Identification of two distinct proteins that are immunologically related to the alpha 1 subunit of the skeletal muscle dihydropyridine-sensitive calcium channel.
The alpha 1 subunit of the dihydropyridine-sensitive calcium channel is a protein which is critical for excitation-contraction coupling and L-type calcium current in skeletal muscle. Using antibodies generated against peptides from three regions of the deduced amino acid sequence of the alpha 1 subunit, we have identified two distinct proteins in rabbit skeletal muscle. Both proteins appeared to be recognized by antibodies against the amino (N) terminus of the alpha 1 subunit sequence. One protein was also recognized by antibodies against an internal (I) region of the predicted sequence but not by antibodies against the carboxyl (C) terminus. In contrast, the other protein was recognized by antibodies against the carboxyl terminus but not by the antibodies against the internal region. We have designated these proteins pNI and pNC based on their patterns of antibody recognition. No protein was detected which was recognized by all three antibodies. pNI is the protein commonly identified as the alpha 1 subunit of the dihydropyridine-sensitive calcium channel. Of note is that pNI, which apparently lacks sequences from the predicted carboxyl tail, is the protein present in preparations which we have previously demonstrated contain dihydropyridine-sensitive calcium channel activity. pNC is herein identified as a skeletal muscle protein that is immunologically related to the alpha 1 subunit of the dihydropyridine-sensitive calcium channel. Its function is unknown. In addition to their distinct patterns of antibody recognition, pNI and pNC were also distinguishable by several other properties. pNC migrated as a protein of approximately 160 kDa in 5% sodium dodecyl sulfate-polyacrylamide gels versus approximately 165 kDa for pNI. pNI was enriched in transverse tubule membranes, whereas pNC was found to be enriched in triad and junctional sarcoplasmic reticulum membrane fractions and was not found in transverse tubule membranes. Under conditions in which pNI bound to wheat germ agglutinin-Sepharose, pNC did not bind. The results demonstrate that there are two proteins in skeletal muscle which are immunologically related to the alpha 1 subunit of the dihydropyridine-sensitive calcium channel but which are distinguishable by several biochemical and immunological characteristics.
Brawley
R M
RM
Department of Pharmacology, Northwestern University Medical School, Chicago, Illinois 60611.
Hosey
M M
MM
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D015220:Calcium Channels / Q000737:chemistry* / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D002846:Chromatography, Affinity; D004095:Dihydropyridines / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D015151:Immunoblotting; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification; D008961:Models, Structural; D008969:Molecular Sequence Data; D009132:Muscles / Q000378:metabolism*; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D011487:Protein Conformation; D011817:Rabbits; D012689:Sequence Homology, Nucleic Acid
D000906:Antibodies; D015220:Calcium Channels; D004095:Dihydropyridines; D046911:Macromolecular Substances; D008565:Membrane Proteins; D010455:Peptides