RESUMEN
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.
Asunto(s)
Venenos de Crotálidos/enzimología , Venenos de Crotálidos/aislamiento & purificación , Metaloendopeptidasas/aislamiento & purificación , Serina Endopeptidasas/aislamiento & purificación , Trimeresurus , Secuencia de Aminoácidos , Animales , Cromatografía DEAE-Celulosa , Cromatografía por Intercambio Iónico , Venenos de Crotálidos/química , Venenos de Crotálidos/metabolismo , Estabilidad de Enzimas , Fibrinógeno/metabolismo , Hemorragia/inducido químicamente , Metaloendopeptidasas/química , Metaloendopeptidasas/metabolismo , Ratones , Datos de Secuencia Molecular , Agregación Plaquetaria/efectos de los fármacos , Conejos , Homología de Secuencia de Aminoácido , Serina Endopeptidasas/química , Serina Endopeptidasas/metabolismo , Inhibidores de Serina Proteinasa/farmacología , Especificidad por SustratoRESUMEN
Several biochemical and biological activities such as phospholipase A2, arginine esterase, proteolytic, L-amino acid oxidase, 5'nucleotidase, acetylcholinesterase, thrombin-like, anticoagulant, and hemorrhagic activities were determined for whole desiccated venom of Trimeresurus jerdonii. An acidic phospholipase (named TJ-PLA2) was purified by anionic exchange chromatography, gel filtration, and reverse phase HPLC. TJ-PLA2 had a molecular weight of 16,000 and a pI of 4.8. TJ-PLA2 was non-lethal to mice up to an i.p. dose of 15 mg/kg body weight and lacked neurotoxicity and myotoxicity. It induced edema in the footpads of mice. The purified enzyme inhibited ADP- and collagen-induced human platelet aggregation in a manner which was both dose- and time-dependent.
