Grafting a new metal ligand in the cocatalytic site of B. cereus metallo-beta-lactamase: structural flexibility without loss of activity.
Metallo-beta-lactamases are zinc enzymes able to hydrolyze the four-membered ring of beta-lactam antibiotics, representing one of the latest generations of beta-lactamases. These enzymes belong to the zinc metallo-hydrolase family of the beta-lactamase fold. Enzymes belonging to this family have a bimetallic active site whose structure varies among different members by point substitutions of the metal ligands. In this work, we have grafted new metal ligands into the metal binding site of BcII from Bacillus cereus that mimic the ligands present in other members of this superfamily. We have characterized spectroscopically and modeled the structure of the redesigned sites, which differ substantially from the wild-type enzyme. Despite the changes introduced in the active site, the mutant enzymes retain almost full activity. These results shed some light on the possible evolutionary origin of these metalloenzymes.
Rasia
Rodolfo M
RM
Molecular Biology Division, Instituto de Biología Molecular y Celular Rosario, Consejo Nacional de Investigaciones Cientificas y Técnicas, University of Rosario, Suipacha 531, S2002LRK Rosario, Argentina.
Ceolín
Marcelo
M
Vila
Alejandro J
AJ
D001409:Bacillus cereus / Q000201:enzymology*; D001665:Binding Sites; D020134:Catalytic Domain; D002511:Cephalosporins / Q000737:chemistry / Q000378:metabolism; D003035:Cobalt / Q000737:chemistry; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D008670:Metals / Q000737:chemistry*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010400:Penicillin G / Q000737:chemistry / Q000378:metabolism; D018583:Pliability; D016415:Sequence Alignment; D013053:Spectrophotometry; D015032:Zinc / Q000032:analysis / Q000737:chemistry; D001618:beta-Lactamases / Q000737:chemistry* / Q000378:metabolism
D002511:Cephalosporins; D008024:Ligands; D008670:Metals; D003035:Cobalt; D001618:beta-Lactamases; C021720:nitrocefin; D015032:Zinc; D010400:Penicillin G