Artigo
A prothrombin activator from Bothrops erythromelas (jararaca-da-seca) snake venom: characterization and molecular cloning
Fecha
2003-01-01Registro en:
Biochemical Journal. London: Portland Press, v. 369, p. 129-139, 2003.
0264-6021
10.1042/BJ20020449
WOS:000180871000014
Autor
Silva, Márcia B.
Schattner, Mirta
Ramos, Celso RR
Junqueira-de-Azevedo, Inácio LM
Guarnieri, Míriam C.
Lazzari, Maria A.
Sampaio, Claudio Augusto Machado [UNIFESP]
Pozner, Roberto G.
Ventura, Janaína S.
Ho, Paulo L.
Chudzinski-Tavassi, Ana M.
Institución
Resumen
A novel prothrombin activator enzyme, which we have named 'berythractivase', was isolated from Bothrops erythromelas (jararaca-da-seca) snake venom. Berythractivase was purified by a single cation-exchange-chromatography step on a Resource S (Amersham Biosciences) column. the overall purification (31-fold) indicates that berythractivase comprises about 5% of the crude venom. It is a single-chain protein with a molecular mass of 78 kDa. SDS/PAGE of prothrombin after activation by berythractivase showed fragment patterns similar to those generated by group A prothrombin activators, which convert prothrombin into meizothrombin, independent of the prothrombinase complex. Chelating agents, such as EDTA and o-phenanthroline, rapidly inhibited the enzymic activity of berythractivase, like a typical metalloproteinase. Human fibrinogen Aalpha-chain was slowly digested only. after longer incubation with berythractivase, and no effect on the beta- or gamma-chains was observed. Berythractivase was also capable of triggering endothelial proinflammatory and procoagulant cell responses. von Willebrand factor-was released, and the surface expression of both intracellular adhesion molecule-1 and E-selectin was up-regulated by berythractivase in cultured human umbilical-vein endothelial cells. the complete berythractivase cDNA was cloned from a B. erythromelas venom-gland cDNA library. the cDNA sequence possesses 2330 bp and encodes a preproprotein with significant sequence similarity to many other mature metalloproteinases reported from snake venoms. Berythractivase contains metalloproteinase, desintegrin-like and cysteine-rich domains. However, berythractivase did not elicit any haemorrhagic response. These results show that, although the primary structure of berythractivase is related to that of snake-venom haemorrhagic metalloproteinases and functionally similar to group A prothrombin activators, it is a prothrombin activator devoid of haemorrhagic activity. This is a feature not observed for most of the snake venom metalloproteinases, including the group A prothrombin activators.