Purification and Characterization of Low Molecular Weight Fibrinolytic/Hemorrhagic Enzymes from Snake (Bothrops jararaca) Venom

Abstract

Two low molecular weight fibrinolytic/hemorrhagic enzymes, jararafibrase III and jararafîbrase IV, were purified from Bothrops jararaca venom using a fast protein liquid chromatography system. The purified jararafibrase III and jararafibrase IV were single chain proteins with molecular weights of 20,400 ± 500 and 21,200 ± 400, respectively, by SDS-PAGE. The isoelectric points of jararafibrase III and jararafibrase IV were 9.4 and 6.9, respectively. The activity of the enzyme was inhibited by 1,10-phenanthroline and EDTA, suggesting that both enzymes were metalloproteinases. The specific fibrinolytic activities of jararafibrase III and jararafibrase IV were 7.5 ±0.4 and 6.5 ±1.6 units/mg protein, respectively. The enzymes induced local hemorrhage in the skin of rats. The minimal hemorrhagic doses of jararafibrase III and IV were 31.0 and 34.0 μg/rat, respectively. The enzymes displayed broad substrate specificities like the previously purified jararafibrases I and II. Jararafibrases III and IV degraded type-IV collagen, gelatin, laminin and fibronectin into smaller fragments. The specific activities of jararafibrase III for type-IV collagen and gelatin were 7.6 ± 0.3 and 43 ±11 units/mg protein, respectively. The specific activities of jararafibrase IV foi type IV-collagen and gelatin were 16.5 ± 1.2 and 112 ± 9 units/ mg protein, respectively.

Modulation of Adhesion Molecules Expression by Different Metalloproteases Isolated from Bothrops Snakes

“…Several hemorrhagic metalloproteases have been isolated from Bothrops jararaca venom (BjV) [12][13][14]. One of the best characterized proteins is jararhagin (Jar), a protein with a molecular weight of 52 kDa that contains PI, metalloproteinase, ECD-disintegrin (ECD: Glu-Cys-Asp) and cysteine-rich domains [14], which are characteristics of an SVMP of the PIII class. In general, the toxins belonging to this class are highly hemorrhagic, and this activity depends on the metalloproteinase domain.…”