|Entry||Database: EMDB / ID: EMD-10834|
|Title||Tobacco mosaic virus (TMV) structure determined by a hybrid STA-SPA workflow|
|Sample||Tobacco mosaic virus:|
|Biological species||Tobacco mosaic virus|
|Method||subtomogram averaging / cryo EM / Resolution: 5.24 Å|
|Authors||Sanchez RM / Zhang Y / Chen W / Dietrich L / Kudryashev M|
|Funding support|| Germany, 1 items |
|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.
|Structure viewer||EM map: |
Downloads & links
|File||Download / File: emd_10834.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 2.2 Å|
|Symmetry||Space group: 1|
CCP4 map header:
|Projections & Slices|
-Half map: half map 1
|Annotation||half map 1|
|Projections & Slices|
-Half map: half map 2
-Entire Tobacco mosaic virus
|Entire||Name: Tobacco mosaic virus / Number of components: 1|
-Component #1: virus, Tobacco mosaic virus
|Virus||Name: Tobacco mosaic virus / Class: VIRION / Empty: No / Enveloped: No / Isolate: STRAIN|
|Species||Species: Tobacco mosaic virus|
|Shell #1||Name of element: Capsid protein / Diameter: 180.0 Å|
|Specimen||Specimen state: helical array / Method: cryo EM|
|Helical parameters||Axial symmetry: C1 (asymmetric) / Delta z: 1.41 Å / Delta phi: 22.033 %deg;|
|Sample solution||Specimen conc.: 33 mg/mL / pH: 7.4|
|Vitrification||Cryogen name: ETHANE|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Imaging||Microscope: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 2.3 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Magnification: 64000 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: -3.5 - -4.5 nm|
|Specimen Holder||Model: FEI TITAN KRIOS AUTOGRID HOLDER|
|Camera||Detector: GATAN K2 SUMMIT (4k x 4k)|
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