%0 Journal Article %1 Hein2005 %A Hein, P. %A Frank, M. %A Hoffmann, C. %A Lohse, M. J. %A Bunemann, M. %D 2005 %J EMBO J %K Bacterial Cell Dyes Energy Fluorescence Fluorescent G-Protein-Coupled/agonists/*metabolism GTP-Binding Humans Kinetics Labeling Line Luminescent Membrane/metabolism Proteins/agonists/*metabolism Proteins/metabolism Resonance Staining Transfer alpha-2/agonists/*metabolism and Receptor Adrenergic %N 23 %P 4106-14 %T Dynamics of receptor/G protein coupling in living cells %U http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16292347 %V 24 %X The interaction of activated G protein-coupled receptors with G proteins is a key event in signal transduction. Here, using a fluorescence resonance energy transfer (FRET)-based assay, we measure directly and in living cells the interaction of YFP-labeled alpha(2A)-adrenergic receptors with CFP-labeled G proteins. Upon agonist stimulation, a small, concentration-dependent increase in FRET was observed. No specific basal FRET was detected in the absence of agonist. Kinetics of the onset of receptor/G protein interaction were <100 ms and depended on expression levels of Galpha. Simultaneously recorded G protein-regulated inwardly rectifying K(+) channel currents revealed a maximal current response already at agonist concentrations producing submaximal FRET amplitudes. By analyzing FRET signals in the presence of a Galpha mutant, which dissociates more slowly from activated receptors, it was demonstrated that only a fraction of wild-type G proteins interacts with the activated receptor at any time. Our data suggest that alpha(2A)-adrenergic receptors and G proteins interact by rapid collision coupling and indicate that there is no significant precoupling between these receptors and G proteins.

@article{Hein2005, abstract = {The interaction of activated G protein-coupled receptors with G proteins is a key event in signal transduction. Here, using a fluorescence resonance energy transfer (FRET)-based assay, we measure directly and in living cells the interaction of YFP-labeled alpha(2A)-adrenergic receptors with CFP-labeled G proteins. Upon agonist stimulation, a small, concentration-dependent increase in FRET was observed. No specific basal FRET was detected in the absence of agonist. Kinetics of the onset of receptor/G protein interaction were <100 ms and depended on expression levels of Galpha. Simultaneously recorded G protein-regulated inwardly rectifying K(+) channel currents revealed a maximal current response already at agonist concentrations producing submaximal FRET amplitudes. By analyzing FRET signals in the presence of a Galpha mutant, which dissociates more slowly from activated receptors, it was demonstrated that only a fraction of wild-type G proteins interacts with the activated receptor at any time. Our data suggest that alpha(2A)-adrenergic receptors and G proteins interact by rapid collision coupling and indicate that there is no significant precoupling between these receptors and G proteins.}, added-at = {2010-12-14T18:12:02.000+0100}, author = {Hein, P. and Frank, M. and Hoffmann, C. and Lohse, M. J. and Bunemann, M.}, biburl = {https://www.bibsonomy.org/bibtex/219616515e3dcc662c100ff979ea15eb2/pharmawuerz}, endnotereftype = {Journal Article}, interhash = {11f312bc208fb62203e0b4941611e389}, intrahash = {19616515e3dcc662c100ff979ea15eb2}, issn = {0261-4189 (Print) 0261-4189 (Linking)}, journal = {EMBO J}, keywords = {Bacterial Cell Dyes Energy Fluorescence Fluorescent G-Protein-Coupled/agonists/*metabolism GTP-Binding Humans Kinetics Labeling Line Luminescent Membrane/metabolism Proteins/agonists/*metabolism Proteins/metabolism Resonance Staining Transfer alpha-2/agonists/*metabolism and Receptor Adrenergic}, month = {Dec 7}, note = {Hein, Peter Frank, Monika Hoffmann, Carsten Lohse, Martin J Bunemann, Moritz Research Support, Non-U.S. Gov't England The EMBO journal EMBO J. 2005 Dec 7;24(23):4106-14. Epub 2005 Nov 17.}, number = 23, pages = {4106-14}, shorttitle = {Dynamics of receptor/G protein coupling in living cells}, timestamp = {2010-12-14T18:22:40.000+0100}, title = {Dynamics of receptor/G protein coupling in living cells}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16292347}, volume = 24, year = 2005 }