Synthetic peptides used to locate the .alpha.-bungarotoxin binding site and immunogenic regions on .alpha. subunits of the nicotinic acetylcholine receptor

Abstract

Synthetic peptides corresponding to 57% of the sequence of alpha subunits of acetylcholine receptors from Torpedo californica electric organ and extending from the NH2 to the COOCH terminus have been synthesized. The alpha-bungarotoxin binding site on denatured alpha subunits was mapped within the sequence alpha 185-199 by assaying binding of 125I-alpha-bungarotoxin to slot blots of synthetic peptides. Further studies showed that residues in the sequence alpha 190-194, especially cysteines-alpha 192, 193, were critical for binding alpha-bungarotoxin. Reduction and alkylation studies suggested that these cysteines must be disulfide linked for alpha-bungarotoxin to bind. Binding sites for serum antibodies to native receptors or alpha subunits were mapped by indirect immunoprecipitation of 125I-peptides. Several antigenic sequences were identified, but a synthetic peptide corresponding to the main immunogenic region (which is highly conformation dependent) was not identified.