Martin_1996_J.Biol.Chem_271_13497

The acetylcholine (ACh) receptors in muscle have the composition alpha2betagammadelta and contain two ACh binding sites. One is formed between an alpha subunit and the gamma subunit, and the other is formed between an alpha subunit and the delta subunit. Among the residues in the ACh binding sites are alphaCys-192 and alphaCys-193. The negatively charged deltaAsp-180 is at an appropriate distance from alphaCys-192/193 also to be in the ACh binding site and to interact electrostatically with the positively charged ammonium group common to agonists and competitive antagonists. Mutation to Asn of either deltaAsp-180 or the aligned residue in the gamma subunit, gammaAsp-174, decreased the affinities of three agonists, acetylcholine, tetramethylammonium, and succinyldicholine 170-560-fold. By contrast, these mutations decreased the affinities of three competitive antagonists, (+)-tubocurarine, hexamethonium, and dihydro-beta-erythroidine, only 2-15-fold. Agonists, but not antagonists, promote the transitions of the receptor from the resting state to the higher affinity active and desensitized states, and the greater effects of the mutations of gammaAsp-174 and deltaAsp-180 on the apparent affinities of agonists could reflect the involvement of these residues in the conformational changes of the receptor corresponding to its transitions to higher affinity states. In these transitions, one possibility is that gammaAsp-174 and deltaAsp-180 move closer to bound agonist.