title: Barbiturates Bind in the GLIC Ion Channel Pore and Cause Inhibition by Stabilizing a Closed State.
abstract: Barbiturates induce anesthesia by modulating the activity of anionic and cationic pentameric ligand-gated ion channels (pLGICs). Despite more than a century of use in clinical practice, the prototypic binding site for this class of drugs within pLGICs is yet to be described. In this study, we present the first X-ray structures of barbiturates bound to GLIC, a cationic prokaryotic pLGIC with excellent structural homology to other relevant channels sensitive to general anesthetics and, as shown here, to barbiturates, at clinically relevant concentrations. Several derivatives of barbiturates containing anomalous scatterers were synthesized, and these derivatives helped us unambiguously identify a unique barbiturate binding site within the central ion channel pore in a closed conformation. In addition, docking calculations around the observed binding site for all three states of the receptor, including a model of the desensitized state, showed that barbiturates preferentially stabilize the closed state. The identification of this pore binding site sheds light on the mechanism of barbiturate inhibition of cationic pLGICs and allows the rationalization of several structural and functional features previously observed for barbiturates.
authors.affiliation: From the Unité de Dynamique Structurale des Macromolécules, UMR 3528 du CNRS, Institut Pasteur, 75015 Paris, France.
authors.forename: Zaineb
authors.initials: Z
authors.lastname: Fourati
authors.affiliation: From the Unité de Dynamique Structurale des Macromolécules, UMR 3528 du CNRS, Institut Pasteur, 75015 Paris, France.
authors.forename: Reinis Reinholds
authors.initials: RR
authors.lastname: Ruza
authors.affiliation: the Department of Neuroscience, Physiology and Pharmacology, University College London, London WC1E 6BT, United Kingdom.
authors.forename: Duncan
authors.initials: D
authors.lastname: Laverty
authors.affiliation: the UMR 8076 du CNRS, BioCIS, Faculté de Pharmacie, Université Paris Sud, 92296 Chatenay-Malabry, France.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Drège
authors.affiliation: the UMR 8076 du CNRS, BioCIS, Faculté de Pharmacie, Université Paris Sud, 92296 Chatenay-Malabry, France.
authors.forename: Sandrine
authors.initials: S
authors.lastname: Delarue-Cochin
authors.affiliation: the UMR 8076 du CNRS, BioCIS, Faculté de Pharmacie, Université Paris Sud, 92296 Chatenay-Malabry, France.
authors.forename: Delphine
authors.initials: D
authors.lastname: Joseph
authors.affiliation: the Department of Computer Science, University of California, Davis, California 95616.
authors.forename: Patrice
authors.initials: P
authors.lastname: Koehl
authors.affiliation: the Department of Neuroscience, Physiology and Pharmacology, University College London, London WC1E 6BT, United Kingdom. Electronic address: t.smart@ucl.ac.uk.
authors.forename: Trevor
authors.initials: T
authors.lastname: Smart
authors.affiliation: From the Unité de Dynamique Structurale des Macromolécules, UMR 3528 du CNRS, Institut Pasteur, 75015 Paris, France. Electronic address: delarue@pasteur.fr.
authors.forename: Marc
authors.initials: M
authors.lastname: Delarue
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001463:Barbiturates / Q000737:chemistry* / Q000494:pharmacology; D001665:Binding Sites; D018360:Crystallography, X-Ray; D000458:Cyanobacteria; D007473:Ion Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D020836:Protein Structure, Quaternary; D014982:Xenopus laevis
chemical_list: D001426:Bacterial Proteins; D001463:Barbiturates; D007473:Ion Channels
keywords: anesthesia; barbiturates; crystallography; electrophysiology; ligand-gated ion channels; membrane protein; structural biology; x-ray crystallography