Identification and characterization of the protein components of the skeletal muscle receptor for the 1,4-dihydropyridine Ca sup 2+ channel blockers

Thesis/Dissertation · Fri Jan 01 04:00:00 EST 1988

OSTI ID:6948601

Sharp, A H

In these studies, photoaffinity labeling and immunolabeling approaches were used to identify and characterize components of the skeletal muscle receptor for the 1,4-dihydropyridine Ca{sup 2+} channel blockers. The 1,4-dihydropyridine receptor purified from rabbit skeletal muscle consists of proteins of 175,000, 170,000, 52,000, and 32,000 Da when analyzed by SDS-PAGE under nonreducing conditions and stained with Coomassie Blue dye. After reduction of disulfide bonds, the 175,000 Da protein shifts in apparent molecular mass to 150,000 Da. Photoaffinity labeling using the dihydropyridine ligands ({sup 3}H)azidopine and ({sup 3}H)PN200-110 identified a protein of 170,000 Da as the dihydropyridine binding component of the receptor. Specific polyclonal antibodies were developed against both the nonreduced and reduced forms of the 175/150,000 and 32,000 Da proteins and were used to show that the 150,000 and 32,000 Da proteins are distinct from each other and from other components of the receptor and that they copurify with the 170,000 Da protein at each step of purification. In addition, monoclonal antibodies against the 170,000 and 52,000 Da polypeptides were shown to coimmunoprecipitate the 150,000 and 32,000 Da polypeptides from solubilized skeletal muscle triads.

• Research Organization: Iowa Univ., Iowa City, IA (USA)
• OSTI ID: 6948601
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ALKALINE EARTH METAL COMPOUNDS
ANIMALS
ANTIBODIES
CALCIUM COMPOUNDS
CHEMICAL COMPOSITION
ENZYME INHIBITORS
FRACTIONATION
HYDROGEN COMPOUNDS
ISOTOPE APPLICATIONS
LABELLING
MAMMALS
MEMBRANE PROTEINS
MOLECULAR WEIGHT
MUSCLES
ORGANIC COMPOUNDS
PORINS
PROTEINS
RABBITS
RECEPTORS
SEPARATION PROCESSES
TRACER TECHNIQUES
TRITIUM COMPOUNDS
VERTEBRATES