Evidence for structural dissimilarity in the neurotransmitter binding region of purified acetylcholine receptors from human muscle and Torpedo electric organ.
Acetylcholine receptor (AChR) purified from human skeletal muscle affinity-alkylated with bromoacetyl[methyl-3H]choline bromide ([3H]BAC) in mildly reducing conditions to yield a specifically radiolabeled polypeptide, Mr 44,000, the alpha-subunit. The binding of [125I]alpha-bungarotoxin to AChR was completely inhibited by affinity-alkylation, indicating that the human AChR's binding site for alpha-bungarotoxin is closely associated with the alpha-subunit's acetylcholine binding site. Structures in the vicinity of the alpha-bungarotoxin binding sites of AChRs from human muscle and Torpedo electric organ were compared by varying the conditions of alkylation. Under optimal conditions of reduction and alkylation, both human and Torpedo AChR incorporated BAC in equivalence to the number of alpha-bungarotoxin binding sites. However, with limited conditions of reduction but sufficient BAC to alkylate 100% of the alpha-bungarotoxin binding sites of human AChR, only 71% of the Torpedo AChR's binding sites were alkylated. In optimal conditions of reduction but with the minimal concentration of BAC that permitted 100% alkylation of the human AChR's alpha-bungarotoxin sites, only 74% of the Torpedo AChR's binding sites were alkylated. These data suggest that the neurotransmitter binding region of human muscle AChR is structurally dissimilar from that of Torpedo electric organ, having a higher binding affinity for BAC and an adjacent disulfide bond that is more readily accessible to reducing agents.
D000109:Acetylcholine / Q000031:analogs & derivatives / Q000378:metabolism; D000478:Alkylation; D000818:Animals; D001665:Binding Sites; D002038:Bungarotoxins / Q000378:metabolism; D004229:Dithiothreitol / Q000378:metabolism; D004557:Electric Organ / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D009132:Muscles / Q000378:metabolism*; D010084:Oxidation-Reduction; D051381:Rats; D011950:Receptors, Cholinergic / Q000378:metabolism*; D014101:Torpedo
D002038:Bungarotoxins; D011950:Receptors, Cholinergic; D000109:Acetylcholine; D004229:Dithiothreitol