ACETYLCHOLINE INTERACTIONS WITH TRYPTOPHAN-184 OF THE ALPHA-SUBUNIT OF THE NICOTINIC ACETYLCHOLINE-RECEPTOR REVEALED BY TRANSFERRED NUCLEAR OVERHAUSER EFFECT

Acetylcholine interactions with three genetically engineered fusion proteins containing peptides from the nicotinic acetylcholine receptor were studied by 1D and 2D nuclear magnetic resonance methods. The three proteins were Torpedo alpha-184-200, Torpedo alpha-186-198, and human alpha-183-204 of the acetylcholine receptor fused to the first 323 residues of the E. coli protein trpE. Nuclear Overhauser effect studies revealed interactions of bound acetylcholine with tryptophan-184 present in the Torpedo alpha-184-200, and the human alpha-183-204 sequences. These interactions are between the N(CH3)3+ and CH3 groups of acetylcholine with the aromatic protons of tryptophan. The appearance of these cross-peaks indicates a distance of less than 5 angstrom between tryptophan and the bound ligand; however, direct contact has yet to be proven.