Cloning and characterization of penicillin V acylase from Streptomyces mobaraensis.
We report on the molecular cloning and characterization of penicillin V acylase (PVA) from an actinomycete, Streptomyces mobaraensis (Sm-PVA), which was originally isolated as an acylase that efficiently hydrolyzes the amide bond of various N-fatty-acyl-l-amino acids and N-fatty-acyl-peptides as well as capsaicin (8-methyl-N-vanillyl-6-nonenamide). In addition, the purified Sm-PVA hydrolyzed penicillin V with the highest activity (k(cat)) among the PVAs so far reported, penicillin G, and 2-nitro-5-phenoxyacetamide benzoic acid. The BLAST search revealed that the Sm-PVA precursor is composed of a polypeptide that is characteristic of enzymes belonging to the beta-lactam acylase family with four distinct segments; a signal sequence (43 amino acids), an alpha subunit (173 amino acids), a linker peptide (28 amino acids), and a beta subunit (570 amino acids). The mature, active Sm-PVA is a heterodimeric protein with alpha and beta subunits, in contrast to PVAs isolated from Bacillus sphaericus and B. subtilis, which have a homotetrameric structure. The amino acid sequence of Sm-PVA showed identities to PVA from S. lavendulae, N-acylhomoserine lactone-degrading acylase from Streptomyces sp., cyclic lipopeptide acylase from Streptomyces sp., and aculeacin A acylase from Actinoplanes utahensis with 68, 67, 67, and 41% identities, respectively.
Zhang
Demin
D
Department of Bioscience and Biotechnology, Faculty of Engineering, Okayama University, 3-1-1, Tsushima-Naka, Okayama 700-8530, Japan.
Koreishi
Mayuko
M
Imanaka
Hiroyuki
H
Imamura
Koreyoshi
K
Nakanishi
Kazuhiro
K
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D008969:Molecular Sequence Data; D010399:Penicillin Amidase / Q000235:genetics* / Q000302:isolation & purification; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics
D004269:DNA, Bacterial; D010399:Penicillin Amidase