Subunit structure of dihydropyridine-sensitive calcium channels from skeletal muscle

Abstract

Purified dihydropyridine-sensitive calcium channels from rabbit transverse-tubule membranes consist of three noncovalently associated classes of subunits: α (167 kDa), β (54 kDa), and γ (30 kDa). Cleavage of disulfide bonds reveals two distinct α polypeptides and an additional component, δ. The α1 subunit, a 175-kDa polypeptide that is not N-glycosylated, contains the dihydropyridine binding site, cAMP-dependent protein kinase phosphorylation site(s), and substantial hydrophobic domain(s). α2, a 143-kDa glycoprotein, has none of the properties characteristic of α1 but binds lectins and contains about 25% N-linked carbohydrate. α2 is disulfide-linked to δ, a 24- to 27-kDa glycopeptide. β (54 kDa) contains a cAMP-dependent phosphorylation site but is not N-glycosylated and does not have a hydrophobic domain. γ (30 kDa) has a carbohydrate content of about 30% and extensive hydrophobic domain(s). Precipitation with affinity-purified anti-α1 antibodies or α2-specific lentil lectin-agarose demonstrated that α1α2βγδ behaves as a complex in the presence of digitonin or 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate, whereas the α2δ complex dissociates from α1βγ in the presence of Triton X-100. A model for subunit interaction and membrane insertion is proposed on the basis of these observations.