Fig. 5: Models of EPAC1 activation by cAMP and membranes and inhibition by CE3F4.

a The conformational ensemble of EPAC1. EPAC1 exists in at least four states in equilibrium (arrows) with distinct GEF activities and cAMP affinities. Because of its low affinity for cAMP in solution, the soluble cAMP-bound EPAC1 intermediate (light colors, top right) is predicted to be a negligible species in cells. For the same reason, it is also predicted that in cells, CE3F4 targets only membrane-associated, cAMP-activated EPAC1 (bottom right). b Electrostatic coincidence detection of EPAC1 and Rap1A on anionic membranes. Positive charges in proteins (blue crosses), negative charges in the membrane (red circles), and the lipidated C-terminus of the Rap GTPase are indicated. c β-hairpin-mediated allosteric inhibition of EPAC1 by CE3F4. The binding of CE3F4 to the hinge located between the CNB and GEF domains (in black) is proposed to displace a conserved β-hairpin (in red) into the Rap-binding site. The hinge and the β-hairpin structures are from the crystal structure of cAMP-activated EPAC2²⁰ (from PDB entry: 3CF6). d An equivalent inhibitory β hairpin in the related RasGEF SOS. In inactive SOS, the β-hairpin protrudes into the Ras-binding site (orange). In Ras-GTP-activated SOS, it has relocated outside the Ras-binding site (in red). The relative positions of the β-hairpin are based on the superimposition of the Ras-binding site in inactive SOS and Ras-GTP-activated SOS structures (ref. ⁶⁵, from PDB entries: 1NXV and 2II0).