Biochemical and ultrastructural characterization of the 1,4-dihydropyridine receptor from rabbit skeletal muscle. Evidence for a 52,000 Da subunit.
The 1,4-dihydropyridine receptor purified from rabbit skeletal muscle contains four polypeptide components of 175,000 Da (nonreduced)/150,000 Da (reduced), 170,000, 52,000, and 32,000 Da (Leung, A. T., Imagawa, T., and Campbell, K. P. (1987) J. Biol. Chem. 262, 7943-7946). A monoclonal antibody specific to the 52,000-Da polypeptide component of the dihydropyridine receptor has been produced and used in immunoprecipitation and immunoblotting experiments to demonstrate that the 52,000-Da polypeptide is an integral subunit of the purified dihydropyridine receptor. Peptide mapping experiments with 32P-labeled dihydropyridine receptor have also demonstrated that the 52,000-Da polypeptide is distinct from and not a proteolytic fragment of the 170,000-Da subunit. Densitometric scanning of Coomassie Blue-stained sodium dodecyl sulfate-polyacrylamide gels of the purified dihydropyridine receptor has demonstrated that the 52,000-Da polypeptide exists in a 1:1 stoichiometric ratio with the 170,000-, 175,000/150,000-, and 32,000-Da subunits of the dihydropyridine receptor. Electron microscopy of the freeze-dried, rotary-shadowed dihydropyridine receptor has shown that the preparation contains a homogeneous population of 16 x 22-nm ovoidal particles large enough to contain all four polypeptides of the dihydropyridine receptor. The particles have two distinct components of similar size which may represent the location in the molecule of the two larger subunits.
Leung
A T
AT
Department of Physiology and Biophysics, University of Iowa, Iowa City 52242.
Imagawa
T
T
Block
B
B
Franzini-Armstrong
C
C
Campbell
K P
KP
D000818:Animals; D000911:Antibodies, Monoclonal; D015220:Calcium Channels; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008970:Molecular Weight; D009132:Muscles / Q000378:metabolism*; D010449:Peptide Mapping; D010766:Phosphorylation; D011817:Rabbits; D011978:Receptors, Nicotinic / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D015220:Calcium Channels; D046911:Macromolecular Substances; D011978:Receptors, Nicotinic