Bacillus cereus metallo-beta-lactamase is active both in the mono- and binuclear forms

TIONI, M.F.; LLARRULL, L.I.; BENNETT, B.; VILA, A.J.

Rosario

Congreso; XXXV Reunión Anual Sociedad Argentina de Biofísica; 2006

Sociedad Argentina de Biofísica

The mechanism of hydrolysis of imipenem by the metallo-beta-lactamase BcII from Bacillus cereus was studied by stopped-flow and rapid Freeze Quench-EPR. The changes in the metal coordination of the enzyme substituted with different Co(II)/enzyme ratios were evaluated during the reaction, which allowed us to conclude that BcII uses a branched mechanism for imipenem hydrolysis, that the branching point involves metal dissociation and that the mono-Co(II) enzyme is active. The RFQ-EPR assays provide support to the hypothesis that at substoichiometric Co(II)/enzyme ratios, the mono-substituted enzyme coexists with considerable amounts of binuclear, fully loaded enzyme. Together, these results indicate that BcII is active when it is loaded with either one or two metal ions. The EPR results also indicate that a distint product-enzyme complex is formed.