Purification and preliminary characterization of a frog-derived proteinaceous chemoattractant eliciting prey attack by checkered garter snakes (Thamnophis marcianus)

A potent proteinaceous chemoattractant, eliciting prey attack by checkered garter snakes (Thamnophis marcianus) was isolated from aqueous washes of the common frogRana temporaria and purified by preparative continuous-elution electrophoresis. The biological activity of the frog crude extract or of the purified chemoattractive protein, measured by a snake bioassay, was unaffected by freezing, lyophilization, or dialysis but was lost after proteolytic digestion. The purified chemoattractant is glycosylated, has an apparent molecular mass of 24 kDa, estimated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE), and a pI of 4.8. It gave one spot in two-dimensional electrophoresis. The bioassay showed that this protein is highly attractive to snakes. The lowest concentration yielding positive responses in the snake bioassay was approximately 25 µg/ml. These results suggest that a water-soluble Mr 24 kDa glycoprotein molecule produced by the common frog may be a vomeronasal stimulus used by checkered garter snakes for prey recognition.