title: The 165-kDa peptide of the purified skeletal muscle dihydropyridine receptor contains the known regulatory sites of the calcium channel.
abstract: The dihydropyridine receptor purified from rabbit skeletal muscle yields in the presence of dithiothreitol and sodium dodecyl sulfate on polyacrylamide gels bands of apparent molecular mass 165 +/- 5, 130 +/- 5, 55 +/- 3, 32 +/- 2 and 28 +/- 1 kDa (chi +/- SEM, n = 12). Under nonreducing conditions, the 130 kDa and 28-kDa peptides migrate as a single peptide of 165 kDa. These peptides were separated on a HPLC size-exclusion column. The specific absorption coefficients of the isolated peptides were determined. From these a stoichiometry of 1:1.7 +/- 0.2:1.4 +/- 0.3 (chi +/- SEM of 12 experiments with three different preparations) was calculated for the 165-kDa, 55-kDa and 32-kDa peptides. The relative amount of the 130/28-kDa peptide varied with different preparations. Tryptic, chymotryptic and V-8 protease peptides of the isolated proteins suggested that the 130/28-kDa peptide was not related to the 165-kDa peptide. The dihydropyridine photoaffinity analog (+/-)-azidopine was specifically incorporated only into the 165-kDa peptide with an efficiency of about 2.4%. The azido analog of desmethoxyverapamil, LU 49888, was specifically incorporated into the same peptide with an efficiency of 1.5%. These results suggest that only the 165-kDa peptide contains the regulatory sites detected so far in the voltage-operated L-type calcium channel. They suggest further that the 130/28-kDa peptide, which migrates as a 165-kDa peptide under nonreducing conditions, does not contain high-affinity binding sites for the calcium channel blockers.
mesh_terms: D000345:Affinity Labels / Q000378:metabolism; D000818:Animals; D001386:Azides / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels; D004095:Dihydropyridines; D007473:Ion Channels / Q000378:metabolism*; D017275:Isradipine; D007700:Kinetics; D008970:Molecular Weight; D009132:Muscles / Q000032:analysis*; D010069:Oxadiazoles / Q000378:metabolism; D011725:Pyridines / Q000378:metabolism; D011817:Rabbits; D011978:Receptors, Nicotinic / Q000032:analysis*; D014700:Verapamil / Q000031:analogs & derivatives / Q000378:metabolism
chemical_list: D000345:Affinity Labels; D001386:Azides; D015220:Calcium Channels; D004095:Dihydropyridines; D007473:Ion Channels; D010069:Oxadiazoles; D011725:Pyridines; D011978:Receptors, Nicotinic; C051537:LU 49888; C041130:azidopine; C043184:4-desmethoxyverapamil; D014700:Verapamil; D002118:Calcium; D017275:Isradipine