directed mutagenesis. The substrate-binding groove is enclosed at one end by two residues, Glu64 and Tyr66, which contribute to the recognition of the nonreducing chain end of the polysaccharide. A comparison with the related enzyme Arb93A which has a quite similar overall structure suggested that Abnx has different mechanisms to funnel substrates to the active site and/or to stabilize the transition state.
KeywordMeSH Terms
48. Henk  DA, Fisher  MC,     ( 2011 )

Genetic diversity, recombination, and divergence in animal associated Penicillium dipodomyis.

PloS one 6 (8)
PMID : 21850241  :   DOI  :   10.1371/journal.pone.0022883     PMC  :   PMC3151277    
Abstract >>
Penicillium dipodomyis is thought to be an exclusively asexual fungus associated with Kangaroo Rats, Dipodomys species, and is unique among Penicillium species in growing at 37�XC but producing no known toxins. Lack of recombination within P. dipodomyis would result in limited adaptive flexibility but possibly enhance local adaptation and host selection via maintenance of favourable genotypes. Here, analysis of DNA sequence data from five protein-coding genes shows that recombination occurs within P. dipodomyis on a small spatial scale. Furthermore, detection of mating-type alleles supports outcrossing and a sexual cycle in P. dipodomyis. P. dipodomyis was a weaker competitor in in vitro assays with other Penicillium species found in association with Kanagaroo rats. Bayesian species level analysis suggests that the P. dipodomyis lineage diverged from closely related species also found in cheek pouches of Kangaroo Rats and their stored seeds about 11 million years ago, a similar divergence time as Dipodomys from its sister rodent taxa.
KeywordMeSH Terms
49. Davolos  D, Pietrangeli  B, Persiani  AM, Maggi  O,     ( 2012 )

Penicillium simile sp. nov. revealed by morphological and phylogenetic analysis.

International journal of systematic and evolutionary microbiology 62 (Pt 2)
PMID : 21460135  :   DOI  :   10.1099/ijs.0.031682-0    
Abstract >>
The morphology of three phenetically identical Penicillium isolates, collected from the bioaerosol in a restoration laboratory in Italy, displayed macro- and microscopic characteristics that were similar though not completely ascribable to Penicillium raistrickii. For this reason, a phylogenetic approach based on DNA sequencing analysis was performed to establish both the taxonomic status and the evolutionary relationships of these three peculiar isolates in relation to previously described species of the genus Penicillium. We used four nuclear loci (both rRNA and protein coding genes) that have previously proved useful for the molecular investigation of taxa belonging to the genus Penicillium at various evolutionary levels. The internal transcribed spacer region (ITS1-5.8S-ITS2), domains D1 and D2 of the 28S rDNA, a region of the tubulin beta chain gene (benA) and part of the calmodulin gene (cmd) were amplified by PCR and sequenced. Analysis of the rRNA genes and of the benA and cmd sequence data indicates the presence of three isogenic isolates belonging to a genetically distinct species of the genus Penicillium, here described and named Penicillium simile sp. nov. (ATCC MYA-4591(T) = CBS 129191(T)). This novel species is phylogenetically different from P. raistrickii and other related species of the genus Penicillium (e.g. Penicillium scabrosum), from which it can be distinguished on the basis of morphological trait analysis.
KeywordMeSH Terms
Air Microbiology
Laboratories
Phylogeny
50. Whiteman  PA, Abraham  EP, Baldwin  JE, Fleming  MD, Schofield  CJ, Sutherland  JD, Willis  AC,     ( 1990 )

Acyl coenzyme A: 6-aminopenicillanic acid acyltransferase from Penicillium chrysogenum and Aspergillus nidulans.

FEBS letters 262 (2)
PMID : 2110531  :   DOI  :   10.1016/0014-5793(90)80224-7    
Abstract >>
A study of the final stages of the biosynthesis of the penicillins in Penicillium chrysogenum has revealed two types of enzyme. One hydrolyses phenoxymethyl penicillin to 6-aminopenicillanic acid (6-APA). The other, also obtained from Aspergillus nidulans, transfers a phenylacetyl group from phenylacetyl CoA to 6-APA. The acyltransferase, purified to apparent homogeneity, had a molecular mass of 40 kDa. It also catalyses the conversion of isopenicillin N (IPN) to benzylpenicillin (Pen G) and hydrolyses IPN to 6-APA. In the presence of SDS it dissociates, with loss of activity, into fragments of ca 30 and 10.5 kDa, but activity is regained when these fragments recombine in the absence of SDS.
KeywordMeSH Terms
Penicillin-Binding Proteins
51. Tobin  MB, Fleming  MD, Skatrud  PL, Miller  JR,     ( 1990 )

Molecular characterization of the acyl-coenzyme A:isopenicillin N acyltransferase gene (penDE) from Penicillium chrysogenum and Aspergillus nidulans and activity of recombinant enzyme in Escherichia coli.

Journal of bacteriology 172 (10)
PMID : 2120195  :   DOI  :   10.1128/jb.172.10.5908-5914.1990     PMC  :   PMC526911    
Abstract >>
The final step in the biosynthesis of beta-lactam antibiotics in Penicillium chrysogenum and Aspergillus nidulans involves removal of the L-alpha-aminoadipyl side chain from isopenicillin N (IPN) and exchange with a nonpolar side chain. The enzyme catalyzing this reaction, acyl-coenzyme A:isopenicillin N acyltransferase (acyltransferase), was purified from P. chrysogenum and A. nidulans. Based on NH2-terminal amino acid sequence information, the acyltransferase gene (penDE) from P. chrysogenum and A. nidulans were cloned. In both organisms, penDE was located immediately downstream from the isopenicillin N synthetase gene (pcbC) and consisted of four exons encoding an enzyme of 357 amino acids (approximately 40 kilodaltons [kDa]). The DNA coding sequences showed approximately 73% identity, while the amino acid sequences were approximately 76% identical. Noncoding DNA regions (including the region between pcbC and penDE) were not conserved. Acyltransferase activity from Escherichia coli producing the 40-kDa protein accepted either 6-aminopenicillanic acid or IPN as the substrate and made a penicillinase-sensitive antibiotic in the presence of phenylacetyl coenzyme A. Therefore, a single gene is responsible for converting IPN to penicillin G. The active form of the enzyme may result from processing of the 40-kDa monomeric precursor to a heterodimer containing subunits of 11 and 29 kDa.
KeywordMeSH Terms
Genes, Fungal
Penicillin-Binding Proteins
52. Díez  B, Gutiérrez  S, Barredo  JL, van Solingen  P, van der Voort  LH, Martín  JF,     ( 1990 )

The cluster of penicillin biosynthetic genes. Identification and characterization of the pcbAB gene encoding the alpha-aminoadipyl-cysteinyl-valine synthetase and linkage to the pcbC and penDE genes.

The Journal of biological chemistry 265 (27)
PMID : 2129535  :  
Abstract >>
Penicillium chrysogenum DNA fragments cloned in EMBL3 or cosmid vectors from the upstream region of the pcbC-penDE cluster carry a gene (pcbAB) that complemented the deficiency of alpha-aminoadipyl-cysteinyl-valine synthetase of mutants npe5 and npe10, and restored penicillin production to mutant npe5. A protein of about 250 kDa was observed in sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels of cell-free extracts of complemented strains that was absent in the npe5 and npe10 mutants but exists in the parental strain from which the mutants were obtained. Transcriptional mapping studies showed the presence of one long transcript of about 11.5 kilobases that hybridized with several probes internal to the pcbAB gene, and two small transcripts of 1.15 kilobases that hybridized with the pcbC or the penDE gene, respectively. The transcription initiation and termination regions of the pcbAB gene were mapped by hybridization with several small probes. The region has been completely sequenced. It includes an open reading frame of 11,376 nucleotides that encodes a protein with a deduced Mr of 425,971. Three repeated dominia were found in the alpha-aminoadipyl-cysteinyl-valine synthetase which have high homology with the gramicidin synthetase I and tyrocidine synthetase I. The pcbAB is linked to the pcbC and penDE genes and is transcribed in the opposite orientation to them.
KeywordMeSH Terms
Genes, Fungal
Multigene Family
53. Scott  J, Untereiner  WA, Wong  B, Straus  NA, Malloch  D,     ( N/A )

Genotypic variation in Penicillium chysogenum from indoor environments.

Mycologia 96 (5)
PMID : 21148929  :  
Abstract >>
We examined 198 isolates of P. chysogenum recovered from 109 houses in Wallaceburg, Ontario, and 25 culture collection isolates including seven ex-type strains. Multilocus genotypes were determined by heteroduplex mobility assay of regions spanning introns in acetyl co-enzyme A synthase, beta-tubulin, thioredoxin reductase and the internal transcribed spacer regions of the nuclear ribosomal subrepeat. Five unique multilocus haplotypes were revealed without evidence of recombination, indicating strictly clonal population structures. Phylogenetic analysis of allele sequences using maximum parsimony resolved three strongly supported lineages. The dominant clade included more than 90% of house isolates in addition to the notable laboratory contaminant isolated by Alexander Fleming in 1929 in Britain. A second clade contained more than 5% of house isolates clustered with the ex-type strains of P. chysogenum and P. notatum. Follow-up sampling of outdoor air in the locality failed to reveal P. chysogenum, confirming the rarity of this fungus in outdoor air.
KeywordMeSH Terms
54. Sakamoto  T, Ogura  A, Inui  M, Tokuda  S, Hosokawa  S, Ihara  H, Kasai  N,     ( 2011 )

Identification of a GH62 �\-L-arabinofuranosidase specific for arabinoxylan produced by Penicillium chrysogenum.

Applied microbiology and biotechnology 90 (1)
PMID : 21181156  :   DOI  :   10.1007/s00253-010-2988-2    
Abstract >>
An arabinoxylan arabinofuranohydrolase (AXS5) was purified from the culture filtrate of Penicillium chrysogenum 31B. A cDNA encoding AXS5 (axs5) was isolated by in vitro cloning using the N-terminal amino acid sequence of the native enzyme as a starting point. The deduced amino acid sequence of the axs5 gene has high similarities with those of arabinoxylan arabinofuranohydrolases of Aspergillus niger, Aspergillus tubingensis, and Aspergillus sojae. Module sequence analysis revealed that a "Glyco_hydro_62" was present at position 28-299 of AXS5. This is a family of �\-L-arabinofuranosidases which are all members of glycoside hydrolase family 62. Recombinant AXS5 (rAXS5) expressed in Escherichia coli was highly active on arabinoxylan but not on branched sugar beet arabinan. (1)H-NMR analysis revealed that the rAXS5 cleaved arabinosyl side-chains linked to C-2 and C-3 of single-substituted xylose residues in arabinoxylan. Semi-quantitative RT-PCR analysis indicated that expression of the axs5 gene in P. chrysogenum 31B was strongly induced by adding D-xylose and arabinoxylan to the culture medium. Moreover, two binding sites of XlnR, a transcriptional activator that regulates the expression of the genes encoding xylanolytic enzymes, are present in the upstream region of the axs5 gene. These results suggest that AXS5 is involved in xylan degradation.
KeywordMeSH Terms
55. Smith  DJ, Earl  AJ, Turner  G,     ( 1990 )

The multifunctional peptide synthetase performing the first step of penicillin biosynthesis in Penicillium chrysogenum is a 421,073 dalton protein similar to Bacillus brevis peptide antibiotic synthetases.

The EMBO journal 9 (9)
PMID : 2118102  :   PMC  :   PMC551982    
Abstract >>
The nucleotide sequence of the Penicillium chrysogenum Oli13 acvA gene encoding delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase, which performs the first step in penicillin biosynthesis, has been determined. The acvA gene contains an open reading frame of 11,238 bp encoding a protein of 3746 amino acids with a predicted mol. wt of 421,073 dalton. Three domains within the protein of approximately 570 amino acids have between 38% and 43% identity with each other and share similarity with two antibiotic peptide synthetases from Bacillus brevis as well as two other enzymes capable of performing ATP-pyrophosphate exchange reactions. The acvA gene is located close to the pcbC gene encoding isopenicillin N synthetase, the enzyme for the second step of beta-lactam biosynthesis, and is transcribed in the opposite orientation to it. The intergenic region of 1107 bp from which the acvA and pcbC genes are divergently transcribed has also been sequenced.
KeywordMeSH Terms
Genes, Fungal
56. Glenn  AE, Karagianni  EP, Ulndreaj  A, Boukouvala  S,     ( 2010 )

Comparative genomic and phylogenetic investigation of the xenobiotic metabolizing arylamine N-acetyltransferase enzyme family.

FEBS letters 584 (14)
PMID : 20621844  :   DOI  :   10.1016/j.febslet.2010.05.063    
Abstract >>
Arylamine N-acetyltransferases (NATs) are xenobiotic metabolizing enzymes characterized in several bacteria and eukaryotic organisms. We report a comprehensive phylogenetic analysis employing an exhaustive dataset of NAT-homologous sequences recovered through inspection of 2445 genomes. We describe the first NAT homologues in viruses, archaea, protists, many fungi and invertebrates, providing complete annotations in line with the consensus nomenclature. Contrary to the NAT genes of vertebrates, introns are commonly found within the homologous coding regions of lower eukaryotes. The NATs of fungi and higher animals are distinctly monophyletic, but evidence supports a mixed phylogeny of NATs among bacteria, protists and possibly some invertebrates.
KeywordMeSH Terms
57. Rodríguez-Martín  A, Acosta  R, Liddell  S, Núñez  F, Benito  MJ, Asensio  MA,     ( 2010 )

Characterization of the novel antifungal protein PgAFP and the encoding gene of Penicillium chrysogenum.

Peptides 31 (4)
PMID : 19914321  :   DOI  :   10.1016/j.peptides.2009.11.002    
Abstract >>
The strain RP42C from Penicillium chrysogenum produces a small protein PgAFP that inhibits the growth of some toxigenic molds. The molecular mass of the protein determined by electrospray ionization mass spectrometry (ESI-MS) was 6 494Da. PgAFP showed a cationic character with an estimated pI value of 9.22. Upon chemical and enzymatic treatments of PgAFP, no evidence for N- or O-glycosylations was obtained. Five partial sequences of PgAFP were obtained by Edman degradation and by ESI-MS/MS after trypsin and chymotrypsin digestions. Using degenerate primers from these peptide sequences, a segment of 70bp was amplified by PCR from pgafp gene. 5'- and 3'-ends of pgafp were obtained by RACE-PCR with gene-specific primers designed from the 70bp segment. The complete pgafp sequence of 404bp was obtained using primers designed from 5'- and 3'-ends. Comparison of genomic and cDNA sequences revealed a 279bp coding region interrupted by two introns of 63 and 62bp. The precursor of the antifungal protein consists of 92 amino acids and appears to be processed to the mature 58 amino acids PgAFP. The deduced amino acid sequence of the mature protein shares 79% identity to the antifungal protein Anafp from Aspergillus niger. PgAFP is a new protein that belongs to the group of small, cysteine-rich, and basic proteins with antifungal activity produced by ascomycetes. Given that P. chrysogenum is regarded as safe mold commonly found in foods, PgAFP may be useful to prevent growth of toxigenic molds in food and agricultural products.
KeywordMeSH Terms
58. Batta  G, Barna  T, Gáspári  Z, Sándor  S, Kövér  KE, Binder  U, Sarg  B, Kaiserer  L, Chhillar  AK, Eigentler  A, Leiter  E, Hegedüs  N, Pócsi  I, Lindner  H, Marx  F,     ( 2009 )

Functional aspects of the solution structure and dynamics of PAF--a highly-stable antifungal protein from Penicillium chrysogenum.

The FEBS journal 276 (10)
PMID : 19459942  :   DOI  :   10.1111/j.1742-4658.2009.07011.x     PMC  :   PMC4290664    
Abstract >>
Penicillium antifungal protein (PAF) is a promising antimycotic without toxic effects on mammalian cells and therefore may represent a drug candidate against the often lethal Aspergillus infections that occur in humans. The pathogenesis of PAF on sensitive fungi involves G-protein coupled signalling followed by apoptosis. In the present study, the solution structure of this small, cationic, antifungal protein from Penicillium chrysogenum is determined by NMR. We demonstrate that PAF belongs to the structural classification of proteins fold class of its closest homologue antifungal protein from Aspergillus giganteus. PAF comprises five beta-strands forming two orthogonally packed beta-sheets that share a common interface. The ambiguity in the assignment of two disulfide bonds out of three was investigated by NMR dynamics, together with restrained molecular dynamics calculations. The clue could not be resolved: the two ensembles with different disulfide patterns and the one with no S-S bond exhibit essentially the same fold. (15)N relaxation dispersion and interference experiments did not reveal disulfide bond rearrangements via slow exchange. The measured order parameters and the 3.0 ns correlation time are appropriate for a compact monomeric protein of this size. Using site-directed mutagenesis, we demonstrate that the highly-conserved and positively-charged lysine-rich surface region enhances the toxicity of PAF. However, the binding capability of the oligosaccharide/oligonucleotide binding fold is reduced in PAF compared to antifungal protein as a result of less solvent-exposed aromatic regions, thus explaining the absence of chitobiose binding. The present study lends further support to the understanding of the documented substantial differences between the mode of action of two highly homologous antifungal proteins.
KeywordMeSH Terms
59. Acosta  R, Rodríguez-Martín  A, Martín  A, Núñez  F, Asensio  MA,     ( 2009 )

Selection of antifungal protein-producing molds from dry-cured meat products.

International journal of food microbiology 135 (1)
PMID : 19683356  :   DOI  :   10.1016/j.ijfoodmicro.2009.07.020    
Abstract >>
To control unwanted molds in dry-cured meats it is necessary to allow the fungal development essential for the desired characteristics of the final product. Molds producing antifungal proteins could be useful to prevent hazards due to the growth of mycotoxigenic molds. The objective has been to select Penicillium spp. that produce antifungal proteins against toxigenic molds. To obtain strains adapted to these products, molds were isolated from dry-cured ham. A first screening with 281 isolates by the radial inhibition assay revealed that 166 were active against some of the toxigenic P. echinulatum, P. commune, and Aspergillusniger used as reference molds. The activity of different extracts from cultured medium was evaluated by a microspectroscopic assay. Molds producing active chloroform extracts were eliminated from further consideration. A total of 16 Penicillium isolates were screened for antifungal activity from both cell-free media and the aqueous residues obtained after chloroform extraction. The cell-free media of 10 isolates that produced a strong inhibition of the three reference molds were fractionated by FPLC on a cationic column. For protein purification, the fractions of the three molds that showed high inhibitory activity were further chromatographed on a gel filtration column, and the subfractions containing the highest absorbance peaks were assayed against the most sensitive reference molds. One subfraction each from strains AS51D and RP42C from Penicilliumchrysogenum confirmed the inhibitory activity against the reference molds. SDS-PAGE revealed a single band from each subfraction, with estimated molecular masses of 37kDa for AS51D and 9kDa for RP42C. Although further characterisation is required, both these proteins and the producing strains can be of interest to control unwanted molds on foods.
KeywordMeSH Terms
60. Cheng  TT, Tam  MF, Chou  H, Tai  HY, Shen  HD,     ( 2008 )

Lys89, Lys90, and Phe91 are critical core amino acid residues of the Pen ch 18 major fungal allergen recognized by human IgE antibodies.

Biochemical and biophysical research communications 375 (4)
PMID : 18760997  :   DOI  :   10.1016/j.bbrc.2008.08.097    
Abstract >>
A vacuolar serine protease (Pen ch 18) has been identified as a major allergen of Penicillium chrysogenum. The molecular features of antigenic determinant(s) on Pen ch 18 recognized by human IgE antibodies, however, have remained unclear. Here, we show that a dominant IgE epitope on the N-terminally processed Pen ch 18 allergen was narrowed down to residues 83-91. In addition, Lys89, Lys90, and possibly Phe91 were identified as the core residues. Substitution of Lys89, Lys90, or Phe91 with alanine can significantly reduce IgE-binding to Pen ch 18. Immunoblot inhibition confirmed that Lys89 and Phe91 played a significant role in IgE-binding against Pen ch 18. Molecular modeling suggests they are located on a loop-like structure at or near the surface of the major fungal allergen.
KeywordMeSH Terms
61. Wang  FQ, Zhao  Y, Dai  M, Liu  J, Zheng  GZ, Ren  ZH, He  JG,     ( 2008 )

Cloning and functional identification of C-4 methyl sterol oxidase genes from the penicillin-producing fungus Penicillium chrysogenum.

FEMS microbiology letters 287 (1)
PMID : 18707625  :   DOI  :   10.1111/j.1574-6968.2008.01294.x    
Abstract >>
Two C-4 methyl sterol oxidase genes (Pcerg25A and Pcerg25B) that are involved in ergosterol biosynthesis have been cloned from the penicillin-producing fungus Penicillium chrysogenum. cDNAs of both Pcerg25A and Pcerg25B have an ORF 885 bp in length, encoding a peptide of 295 residues. The deduced amino acid sequences of PcErg25A and PcErg25B show 86% identity, and have high identities to the characterized C-4 methyl sterol oxidases from Candida albicans and Saccharomyces cerevisiae. The function of Pcerg25A and Pcerg25B was identified by complementation of a yeast erg25-deficient strain. Pcerg25A is located in the DNA region containing the penicillin gene cluster, and thus its copy number is dependent on the patterns of the cluster region. Up to eight copies of Pcerg25A were found in the high-productivity strain NCPC 10086. By contrast, Pcerg25B was present in just a single copy in all tested P. chrysogenum genomes. Differences in the transcript level of either Pcerg25A or Pcerg25B were observed in different P. chrysogenum strains by real-time quantitative reverse transcriptase PCR analysis.
KeywordMeSH Terms
62. Serra  R, Peterson  S, N/A  N/A, Venâncio  A,     ( 2008 )

Multilocus sequence identification of Penicillium species in cork bark during plank preparation for the manufacture of stoppers.

Research in microbiology 159 (3)
PMID : 18321681  :   DOI  :   10.1016/j.resmic.2007.12.009    
Abstract >>
Despite several studies reporting Penicillium as one of the most frequent fungal genera in cork planks, the isolates were rarely identified to species level. We conducted a detailed study to identify Penicillium species from the field to the factory environment prior to and after boiling the cork planks. A total of 84 samples were analyzed. Of the 486 Penicillium isolates phenotypically identified, 32 representative or unusual strains were selected for identification by multilocus DNA sequence type. Cork proved to be a rich source of Penicillium biodiversity. A total of 30 taxa were recognized from cork including rarely seen species and 6 phylogenetically unique groups. Spores of some species lodged deep in cork can survive the boiling process. P. glabrum, P. glandicola and P. toxicarium, species with high CFU numbers in the field, are still frequently present in cork after boiling. Other species are killed by the boiling treatment and replaced by Penicillium species originating from the factory environment. Species known to contribute to cork taint were isolated at all stages. Good manufacturing practices are necessary at all stages in the preparation of cork planks to minimize the load of Penicillium species that produce cork taint.
KeywordMeSH Terms
Industrial Microbiology
63. Hoff  B, Pöggeler  S, Kück  U,     ( 2008 )

Eighty years after its discovery, Fleming's Penicillium strain discloses the secret of its sex.

Eukaryotic cell 7 (3)
PMID : 18223118  :   DOI  :   10.1128/EC.00430-07     PMC  :   PMC2268512    
Abstract >>
Eighty years ago, Alexander Fleming discovered antibacterial activity in the asexual mold Penicillium, and the strain he studied later was replaced by an overproducing isolate still used for penicillin production today. Using a heterologous PCR approach, we show that these strains are of opposite mating types and that both have retained transcriptionally expressed pheromone and pheromone receptor genes required for sexual reproduction. This discovery extends options for industrial strain improvement programs using conventional genetical approaches.
KeywordMeSH Terms
Genes, Mating Type, Fungal
64. Wang  L, Zhang  XM, Zhuang  WY,     ( 2007 )

Penicillium macrosclerotiorum, a new species producing large sclerotia discovered in south China.

Mycological research 111 (Pt 10)
PMID : 17998158  :   DOI  :   10.1016/j.mycres.2007.06.017    
Abstract >>
A distinctive new penicillium species, Penicillium macrosclerotiorum, isolated from soil in south China is reported. Morphologically, it resembles P. thomii, but is distinguished from the latter by its large, white to ivory coloured sclerotia, smooth-walled stipes, and globose, smooth-walled conidia. Although its rDNA ITS1-5.8S-ITS2 sequence is identical with that of P. thomii, the partial calmodulin gene sequence data show that it is a unique taxon and has no closely related relatives in penicillia.
KeywordMeSH Terms
Soil Microbiology
65. Wang  FQ, Liu  J, Dai  M, Ren  ZH, Su  CY, He  JG,     ( 2007 )

Molecular cloning and functional identification of a novel phenylacetyl-CoA ligase gene from Penicillium chrysogenum.

Biochemical and biophysical research communications 360 (2)
PMID : 17612506  :   DOI  :   10.1016/j.bbrc.2007.06.074    
Abstract >>
A novel phenylacetyl-CoA ligase gene, designated phlB, was cloned and identified from the penicillin producing strain Penicillium chrysogenum based on subtractive suppression hybridization approach. The phlB gene contains a 1686-bp open-reading frame and encodes a protein of approximately 62.6 kDa. The deduced amino acid sequence shows about 35% identity to the characterized P. chrysogenum phenylacetyl-CoA ligase Phl and has a peroxisomal targeting signal on its C-terminal. Recombinant PhlB protein was overexpressed in Escherichia coli and purified by nickel affinity chromatography. Enzymatic assay confirmed that recombinant PhlB can catalyze the reaction of phenylacetic acid (PAA) with CoA to yield phenylacetyl-CoA. The expression level of phlB in the penicillin producing medium supplemented with PAA, the side chain precursor of penicillin G, was about 2.5-fold higher than that in medium without PAA. The study suggested that PhlB might participate in the activation of PAA during penicillin biosynthesis in P. chrysogenum.
KeywordMeSH Terms
66. van den Berg  MA, Westerlaken  I, Leeflang  C, Kerkman  R, Bovenberg  RA,     ( 2007 )

Functional characterization of the penicillin biosynthetic gene cluster of Penicillium chrysogenum Wisconsin54-1255.

Fungal genetics and biology : FG & B 44 (9)
PMID : 17548217  :   DOI  :   10.1016/j.fgb.2007.03.008    
Abstract >>
Industrial strain improvement via classical mutagenesis is a black box approach. In an attempt to learn from and understand the mutations introduced, we cloned and characterized the amplified region of industrial penicillin production strains. Upon amplification of this region Penicillium chrysogenum is capable of producing an increased amount of antibiotics, as was previously reported [Barredo, J.L., Diez, B., Alvarez, E., Mart?n, J.F., 1989a. Large amplification of a 35-kb DNA fragment carrying two penicillin biosynthetic genes in high yielding strains of Penicillium chrysogenum. Curr. Genet. 16, 453-459; Newbert, R.W., Barton, B., Greaves, P., Harper, J., Turner, G., 1997. Analysis of a commercially improved Penicillium chrysogenum strain series, involvement of recombinogenic regions in amplification and deletion of the penicillin gene cluster. J. Ind. Microbiol. 19, 18-27]. Bioinformatic analysis of the central 56.9kb, present as six direct repeats in the strains analyzed in this study, predicted 15 Open Reading Frames (ORFs). Besides the three penicillin biosynthetic genes (pcbAB, pcbC and penDE) only one ORF has an orthologue of known function in the database: the Saccharomyces cerevisiae gene ERG25. Surprisingly, many genes known to encode direct or indirect steps beta-lactam biosynthesis like phenyl acetic acid CoA ligase and transporters are not present. Detailed analyses reveal a detectable transcript for most of the predicted ORFs under the conditions tested. We have studied the role of these in relation to penicillin production and amplification of the biosynthetic gene cluster. In contrast to what was expected, the genes encoding the three penicillin biosynthetic enzymes alone are sufficient to restore full beta-lactam synthesis in a mutant lacking the complete region. Therefore, the role of the other 12 ORFs in this region seems irrelevant for penicillin biosynthesis.
KeywordMeSH Terms
67. Kosalková  K, Rodríguez-Sáiz  M, Barredo  JL, Martín  JF,     ( 2007 )

Binding of the PTA1 transcriptional activator to the divergent promoter region of the first two genes of the penicillin pathway in different Penicillium species.

Current genetics 52 (5��6��)
PMID : 17924108  :   DOI  :   10.1007/s00294-007-0157-7    
Abstract >>
The aim of this work is to establish the correlation between the transcriptional activator PTA1 and the expression of the penicillin genes in different penicillin-producing strains. The level of expression of the first two genes of the penicillin pathway was clearly higher in Penicillium chrysogenum than in Penicillium notatum and Penicillium nalgiovense. The divergent promoter pcbAB-pcbC region contains binding sequences for several transcriptional factors that are conserved in P. notatum and P. chrysogenum, but not in P. nalgiovense. Binding of the purified P. chrysogenum transcriptional activator PTA1 to the palindromic heptamer TTAGTAA took place when the P. chrysogenum 35 bp DNA fragment containing the heptamer was used as a probe, but not when the sequence occurring in P. nalgiovense was used. P. nalgiovense protein fractions purified by heparin agarose chromatography did not bind to the 35-bp DNA fragment either from P. nalgiovense or P. chrysogenum, although some degree of binding was observed when crude extracts were used. This finding may explain the low expression of pcbC in P. nalgiovense. All the P. chrysogenum strains, including the industrial strain E1, showed the same nucleotide sequence, including the consensus PTA1 binding site.
KeywordMeSH Terms
Genes, Fungal
Promoter Regions, Genetic
68. Matsumoto  S, Yamada  H, Kunishige  Y, Takenaka  S, Nakazawa  M, Ueda  M, Sakamoto  T,     ( 2017 )

Identification of a novel Penicillium chrysogenum rhamnogalacturonan rhamnohydrolase and the first report of a rhamnogalacturonan rhamnohydrolase gene.

Enzyme and microbial technology 98 (N/A)
PMID : 28110667  :   DOI  :   10.1016/j.enzmictec.2016.12.008    
Abstract >>
Rhamnogalacturonan (RG) I is one of the main components of pectins in the plant cell wall. We recently reported two RG I-degrading enzymes, endo-RG and exo-RG lyases, secreted by Penicillium chrysogenum 31B. Here, our aims were to purify a RG rhamnohydrolase (PcRGRH78A) from the culture filtrate of this strain and to characterize this enzyme. On the basis of the internal amino acid sequences, the encoding gene, Pcrgrh78A, was cloned and overexpressed in Aspergillus oryzae. The deduced amino acid sequence of PcRGRH78A is highly similar to an uncharacterized protein belonging to glycoside hydrolase family 78. Pfam analysis revealed that PcRGRH78A contains a bacterial �\-l-rhamnosidase (PF05592) domain. PcRGRH78A shows optimal activity at 45�XC and pH 5. The specificity of PcRGRH78A toward rhamnose (Rha)-containing substrates was compared with that of a P. chrysogenum �\-l-rhamnosidase (PcRHA78B) belonging to glycoside hydrolase family 78. PcRGRH78A specifically hydrolyzes RG oligosaccharides that contain Rha at their nonreducing ends, releasing the Rha, but has no activity toward naringin, hesperidin, or rutin. In contrast, PcRHA78B effectively degrades p-nitrophenyl �\-l-rhamnopyranoside and the three flavonoids, but not RG oligosaccharides. When galactosyl RG oligosaccharides were used as the substrate, PcRGRH78A released Rha in 3.5-fold greater amounts in the presence of �]-galactosidase than in its absence, indicating that PcRGRH78A preferentially acts on Rha residues without the galactose moiety at nonreducing ends. To our knowledge, this is the first report of a gene encoding a RG rhamnohydrolase.
KeywordMeSH Terms
Gene cloning
Glycoside hydrolase family 78
Penicillium chrysogenum
Rhamnogalacturonan rhamnohydrolase
α-l-Rhamnosidase
Genes, Fungal
69. Chen  AJ, Sun  BD, Houbraken  J, Frisvad  JC, Yilmaz  N, Zhou  YG, Samson  RA,     ( 2016 )

New Talaromyces species from indoor environments in China.

Studies in mycology 84 (N/A)
PMID : 28070136  :   DOI  :   10.1016/j.simyco.2016.11.003     PMC  :   PMC5219591    
Abstract >>
Talaromyces contains both asexual and sexually reproducing species. This genus is divided in seven sections and currently has 105 accepted species. In this study we investigated the Talaromyces isolates that were obtained during a study of indoor air collected in Beijing, China. These indoor Talaromyces strains are resolved in four sections, seven of them are identified as T. islandicus, T. aurantiacus, T. siamensis and T. albobiverticillius according to BenA sequences, while 14 isolates have divergent sequences and are described here as nine new species. The new species are placed in four sections, namely sections Helici, Islandici, Talaromyces and Trachyspermi. They are described based on sequence data (ITS, BenA, CaM and RPB2) in combination with phenotypic and extrolite characters. Morphological descriptions and notes for distinguishing similar species are provided for each new species. The recently described T. rubrifaciens is synonymised with T. albobiverticillius based on presented phylogenetic results.
KeywordMeSH Terms
Eurotiales
Indoor air
Polyphasic taxonomy
T. adpressus A.J. Chen, Frisvad & Samson
T. beijingensis A.J. Chen, Frisvad & Samson
T. cerinus A.J. Chen, Frisvad & Samson
T. chlamydosporus A.J. Chen, Frisvad & Samson
T. diversiformis A.J. Chen, Frisvad & Samson
T. fusiformis A.J. Chen, Frisvad & Samson
T. neorugulosus A.J. Chen, Frisvad & Samson
T. reverso-olivaceus A.J. Chen, Frisvad & Samson
Talaromyces aerius A.J. Chen, Frisvad & Samson
Talaromyces albobiverticillius
Eurotiales
Indoor air
Polyphasic taxonomy
T. adpressus A.J. Chen, Frisvad & Samson
T. beijingensis A.J. Chen, Frisvad & Samson
T. cerinus A.J. Chen, Frisvad & Samson
T. chlamydosporus A.J. Chen, Frisvad & Samson
T. diversiformis A.J. Chen, Frisvad & Samson
T. fusiformis A.J. Chen, Frisvad & Samson
T. neorugulosus A.J. Chen, Frisvad & Samson
T. reverso-olivaceus A.J. Chen, Frisvad & Samson
Talaromyces aerius A.J. Chen, Frisvad & Samson
Talaromyces albobiverticillius
Eurotiales
Indoor air
Polyphasic taxonomy
T. adpressus A.J. Chen, Frisvad & Samson
T. beijingensis A.J. Chen, Frisvad & Samson
T. cerinus A.J. Chen, Frisvad & Samson
T. chlamydosporus A.J. Chen, Frisvad & Samson
T. diversiformis A.J. Chen, Frisvad & Samson
T. fusiformis A.J. Chen, Frisvad & Samson
T. neorugulosus A.J. Chen, Frisvad & Samson
T. reverso-olivaceus A.J. Chen, Frisvad & Samson
Talaromyces aerius A.J. Chen, Frisvad & Samson
Talaromyces albobiverticillius
Eurotiales
Indoor air
Polyphasic taxonomy
T. adpressus A.J. Chen, Frisvad & Samson
T. beijingensis A.J. Chen, Frisvad & Samson
T. cerinus A.J. Chen, Frisvad & Samson
T. chlamydosporus A.J. Chen, Frisvad & Samson
T. diversiformis A.J. Chen, Frisvad & Samson
T. fusiformis A.J. Chen, Frisvad & Samson
T. neorugulosus A.J. Chen, Frisvad & Samson
T. reverso-olivaceus A.J. Chen, Frisvad & Samson
Talaromyces aerius A.J. Chen, Frisvad & Samson
Talaromyces albobiverticillius
Eurotiales
Indoor air
Polyphasic taxonomy
T. adpressus A.J. Chen, Frisvad & Samson
T. beijingensis A.J. Chen, Frisvad & Samson
T. cerinus A.J. Chen, Frisvad & Samson
T. chlamydosporus A.J. Chen, Frisvad & Samson
T. diversiformis A.J. Chen, Frisvad & Samson
T. fusiformis A.J. Chen, Frisvad & Samson
T. neorugulosus A.J. Chen, Frisvad & Samson
T. reverso-olivaceus A.J. Chen, Frisvad & Samson
Talaromyces aerius A.J. Chen, Frisvad & Samson
Talaromyces albobiverticillius
Eurotiales
Indoor air
Polyphasic taxonomy
T. adpressus A.J. Chen, Frisvad & Samson
T. beijingensis A.J. Chen, Frisvad & Samson
T. cerinus A.J. Chen, Frisvad & Samson
T. chlamydosporus A.J. Chen, Frisvad & Samson
T. diversiformis A.J. Chen, Frisvad & Samson
T. fusiformis A.J. Chen, Frisvad & Samson
T. neorugulosus A.J. Chen, Frisvad & Samson
T. reverso-olivaceus A.J. Chen, Frisvad & Samson
Talaromyces aerius A.J. Chen, Frisvad & Samson
Talaromyces albobiverticillius
70. Sonderegger  C, Fizil  ?, Burtscher  L, Hajdu  D, Muñoz  A, Gáspári  Z, Read  ND, Batta  G, Marx  F,     ( 2017 )

D19S Mutation of the Cationic, Cysteine-Rich Protein PAF: Novel Insights into Its Structural Dynamics, Thermal Unfolding and Antifungal Function.

PloS one 12 (1)
PMID : 28072824  :   DOI  :   10.1371/journal.pone.0169920     PMC  :   PMC5224997    
Abstract >>
The cysteine-rich, cationic, antifungal protein PAF is abundantly secreted into the culture supernatant of the filamentous Ascomycete Penicillium chrysogenum. The five �]-strands of PAF form a compact �]-barrel that is stabilized by three disulphide bonds. The folding of PAF allows the formation of four surface-exposed loops and distinct charged motifs on the protein surface that might regulate the interaction of PAF with the sensitive target fungus. The growth inhibitory activity of this highly stable protein against opportunistic fungal pathogens provides great potential in antifungal drug research. To understand its mode of action, we started to investigate the surface-exposed loops of PAF and replaced one aspartic acid at position 19 in loop 2 that is potentially involved in PAF active or binding site, with a serine (Asp19 to Ser19). We analysed the overall effects, such as unfolding, electrostatic changes, sporadic conformers and antifungal activity when substituting this specific amino acid to the fairly indifferent amino acid serine. Structural analyses revealed that the overall 3D solution structure is virtually identical with that of PAF. However, PAFD19S showed slightly increased dynamics and significant differences in the surface charge distribution. Thermal unfolding identified PAFD19S to be rather a two-state folder in contrast to the three-state folder PAF. Functional comparison of PAFD19S and PAF revealed that the exchange at residue 19 caused a dramatic loss of antifungal activity: the binding and internalization of PAFD19S by target cells was reduced and the protein failed to trigger an intracellular Ca2+ response, all of which are closely linked to the antifungal toxicity of PAF. We conclude that the negatively charged residue Asp19 in loop 2 is essential for full function of the cationic protein PAF.
KeywordMeSH Terms
Molecular Dynamics Simulation
Mutation, Missense
Protein Denaturation
71. Garnier  L, Valence  F, Pawtowski  A, Auhustsinava-Galerne  L, Frotté  N, Baroncelli  R, Deniel  F, Coton  E, Mounier  J,     ( 2017 )

Diversity of spoilage fungi associated with various French dairy products.

International journal of food microbiology 241 (N/A)
PMID : 27794247  :   DOI  :   10.1016/j.ijfoodmicro.2016.10.026    
Abstract >>
Yeasts and molds are responsible for dairy product spoilage, resulting in significant food waste and economic losses. Yet, few studies have investigated the diversity of spoilage fungi encountered in dairy products. In the present study, 175 isolates corresponding to 105 from various spoiled dairy products and 70 originating from dairy production environments, were identified using sequencing of the ITS region, the partial �]-tubulin, calmodulin and/or EF�\ genes, and the D1-D2 domain of the 26S rRNA gene for filamentous fungi and yeasts, respectively. Among the 41 species found in spoiled products, Penicillium commune and Penicillium bialowiezense were the most common filamentous fungi, representing around 10% each of total isolates while Meyerozyma guilliermondii and Trichosporon asahii were the most common yeasts (4.8% each of total isolates). Several species (e.g. Penicillium antarcticum, Penicillium salamii and Cladosporium phyllophilum) were identified for the first time in dairy products or their environment. In addition, numerous species were identified in both spoiled products and their corresponding dairy production environment suggesting that the latter acts as a primary source of contamination. Secondly, the resistance to chemical preservatives (sodium benzoate, calcium propionate, potassium sorbate and natamycin) of 10 fungal isolates representative of the observed biodiversity was also evaluated. Independently of the fungal species, natamycin had the lowest minimum inhibitory concentration (expressed in gram of preservative/l), followed by potassium sorbate, sodium benzoate and calcium propionate. In the tested conditions, Cladosporium halotolerans and Didymella pinodella were the most sensitive fungi while Yarrowia lipolytica and Candida parapsilosis were the most resistant towards the tested preservatives. This study provides interesting information on the occurrence of fungal contaminants in dairy products and environments that may help developing adequate strategies for fungal spoilage control.
KeywordMeSH Terms
Dairy products
Diversity
Fungi
Preservative resistance
Spoilage
Dairy products
Diversity
Fungi
Preservative resistance
Spoilage
Dairy products
Diversity
Fungi
Preservative resistance
Spoilage
Dairy products
Diversity
Fungi
Preservative resistance
Spoilage
Dairy products
Diversity
Fungi
Preservative resistance
Spoilage
Dairy products
Diversity
Fungi
Preservative resistance
Spoilage
Dairy products
Diversity
Fungi
Preservative resistance
Spoilage
Dairy products
Diversity
Fungi
Preservative resistance
Spoilage
72. Abastabar  M, Mirhendi  H, Hedayati  MT, Shokohi  T, Rezaei-Matehkolaei  A, Mohammadi  R, Badali  H, Moazeni  M, Haghani  I, Ghojoghi  A, Akhtari  J,     ( 2016 )

Genetic and Morphological Diversity of the Genus Penicillium From Mazandaran and Tehran Provinces, Iran.

Jundishapur journal of microbiology 9 (1)
PMID : 27099684  :   DOI  :   10.5812/jjm.28280     PMC  :   PMC4833887    
Abstract >>
The genus Penicillium contains a large number of ubiquitous environmental taxa, of which some species are clinically important. Identification of Penicillium down to the species level is currently based on polyphasic criteria, including phenotypic features and genetic markers. Biodiversity of the genus Penicillium from Mazandaran and Tehran provinces has not been described. The current paper focused on the environmental biodiversity of Penicillium isolates within some areas of Mazandaran and Tehran provinces, based on morphological traits and the molecular data from partial sequence of the �]-tubulin (BT2) gene. A total of 400 strains were isolated from the environment and investigated using morphological tests and sequencing of BT2, in order to characterize the spectrum of the Penicillium species. Sequence analysis of BT2 and morphological criteria of 20 strains representative of 10 species showed that Penicillium chrysogenum was the most prevalent species (n = 6), followed by P. polonicum (n = 3), P. glabrum (n = 2), P. palitans (n = 2), P. melanoconidium (n = 2), and other species, including P. expansum, P. canescense, P. griseofulvum, P. italicum, and P. raistrickii with one case each. It was shown that partial �]-tubulin sequence, as a reliable genetic target, supported specific morphological criteria for identification of the Penicillium species. Like other assessments throughout the world, P. chrysogenum remains the most frequent environmental Penicillium species in Mazandaran and Tehran Provinces.
KeywordMeSH Terms
Beta-Tubulin
DNA Sequencing
PCR
Penicillium
Beta-Tubulin
DNA Sequencing
PCR
Penicillium
Beta-Tubulin
DNA Sequencing
PCR
Penicillium
73. Poli  A, Lazzari  A, Prigione  V, Voyron  S, Spadaro  D, Varese  GC,     ( N/A )

Influence of plant genotype on the cultivable fungi associated to tomato rhizosphere and roots in different soils.

Fungal biology 120 (6��7��)
PMID : 27268246  :   DOI  :   10.1016/j.funbio.2016.03.008    
Abstract >>
Rhizosphere and root-associated microbiota are crucial in determining plant health and in increasing productivity of agricultural crops. To date, research has mainly focused on the bacterial dimension of the microbiota. However, interest in the mycobiota is increasing, since fungi play a key role in soil ecosystems. We examined the effect of plant genotype, soil, and of Fusarium oxysporum f. sp. lycopersici (Fol) on the cultivable component of rhizosphere and root-associated mycobiota of tomato. Resistant and susceptible varieties were cultivated on two different soils (A and B), under glasshouse conditions. Isolated fungi were identified by morphological and molecular approaches. Differences were found between the rhizosphere and the roots, which in general displayed a lower number of species. The structure of the mycobiota was significantly affected by the soil type in the rhizosphere as well as by the plant genotype within the roots (NPERMANOVA, p < 0.05). The addition of Fol changed the community structure, particularly in soil A, where Penicillium spp. and Fusarium spp. were the dominant responding fungi. Overall, the results indicated that i) soil type and plant genotype affect the fungal communities; ii) plant roots select few species from the rhizosphere; and iii) the fungal community structure is influenced by Fol.
KeywordMeSH Terms
Fusarium wilt
Genetic diversity
Mycobiota
Soil type
Fusarium wilt
Genetic diversity
Mycobiota
Soil type
Fusarium wilt
Genetic diversity
Mycobiota
Soil type
Fusarium wilt
Genetic diversity
Mycobiota
Soil type
Fusarium wilt
Genetic diversity
Mycobiota
Soil type
Fusarium wilt
Genetic diversity
Mycobiota
Soil type
Fusarium wilt
Genetic diversity
Mycobiota
Soil type
Fusarium wilt
Genetic diversity
Mycobiota
Soil type
Biota
Genotype
Rhizosphere
Soil Microbiology
74. Iwai  M, Yamada  H, Ikemoto  T, Matsumoto  S, Fujiwara  D, Takenaka  S, Sakamoto  T,     ( 2015 )

Biochemical Characterization and Overexpression of an Endo-rhamnogalacturonan Lyase from Penicillium chrysogenum.

Molecular biotechnology 57 (6)
PMID : 25666014  :   DOI  :   10.1007/s12033-015-9847-4    
Abstract >>
Rhamnogalacturonan lyase (PcRGL4A) was purified from the culture supernatant of Penicillium chrysogenum 31B. PcRGL4A optimal activity occurred between pH 7-8 and at 40 �XC. Conserved Domain Search analysis identified PcRGL4A as a member of Polysaccharide Lyase family 4. PcRGL4A contains two conserved catalytic and four conserved substrate-binding residues as determined by X-ray crystallography of the Aspergillus aculeatus RG lyase. Recombinant PcRGL4A (rPcRGL4A) expressed in Escherichia coli demonstrated specific activity against rhamnogalacturonan (RG) but not homogalacturonan. Analysis of the RG reaction products by high-performance anion-exchange chromatography revealed that rPcRGL4A cleaved the substrate in an endo-manner and that the major final product was an RG tetrasaccharide with 4-deoxy-4,5-unsaturated galacturonic acid at the nonreducing end. Based on these results, PcRGL4A was classified as an endo-acting RG lyase (EC 4.2.2.23). Divalent cations were not essential for the enzymatic activity of rPcRGL4A, but addition of calcium ions to the reaction mixture increased enzymatic activity. rPcRGL4A demonstrated a preference for RG lacking galactose decoration.
KeywordMeSH Terms
75. Barredo  JL, van Solingen  P, Díez  B, Alvarez  E, Cantoral  JM, Kattevilder  A, Smaal  EB, Groenen  MA, Veenstra  AE, Martín  JF,     ( 1989 )

Cloning and characterization of the acyl-coenzyme A: 6-aminopenicillanic-acid-acyltransferase gene of Penicillium chrysogenum.

Gene 83 (2)
PMID : 2555269  :   DOI  :   10.1016/0378-1119(89)90115-7    
Abstract >>
A gene, aat, encoding acyl-CoA: 6-aminopenicillanic acid acyltransferase (AAT), the last enzyme of the penicillin (Pn) biosynthetic pathway, has been cloned from the genome of Penicillium chrysogenum AS-P-78. The gene contains three introns in the 5'-region and encodes a protein of 357 amino acids with an Mr of 39,943. It complements mutants of P. chrysogenum deficient in AAT activity. The aat gene is expressed as a 1.15-kb transcript and the encoded protein appears to be processed post-translationally into two nonidentical polypeptides of 102 and 255 aa, with Mrs of 11,498 and 28,461, respectively. Three proteins of 40, 11, and 29 kDa (the last one corresponding to the previously purified AAT), were identified in extracts of P. chrysogenum. The aa sequence of the N-terminal end of the 11-kDa polypeptide matched the nucleotide (nt) sequence of the 5'-region of aat. The N-terminal end of the 29-kDa polypeptide corresponded to the sequence beginning at nt position 916 of the sequenced DNA fragment (nt 441 of aat gene). The aat gene of P. chrysogenum resembles the genes encoding Pn acylases of Escherichia coli, Proteus rettgeri and Pseudomonas sp., all of which encode two nonidentical subunits derived from a common precursor, encoded by a single open reading frame.
KeywordMeSH Terms
Cloning, Molecular
Genes, Fungal
Penicillin-Binding Proteins
76. Visagie  CM, Houbraken  J, Frisvad  JC, Hong  SB, Klaassen  CH, Perrone  G, Seifert  KA, Varga  J, Yaguchi  T, Samson  RA,     ( 2014 )

Identification and nomenclature of the genus Penicillium.

Studies in mycology 78 (N/A)
PMID : 25505353  :   DOI  :   10.1016/j.simyco.2014.09.001     PMC  :   PMC4261876    
Abstract >>
Penicillium is a diverse genus occurring worldwide and its species play important roles as decomposers of organic materials and cause destructive rots in the food industry where they produce a wide range of mycotoxins. Other species are considered enzyme factories or are common indoor air allergens. Although DNA sequences are essential for robust identification of Penicillium species, there is currently no comprehensive, verified reference database for the genus. To coincide with the move to one fungus one name in the International Code of Nomenclature for algae, fungi and plants, the generic concept of Penicillium was re-defined to accommodate species from other genera, such as Chromocleista, Eladia, Eupenicillium, Torulomyces and Thysanophora, which together comprise a large monophyletic clade. As a result of this, and the many new species described in recent years, it was necessary to update the list of accepted species in Penicillium. The genus currently contains 354 accepted species, including new combinations for Aspergillus crystallinus, A. malodoratus and A. paradoxus, which belong to Penicillium section Paradoxa. To add to the taxonomic value of the list, we also provide information on each accepted species MycoBank number, living ex-type strains and provide GenBank accession numbers to ITS, �]-tubulin, calmodulin and RPB2 sequences, thereby supplying a verified set of sequences for each species of the genus. In addition to the nomenclatural list, we recommend a standard working method for species descriptions and identifications to be adopted by laboratories working on this genus.
KeywordMeSH Terms
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
Aspergillaceae
Fungal identification
media
nomenclature
phylogeny
77. Gonçalves  VN, Cantrell  CL, Wedge  DE, Ferreira  MC, Soares  MA, Jacob  MR, Oliveira  FS, Galante  D, Rodrigues  F, Alves  TM, Zani  CL, Junior  PA, Murta  S, Romanha  AJ, Barbosa  EC, Kroon  EG, Oliveira  JG, Gomez-Silva  B, Galetovic  A, Rosa  CA, Rosa  LH,     ( 2016 )

Fungi associated with rocks of the Atacama Desert: taxonomy, distribution, diversity, ecology and bioprospection for bioactive compounds.

Environmental microbiology 18 (1)
PMID : 26235221  :   DOI  :   10.1111/1462-2920.13005    
Abstract >>
This study assessed the diversity of cultivable rock-associated fungi from Atacama Desert. A total of 81 fungal isolates obtained were identified as 29 Ascomycota taxa by sequencing different regions of DNA. Cladosporium halotolerans, Penicillium chrysogenum and Penicillium cf. citrinum were the most frequent species, which occur at least in four different altitudes. The diversity and similarity indices ranged in the fungal communities across the latitudinal gradient. The Fisher-�\ index displayed the higher values for the fungal communities obtained from the siltstone and fine matrix of pyroclastic rocks with finer grain size, which are more degraded. A total of 23 fungal extracts displayed activity against the different targets screened. The extract of P. chrysogenum afforded the compounds �\-linolenic acid and ergosterol endoperoxide, which were active against Cryptococcus neoformans and methicillin-resistance Staphylococcus aureus respectively. Our study represents the first report of a new habitat of fungi associated with rocks of the Atacama Desert and indicated the presence of interesting fungal community, including species related with saprobes, parasite/pathogen and mycotoxigenic taxa. The geological characteristics of the rocks, associated with the presence of rich resident/resilient fungal communities suggests that the rocks may provide a favourable microenvironment fungal colonization, survival and dispersal in extreme conditions.
KeywordMeSH Terms
78. Fizil  ?, Gáspári  Z, Barna  T, Marx  F, Batta  G,     ( 2015 )

Invisible conformers of an antifungal disulfide protein revealed by constrained cold and heat unfolding, CEST-NMR experiments, and molecular dynamics calculations.

Chemistry (Weinheim an der Bergstrasse, Germany) 21 (13)
PMID : 25676351  :   DOI  :   10.1002/chem.201404879     PMC  :   PMC4464532    
Abstract >>
Transition between conformational states in proteins is being recognized as a possible key factor of function. In support of this, hidden dynamic NMR structures were detected in several cases up to populations of a few percent. Here, we show by two- and three-state analysis of thermal unfolding, that the population of hidden states may weight 20-40 % at 298 K in a disulfide-rich protein. In addition, sensitive (15) N-CEST NMR experiments identified a low populated (0.15 %) state that was in slow exchange with the folded PAF protein. Remarkably, other techniques failed to identify the rest of the NMR "dark matter". Comparison of the temperature dependence of chemical shifts from experiments and molecular dynamics calculations suggests that hidden conformers of PAF differ in the loop and terminal regions and are most similar in the evolutionary conserved core. Our observations point to the existence of a complex conformational landscape with multiple conformational states in dynamic equilibrium, with diverse exchange rates presumably responsible for the completely hidden nature of a considerable fraction.
KeywordMeSH Terms
NMR spectroscopy
conformation analysis
molecular modeling
protein folding
protein structures
79. Martin  RL, Daley  LA, Lovric  Z, Wailes  LM, Renosto  F, Segel  IH,     ( 1989 )

The "regulatory" sulfhydryl group of Penicillium chrysogenum ATP sulfurylase. Cooperative ligand binding after SH modification; chemical and thermodynamic properties.

The Journal of biological chemistry 264 (20)
PMID : 2545683  :  
Abstract >>
ATP sulfurylase from Penicillium chrysogenum is a homohexamer that contains three free sulfhydryl groups/subunit, only one of which (designated SH-1) can be modified by disulfide, maleimide, and halide reagents under nondenaturing conditions. Modification of SH-1 has only a small effect on kcat but causes the [S]0.5 values for MgATP and SO4(2-) (or MoO4(2-) to increase by an order of magnitude. Additionally, the velocity curves become sigmoidal with a Hill coefficient (nH) of about 2 (Renosto, F., Martin, R. L., and Segel, I. H. (1987) J. Biol. Chem. 262, 16279-16288). Direct equilibrium binding measurements confirmed that [32P]MgATP binds to the SH-modified enzyme in a positively cooperative fashion (nH = 2.0) if a sulfate subsite ligand (e.g. FSO3-) is also present. [35S]Adenosine 5'-phosphosulfate (APS) binding to the SH-modified enzyme displayed positive cooperativity (nH = 1.9) in the absence of a PPi subsite ligand. The results indicate that positive cooperativity requires occupancy of the adenylyl and sulfate (but not the pyrophosphate) subsites. [35S]APS binding to the native enzyme displayed negative cooperativity (or binding to at least two classes of sites). Isotope trapping profiles for the single turnover of [35S]APS: (a) confirmed the equilibrium binding curves, (b) indicated that all six sites/hexamer are catalytically active, and (c) showed that APS does not dissociate at a significant rate from E.APS.PPi. The MgPPi concentration dependence of [35S]APS trapping was indicative of MgPPi binding to two classes of sites on both the native and SH-modified enzyme. Inactivation of the native or SH-modified enzyme by phenylglyoxal in the presence of saturating APS was biphasic. The semilog plots suggested that only half of the sites were highly protected. The cumulative data suggest a model in which pairs of sites or subunits can exist in three different states designated HH (both sites have a high APS affinity, as in the native free enzyme), LL (both sites have a low APS affinity as in the SH-modified enzyme), and LH (as in the APS-occupied native or SH-modified enzyme). Thus, the HH----LH transition displays negative cooperativity for APS binding while the LL----LH transition displays positive cooperativity. The relative reactivities of like-paired SH-reactive reagents were in the order: N-phenylmaleimide greater than N-ethylmaleimide; dithionitropyridine greater than dithionitrobenzoate; thiolyte-MQ greater than thiolyte-MB. The log kmod versus pH curve indicates that the pKa of SH-1 is greater than 9.(ABSTRACT TRUNCATED AT 400 WORDS)
KeywordMeSH Terms
80. Iwai  M, Kawakami  T, Ikemoto  T, Fujiwara  D, Takenaka  S, Nakazawa  M, Ueda  M, Sakamoto  T,     ( 2015 )

Molecular characterization of a Penicillium chrysogenum exo-rhamnogalacturonan lyase that is structurally distinct from other polysaccharide lyase family proteins.

Applied microbiology and biotechnology 99 (20)
PMID : 25921806  :   DOI  :   10.1007/s00253-015-6600-7    
Abstract >>
We previously described an endo-acting rhamnogalacturonan (RG) lyase, termed PcRGL4A, of Penicillium chrysogenum 31B. Here, we describe a second RG lyase, called PcRGLX. We determined the cDNA sequence of the Pcrglx gene, which encodes PcRGLX. Based on analyses using a BLAST search and a conserved domain search, PcRGLX was found to be structurally distinct from known RG lyases and might belong to a new polysaccharide lyase family together with uncharacterized fungal proteins of Nectria haematococca, Aspergillus oryzae, and Fusarium oxysporum. The Pcrglx cDNA gene product (rPcRGLX) expressed in Escherichia coli demonstrated specific activity against RG but not against homogalacturonan. Divalent cations were not essential for the enzymatic activity of rPcRGLX. rPcRGLX mainly released unsaturated galacturonosyl rhamnose (�GGR) from RG backbones used as the substrate from the initial stage of the reaction, indicating that the enzyme can be classified as an exo-acting RG lyase (EC 4.2.2.24). This is the first report of an RG lyase with this mode of action in Eukaryota. rPcRGLX acted synergistically with PcRGL4A to degrade soybean RG and released �GGR. This �GGR was partially decorated with galactose (Gal) residues, indicating that rPcRGLX preferred oligomeric RGs to polymeric RGs, that the enzyme did not require Gal decoration of RG backbones for degradation, and that the enzyme bypassed the Gal side chains of RG backbones. These characteristics of rPcRGLX might be useful in the determination of complex structures of pectins.
KeywordMeSH Terms
Exo-acting rhamnogalacturonan lyase
Nucleotide sequence
Overexpression
Penicillium chrysogenum
Unsaturated galacturonosyl rhamnose
81. Shinozaki  A, Hosokawa  S, Nakazawa  M, Ueda  M, Sakamoto  T,     ( 2015 )

Identification and characterization of three Penicillium chrysogenum �\-l-arabinofuranosidases (PcABF43B, PcABF51C, and AFQ1) with different specificities toward arabino-oligosaccharides.

Enzyme and microbial technology 73-74 (N/A)
PMID : 26002506  :   DOI  :   10.1016/j.enzmictec.2015.04.003    
Abstract >>
We previously described four �\-l-arabinofuranosidases (ABFs) secreted by Penicillium chrysogenum 31B. Here, we cloned the fifth and sixth genes (Pcabf43B and Pcabf51C) encoding the ABFs PcABF43B and PcABF51C in this strain and overexpressed these genes in Escherichia coli. The deduced amino acid sequences of PcABF43B and PcABF51C were highly similar to putative ABFs belonging to glycoside hydrolase families 43 and 51, respectively. Semiquantitative reverse transcription polymerase chain reaction indicated that both genes were induced by arabinose, arabinitol, arabinan, and arabinoxylan; however, the Pcabf51C gene was constitutively expressed at low levels in P. chrysogenum 31B. PcABF43B had optimal activity at 20�XC and pH 5-6, indicating that this enzyme was psychrophilic and had the lowest optimal temperature reported for ABFs. PcABF51C had optimal activity at 45�XC and pH 6-7. Both recombinant enzymes showed high activity on arabino-oligosaccharides, but little activity on arabinose-containing polysaccharides, such as l-arabinan. Next, we compared the substrate specificities of PcABF43B, PcABF51C, and AFQ1, a P. chrysogenum ABF that preferentially degraded oligosaccharides over polysaccharides. PcABF43B was found to preferentially hydrolyze (1��3)-linkages in branched arabino-oligosaccharides and released only a small amount of arabinose from linear �\-1,5-arabino-oligosaccharides. In contrast, AFQ1 and PcABF51C showed higher activities on linear arabino-oligosaccharides than on branched arabino-oligosaccharides. AFQ1 showed high catalytic efficiencies for �\-1,5-l-arabinofuranobiose (�\-1,5-Ara2) and �\-1,5-l-arabinofuranotriose (�\-1,5-Ara3) at the same level. In contrast, intracellular PcABF51C showed much higher catalytic efficiency for �\-1,5-Ara2 than for �\-1,5-Ara3.
KeywordMeSH Terms
Arabino-oligosaccharides
Expression profile
Overexpression
Penicillium chrysogenum
α-l-Arabinofuranosidases
Genes, Fungal
82. El-Sayed  AS, Shindia  AA, Diab  AA, Rady  AM,     ( 2014 )

Purification and immobilization of L-arginase from thermotolerant Penicillium chrysogenum KJ185377.1; with unique kinetic properties as thermostable anticancer enzyme.

Archives of pharmacal research N/A (N/A)
PMID : 25322968  :   DOI  :   10.1007/s12272-014-0498-y    
Abstract >>
L-Arginase, hydrolyzing L-arginine to L-ornithine and urea, is a powerful anticancer, L-arginine-depleting agent, against argininosuccinate synthase expressing tumors. Otherwise, the higher antigenicity and lower thermal stability of this enzyme was the main biochemical hurdles. Since, the intrinsic thermal stability of enzymes follow the physiological temperature of their producer, thus, characterization of L-arginase from thermotolerant Penicillium chrysogenum was the objective of this study. L-Arginase (Arg) was purified to its homogeneity from P. chrysogenum by 10.1-fold, with 37.0 kDa under denaturing PAGE, optimum reaction at 50 �XC, pH stability (6.8-7.9), with highest molar ratio of constitutional arginine, glutamic acid, lysine and aspartic acid. The purified enzyme was PEGylated and immobilized on chitosan, with 41.9 and 22.1 % yield of immobilization. At 40 �XC, the T1/2 value of free-Arg, PEG-Arg and Chit-Arg was 10.4, 15.6, 20.5 h, respectively. The free-Arg and Chit-Arg have a higher affinity to L-arginine (K m 4.8 mM), while, PEG-Arg affinity was decreased by about 3 fold (K m 15.2 mM). The inhibitory constants to the free and PEG-Arg were relatively similar towards HA and PPG. The IC50 for the free enzyme against HEPG-2 and A549 tumor cells was 0.136 and 0.165 U/ml, comparing to 0.232 and 0.496 U/ml for PEG-Arg, respectively. The in vivo T1/2 to the free Arg and PEG-Arg was 16.4 and 20.4 h, respectively as holo-enzyme. The residual L-arginine level upon using free Arg was 156.9 and 144.5 ?M, after 6 and 8 h, respectively, regarding to initials at 253.6 ?M, while for Peg-Arg the level of L-arginine was nil till 7 h of initial dosing. The titer of IgG was induced by 10-15 % in response to free-Arg after 28 days comparing to IgG titer for PEG-Arg.
KeywordMeSH Terms
83. Shinozaki  A, Kawakami  T, Hosokawa  S, Sakamoto  T,     ( 2014 )

A novel GH43 �\-l-arabinofuranosidase of Penicillium chrysogenum that preferentially degrades single-substituted arabinosyl side chains in arabinan.

Enzyme and microbial technology 58-59 (N/A)
PMID : 24731829  :   DOI  :   10.1016/j.enzmictec.2014.03.005    
Abstract >>
We previously described three �\-l-arabinofuranosidases (ABFs) secreted by Penicillium chrysogenum 31B. Here, we purified a fourth ABF, termed PcABF43A, from the culture filtrate. The molecular mass of the enzyme was estimated to be 31kDa. PcABF43A had the highest activity at 35�XC and at around pH 5. The enzyme activity was strong on sugar beet l-arabinan but weak on debranched arabinan and arabinoxylan. Low molecular-mass substrates such as p-nitrophenyl �\-l-arabinofuranoside, �\-1,5-l-arabinooligosaccharides, and branched arabinotriose were highly resistant to the action of PcABF43A. (1)H-NMR analysis revealed that PcABF43A hydrolyzed arabinosyl side chains linked to C-2 or C-3 of single-substituted arabinose residues in l-arabinan. Reports concerning enzymes specific for l-arabinan are quite limited. Pcabf43A cDNA encoding PcABF43A was isolated by in vitro cloning. The deduced amino acid sequence of the enzyme shows high similarities with the sequences of other fungal uncharacterized proteins. Semi-quantitative RT-PCR analysis indicated that the Pcabf43A gene was constitutively expressed in P. chrysogenum 31B at a low level, although the expression was induced with pectic components such as l-arabinose, l-rhamnose, and d-galacturonic acid. Analysis of enzymatic characteristics of PcABF43A, GH51 ABF (AFQ1), and GH54 ABF (AFS1) from P. chrysogenum suggested that PcABF43A and AFS1 function as debranching enzymes and AFQ1 plays a role of saccharification in the degradation of l-arabinan by this fungus.
KeywordMeSH Terms
Expression profile
Glycoside hydrolase family 43
Penicillium chrysogenum
l-Arabinan
α-l-Arabinofuranosidase
84. Barredo  JL, Cantoral  JM, Alvarez  E, Díez  B, Martín  JF,     ( 1989 )

Cloning, sequence analysis and transcriptional study of the isopenicillin N synthase of Penicillium chrysogenum AS-P-78.

Molecular & general genetics : MGG 216 (1)
PMID : 2499766  :   DOI  :   10.1007/bf00332235    
Abstract >>
A gene (ips) encoding the isopenicillin N synthase of Penicillium chrysogenum AS-P-78 was cloned in a 3.9 kb SalI fragment using a probe corresponding to the amino-terminal end of the enzyme. The SalI fragment was trimmed down to a 1.3 kb NcoI-BglII fragment that contained an open reading frame of 996 nucleotides encoding a polypeptide of 331 amino acids with an Mr of 38012 dalton. The predicted polypeptide encoded by the ips gene of strain AS-P-78 contains a tyrosine at position 195, whereas the gene of the high penicillin producing strain 23X-80-269-37-2 shows an isoleucine at the same position. The ips gene is expressed in Escherichia coli minicells using the lambda phage PL promoter. Some similar sequence motifs were found in the upstream region of the ips gene of P. chrysogenum when compared with the upstream sequences of the ips genes of Cephalosporium acremonium and Aspergillus nidulans. Primer extension studies indicated that the start of the mRNA coincides with a T in position -11 which is located in a conserved pyrimidine-rich sequence, near two CAAG boxes. Clones of P. chrysogenum Wis 54-1255 transformed with the ips gene showed a five-fold higher isopenicillin N synthase activity than the untransformed cultures.
KeywordMeSH Terms
Genes, Fungal
Oxidoreductases
85. Park  MS, Fong  JJ, Oh  SY, Kwon  KK, Sohn  JH, Lim  YW,     ( 2014 )

Marine-derived Penicillium in Korea: diversity, enzyme activity, and antifungal properties.

Antonie van Leeuwenhoek 106 (2)
PMID : 24908060  :   DOI  :   10.1007/s10482-014-0205-5    
Abstract >>
The diversity of marine-derived Penicillium from Korea was investigated using morphological and multigene phylogenetic approaches, analyzing sequences of the internal transcribed spacer region, �]-tubulin gene, and RNA polymerase subunit II gene. In addition, the biological activity of all isolated strains was evaluated. We tested for the extracellular enzyme activity of alginase, endoglucanase, and �]-glucosidase, and antifungal activity against two plant pathogens (Colletotrichum acutatum and Fusarium oxysporum). A total of 184 strains of 36 Penicillium species were isolated, with 27 species being identified. The most common species were Penicillium polonicum (19.6 %), P. rubens (11.4 %), P. chrysogenum (11.4 %), and P. crustosum (10.9 %). The diversity of Penicillium strains isolated from soil (foreshore soil and sand) and marine macroorganisms was higher than the diversity of strains isolated from seawater. While many of the isolated strains showed alginase and �]-glucosidase activity, no endoglucanase activity was found. More than half the strains (50.5 %) showed antifungal activity against at least one of the plant pathogens tested. Compared with other strains in this study, P. citrinum (strain SFC20140101-M662) showed high antifungal activity against both plant pathogens. The results reported here expand our knowledge of marine-derived Penicillium diversity. The relatively high proportion of strains that showed antifungal and enzyme activity demonstrates that marine-derived Penicillium have great potential to be used in the production of natural bioactive products for pharmaceutical and/or industrial use.
KeywordMeSH Terms
86. Specht  T, Dahlmann  TA, Zadra  I, Kürnsteiner  H, Kück  U,     ( 2014 )

Complete Sequencing and Chromosome-Scale Genome Assembly of the Industrial Progenitor Strain P2niaD18 from the Penicillin Producer Penicillium chrysogenum.

Genome announcements 2 (4)
PMID : 25059858  :   DOI  :   10.1128/genomeA.00577-14     PMC  :   PMC4110216    
Abstract >>
Penicillium chrysogenum is the major industrial producer of the �]-lactam antibiotic penicillin. Here, we report the complete genome sequence of the industrial progenitor strain P. chrysogenum P2niaD18 in a chromosome-scale genome assembly. P2niaD18 is distinguished from the recently sequenced P. chrysogenum Wisconsin 54-1255 strain by major chromosomal rearrangements leading to a modified chromosomal architecture.
KeywordMeSH Terms
87. Lecellier  A, Mounier  J, Gaydou  V, Castrec  L, Barbier  G, Ablain  W, Manfait  M, Toubas  D, Sockalingum  GD,     ( 2014 )

Differentiation and identification of filamentous fungi by high-throughput FTIR spectroscopic analysis of mycelia.

International journal of food microbiology 168-169 (N/A)
PMID : 24231128  :   DOI  :   10.1016/j.ijfoodmicro.2013.10.011    
Abstract >>
Routine identification of fungi based on phenotypic and genotypic methods can be fastidious and time-consuming. In this context, there is a constant need for new approaches allowing the rapid identification of molds. Fourier-transform infrared (FTIR) spectroscopy appears as such an indicated method. The objective of this work was to evaluate the potential of FTIR spectroscopy for an early differentiation and identification of filamentous fungi. One hundred and thirty-one strains identified using DNA sequencing, were analyzed using FTIR spectroscopy of the mycelia obtained after a reduced culture time of 48 h compared to current conventional methods. Partial least square discriminant analysis was used as a chemometric method to analyze the spectral data and for identification of the fungal strains from the phylum to the species level. Calibration models were constructed using 106 strains pertaining to 14 different genera and 32 species and were used to identify 25 fungal strains in a blind manner. Identification levels of 98.97% and 98.77% achieved were correctly assigned to the genus and species levels respectively. FTIR spectroscopy with its high discriminating power and rapidity therefore shows strong promise for routine fungal identification. Upgrading of our database is ongoing to test the technique's robustness.
KeywordMeSH Terms
Fungi
High-throughput FTIR spectroscopy
Identification
PLS-DA
Fungi
High-throughput FTIR spectroscopy
Identification
PLS-DA
Fungi
High-throughput FTIR spectroscopy
Identification
PLS-DA
Fungi
High-throughput FTIR spectroscopy
Identification
PLS-DA
Fungi
High-throughput FTIR spectroscopy
Identification
PLS-DA
Fungi
High-throughput FTIR spectroscopy
Identification
PLS-DA
Spectroscopy, Fourier Transform Infrared
88. Lopes  FC, Tichota  DM, Pereira  JQ, Segalin  J, Rios  Ade O, Brandelli  A,     ( 2013 )

Pigment production by filamentous fungi on agro-industrial byproducts: an eco-friendly alternative.

Applied biochemistry and biotechnology 171 (3)
PMID : 23873642  :   DOI  :   10.1007/s12010-013-0392-y    
Abstract >>
The search for new sources of natural pigments has increased, mainly because of the toxic effects caused by synthetic dyes used in food, pharmaceutical, textile, and cosmetic industries. Fungi provide a readily available alternative source of natural pigments. In this context, the fungi Penicillium chrysogenum IFL1 and IFL2, Fusarium graminearum IFL3, Monascus purpureus NRRL 1992, and Penicillium vasconiae IFL4 were selected as pigments producers. The fungal identification was performed using ITS and part of the �]-tubulin gene sequencing. Almost all fungi were able to grow and produce water-soluble pigments on agro-industrial residues, with the exception of P. vasconiae that produced pigments only on potato dextrose broth. The production of yellow pigments was predominant and the two strains of P. chrysogenum were the largest producers. In addition, the production of pigments and mycotoxins were evaluated in potato dextrose agar using TOF-MS and TOF-MS/MS. Metabolites as roquefortine C, chrysogine were found in both extracts of P. chrysogenum, as well fusarenone X, diacetoxyscirpenol, and neosolaniol in F. graminearum extract. In the M. purpureus extract, the pigments monascorubrin, rubropunctatin, and the mycotoxin citrinin were found. The crude filtrates have potential to be used in the textile industry; nevertheless, additional pigment purification is required for food and pharmaceutical applications.
KeywordMeSH Terms
89. Chen  AJ, Tang  D, Zhou  YQ, Sun  BD, Li  XJ, Wang  LZ, Gao  WW,     ( 2013 )

Identification of ochratoxin A producing fungi associated with fresh and dry liquorice.

PloS one 8 (10)
PMID : 24205182  :   DOI  :   10.1371/journal.pone.0078285     PMC  :   PMC3804526    
Abstract >>
The presence of fungi on liquorice could contaminate the crop and result in elevated levels of mycotoxin. In this study, the mycobiota associated with fresh and dry liquorice was investigated in 3 producing regions of China. Potential toxigenic fungi were tested for ochratoxin A (OTA) and aflatoxin B1 (AFB1) production using liquid chromatography/mass spectrometry/mass spectrometry. Based on a polyphasic approach using morphological characters, �]-tubulin and RNA polymerase II second largest subunit gene phylogeny, a total of 9 genera consisting of 22 fungal species were identified, including two new Penicillium species (Penicillium glycyrrhizacola sp. nov. and Penicillium xingjiangense sp. nov.). The similarity of fungal communities associated with fresh and dry liquorice was low. Nineteen species belonging to 8 genera were detected from fresh liquorice with populations affiliated with P. glycyrrhizacola, P. chrysogenum and Aspergillus insuetus comprising the majority (78.74%, 33.33% and 47.06% of total) of the community from Gansu, Ningxia and Xinjiang samples, respectively. In contrast, ten species belonging to 4 genera were detected from dry liquorice with populations affiliated with P. chrysogenum, P. crustosum and Aspergillus terreus comprising the majority (64.00%, 52.38% and 90.91% of total) of the community from Gansu, Ningxia and Xinjiang samples, respectively. Subsequent LC/MS/MS analysis indicated that 5 fungal species were able to synthesize OTA in vitro including P. chrysogenum, P. glycyrrhizacola, P. polonicum, Aspergillus ochraceus and A. westerdijkiae, the OTA concentration varied from 12.99 to 39.03 ?g/kg. AFB1 was absent in all tested strains. These results demonstrate the presence of OTA producing fungi on fresh liquorice and suggest that these fungi could survive on dry liquorice after traditional sun drying. Penicillium chrysogenum derived from surrounding environments is likely to be a stable contributor to high OTA level in liquorice. The harvesting and processing procedure needs to be monitored in order to keep liquorice free of toxigenic fungi.
KeywordMeSH Terms
90. Walker  DM, White  JF, Rossman  AY,     ( 2012 )

New molecular markers for fungal phylogenetics: two genes for species-level systematics in the Sordariomycetes (Ascomycota).

Molecular phylogenetics and evolution 64 (3)
PMID : 22626621  :   DOI  :   10.1016/j.ympev.2012.05.005    
Abstract >>
Although significant progress has been made resolving deep branches of the fungal tree of life, many fungal systematists are interested in species-level questions to both define species and assess fungal biodiversity. Fungal genome sequences are a useful resource to systematic biologists for developing new phylogenetic markers that better represent the whole genome. Here we report primers for two newly identified single-copy protein-coding genes, FG1093 and MS204, for use with ascomycetes. Although fungi were the focus of this study, this methodological approach could be easily applied to marker development for studies of other organisms. The tests used here to assess phylogenetic informativeness are computationally rapid, require only rudimentary datasets to evaluate existing or newly developed markers, and can be applied to other non-model organisms to assist in experimental design of phylogenetic studies. Phylogenetic utility of the markers was tested in two genera, Gnomoniopsis and Ophiognomonia (Gnomoniaceae, Diaporthales). The phylogenetic performance of �]-tubulin, ITS, and tef-1�\ was compared with FG1093 and MS204. Phylogenies inferred from FG1093 and MS204 were largely in agreement with �]-tubulin, ITS, and tef-1�\ although some topological conflict was observed. Resolution and support for branches differed based on the combination of markers used for each genus. Based on two independent tests of phylogenetic performance, FG1093 and MS204 were determined to be equal to or better than �]-tubulin, ITS, and tef-1�\ in resolving species relationships. Differences were found in site-specific rate of evolution in all five markers. In addition, isolates from 15 orders and 22 families of Ascomycota were screened using primers for FG1093 and MS204 to demonstrate primer utility across a wide diversity of ascomycetes. The primer sets for the newly identified genes FG1093 and MS204 and methods used to develop them are useful additions to the ascomycete systematists' toolbox.
KeywordMeSH Terms
Evolution, Molecular
Genes, Fungal
Phylogeny
91. Houbraken  J, Samson  RA,     ( 2011 )

Phylogeny of Penicillium and the segregation of Trichocomaceae into three families.

Studies in mycology 70 (1)
PMID : 22308045  :   DOI  :   10.3114/sim.2011.70.01     PMC  :   PMC3233907    
Abstract >>
Species of Trichocomaceae occur commonly and are important to both industry and medicine. They are associated with food spoilage and mycotoxin production and can occur in the indoor environment, causing health hazards by the formation of �]-glucans, mycotoxins and surface proteins. Some species are opportunistic pathogens, while others are exploited in biotechnology for the production of enzymes, antibiotics and other products. Penicillium belongs phylogenetically to Trichocomaceae and more than 250 species are currently accepted in this genus. In this study, we investigated the relationship of Penicillium to other genera of Trichocomaceae and studied in detail the phylogeny of the genus itself. In order to study these relationships, partial RPB1, RPB2 (RNA polymerase II genes), Tsr1 (putative ribosome biogenesis protein) and Cct8 (putative chaperonin complex component TCP-1) gene sequences were obtained. The Trichocomaceae are divided in three separate families: Aspergillaceae, Thermoascaceae and Trichocomaceae. The Aspergillaceae are characterised by the formation flask-shaped or cylindrical phialides, asci produced inside cleistothecia or surrounded by H?lle cells and mainly ascospores with a furrow or slit, while the Trichocomaceae are defined by the formation of lanceolate phialides, asci borne within a tuft or layer of loose hyphae and ascospores lacking a slit. Thermoascus and Paecilomyces, both members of Thermoascaceae, also form ascospores lacking a furrow or slit, but are differentiated from Trichocomaceae by the production of asci from croziers and their thermotolerant or thermophilic nature. Phylogenetic analysis shows that Penicillium is polyphyletic. The genus is re-defined and a monophyletic genus for both anamorphs and teleomorphs is created (Penicillium sensu stricto). The genera Thysanophora, Eupenicillium, Chromocleista, Hemicarpenteles and Torulomyces belong in Penicilliums. str. and new combinations for the species belonging to these genera are proposed. Analysis of Penicillium below genus rank revealed the presence of 25 clades. A new classification system including both anamorph and teleomorph species is proposed and these 25 clades are treated here as sections. An overview of species belonging to each section is presented. New sections, all in Penicillium: sect. Sclerotiora Houbraken & Samson, sect. Charlesia Houbraken & Samson, sect. Thysanophora Houbraken & Samson,sect. Ochrosalmonea Houbraken & Samson, sect. Cinnamopurpurea Houbraken & Samson, Fracta Houbraken & Samson, sect. Stolkia Houbraken & Samson, sect. Gracilenta Houbraken & Samson, sect. Citrina Houbraken & Samson, sect. Turbata Houbraken & Samson, sect. Paradoxa Houbraken & Samson, sect. Canescentia Houbraken & Samson. New combinations:Penicillium asymmetricum (Subramanian & Sudha) Houbraken & Samson, P. bovifimosum (Tuthill & Frisvad) Houbraken & Samson, P. glaucoalbidum (Desmazi?res) Houbraken & Samson, P. laeve (K. Ando & Manoch) Houbraken & Samson, P. longisporum (Kendrick) Houbraken & Samson, P. malachiteum (Yaguchi & Udagawa) Houbraken & Samson, P. ovatum (K. Ando & Nawawi) Houbraken & Samson, P. parviverrucosum (K. Ando & Pitt) Houbraken & Samson, P. saturniforme (Wang & Zhuang) Houbraken & Samson, P. taiwanense (Matsushima) Houbraken & Samson. New names:Penicillium coniferophilum Houbraken & Samson, P. hennebertii Houbraken & Samson, P. melanostipe Houbraken & Samson, P. porphyreum Houbraken & Samson.
KeywordMeSH Terms
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
Aspergillus
Eupenicillium
Penicillium
Talaromyces
nomenclature
taxonomy.
92. Schoch  CL, Seifert  KA, Huhndorf  S, Robert  V, Spouge  JL, Levesque  CA, Chen  W, N/A  N/A, N/A  N/A,     ( 2012 )

Nuclear ribosomal internal transcribed spacer (ITS) region as a universal DNA barcode marker for Fungi.

Proceedings of the National Academy of Sciences of the United States of America 109 (16)
PMID : 22454494  :   DOI  :   10.1073/pnas.1117018109     PMC  :   PMC3341068    
Abstract >>
Six DNA regions were evaluated as potential DNA barcodes for Fungi, the second largest kingdom of eukaryotic life, by a multinational, multilaboratory consortium. The region of the mitochondrial cytochrome c oxidase subunit 1 used as the animal barcode was excluded as a potential marker, because it is difficult to amplify in fungi, often includes large introns, and can be insufficiently variable. Three subunits from the nuclear ribosomal RNA cistron were compared together with regions of three representative protein-coding genes (largest subunit of RNA polymerase II, second largest subunit of RNA polymerase II, and minichromosome maintenance protein). Although the protein-coding gene regions often had a higher percent of correct identification compared with ribosomal markers, low PCR amplification and sequencing success eliminated them as candidates for a universal fungal barcode. Among the regions of the ribosomal cistron, the internal transcribed spacer (ITS) region has the highest probability of successful identification for the broadest range of fungi, with the most clearly defined barcode gap between inter- and intraspecific variation. The nuclear ribosomal large subunit, a popular phylogenetic marker in certain groups, had superior species resolution in some taxonomic groups, such as the early diverging lineages and the ascomycete yeasts, but was otherwise slightly inferior to the ITS. The nuclear ribosomal small subunit has poor species-level resolution in fungi. ITS will be formally proposed for adoption as the primary fungal barcode marker to the Consortium for the Barcode of Life, with the possibility that supplementary barcodes may be developed for particular narrowly circumscribed taxonomic groups.
KeywordMeSH Terms
93. Yu  ZL, Liu  J, Wang  FQ, Dai  M, Zhao  BH, He  JG, Zhang  H,     ( 2011 )

Cloning and characterization of a novel CoA-ligase gene from Penicillium chrysogenum.

Folia microbiologica 56 (3)
PMID : 21625874  :   DOI  :   10.1007/s12223-011-0044-y    
Abstract >>
A novel phenylacetic acid (PAA)-induced CoA-ligase-encoding gene, designated as phlC, has been cloned from penicillin-producing fungus Penicillium chrysogenum. The open reading frame of phlC cDNA was 1671 bp and encoded a 556 amino acid residues protein with the consensus AMP binding site and a peroxisomal targeting signal 1 on its C terminus. The deduced amino acid sequence showed 37% and 38% identity with characterized P. chrysogenum Phl and PhlB protein, respectively. Functional recombinant PhlC protein was overexpressed in Escherichia coli. The purified recombinant enzyme was capable to convert PAA into its corresponding CoA ester with a specific activity of 129.5 �� 3.026 pmol/min per mg protein. Similar to Phl and PhlB, PhlC displayed broad substrate spectrum and showed higher activities to medium- and long-chain fatty acids. The catalytic properties of PhlC have been determined and compared to those of Phl and PhlB.
KeywordMeSH Terms
94. Henk  DA, Eagle  CE, Brown  K, Van Den Berg  MA, Dyer  PS, Peterson  SW, Fisher  MC,     ( 2011 )

Speciation despite globally overlapping distributions in Penicillium chrysogenum: the population genetics of Alexander Fleming's lucky fungus.

Molecular ecology 20 (20)
PMID : 21951491  :   DOI  :   10.1111/j.1365-294X.2011.05244.x    
Abstract >>
Eighty years ago, Alexander Fleming described the antibiotic effects of a fungus that had contaminated his bacterial culture, kick starting the antimicrobial revolution. The fungus was later ascribed to a putatively globally distributed asexual species, Penicillium chrysogenum. Recently, the species has been shown to be genetically diverse, and possess mating-type genes. Here, phylogenetic and population genetic analyses show that this apparently ubiquitous fungus is actually composed of at least two genetically distinct species with only slight differences detected in physiology. We found each species in air and dust samples collected in and around St Mary's Hospital where Fleming worked. Genotyping of 30 markers across the genome showed that preserved fungal material from Fleming's laboratory was nearly identical to derived strains currently in culture collections and in the same distinct species as a wild progenitor strain of current penicillin producing industrial strains rather than the type species P. chrysogenum. Global samples of the two most common species were found to possess mating-type genes in a near 1:1 ratio, and show evidence of recombination with little geographic population subdivision evident. However, no hybridization was detected between the species despite an estimated time of divergence of less than 1MYA. Growth studies showed significant interspecific inhibition by P. chrysogenum of the other common species, suggesting that competition may facilitate species maintenance despite globally overlapping distributions. Results highlight under-recognized diversity even among the best-known fungal groups and the potential for speciation despite overlapping distribution.
KeywordMeSH Terms
Alexander Fleming
Penicillium flemingii
asexual
microsatellite
speciation
Genetic Speciation
Phylogeny
95. Balabanova  LA, Gafurov  YM, Pivkin  MV, Terentyeva  NA, Likhatskaya  GN, Rasskazov  VA,     ( 2012 )

An extracellular S1-type nuclease of marine fungus Penicillium melinii.

Marine biotechnology (New York, N.Y.) 14 (1)
PMID : 21647618  :   DOI  :   10.1007/s10126-011-9392-5    
Abstract >>
An extracellular nuclease was purified 165-fold with a specific activity of 41,250 U/mg poly(U) by chromatography with modified chitosan from the culture of marine fungus Penicillium melinii isolated from colonial ascidium collected near Shikotan Island, Sea of Okhotsk, at a depth of 123 m. The purified nuclease is a monomer with the molecular weight of 35 kDa. The enzyme exhibits maximum activity at pH 3.7 for DNA and RNA. The enzyme is stable until 75�XC and in the pH range of 2.5-8.0. The enzyme endonucleolytically degrades ssDNA and RNA by 3'-5' mode to produce 5'-oligonucleotides and 5'-mononucleotides; however, it preferentially degrades poly(U). The enzyme can digest dsDNA in the presence of pregnancy-specific beta-1-glycoprotein-1. The nuclease acts on closed circular double-stranded DNA to produce opened circular DNA and then the linear form DNA by single-strand scission. DNA sequence encoding the marine fungus P. melinii endonuclease revealed homology to S1-type nucleases. The tight correlation found between the extracellular endonuclease activity and the rate of H?-thymidine uptake by actively growing P. melinii cells suggests that this nuclease is required for fulfilling the nucleotide pool of precursors of DNA biosynthesis during the transformation of hyphae into the aerial mycelium and conidia in stressful environmental conditions.
KeywordMeSH Terms
96. Gao  Z, Li  Z, Zhang  Y, Huang  H, Li  M, Zhou  L, Tang  Y, Yao  B, Zhang  W,     ( 2012 )

High-level expression of the Penicillium notatum glucose oxidase gene in Pichia pastoris using codon optimization.

Biotechnology letters 34 (3)
PMID : 22052258  :   DOI  :   10.1007/s10529-011-0790-6    
Abstract >>
The glucose oxidase (GOD) gene from Penicillium notatum was expressed in Pichia pastoris. The 1,815 bp gene, god-w, encodes 604 amino acids. Recombinant GOD-w had optimal activity at 35-40�XC and pH 6.2 and was stable, from pH 3 to 7 maintaining >75% maximum activity after incubation at 50�XC for 1 h. GOD-w worked as well as commercial GODs to improve bread making. To achieve high-level expression of recombinant GOD in P. pastoris, 272 nucleotides involving 228 residues were mutated, consistent with the codon bias of P. pastoris. The optimized recombinant GOD-m yielded 615 U ml(-1) (2.5 g protein l(-1)) in a 3 l fermentor--410% higher than GOD-w (148 U ml(-1)), and thus is a low-cost alternative for the bread baking industry.
KeywordMeSH Terms
Gene Expression
97. Veiga  T, Solis-Escalante  D, Romagnoli  G, ten Pierick  A, Hanemaaijer  M, Deshmukh  AT, Deshmuhk  A, Wahl  A, Pronk  JT, Daran  JM,     ( 2012 )

Resolving phenylalanine metabolism sheds light on natural synthesis of penicillin G in Penicillium chrysogenum.

Eukaryotic cell 11 (2)
PMID : 22158714  :   DOI  :   10.1128/EC.05285-11     PMC  :   PMC3272894    
Abstract >>
The industrial production of penicillin G by Penicillium chrysogenum requires the supplementation of the growth medium with the side chain precursor phenylacetate. The growth of P. chrysogenum with phenylalanine as the sole nitrogen source resulted in the extracellular production of phenylacetate and penicillin G. To analyze this natural pathway for penicillin G production, chemostat cultures were switched to [U-(13)C]phenylalanine as the nitrogen source. The quantification and modeling of the dynamics of labeled metabolites indicated that phenylalanine was (i) incorporated in nascent protein, (ii) transaminated to phenylpyruvate and further converted by oxidation or by decarboxylation, and (iii) hydroxylated to tyrosine and subsequently metabolized via the homogentisate pathway. The involvement of the homogentisate pathway was supported by the comparative transcriptome analysis of P. chrysogenum cultures grown with phenylalanine and with (NH(4))(2)SO(4) as the nitrogen source. This transcriptome analysis also enabled the identification of two putative 2-oxo acid decarboxylase genes (Pc13g9300 and Pc18g01490). cDNAs of both genes were cloned and expressed in the 2-oxo-acid-decarboxylase-free Saccharomyces cerevisiae strain CEN.PK711-7C (pdc1 pdc5 pdc6�G aro10�G thi3�G). The introduction of Pc13g09300 restored the growth of this S. cerevisiae mutant on glucose and phenylalanine, thereby demonstrating that Pc13g09300 encodes a dual-substrate pyruvate and phenylpyruvate decarboxylase, which plays a key role in an Ehrlich-type pathway for the production of phenylacetate in P. chrysogenum. These results provide a basis for the metabolic engineering of P. chrysogenum for the production of the penicillin G side chain precursor phenylacetate.
KeywordMeSH Terms
98.     ( 1997 )

Molecular analysis of a Penicillium chrysogenum GATA factor encoding gene (sreP) exhibiting significant homology to the Ustilago maydis urbs1 gene.

Gene 184 (1)
PMID : 9016950  :   DOI  :   10.1016/s0378-1119(96)00570-7    
Abstract >>
Employing a PCR-aided strategy, a Penicillium chrysogenum gene (sreP) encoding a putative GATA-transcription factor has been cloned and characterized. Comparison of the genomic and cDNA sequences revealed the presence of an open reading frame (ORF) encoding a protein of 532 amino acids (aa) which is interrupted by two introns. The deduced aa sequence of sreP reveals 50% identity to a regulator of siderophore biosynthesis (URBS1) from Ustilago maydis over a stretch of 200 aa containing two GATA-type zinc finger motifs and a Cys-rich intervening sequence. Northern blot analysis indicated two transcripts of 2.2 and 2.7 kb in approximately equivalent amount. due to two major transcription start sites.
KeywordMeSH Terms
CCAAT-Enhancer-Binding Proteins
99.     ( 1996 )

Cloning and characterization of chitin synthase gene fragments from Penicillium chrysogenum.

FEMS microbiology letters 145 (1)
PMID : 8931329  :   DOI  :   10.1111/j.1574-6968.1996.tb08558.x    
Abstract >>
DNA fragments homologous to chitin synthase were amplified from the genomic DNA of Penicillium chrysogenum by PCR. Cloning and sequencing of the PCR-amplified fragments led to the identification of four different genes, designated PcCHS1, PcCHS2, PcCHS3, and PcCHS4. By comparison of the deduced amino acid sequences, PcCHS1 was identified as a gene for class I chitin synthase, PcCHS2 and PcCHS3 were for class II, and PcCHS4 was for class III. Among these only PcCHS4 includes an intervening sequence of 56 bp. The analysis of the deduced amino acid sequences revealed a close evolutionary relationship between Penicillium and ascomycetous fungi.
KeywordMeSH Terms
100.     ( 1993 )

Genomic sequence of mitochondrial genes coding for ATPase subunit 6 and small subunit ribosomal RNA from Penicillium chrysogenum: a key for molecular systematics on fungi.

Nucleic acids research 21 (18)
PMID : 8414999  :   DOI  :   10.1093/nar/21.18.4393     PMC  :   PMC310079    
Abstract >>
N/A
KeywordMeSH Terms
101.     ( 1995 )

NRE, the major nitrogen regulatory protein of Penicillium chrysogenum, binds specifically to elements in the intergenic promoter regions of nitrate assimilation and penicillin biosynthetic gene clusters.

Current genetics 28 (2)
PMID : 8590470  :  
Abstract >>
NRE, the nitrogen regulatory protein of Penicillium chrysogenum, contains a single Cys2/Cys2-type zinc-finger motif followed immediately by a highly basic region. The zinc-finger domain was expressed to Escherichia coli as a fusion protein with beta-galactosidase. In order to test the putative DNA-binding ability of NRE, the intergenic promoter region of the nitrate reductase/nitrite reductase gene cluster (niiA-niaD) of Penicillium was sequenced. Our results show that NRE is a DNA-binding protein and binds to the intergenic promoter regions of the P. chrysogenum niiA-niaD and acvA-pcbC gene cluster, encoding the first two enzymes in penicillin biosynthesis. Three of the four high-affinity NRE-binding sites contained two GATA core elements. In one of the recognition sites for NRE, one GATA motif was replaced by GATT. The two GATA elements showed all possible orientations, head-to-head, head-to-tail and tail-to-tail, and were separated by between 4 and 27 bp. Missing-contact analysis showed that all three purines in both of the GATA core sequences and the single adenine residue in each of the complementary TATC sequences were involved in the binding of NRE. Moreover, loss of purines in the flanking regions of the GATA elements also affect binding of NRE, as their loss causes reduced affinity.
KeywordMeSH Terms
Promoter Regions, Genetic
102.     ( 1996 )

Characterization of a Penicillium chrysogenum gene encoding a PacC transcription factor and its binding sites in the divergent pcbAB-pcbC promoter of the penicillin biosynthetic cluster.

Molecular microbiology 20 (3)
PMID : 8736532  :   DOI  :   10.1046/j.1365-2958.1996.5421065.x    
Abstract >>
Previous work established that pH regulation of gene expression in Aspergillus nidulans, a major determinant of penicillin biosynthesis, is mediated by the zinc-finger transcription factor PacC, an activator of transcription of the isopenicillin N synthase gene. We characterize here the pacC gene from the efficient penicillin producer Penicillium chrysogenum, which functionally complements an A. nidulans pacC null mutation. It encodes a 641-residue polypeptide showing 64% identify to A. nidulans PacC and containing three putative zinc fingers specifically recognizing a 5'-GCCARG-3' hexanucleotide. Penicillium pacC transcript levels are higher under alkaline than under acidic growth conditions and elevate at late stages of growth. The gene contains three PacC-binding sites in its 5'-upstream region. Transcript levels of pcbC (encoding P. chrysogenum isopenicillin N synthase) are low on a repressing carbon source and elevated on a derepressing carbon source. With either carbon source, alkaline pH elevates pcbC transcript levels, correlating with the presence of seven PacC-binding sites in the 1.1 kb pcbAB-pcbC intergenic region and strongly suggesting that pcbC is under direct pacC control. However, in contrast to the situation in A. nidulans, alkaline pH does not override the negative effects of a repressing carbon source, revealing differences in the regulation of the penicillin pathway between Penicillium and Aspergillus.
KeywordMeSH Terms
Fungal Proteins
Multigene Family
Promoter Regions, Genetic
103.     ( 1993 )

Investigations into the post-translational modification and mechanism of isopenicillin N:acyl-CoA acyltransferase using electrospray mass spectrometry.

The Biochemical journal 294 (Pt 2) (N/A)
PMID : 8396910  :   DOI  :   10.1042/bj2940357     PMC  :   PMC1134462    
Abstract >>
Electrospray mass spectrometry (e.s.m.s.) was used to confirm the position of the post-translational cleavage of the isopenicillin N:acyl-CoA acyltransferase preprotein to give the alpha- and beta-subunits. The e.s.m.s. studies suggested partial modification of the alpha-subunit in vivo by exogenously added substituted acetic acids. E.s.m.s. has also allowed the observation in vitro of the transfer of the acyl group from several acyl-CoAs to the beta-subunit. N.m.r. data for the CoA species have been deposited as Supplementary Publication SUP 500173 (2 pages) at the British Library Document Supply Centre (DSC), Boston Spa, Wetherby, West Yorkshire LS23 7BQ, from whom copies can be obtained on the terms indicated in Biochem. J. (1993) 289, 9.
KeywordMeSH Terms
Mass Spectrometry
Penicillin-Binding Proteins
Protein Processing, Post-Translational
104.     ( 1994 )

The thioredoxin system of Penicillium chrysogenum and its possible role in penicillin biosynthesis.

Journal of bacteriology 176 (4)
PMID : 8106340  :   DOI  :   10.1128/jb.176.4.973-984.1994     PMC  :   PMC205147    
Abstract >>
Penicillium chrysogenum is an important producer of penicillin antibiotics. A key step in their biosynthesis is the oxidative cyclization of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine (ACV) to isopenicillin N by the enzyme isopenicillin N synthase (IPNS). bis-ACV, the oxidized disulfide form of ACV is, however, not a substrate for IPNS. We report here the characterization of a broad-range disulfide reductase from P. chrysogenum that efficiently reduces bis-ACV to the thiol monomer. When coupled in vitro with IPNS, it converts bis-ACV to isopenicillin N and may therefore play a role in penicillin biosynthesis. The disulfide reductase consists of two protein components, a 72-kDa NADPH-dependent reductase, containing two identical subunits, and a 12-kDa general disulfide reductant. The latter reduces disulfide bonds in low-molecular-weight compounds and in proteins. The genes coding for the reductase system were cloned and sequenced. Both possess introns. A comparative analysis of their predicted amino acid sequences showed that the 12-kDa protein shares 26 to 60% sequence identity with thioredoxins and that the 36-kDa protein subunit shares 44 to 49% sequence identity with the two known bacterial thioredoxin reductases. In addition, the P. chrysogenum NADPH-dependent reductase is able to accept thioredoxin as a substrate. These results establish that the P. chrysogenum broad-range disulfide reductase is a member of the thioredoxin family of oxidoreductases. This is the first example of the cloning of a eucaryotic thioredoxin reductase gene.
KeywordMeSH Terms
105.     ( 1993 )

Cloning and structural organization of a xylanase-encoding gene from penicillium chrysogenum.

Gene 126 (2)
PMID : 8482539  :   DOI  :   10.1016/0378-1119(93)90372-a    
Abstract >>
The filamentous fungus, Penicillium chrysogenum, is able to grow on xylan as a sole carbon source. Under these conditions, high levels of a xylanase (XYLP) are secreted into the medium. After purification and characterization of this enzyme, we have isolated both the encoding cDNA and the genomic sequence by using oligodeoxyribonucleotides derived from partial amino acid (aa) sequences of the purified enzyme. The gene is approximately 1.6 kb in length, and comparison of the nucleotide (nt) sequence of the genomic and the cDNA clone revealed the presence of ten exons and nine introns. All intron/exon splice junctions exactly follow the GT/AG rule, except for the seventh intron which shows atypical AT/AC splice sites. The immediate 5'-flanking region of the first exon contains one putative CCAAT consensus sequence and a perfect TATA box. Primer extension analysis revealed two transcription start points located 38 and 34 nt upstream from the ATG start codon. A sequence of 23 aa representing a typical signal peptide is present at the N terminus of the deduced aa sequence. Northern blot analysis of total cellular RNA indicated that xylP encodes a 1.3-kb transcript which is induced by xylan. The aa sequence of XYLP shows considerable homology to high-M(r) acidic xylanases (Xln) and cellulases from different bacteria, yeasts and fungi.
KeywordMeSH Terms
106.     ( 1995 )

Cloning, structural organization and regulation of expression of the Penicillium chrysogenum paf gene encoding an abundantly secreted protein with antifungal activity.

Gene 167 (1��2��)
PMID : 8566771  :   DOI  :   10.1016/0378-1119(95)00701-6    
Abstract >>
An abundantly secreted, highly basic 12-kDa protein (PAF) was purified from the culture medium of Penicillium chrysogenum (Pc). Based on the N-terminal amino acid (aa) sequence of the protein, an oligodeoxyribonucleotide probe was derived and used for amplification of the encoding cDNA by PCR. This cDNA fragment encodes a Cys-rich preproprotein of 92 aa which appears to be processed to a mature product of 55 aa. The deduced aa sequence of the preproprotein reveals 42.6% identity to an antifungal protein (AFP) of Aspergillus giganteus. Agar diffusion tests confirmed that the Pc protein exhibits antifungal activity. In order to investigate the promoter region and the structural organization of the paf gene, a genomic 6-kb fragment was isolated and partially sequenced. Comparison of the nucleotide sequence of the genomic fragment and the cDNA clone revealed the presence of a coding region of 279 bp which is interrupted by two introns of 76 and 68 bp in length. In the promoter region, a typical TATA box, a motif resembling the fungal carbon catabolite repression element, as well as several putative GATA factor binding motifs, were found. Northern blot analysis indicated that the regulation of paf expression occurs at the level of mRNA transcription and is under control of carbon catabolite and nitrogen metabolite repression regulatory circuits.
KeywordMeSH Terms
Antifungal Agents
107.     ( 1994 )

Expression of the 3-phosphoglycerate kinase gene (pgkA) of Penicillium chrysogenum.

Molecular & general genetics : MGG 243 (3)
PMID : 8190080  :   DOI  :   10.1007/bf00301062    
Abstract >>
The sequence of the Penicillium chrysogenum pgkA gene promoter was determined up to 952 nucleotides (nt) 5' to the major transcriptional start point (position +1), and contains a 38 bp pyrimidine-rich region within which transcription initiates at this and two minor sites (-11, -23). A 21 bp segment (-99 to -79) closely matches a region which is essential for the expression of the Aspergillus nidulans pgkA gene. A further region was found with similarity to sequences in other A. nidulans promoters possibly effecting response to carbon source. The terminator region of the P. chrysogenum pgkA gene was sequenced as far as 192 nt 3' to the stop codon and three polyadenylation sites were found at 94, 103 and 107 nt from this point, the first preceded by a possible polyadenylation signal. No transcription termination signal was found but several regions potentially forming stem-loop-structures were noted. A single 1.3 kb pgkA mRNA was readily detected by Northern blot analysis of total cellular RNA. Steady-state levels of pgkA mRNA were 1.5 to 2.0 times greater in mycelium harvested at similar stages of growth from medium containing the carbon sources acetate or quinate compared to glucose. A transformed strain of P. chrysogenum containing a fusion of the pgkA promoter to the Escherichia coli lacZ reporter gene integrated at the oliC locus was constructed, and beta-galactosidase activity monitored during growth of batch cultures in defined media. The pgkA promoter activity increased during exponential growth and was 2-3 times greater and increased most rapidly in mycelium grown on quinate or acetate compared to glucose.(ABSTRACT TRUNCATED AT 250 WORDS)
KeywordMeSH Terms
Genes, Fungal
108.     ( 1993 )

Characterisation of the gene encoding acetyl-CoA synthetase in Penicillium chrysogenum: conservation of intron position in plectomycetes.

Gene 130 (2)
PMID : 8103029  :   DOI  :   10.1016/0378-1119(93)90429-7    
Abstract >>
Acetyl-coenzyme A synthetase (ACS; EC 6.2.1.1) from some plectomycete fungi is possibly involved in an accessory step of penicillin biosynthesis, in addition to its role in primary metabolism. We present the characterisation of the gene encoding this enzyme in Penicillium chrysogenum, which we designated acuA. Sequencing of genomic and cDNA clones showed that the coding region was interrupted by five introns, located at the same positions as those present in the Aspergillus nidulans homologue. This supports the possibility that the gene acquired its definitive mosaic organisation before the Penicillium/Aspergillus divergence. The mature transcript encodes a polypeptide with an M(r) of 74,287 which is 89.4% identical to its A. nidulans counterpart.
KeywordMeSH Terms
Genes, Fungal
Introns
109.     ( 1994 )

Cloning and sequencing of ATP sulfurylase from Penicillium chrysogenum. Identification of a likely allosteric domain.

The Journal of biological chemistry 269 (31)
PMID : 8051058  :  
Abstract >>
Fungal (Penicillium chrysogenum) and yeast (Saccharomyces cerevisiae) ATP sulfurylases were shown to have very similar kinetic and chemical properties except that the fungal enzyme (a) contains a highly reactive Cys residue (SH-1) whose modification results in sigmoidal velocity curves (Renosto, F., Martin, R. L., and Segel, I. H. (1987) J. Biol. Chem. 262, 16279-16288) and (b) is allosterically inhibited by 3'-phosphoadenosine 5'-phosphosulfate (PAPS), while the yeast enzyme displays neither of these properties. The fungal enzyme subunit (64.3 kDa, 572 amino acids) is also larger than the yeast enzyme subunit (59.3 kDa, 521 amino acids). To correlate the unique allosteric properties of the fungal enzyme with specific structural features, we cloned and sequenced the ATP sulfurylase gene (aps) from P. chrysogenum. The yeast and fungal enzymes are homologous over the first 400 amino acids and contain two regions high in basic residues which are conserved in sulfurylases from Arabidopsis and the Riftia pachyptila (hydrothermal vent tube worm) chemolithotrophic symbiont. These regions may participate in forming the binding sites for MgATP2- and SO4(2-). The fungal enzyme has no sites for MgATP2- and SO4(2-). The fungal enzyme has no significant sequence homology to the yeast enzyme in the C-terminal 172 amino acids. This C-terminal region contains SH-1 (Cys-508) and has homology to MET14 (S. cerevisiae), CYSC (E. coli), and NODQ (Rhizobium meliloti), i.e. adenosine 5'-phosphosulfate (APS) kinase. The cumulative results suggest that (a) the allosteric PAPS binding site of P. chrysogenum ATP sulfurylase is located in the C-terminal domain of the protein and (b) that this domain may have evolved from APS kinase. In spite of the homology, this C-terminal region does not account for the APS kinase activity of P. chrysogenum. Fungal ATP sulfurylase has no significant homology to (or regulatory properties in common with) CYSD or CYSN, proteins reported to comprise E. coli ATP sulfurylase (Leyh, T., Vogt, T. F., and Suo, Y. (1992) J. Biol. Chem. 267, 10405-10410).
KeywordMeSH Terms
110. Shen  HD, Liaw  SF, Lin  WL, Ro  LH, Yang  HL, Han  SH,     ( 1995 )

Molecular cloning of cDNA coding for the 68 kDa allergen of Penicillium notatum using MoAbs.

Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology 25 (4)
PMID : 7600381  :   DOI  :   10.1111/j.1365-2222.1995.tb01053.x    
Abstract >>
To characterize the 68 kDa allergen of Penicillium notatum (also known as P. chrysogenum), a molecular antibody (MoAb) (P40) was previously generated. For cDNA cloning, three more MoAbs (3F, 5A3, 5G2) were generated in the present study. A mixture of all the four MoAbs was used in cloning of the gene coding for the 68 kDa allergen from a lambda gt11 cDNA library of P. chrysogenum. A cDNA clone (A6) with DNA insert of about 0.5 kb which encodes for the 3'-terminal nucleotide sequence of the 68 kDa allergen was obtained. The cloned sequence contained two putative N-glycosylation sites. The reduction in molecular weight from 68 to 62 kDa in immunoblotting after treatment of the crude extract of P. notatum with N-glycosidase F indicates that the 68 kDa allergen is a glycoprotein. Nucleotide sequence determination showed that 188 (54%) of the 348 nucleotides of the cDNA sequence obtained were identical to the same region of the nucleotide sequence of the beta-N-acetylglucosaminidase gene of Candida albicans. Although the cDNA clone obtained did not encode the full-length gene of the 68 kDa allergen, polypeptide expressed from the A6 cDNA showed positive immunological reactivities to all four MoAbs used in the cloning experiment and to IgE antibodies in sera of asthmatic patients. There was a loss of immunoblotting activity to the 68 kDa component after absorption of MoAb P40-containing culture supernatant with filters blotted on plaque lawns of cDNA clone A6. Moreover, the immunoblotting activity remained when the MoAbs affinity-purified with filters containing polypeptides encoded by the cDNA insert of clone A6 were used.(ABSTRACT TRUNCATED AT 250 WORDS)
KeywordMeSH Terms
Antibodies, Monoclonal
Cloning, Molecular
111. Haas  H, Bauer  B, Redl  B, Stöffler  G, Marzluf  GA,     ( 1995 )

Molecular cloning and analysis of nre, the major nitrogen regulatory gene of Penicillium chrysogenum.

Current genetics 27 (2)
PMID : 7788718  :  
Abstract >>
We have isolated the Penicillium chrysogenum nre gene which is homologous to the major nitrogen regulatory genes areA from Aspergillus nidulans and nit-2 from Neurospora crassa. Overall, nre shows 60% identity to areA and 30% identity to nit-2 at the amino-acid level. The gene encodes a protein of 835 amino-acid residues and contains a single Cys2/Cys2-type zinc finger with an adjacent basic region and a putative acidic activation region. In the DNA-binding domain, 98% of the amino-acid residues are identical in nre, areA and nit-2. The nre gene has been shown to be functional in N. crassa by heterologous complementation of a nit-2 mutant. Growth tests indicated that transformants could utilize nitrate, amino-acids, purines and amides as sole nitrogen sources. Nitrate reductase activity assays performed with transformants demonstrated that nitrogen control was completely normal. Complementation of N. crassa nit-2 mutants with 5'-deletion clones of nre suggests the possible presence of an internal promoter within the coding region. Northern analysis and ribonuclease protection assays of total cellular RNA indicated that nre encodes a 3.2-kb transcript which is reduced in content under conditions of nitrogen repression.
KeywordMeSH Terms
Gene Expression Regulation, Fungal
Genes, Fungal
112. Marx  F, Haas  H, Hofer  S, Stöffler  G, Redl  B,     ( 1995 )

Sequence and structure of Penicillium chrysogenum phoG, homologous to an acid phosphatase-encoding gene of Aspergillus nidulans.

Gene 160 (1)
PMID : 7628710  :   DOI  :   10.1016/0378-1119(95)00245-2    
Abstract >>
A Penicillium chrysogenum (Pc) gene (phoG), homologous to an Aspergillus nidulans (An) gene which confers phosphate-non-repressible acid phosphatase (APase) activity, has been cloned and sequenced. The 2.9-kb genomic sequence corresponds to two ORFs of 149 and 1630 bp encoding a protein of 593 amino acids (aa). As verified by cDNA sequencing, the coding region is interrupted by an 85-bp intron. The deduced aa sequence of phoG reveals 61% aa identity to the translated long ORF of the An APase-encoding gene. Northern blot analysis indicated a 2.3-kb transcript in approximately equivalent amount in mycelia grown under different phosphate concentrations.
KeywordMeSH Terms
Genes, Fungal
113. Gouka  RJ, van Hartingsveldt  W, Bovenberg  RA, van Zeijl  CM, van den Hondel  CA, van Gorcom  RF,     ( 1993 )

Development of a new transformant selection system for Penicillium chrysogenum: isolation and characterization of the P. chrysogenum acetyl-coenzyme A synthetase gene (facA) and its use as a homologous selection marker.

Applied microbiology and biotechnology 38 (4)
PMID : 7765289  :  
Abstract >>
A new transformation system for the filamentous fungus Penicillium chrysogenum is described, based on the use of the homologous acetyl-coenzyme A synthetase (facA) gene as a selection marker. Acetate-non-utilizing (Fac-) strains of P. chrysogenum were obtained by positive selection for spontaneous resistance to fluoroacetate. Among these fac mutants putative facA strains were selected for a loss of acetyl-coenzyme A (CoA) synthetase activity. The facA gene, coding for the enzyme acetyl-CoA synthetase, was isolated from a P. chrysogenum genomic library using synthetic oligonucleotides derived from conserved regions from the corresponding genes of Aspergillus nidulans and Neurospora crassa. Vector pPC2-3, comprising a genomic 6.5 kb PstI fragment, was able to complement P. chrysogenum facA strains with frequencies up to 27 transformants.micrograms-1 DNA. Direct selection of transformants was accomplished using acetate and low amounts (0.001%) of glucose as carbon sources. About 50% of the transformants arose by integration of pPC2-3 DNA at the homologous facA locus and 50% by integration elsewhere in the genome. Determination of the nucleotide sequence of part of the cloned fragment showed the presence of an open reading frame of 2007 nucleotides, interrupted by five putative introns. Comparison of the nucleotide and the amino acid sequence of the facA gene of P. chrysogenum with the facA gene of A. nidulans reveals similarities of 80% and 89%, respectively. The putative introns present in the P. chrysogenum facA gene appear at identical positions as those in the A. nidulans facA gene, but show no significant sequence similarity.
KeywordMeSH Terms
Transformation, Genetic
114. Naruse  A, Yamamoto  H, Sekiguchi  J,     ( 1993 )

Nucleotide sequence of the large mitochondrial rRNA gene of Penicillium chrysogenum.

Biochimica et biophysica acta 1172 (3)
PMID : 7680578  :   DOI  :   10.1016/0167-4781(93)90231-2    
Abstract >>
The nucleotide sequence of a large rRNA (1-rRNA) gene and its flanking regions in the cloned fragments of mitochondrial (mt) DNA from Penicillium chrysogenum NRRL1951 (Sekiguchi J., Ohsaki, T., Yamamoto, H., Koichi, K. and Shida, T. (1990) J. Gen. Microbiol. 136, 535-543) was determined and compared with those in Aspergillus nidulans and Neurospora crassa mitochondrial DNAs. The P. chrysogenum mt 1-rRNA gene has a 1678 bp intron which intervenes between a 2835 bp 5' exon and a 581 bp 3' exon, and extensive homology exists between overall sequences of mt 1-rRNA genes of P. chrysogenum and A. nidulans. The P. chrysogenum intron contains a large open reading frame which encodes a polypeptide comprising of 399 amino acids. The intron sequence also suggests that the intron belongs to a self-splicing group IA.
KeywordMeSH Terms
Genes, Fungal
Mitochondria
115. Alves  IMS, Gonçalves  VN, Oliveira  FS, Schaefer  CEGR, Rosa  CA, Rosa  LH,     ( 2019 )

The diversity, distribution, and pathogenic potential of cultivable fungi present in rocks from the South Shetlands archipelago, Maritime Antarctica.

Extremophiles : life under extreme conditions 23 (3)
PMID : 30852677  :   DOI  :   10.1007/s00792-019-01086-8    
Abstract >>
We studied the molecular taxonomy and diversity of cultivable rock fungi from Antarctic islands. From 50 rock samples, 386 fungal isolates were obtained and identified as 33 taxa of 20 genera. The genera Cladophialophora, Cladosporium, Cyphellophora, Eichleriella, Paracladophialophora, and Penicillium displayed the highest densities. Ecological diversity indices showed that the fungal assemblages are diverse and rich with low dominance. The genera Cladophialophora, Cladosporium, and Penicillium showed a broad distribution from rocks of the various islands. One hundred and fifty-nine fungi, grown at 37 �XC, were identified as Penicillium chrysogenum, Fusarium sp., and Rhodotorula mucilaginosa. One hundred and three fungi displayed haemolytic activity, 81 produced proteinase, 9 produced phospholipase, and 25 presented dimorphism and a spore diameter ? 4 ?m. The Antarctic Peninsula region appears to be under the effects of global climate changes, which may expose and accelerate the rock's weathering processes, and expose and release cryptic fungi and other microbes, especially those with innate pathogenic potential, previously arrested in rocks. Consequently, these rocks and their particles may represent a vehicle for the dispersal of microbial propagules, including those able to spread pathogens, along, across, and out of Antarctica.
KeywordMeSH Terms
Antarctica
Extremophile
Rocks
Taxonomy
Biodiversity
Fungi
Phylogeny
Soil Microbiology
116. Borman  AM, Fraser  M, Szekely  A, Johnson  EM,     ( 2019 )

Rapid and robust identification of clinical isolates of Talaromyces marneffei based on MALDI-TOF mass spectrometry or dimorphism in Galleria mellonella.

Medical mycology N/A (N/A)
PMID : 30649411  :   DOI  :   10.1093/mmy/myy162    
Abstract >>
Talaromyces marneffei is a thermally dimorphic fungal pathogen that causes serious infections particularly in patients with human immunodeficiency virus (HIV). Although the mould form typically produces a characteristic red-diffusing pigment, and conidia from penicillate heads, several nonpathogenic Talaromyces/Penicillium species are morphologically and phenotypically similar. While those other species do not exhibit thermal dimorphism, conversion of T. marneffei to the distinctive fission yeast form in vitro is arduous and frequently incomplete. Here we show that T. marneffei can be rapidly and unambiguously discriminated from related nonpathogenic Talaromyces/Penicillium spp., either by matrix-assisted laser desorption ionisation time-of-flight (MALDI-TOF) mass spectrometry or conversion to fission yeast after introduction into Galleria mellonella. Conversion of T. marneffei conidia to the fission yeast form in G. mellonella larvae occurred as early as 24 h post inoculation at 37oC. Identification by MALDI-TOF was possible after supplementation of the commercial Bruker database with in-house mass spectral profiles created from either the yeast or mycelial phase of T. marneffei. In addition, we show that in-house generated mass spectral profiles could be successfully used to identify T. marneffei with a recently published on-line MALDI-TOF database, circumventing the need to create extensive in-house additional databases for rarely encountered fungal pathogens.
KeywordMeSH Terms
117. Ullah  SF, Souza  AA, Hamann  PRV, Ticona  ARP, Oliveira  GM, Barbosa  JARG, Freitas  SM, Noronha  EF,     ( 2019 )

Structural and functional characterisation of xylanase purified from Penicillium chrysogenum produced in response to raw agricultural waste.

International journal of biological macromolecules 127 (N/A)
PMID : 30654038  :   DOI  :   10.1016/j.ijbiomac.2019.01.057    
Abstract >>
Commercial interest in plant cell wall degrading enzymes (PCWDE) is motivated by their potential for energy or bioproduct generation that reduced dependency on non-renewable (fossil-derived) feedstock. Therefore, underlying work analysed the Penicillium chrysogenum isolate for PCWDE production by employing different biomass as a carbon source. Among the produced enzymes, three xylanase isoforms were observed in the culture filtrate containing sugarcane bagasse. Xylanase (PcX1) presenting 35 kDa molecular mass was purified by gel filtration and anion exchange chromatography. Unfolding was probed and analysed using fluorescence, circular dichroism and enzyme assay methods. Secondary structure contents were estimated by circular dichroism 45% �\-helix and 10% �]-sheet, consistent with the 3D structure predicted by homology. PcX1 optimally active at pH 5.0 and 30 �XC, presenting t1/2 19 h at 30 �XC and 6 h at 40 �XC. Thermodynamic parameters/melting temperature 51.4 �XC confirmed the PcX1 stability at pH 5.0. PcX1 have a higher affinity for oat spelt xylan, KM 1.2 mg�PmL-1, in comparison to birchwood xylan KM 29.86 mg�PmL-1, activity was inhibited by Cu+2 and activated by Zn+2. PcX1 exhibited significant tolerance for vanillin, trans-ferulic acid, �l-coumaric acid, syringaldehyde and 4-hydroxybenzoic acid, activity slightly inhibited (17%) by gallic and tannic acid.
KeywordMeSH Terms
Biotechnological products
Lignocellulosic biomass
Phenolic compounds
Protein structural stability
Secondary structure
Xylanase
118. Carr  LG, Skatrud  PL, Scheetz  ME, Queener  SW, Ingolia  TD,     ( 1986 )

Cloning and expression of the isopenicillin N synthetase gene from Penicillium chrysogenum.

Gene 48 (2��3��)
PMID : 3104145  :   DOI  :   10.1016/0378-1119(86)90084-3    
Abstract >>
The isopenicillin N synthetase (IPS) gene from Penicillium chrysogenum was isolated from a recombinant bacteriophage lambda library using the Cephalosporium acremonium IPS (cIPS) gene as a heterologous hybridization probe. The protein coding region of the P. chrysogenum IPS (pIPS) gene was about 74% homologous to the cIPS gene, and the predicted amino acid sequences of the encoded proteins were about 73% homologous. Escherichia coli cells with the pIPS gene contained IPS activity whereas untransformed cells were completely devoid of this enzymatic activity. The transformed cells were also shown to contain an abundant protein accounting for about 10% of total cell protein which reacted strongly with anti-cIPS antiserum.
KeywordMeSH Terms
Genes, Fungal
Oxidoreductases
119. van Solingen  P, Muurling  H, Koekman  B, van den Berg  J,     ( 1988 )

Sequence of the Penicillium chrysogenum phosphoglycerate kinase gene.

Nucleic acids research 16 (24)
PMID : 3145495  :   DOI  :   10.1093/nar/16.24.11823     PMC  :   PMC339123    
Abstract >>
N/A
KeywordMeSH Terms
Genes, Fungal
120. Peñalva  MA, Sánchez  F,     ( 1987 )

The complete nucleotide sequence of the trpC gene from Penicillium chrysogenum.

Nucleic acids research 15 (4)
PMID : 3103104  :   DOI  :   10.1093/nar/15.4.1874     PMC  :   PMC340590    
Abstract >>
N/A
KeywordMeSH Terms
Genes
Genes, Fungal
121. Shlyapnikov  SV, Bezborodova  SI, Kulikov  VA, Yakovlev  GI,     ( 1986 )

Express analysis of protein amino acid sequences. Primary structure of Penicillium chrysogenum 152A guanyl-specific ribonuclease.

FEBS letters 196 (1)
PMID : 3080339  :   DOI  :   10.1016/0014-5793(86)80208-3    
Abstract >>
The complete amino acid sequence of Penicillium chrysogenum 152A guanyl-specific RNase has been established using automated Edman degradation of two non-fractionated peptide mixtures produced by tryptic and staphylococcal protease digests of the protein. The RNase contains 102 amino acid residues: His2, Arg3, Asp7, Asn8, Thr5, Ser11, Glu4, Gln2, Pro4, Gly11, Ala13, Cys4, Val8, Ile3, Leu3, Tyr9, Phe5 (Mr 10 747).
KeywordMeSH Terms
Metalloendopeptidases
122. Cantoral  JM, Barredo  JL, Alvarez  E, Díez  B, Martín  JF,     ( 1988 )

Nucleotide sequence of the Penicillium chrysogenum pyrG (orotidine-5'-phosphate decarboxylase) gene.

Nucleic acids research 16 (16)
PMID : 3138658  :   DOI  :   10.1093/nar/16.16.8177     PMC  :   PMC338522    
Abstract >>
N/A
KeywordMeSH Terms
Genes, Fungal
123. Kunishige  Y, Iwai  M, Nakazawa  M, Ueda  M, Tada  T, Nishimura  S, Sakamoto  T,     ( 2018 )

Crystal structure of exo-rhamnogalacturonan lyase from Penicillium chrysogenum as a member of polysaccharide lyase family 26.

FEBS letters 592 (8)
PMID : 29574769  :   DOI  :   10.1002/1873-3468.13034    
Abstract >>
Exo-rhamnogalacturonan lyase from Penicillium chrysogenum 31B (PcRGLX) was recently classified as a member of polysaccharide lyase (PL) family 26 along with hypothetical proteins derived from various organisms. In this study, we determined the crystal structure of PcRGLX as the first structure of a member of this family. Based on the substrate-binding orientation and substrate specificity, PcRGLX is an exo-type PL that cleaves rhamnogalacturonan from the reducing end. Analysis of PcRGLX-complex structures with reaction products indicate that the active site possesses an L-shaped cleft that can accommodate galactosyl side chains, suggesting side-chain-bypassing activity in PcRGLX. Furthermore, we determined the residues critical for catalysis by analyzing the enzyme activities of inactive variants.
KeywordMeSH Terms
crystal structure
exo-rhamnogalacturonan lyase
polysaccharide lyase family 26
124. Gomes  ECQ, Godinho  VM, Silva  DAS, de Paula  MTR, Vitoreli  GA, Zani  CL, Alves  TMA, Junior  PAS, Murta  SMF, Barbosa  EC, Oliveira  JG, Oliveira  FS, Carvalho  CR, Ferreira  MC, Rosa  CA, Rosa  LH,     ( 2018 )

Cultivable fungi present in Antarctic soils: taxonomy, phylogeny, diversity, and bioprospecting of antiparasitic and herbicidal metabolites.

Extremophiles : life under extreme conditions 22 (3)
PMID : 29332141  :   DOI  :   10.1007/s00792-018-1003-1    
Abstract >>
Molecular biology techniques were used to identify 218 fungi from soil samples collected from four islands of Antarctica. These consisted of 22 taxa of 15 different genera belonging to the Zygomycota, Ascomycota, and Basidiomycota. Mortierella, Antarctomyces, Pseudogymnoascus, and Penicillium were the most frequently isolated genera and Penicillium tardochrysogenum, Penicillium verrucosus, Goffeauzyma gilvescens, and Mortierella sp. 2 the most abundant taxa. All fungal isolates were cultivated using solid-state fermentation to obtain their crude extracts. Pseudogymnoascus destructans, Mortierella parvispora, and Penicillium chrysogenum displayed antiparasitic activities, whilst extracts of P. destructans, Mortierella amoeboidea, Mortierella sp. 3, and P. tardochrysogenum showed herbicidal activities. Reported as pathogenic for bats, different isolates of P. destructans exhibited trypanocidal activities and herbicidal activity, and may be a source of bioactive molecules to be considered for chemotherapy against neglected tropical diseases. The abundant presence of P. destructans in soils of the four islands gives evidence supporting that soils in the Antarctic Peninsula constitute a natural source of strains of this genus, including some P. destructans strains that are phylogenetically close to those that infect bats in North America and Europe/Palearctic Asia.
KeywordMeSH Terms
Antarctica
Fungi
Mortierella
Neglected tropical diseases
Pseudogymnoascus
Microbiota
Phylogeny
Soil Microbiology
125. Coronado-Ruiz  C, Avendaño  R, Escudero-Leyva  E, Conejo-Barboza  G, Chaverri  P, Chavarría  M,     ( 2018 )

Two new cellulolytic fungal species isolated from a 19th-century art collection.

Scientific reports 8 (1)
PMID : 29748544  :   DOI  :   10.1038/s41598-018-24934-7     PMC  :   PMC5945893    
Abstract >>
The archive of the Universidad de Costa Rica maintains a nineteenth-century French collection of drawings and lithographs in which the biodeterioration by fungi is rampant. Because of nutritional conditions in which these fungi grew, we suspected that they possessed an ability to degrade cellulose. In this work our goal was to isolate and identify the fungal species responsible for the biodegradation of a nineteenth-century art collection and determine their cellulolytic activity. Fungi were isolated using potato-dextrose-agar (PDA) and water-agar with carboxymethyl cellulose (CMC). The identification of the fungi was assessed through DNA sequencing (nrDNA ITS and �\-actin regions) complemented with morphological analyses. Assays for cellulolytic activity were conducted with Gram's iodine as dye. Nineteen isolates were obtained, of which seventeen were identified through DNA sequencing to species level, belonging mainly to genera Arthrinium, Aspergillus, Chaetomium, Cladosporium, Colletotrichum, Penicillium and Trichoderma. For two samples that could not be identified through their ITS and �\-actin sequences, a morphological analysis was conducted; they were identified as new species, named Periconia epilithographicola sp. nov. and Coniochaeta cipronana sp. nov. Qualitative tests showed that the fungal collection presents important cellulolytic activity.
KeywordMeSH Terms
126. Corral  P, Esposito  FP, Tedesco  P, Falco  A, Tortorella  E, Tartaglione  L, Festa  C, D'Auria  MV, Gnavi  G, Varese  GC, de Pascale  D,     ( 2018 )

Identification of a Sorbicillinoid-Producing Aspergillus Strain with Antimicrobial Activity Against Staphylococcus aureus: a New Polyextremophilic Marine Fungus from Barents Sea.

Marine biotechnology (New York, N.Y.) 20 (4)
PMID : 29651633  :   DOI  :   10.1007/s10126-018-9821-9    
Abstract >>
The exploration of poorly studied areas of Earth can highly increase the possibility to discover novel bioactive compounds. In this study, the cultivable fraction of fungi and bacteria from Barents Sea sediments has been studied to mine new bioactive molecules with antibacterial activity against a panel of human pathogens. We isolated diverse strains of psychrophilic and halophilic bacteria and fungi from a collection of nine samples from sea sediment. Following a full bioassay-guided approach, we isolated a new promising polyextremophilic marine fungus strain 8Na, identified as Aspergillus protuberus MUT 3638, possessing the potential to produce antimicrobial agents. This fungus, isolated from cold seawater, was able to grow in a wide range of salinity, pH and temperatures. The growth conditions were optimised and scaled to fermentation, and its produced extract was subjected to chemical analysis. The active component was identified as bisvertinolone, a member of sorbicillonoid family that was found to display significant activity against Staphylococcus aureus with a minimum inhibitory concentration (MIC) of 30 �gg/mL.
KeywordMeSH Terms
Antimicrobial activity
Aspergillus protuberus
Bisvertinolone
MDR
Marine fungi
Sediments
Antimicrobial activity
Aspergillus protuberus
Bisvertinolone
MDR
Marine fungi
Sediments
127.     ( 2012 )

New penicillin-producing Penicillium species and an overview of section Chrysogena.

Persoonia 29 (N/A)
PMID : 23606767  :   DOI  :   10.3767/003158512X660571     PMC  :   PMC3589797    
Abstract >>
Species classified in Penicillium sect. Chrysogena are primary soil-borne and the most well-known members are P. chrysogenum and P. nalgiovense. Penicillium chrysogenum has received much attention because of its role in the production on penicillin and as a contaminant of indoor environments and various food and feedstuffs. Another biotechnologically important species is P. nalgiovense, which is used as a fungal starter culture for the production of fermented meat products. Previous taxonomic studies often had conflicting species circumscriptions. Here, we present a multigene analysis, combined with phenotypic characters and extrolite data, demonstrating that sect. Chrysogena consists of 18 species. Six of these are newly described here (P. allii-sativi, P. desertorum, P. goetzii, P. halotolerans, P. tardochrysogenum, P. vanluykii) and P. lanoscoeruleum was found to be an older name for P. aethiopicum. Each species produces a unique extrolite profile. The species share phenotypic characters, such as good growth on CYA supplemented with 5 % NaCl, ter- or quarterverticillate branched conidiophores and short, ampulliform phialides (< 9 �gm). Conidial colours, production of ascomata and ascospores, shape and ornamentation of conidia and growth rates on other agar media are valuable for species identification. Eight species (P. allii-sativi, P. chrysogenum, P. dipodomyis, P. flavigenum, P. nalgiovense, P. rubens, P. tardochrysogenum and P. vanluykii) produce penicillin in culture.
KeywordMeSH Terms
Fleming
P. chrysogenum
P. rubens
phylogeny
taxonomy
Fleming
P. chrysogenum
P. rubens
phylogeny
taxonomy
Fleming
P. chrysogenum
P. rubens
phylogeny
taxonomy
128. Stielow  JB, Lévesque  CA, Seifert  KA, Meyer  W, Iriny  L, Smits  D, Renfurm  R, Verkley  GJ, Groenewald  M, Chaduli  D, Lomascolo  A, Welti  S, Lesage-Meessen  L, Favel  A, Al-Hatmi  AM, Damm  U, Yilmaz  N, Houbraken  J, Lombard  L, Quaedvlieg  W, Binder  M, Vaas  LA, Vu  D, Yurkov  A, Begerow  D, Roehl  O, Guerreiro  M, Fonseca  A, Samerpitak  K, van Diepeningen  AD, Dolatabadi  S, Moreno  LF, Casaregola  S, Mallet  S, Jacques  N, Roscini  L, Egidi  E, Bizet  C, Garcia-Hermoso  D, Martín  MP, Deng  S, Groenewald  JZ, Boekhout  T, de Beer  ZW, Barnes  I, Duong  TA, Wingfield  MJ, de Hoog  GS, Crous  PW, Lewis  CT, Hambleton  S, Moussa  TA, Al-Zahrani  HS, Almaghrabi  OA, Louis-Seize  G, Assabgui  R, McCormick  W, Omer  G, Dukik  K, Cardinali  G, Eberhardt  U, de Vries  M, Robert  V,     ( 2015 )

One fungus, which genes? Development and assessment of universal primers for potential secondary fungal DNA barcodes.

Persoonia 35 (N/A)
PMID : 26823635  :   DOI  :   10.3767/003158515X689135     PMC  :   PMC4713107    
Abstract >>
KeywordMeSH Terms
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
DNA barcoding
ITS supplement
molecular taxonomy
phylogeny
species identification
universal primers
129. Crutcher  FK, Puckhaber  LS, Stipanovic  RD, Bell  AA, Nichols  RL, Lawrence  KS, Liu  J,     ( 2017 )

Microbial Resistance Mechanisms to the Antibiotic and Phytotoxin Fusaric Acid.

Journal of chemical ecology 43 (10)
PMID : 28986689  :   DOI  :   10.1007/s10886-017-0889-x    
Abstract >>
Fusaric acid (FA) produced by Fusarium oxysporum plays an important role in disease development in plants, including cotton. This non-specific toxin also has antibiotic effects on microorganisms. Thus, one expects a potential pool of diverse detoxification mechanisms of FA in nature. Bacteria and fungi from soils infested with Fusarium and from laboratory sources were evaluated for their ability to grow in the presence of FA and to alter the structure of FA into less toxic compounds. None of the bacterial strains were able to chemically modify FA. Highly FA-resistant strains were found only in Gram-negative bacteria, mainly in the genus of Pseudomonas. The FA resistance of the Gram-negative bacteria was positively correlated with the number of predicted genes for FA efflux pumps present in the genome. Phylogenetic analysis of predicted FA resistance proteins (FUSC, an inner membrane transporter component of the efflux pump) revealed that FUSC proteins having high sequence identities with the functionally characterized FA resistance protein FusC or Fdt might be the major contributors of FA resistance. In contrast, most fungi converted FA to less toxic compounds regardless of the level of FA resistance they exhibited. Five derivatives were detected, and the detoxification of FA involved either oxidative reactions on the butyl side chain or reductive reactions on the carboxylic acid group. The production of these metabolites from widely different phyla indicates that resistance to FA by altering its structure is highly conserved. A few FA resistant saprophytic or biocontrol strains of fungi were incapable of altering FA, indicating a possible involvement of efflux transporters. Deployment of both efflux and derivatization mechanisms may be a common feature of fungal FA resistance.
KeywordMeSH Terms
Antibiotic resistance
Bacteria
Detoxification
Efflux pump
FUSC
Fdt
Fungi
Fusaric acid
Resistance
Soil microbiome
Antibiotic resistance
Bacteria
Detoxification
Efflux pump
FUSC
Fdt
Fungi
Fusaric acid
Resistance
Soil microbiome
Antibiotic resistance
Bacteria
Detoxification
Efflux pump
FUSC
Fdt
Fungi
Fusaric acid
Resistance
Soil microbiome
Antibiotic resistance
Bacteria
Detoxification
Efflux pump
FUSC
Fdt
Fungi
Fusaric acid
Resistance
Soil microbiome
Soil Microbiology
130. Gonçalves  VN, Vitoreli  GA, de Menezes  GCA, Mendes  CRB, Secchi  ER, Rosa  CA, Rosa  LH,     ( 2017 )

Taxonomy, phylogeny and ecology of cultivable fungi present in seawater gradients across the Northern Antarctica Peninsula.

Extremophiles : life under extreme conditions 21 (6)
PMID : 28856503  :   DOI  :   10.1007/s00792-017-0959-6    
Abstract >>
Thirty-six seawater samples collected at different depths of the Gerlache and Bransfield Straits in the Northern Antarctic Peninsula were analyzed, and the average of the total fungal counts ranged from 0.3 to >300 colony forming units per liter (CFU/L) in density. The fungal were purified and identified as 15 taxa belonged to the genera Acremonium, Aspergillus, Cladosporium, Cystobasidium, Exophiala, Glaciozyma, Graphium, Lecanicillium, Metschnikowia, Penicillium, Purpureocillium and Simplicillium. Penicillium chrysogenum, Cladosporium sphaerospermum, and Graphium rubrum were found at high densities in at least two different sites and depths. Our results show at the first time that in the seawater of Antarctic Ocean occur diverse fungal assemblages despite extreme conditions, which suggests the presence of a complex aquatic fungi food web, including species reported as barophiles, symbionts, weak and strong saprobes, parasites and pathogens, as well as those found in the polluted environments of the world. Additionally, some taxa were found in different sites, suggesting that the underwater current might contribute to fungal (and microbial) dispersal across the Antarctic Ocean, and nearby areas such as South America and Australia.
KeywordMeSH Terms
Antarctic Peninsula
Extremophile
Fungi
Seawater
Taxonomy
Antarctic Peninsula
Extremophile
Fungi
Seawater
Taxonomy
Antarctic Peninsula
Extremophile
Fungi
Seawater
Taxonomy
Extreme Cold
Mycobiome
Phylogeny
131.     ( 2012 )

Amplification of an MFS transporter encoding gene penT significantly stimulates penicillin production and enhances the sensitivity of Penicillium chrysogenum to phenylacetic acid.

Journal of genetics and genomics = Yi chuan xue bao 39 (11)
PMID : 23177147  :   DOI  :   10.1016/j.jgg.2012.08.004    
Abstract >>
Penicillin is historically important as the first discovered drug against bacterial infections in human. Although the penicillin biosynthetic pathway and regulatory mechanism have been well studied in Penicillium chrysogenum, the compartmentation and molecular transport of penicillin or its precursors are still poorly understood. In search of the genomic database, more than 830 open reading frames (ORFs) were found to encode transmembrane proteins of P. chrysogenum. In order to investigate their roles on penicillin production, one of them (penT) was selected and cloned. The deduced protein of penT belongs to the major facilitator superfamily (MFS) and contains 12 transmembrane spanning domains (TMS). During fermentation, the transcription of penT was greatly induced by penicillin precursors phenylacetic acid (PAA) and phenoxyacetic acid (POA). Knock-down of penT resulted in significant decrease of penicillin production, while over-expression of penT under the promoter of trpC enhanced the penicillin production. Introduction of an additional penT in the wild-type strain of P. chrysogenum doubled the penicillin production and enhanced the sensitivity of P. chrysogenum to the penicillin precursors PAA or POA. These results indicate that penT stimulates penicillin production probably through enhancing the translocation of penicillin precursors across fungal cellular membrane.
KeywordMeSH Terms
132.     ( 1998 )

Characterization of the lys2 gene of Penicillium chrysogenum encoding alpha-aminoadipic acid reductase.

Molecular & general genetics : MGG 259 (5)
PMID : 9790587  :   DOI  :   10.1007/s004380050847    
Abstract >>
A DNA fragment containing a gene homologous to LYS2 gene of Saccharomyces cerevisiae was cloned from a genomic DNA library of Penicillium chrysogenum AS-P-78. It encodes a protein of 1409 amino acids (Mr 154859) with strong similarity to the S. cerevisiae (49.9% identity) Schizosaccharomyces pombe (51.3% identity) and Candida albicans (48.12% identity) alpha-aminoadipate reductases and a lesser degree of identity to the amino acid-activating domains of the non-ribosomal peptide synthetases, including the alpha-aminoadipate-activating domain of the alpha-aminoadipyl-cysteinyl-valine synthetase of P. chrysogenum (12.4% identical amino acids). The lys2 gene contained one intron in the 5'-region and other in the 3'-region, as shown by comparing the nucleotide sequences of the cDNA and genomic DNA, and was transcribed as a 4.7-kb monocistronic mRNA. The lys2 gene was localized on chromosome III (7.5 Mb) in P. chrysogenum AS-P-78 and on chromosome IV (5.6 Mb) in strain P2, whereas the penicillin gene cluster is known to be located in chromosome I in both strains. The lys2-encoded protein is a member of the aminoacyladenylate-forming enzyme family with a reductase domain in its C-terminal region.
KeywordMeSH Terms
Repressor Proteins
Saccharomyces cerevisiae Proteins
133.     ( 1998 )

The manganese superoxide dismutase from the penicillin producer Penicillium chrysogenum.

Current genetics 33 (6)
PMID : 9644201  :  
Abstract >>
The antioxidant enzyme superoxide dismutase has been studied in order to define mechanisms for the influence of oxygen on penicillin production. Manganese-containing SOD activity was purified from penicillin-producing cultures of the filamentous fungus Penicillium chrysogenum and reverse genetics was used to identify full-length cDNA and genomic clones. Sequence analysis revealed a 630-bp ORF containing three exons and two introns with fungal consensus splice-site junctions. The deduced amino-acid sequence (210 amino acids; 23.13 kDa) includes conserved residues required for enzymatic activity and metal binding, and shares significant similarity with Mn- and Fe-containing superoxide dismutases. The sod gene is present as a single copy in the genome of different P. chrysogenum strains and its expression level is not correlated with penicillin-G productivity.
KeywordMeSH Terms

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