title: Rhodopsin activation blocked by metal-ion-binding sites linking transmembrane helices C and F.
abstract: A large superfamily of receptors containing seven transmembrane (TM) helices transmits hormonal and sensory signals across the plasma membrane to heterotrimeric G proteins at the cytoplasmic face of the membrane. To investigate how G-protein-coupled receptors work at the molecular level, we have engineered metal-ion-binding sites between TM helices to restrain activation-induced conformational change in specific locations. In rhodopsin, the photoreceptor of retinal rod cells, we substituted histidine residues for natural amino acids at the cytoplasmic ends of the TM helices C and F. The resulting mutant proteins were able to activate the visual G protein transducin in the absence but not in the presence of metal ions. These results indicate that the TM helices C and F are in close proximity and suggest that movements of these helices relative to one another are required for transducin activation. Thus a change in the orientations of TM helices C and F is likely to be a key element in the mechanism for coupling binding of ligands (or isomerization of retinal) to the activation of G-protein-coupled receptors.
authors.affiliation: Department of Cellular and Molecular Biology and the Cardiovascular Research Institute, University of California, San Francisco 94143, USA.
authors.forename: S P
authors.initials: SP
authors.lastname: Sheikh
authors.forename: T A
authors.initials: TA
authors.lastname: Zvyaga
authors.forename: O
authors.initials: O
authors.lastname: Lichtarge
authors.forename: T P
authors.initials: TP
authors.lastname: Sakmar
authors.forename: H R
authors.initials: HR
authors.lastname: Bourne
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011956:Receptors, Cell Surface / Q000378:metabolism*; D012243:Rhodopsin / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013056:Spectrophotometry, Ultraviolet; D015293:Transducin / Q000378:metabolism*; D015032:Zinc / Q000378:metabolism*
chemical_list: D011956:Receptors, Cell Surface; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006639:Histidine; D012243:Rhodopsin; D015293:Transducin; D015032:Zinc