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- PDB-3cf6: Structure of Epac2 in complex with cyclic-AMP and Rap -

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Basic information

Entry
Database: PDB / ID: 3cf6
TitleStructure of Epac2 in complex with cyclic-AMP and Rap
Components
  • Rap guanine nucleotide exchange factor (GEF) 4
  • Ras-related protein Rap-1b
KeywordsSIGNALING PROTEIN/GTP-BINDING PROTEIN / EPAC / RAPGEF4 / RAP / RAP1B / CAMP / SP-CAMPS / GEF / GUNANINE NUCLEOTIDE EXCHANGE FACTOR / G-PROTEIN / GTP-BINDING / NUCLEOTIDE-BINDING / SIGNALING PROTEIN REGULATOR-GTP-BINDING PROTEIN COMPLEX / SIGNALING PROTEIN-GTP-BINDING PROTEIN COMPLEX
Function / homology
Function and homology information


Rap protein signal transduction / modification of postsynaptic structure / regulation of cell junction assembly / negative regulation of calcium ion-dependent exocytosis / positive regulation of integrin activation / negative regulation of synaptic vesicle exocytosis / calcium-ion regulated exocytosis / Rap1 signalling / establishment of endothelial barrier / regulation of establishment of cell polarity ...Rap protein signal transduction / modification of postsynaptic structure / regulation of cell junction assembly / negative regulation of calcium ion-dependent exocytosis / positive regulation of integrin activation / negative regulation of synaptic vesicle exocytosis / calcium-ion regulated exocytosis / Rap1 signalling / establishment of endothelial barrier / regulation of establishment of cell polarity / MET activates RAP1 and RAC1 / azurophil granule membrane / p130Cas linkage to MAPK signaling for integrins / GRB2:SOS provides linkage to MAPK signaling for Integrins / cellular response to cAMP / Gene and protein expression by JAK-STAT signaling after Interleukin-12 stimulation / lipid droplet / Integrin signaling / small monomeric GTPase / G protein activity / establishment of localization in cell / Signaling by high-kinase activity BRAF mutants / MAP2K and MAPK activation / GDP binding / Signaling by RAF1 mutants / Signaling by moderate kinase activity BRAF mutants / Paradoxical activation of RAF signaling by kinase inactive BRAF / Signaling downstream of RAS mutants / cell-cell junction / Signaling by BRAF and RAF1 fusions / cell population proliferation / positive regulation of ERK1 and ERK2 cascade / GTPase activity / glutamatergic synapse / Neutrophil degranulation / protein-containing complex binding / GTP binding / extracellular exosome / membrane / plasma membrane / cytosol
Similarity search - Function
Helicase, Ruva Protein; domain 3 - #1240 / Ras-related protein Rap1 / Son of sevenless (SoS) protein; Chain S, domain 1 / Son of sevenless (SoS) protein Chain: S domain 1 / Son of Sevenless (SoS) protein; Chain S, domain 2 / Ras guanine-nucleotide exchange factors catalytic domain / Small GTPase, Ras-type / small GTPase Ras family profile. / Ran (Ras-related nuclear proteins) /TC4 subfamily of small GTPases / Jelly Rolls ...Helicase, Ruva Protein; domain 3 - #1240 / Ras-related protein Rap1 / Son of sevenless (SoS) protein; Chain S, domain 1 / Son of sevenless (SoS) protein Chain: S domain 1 / Son of Sevenless (SoS) protein; Chain S, domain 2 / Ras guanine-nucleotide exchange factors catalytic domain / Small GTPase, Ras-type / small GTPase Ras family profile. / Ran (Ras-related nuclear proteins) /TC4 subfamily of small GTPases / Jelly Rolls / Phosphatidylinositol 3-kinase Catalytic Subunit; Chain A, domain 1 / Helicase, Ruva Protein; domain 3 / Rho (Ras homology) subfamily of Ras-like small GTPases / Ras subfamily of RAS small GTPases / Small GTPase / Ras family / Rab subfamily of small GTPases / Ubiquitin-like (UB roll) / Small GTP-binding protein domain / P-loop containing nucleotide triphosphate hydrolases / Jelly Rolls / Roll / Up-down Bundle / Sandwich / P-loop containing nucleoside triphosphate hydrolase / Rossmann fold / Orthogonal Bundle / 3-Layer(aba) Sandwich / Mainly Beta / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
Chem-SP1 / : / Ras-related protein Rap-1b
Similarity search - Component
Biological speciesMus musculus (house mouse)
Homo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.2 Å
AuthorsRehmann, H. / Arias-Palomo, E. / Hadders, M.A. / Schwede, F. / Llorca, O. / Bos, J.L.
Citation
Journal: Nature / Year: 2008
Title: Structure of Epac2 in complex with a cyclic AMP analogue and RAP1B.
Authors: Holger Rehmann / Ernesto Arias-Palomo / Michael A Hadders / Frank Schwede / Oscar Llorca / Johannes L Bos /
Abstract: 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 ...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.
#1: Journal: Nature / Year: 2006
Title: Structure of the cyclic-AMP-responsive exchange factor Epac2 in its auto-inhibited state
Authors: Rehmann, H. / Das, J. / Knipscheer, P. / Wittinghofer, A. / Bos, J.L.
#2: Journal: Nat.Struct.Biol. / Year: 2003
Title: Structure and regulation of the cAMP-binding domains of Epac2
Authors: Rehmann, H. / Prakash, B. / Wolf, E. / Rueppel, A. / de Rooij, J. / Bos, J.L. / Wittinghofer, A.
History
DepositionMar 2, 2008Deposition site: RCSB / Processing site: PDBJ
Revision 1.0Jul 29, 2008Provider: repository / Type: Initial release
Revision 1.1Jul 13, 2011Group: Version format compliance
Revision 1.2Feb 5, 2014Group: Derived calculations
Revision 1.3Aug 4, 2021Group: Database references / Derived calculations / Refinement description
Category: refine / struct_ref_seq_dif / struct_site
Item: _refine.ls_percent_reflns_obs / _struct_ref_seq_dif.details ..._refine.ls_percent_reflns_obs / _struct_ref_seq_dif.details / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id
Revision 1.4Nov 1, 2023Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
E: Rap guanine nucleotide exchange factor (GEF) 4
R: Ras-related protein Rap-1b
hetero molecules


Theoretical massNumber of molelcules
Total (without water)99,0855
Polymers98,2982
Non-polymers7873
Water4,936274
1
E: Rap guanine nucleotide exchange factor (GEF) 4
R: Ras-related protein Rap-1b
hetero molecules

E: Rap guanine nucleotide exchange factor (GEF) 4
R: Ras-related protein Rap-1b
hetero molecules


Theoretical massNumber of molelcules
Total (without water)198,17010
Polymers196,5974
Non-polymers1,5736
Water724
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation8_554x,-y,-z-1/21
Buried area11990 Å2
ΔGint-97 kcal/mol
Surface area67380 Å2
MethodPISA
2


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4280 Å2
ΔGint-43 kcal/mol
Surface area35410 Å2
MethodPISA
Unit cell
Length a, b, c (Å)125.367, 149.025, 225.291
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number24
Space group name H-MI212121
Components on special symmetry positions
IDModelComponents
11E-998-

HOH

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Components

#1: Protein Rap guanine nucleotide exchange factor (GEF) 4


Mass: 79277.836 Da / Num. of mol.: 1 / Fragment: UNP residues 306-993
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Description: GST fusion / Gene: Rapgef4 / Plasmid: pGEX4T1 / Production host: Escherichia coli (E. coli) / Strain (production host): CK600K / References: UniProt: A2ASW3
#2: Protein Ras-related protein Rap-1b / GTP-binding protein smg p21B / Rap1B


Mass: 19020.508 Da / Num. of mol.: 1 / Fragment: UNP residues 1-167
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: RAP1B / Plasmid: pTac / Production host: Escherichia coli (E. coli) / Strain (production host): CK600K / References: UniProt: P61224
#3: Chemical ChemComp-SP1 / 6-(6-AMINO-PURIN-9-YL)-2-THIOXO-TETRAHYDRO-2-FURO[3,2-D][1,3,2]DIOXAPHOSPHININE-2,7-DIOL / SP-ADENOSINE-3',5'-CYCLIC-MONOPHOSPHOROTHIOATE


Mass: 345.272 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H12N5O5PS
#4: Chemical ChemComp-SO4 / SULFATE ION / Sulfate


Mass: 96.063 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: SO4
#5: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 274 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 5.352371 Å3/Da / Density % sol: 77.019531 %
Crystal growTemperature: 277 K / Method: vapor diffusion, sitting drop / pH: 5.6
Details: 0.4M (NH4)2SO4, 1.2M Li2SO4, 0.1M citrate, pH 5.6, VAPOR DIFFUSION, SITTING DROP, temperature 277K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: ESRF / Beamline: ID23-1 / Wavelength: 0.9732 Å
DetectorType: ADSC QUANTUM 315 / Detector: CCD / Date: Oct 7, 2007 / Details: mirror
RadiationMonochromator: Silicon (1 1 1) channel-cut / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9732 Å / Relative weight: 1
ReflectionResolution: 2.2→50 Å / Num. all: 104773 / Num. obs: 102111 / % possible obs: 95.6 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 3.62 % / Rmerge(I) obs: 0.084
Reflection shellResolution: 2.2→2.3 Å / Rmerge(I) obs: 0.49 / Mean I/σ(I) obs: 2.96 / % possible all: 87.3

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Processing

Software
NameVersionClassification
ADSCQuantumdata collection
MOLREPphasing
REFMAC5.2.0019refinement
XDSdata reduction
XSCALEdata scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: pdb entry 2byv, pdb entry 1BKD

2byv

1bkd


Resolution: 2.2→48.56 Å / Isotropic thermal model: isotropic / Cross valid method: THROUGHOUT / σ(F): 0 / Stereochemistry target values: Engh & Huber
RfactorNum. reflection% reflectionSelection details
Rfree0.26503 5214 -random
Rwork0.2434 ---
all0.24446 100557 --
obs0.24446 100557 95.6 %-
Displacement parametersBiso mean: 48.187 Å2
Baniso -1Baniso -2Baniso -3
1-0.11 Å20 Å20 Å2
2--1.41 Å20 Å2
3----1.52 Å2
Refinement stepCycle: LAST / Resolution: 2.2→48.56 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6238 0 49 274 6561
Refine LS restraints
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONr_bond_refined_d0.012
X-RAY DIFFRACTIONr_angle_refined_deg1.418
X-RAY DIFFRACTIONr_chiral_restr0.096
X-RAY DIFFRACTIONr_gen_planes_refined0.005

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