AMIDINO-SUBSTITUTED AROMATIC HETEROCYCLES AS PROBES OF THE SPECIFICITY POCKET OF TRYPSIN-LIKE PROTEASES

A number of amidino-substituted heterocycles were synthesized and examined for their inhibitory effect against arginine-directed esteroproteases, including human and bovine thrombin (EC 3.4.21.5), human factor Xa and plasmin (EC 3.4.21.7), bovine trypsin (EC 3.4.21.4), porcine pancreatic Kallikrein (EC 3.4.21.8), and boar acrosin (EC 3.4.21.10). Inhibition was competitive and reversible in all cases, and the Ki values were taken to reflect binding conditions in the specificity pockets of the enzymes. A remarkably potent blocking agent was found in 5-amidinoindole. Compared to the established inhibitor benzamidine, it proved one to two orders of magnitude more effective against the majority of the proteases, the exceptions being trypsin and plasmin. Improved hydrophobic interaction, fortuitous hydrogen bonding, and charge transfer complex formation were considered in accounting for the considerable activity of the compound. Placement of the amidino moiety in the 6-position rather than in the 5-position on the indole ring resulted in a striking loss of inhibitory potency against human and bovine thrombin, factor Xa, and Kallikrein, yet slightly improved the affinity with respect to acrosin. Tightness of binding of the inhibitors to the enzymes was pH dependent and was significantly reduced below pH 7.0. © 1979.