Hydrolysis of 3-substituted cephalosporins catalysed by ?-lactamases I and II from Bacillus cereus and by hydroxide ion

Second-order rate constants for the alkaline hydrolysis of 3-thiol substituted cephalosporins are independent of the pKa of the thiol over a pKa range of 9. If there is a leaving group at C-3 it is expelled after the -lactam ring is opened and the expulsion of the leaving group does not enhance the rate of -lactam C–N bond fission. The zinc enzyme -lactamase II is about a 100-fold better catalyst than the serine enzyme -lactamase I for the hydrolysis of the same cephalosporin. The second-order rate constant kcat/Km for both -lactamase enzymes shows no dependence on the nature of the substituent at C-3 which is not explicable by the different chemical reactivity of the cephalosporins. There is no evidence for a significant recognition site in either enzyme for the C-3 substituent. The kinetic parameters kcat and Km for the -lactamase I-catalysed hydrolysis may be complicated by the formation of intermediates.