Characterization Of Two Proteolytic Enzymes Purified From Bothrops alternatus Snake Venom.

¹Costa, J. de O.; ¹Izidoro, L. F. M.; ¹Hamaguchi, A.; ¹Homsi-Brandeburgo, M. I. and ²Oliveira, F.

1 Instituto de Genética e Bioquímica, 2 Departamento de Ciências Fisiológicas/Instituto de Ciências Biomédicas, Universidade Federal de Uberlândia, Minas Gerais, Brasil.

The present work describes the purification and characterization of two fibrinogenolytic enzymes, called MPA1 and MPA2, from Bothropsalternatus snake venom. Both enzymes were obtained by DEAE-Sephacel, Sephadex G-75 and Heparin-Agarose chromatographies. MPA1 and MPA2 showed each one a single protein band on polyacrylamide gel electrophoresis (SDS-PAGE) and apparent mol. weight of 29,000 and 36,000, respectively. Fibrinogenolytic activity upon fibrinogen were determinate. Samples of 50µL of bovine fibrinogen (1mg/mL PBS) were incubated with enzymes (1:100 w/w) at 37ºC for 1h, pH 8.0. This activity were analyzed by 14% SDS-PAGE Both enzymes were proteolytically active against bovine fibrinogen. MPA2 cleaved the Aa-chain first, then the Bb-chain. The action of MPA1 was similar, but slower. None of the proteases degraded the g-chain of fibrinogen. The fibrinogenolytic activity of the enzymes was inhibited by 1,10-phenanthroline, suggesting that both enzymes are metalloproteases or -fibrinogenases. Since the both enzymes were found to cause blood incoagulability, when administered i. p. via in mice, it is expected that it may be of medical interest as a therapeutic agent in the treatment and prevention of arterial thrombosis.

Support: CAPES, CNPq, FAPEMIG and UFU.

Correspondence to: lfmizidoro@zipmail.com.br