Figure 6

Ligand Observed NMR reveals Hit compounds Interact Directly with EPAC1-CNB and EPAC2-CNB, but not GST alone. (A) One dimensional proton (1D) and waterLOGSY (WL) spectra were performed on samples containing hit compound I942 (50 µM) in the absence and presence of EPAC1-CNB, EPAC2-CNB or GST alone, as indicated (2.5 µM). A representative region of the I942 spectrum is shown (7.1–7.75 ppm). Interaction between compound and protein was assessed through comparison of the compound alone and protein-compound sample spectra. The NMR data are representative of data collected at several protein and ligand concentrations. (B) 1D and WL spectra were performed on samples containing I178 (50 µM) in the absence and presence of EPAC1-CNB, EPAC2-CNB or GST alone, as indicated (2.5 µM). A representative region of the I178 spectrum is shown (7.1–7.51 ppm). (C) 1D and WL spectra were performed on samples containing the commercially available EPAC selective inhibitor ESI-09 (50 µM) in the absence and presence of EPAC1-CNB, EPAC2-CNB or GST alone, as indicated (2.5 µM). A representative region of the ESI-09 spectrum is shown (0.6–1.2 ppm).