Rattlesnake Neurotoxin Structure, Mechanism of Action, Immunology and MolecularBiology

摘要

This project was designed to provide a better understanding of rattlesnakeneurotoxin structure, mechanism of action, immunology and molecular biology. We demonstrated the structural and functional similarities of all rattlesnake neurotoxins, showed that the acidic subunit plays more than a chaperone role for the basic subunit and is clearly involved in the targeting of crotoxin. Sequencing studies have provided the primary sequence of the basic subunit of Mojave toxin, identified phospholipase A2-like molecules in Bothrops asper venom, identified isoforms of notexin, and expanded our phospholipase A2 sequence database. In studies on the mechanism of action we identified and partially characterized the binding properties of crotoxin receptors on brain synaptosomes. In collaborative experiments we examined crotoxin's neuromuscular blocking properties on isolated phrenic nerve-hemidiaphragm preparations under a variety of conditions and have preliminary evidence suggesting that crotoxin does not become internalized to exert its poisoning effects. Characterization of four different monoclonal antibodies raised against the basic subunit of crotoxin were examined for their cross-reactivity and shown to react with unique, continuous epitopes. Polyclonal antibodies raised to the same antigen indicated that four regions of the basic subunit were antigenic. In collaborative experiments we have successfully constructed both cDNA and genomic libraries of Crotalus scutulatus scutulatus. Acidic and basic subunit clones have been identified in both libraries. Both cDNA clones and the acidic subunit genomic clone have been sequenced.