Abstract
Cyclic adenosine monophosphate (cAMP) is a universal second messenger that, in eukaryotes, was believed to act only on cAMP-dependent protein kinase A (PKA) and cyclic nucleotide-regulated ion channels. Recently, guanine nucleotide exchange factors specific for the small GTP-binding proteins Rap1 and Rap2 (Epacs) were described, which are also activated directly by cAMP. Here, we have determined the three-dimensional structure of the regulatory domain of Epac2, which consists of two cyclic nucleotide monophosphate (cNMP)-binding domains and one DEP (Dishevelled, Egl, Pleckstrin) domain. This is the first structure of a cNMP-binding domain in the absence of ligand, and comparison with previous structures, sequence alignment and biochemical experiments allow us to delineate a mechanism for cyclic nucleotide-mediated conformational change and activation that is most likely conserved for all cNMP-regulated proteins. We identify a hinge region that couples cAMP binding to a conformational change of the C-terminal regions. Mutations in the hinge of Epac can uncouple cAMP binding from its exchange activity.
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Acknowledgements
We thank I. Vetter, I. Schlichting, M. Weyand and A. Scheidig for help during data collection and interesting discussions. We thank D. Kuehlmann, C. Koerner and A. Gerhards for excellent technical support, R. Schebaum for secretarial assistance and M. Würtele and M. Hess for help with figure preparation. We thank the staff of ESRF Grenoble, particularly M. Roth, for beam time allocation and assistance with X-ray data collection. H.R. was supported by a grant from the Council of Earth and Life Science of the Netherlands Organization for Scientific Research (NWO-ALW) and B.P. from an HFSP grant to A.W.
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Rehmann, H., Prakash, B., Wolf, E. et al. Structure and regulation of the cAMP-binding domains of Epac2. Nat Struct Mol Biol 10, 26–32 (2003). https://doi.org/10.1038/nsb878
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DOI: https://doi.org/10.1038/nsb878
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