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- EMDB-3365: electron density map of murine leukaemia virus envelope glycoprot... -

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Basic information

Entry
Database: EMDB / ID: EMD-3365
Titleelectron density map of murine leukaemia virus envelope glycoprotein as reconstructed by subtomogram averaging on murine leukemia virus virus like particles (3 plasmid system) produced in COS1 cells
Map datareconstruction of virus like particle bound murine leukemia virus Env protein
Sample
  • Sample: murine leukemia virus Env protein on murine leukemia virus virus like particles
  • Protein or peptide: murine leukemia virus Env protein
Keywordsmurine leukemia virus / retrovirus / envelope glycoprotein / cryo electron tomography / subtomogram averaging
Biological speciesMurine leukemia virus
Methodsubtomogram averaging / cryo EM / Resolution: 22.0 Å
AuthorsRiedel C / Vasishtan D / Siebert CA / Whittle C / Lehmann MJ / Mothes W / Grunewald K
CitationJournal: J Struct Biol / Year: 2017
Title: Native structure of a retroviral envelope protein and its conformational change upon interaction with the target cell.
Authors: Christiane Riedel / Daven Vasishtan / C Alistair Siebert / Cathy Whittle / Maik J Lehmann / Walther Mothes / Kay Grünewald /
Abstract: Enveloped viruses enter their host cells by membrane fusion. The process of attachment and fusion in retroviruses is mediated by a single viral envelope glycoprotein (Env). Conformational changes of ...Enveloped viruses enter their host cells by membrane fusion. The process of attachment and fusion in retroviruses is mediated by a single viral envelope glycoprotein (Env). Conformational changes of Env in the course of fusion are a focus of intense studies. Here we provide further insight into the changes occurring in retroviral Env during its initial interaction with the cell, employing murine leukemia virus (MLV) as model system. We first determined the structure of both natively membrane anchored MLV Env and MLV Env tagged with YFP in the proline rich region (PRR) by electron cryo tomography (cET) and sub-volume averaging. At a resolution of ∼20Å, native MLV Env presents as a hollow trimer (height ∼85Å, diameter ∼120Å) composed of step-shaped protomers. The major difference to the YFP-tagged protein was in regions outside of the central trimer. Next, we focused on elucidating the changes in MLV Env upon interaction with a host cell. Virus interaction with the plasma membrane occurred over a large surface and Env clustering on the binding site was observed. Sub-volume averaging did yield a low-resolution structure of Env interacting with the cell, which had lost its threefold symmetry and was elongated by ∼35Å in comparison to the unbound protein. This indicates a major rearrangement of Env upon host cell binding. At the site of virus interaction, the otherwise clearly defined bilayer structure of the host cell plasma membrane was much less evident, indicative of integral membrane protein accumulation and/or a change in membrane lipid composition.
History
DepositionMar 8, 2016-
Header (metadata) releaseApr 13, 2016-
Map releaseJul 20, 2016-
UpdateJul 26, 2017-
Current statusJul 26, 2017Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 8.65
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 8.65
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3365.tif

Downloads & links

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Map

FileDownload / File: emd_3365.map.gz / Format: CCP4 / Size: 825.2 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationreconstruction of virus like particle bound murine leukemia virus Env protein
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
4.6 Å/pix.
x 60 pix.
= 276. Å		4.6 Å/pix.
x 60 pix.
= 276. Å		4.6 Å/pix.
x 60 pix.
= 276. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 4.6 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 8.65 / Movie #1: 8.65
Minimum - Maximum0.00273132 - 14.4690361
Average (Standard dev.)6.44099903 (±1.24479997)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions606060
Spacing606060
CellA=B=C: 276.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.64.64.6
M x/y/z606060
origin x/y/z0.0000.0000.000
length x/y/z276.000276.000276.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS606060
D min/max/mean0.00314.4696.441

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Supplemental data

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Sample components

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Entire : murine leukemia virus Env protein on murine leukemia virus virus ...

EntireName: murine leukemia virus Env protein on murine leukemia virus virus like particles
Components
  • Sample: murine leukemia virus Env protein on murine leukemia virus virus like particles
  • Protein or peptide: murine leukemia virus Env protein

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Supramolecule #1000: murine leukemia virus Env protein on murine leukemia virus virus ...

SupramoleculeName: murine leukemia virus Env protein on murine leukemia virus virus like particles
type: sample / ID: 1000
Details: purified virus like particles produced in COS1 cells
Oligomeric state: trimer / Number unique components: 1
Molecular weightTheoretical: 226 KDa

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Macromolecule #1: murine leukemia virus Env protein

MacromoleculeName: murine leukemia virus Env protein / type: protein_or_peptide / ID: 1 / Details: imaged on intact virus like particles / Number of copies: 3 / Oligomeric state: trimer / Recombinant expression: No
Source (natural)Organism: Murine leukemia virus / Strain: Friend's murine leukemia virus
Molecular weightTheoretical: 226 KDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferpH: 7.4 / Details: DMEM + 10% FSC
GridDetails: C-flat copper grids (Protochips, CF-2/1-2C)
VitrificationCryogen name: ETHANE-PROPANE MIXTURE / Instrument: OTHER / Method: manually blotted for 3sec

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Electron microscopy

MicroscopeFEI POLARA 300
Specialist opticsEnergy filter - Name: Gatan QUANTUM 964 postcolumn energy filter
Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV
DateAug 6, 2014
Image recordingCategory: CCD / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 60 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 95000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.0 µm / Nominal defocus min: 1.5 µm
Sample stageSpecimen holder model: OTHER / Tilt series - Axis1 - Min angle: -45 ° / Tilt series - Axis1 - Max angle: 45 °
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Detailsparticles were picked manually
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 22.0 Å / Resolution method: OTHER / Software - Name: motioncorr, IMOD, TomoCTF, PEET, bsoft
Details: Images were aligned using motioncorr. Tomograms were reconstructed using weighted back projection. CTF correction was performed employing TomoCTF. PEET was used for the generation of the ...Details: Images were aligned using motioncorr. Tomograms were reconstructed using weighted back projection. CTF correction was performed employing TomoCTF. PEET was used for the generation of the subtomogram average, FSC was calculated using bsoft.
Number subtomograms used: 2358
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

1aol


Chain - Chain ID: A
SoftwareName: Chimera
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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