Inhibition of Macromolecular Synthesis in Escherichia coli by Protease Inhibitors

A number of chloromethyl ketone protease inhibitors reversibly inhibit β-galactosidase induction, growth, RNA synthesis, and protein synthesis in Escherichia coli at concentrations (0.1 to 1.0 mm) which are not toxic to the cells. The competitive protease inhibitor pentamidine isothionate inhibits β-galactosidase induction at a concentration (0.01 mm) which has little effect on general macromolecular synthesis. The action of chloromethyl ketone compounds is specifically blocked by reduced glutathione. Neither oxidized glutathione, Cleland's reagent, cysteine, nor mercaptoethanol were able to replace glutathione. Although there is a structural similarity between chloromethyl ketones and iodoacetamide, our studies indicate that chloromethyl ketones do not act as general sulfhydryl inhibitors. While tosyl lysine chloromethyl ketone can react with glutathione under neutral or alkaline conditions it does not inactivate RNase, as does iodoacetamide. Furthermore, glutathione was able to reverse the inhibition caused by tosyl-lysine chloromethyl ketone even when added 15 min after inhibition has been established. Glutathione is not able to reverse the inhibition caused by iodoacetamide. In E. coli, the inhibition of macromolecular synthesis by chloromethyl ketones may be the result of reactions with intracellular glutathione.