title: Catalytic mechanism of class B2 metallo-beta-lactamase.
abstract: The initial nucleophilic substitution step of biapenem hydrolysis catalyzed by a subclass B2 metallo-beta-lactamase (CphA from Aeromonas hydrophila) is investigated using hybrid quantum mechanical/molecular mechanical methods and density functional theory. We focused on a recently proposed catalytic mechanism that involves a non-metal-binding water nucleophile in the active site of the monozinc CphA. Both theoretical models identified a single transition state featuring nearly concomitant nucleophilic addition and elimination steps, and the activation free energy from the potential of mean force calculations was estimated to be approximately 14 kcal/mol. The theoretical results also identified the general base for activating the water nucleophile to be the metal-binding Asp-120 rather than His-118, as suggested earlier. The protonation of Asp-120 leads to cleavage of the O(delta2)-Zn coordination bond, whereas the negatively charged nitrogen leaving group resulting from the ring opening replaces Asp-120 as the fourth ligand of the sole zinc ion. The electrophilic catalysis by the metal ion provides sufficient stabilization for the leaving group to avoid a tetrahedral intermediate. The theoretical studies provided detailed insights into the catalytic strategy of this unique metallo-beta-lactamase.
authors.affiliation: Department of Chemistry, University of New Mexico, Albuquerque, New Mexico 87131.
authors.forename: Dingguo
authors.initials: D
authors.lastname: Xu
authors.forename: Daiqian
authors.initials: D
authors.lastname: Xie
authors.forename: Hua
authors.initials: H
authors.lastname: Guo
mesh_terms: D016980:Aeromonas hydrophila / Q000201:enzymology; D001224:Aspartic Acid / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry / Q000145:classification* / Q000378:metabolism*; D001665:Binding Sites; D015780:Carbapenems / Q000378:metabolism; D002384:Catalysis; D006868:Hydrolysis; D008024:Ligands; D008956:Models, Chemical; D008958:Models, Molecular; D008962:Models, Theoretical; D011485:Protein Binding; D011487:Protein Conformation; D011522:Protons; D011789:Quantum Theory; D013816:Thermodynamics; D013845:Thienamycins / Q000378:metabolism; D014867:Water / Q000737:chemistry; D015032:Zinc / Q000737:chemistry; D001618:beta-Lactamases / Q000737:chemistry / Q000145:classification* / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D015780:Carbapenems; D008024:Ligands; D011522:Protons; D013845:Thienamycins; D014867:Water; D001224:Aspartic Acid; D001618:beta-Lactamases; D015032:Zinc; C065257:biapenem