Structural and Functional Characterization of Crotoxin and its Basic Subunit in the Neuromuscular Junction and Influence of Neutralizing Agents

Abstract

The crotoxin is the main toxic fraction present in the venom of Crotalus durissus terrificus. This protein is a heterodimeric complex consisting of a basic phospholipase A2 (PLA2),crotoxin B (CB), associated with a chaperon and nonenzymatic subunit, crotapotina (CA). Pharmacological studies have revealed that the neuromuscular junction is a important site of action of this toxin, which causes muscle paralysis due to a sum of pre and postsynaptic effects. Recently, crystallographic studies have elucidated the three dimensional structure of the CB (from the venon of Crotalus durissus terrificus), which has advanced the current knowledge of the molecular toxic sites of this toxin. Unfortunately, despite the efforts devoted to this subject, the mechanism of action and the molecular site of action of crotoxin are unknown. Thus, the objective of this project is to identify potential inhibitors of crotoxin (or its subunit) and to elucidate the theoretical sites of interaction between the inhibitor and the CB. In addition, we will investigate the kinects of the neurotransmitter released under the influence of crotoxin or CB through the quantification of tritium-labeled with acetylcholine. We will also evaluate the real time exocytosis of the chemical mediator.