Carboxypeptidase Y activity and maintenance is modulated by a large helical structure

• Published in: FEBS open bio Vol. 9; no. 7; pp. 1337 - 1343
• Main Authors: Makino, Mai, Sahara, Takehiko, Morita, Naoki, Ueno, Hiroshi
• Format: Journal Article
• Language: English
• Published: England John Wiley & Sons, Inc 01.07.2019; John Wiley and Sons Inc; Wiley
• Subjects: Amino Acid Sequence - genetics; Binding Sites - genetics; Carboxypeptidase; Carboxypeptidase C; Carboxypeptidases - metabolism; Catalysis; Catalytic Domain; Cathepsin A - genetics; Cathepsin A - metabolism; Cathepsin A - ultrastructure; Chemical bonds; Conformation; Deoxyribonucleic acid; disulfide bond; Disulfide bonds; DNA; Enzymes; Hydrogen bonds; Hydrolase; Mutagenesis; Protein Conformation; Protein Denaturation; Protein Folding; Proteins; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - metabolism; Serine; serine carboxypeptidase; Serine proteinase; Substrate specificity; Substrate Specificity - genetics; α/β hydrolase fold; United States > US; Japan; disulfide bond; protein folding; α/β hydrolase fold; serine carboxypeptidase
• ISSN: 2211-5463; 2211-5463
• DOI: 10.1002/2211-5463.12686

Yeast carboxypeptidase Y (CPY) is a serine protease with broad substrate specificity. Structurally, CPY belongs to the α/β hydrolase fold family and contains characteristic large helices, termed the V‐shape helix, above the active site cavity. Four intramolecular disulfide bonds are located in and around the V‐shape helix. In this study, mutant CPYs were constructed in which one of these disulfide bonds was disrupted. Mutants lacking the C193–C207 bond located at the beginning of the V‐shape helix aggregated easily, while mutants lacking the C262–C268 bond located at the end of the V‐shape helix displayed decreased hydrolytic activity. The results indicate that the V‐shape helix is involved in CPY catalysis and in maintenance of its conformation. Yeast carboxypeptidase Y (CPY) contains characteristic large helices, termed the V‐shape helix, above the active site cavity. When the C193–C207 bond located at the beginning of the V‐shape helix was disrupted, the mutant CPYs were aggregated easily. When the C262–C268 bond located at the end of the V‐shape helix was disrupted, the mutant CPYs displayed decreased hydrolytic activity.