Mechanism of inhibition of LDL phospholipase A2 by monocyclic-β-lactams.: Burst kinetics and the effect of stereochemistry

Investigation of the inhibition of LDL-associated phospholipase A(2) by monocyclic beta-lactams has shown that LDL phospholipase A(2) is capable of hydrolyzing monocyclic-beta-lactams by a mechanism which shares many similarities to the hydrolysis of beta-lactams by beta-lactamases, We believe that this is the first demonstration of a serine-dependent lipase being able to hydrolyze an amide bond. Although 4-(phenylthio)-N-(4-phenyl-2-oxobutyl)az SB-216477, and its enantiomers are relatively modest covalent inactivators with k(obs)/[I] = 46 M-1 s(-1) for the R enantiomer, analysis of the kinetics of inactivation and reactivation shows that these compounds act as slow-turnover substrates, presumably via an acylation-deacylation mechanism. The detection of a suprastoichiometric burst indicates that the pathway must be branched with the branching giving rise to the slow reactivation via a more stable covalent intermediate. Study of the two enantiomers of SE-216477 shows that LDL-associated phospholipase A(2) is sensitive to the p-lactam stereochemistry at C4. However, a common achiral intermediate is formed along the turnover pathway, and this must be at or immediately prior to the branch point.