Metallo-beta-lactamase from Bacillus cereus as a framework for novel metllohydrolases

JAVIER MARCELO GONZALEZ; ALEJANDRO J. VILA

Angra dos Reis

Congreso; First Latin American Protein Society Meeting; 2004

Latin American Protein Society

Metallo-beta-lactamases (MBL) are zinc hydrolases that confer broad-spectrum resistance to beta-lactam antibiotics. To date, no clinically useful inhibitors have been designed for these enzymes. MBLs have been classified into three subclasses based on sequence homology. Plasmid-borne subclass B1 MBLs stand as rapidly evolving resistance mechanisms among pathogenic bacteria. B2 and B3 MBLs are divergent chromosomal enzymes exhibiting dissimilar catalytic mechanisms.   The enzyme BcII from B. cereus, strictly related to B. anthracis, is a subclass B1 chromosomal enzyme related to B1 MBLs. We have generated mutants of BcII that resemble the primary zinc coordination shell of L1 and glyoxalase II; BcII-HS (BcII-R121H/C221S) and BcII-HD (BcII-R121H/C221D), respectively. L1 is a subclass B3 MBL found in S. maltophilia, and glyoxalase II is a thiolesterase structurally related to MBLs. Notably, BcII-HS evinced a diminished catalytic efficiency, about 10 to 100-fold decrease against nitrocefin and cefaloridine, and 103 to 104-fold decrease against benzylpenicillin and imipenem. In addition, BcII-HD showed no glyoxalase II activity and a severely reduced b-lactamase activity, with catalytic efficiencies about 10 to 100 times lower than those determined for BcII-HS.