Biogenesis of the yeast vacuole (lysosome). Active site mutation in the vacuolar aspartate proteinase yscA blocks maturation of vacuolar proteinases.
The activation process of vacuolar (lysosomal) proteinases in the yeast Saccharomyces cerevisiae is initiated by the PRA1 (PEP4) gene product, proteinase yscA. To elucidate the activation process of proteinase yscA the catalytically active amino acid Asp294 of the enzyme was exchanged with Ala294 using site directed mutagenesis. The resulting proteinase yscA-Ala294 showed no proteolytic activity against the substrate hemoglobin. The mutant protein did not undergo processing in vivo. This phenomenon is in good agreement with the hypothesis that maturation of proteinase yscA is due to self-processing of pro-proteinase yscA. Furthermore, proteinase yscA-Ala294 is not able to convert the zymogens pro-proteinase yscB and pro-carboxypeptidase yscY to the mature enzymes.
Rupp
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Institut für Biochemie, Universität Stuttgart, Germany.
Hirsch
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Wolf
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DH
D000595:Amino Acid Sequence; D016282:Aspartic Acid Endopeptidases / Q000096:biosynthesis* / Q000235:genetics; D001665:Binding Sites; D002268:Carboxypeptidases / Q000096:biosynthesis; D002384:Catalysis; D043402:Cathepsin A; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D008247:Lysosomes / Q000201:enzymology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011499:Protein Processing, Post-Translational; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D012697:Serine Endopeptidases / Q000096:biosynthesis; D014617:Vacuoles / Q000201:enzymology*
D029701:Saccharomyces cerevisiae Proteins; D002268:Carboxypeptidases; D043402:Cathepsin A; C499283:PRC1 protein, S cerevisiae; C022633:serine carboxypeptidase; D012697:Serine Endopeptidases; C023302:yeast proteinase B; C022325:aspartic proteinase A; C499267:PEP4 protein, S cerevisiae; D016282:Aspartic Acid Endopeptidases