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- EMDB-4060: Negative Stain electron microscopy of GIF:IL2 complex -

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Basic information

Entry
Database: EMDB / ID: EMD-4060
TitleNegative Stain electron microscopy of GIF:IL2 complex
Map data
Sample
  • Complex: Complex of GIF:IL2
    • Complex: Ovine cytokine Interleukin-2
    • Complex: Orf virus decoy receptor GIF.
Biological speciesOvis aries (sheep) / Orf virus
Methodsingle particle reconstruction / negative staining / Resolution: 17.0 Å
AuthorsKandiah E / Gutsche I
CitationJournal: Nat Commun / Year: 2016
Title: Structural basis of GM-CSF and IL-2 sequestration by the viral decoy receptor GIF.
Authors: Jan Felix / Eaazhisai Kandiah / Steven De Munck / Yehudi Bloch / Gydo C P van Zundert / Kris Pauwels / Ann Dansercoer / Katka Novanska / Randy J Read / Alexandre M J J Bonvin / Bjorn ...Authors: Jan Felix / Eaazhisai Kandiah / Steven De Munck / Yehudi Bloch / Gydo C P van Zundert / Kris Pauwels / Ann Dansercoer / Katka Novanska / Randy J Read / Alexandre M J J Bonvin / Bjorn Vergauwen / Kenneth Verstraete / Irina Gutsche / Savvas N Savvides /
Abstract: Subversion of the host immune system by viruses is often mediated by molecular decoys that sequester host proteins pivotal to mounting effective immune responses. The widespread mammalian pathogen ...Subversion of the host immune system by viruses is often mediated by molecular decoys that sequester host proteins pivotal to mounting effective immune responses. The widespread mammalian pathogen parapox Orf virus deploys GIF, a member of the poxvirus immune evasion superfamily, to antagonize GM-CSF (granulocyte macrophage colony-stimulating factor) and IL-2 (interleukin-2), two pleiotropic cytokines of the mammalian immune system. However, structural and mechanistic insights into the unprecedented functional duality of GIF have remained elusive. Here we reveal that GIF employs a dimeric binding platform that sequesters two copies of its target cytokines with high affinity and slow dissociation kinetics to yield distinct complexes featuring mutually exclusive interaction footprints. We illustrate how GIF serves as a competitive decoy receptor by leveraging binding hotspots underlying the cognate receptor interactions of GM-CSF and IL-2, without sharing any structural similarity with the cytokine receptors. Our findings contribute to the tracing of novel molecular mimicry mechanisms employed by pathogenic viruses.
History
DepositionJul 18, 2016-
Header (metadata) releaseOct 12, 2016-
Map releaseAug 2, 2017-
UpdateAug 2, 2017-
Current statusAug 2, 2017Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.07
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.07
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_4060.png

Downloads & links

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Map

FileDownload / File: emd_4060.map.gz / Format: CCP4 / Size: 1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 3.5 Å
Density
Contour LevelBy AUTHOR: 0.07 / Movie #1: 0.07
Minimum - Maximum-0.117920734 - 0.17561504
Average (Standard dev.)0.0021575294 (±0.01784704)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions646464
Spacing646464
CellA=B=C: 224.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.53.53.5
M x/y/z646464
origin x/y/z0.0000.0000.000
length x/y/z224.000224.000224.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS646464
D min/max/mean-0.1180.1760.002

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Supplemental data

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Sample components

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Entire : Complex of GIF:IL2

EntireName: Complex of GIF:IL2
Components
  • Complex: Complex of GIF:IL2
    • Complex: Ovine cytokine Interleukin-2
    • Complex: Orf virus decoy receptor GIF.

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Supramolecule #1: Complex of GIF:IL2

SupramoleculeName: Complex of GIF:IL2 / type: complex / ID: 1 / Parent: 0
Details: Ovine cytokine Interleukin-2 is complexed with orf virus decoy receptor GIF.
Source (natural)Organism: Ovis aries (sheep)

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Supramolecule #2: Ovine cytokine Interleukin-2

SupramoleculeName: Ovine cytokine Interleukin-2 / type: complex / ID: 2 / Parent: 1
Source (natural)Organism: Ovis aries (sheep)
Recombinant expressionOrganism: HEK293T (human) / Recombinant plasmid: pHLsec

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Supramolecule #3: Orf virus decoy receptor GIF.

SupramoleculeName: Orf virus decoy receptor GIF. / type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Orf virus
Recombinant expressionOrganism: HEK293T (human) / Recombinant plasmid: pHLsec

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5
StainingType: NEGATIVE / Material: Uranyl acetate
Details: Negatively stained EM specimens were prepared using a carbon-sandwich technique and 2% (w/v) uranyl-acetate stain.

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Electron microscopy

MicroscopeJEOL 1200EXII
Electron beamAcceleration voltage: 100 kV / Electron source: LAB6
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal magnification: 40000
Image recordingFilm or detector model: KODAK SO-163 FILM / Average electron dose: 10.0 e/Å2

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Image processing

Startup modelType of model: INSILICO MODEL / In silico model: Ab-inition 3D reconstruction using VIPER
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 17.0 Å / Resolution method: FSC 0.5 CUT-OFF / Number images used: 24324

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Atomic model buiding 1

RefinementProtocol: RIGID BODY FIT

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