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PDBsum entry 3cf6
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Signaling protein/gtp-binding protein
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PDB id
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3cf6
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References listed in PDB file
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Key reference
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Title
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Structure of epac2 in complex with a cyclic AMP analogue and rap1b.
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Authors
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H.Rehmann,
E.Arias-Palomo,
M.A.Hadders,
F.Schwede,
O.Llorca,
J.L.Bos.
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Ref.
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Nature, 2008,
455,
124-127.
[DOI no: ]
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PubMed id
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Abstract
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Epac proteins are activated by binding of the second messenger cAMP and then act
as guanine nucleotide exchange factors for Rap proteins. The Epac proteins are
involved in the regulation of cell adhesion and insulin secretion. 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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Figure 1.
Figure 1: Active Epac 2. a, Domain organization of Epac2.
Residues that were subjected to mutational analysis are
indicated. The same colour code is used throughout the figures.
Hinge (residues 432–445, dark green); helical hairpin
(residues 906 to 946, dark blue). CDC25-HD, CDC25 homology
domain; CNB, cyclic nucleotide binding domain; DEP, Dishevelled,
Egl-10, Pleckstrin domain; RA, Ras-association domain; REM,
Ras-exchange motif. b, Left, inactive Epac2 (first CNB and DEP
domain omitted); right, active Epac2  305
 Sp-cAMPS
RAP1B.
RAP1B is shown orange; Sp-cAMPS and SO[4]^2- are shown in ball
and stick representation. Arrow, movement of the second CNB
domain; straight lines, missing connectivity; dotted lines,
ionic latch (IL); asterisks, interface between the REM and the
CNB domain; HP, helical hairpin; PBC, phosphate binding
cassette. c, RAP1B placed into the inactive structure. d, The
crystal structure of Epac2 305
Sp-cAMPS
RAP1B
was fitted into the EM density reconstruction (grey grid) of
full length Epac2 cAMP
RAP1B.
Yellow surface, difference density.
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Figure 2.
Figure 2: Sp-cAMPS induced conformational changes. a,
Superposition of the active and inactive second CNB domain. The
arrows indicate the movement of the hinge and the lid region.
Light green, active conformation; dark green, inactive
conformation; grey, no difference in conformation. b,
Interactions of Sp-cAMPS with the CNB domain and the REM domain.
Hydrogen bonds are shown by dotted lines; w, water. c,
Interaction of Lys 405 with the hinge-lid region.
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The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
Nature
(2008,
455,
124-127)
copyright 2008.
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Secondary reference #1
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Title
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Structure of the cyclic-Amp-Responsive exchange factor epac2 in its auto-Inhibited state.
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Authors
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H.Rehmann,
J.Das,
P.Knipscheer,
A.Wittinghofer,
J.L.Bos.
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Ref.
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Nature, 2006,
439,
625-628.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1: Structure of Epac2. a, Domain organization of Epac.
The same colour code is used throughout the figures. CDC25-HD,
CDC25-homology domain; cNBD, cyclic-nucleotide-binding domain;
DEP, Dishevelled, Egl-10, Pleckstrin domain; RA, Ras-association
domain; REM, Ras-exchange motif. b, Ribbon diagram of Epac2 in
stereo view. Missing connectivity is indicated by coloured
dotted lines. The green ball indicates the cAMP-binding site in
cNBD-B. HP, helical hairpin of the CDC25-HD (dark blue); SB,
switchboard; IL, ionic latch (doted black lines); CH, connecting
helix (helix H4 in ref 13). c, Surface representation of Epac.
d, Superposition of Epac and the Ras-Sos complex. The RA, DEP
and REM domains are omitted. Only Ras (magenta) from the Ras-Sos
complex is shown.
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Figure 2.
Figure 2: Anchoring points between the regulatory and catalytic
regions. a, The switchboard is formed by strands provided by
cNBD-B (dark green), the REM domain (orange) and the loop of the
helical hairpin (HP) of CDC25-HD (blue). Peptides are reduced to
polyglycine. Hydrogen bonding by main-chain atoms is indicated
by dotted lines. b, The REM domain (orange) interacts tightly
with the C-terminal helix of the helical hairpin (blue). Thick C
 traces
highlight the parts of the switchboard shown in a. c, The ionic
latch between cNBD-B (green) and CDC25-HD (blue). CH, connecting
helix (see Fig. 1b).
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The above figures are
reproduced from the cited reference
with permission from Macmillan Publishers Ltd
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Secondary reference #2
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Title
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Structure and regulation of the camp-Binding domains of epac2.
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Authors
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H.Rehmann,
B.Prakash,
E.Wolf,
A.Rueppel,
J.De rooij,
J.L.Bos,
A.Wittinghofer.
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Ref.
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Nat Struct Biol, 2003,
10,
26-32.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1. Structure of Epac. a, Domain organization of Epac1
and 2. Indicated color code is used throughout the figure. b,
Stereo view of a 2F[o] - F[c] composite omitted electron density
map (contoured at 1.5  ).
The hydrophobic environment of Leu408 and Phe435 is shown.
Different parts of the peptide chain are highlighted by
individual colors. c, Ribbon diagram of the regulatory domain of
Epac2 with N and C termini as indicated. The C-terminal extra
helix of the DEP domain is dark green. d, Amino acid sequence,
with secondary structure annotation. The phosphate-binding
cassette (PBC) is indicated in red letters. Dashed lines
indicate portions of the polypeptide chain not visible in the
electron density. e, The two possible arrangements (1 and 2) for
the first cNMP-binding domain relative to the second (see text).
Arrangement 1 corresponds to (c). Dotted lines, linker 1 and 2,
indicate the minimal path of the polypeptide chain required to
bridge the gap in both arrangements.
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Figure 2.
Figure 2. Ribbon diagram of the DEP domains of Epac and
Dishevelled. The DEP domains of Epac (yellow and green) and
Dishevelled (gray) are superimposed on each other. Secondary
structure elements are labeled as in Fig. 1c. N and C termini,
as well as the position of the residues forming the dipole, are
indicated as follows: '1' corresponds to Asp225^Epac and
Glu448^Dvl1; '2' to Glu222^Epac and Asp445^Dvl1; and '3' to
Lys212^Epac and Lys434^Dvl1.
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The above figures are
reproduced from the cited reference
with permission from Macmillan Publishers Ltd
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