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- EMDB-22164: Assembly intermediates of orthoreovirus captured in the cell -

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Basic information

Entry
Database: EMDB / ID: EMD-22164
TitleAssembly intermediates of orthoreovirus captured in the cell
Map data
Samplereovirus SLP
Function / homology
Function and homology information


host cell surface binding / viral outer capsid / viral entry via permeabilization of inner membrane / permeabilization of host organelle membrane involved in viral entry into host cell / suppression by virus of host PKR activity / host cell mitochondrion / host cell endoplasmic reticulum / viral life cycle / viral capsid / regulation of translation ...host cell surface binding / viral outer capsid / viral entry via permeabilization of inner membrane / permeabilization of host organelle membrane involved in viral entry into host cell / suppression by virus of host PKR activity / host cell mitochondrion / host cell endoplasmic reticulum / viral life cycle / viral capsid / regulation of translation / suppression by virus of host type I interferon-mediated signaling pathway / pathogenesis / host cell plasma membrane / structural molecule activity / RNA binding / membrane / metal ion binding
Mu1 membrane penetration protein, subdomain 1 / Mu1 membrane penetration protein, subdomain 3 / Outer capsid protein Mu1/VP4 / Mu1/VP4 superfamily / Reovirus, outer capsid sigma 3 / Reovirus, outer capsid sigma-3 domain superfamily / Mu1 membrane penetration protein, subdomain 2
Outer capsid protein sigma-3 / Outer capsid protein mu-1
Biological speciesMammalian orthoreovirus 3 Dearing
Methodsubtomogram averaging / cryo EM / Resolution: 5.6 Å
AuthorsSutton G / Sun DP / Fu XF / Kotecha A / Hecksel GW / Clare DK / Zhang P / Stuart D / Boyce M
Funding support United Kingdom, United States, 3 items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MR/N00065X/1 United Kingdom
National Institutes of Health/National Institute of Biomedical Imaging and Bioengineering (NIH/NIBIB)GM082251 United States
Wellcome Trust206422/Z/17/Z United Kingdom
Citation
Journal: Nat Commun / Year: 2020
Title: Assembly intermediates of orthoreovirus captured in the cell.
Authors: Geoff Sutton / Dapeng Sun / Xiaofeng Fu / Abhay Kotecha / Corey W Hecksel / Daniel K Clare / Peijun Zhang / David I Stuart / Mark Boyce /
Abstract: Traditionally, molecular assembly pathways for viruses are inferred from high resolution structures of purified stable intermediates, low resolution images of cell sections and genetic approaches. ...Traditionally, molecular assembly pathways for viruses are inferred from high resolution structures of purified stable intermediates, low resolution images of cell sections and genetic approaches. Here, we directly visualise an unsuspected 'single shelled' intermediate for a mammalian orthoreovirus in cryo-preserved infected cells, by cryo-electron tomography of cellular lamellae. Particle classification and averaging yields structures to 5.6 Å resolution, sufficient to identify secondary structural elements and produce an atomic model of the intermediate, comprising 120 copies each of protein λ1 and σ2. This λ1 shell is 'collapsed' compared to the mature virions, with molecules pushed inwards at the icosahedral fivefolds by ~100 Å, reminiscent of the first assembly intermediate of certain prokaryotic dsRNA viruses. This supports the supposition that these viruses share a common ancestor, and suggests mechanisms for the assembly of viruses of the Reoviridae. Such methodology holds promise for dissecting the replication cycle of many viruses.
#1: Journal: bioRxiv / Year: 2020
Title: Assembly intermediates of orthoreovirus captured in the cell
Authors: Sutton G / Sun DP / Fu XF / Kotecha A / Hecksel GW / Clare DK / Zhang P / Stuart D / Boyce M
Validation ReportPDB-ID: 6zty

SummaryFull reportAbout validation report
History
DepositionJun 15, 2020-
Header (metadata) releaseSep 16, 2020-
Map releaseSep 16, 2020-
UpdateSep 23, 2020-
Current statusSep 23, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 2.8
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 2.8
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6zty
  • Surface level: 2.8
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6zty
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_22164.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.8 Å/pix.
x 256 pix.
= 460.8 Å
1.8 Å/pix.
x 256 pix.
= 460.8 Å
1.8 Å/pix.
x 256 pix.
= 460.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.8 Å
Density
Contour LevelBy AUTHOR: 2.8 / Movie #1: 2.8
Minimum - Maximum-5.3467703 - 8.876596
Average (Standard dev.)0.049125537 (±0.4568551)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 460.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.81.81.8
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z460.800460.800460.800
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ400400400
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-5.3478.8770.049

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Supplemental data

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Sample components

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Entire reovirus SLP

EntireName: reovirus SLP / Number of components: 1

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Component #1: protein, reovirus SLP

ProteinName: reovirus SLP / Recombinant expression: No
SourceSpecies: Mammalian orthoreovirus 3 Dearing

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Experimental details

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Sample preparation

SpecimenSpecimen state: Cell / Method: cryo EM
Sample solutionpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 2 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: subtomogram averaging / Applied symmetry: C6 (6 fold cyclic) / Number of subtomograms: 3039
3D reconstructionSoftware: emClarity / Resolution: 5.6 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement protocol: flexible
Output model

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