Understanding the molecular mechanisms of agonist binding and agonist-dependent conformational changes during activation of neuronal nicotinic acetylcholine receptors (nAChRs) is essential for our understanding nicotine addiction, certain forms of epilepsy, Alzheimer's disease, and other neurological disorders. The research in this proposal focuses on alpha7 neuronal nAChRs because they are important in modulating neurotransmitter release. Specific residues that are important for ligand binding have been identified and characterized, but the conformational changes that couple agonist binding to channel gating are unclear. We use a homology model of the alpha7 receptors based on the crystal structure of the ACh Binding Protein to identify amino acids that may play important roles in agonist- or antagonist-dependent conformational changes. The specific goals of this research are to: (1) test the hypothesis that agonist binding causes a contraction or partial collapse of the ligand-binding pocket; (2) test the hypothesis that the beta9-beta10 hairpin structure, connecting the ligand-binding pocket to the transmembrane pore domain, undergoes conformational changes following the binding of agonists; and (3) determine whether lateral movement of the beta9-beta10 hairpin and/or rotational movements of subunits participate in agonist-driven activation. Several complementary experimental approaches will be employed. We will express alpha7 receptors in Xenopus oocytes and use electrophysiological approaches to probe functional properties. We will use the substituted cysteine accessibility method to identify residues that change accessibility during ligand binding. In addition, we will introduce electrostatic constraints and form disulfides or cysteine cross-links in (and near) the beta9-beta10 hairpin to limit movement and alter receptor activation.
| McLaughlin, James T; Barron, Sean C; See, Jennifer A et al. (2009) Conformational changes in alpha 7 acetylcholine receptors underlying allosteric modulation by divalent cations. BMC Pharmacol 9:1 |
| Barron, Sean C; McLaughlin, James T; See, Jennifer A et al. (2009) An allosteric modulator of alpha7 nicotinic receptors, N-(5-Chloro-2,4-dimethoxyphenyl)-N'-(5-methyl-3-isoxazolyl)-urea (PNU-120596), causes conformational changes in the extracellular ligand binding domain similar to those caused by acetylcholine. Mol Pharmacol 76:253-63 |
| McLaughlin, James T; Fu, Jie; Rosenberg, Robert L (2007) Agonist-driven conformational changes in the inner beta-sheet of alpha7 nicotinic receptors. Mol Pharmacol 71:1312-8 |
