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Jerdonobin and jerdofibrase are two serine proteases purified from the venom of Trimeresurus jerdonii. The Michaelis constant Km and the catalytic rate constant Kcat of jerdonobin or jerdofibrase on three chromogenic substrates, H-D-Pro–Phe–Arg–pNA (S2302), H-D-Phe-pipecolyl-Arg-pNA (S2238), and H-D-Val–Leu–Lys–pNA (S2251) were obtained from lineweaver–Burk plots. Jerdofibrase could hydrolyze all three substrates, but jerdonobin had no detectable activity on S2251, suggesting a relatively broader substrate specificity for jerdofibrase than jerdonobin. By SDS-PAGE, jerdofibrase preferentially degraded Bβ-chain of fibrinogen. It also degraded Aα-chain of fibrinogen with relatively slow activity, but did not act on the γ-chain. In contrast, jerdonobin did not degrade fibrinogen within 12 h. Fibrinopeptides liberation test, identified by HPLC, showed jerdonobin released fibrinopeptide A and a small amount of fibrinopeptide B. Unlike jerdonobin, jerdofibrase mainly released fibrinopeptide B. These results indicate that the two enzymes differ in their ability to hydrolyze chromogenic substrates and in their actions on fibrinogen. [Copyright &y& Elsevier]

A novel disintegrin, jerdonatin, was purified to homogeneity from Trimeresurus jerdonii venom by gel filtration and reversed-phase high-pressure liquid chromatography. We isolated the cDNA encoding jerdonatin from the snake venom gland. Jerdonatin cDNA precursor encoded pre-peptide, metalloprotease and disintegrin domain. Jerdonatin is composed of 72 amino acid residues including 12 cysteines and the tripeptide sequence Arg–Gly–Asp (RGD), a well-known characteristic of the disintegrin family. Molecular mass of jerdonatin was determined to be 8011 Da by matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS). Jerdonatin inhibited ADP- and collagen-induced human platelet aggregation with IC50 of 123 and 135 nM, respectively. We also investigated the effect of jerdonatin on the binding of B6D2F1 hybrid mice spermatozoa to mice zona-free eggs and their subsequent fusion. Jerdonatin significantly inhibited sperm–egg binding in a concentration-dependent manner, but had no effect on the fusion of sperm–egg. These results indicate that integrins on the egg play a role in mammalian fertilization. [Copyright &y& Elsevier]

A chymotrypsin inhibitor, designated NA-CI, was isolated from the venom of the Chinese cobra Naja atra by three-step chromatography. It inhibited bovine α-chymotrypsin with a Ki of 25 nM. The molecular mass of NA-CI was determined to be 6403.8 Da by matrix-assisted laser-desorption ionization time-of-flight (MALDI-TOF) analysis. The complete amino acid sequence was determined after digestion of S-carboxymethylated inhibitor with Staphylococcus aureus V8 protease and porcine trypsin. NA-CI was a single polypeptide chain composed of 57 amino acid residues. The main contact site with the protease (P1) has a Phe, showing the specificity of the inhibitor. NA-CI shared great similarity with the chymotrypsin inhibitor from Naja naja venom (identities=89.5%) and other snake venom protease inhibitors. [Copyright &y& Elsevier]