IQUIFIB   02644
INSTITUTO DE QUIMICA Y FISICOQUIMICA BIOLOGICAS "PROF. ALEJANDRO C. PALADINI"
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Structural insights into the ?ceftazidimase? behavior of CTX-M-96 β-lactamase
Autor/es:
B. GHIGLIONE; M. M. RODRÍGUEZ; L. CURTO; M. GALLENI; P. CHARLIER; G. GUTKIND; E. SAUVAGE; P. POWER
Lugar:
Buenos Aires
Reunión:
Congreso; XLIII Reunión Anual SAB 2014; 2014
Institución organizadora:
Sociedad Argentina de Biofísica
Resumen:
Diversification of the CTX-M β-lactamases led to the emergence of variants responsible for decreased susceptibility to ceftazidime, being the D240G mutation the most prevalent among the so called ?ceftazidimases?. From the recently solved crystallographic structure of CTX-M-96 (1.2 Å), we analyzed the organization of the active site and evaluated the possible role of key amino acid residues in the overall stabilization of the structure and also in the interaction with different β-lactams and mechanism-based inhibitors. Wild-type blaCTX-M-96 and derived mutants in D240 were generated by site-directed mutagenesis. β-Lactamases were purified to homogeneity and kinetic parameters to selected β-lactams and inhibitors were determined. Circular dichroism was performed for all enzymes at near and far UV. CTX-M-96 presents some differences in the disposition of specific amino acids, although none of them seem to impair the interaction with β-lactams. In the absence of oxyimino-cephalosporins (OC), N132, E166, P167 and N170 seem to be shifted up to 0.7 Å away from the catalytic cleft, suggesting that the presence of antibiotic induces the approach of these residues towards the OC through hydrogen bonds. Circular dichroism results suggest that none of the mutations have negative impact in the overall structure. Structural differences do not seem to be conclusive to determine the ?ceftazidimase? behavior, and additional data are still needed to explain the observed in vivo resistance to OC.