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- EMDB-10834: Tobacco mosaic virus (TMV) structure determined by a hybrid STA-S... -

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Basic information

Entry
Database: EMDB / ID: EMD-10834
TitleTobacco mosaic virus (TMV) structure determined by a hybrid STA-SPA workflow
Map datarefine map
Sample
  • Virus: Tobacco mosaic virus
Function / homologyTobacco mosaic virus-like, coat protein / Tobacco mosaic virus-like, coat protein superfamily / Virus coat protein (TMV like) / helical viral capsid / structural molecule activity / identical protein binding / Capsid protein / Capsid protein
Function and homology information
Biological speciesTobacco mosaic virus
Methodsubtomogram averaging / cryo EM / Resolution: 5.24 Å
AuthorsSanchez RM / Zhang Y / Chen W / Dietrich L / Kudryashev M
Funding support Germany, 1 items
OrganizationGrant numberCountry
Alexander von Humboldt FoundationSFB807 Germany
Citation
Journal: Nat Commun / Year: 2020
Title: Subnanometer-resolution structure determination in situ by hybrid subtomogram averaging - single particle cryo-EM.
Authors: Ricardo M Sanchez / Yingyi Zhang / Wenbo Chen / Lea Dietrich / Mikhail Kudryashev /
Abstract: Cryo-electron tomography combined with subtomogram averaging (StA) has yielded high-resolution structures of macromolecules in their native context. However, high-resolution StA is not commonplace ...Cryo-electron tomography combined with subtomogram averaging (StA) has yielded high-resolution structures of macromolecules in their native context. However, high-resolution StA is not commonplace due to beam-induced sample drift, images with poor signal-to-noise ratios (SNR), challenges in CTF correction, and limited particle number. Here we address these issues by collecting tilt series with a higher electron dose at the zero-degree tilt. Particles of interest are then located within reconstructed tomograms, processed by conventional StA, and then re-extracted from the high-dose images in 2D. Single particle analysis tools are then applied to refine the 2D particle alignment and generate a reconstruction. Use of our hybrid StA (hStA) workflow improved the resolution for tobacco mosaic virus from 7.2 to 4.4 Å and for the ion channel RyR1 in crowded native membranes from 12.9 to 9.1 Å. These resolution gains make hStA a promising approach for other StA projects aimed at achieving subnanometer resolution.
#2: Journal: Biorxiv / Year: 2020
Title: Subnanometer-resolution structure determination in situ by a hybrid subtomogram averaging - single particle cryoEM - workflow
Authors: Sanchez RM / Zhang Y / Chen W / Dietrich L / Kudryashev M
History
DepositionApr 6, 2020-
Header (metadata) releaseMay 20, 2020-
Map releaseMay 20, 2020-
UpdateAug 12, 2020-
Current statusAug 12, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.035
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.035
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10834.png

Downloads & links

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Map

FileDownload / File: emd_10834.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationrefine map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.2 Å/pix.
x 200 pix.
= 440. Å		2.2 Å/pix.
x 200 pix.
= 440. Å		2.2 Å/pix.
x 200 pix.
= 440. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.2 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.035 / Movie #1: 0.035
Minimum - Maximum-0.018232355 - 0.060033157
Average (Standard dev.)-0.0005931545 (±0.008035485)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 440.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.22.22.2
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z440.000440.000440.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.0180.060-0.001

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Supplemental data

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Mask #1

Fileemd_10834_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 1

Fileemd_10834_half_map_1.map
Annotationhalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 2

Fileemd_10834_half_map_2.map
Annotationhalf map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Tobacco mosaic virus

EntireName: Tobacco mosaic virus
Components
  • Virus: Tobacco mosaic virus

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Supramolecule #1: Tobacco mosaic virus

SupramoleculeName: Tobacco mosaic virus / type: virus / ID: 1 / Parent: 0 / NCBI-ID: 12242 / Sci species name: Tobacco mosaic virus / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: No / Virus empty: No
Virus shellShell ID: 1 / Name: Capsid protein / Diameter: 180.0 Å

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statehelical array

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Sample preparation

Concentration33 mg/mL
BufferpH: 7.4
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 2.3 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: -0.0045 µm / Nominal defocus min: -0.0035 µm / Nominal magnification: 64000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 1.41 Å
Applied symmetry - Helical parameters - Δ&Phi: 22.033 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 5.24 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number subtomograms used: 17998
ExtractionNumber tomograms: 4 / Number images used: 20214
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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