Purification and characterization of 94kDa and 80kDa forms of the muscarinic cholinergic receptor
Two molecular forms of the muscarinic cholinergic receptor have been consistently observed in a variety of species, albeit in variable amounts. Proteins which are specifically labeled by (/sup 3/H)propylbenzilylcholine mustard ((/sup 3/H)PrBCM) were observed at 94kDa and 80kDa upon SDS-PAGE of membrane proteins prepared from brains and hearts of trout, frog, turtle, chicken, rat, and pig. They have developed a purification procedure which yields each of these proteins in a homogeneous form suitable for structural analysis. The four step procedure involves affinity chromatography on 3-(2'-aminobenzhydryloxy)tropane-sepharose, concentration on hydroxylapatite, preparative SDS-PAGE and extraction of individual bands from the gel. Limited tryptic digestion of purified (/sup 3/H)PrBCM-labeled porcine atrial muscarinic receptor yields (/sup 3/H)-labeled fragments of 75, 65, 52, 40, 35, 30, 25, and 20kDa, in close agreement with results of analogous digestions of muscarinic receptor from other species and tissues. Complete tryptic digestion and subsequent mapping by reverse-phase HPLC yields very similar profiles for (/sup 125/I)-labeled 94kDa and 80kDA receptor forms. Most peaks which elute in the hydrophobic region of the profile overlap for the two proteins while the 94kDa protein contains several additional peaks of apparent low hydrophobicity.
- Research Organization:
- National Institute of Health, Bethesda, MD
- OSTI ID:
- 5020666
- Report Number(s):
- CONF-8606151-
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Vol. 45:6; Conference: 76. annual meeting of the Federation of American Society for Experimental Biology, Washington, DC, USA, 8 Jun 1986
- Country of Publication:
- United States
- Language:
- English
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PROTEINS
RECEPTORS
CHEMICAL COMPOSITION
PURIFICATION
ACETYLCHOLINE
BRAIN
CELL MEMBRANES
CHICKENS
CHROMATOGRAPHY
FROGS
HEART
IODINE 125
RATS
SWINE
TRACER TECHNIQUES
TRITIUM COMPOUNDS
TROUT
AMINES
AMMONIUM COMPOUNDS
AMPHIBIANS
ANIMALS
AQUATIC ORGANISMS
AUTONOMIC NERVOUS SYSTEM AGENTS
BETA DECAY RADIOISOTOPES
BIRDS
BODY
CARDIOVASCULAR SYSTEM
CELL CONSTITUENTS
CENTRAL NERVOUS SYSTEM
DAYS LIVING RADIOISOTOPES
DOMESTIC ANIMALS
DRUGS
ELECTRON CAPTURE RADIOISOTOPES
ESTERS
FISHES
FOWL
INTERMEDIATE MASS NUCLEI
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
LABELLED COMPOUNDS
MAMMALS
MEMBRANE PROTEINS
MEMBRANES
NERVOUS SYSTEM
NEUROREGULATORS
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANS
PARASYMPATHOMIMETICS
QUATERNARY COMPOUNDS
RADIOISOTOPES
RODENTS
SEPARATION PROCESSES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques