Purification and partial characterization of two phospholipases A2 from Bothrops leucurus (white-tailed-jararaca) snake venom

Abstract

Two proteins with phospholipase A(2) (PLA(2)) activity were purified to homogeneity from Bothrops leucurus (white-tailed-jararaca) snake venom through three chromatographic steps: Conventional gel filtration on Sephacryl S-200, ion-exchange on Q-Sepharose and reverse phase on Vydac C4 HPLC column. The molecular mass for both enzymes was estimated to be approximately 14 kDa by SDS-PAGE. The N-terminal sequences (48 residues) show that one enzyme presents lysine at position 48 and the other an aspartic acid in this position, and therefore they were designated blK-PLA(2) and blD-PLA(2) respectively. blK-PLA(2) presented negligible levels of PLA(2) activity as compared to that of blD-PLA(2). The PLA(2) activity of both enzymes is Ca(2+)-dependent. blD-PLA(2) did not have any effect upon platelet aggregation induced by arachidonic acid, ADP or collagen, but strongly inhibits coagulation and is able to stimulate Ehrlich tumor growth but not angiogenesis.