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- PDB-6tgb: CryoEM structure of the binary DOCK2-ELMO1 complex -

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Basic information

Entry
Database: PDB / ID: 6tgb
TitleCryoEM structure of the binary DOCK2-ELMO1 complex
Components
  • Dedicator of cytokinesis protein 2
  • Engulfment and cell motility protein 1ELMO1
KeywordsSIGNALING PROTEIN / guanine nucleotide exchange factor / cytoskeleton / actin / cryoEM
Function / homology
Function and homology information


alpha-beta T cell proliferation / membrane raft polarization / myeloid dendritic cell activation involved in immune response / establishment of T cell polarity / guanyl-nucleotide exchange factor complex / macropinocytosis / Rac guanyl-nucleotide exchange factor activity / immunological synapse formation / Rac protein signal transduction / negative thymic T cell selection ...alpha-beta T cell proliferation / membrane raft polarization / myeloid dendritic cell activation involved in immune response / establishment of T cell polarity / guanyl-nucleotide exchange factor complex / macropinocytosis / Rac guanyl-nucleotide exchange factor activity / immunological synapse formation / Rac protein signal transduction / negative thymic T cell selection / positive thymic T cell selection / small GTPase mediated signal transduction / cell motility / positive regulation of phagocytosis / GTPase activator activity / T cell receptor binding / phagocytosis, engulfment / actin cytoskeleton organization / regulation of defense response to virus by virus / SH3 domain binding / vascular endothelial growth factor receptor signaling pathway / Fc-gamma receptor signaling pathway involved in phagocytosis / specific granule lumen / chemotaxis / cell migration / cytoskeleton / apoptotic process / neutrophil degranulation / extracellular exosome / membrane / extracellular region / plasma membrane / cytosol / cytoplasm
Dedicator of cytokinesis / Armadillo-like helical / PH-like domain superfamily / Armadillo-type fold / ELMO domain / Domain of unknown function DUF3361 / Dedicator of cytokinesis protein 2 / DHR-1 domain / DHR-2 domain / Engulfment and cell motility protein 1 ...Dedicator of cytokinesis / Armadillo-like helical / PH-like domain superfamily / Armadillo-type fold / ELMO domain / Domain of unknown function DUF3361 / Dedicator of cytokinesis protein 2 / DHR-1 domain / DHR-2 domain / Engulfment and cell motility protein 1 / Dedicator of cytokinesis, N-terminal domain / C2 domain superfamily / SH3-like domain superfamily / Dedicator of cytokinesis, N-terminal, subdomain 1 / Dedicator of cytokinesis, C-terminal, lobe A / Dedicator of cytokinesis, C-terminal, lobe C / Pleckstrin homology domain / SH3 domain / Dedicator of cytokinesis, C-terminal
Engulfment and cell motility protein 1 / Dedicator of cytokinesis protein 2
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 5.5 Å
AuthorsChang, L. / Yang, J. / Chang, J.H. / Zhang, Z. / Boland, A. / McLaughlin, S.H. / Abu-Thuraia, A. / Killoran, R.C. / Smith, M.J. / Cote, J.F. / Barford, D.
Funding support United Kingdom, 3items
OrganizationGrant numberCountry
Medical Research Council (United Kingdom)MC_UP_1201/6 United Kingdom
Cancer Research UKC576/A14109 United Kingdom
Marie Sklodowska-Curie Actions, FragNET ITN657725 United Kingdom
CitationJournal: Nat Commun / Year: 2020
Title: Structure of the DOCK2-ELMO1 complex provides insights into regulation of the auto-inhibited state.
Authors: Leifu Chang / Jing Yang / Chang Hwa Jo / Andreas Boland / Ziguo Zhang / Stephen H McLaughlin / Afnan Abu-Thuraia / Ryan C Killoran / Matthew J Smith / Jean-Francois Côté / David Barford /
Abstract: DOCK (dedicator of cytokinesis) proteins are multidomain guanine nucleotide exchange factors (GEFs) for RHO GTPases that regulate intracellular actin dynamics. DOCK proteins share catalytic (DOCK) ...DOCK (dedicator of cytokinesis) proteins are multidomain guanine nucleotide exchange factors (GEFs) for RHO GTPases that regulate intracellular actin dynamics. DOCK proteins share catalytic (DOCK) and membrane-associated (DOCK) domains. The structurally-related DOCK1 and DOCK2 GEFs are specific for RAC, and require ELMO (engulfment and cell motility) proteins for function. The N-terminal RAS-binding domain (RBD) of ELMO (ELMO) interacts with RHOG to modulate DOCK1/2 activity. Here, we determine the cryo-EM structures of DOCK2-ELMO1 alone, and as a ternary complex with RAC1, together with the crystal structure of a RHOG-ELMO2 complex. The binary DOCK2-ELMO1 complex adopts a closed, auto-inhibited conformation. Relief of auto-inhibition to an active, open state, due to a conformational change of the ELMO1 subunit, exposes binding sites for RAC1 on DOCK2, and RHOG and BAI GPCRs on ELMO1. Our structure explains how up-stream effectors, including DOCK2 and ELMO1 phosphorylation, destabilise the auto-inhibited state to promote an active GEF.
Validation Report
SummaryFull reportAbout validation report
History
DepositionNov 15, 2019Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jul 29, 2020Provider: repository / Type: Initial release

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Structure visualization

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Assembly

Deposited unit
A: Dedicator of cytokinesis protein 2
B: Engulfment and cell motility protein 1
D: Dedicator of cytokinesis protein 2


Theoretical massNumber of molelcules
Total (without water)475,6963
Polymers475,6963
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: cross-linking, 0.025% glutaraldehyde for 10 min
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TypeNameSymmetry operationNumber
identity operation1_5551
Buried area6480 Å2
ΔGint-51 kcal/mol
Surface area128600 Å2
MethodPISA

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Components

#1: Protein Dedicator of cytokinesis protein 2


Mass: 195902.516 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DOCK2, KIAA0209 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q92608
#2: Protein Engulfment and cell motility protein 1 / ELMO1 / Protein ced-12 homolog / ELMO1


Mass: 83891.328 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ELMO1, KIAA0281 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q92556

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Binary complex of DOCK2-ELMO1 / Type: COMPLEX / Entity ID: 1,2 / Source: RECOMBINANT
Molecular weightValue: 0.55 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHepesC8H18N2O4S1
2200 mMsodium chlorideNaClSodium chloride1
SpecimenConc.: 0.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
EM imaging opticsEnergyfilter name: GIF Quantum ER / Energyfilter slit width: 20 eV
Image scansMovie frames/image: 20

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.16_3549refinement
PHENIX1.16_3549refinement
EM software
IDNameVersionCategory
1RELION1.4particle selection
2EPUimage acquisition
4GctfCTF correction
12RELION1.43D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 5.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 154428 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 304.8 Å2
Refine LS restraints
Refinement-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.02119006
ELECTRON MICROSCOPYf_angle_d1.345525794
ELECTRON MICROSCOPYf_chiral_restr0.05912977
ELECTRON MICROSCOPYf_plane_restr0.00813331
ELECTRON MICROSCOPYf_dihedral_angle_d6.410211476

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