title: Role of dynamics in the autoinhibition and activation of the exchange protein directly activated by cyclic AMP (EPAC).
abstract: The exchange protein directly activated by cAMP (EPAC) is a key receptor of cAMP in eukaryotes and controls critical signaling pathways. Currently, no residue resolution information is available on the full-length EPAC dynamics, which are known to be pivotal determinants of allostery. In addition, no information is presently available on the intermediates for the classical induced fit and conformational selection activation pathways. Here these questions are addressed through molecular dynamics simulations on five key states along the thermodynamic cycle for the cAMP-dependent activation of a fully functional construct of EPAC2, which includes the cAMP-binding domain and the integral catalytic region. The simulations are not only validated by the agreement with the experimental trends in cAMP-binding domain dynamics determined by NMR, but they also reveal unanticipated dynamic attributes, rationalizing previously unexplained aspects of EPAC activation and autoinhibition. Specifically, the simulations show that cAMP binding causes an extensive perturbation of dynamics in the distal catalytic region, assisting the recognition of the Rap1b substrate. In addition, analysis of the activation intermediates points to a possible hybrid mechanism of EPAC allostery incorporating elements of both the induced fit and conformational selection models. In this mechanism an entropy compensation strategy results in a low free-energy pathway of activation. Furthermore, the simulations indicate that the autoinhibitory interactions of EPAC are more dynamic than previously anticipated, leading to a revised model of autoinhibition in which dynamics fine tune the stability of the autoinhibited state, optimally sensitizing it to cAMP while avoiding constitutive activation.
authors.affiliation: Departments of Chemistry and Chemical Biology, McMaster University, Hamilton, Ontario L8S 4M1, Canada.
authors.forename: Bryan
authors.initials: B
authors.lastname: VanSchouwen
authors.affiliation: Departments of Chemistry and Chemical Biology, McMaster University, Hamilton, Ontario L8S 4M1, Canada.
authors.forename: Rajeevan
authors.initials: R
authors.lastname: Selvaratnam
authors.affiliation: Department of Biomedical Science and Technology, University of Udine, Piazzale Kolbe 4, 33100 Udine, Italy.
authors.forename: Federico
authors.initials: F
authors.lastname: Fogolari
authors.affiliation: Departments of Chemistry and Chemical Biology, McMaster University, Hamilton, Ontario L8S 4M1, Canada; Departments of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4M1, Canada. Electronic address: melacin@mcmaster.ca.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Melacini
mesh_terms: D000495:Allosteric Site; D000818:Animals; D003198:Computer Simulation; D000242:Cyclic AMP / Q000737:chemistry*; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D051379:Mice; D008958:Models, Molecular; D008968:Molecular Conformation; D056004:Molecular Dynamics Simulation; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D015398:Signal Transduction; D012997:Solvents
chemical_list: C482601:Epac protein, mouse; D020662:Guanine Nucleotide Exchange Factors; C486707:Rapgef4 protein, mouse; D012997:Solvents; D000242:Cyclic AMP