title: Dramatic broadening of the substrate profile of the Aeromonas hydrophila CphA metallo-beta-lactamase by site-directed mutagenesis.
abstract: Among class B beta-lactamases, the subclass B2 CphA enzyme is characterized by a unique specificity profile. CphA efficiently hydrolyzes only carbapenems. In this work, we generated site-directed mutants that possess a strongly broadened activity spectrum when compared with the WT CphA. Strikingly, the N116H/N220G double mutant exhibits a substrate profile close to that observed for the broad spectrum subclass B1 enzymes. The double mutant is significantly activated by the binding of a second zinc ion under conditions where the WT enzyme is non-competitively inhibited by the same ion.
authors.affiliation: Centre d'Ingénierie des Protéines, Université de Liège, Allée de 6 Août B6, Sart-Tilmanm, 4000 Liège, Belgium.
authors.forename: Carine
authors.initials: C
authors.lastname: Bebrone
authors.forename: Christine
authors.initials: C
authors.lastname: Anne
authors.forename: Kris
authors.initials: K
authors.lastname: De Vriendt
authors.forename: Bart
authors.initials: B
authors.lastname: Devreese
authors.forename: Gian Maria
authors.initials: GM
authors.lastname: Rossolini
authors.forename: Jozef
authors.initials: J
authors.lastname: Van Beeumen
authors.forename: Jean-Marie
authors.initials: JM
authors.lastname: Frère
authors.forename: Moreno
authors.initials: M
authors.lastname: Galleni
mesh_terms: D016980:Aeromonas hydrophila / Q000201:enzymology*; D000900:Anti-Bacterial Agents / Q000493:pharmacokinetics; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D017931:DNA Primers; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D006868:Hydrolysis; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity; D001618:beta-Lactamases / Q000235:genetics* / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D017931:DNA Primers; D011994:Recombinant Proteins; D001618:beta-Lactamases; C082063:cphA protein, Aeromonas hydrophila