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 and reconstituted ryanodine receptors partially purified from rabbit skeletal muscle. In [3H]ryanodine binding experiments, the toxin increased the affinity of ryanodine for the receptor, from a Kd of 24.3 nM to 2.9 nM, which is an enhancement similar to that seen with known receptor activators, such 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 single channel recordings of Ca2+-release activity, the toxin increased the open channel probability (Po) from 0.019 to 0.043 (226% of control) in triad preparations. Further toxin enhancement of Po from 0.07 to 0.37 (529% of control) was observed using partially-purified receptors in the presence of ATP. When purified receptors were assayed in the presence of ATP, however, they showed a high value of Po (0.33) and no further increase was observed following application of the toxin. Results derived from two different experimental methods consistently suggest that a molecule(s) required for toxin-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. The scorpion toxin minimally enhanced Ca2+-release channel activity of cardiac preparations. From these results, we conclude that the toxin preferentially increases the activity of skeletal-muscle ryanodine receptors by an indirect mechanism, possibly binding to associated protein molecule(s). Triadin is a strong candidate for such a molecule.
Kuniyasu
A
A
Faculty of Pharmaceutical Sciences, Kumamoto University, 5-1 Ohe-Honmachi, Kumamoto 862-0973, Japan.
Kawano
S
S
Hirayama
Y
Y
Ji
Y H
YH
Xu
K
K
Ohkura
M
M
Furukawa
K
K
Ohizumi
Y
Y
Hiraoka
M
M
Nakayama
H
H
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002120:Calcium Channel Agonists / Q000737:chemistry / Q000494:pharmacology*; D008051:Lipid Bilayers; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000187:drug effects* / Q000378:metabolism; D011817:Rabbits; D012433:Ryanodine / Q000378:metabolism; D019837:Ryanodine Receptor Calcium Release Channel / Q000187:drug effects* / Q000302:isolation & purification / Q000378:metabolism; D012604:Scorpion Venoms / Q000737:chemistry*
D002120:Calcium Channel Agonists; D008051:Lipid Bilayers; D019837:Ryanodine Receptor Calcium Release Channel; D012604:Scorpion Venoms; D012433:Ryanodine