Structure and regulation of the cAMP-binding domains of Epac2.
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.
Rehmann
Holger
H
Max-Planck Institut für Molekulare Physiologie, Otto Hahn Strasse 11, D-44227, Dortmund, Germany.
Prakash
Balaji
B
Wolf
Eva
E
Rueppel
Alma
A
de Rooij
Johan
J
Bos
Johannes L
JL
Wittinghofer
Alfred
A
D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D000242:Cyclic AMP / Q000737:chemistry / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D020668:rap GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism
D020662:Guanine Nucleotide Exchange Factors; C442864:RAPGEF4 protein, human; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D020668:rap GTP-Binding Proteins