The purified Ca2+ antagonist receptor from skeletal muscle: subunit structure, photoaffinity labeling and endogenous protein kinase activity.
Tritiated analogues of the Ca2+ channel blockers such as [3H] PN200-110, [3H] verapamil and [3H] diltiazem have been used to identify and isolate Ca2+ antagonist receptors. The Ca2+ antagonist binding sites were solubilized from skeletal muscle transverse tubules with the detergent CHAPS and purified by wheat germ lectin column chromatography and sucrose density gradient centrifugation. The isolated proteins retained their ability to bind the various classes of Ca2+ channel blockers. Polypeptides of 170, 150, 108, 56, and 32 kDa were found to be present in the purified receptor fraction when analysed by sodium dodecyl sulfate polyacrylamide gel electrophoresis under non-reducing conditions. The apparent molecular weight of the 170 kDa polypeptide changed to 145 kDa in the presence of reducing agents, as where the apparent molecular weight of the 150, 108, 56 and 32 kDa peptides remained unchanged. An endogenous protein-kinase present in the original membranes, co-purified with the receptor and stimulated the phosphorylation of the 150 and 56 kDa polypeptides in the isolated fraction.
Tuana
B S
BS
Department of pharmacology, Faculty of Medicine, University of Toronto, Canada.
Murphy
B J
BJ
Yi
Q
Q
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000378:metabolism*; D015220:Calcium Channels / Q000032:analysis* / Q000187:drug effects / Q000378:metabolism; D002793:Cholic Acids; D004110:Diltiazem / Q000378:metabolism; D008970:Molecular Weight; D009132:Muscles / Q000378:metabolism*; D011494:Protein Kinases / Q000302:isolation & purification*; D011817:Rabbits; D011978:Receptors, Nicotinic / Q000378:metabolism*; D014700:Verapamil / Q000378:metabolism
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D002793:Cholic Acids; D011978:Receptors, Nicotinic; D014700:Verapamil; D011494:Protein Kinases; D004110:Diltiazem; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate; D002118:Calcium