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- EMDB-8071: Map of Venezuelan Equine Encephalitis Virus by cryoSPT (subtomogr... -

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Entry
Database: EMDB / ID: EMD-8071
TitleMap of Venezuelan Equine Encephalitis Virus by cryoSPT (subtomogram averaging)
Map dataNone
SampleVenezuelan Equine Encephalitis Virus != Venezuelan equine encephalitis virus

Venezuelan Equine Encephalitis Virus

  • Virus: Venezuelan equine encephalitis virus
Biological speciesVenezuelan equine encephalitis virus
Methodsubtomogram averaging / cryo EM / Resolution: 13.0 Å
AuthorsGalaz-Montoya JG
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P41GM103832 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI057156 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM080139 United States
Cancer Prevention and Research Institute of Texas (CPRIT)RP140113 United States
Robert WelchQ1242 United States
CitationJournal: J Struct Biol / Year: 2016
Title: Alignment algorithms and per-particle CTF correction for single particle cryo-electron tomography.
Authors: Jesús G Galaz-Montoya / Corey W Hecksel / Philip R Baldwin / Eryu Wang / Scott C Weaver / Michael F Schmid / Steven J Ludtke / Wah Chiu /
Abstract: Single particle cryo-electron tomography (cryoSPT) extracts features from cryo-electron tomograms, followed by 3D classification, alignment and averaging to generate improved 3D density maps of such ...Single particle cryo-electron tomography (cryoSPT) extracts features from cryo-electron tomograms, followed by 3D classification, alignment and averaging to generate improved 3D density maps of such features. Robust methods to correct for the contrast transfer function (CTF) of the electron microscope are necessary for cryoSPT to reach its resolution potential. Many factors can make CTF correction for cryoSPT challenging, such as lack of eucentricity of the specimen stage, inherent low dose per image, specimen charging, beam-induced specimen motions, and defocus gradients resulting both from specimen tilting and from unpredictable ice thickness variations. Current CTF correction methods for cryoET make at least one of the following assumptions: that the defocus at the center of the image is the same across the images of a tiltseries, that the particles all lie at the same Z-height in the embedding ice, and/or that the specimen, the cryo-electron microscopy (cryoEM) grid and/or the carbon support are flat. These experimental conditions are not always met. We have developed a CTF correction algorithm for cryoSPT without making any of the aforementioned assumptions. We also introduce speed and accuracy improvements and a higher degree of automation to the subtomogram averaging algorithms available in EMAN2. Using motion-corrected images of isolated virus particles as a benchmark specimen, recorded with a DE20 direct detection camera, we show that our CTF correction and subtomogram alignment routines can yield subtomogram averages close to 4/5 Nyquist frequency of the detector under our experimental conditions.
History
DepositionFeb 7, 2016-
Header (metadata) releaseApr 13, 2016-
Map releaseApr 13, 2016-
UpdateJan 29, 2020-
Current statusJan 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 1
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_8071.png

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Map

FileDownload / File: emd_8071.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNone
Voxel sizeX=Y=Z: 5 Å
Density
Contour LevelBy AUTHOR: 2 / Movie #1: 1
Minimum - Maximum-8.703346 - 10.424462
Average (Standard dev.)0.110686526 (±0.7516296)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-128-128-128
Dimensions256256256
Spacing256256256
CellA=B=C: 1280.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z555
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z1280.0001280.0001280.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-38-19-20
NX/NY/NZ858082
MAP C/R/S123
start NC/NR/NS-128-128-128
NC/NR/NS256256256
D min/max/mean-8.70310.4240.111

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Supplemental data

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Sample components

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Entire : Venezuelan Equine Encephalitis Virus

EntireName: Venezuelan Equine Encephalitis Virus
Components
  • Virus: Venezuelan equine encephalitis virus

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Supramolecule #1: Venezuelan equine encephalitis virus

SupramoleculeName: Venezuelan equine encephalitis virus / type: virus / ID: 1 / Parent: 0 / NCBI-ID: 11036 / Sci species name: Venezuelan equine encephalitis virus / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: No
Host (natural)Organism: Horses (horses)
Virus shellShell ID: 1 / Name: Capsid / Diameter: 700.0 Å / T number (triangulation number): 4

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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
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - #0 - Film type ID: 1 / Support film - #0 - Material: CARBON / Support film - #0 - topology: HOLEY / Support film - #1 - Film type ID: 2 / Support film - #1 - Material: CARBON / Support film - #1 - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Instrument: GATAN CRYOPLUNGE 3

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Electron microscopy

MicroscopeJEOL 3200FSC
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Specialist opticsEnergy filter - Name: JEOL Omega / Energy filter - Upper energy threshold: 25 eV
Sample stageSpecimen holder model: JEOL / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: DIRECT ELECTRON DE-20 (5k x 3k) / Detector mode: INTEGRATING / Average exposure time: 0.5 sec. / Average electron dose: 3.0 e/Å2

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Image processing

ExtractionNumber tomograms: 6 / Number images used: 516 / Method: Manual picking / Software - Name: EMAN2 (ver. 2.1)
CTF correctionSoftware - Name: EMAN2 (ver. 2.1)
Final 3D classificationNumber classes: 1 / Avg.num./class: 387 / Software - Name: EMAN2 (ver. 2.1)
Final angle assignmentType: OTHER / Software - Name: EMAN2 (ver. 2.1)
Details: Cross correlation of volumes to find best relative orientations
Final reconstructionNumber classes used: 1 / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 13.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: EMAN2 (ver. 2.1) / Number subtomograms used: 387
FSC plot (resolution estimation)

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