Snake venom proteinase inhibitors: I. isolation and properties of two inhibitors of kallikrein, trypsin, plasmin, and α-chymotrypsin from the venom of russell's viper (vipera russelli)

Abstract

Several substances which inhibit the activities of bovine plasma kallikrein [EC 3.4.21.8], trypsin [EC 3.4.21.4], plasmin [EC 3.4.21.7], and α-chymotrypsin [EC 3.4.21.1], were separated from crude venom of Russell's viper (Vipera russelli). All those inhibitors were polypeptidic substances having a molecular weight of about 6,000 to 7,000 and two of them, designated as inhibitors I and II, were purified extensively and their properties examined. The final preparations appeared pure on disc polyacrylamide gel electrophoresis with and without sodium dodecylsulfate (SDS). Their molecular weights were estimated by SDS-gel electrophoresis to be about 7,200 (±1,000). Inhibitors I and II contained 52 and 60 amino acid residues, respectively. The amino-terminal residue of inhibitor II was a single histidine and the carboxyl-terminus was lysine. The N-terminal of inhibitor I did not react with phenylisothiocyanate, suggesting that it was in a masked form. The carboxyl-terminus of inhibitor I was lysine.Both inhibitors inactivated bovine trypsin, probably by formation an enzyme-inhibitor complex in a molar ratio of 1: 1. The K1 values of inhibitor II, measured using synthetic ester substrates, were 7.6×10-10 M for bovine trypsin, 1.4×10-10 M for bovine α-chymotrypsin, 2.9×10-10M for bovine plasma kallikrein, and 1.0×10-9 M for bovine plasmin. Inhibitor II has no effect on the activities of thrombin [EC 3.4.21.5], thiol proteinases (bromelain [EC 3.4.22.4], papain [EC 3.4.22.2], and ficin [EC 3.4.22.3]), metal proteinases (thermolysin [EC 3.4.24.4]) or exopeptidases (carboxy-peptidases A and B [EC 3.4.12.2 and 3]). Thus, its inhibition spectrum was very similar to that of pancreatic basic trypsin inhibitor (Kunitz type). © 1974 THE JOURNAL OF BIOCHEMISTRY.