title: Varenicline Interactions at the 5-HT3 Receptor Ligand Binding Site are Revealed by 5-HTBP.
abstract: Cys-loop receptors are the site of action of many therapeutic drugs. One of these is the smoking cessation agent varenicline, which has its major therapeutic effects at nicotinic acetylcholine (nACh) receptors but also acts at 5-HT3 receptors. Here, we report the X-ray crystal structure of the 5-HT binding protein (5-HTBP) in complex with varenicline, and test the predicted interactions by probing the potency of varenicline in a range of mutant 5-HT3 receptors expressed in HEK293 cells and Xenopus oocytes. The structure reveals a range of interactions between varenicline and 5-HTBP. We identified residues within 5 Å of varenicline and substituted the equivalent residues in the 5-HT3 receptor with Ala or a residue with similar chemical properties. Functional characterization of these mutant 5-HT3 receptors, using a fluorescent membrane potential dye in HEK cells and voltage clamp in oocytes, supports interactions between varenicline and the receptor that are similar to those in 5-HTBP. The structure also revealed C-loop closure that was less than in the 5-HT-bound 5-HTBP, and hydrogen bonding between varenicline and the complementary face of the binding pocket via a water molecule, which are characteristics consistent with partial agonist behavior of varenicline in the 5-HT3 receptor. Together, these data reveal detailed insights into the molecular interaction of varenicline in the 5-HT3 receptor.
authors: †Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QW, United Kingdom.
authors: Kerry L
authors: KL
authors: Price
authors: †Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QW, United Kingdom.
authors: Reidun K
authors: RK
authors: Lillestol
authors: ‡The Laboratory of Structural Neurobiology, Department of Cellular and Molecular Medicine, KU Leuven, Herestraat 49, PB 601, B-3000 Leuven, Belgium.
authors: Chris
authors: C
authors: Ulens
authors: †Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QW, United Kingdom.
authors: Sarah C R
authors: SC
authors: Lummis
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D057809:HEK293 Cells; D006801:Humans; D006860:Hydrogen Bonding; D051379:Mice; D008958:Models, Molecular; D009154:Mutation; D009865:Oocytes; D018408:Patch-Clamp Techniques; D017433:Protein Structure, Secondary; D044406:Receptors, Serotonin, 5-HT3 / Q000235:genetics / Q000378:metabolism*; D012701:Serotonin / Q000378:metabolism; D018490:Serotonin Agents / Q000378:metabolism* / Q000494:pharmacology; D000068580:Varenicline / Q000378:metabolism* / Q000494:pharmacology; D014867:Water / Q000378:metabolism; D014981:Xenopus
chemical_list: D002352:Carrier Proteins; C511230:Htr3a protein, mouse; D044406:Receptors, Serotonin, 5-HT3; D018490:Serotonin Agents; D014867:Water; D012701:Serotonin; D000068580:Varenicline
keywords: Cys-loop receptor; Ligand-gated ion channel; serotonin receptor
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