Molecular characterization of a PDI-related gene prpA in Aspergillus niger var. awamori.
A gene (prpA) homologous to the protein disulfide isomerase gene was isolated from Aspergillus niger by Southern hybridization using the pdi1 gene isolated from Trichoderma reesei as a DNA probe. The corresponding cDNA of the prpA gene has also been isolated from an A. niger var. awamori cDNA library. The prpA gene does not belong to any currently recognized family of protein disulfide isomerases since it contains only a single conserved thioredoxin domain at the N-terminus of the protein. The C-terminal two-thirds of the protein has no homology to any known proteins in the database. The PRPA protein contains an ER retention signal (HDEL) at its C-terminal end suggesting that it is located in the ER. Southern hybridization at high stringency showed that it was present as a single copy in the genome. Northern hybridization indicated that the transcript level of the prpA gene was higher if the cells were secreting a heterologous protein, bovine prochymosin. However, over-expression of the prpA gene from a multicopy integrated vector had little effect on chymosin secretion. A strain containing a deletion of the prpA gene was viable. However, deletion of the prpA gene appeared to cause a reduction of bovine chymosin production.
Wang
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Genencor International, California, CA 94304-1013, USA. hwang@genencor.com
Ward
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D000595:Amino Acid Sequence; D000818:Animals; D001234:Aspergillus niger / Q000145:classification / Q000166:cytology / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D002417:Cattle; D002455:Cell Division; D012085:Chymosin / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D017353:Gene Deletion; D018628:Gene Dosage; D015870:Gene Expression; D005800:Genes, Fungal / Q000235:genetics* / Q000502:physiology; D014644:Genetic Variation / Q000235:genetics*; D005087:Glucan 1,4-alpha-Glucosidase / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry / Q000235:genetics; D019704:Protein Disulfide-Isomerases / Q000737:chemistry / Q000235:genetics*; D021382:Protein Sorting Signals / Q000737:chemistry / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013879:Thioredoxins / Q000737:chemistry / Q000235:genetics; D014242:Trichoderma / Q000235:genetics
D004792:Enzyme Precursors; D005656:Fungal Proteins; D009842:Oligopeptides; C404324:PRPA protein, Aspergillus niger; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C077480:histidyl-aspartyl-glutamyl-leucine; D013879:Thioredoxins; D005087:Glucan 1,4-alpha-Glucosidase; C032443:prorennin; D012085:Chymosin; D019704:Protein Disulfide-Isomerases