Efficient inhibition of class A and class D β-lactamases by Michaelis complexes

A 6-alkylidiene penam sulfone, SA-1-204, is an efficient inhibitor of both SHV-1 and OXA-1 beta-lactamases with K-I = 42 +/- 4nM and 1.0 +/- 0.1 mu M, respectively. To gain insight into the reaction chemistry of SA-1-204, the reactions between this inhibitor and SHV-1 and OXA-1 were studied by Raman spectroscopy in single crystals and in solution. Raman signatures characteristic of the unreacted beta-lactam ring show that in both phases the inhibitor binds as a noncovalent Michaelis-like complex. This complex is present as the major population for periods of up to an hour. On longer time scales, the Raman data show that beta-lactam ring opening eventually leads to a complex mixture of reaction products. However, the data clearly demonstrate that the key species for inhibition on the time scale of bacterial half-lives is the noncovalent complex preceding acylation.