Ligand-receptor interactions in the nicotinic acetylcholine receptor probed using multiple substitutions at conserved tyrosines on the alpha subunit.

Affinity labeling studies have identified several conserved tyrosine residues in the alpha subunit of the nicotinic acetylcholine receptor (alpha Y93, alpha Y190, and alpha Y198) as being in or near the ligand binding site. Mutagenesis studies from several laboratories have shown that substitution of phenylalanine for tyrosine at these positions reduces the apparent affinity for ACh. We have examined this apparent reduction in affinity further through the use of multiple substitutions at each position. Substitution of either phenylalanine, tryptophan, or serine resulted in an apparent decrease in agonist affinity, but the degree of reduction depended on both the position and the nature of the substitution. Analysis of the effects of each substitution suggests that each residue interacts with the quaternary N of ACh, and that each residue may make a different type of interaction with the agonist.