Evaluation of inhibition of the carbenicillin-hydrolyzing beta-lactamase PSE-4 by the clinically used mechanism-based inhibitors

Characterization of the biochemical steps in the inactivation chemistry of clavulanic acid, sulbactam and tazobactam with the carbenicillin-hydrolyzin g beta-lactamase PSE-4 from Pseudomonas aeruginosa is described, Although t azobactam showed the highest affinity to the enzyme, all three inactivators were excellent inhibitors for this enzyme, Transient inhibition was observ ed for the three inactivators before the onset of irreversible inactivation of the enzyme. Partition ratios (k(cat)/k(inact)) of 11, 41 and 131 were o btained with clavulanic acid, tazobactam and sulbactam, respectively. Furth ermore, these values were found to be 14-fold, 3-fold and 80-fold lower, re spectively, than the values obtained for the clinically important TEM-I bet a-lactamase, The kinetic findings were put in perspective by determining th e computational models for the pre-acylation complexes and the immediate ac yl-enzyme intermediates for all three inactivators, A discussion of the per tinent structural factors is presented, with PSE-4 showing subtle differenc es in interactions with the three inhibitors compared to the TEM-I enzyme. (C) 2000 Federation of European Biochemical Societies.