Synthesis of peptide fluoromethyl ketones and the inhibition of human cathepsin B

Peptide fluoromethyl ketones have been synthesized for the first time. The inhibitor 3-(N-benzyloxycarbonylphenylalanylamido)-DL-1-fluoro-2-butanone (Z-Phe-AlaCH2F) was found to be a 30-fold more potent inactivator of human cathepsin B than 3-(N-benzyloxycarbonylphenylalanylamido)-L-1-diazo-2-butanone (Z-Phe-AlaCHN2), but less reactive than 3-(N-benzyloxycarbonylphenylalanylamido)-L-1-chloro-2-butanone (Z-Phe-AlaCH2Cl). The fluoromethyl ketone's increased potency over the diazomethyl ketone is mainly due to its tighter binding to cathepsin B, with little difference between their respective k3 values. Both Z-Phe-AlaCHN2 and Z-Phe-AlaCH2F were quite stable to high concentrations of dithiothreitol, while Z-Phe-AlaCH2Cl was rapidly destroyed by the thiol.