title: Structural requirements in the transmembrane domain of GLIC revealed by incorporation of noncanonical histidine analogs.
abstract: The cyanobacterial pentameric ligand-gated ion channel GLIC, a homolog of the Cys-loop receptor superfamily, has provided useful structural and functional information about its eukaryotic counterparts. X-ray diffraction data and site-directed mutagenesis have previously implicated a transmembrane histidine residue (His234) as essential for channel function. Here, we investigated the role of His234 via synthesis and incorporation of histidine analogs and α-hydroxy acids using in vivo nonsense suppression. Receptors were expressed heterologously in Xenopus laevis oocytes, and whole-cell voltage-clamp electrophysiology was used to monitor channel activity. We show that an interhelix hydrogen bond involving His234 is important for stabilization of the open state, and that the shape and basicity of its side chain are highly sensitive to perturbations. In contrast, our data show that two other His residues are not involved in the acid-sensing mechanism.
authors.affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
authors.forename: Matthew
authors.initials: M
authors.lastname: Rienzo
authors.affiliation: Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1GA, UK.
authors.forename: Sarah C R
authors.initials: SC
authors.lastname: Lummis
authors.affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA. Electronic address: dadougherty@caltech.edu.
authors.forename: Dennis A
authors.initials: DA
authors.lastname: Dougherty
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D000458:Cyanobacteria; D006639:Histidine / Q000031:analogs & derivatives*; D006860:Hydrogen Bonding; D006880:Hydroxy Acids / Q000737:chemistry; D058446:Ligand-Gated Ion Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D017434:Protein Structure, Tertiary
chemical_list: D001426:Bacterial Proteins; D006880:Hydroxy Acids; D058446:Ligand-Gated Ion Channels; D006639:Histidine