Biochemical characterization of two crotamine isoforms isolated by a single step RP-HPLC from Crotalus durissus terrificus (South American rattlesnake) venom and their action on insulin secretion by pancreatic islets

Crotamine. a neurotoxin present in the venom of the South American rattlesnake Crotalus durrisus terrificus exists as several polymorphic variants, as demonstrated by recombinant DNA technology (Smith and Schmidt, Toxicon 28 (1990) 575-585). We have isolated native crotamine by chromatography on Sephadex G75, and have purified two crotamine isoforms (F2 and F3) by a single step of RP-HPLC. Native crotamine and RP-HPLC fractions F2 and F3 produced skeletal muscle spasms and spastic paralysis in mice. At low glucose concentrations (2.8-5.6 mmol/l), none of the crotamines altered the insulin secretion by rat isolated islets. In the presence of 16.7 mmol glucose/l, F2 (5 mu g/ml), but not F3, increased insulin secretion two-fold, whereas native crotamine (1.5, 5 and 16.5 mu g/ml) potentiated the secretion dose-dependently. The increase in insulin secretion induced by F2 fraction (5 mu g/ml) was similar to that obtained with 16.5 mu g of native crotamine/ml. These results indicate that the mode of action of the F2 and F3 isoforms in beta-cells is different from that in muscle cells. This difference may be related to the binding affinity of each isoform for the Na+ channels located in the beta-cell membrane. Crotamine isoforms may be valuable tools for studying the involvement of Na+ channels in the mechanism of insulin secretion. (C) 2000 Elsevier Science B.V. All rights reserved.