Department of Pharmacology, University of Washington, Seattle, Washington 98195 Received February 9, 1987; Revised Manuscript Received March 31, 1987 abstract: Polyclonal antibodies (PAC-2) against the purified skeletal muscle calcium channel were prepared and shown to be directed against a subunits of this protein by immunoblotting and immunoprecipitation. These polypeptides have an apparent molecular weight of 162000 without reduction of disulfide bonds. Under conditions where the functional properties of the purified skeletal muscle calcium channel are retained,{3 subunits (Mr 50000) and 7 subunits (Mr 33000) are coprecipitated, demonstrating specific noncovalent association of these three polypeptides in the purified skeletal muscle channel. PAC-2 immunoprecipitated cardiac calcium channels labeled with [3H] isopropyl 4-(2, l, 3-benzoxadiazol-4-yl)-l, 4-dihydro-2, 6-dimethyl-5-(methoxycarbonyl) pyridine-3-carboxylate ([3H] PN200-110) at a 3-fold higher concentration than skeletal muscle channels. Preincubationwith cardiac calcium channels blocked only 49% of the immunoreactivity of PAC-2 toward skeletalmuscle channels, indicating that these two proteins have both ho-mologous and distinct epitopes. The immunoreactive component of the cardiac calcium channel was identified by immunoprecipitation and polyacrylamide gel electrophoresis as a polypeptide with an apparent molecular weight of 170000 before reduction of disulfide bonds and 141 000 after reduction, in close analogy with the properties of the a2 subunits of the skeletal muscle channel. It is concluded that these two calcium channels have a homologous, butdistinct, a subunit as a major polypeptide component.

Toltage-sensitive calcium channels play important roles in the regulation of the calcium-linked cellular functions including muscle contraction, neurotransmitter and hormone release, and calcium-dependent phosphorylation of intracellular proteins by controlling calcium influx from the extracellular environ-ment (Hagiwara & Byerly, 1981; Tsien, 1983). Dihydro-