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. 2010 Sep;78(3):524–533. doi: 10.1124/mol.110.065128

Fig. 1.

Fig. 1.

Characterization of the RGS4 TR-FRET high-throughput assay. A, schematic of RGS4-Gαo TR-FRET assay. Gαo is labeled with the LanthaScreen Tb-chelate donor fluorophore, and RGS4 is labeled with an Alexa Fluor 488 acceptor fluorophore. Excitation and emission maxima are listed for each fluorophore. B, representative data showing the AlF4−/GDP dependence of the interaction between RGS4-AF488 and 10 nM Tb-Gαo. This saturable interaction has a Kd of 35 ± 4 nM. C, two compounds identified in the high-throughput screen, CCG-63802 and CCG-63808, dose-dependently inhibit the TR-FRET signal between RGS4-AF488 and Tb-Gαo with IC50 values of 1.4 (0.76; 2.6 μM) and 1.9 μM (1.02; 3.5 μM), respectively. Data (n = 3 for all data) are presented as mean ± S.E.M. or mean (95% confidence interval) in B and C, respectively. D, the chemical structures of CCG-63802 and CCG-63808.