Stoichiometry of the Ligand-Binding Sites in the Acetylcholine-Receptor Oligomer from Muscle and from Electric Organ. Measurement by Affinity Alkylation with Bromoacetylcholine
- 1 August 1980
- journal article
- research article
- Published by Wiley in European Journal of Biochemistry
- Vol. 109 (2), 495-505
- https://doi.org/10.1111/j.1432-1033.1980.tb04821.x
Abstract
The affinity alkylation reaction of the cholinergic, depolarizing ligand, bromoacetylcholine with reduced acetylcholine receptor in the membrane fragments of Torpedo marmorata and in Triton-solubilized receptor from cat deneravated muscle was studied. Brief pretreatment with 100 .mu.M bromoacetylcholine abolishes all [3H].alpha.-neurotoxin binding in both cases. 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 are specifically alkylated by bromol[3H]acetylcholine, at saturation of either ligand. The concentration-dependence of specific bromo[3H]acetylcholine binding is biphasic. A 1st phase is clearly discerned in which 1/2 of the total specific ligand-binding sites are alkylated readily. In a 2nd phase, the remainder react at higher reagent concentrations. The same discrimination of 2 equal sets of ligand sites are obtained by preblockade using low concentrations of unlabeled bromoacetylcholine followed by reaction with [3H].alpha.-neurotoxin or bromo[3H]acetylcholine. In both phases, a single subunit of Mr about 43,000 is the sole site of specific alkylation in Torpedo and muscle. The reasons for the 2 equal but distinct populations in the ligand binding sites in the receptors are discussed.This publication has 29 references indexed in Scilit:
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