INHIBITION OF CATHEPSIN-B AND PAPAIN BY PEPTIDYL, ALPHA-KETO ESTERS, ALPHA-KETO AMIDES, ALPHA-DIKETONES, AND ALPHA-KETO ACIDS

A series of peptidyl α-keto esters, α-keto amides, α-keto acids, and α-diketones were synthesized which reversibly inhibit papain and cathepsin B. Methyl 3-(N-benzyloxycarbonyl - l- phenylalanyl)amino-2-oxopro-pionate (a dicarbonyl compound) inhibits papain with a ki value of 1 μm, whereas the Ki of 3-(N-acetyl-l-phenylalanyl)aminopropanone (a monocarbonyl compound) is 1.5 mm (M. R. Bendall et al., 1979. Eur. J. Biochem. 79, 201-209). Both carbonyl groups are required for effective inhibition. Extension of these inhibitors by addition of P′ substituents (e.g., hexyl) does not affect the Ki for papain, but reduces Ki for cathepsin B 33-fold. For these two enzymes slow binding inhibition was observed with slow on rates (kon, 5.2 × 102 M-1 s-1 for papain, and 2.7 × 103 m- s- for cathepsin B). Addition of a P3 substituent (glycine) has no effect on Ki. We propose that the mechanism of inhibition involves the formation of a hemithioketal by addition of the active-site thiol to the carbonyl group of the inhibitor closer to the N-terminus. The hemithioketal intermediate is most likely stabilized by the electron withdrawing effect of the second carbonyl group. © 1990.