To further analyse the conformational changes induced by cAMP, 10,000 frames were sampled from the seeded MD data pool of apo EPAC1, as well as EPAC1-cAMP, according to the MSM state probabilities. Analysis of the resulting conformational ensemble confirms that cAMP maintains hydrogen bond interactions with the phosphate binding cassette (PBC) throughout the seeded MD simulations, as shown in Figure . The distance between these two ends of the PBC, shown in Figure as distance d, can be used to monitor the "in"/"out" conformation of the PBC. Apo EPAC1 shows large fluctuation between the two conformations, with the distance varying between 8 Å and 11 Å (Figure ). This agrees with the high population of the intermediate state seen in apo EPAC1 (Figures and).