Benians, A; Leaney, JL; Milligan, G; Tinker, A; (2003) The dynamics of formation and action of the ternary complex revealed in living cells using a G-protein-gated K+ channel as a biosensor. J BIOL CHEM , 278 (12) 10851 - 10858. 10.1074/jbc.M212299200.
Traditionally the consequences of activation of G-protein-coupled receptors (GPCRs) by an agonist are studied using biochemical assays. In this study we use live cells and take advantage of a G-protein-gated inwardly rectifying potassium channel (Kir3.1+3.2A) that is activated by the direct binding of Gbetagamma subunit to the channel complex to report, in real-time, using the patch clamp technique the activity of the "ternary complex" of agonist/receptor/G-protein. This analysis is further facilitated by the use of pertussis toxin-resistant fluorescent and non-fluorescent Galpha(i/o) subunits and a series of HEK293 cell lines stably expressing both channel and receptors (including the adenosine A(1) receptor, the adrenergic alpha(2A) receptor, the dopamine D-2S receptor, the M4 muscarinic receptor, and the dimeric GABA-B-1b/2 receptor). We systematically analyzed the contribution of the various inputs to the observed kinetic response of channel activation. Our studies indicate that the combination of agonist, GPCR, and G-protein isoform uniquely specify the behavior of these channels and thus support the importance of the whole ternary complex at a kinetic level.
|Title:||The dynamics of formation and action of the ternary complex revealed in living cells using a G-protein-gated K+ channel as a biosensor|
|Open access status:||An open access publication|
|Keywords:||GTP-BINDING PROTEINS, MUSCARINIC POTASSIUM CHANNEL, HIPPOCAMPAL CA3 NEURONS, FUNCTIONAL EXPRESSION, COUPLED RECEPTORS, FUSION PROTEINS, ALPHA-SUBUNITS, ION CHANNELS, C-PROTEIN, I-KACH|
|UCL classification:||UCL > School of Life and Medical Sciences > Faculty of Medical Sciences > Medicine (Division of)|
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