title: Crystal structure of the extended-spectrum β-lactamase PER-2 and insights into the role of specific residues in the interaction with β-lactams and β-lactamase inhibitors.
abstract: PER-2 belongs to a small (7 members to date) group of extended-spectrum β-lactamases. It has 88% amino acid identity with PER-1 and both display high catalytic efficiencies toward most β-lactams. In this study, we determined the X-ray structure of PER-2 at 2.20 Å and evaluated the possible role of several residues in the structure and activity toward β-lactams and mechanism-based inhibitors. PER-2 is defined by the presence of a singular trans bond between residues 166 to 167, which generates an inverted Ω loop, an expanded fold of this domain that results in a wide active site cavity that allows for efficient hydrolysis of antibiotics like the oxyimino-cephalosporins, and a series of exclusive interactions between residues not frequently involved in the stabilization of the active site in other class A β-lactamases. PER β-lactamases might be included within a cluster of evolutionarily related enzymes harboring the conserved residues Asp136 and Asn179. Other signature residues that define these enzymes seem to be Gln69, Arg220, Thr237, and probably Arg/Lys240A ("A" indicates an insertion according to Ambler's scheme for residue numbering in PER β-lactamases), with structurally important roles in the stabilization of the active site and proper orientation of catalytic water molecules, among others. We propose, supported by simulated models of PER-2 in combination with different β-lactams, the presence of a hydrogen-bond network connecting Ser70-Gln69-water-Thr237-Arg220 that might be important for the proper activity and inhibition of the enzyme. Therefore, we expect that mutations occurring in these positions will have impacts on the overall hydrolytic behavior.
authors: Laboratorio de Resistencia Bacteriana, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina.
authors: Melina
authors: M
authors: Ruggiero
authors: Centre d'Ingénierie des Protéines, Université de Liège, Liège, Belgium.
authors: Frédéric
authors: F
authors: Kerff
authors: Centre d'Ingénierie des Protéines, Université de Liège, Liège, Belgium.
authors: Raphaël
authors: R
authors: Herman
authors: Centre d'Ingénierie des Protéines, Université de Liège, Liège, Belgium.
authors: Frédéric
authors: F
authors: Sapunaric
authors: Centre d'Ingénierie des Protéines, Université de Liège, Liège, Belgium.
authors: Moreno
authors: M
authors: Galleni
authors: Laboratorio de Resistencia Bacteriana, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina.
authors: Gabriel
authors: G
authors: Gutkind
authors: Centre d'Ingénierie des Protéines, Université de Liège, Liège, Belgium.
authors: Paulette
authors: P
authors: Charlier
authors: Centre d'Ingénierie des Protéines, Université de Liège, Liège, Belgium.
authors: Eric
authors: E
authors: Sauvage
authors: Laboratorio de Resistencia Bacteriana, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina ppower@ffyb.uba.ar.
authors: Pablo
authors: P
authors: Power
mesh_terms: D000595:Amino Acid Sequence; D002511:Cephalosporins / Q000737:chemistry / Q000378:metabolism; D018360:Crystallography, X-Ray / Q000379:methods*; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017386:Sequence Homology, Amino Acid; D065093:beta-Lactamase Inhibitors / Q000378:metabolism*; D001618:beta-Lactamases / Q000737:chemistry* / Q000378:metabolism*; D047090:beta-Lactams / Q000378:metabolism*
chemical_list: D002511:Cephalosporins; D065093:beta-Lactamase Inhibitors; D047090:beta-Lactams; D001618:beta-Lactamases