Dynamics of receptor/G protein coupling in living cells
P. Hein, M. Frank, C. Hoffmann, M. Lohse, and M. Bunemann. EMBO J, 24 (23):
4106-14(December 2005)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..
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.
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.
%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
}