Stoichiometry of the ligand-binding sites in the acetylcholine-receptor oligomer from muscle and from electric organ. Measurement by affinity alkylation with bromoacetylcholine.
1. The affinity alkylation reaction of the cholinergic, depolarising ligand, bromoacetylcholine with reduced acetylcholine receptor in the membrane fragments of Torpedo marmorata and in Triton-solubilised receptor from cat denervated muscle has been studied. 2. Brief pretreatment with 100 microM bromoacetylcholine abolishes all [3H]alpha-neurotoxin binding in both cases. 3. In the receptor from each of these sources, the number of sites of specific alpha-neurotoxin binding is exactly equal to the number of sites that can be specifically alkylated by bromol[3H]acetytlcholine, at saturation of either ligand. 4. The concentration-dependence of specific bromo[3H]acetylcholine binding is found to be biphasic. A first phase can be clearly discerned in which one-half of the total specific ligand-binding sites are alkylated readily, and a second phase in which the remainder react at higher reagent concentrations. The same discrimination of two equal sets of ligand sites can be obtained by pre-blockade using low concentrations of unlabelled bromoacetylcholine followed by reaction with [3H]alpha-neurotoxin or bromo[3H]acetylcholine. 5. In both phases, a single subunit of Mr about 43000 is the sole site of specific alkylation in both Torpedo and muscle. The reasons for the appearance of two equal but distinct populations in the ligand binding sites in the receptors are discussed.
D000109:Acetylcholine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000378:metabolism; D000478:Alkylation; D000818:Animals; D004557:Electric Organ / Q000378:metabolism*; D005399:Fishes; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009132:Muscles / Q000378:metabolism*; D011950:Receptors, Cholinergic / Q000378:metabolism*
D046911:Macromolecular Substances; D011950:Receptors, Cholinergic; D000109:Acetylcholine