The long term objective of the proposed research are to determine the three dimensional structural features of myotoxins from rattlesnake venoms and to understand how the secondary and tertiary structural features relate to biological activity of this unique group of toxins at the biochemical and physiological levels.
One specific aim i s to derive a three dimensional structure that can be used as a working model for this group of toxins. This will be done by using high resolution nuclear magnetic resonance data and appropriate computer algorithms to predict spatial arrangements from the amino acids that constitute the primary structure. A second specific aim is to discover which amino acid side chain residues are mandatory for biological activity. This problem will be addressed by specific chemical modification of reactive, highly conserved amino acid residues in the myotoxin sequence and subsequent assays of biological activities. A third specific goal is to elucidate the biochemical basis for the adverse effects that result when muscle cells are treated with myotoxin. This will be accomplished by measuring changes in membrane metabolic events and ion channels using radioisotopes and patch clamp techniques. The binding of radiolabelled toxin will also be used to explore interactions between myotoxins and cell membranes. The fourth specific aim pertains to the relatedness of myotoxins obtained from venoms of different species of rattlesnake. Hybridoma cell liens will be developed to produce groups of monoclonal antibodies that are specific for epitopes in the myotoxin structures. Surface characteristics of the five myotoxins of known sequence will be explored with selected monoclonal antibodies. The health related aspects of the research derive from the fact that these toxins are present in rattlesnake venoms and cause adverse effects in the envenomated host, particularity on muscle tissue. This basic research effort will lead to a much better understanding of toxin induced damage to membrane properties and may provide a useful model for degenerative muscle disease. The monoclonal antibodies could become useful diagnostic tools for screening rattlesnake venoms and have the potential to become therapeutic agents.
