Topographical studies of the nicotinic acetylcholine receptor. [Torpedo californica]
All four subunits of the nicotinic acetylcholine receptor in membrane vesicles isolated from Torpedo californica have been labeled with the photoactivated hydrophobic probe, (/sup 3/H)adamantanediazirine, which selectively labels regions of integral membrane proteins in contact with the hydrocarbon core of the lipid bilayer. All four subunits of the acetylcholine receptor in membrane vesicles isolated from Torpedo californica have been labeled with (/sup 3/H)cholesteryl diazoacetate. As this probe incorporates into lipid bilayers analogously to cholesterol, this result indicates that acetylcholine receptor interacts with cholesterol. Since the photogenerated carbene is situated near the lipid-water interface, this probe has potential as a topographic tool for mapping membrane protein structure. The labeling studies with both (/sup 3/H)adamantanediazirine and (/sup 3/H)cholesteryl diazoacetate support the concept that the acetylcholine receptor is a pseudosymmetric complex of homologous subunits, all of which interact with and span the membrane. The synthesis of the fluorine-containing agonists for the Torpedo californica nicotinic acetylcholine receptor, fluoroacetylcholine bromide and p-fluorophenyltrimethylammonium iodide, are described. It is demonstrated that both are agonists using a cation flux assay with acetylcholine receptor enriched membrane vesicles. The affinity cleavage reagent, p-thiocyanophenyltrimethylammonium iodide, specifically cleaves a peptide bond of the nicotinic acetylcholine receptor in membrane vesicles isolated from Torpedo californica. It is demonstrated that this reagent is an agonist using a cation flux assay. The cleavage is blocked by stoichiometric quantities of ..cap alpha..-bungarotoxin.
- Research Organization:
- California Inst. of Tech., Pasadena (USA)
- OSTI ID:
- 7140814
- Resource Relation:
- Other Information: Thesis (Ph. D.)
- Country of Publication:
- United States
- Language:
- English
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ACETYLCHOLINE
RECEPTORS
FISHES
LIPOSOMES
BIOCHEMICAL REACTION KINETICS
STRUCTURAL CHEMICAL ANALYSIS
BIOELECTRICITY
CARBENES
CATIONS
CHOLESTEROL
PHOTOCHEMICAL REACTIONS
PROTEINS
TRACER TECHNIQUES
TRITIUM COMPOUNDS
AMINES
AMMONIUM COMPOUNDS
ANIMALS
AQUATIC ORGANISMS
AUTONOMIC NERVOUS SYSTEM AGENTS
CELL CONSTITUENTS
CHARGED PARTICLES
CHEMICAL REACTIONS
DRUGS
ELECTRICITY
ESTERS
HYDROXY COMPOUNDS
IONS
ISOTOPE APPLICATIONS
KINETICS
LABELLED COMPOUNDS
MEMBRANE PROTEINS
NEUROREGULATORS
ORGANIC COMPOUNDS
ORGANOIDS
PARASYMPATHOMIMETICS
QUATERNARY COMPOUNDS
RADICALS
REACTION KINETICS
STEROIDS
STEROLS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques