title: In vivo impact of Met221 substitution in GOB metallo-β-lactamase.
abstract: Metallo-β-lactamases (MβLs) represent one of the main mechanisms of bacterial resistance against β-lactam antibiotics. The elucidation of their mechanism has been limited mostly by the structural diversity among their active sites. All MβLs structurally characterized so far present a Cys or a Ser residue at position 221, which is critical for catalysis. GOB lactamases stand as an exception within this picture, possessing a Met residue in this location. We studied different mutants in this position, and we show that Met221 is essential for protein stability, most likely due to its involvement in a hydrophobic core. In contrast to other known MβLs, residue 221 is not involved in metal binding or in catalysis in GOB enzymes, further highlighting the structural diversity of MβLs. We also demonstrate the usefulness of protein periplasmic profiles to assess the contribution of protein stability to antibiotic resistance.
authors.affiliation: Departamento de Química Biológica and Instituto de Biología Molecular y Celular de Rosario-IBR, CONICET-UNR, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Rosario, Argentina.
authors.forename: Jorgelina
authors.initials: J
authors.lastname: Morán-Barrio
authors.forename: María-Natalia
authors.initials: MN
authors.lastname: Lisa
authors.forename: Alejandro J
authors.initials: AJ
authors.lastname: Vila
mesh_terms: D019943:Amino Acid Substitution; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D024881:Drug Resistance, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics / Q000472:pathogenicity; D008715:Methionine / Q000235:genetics*; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics / Q000502:physiology; D010957:Plasmids / Q000235:genetics; D011487:Protein Conformation; D001618:beta-Lactamases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D008715:Methionine; D001618:beta-Lactamases