The nicotinic acetylcholine receptor is the prototype ligand-gated ion channel. A number of aromatic amino acids have been identified as contributing to the agonist binding site, suggesting that cation-p interactions may be involved in binding the quaternary ammonium group of the agonist, acetylcholine. Here we show a compelling correlation between: (i) ab initio quantum mechanical predictions of cation-p binding abilities and (ii )E C 50 values for acetylcho- line at the receptor for a series of tryptophan derivatives that were incorporated into the receptor by using the in vivo nonsense-suppression method for unnatural amino acid in- corporation. Such a correlation is seen at one, and only one, of the aromatic residues—tryptophan-149 of the a subunit. This finding indicates that, on binding, the cationic, quater- nary ammonium group of acetylcholine makes van der Waals contact with the indole side chain of a tryptophan-149, providing the most precise structural information to date on this receptor. Consistent with this model, a tethered quater- nary ammonium group emanating from position a149 pro- duces a constitutively active receptor.