The inactivation of bovine cathepsin B by novel N-chloro-acetyl-dipeptides: application of the Houghten 'tea bag' methodology to inhibitor synthesis.
In this study, a series of N-chloro-acetylated dipeptides were synthesised by the application of Houghten's methodology of multiple analog peptide syntheses (MAPS). The peptides, all of which contain a C-terminal free acid, were tested as inactivators of bovine cathepsin B, in an attempt at exploiting the known and, amongst the cysteine proteinases, unique carboxy dipeptidyl peptidase activity of the protease. We have succeeded in obtaining a number of effective inactivators, the most potent of which-chloroacetyl-Leu-Leu-OH, inactivates the enzyme with an apparent second-order rate constant of 3.8 x 10(4)M(-1)min(-1). In contrast, the esterified analog, chloroacetyl-Leu-Leu-OMe, inactivates the enzyme some three orders of magnitude less efficiently, lending credence to our thesis that a free carboxylic acid moiety is an important determinant for inhibitor effectiveness. This preliminary study has highlighted a number of interesting features about the specificity requirements of the bovine proteinase and we believe that our approach has great potential for the rapid delineation of the subsite specificities of cathepsin B-like proteases from various species.
Biomolecular Sciences Group, The School of Pharmacy, McClay Research Centre Queen's University Belfast, 97 Lisburn Road, Belfast BT9 7BL, UK.
B F
BF
Gilmore
P
P
Harriott
B
B
Walker
D000107:Acetylation; D000818:Animals; D001665:Binding Sites; D002401:Cathepsin B / Q000037:antagonists & inhibitors / Q000737:chemistry*; D002417:Cattle; D017606:Chlorine Compounds / Q000737:chemistry; D004151:Dipeptides / Q000737:chemistry*; D015195:Drug Design; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000737:chemistry; D007700:Kinetics; D011485:Protein Binding
D017606:Chlorine Compounds; D004151:Dipeptides; D004791:Enzyme Inhibitors; D002401:Cathepsin B