Solubilization of the omega-conotoxin receptor associated with voltage-sensitive calcium channels from bovine brain.
The omega-conotoxin receptor in brain membranes contains components of Mr approximately equal to 310,000, approximately equal to 230,000, and 37,000 as identified by photoaffinity labeling. The toxin specifically bound to two sites with apparent dissociation constants (Kd) of approximately 3 pM and 3.5 nM under the conditions employed. There was about 8 times more of the low affinity site than the high affinity site. Binding was not affected by dihydropyridines or verapamil. However, diltiazem stereospecifically inhibited the binding to the high affinity site. Dissociation of the toxin from the membranes was very slow and only partial. Among the detergents tested, digitonin solubilized the highest toxin-binding activity. The digitonin extract contained only a single class of binding sites with an apparent Kd of about 0.46 nM. Probably only the high affinity binding site was recovered in active form in digitonin extract. The properties of the toxin binding to digitonin extract were in good agreement with those of the binding to the high affinity site in the original membranes. Photoaffinity labeling of the digitonin extract indicated that the solubilized toxin receptor contained the two large components (Mr congruent 310,000 and approximately equal to 230,000) observed in the membranes.
Yamaguchi
T
T
Department of Neurochemistry, Niigata University, Japan.
Saisu
H
H
Mitsui
H
H
Abe
T
T
D000818:Animals; D001921:Brain / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels; D002417:Cattle; D002499:Centrifugation, Density Gradient; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D007473:Ion Channels / Q000378:metabolism*; D007700:Kinetics; D008970:Molecular Weight; D017981:Receptors, Neurotransmitter / Q000302:isolation & purification / Q000378:metabolism*; D012995:Solubility
D015220:Calcium Channels; D007473:Ion Channels; D017981:Receptors, Neurotransmitter; C053157:omega-conotoxin receptor; D002118:Calcium