title: Properties of IRT-14 (TEM-45), a newly characterized mutant of TEM-type beta-lactamases.
abstract: IRT-14 (TEM-45) is a new mutant TEM-type beta-lactamase that was isolated from clinical Escherichia coli P37 and that confers resistance to broad-spectrum penicillins with reduced sensitivity to beta-lactamase inhibitors. The MICs of amoxicillin alone and of amoxicillin combined with 2 micrograms of clavulanic acid or 2 micrograms of tazobactam per ml were 4,096, 2,048, and 1,024 micrograms/ml, respectively. The strain was susceptible to cephalosporins, aztreonam, moxalactam, and imipenem. The enzyme was purified to homogeneity, and values of the kinetic parameters Kcat, Km, and Kcat/Km were determined for different substrates. This enzyme, with a pI of 5.2, was found to have reduced affinity for broad-spectrum penicillins and cephalosporins. The values of 50% inhibitory concentrations of clavulanic acid, sulbactam, tazobactam, and brobactam are correlated with the higher KmS for substrates. The resistance of E. coli P37 to mechanism-based inactivators results from a higher level of production of the TEM-derived enzyme due to the G-to-T substitution at position 162 (G-162-->T) in the promoter region of blaTEM and from the structural modifications resulting from the Met-69-->Leu and Arg-275-->Gln substitutions that characterize IRT-14 beta-lactamase.
authors.affiliation: INSERM U 120, Hôpital Robert Debré, Paris, France.
authors.forename: M M
authors.initials: MM
authors.lastname: Caniça
authors.forename: M
authors.initials: M
authors.lastname: Barthélémy
authors.forename: L
authors.initials: L
authors.lastname: Gilly
authors.forename: R
authors.initials: R
authors.lastname: Labia
authors.forename: R
authors.initials: R
authors.lastname: Krishnamoorthy
authors.forename: G
authors.initials: G
authors.lastname: Paul
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D004352:Drug Resistance, Microbial; D018432:Drug Resistance, Multiple; D004926:Escherichia coli / Q000187:drug effects* / Q000201:enzymology / Q000302:isolation & purification; D007525:Isoelectric Focusing; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D013379:Substrate Specificity; D001618:beta-Lactamases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; C105418:beta-lactamase IRT-14; D001618:beta-Lactamases