CYSTATIN-C BASED PEPTIDYL DIAZOMETHANES AS CYSTEINE PROTEINASE-INHIBITORS - INFLUENCE OF THE PEPTIDYL CHAIN-LENGTH

The peptidyl diazomethanes Cbz-Gly-CHN2, Boc-Val-Gly-CHN2, H-Leu-Val-Gly-CHN2, Cbz-Leu-Val-Gly-CHN2 and Cbz-Arg-Leu-Val-Gly-CHN2, with peptidyl portions modelled after the proposed cysteine proteinase interacting N-terminal segment of human cystatin C, were synthesized. Their efficiency as cysteine proteinase inhibitors was tested against papain, human cathepsin B and bovine cathepsin B. All, except Cbz-Gly-CHN2, were found to be irreversible inhibitors of the tested enzymes. Each addition of an amino acid residue to their peptidyl portions resulted in an increased inhibition rate of all three enzymes. These data suggest that the arginyl residue of the tetrapeptidyl diazomethane, and also the corresponding arginyl residue in native cystatin C, interact with a S4 substrate pocket subsite of both papain and cathepsin B. The most efficient inhibitor, Cbz-Arg-Leu-Val-Gly-CHN2, inhibited papain and cathepsin B with rate constants of the same order of magnitude as those for L-3-carboxy-trans-2,3-epoxypropionyl-leucylamido(4-guanidino)butane (E-64). The high water-solubility of Cbz-Arg-Leu-Val-Gly-CHN, allowing it to be dissolved to molar concentrations without use of non-physiological additives, makes it suitable for in vitro and in vivo cysteine proteinase inhibition studies.