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A fibrinogen-clotting enzyme designed as jerdonobin-II was isolated from the venom of Trimeresurus jerdonii. It differed in molecular weight and N-terminal sequence with the previously isolated jerdonobin, a thrombin-like enzyme from the same venom. The enzyme consists of a single polypeptide chain with molecular weights of 30,000 and 32,000 under non-reducing and reducing conditions, respectively. Jerdonobin-II showed weak fibrinogen clotting activity and its activity unit on fibrinogen was calculated to be less than one unit using human thrombin as standard. The precursor protein sequence of jerodonobin-II was deduced from cloned cDNA sequence. The sequence shows high similarity (identity=89%) to TSV-PA, a specific plasminogen activator from venom of T. stejnegeri. Despite of the sequence similarity, jerdonobin-II was found devoid of plasminogen activating effect. Sequence alignment analysis suggested that the replacement of Lys239 in TSV-PA to Gln239 in jerdonobin-II might play an important role on their plasminogen activating activity difference.

A novel fibrinogenolytic protease, named alpha-mucrofibrase, was purified from the venom of Chinese Habu (Trimeresurus mucrosquamatus) by DEAE-Sephadex A-50 ion-exchange chromatography and Sephadex G-100 (super fine) gel filtration alpha-Mucrofibrase is a single-chain polypeptide of approximately 29 kDa. It is stable even at 95 degrees C, and the most susceptible hydrolysis substrate is S-2302. It cleaved primarily the Aalpha chain of fibrinogen followed by the Bbeta chain, while the gamma chain was partially affected. N-terminal sequence of this fibrinogenolytic enzyme has great homology with those of other snake venom serine proteases. The esterase activity of alpha-mucrofibrase is inhibited by phenylmethylsulfonyl fluoride (PMSF) but not by metal chelator (EDTA), suggesting this fibrinogenase belongs to the venom serine protease family.

Jerdonobin and jerdofibrase are two serine proteases purified from the venom of Trimeresurus jerdonii. The Michaelis constant K(m) and the catalytic rate constant K(cat) 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 Bbeta-chain of fibrinogen. It also degraded Aalpha-chain of fibrinogen with relatively slow activity, but did not act on the gamma-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.

A fibrin(ogen)olytic serine protease from Trimeresurus jerdonii venom was identified and purified to SDS-polyacrylamide gel electrophoresis homogeneity. It is a single chain polypeptide with a molecular weight of 32kDa under reduced condition and 28kDa under non-reduced condition, respectively. The venom protease catalysed the hydrolysis of some chromogenic substrates such as S2238, S2160, S2302 and S2251. It degraded Bbeta-chain of human fibrinogen preferentially. Also the enzyme degraded fibrin directly. Its enzymatic activity was completely inhibited by phenylmethylsulfonyl fluoride (PMSF), but not affected by EDTA. That suggested it was a serine protease. N-terminal sequence of the purified component showed high homology with other snake venom serine proteases.

From the venom of Trimeresurus jerdonii, a distinct thrombin-like enzyme, called jerdonobin, was purified by DEAE A-25 ion-exchange chromatography, Sephadex G-75 gel filtration, and fast protein liquid chromatography (FPLC). SDS-PAGE analysis of this enzyme shows that it consists of a single polypeptide chain with a molecular weight of 38,000. The NH(2)-terminal amino acid sequence of jerdonobin has great homology with venom thrombin-like enzymes documented. Jerdonobin is able to hydrolyze several chromogenic substrates. The enzyme directly clots fibrinogen with an activity of 217 NIH units/mg. The fibrinopeptides released, identified by HPLC, consisted of fibrinopeptide A and a small amount of fibrinopepide B. The activities of the enzyme were inhibited by phenylmethylsulfonyl fluoride (PMSF) and p-nitrophenyl-p-guanidinobenzoate (NPGB). However, metal chelator (EDTA) had no effect on it, indicating it is venom serine protease.