Mechanism of substrate-induced inactivation of beta-lactamase I

Abstract

beta-Lactamase I (from Bacillus cereus 569/H) is inactivated by certain substrates (e.g. methicillin or cloxacillin) but not by others (e.g. benzylpenicillin). Emzyme that had been inactivated was found to be labelled stoichiometrically, as shown by the use of radioactive methicillin. Use of the penamaldate reaction showed the presence of a penicilloyl group in the enzyme inactivated by either methicillin or cloxacillin. In conditions under which enzymic activity was regained the penicilloyl group was shed. When the activity of beta-lactamase I was measured in 0.3-1.2 M guanidinium chloride the rates of hydrolysis of methicillin or cloxacillin (but not benzylpenicillin) were greatly reduced. The unliganded enzyme was stable. The results are explained by supposing that a normal intermediate, the acyl enzyme, is prone to unfold.