title: Purification and characterization of serine carboxypeptidases from Absidia zychae.
abstract: Among 29 strains of zygomycetes screened for serine carboxypeptidases, Absidia zychae NRIC 1199 showed the highest enzyme production. Two serine carboxypeptidases, CPZ-1 and CPZ-2, were purified to a homogeneous state from an extract of koji culture of A. zychae NRIC 1199. Purified CPZ-1 and CPZ-2 showed similar properties except the isoelectric point (pI); The pI of CPZ-1 and CPZ-2 were 4.50 and 4.65, respectively. The molecular weights of the CPZ-1 and CPZ-2 were 48,000 by SDS-PAGE and gel filtration. Among the proteinase inhibitors tested, phenylmethylsulfonyl fluoride and monoiodoacetic acid strongly inhibited the enzyme activity. The optimum pHs of CPZ-1 and CPZ-2 were 4.2 towards Z-Glu-Tyr. It is shown that the substrate specificities of CPZ-1 and CPZ-2 were dependent on the presence of bulky amino acid residues in the penultimate position (P1) for the small Z-peptides. However, in spite of the presence of Gly, Asp, Arg, or Pro in the P1 position, oligopeptides were hydrolyzed rapidly. CPZ-1 and CPZ-2 had not only carboxypeptidase but also carboxyamidase and amidase activities, and acted preferentially as a carboxyamidase for C-terminal amidated peptides. The hydrophobicity of P2 and P3 positions and the bulkiness of P1 and P'1 positions of the substrate may be important for carboxyamidase and amidase activities.
authors.affiliation: Laboratory of Molecular Biology and Microbial Chemistry, Tokyo University of Agriculture and Technology, Japan.
authors.forename: B R
authors.initials: BR
authors.lastname: Lee
authors.forename: M
authors.initials: M
authors.lastname: Takeuchi
authors.forename: Y
authors.initials: Y
authors.lastname: Kobayashi
mesh_terms: D000595:Amino Acid Sequence; D002268:Carboxypeptidases / Q000032:analysis / Q000302:isolation & purification*; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D005656:Fungal Proteins / Q000032:analysis; D005658:Fungi / Q000201:enzymology*; D007700:Kinetics; D008969:Molecular Sequence Data; D013379:Substrate Specificity
chemical_list: D005656:Fungal Proteins; D002268:Carboxypeptidases; C022633:serine carboxypeptidase; C499287:carboxypeptidase Z