Puskar_2011_J.Biol.Chem_286_14618

Nicotinic acetylcholine receptors (nAChRs) are pentameric, neurotransmitter-gated ion channels responsible for rapid excitatory neurotransmission in the central and peripheral nervous systems, resulting in skeletal muscle tone and various cognitive effects in the brain. These complex proteins are activated by the endogenous neurotransmitter ACh as well as by nicotine and structurally related agonists. Activation and modulation of nAChRs has been implicated in the pathology of multiple neurological disorders, and as such, these proteins are established therapeutic targets. Here we use unnatural amino acid mutagenesis to examine the ligand binding mechanisms of two homologous neuronal nAChRs: the alpha4beta4 and alpha7 receptors. Despite sequence identity among the residues that form the core of the agonist-binding site, we find that the alpha4beta4 and alpha7 nAChRs employ different agonist-receptor binding interactions in this region. The alpha4beta4 receptor utilizes a strong cation-pi interaction to a conserved tryptophan (TrpB) of the receptor for both ACh and nicotine, and nicotine participates in a strong hydrogen bond with a backbone carbonyl contributed by TrpB. Interestingly, we find that the alpha7 receptor also employs a cation-pi interaction for ligand recognition, but the site has moved to a different aromatic amino acid of the agonist-binding site depending on the agonist. ACh participates in a cation-pi interaction with TyrA, whereas epibatidine participates in a cation-pi interaction with TyrC2.