title: Identification of the catalytic triad of family S46 exopeptidases, closely related to clan PA endopeptidases.
abstract: The exopeptidases of family S46 are exceptional, as the closest homologs of these enzymes are the endopeptidases of clan PA. The three-dimensional structure of S46 enzymes is unknown and only one of the catalytic residues, the serine, has been identified. The catalytic histidine and aspartate residues are not experimentally identified. Here we present phylogenetic and experimental data that identify all residues of the catalytic triad of S46 peptidase, dipeptidyl aminopeptidase BII (DAP BII) from Pseudoxanthomonas mexicana WO24. Phylogenetic comparison with the protein and S46 peptidases, revealed His-86, Ser-657, and five aspartate residues as possible catalytic residues. Mutation studies identified the catalytic triad of DAP BII as His-86, Asp-224, and Ser-657, while secondary structure analysis predicted an extended alpha-helical domain in between Asp-224 and Ser-657. This domain is unique for family S46 exopeptidases and its absence from the endopeptidases of clan PA might be key to their different hydrolysis activities.
authors.affiliation: The Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata 940-2188, Japan.
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Suzuki
authors.affiliation: School of Pharmacy, Iwate Medical University, 2-1-1 Nishitokuta, Yahaba, Iwate 028-3694, Japan.
authors.forename: Yasumitsu
authors.initials: Y
authors.lastname: Sakamoto
authors.affiliation: School of Pharmacy, Showa University, 1-5-8 Hatanodai, Shinagawa-ku, Tokyo 142-8555, Japan.
authors.forename: Nobutada
authors.initials: N
authors.lastname: Tanaka
authors.affiliation: The Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata 940-2188, Japan.
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Okada
authors.affiliation: The Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata 940-2188, Japan.
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Morikawa
authors.affiliation: The Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata 940-2188, Japan.
authors.forename: Wataru
authors.initials: W
authors.lastname: Ogasawara
mesh_terms: D000595:Amino Acid Sequence; D002384:Catalysis; D003001:Cloning, Molecular; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020563:Gammaproteobacteria / Q000145:classification / Q000201:enzymology* / Q000235:genetics; D015870:Gene Expression; D008969:Molecular Sequence Data; D005810:Multigene Family; D010802:Phylogeny; D017433:Protein Structure, Secondary; D016415:Sequence Alignment
chemical_list: D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; C059010:dipeptidyl aminopeptidase B