Isolation and functional characterization of a new myotoxic acidic phospholipase A2 from Bothrops pauloensis snake venom

This article reports the purification procedure and the biochemical/functional characterization of Bp-PLA2, a new myotoxic acidic phospholipase A2 from Bothrops pauloensis snake venom. It was highly purified through three chromatographic steps (ion-exchange on CM-Sepharose, hydrophobic chromatography on Phenyl-Sepharose and RP-HPLC on a C8 column). Bp-PLA2 is a single-chain protein of 15.8 kDa and pI 4.3. Its N-terminal sequence revealed a high homology with other Asp49 acidic PLA2s from snake venoms. Its specific activity was 585.3 U/mg. It displayed a high indirect hemolytic activity and inhibited platelet aggregation induced by collagen or ADP. It also induced in vivo edema and myotoxicity. Pretreatment of Bp-PLA2 with BPB reduced the enzymatic activity, the inhibitory action on platelet aggregation and myotoxicity in vitro. Morphological analyses indicated that Bp-PLA2 induced an intense edema, with visible leukocyte infiltrate and damaged muscle cells 24 h after injection. Acidic myotoxic PLA2s from Bothrops snake venoms are still not extensively explored and knowledge of their structural and functional features will contribute for a better understanding of their action mechanism regarding enzymatic and toxic activities.