A new scorpion toxin (BmK-PL) stimulates Ca2+-release channel activity of the skeletal-muscle ryanodine receptor by an indirect mechanism

A peptide toxin isolated from the Chinese scorpion Buthus martensi Karsch ( BmK-PL) stimulated Ca2+-release channel activity in both triad membranes an d reconstituted ryanodine receptors partially purified from rabbit skeletal muscle. In [H-3]ryanodine binding experiments, the toxin increased the aff inity of ryanodine for the receptor, from a K-d of 24.3 nM to 2.9 nM, which is an enhancement similar to that seen with known receptor activators, suc h as ATP and high concentrations of KCl. In contrast, toxin enhancement was not observed with purified receptors, although intrinsic binding activity and stimulation by the conventional receptor activators were retained. In s ingle channel recordings of Ca2+-release activity, the toxin increased the open channel probability (P-o) from 0.019 to 0.043 (226% of control) in tri ad preparations. Further toxin enhancement of P-o from 0.07 to 0.37 (529% o f control) was observed using partially-purified receptors in the presence of ATP. When purified receptors were assayed in the presence of ATP, howeve r, they showed a high value of P-o (0.33) and no further increase was obser ved following application of the toxin, Results derived from two different experimental methods consistently suggest that a molecule(s) required for t oxin-induced enhancement is absent from the purified receptor preparation. Western blot analysis of receptors prepared using three different protocols showed that triadin was missing from the purified receptor preparation. Th e scorpion toxin minimally enhanced Ca2+-release channel activity of cardia c preparations. From these results, we conclude that the toxin preferential ly increases the activity of skeletal-muscle ryanodine receptors by an indi rect mechanism, possibly binding to associated protein molecule(s). Triadin is a strong candidate for such a molecule.