title: Monoclonal antibodies that coimmunoprecipitate the 1,4-dihydropyridine and phenylalkylamine receptors and reveal the Ca2+ channel structure.
abstract: Monoclonal hybridoma cell lines secreting antibodies against the (+)-PN 200-110 and the (-)-demethoxyverapamil binding components of the voltage-dependent calcium channel from rabbit transverse-tubule membranes have been isolated. The specificity of these monoclonal antibodies was established by their ability to coimmunoprecipitate (+)-[3H]PN 200-110 and (-)-[3H]demethoxyverapamil receptors. Monoclonal antibodies described in this work cross-reacted with rat, mouse, chicken, and frog skeletal muscle Ca2+ channels but not with crayfish muscle Ca2+ channels. Cross-reactivity was also detected with membranes prepared from rabbit heart, brain, and intestinal smooth muscle. These antibodies were used in immunoprecipitation experiments with 125I-labeled detergent [3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) and digitonin] solubilized membranes. They revealed a single immunoprecipitating component of molecular weight (Mr) 170,000 in nonreducing conditions. After disulfide bridge reduction the CHAPS-solubilized (+)-PN 200-110-(-)-demethoxyverapamil binding component gave rise to a large peptide of Mr 140,000 and to smaller polypeptides of Mr 30,000 and 26,000 whereas the digitonin-solubilized receptor appeared with subunits at Mr 170,000, 140,000, 30,000, and 26,000. All these results taken together are interpreted as showing that both the 1,4-dihydropyridine and the phenylalkylamine receptors are part of a single polypeptide chain of Mr 170,000.
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000378:metabolism*; D015220:Calcium Channels; D007473:Ion Channels / Q000378:metabolism*; D007700:Kinetics; D008870:Microtubules / Q000378:metabolism; D008970:Molecular Weight; D009132:Muscles / Q000378:metabolism*; D010449:Peptide Mapping; D011817:Rabbits; D011978:Receptors, Nicotinic / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D007473:Ion Channels; D011978:Receptors, Nicotinic; D002118:Calcium