title: Structural and Mutagenic Analysis of Metallo-β-Lactamase IMP-18.
abstract: IMP-type metallo-β-lactamases (MBLs) are exogenous zinc metalloenzymes that hydrolyze a broad range of β-lactams, including carbapenems. Here we report the crystal structure of IMP-18, an MBL cloned from Pseudomonas aeruginosa, at 2.0-Å resolution. The overall structure of IMP-18 resembles that of IMP-1, with an αβ/βα "folded sandwich" configuration, but the loop that covers the active site has a distinct conformation. The relationship between IMP-18's loop conformation and its kinetic properties was investigated by replacing the amino acid residues that can affect the loop conformation (Lys44, Thr50, and Ile69) in IMP-18 with those occupying the corresponding positions in the well-described enzyme IMP-1. The replacement of Thr50 with Pro considerably modified IMP-18's kinetic properties, specifically those pertaining to meropenem, with the kcat/Km value increased by an order of magnitude. The results indicate that this is a key residue that defines the kinetic properties of IMP-type β-lactamases.
authors.affiliation: Department of Material and Biological Chemistry, Faculty of Science, Yamagata University, Yamagata, Japan.
authors.forename: Takamitsu
authors.initials: T
authors.lastname: Furuyama
authors.affiliation: Faculty of Applied Life Sciences, Niigata University of Pharmacy and Applied Life Sciences, Akiha-ku, Niigata, Japan.
authors.forename: Haruka
authors.initials: H
authors.lastname: Nonomura
authors.affiliation: Department of Microbiology and Infectious Diseases, Toho University School of Medicine, Ota-ku, Tokyo, Japan.
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Ishii
authors.affiliation: Department of Medical Microbiology and Immunology, Creighton University, Omaha, Nebraska, USA.
authors.forename: Nancy D
authors.initials: ND
authors.lastname: Hanson
authors.affiliation: Department of Material and Biological Chemistry, Faculty of Science, Yamagata University, Yamagata, Japan Faculty of Applied Life Sciences, Niigata University of Pharmacy and Applied Life Sciences, Akiha-ku, Niigata, Japan ibuka@nupals.ac.jp.
authors.forename: Akiko
authors.identifier: 0000-0002-5310-9216
authors.initials: A
authors.lastname: Shimizu-Ibuka
mesh_terms: D015780:Carbapenems / Q000494:pharmacology; D020134:Catalytic Domain / Q000235:genetics; D007700:Kinetics; D000077731:Meropenem; D009153:Mutagens; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000235:genetics*; D013845:Thienamycins / Q000494:pharmacology; D001618:beta-Lactamases / Q000235:genetics*; D047090:beta-Lactams / Q000494:pharmacology
chemical_list: D015780:Carbapenems; D009153:Mutagens; D013845:Thienamycins; D047090:beta-Lactams; C409201:beta-lactamase IMP-1; D001618:beta-Lactamases; C527279:beta-lactamase IMP-18, Pseudomonas aeruginosa; D000077731:Meropenem