title: Characterization and kinetic analysis of enzyme-substrate recognition by three recombinant lactococcal PepVs.
abstract: The dipeptidases (PepVs) from three typical lactococcal strains, Lactococcus lactis subsp. lactis (L9), L. lactis subsp. cremoris (L6) and L. lactis subsp. hordniae (hT) were cloned and characterized. The metal-binding, catalytic, and substrate-binding sites are highly conserved among of them. A computer-generated three-dimensional model suggested that the amino acid differences between these PepVs were mostly located away from the active center. L9 PepV does not hydrolyze dipeptides bearing Pro or D-amino acid at the C-terminal amino acid. Unlike PepV from Lactobacillus delbrueckii, L9 PepV does not cleave beta-Asp-His, and has little ability to cleave dipeptides containing a beta-alanine. In addition, L9 PepV has a much higher kcat for dipeptides with an N-terminal Ala but a significantly higher Km when the N-terminal amino acid is Gly. The substrate recognition profile of PepV is further discussed on the basis of the kinetic analysis and the structural model.
authors.affiliation: Biological Function Division, National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642, Japan.
authors.forename: Sumiko
authors.initials: S
authors.lastname: Mori
authors.forename: Maki
authors.initials: M
authors.lastname: Miyamoto
authors.forename: Satoshi
authors.initials: S
authors.lastname: Kaneko
authors.forename: Satoru
authors.initials: S
authors.lastname: Nirasawa
authors.forename: Shiro
authors.initials: S
authors.lastname: Komba
authors.forename: Takafumi
authors.initials: T
authors.lastname: Kasumi
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D001665:Binding Sites; D004150:Dipeptidases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D004151:Dipeptides / Q000378:metabolism*; D019075:Electrophoresis, Capillary; D007700:Kinetics; D013294:Lactococcus lactis / Q000201:enzymology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D004151:Dipeptides; D011994:Recombinant Proteins; D004150:Dipeptidases