Potent Inactivation of Cathepsins S and L by Peptidyl (Acyloxy)methyl Ketones

Peptidyl (acyloxy)methyl ketones (Z-Aa-Aa-CH2-O-CO-R), a new class of irreversible inhibitors whose chemical reactivity can be modulated by varying the substitution pattern of the carboxylate leaving group, are shown to be extremely potent inactivators of the lysosomal cysteine proteinases cathepsin L and cathepsin S. The highest k2/Ki values measured were found to exceed 10(6) M-1s-1 for both cathepsin L and cathepsin S. The rate of inactivation can be controlled by varying the dipeptidyl moiety or the carboxylate leaving group, with the second-order rate constants for both enzymes found to be strongly dependent on the pKa values of the leaving group. The specificities of the cathepsins S and L reveal a different selectivity towards the nature of substitution of the aryl P' leaving group of the inhibitor. This new inhibitor class opens the possibility of the design of selective and specific inhibitors for lysosomal cysteine proteinases.