title: The role of the nonconserved residues at position 167 of class A beta-lactamases in susceptibility to mechanism-based inhibitors.
abstract: Differences in specificities between the class A beta-lactamases for both substrate and inhibitors are known. The role of the nonconserved amino acid residue at position 167 of the class A enzyme, which forms a cis bond with the catalytically essential Glu-166 residue, in both the hydrolysis of beta-lactam substrates and inactivation by mechanism-based inhibitors, was investigated. Site-directed mutagenesis was performed on the penPC gene encoding the Bacillus cereus 569/H beta-lactamase I to replace thr-167 with the corresponding Staphylococcus aureus PC1 residue Ile. Kinetic data obtained from the purified Thr-167-Ile B. cereus 569/H beta-lactamase was compared to that obtained from the wild-type B. cereus and S. aureus enzymes and indicated that the replacement had little effect on the Michaelis parameters for the hydrolysis of S- and A-type penicillins. However, the Thr-167-Ile enzymes became more S. aureus PC1-like in its response to the mechanism-based inhibitors clavulanic acid and 6-beta-(trifluoromethane sulfonyl)amidopenicillanic acid sulfone. A model for the role of this nonconserved residue at position 167 in the mechanism of inactivation by mechanism-based inhibitors is proposed.
authors.affiliation: Department of Chemistry, University of Waterloo, Ontario, Canada.
authors.forename: F
authors.initials: F
authors.lastname: Guo
authors.forename: G I
authors.initials: GI
authors.lastname: Dmitrienko
authors.forename: A J
authors.initials: AJ
authors.lastname: Clarke
authors.forename: T
authors.initials: T
authors.lastname: Viswanatha
mesh_terms: D000595:Amino Acid Sequence; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005798:Genes, Bacterial / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010406:Penicillins / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D013379:Substrate Specificity; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases / Q000096:biosynthesis / Q000235:genetics*
chemical_list: D004791:Enzyme Inhibitors; D010406:Penicillins; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases