Abstract
Epac proteins are activated by binding of the second messenger cAMP and then act as guanine nucleotide exchange factors for Rap proteins1,2. The Epac proteins are involved in the regulation of cell adhesion3 and insulin secretion4. Here we have determined the structure of Epac2 in complex with a cAMP analogue (Sp-cAMPS) and RAP1B by X-ray crystallography and single particle electron microscopy. The structure represents the cAMP activated state of the Epac2 protein with the RAP1B protein trapped in the course of the exchange reaction. Comparison with the inactive conformation reveals that cAMP binding causes conformational changes that allow the cyclic nucleotide binding domain to swing from a position blocking the Rap binding site towards a docking site at the Ras exchange motif domain.
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Primary accessions
Protein Data Bank
Data deposits
Co-ordinates have been deposited with the Protein Data Bank under accession number 3cf6. The EM map of activated full length Epac2 bound to RAP1B has been deposited in the 3D EM database (http://www.ebi.ac.uk/msd/) under accession code EMD-1510.
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Acknowledgements
We thank P. Gros for access to crystallization robots, M. Rensen-De Leeuw for technical assistance, A. Wittinghofer for reading the manuscript, the European Synchrotron Radiation Facility for providing synchrotron facilities, the scientists at ID23-1 for help with data collection, and the Galicia Supercomputer Centre (CESGA) and the Barcelona Supercomputing Centre for computing resources. H.R. is a recipient of the Hendrik Casimir-Karl Ziegler-Forschungspreis of the Nordrhein-Westfälischen Akademie der Wissenschaften and the Koninklijke Nederlandse Akademie van Wetenschappen. E.A.-P. and O.L. are supported by the Autonomous Region of Madrid, F.S. by the Bremer Innovationsagentur, O.L. by the Spanish Ministry of Education and Science (MEC) and the Red Temática de Investigación cooperativa en Cáncer (RTICC), and J.L.B. by the Chemical Sciences and the Netherlands Genomic Initiative of the Netherlands Organisation for Scientific Research (NWO).
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Supplementary Information 1
This file contains Supplementary Table 1 describing the data collections and refinement statistics, a legend to Supplementary Movie describing the content of the movie in detail and Supplementary Figures S1 to S9 showing detail aspect of the EM and the crystal structures as well as a biochemical data. (PDF 5084 kb)
Supplementary Information 2
This file contains Supplementary Movie 1. This movie illustrates in cartoon style how cAMP induces the activation of Epac. The conformational changes within the CNB domain are shown left and the resulting structural consequences for the domain organisation of Epac right. A detailed description can be found in the Supplementary Information file. (MOV 2324 kb)
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Rehmann, H., Arias-Palomo, E., Hadders, M. et al. Structure of Epac2 in complex with a cyclic AMP analogue and RAP1B. Nature 455, 124–127 (2008). https://doi.org/10.1038/nature07187
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DOI: https://doi.org/10.1038/nature07187
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