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- EMDB-3874: Cryo-electron tomogram of Ebola virus -

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Basic information

Entry
Database: EMDB / ID: EMD-3874
TitleCryo-electron tomogram of Ebola virus
Map dataA 4x binned representative tomogram of intact Ebola virus virions
Sample
  • Virus: Ebola virus - Mayinga, Zaire, 1976
Biological speciesEbola virus - Mayinga, Zaire, 1976
Methodelectron tomography / cryo EM
AuthorsWan W / Kolesnikova L / Clarke M / Koehler A / Noda T / Becker S / Briggs JAG
Funding support Germany, 3 items
OrganizationGrant numberCountry
European Research CouncilERC-CoG-648432 MEMBRANEFUSION Germany
German Research FoundationSonderforschungsbereich 1021 Germany
European Molecular Biology OrganizationALTF 748-2014 Germany
CitationJournal: Nature / Year: 2017
Title: Structure and assembly of the Ebola virus nucleocapsid.
Authors: William Wan / Larissa Kolesnikova / Mairi Clarke / Alexander Koehler / Takeshi Noda / Stephan Becker / John A G Briggs /
Abstract: Ebola and Marburg viruses are filoviruses: filamentous, enveloped viruses that cause haemorrhagic fever. Filoviruses are within the order Mononegavirales, which also includes rabies virus, measles ...Ebola and Marburg viruses are filoviruses: filamentous, enveloped viruses that cause haemorrhagic fever. Filoviruses are within the order Mononegavirales, which also includes rabies virus, measles virus, and respiratory syncytial virus. Mononegaviruses have non-segmented, single-stranded negative-sense RNA genomes that are encapsidated by nucleoprotein and other viral proteins to form a helical nucleocapsid. The nucleocapsid acts as a scaffold for virus assembly and as a template for genome transcription and replication. Insights into nucleoprotein-nucleoprotein interactions have been derived from structural studies of oligomerized, RNA-encapsidating nucleoprotein, and cryo-electron microscopy of nucleocapsid or nucleocapsid-like structures. There have been no high-resolution reconstructions of complete mononegavirus nucleocapsids. Here we apply cryo-electron tomography and subtomogram averaging to determine the structure of Ebola virus nucleocapsid within intact viruses and recombinant nucleocapsid-like assemblies. These structures reveal the identity and arrangement of the nucleocapsid components, and suggest that the formation of an extended α-helix from the disordered carboxy-terminal region of nucleoprotein-core links nucleoprotein oligomerization, nucleocapsid condensation, RNA encapsidation, and accessory protein recruitment.
History
DepositionSep 13, 2017-
Header (metadata) releaseNov 22, 2017-
Map releaseNov 22, 2017-
UpdateDec 4, 2019-
Current statusDec 4, 2019Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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  • Simplified surface model
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3874.png

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Map

FileDownload / File: emd_3874.map.gz / Format: CCP4 / Size: 409.8 MB / Type: IMAGE STORED AS SIGNED INTEGER (2 BYTES)
AnnotationA 4x binned representative tomogram of intact Ebola virus virions
Voxel sizeX=Y=Z: 7.12 Å
Density
Minimum - Maximum-40 - 59
Average (Standard dev.)3.0713525 (±3.417756)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin0042
Dimensions927927250
Spacing927927250
CellA: 6600.2397 Å / B: 6600.2397 Å / C: 1780.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Integer*27
Å/pix. X/Y/Z7.127.127.12
M x/y/z927927250
origin x/y/z0.0000.0000.000
length x/y/z6600.2406600.2401780.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ300300300
MAP C/R/S123
start NC/NR/NS0042
NC/NR/NS927927250
D min/max/mean-40.00059.0003.071

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Supplemental data

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Sample components

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Entire : Ebola virus - Mayinga, Zaire, 1976

EntireName: Ebola virus - Mayinga, Zaire, 1976
Components
  • Virus: Ebola virus - Mayinga, Zaire, 1976

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Supramolecule #1: Ebola virus - Mayinga, Zaire, 1976

SupramoleculeName: Ebola virus - Mayinga, Zaire, 1976 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Details: Virus was isolated from infected VeroE6 cells. Purified viruses were fixed with paraformaldehyde.
NCBI-ID: 128952 / Sci species name: Ebola virus - Mayinga, Zaire, 1976 / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: No
Virus shellShell ID: 1 / Name: Nucleocapsid / Diameter: 280.0 Å

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation statehelical array

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Sample preparation

BufferpH: 7.4
Details: Virus was purified into Dulbecco's modified Eagle's medium (DMEM) with 4% paraformaldehyde
GridModel: C-flat 2/1 3C / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 20.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.039 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Instrument: FEI VITROBOT MARK II
SectioningOther: NO SECTIONING
Fiducial markerManufacturer: UMC Utrecht / Diameter: 10 nm

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 81000
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Lower energy threshold: -10 eV / Energy filter - Upper energy threshold: 10 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 3708 pixel / Digitization - Dimensions - Height: 3708 pixel / Digitization - Frames/image: 1-5 / Average exposure time: 2.0 sec. / Average electron dose: 2.9 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware:
Namedetails
CTFFIND (ver. 4)defocus determination
CTFPHASEFLIPCTF-correction

Details: CTF amplitude correction was performed during the wedge-weighted subtomogram averaging step.
Final reconstructionAlgorithm: BACK PROJECTION / Software - Name: IMOD / Number images used: 41
DetailsFrames were aligned using K2Align software, based off the MotionCorr algorithm. Tomograms were reconstructed with IMOD, using stripwise CTF-correction and weighted back projection.

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