Fig. 2.
Deoxyforskolin-stimulated complementation in SYNAC correlates with activity. (A–C) Effects of the indicated dFSK molecule on SYNAC activity. Insets represent structure of the indicated dFSK molecule with red boxes indicating differences from the structure of FSK. (D) Effects of the indicated dFSK molecules on SYNAC FRET at the indicated concentrations. (E) Activity of SYNAC plotted against FRET readout at 100 μM of the indicated compound. R2 of 0.94 indicates a highly linear correlation. (F) Inhibition of 10 μM FSK-stimulated SYNAC activity by 1-day FSK at the indicated concentrations. The red line indicates basal activity levels for SYNAC. The solid black line indicates 10 μM FSK-stimulated SYNAC activity, with the dashed lines representing S.E.M. Data for this panel was normalized to FSK-stimulated activity, and significance was assessed by Student’s t test (paired) to account for a systematic bias in data. Activity data are mean ± S.E.M. of three independent repeats using three different preparations of protein. FRET data are mean ± S.D. of three spectra. Significance assessed by Student’s t test (unpaired) unless otherwise indicated. ***P < 0.001.