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Gs Activation Is Time-limiting in Initiating Receptor-mediated Signaling*.
- Source :
-
Journal of Biological Chemistry . 11/3/2006, Vol. 281 Issue 44, p33345-33351. 7p. 4 Graphs. - Publication Year :
- 2006
-
Abstract
- To analyze individual steps of GS-linked signaling in intact cells, we used fluorescence resonance energy transfer (FRET)-based assays for receptor-G protein interaction, G protein activation, and cAMP effector activation. To do so, we developed a FRET-based sensor to directly monitor GS activation in living cells. This was done by coexpressing a Gαs mutant, in which a yellow fluorescent protein was inserted, together with cyan fluorescent protein-tagged Gβγ subunits and appropriate receptors in HEK293 cells. Together with assays for receptor activation and receptor-G protein interaction, it is possible to characterize large parts of the GS signaling cascade. When A2A-adenosine or β1-adrenergic receptors are coexpressed with GS in HEK293T cells, the receptor-GS interaction was on the same time scale as A2A receptor activation with a time constant of <50 ms. GS activation was markedly slower and around 450 ms with similar kinetics following activation of A2A- or β1-receptors. Taken together, our kinetic measurements demonstrate that the rate of GS activation limits initiation of GS-coupled receptor signaling. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 00219258
- Volume :
- 281
- Issue :
- 44
- Database :
- Academic Search Index
- Journal :
- Journal of Biological Chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 23851009
- Full Text :
- https://doi.org/10.1074/jbc.M606713200