title: Unorthodox Acetylcholine Binding Sites Formed by α5 and β3 Accessory Subunits in α4β2* Nicotinic Acetylcholine Receptors.
abstract: All nicotinic acetylcholine receptors (nAChRs) evolved from homomeric nAChRs in which all five subunits are involved in forming acetylcholine (ACh) binding sites at their interfaces. Heteromeric α4β2* nAChRs typically have two ACh binding sites at α4/β2 interfaces and a fifth accessory subunit surrounding the central cation channel. β2 accessory subunits do not form ACh binding sites, but α4 accessory subunits do at the α4/α4 interface in (α4β2)2α4 nAChRs. α5 and β3 are closely related subunits that had been thought to act only as accessory subunits and not take part in forming ACh binding sites. The effect of agonists at various subunit interfaces was determined by blocking homologous sites at these interfaces using the thioreactive agent 2-((trimethylammonium)ethyl) methanethiosulfonate (MTSET). We found that α5/α4 and β3/α4 interfaces formed ACh binding sites in (α4β2)2α5 and (α4β2)2β3 nAChRs. The α4/α5 interface in (β2α4)2α5 nAChRs also formed an ACh binding site. Blocking of these sites with MTSET reduced the maximal ACh evoked responses of these nAChRs by 30-50%. However, site-selective agonists NS9283 (for the α4/α4 site) and sazetidine-A (for the α4/β2 site) did not act on the ACh sites formed by the α5/α4 or β3/α4 interfaces. This suggests that unorthodox sites formed by α5 and β3 subunits have unique ligand selectivity. Agonists or antagonists for these unorthodox sites might be selective and effective drugs for modulating nAChR function to treat nicotine addiction and other disorders.
authors.affiliation: From the Department of Neuroscience, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Akansha
authors.initials: A
authors.lastname: Jain
authors.affiliation: From the Department of Neuroscience, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Alexander
authors.initials: A
authors.lastname: Kuryatov
authors.affiliation: the Division of Pharmacology and Toxicology, College of Pharmacy, The University of Texas at Austin, Austin, Texas 78712, and.
authors.forename: Jingyi
authors.initials: J
authors.lastname: Wang
authors.affiliation: the Department of Molecular Therapeutics, Scripps Research Institute, Jupiter, Florida 33458.
authors.forename: Theodore M
authors.initials: TM
authors.lastname: Kamenecka
authors.affiliation: From the Department of Neuroscience, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania 19104, JSLKK@mail.med.upenn.edu.
authors.forename: Jon
authors.initials: J
authors.lastname: Lindstrom
mesh_terms: D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D006801:Humans; D055503:Protein Multimerization; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D011978:Receptors, Nicotinic / Q000737:chemistry / Q000378:metabolism*; D014981:Xenopus
chemical_list: C539371:CHRNB3 protein, human; D021122:Protein Subunits; D011978:Receptors, Nicotinic; C403531:nicotinic receptor alpha4beta2; C577601:nicotinic receptor alpha5 subunit, human; D000109:Acetylcholine
keywords: Xenopus; channel activation; electrophysiology; ion channel; nicotinic acetylcholine receptors (nAChR); site-directed mutagenesis