title: Structural Insights into the Atomistic Mechanisms of Action of Small Molecule Inhibitors Targeting the KCa3.1 Channel Pore.
abstract: The intermediate-conductance Ca2+-activated K+channel (KCa3.1) constitutes an attractive pharmacological target for immunosuppression, fibroproliferative disorders, atherosclerosis, and stroke. However, there currently is no available crystal structure of this medically relevant channel that could be used for structure-assisted drug design. Using the Rosetta molecular modeling suite we generated a molecular model of the KCa3.1 pore and tested the model by first confirming previously mapped binding sites and visualizing the mechanism of TRAM-34 (1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole), senicapoc (2,2-bis-(4-fluorophenyl)-2-phenylacetamide), and NS6180 (4-[[3-(trifluoromethyl)phenyl]methyl]-2H-1,4-benzothiazin-3(4H)-one) inhibition at the atomistic level. All three compounds block ion conduction directly by fully or partially occupying the site that would normally be occupied by K+before it enters the selectivity filter. We then challenged the model to predict the receptor sites and mechanisms of action of the dihydropyridine nifedipine and an isosteric 4-phenyl-pyran. Rosetta predicted receptor sites for nifedipine in the fenestration region and for the 4-phenyl-pyran in the pore lumen, which could both be confirmed by site-directed mutagenesis and electrophysiology. While nifedipine is thus not a pore blocker and might be stabilizing the channel in a nonconducting conformation or interfere with gating, the 4-phenyl-pyran was found to be a classical pore blocker that directly inhibits ion conduction similar to the triarylmethanes TRAM-34 and senicapoc. The Rosetta KCa3.1 pore model explains the mechanism of action of several KCa3.1 blockers at the molecular level and could be used for structure-assisted drug design.
authors.affiliation: Department of Pharmacology (H.M.N, V.S., B.P., D.P.J., H.W.) and Department of Physiology and Membrane Biology (V. Y.-Y.), School of Medicine, University of California at Davis, Davis, California.
authors.forename: Hai M
authors.initials: HM
authors.lastname: Nguyen
authors.affiliation: Department of Pharmacology (H.M.N, V.S., B.P., D.P.J., H.W.) and Department of Physiology and Membrane Biology (V. Y.-Y.), School of Medicine, University of California at Davis, Davis, California.
authors.forename: Vikrant
authors.initials: V
authors.lastname: Singh
authors.affiliation: Department of Pharmacology (H.M.N, V.S., B.P., D.P.J., H.W.) and Department of Physiology and Membrane Biology (V. Y.-Y.), School of Medicine, University of California at Davis, Davis, California.
authors.forename: Brandon
authors.initials: B
authors.lastname: Pressly
authors.affiliation: Department of Pharmacology (H.M.N, V.S., B.P., D.P.J., H.W.) and Department of Physiology and Membrane Biology (V. Y.-Y.), School of Medicine, University of California at Davis, Davis, California.
authors.forename: David Paul
authors.initials: DP
authors.lastname: Jenkins
authors.affiliation: Department of Pharmacology (H.M.N, V.S., B.P., D.P.J., H.W.) and Department of Physiology and Membrane Biology (V. Y.-Y.), School of Medicine, University of California at Davis, Davis, California yarovoy@ucdavis.edu hwulff@ucdavis.edu.
authors.forename: Heike
authors.initials: H
authors.lastname: Wulff
authors.affiliation: Department of Pharmacology (H.M.N, V.S., B.P., D.P.J., H.W.) and Department of Physiology and Membrane Biology (V. Y.-Y.), School of Medicine, University of California at Davis, Davis, California yarovoy@ucdavis.edu hwulff@ucdavis.edu.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Yarov-Yarovoy
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D006801:Humans; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels / Q000737:chemistry / Q000494:pharmacology; D008024:Ligands; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D009543:Nifedipine / Q000737:chemistry / Q000494:pharmacology; D026902:Potassium Channel Blockers / Q000737:chemistry / Q000494:pharmacology*; D000072417:Protein Domains; D016415:Sequence Alignment; D054852:Small Molecule Libraries / Q000737:chemistry* / Q000494:pharmacology*; D040681:Structural Homology, Protein; D013329:Structure-Activity Relationship; D013843:Thiazines / Q000737:chemistry / Q000494:pharmacology
chemical_list: C581707:4-((3-(trifluoromethyl)phenyl)methyl)-2H-1,4-benzothiazin-3(4H)-one; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels; C496923:KCNN4 protein, human; D008024:Ligands; D026902:Potassium Channel Blockers; D054852:Small Molecule Libraries; D013843:Thiazines; D009543:Nifedipine