title: Characterization of a ligand binding site in the human transient receptor potential ankyrin 1 pore.
abstract: The pharmacology and regulation of Transient Receptor Potential Ankyrin 1 (TRPA1) ion channel activity is intricate due to the physiological function as an integrator of multiple chemical, mechanical, and temperature stimuli as well as differences in species pharmacology. In this study, we describe and compare the current inhibition efficacy of human TRPA1 on three different TRPA1 antagonists. We used a homology model of TRPA1 based on Kv1.2 to select pore vestibule residues available for interaction with ligands entering the vestibule. Site-directed mutation constructs were expressed in Xenopus oocytes and their functionality and pharmacology assessed to support and improve our homology model. Based on the functional pharmacology results we propose an antagonist-binding site in the vestibule of the TRPA1 ion channel. We use the results to describe the proposed intravestibular ligand-binding site in TRPA1 in detail. Based on the single site substitutions, we designed a human TRPA1 receptor by substituting several residues in the vestibule and adjacent regions from the rat receptor to address and explain observed species pharmacology differences. In parallel, the lack of effect on HC-030031 inhibition by the vestibule substitutions suggests that this molecule interacts with TRPA1 via a binding site not situated in the vestibule.
authors: Neuroscience Department, CNS & Pain Innovative Medicines, AstraZeneca R&D Södertälje, Södertälje, Sweden.
authors: Göran
authors: G
authors: Klement
authors: Lina
authors: L
authors: Eisele
authors: David
authors: D
authors: Malinowsky
authors: Andreas
authors: A
authors: Nolting
authors: Mats
authors: M
authors: Svensson
authors: Gitte
authors: G
authors: Terp
authors: Dirk
authors: D
authors: Weigelt
authors: Michael
authors: M
authors: Dabrowski
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D015220:Calcium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051663:Kv1.2 Potassium Channel / Q000737:chemistry / Q000235:genetics; D008024:Ligands; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010091:Oximes / Q000494:pharmacology; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D051381:Rats; D017385:Sequence Homology; D000074025:TRPA1 Cation Channel; D050051:Transient Receptor Potential Channels / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014981:Xenopus
chemical_list: C560402:A 967079; D015220:Calcium Channels; D051663:Kv1.2 Potassium Channel; D008024:Ligands; D009419:Nerve Tissue Proteins; D010091:Oximes; D000074025:TRPA1 Cation Channel; C473350:TRPA1 protein, human; D050051:Transient Receptor Potential Channels