Characterization of Fibrinolytic Proteases from Gloydius Blomhoffii Siniticus Venom

摘要

Objectives: This study was undertaken to identify fibrinolytic proteases from gloydius blomhoffii siniticus venom and tocharacterize the major fibrinolytic protease purified from the venom.Methods: The venom was subjected to chromatography using columns of Q-Sepharose and Sephadex G-75. The molecularweights of fibrinolytic proteases showing a fibrinolytic zone on the fibrin plate assay were determined in SDS-PAGE (sodiumdodecyl sulfate-polyacrylamide gel electrophoresis) The effects of inhibitors and metal ions on fibrinolytic protease and theproteolysis patterns of fibrinogen, gelatin, and bovine serum albumin were investigated.Results:1. The fibrinolytic fractions of the three peaks isolated from gloydius blomhoffii siniticus venom contained two poly-peptides each of 46 and 59 kDa and three polypeptides each of 32, 18, and 15 kDa and one major polypeptide of 54 kDa.2. The fibrinolytic activity of the purified protease of 54 kDA was inhibited by metal chelators, such as ethyl-enediaminetetraacetic acid (EDTA), ethylene glycol tetraacetic acid (EGTA), and 1, 10-phenanthroline, and by disul-fhydryl-reducing compounds, such as dithiothreitol and cysteine.3. Calcium chloride promoted the fibrinolytic activity of the protease, but mercuric chloride and cobalt(II) chlorideinhibited it.4. The fibrinolytic protease preferentially cleaved Aa-chains and slowly elevated Bb-chains of fibrinogen. It also hydro-lyzed gelatin, but not bovine serum albumin.Conclusions: The gloydius blomhoffii siniticus venom contained more than three fibrinolytic proteases. The major fibri-nolytic protease was a metalloprotease that hydrolyzed both fibrinogen and gelatin, but not bovine serum albumin.