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- EMDB-5204: Average of 50 Epsilon15 subtomograms using electron cryo-tomography -

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Basic information

Entry
Database: EMDB / ID: EMD-5204
TitleAverage of 50 Epsilon15 subtomograms using electron cryo-tomography
Map dataThis is an image of a surface rendered top-view of Epsilon15
Sample
  • Sample: Epsilon15Salmonella virus Epsilon15
  • Virus: epsilon15 (virus)
KeywordsElectron tomography / Phase plate / Bacteriophage
Biological speciesepsilon15 (virus)
Methodsubtomogram averaging / cryo EM / Resolution: 38.0 Å
AuthorsMurata K / Liu X / Danev R / Jakana J / Schmid MF / King J / Nagayama K / Chiu W
CitationJournal: Structure / Year: 2010
Title: Zernike phase contrast cryo-electron microscopy and tomography for structure determination at nanometer and subnanometer resolutions.
Authors: Kazuyoshi Murata / Xiangan Liu / Radostin Danev / Joanita Jakana / Michael F Schmid / Jonathan King / Kuniaki Nagayama / Wah Chiu /
Abstract: Zernike phase contrast cryo-electron microscopy (ZPC-cryoEM) is an emerging technique that is capable of producing higher image contrast than conventional cryoEM. By combining this technique with ...Zernike phase contrast cryo-electron microscopy (ZPC-cryoEM) is an emerging technique that is capable of producing higher image contrast than conventional cryoEM. By combining this technique with advanced image processing methods, we achieved subnanometer resolution for two biological specimens: 2D bacteriorhodopsin crystal and epsilon15 bacteriophage. For an asymmetric reconstruction of epsilon15 bacteriophage, ZPC-cryoEM can reduce the required amount of data by a factor of approximately 3, compared with conventional cryoEM. The reconstruction was carried out to 13 A resolution without the need to correct the contrast transfer function. New structural features at the portal vertex of the epsilon15 bacteriophage are revealed in this reconstruction. Using ZPC cryo-electron tomography (ZPC-cryoET), a similar level of data reduction and higher resolution structures of epsilon15 bacteriophage can be obtained relative to conventional cryoET. These results show quantitatively the benefits of ZPC-cryoEM and ZPC-cryoET for structural determinations of macromolecular machines at nanometer and subnanometer resolutions.
History
DepositionJun 30, 2010-
Header (metadata) releaseSep 1, 2010-
Map releaseSep 1, 2010-
UpdateJul 8, 2011-
Current statusJul 8, 2011Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.2
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.2
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_5204.map.gz / Format: CCP4 / Size: 37.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThis is an image of a surface rendered top-view of Epsilon15
Voxel sizeX=Y=Z: 4.88 Å
Density
Contour LevelBy AUTHOR: 0.2 / Movie #1: 0.2
Minimum - Maximum-2.66289 - 5.11135
Average (Standard dev.)0.0795056 (±0.424669)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions216216216
Spacing216216216
CellA=B=C: 1054.08 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.884.884.88
M x/y/z216216216
origin x/y/z0.0000.0000.000
length x/y/z1054.0801054.0801054.080
α/β/γ90.00090.00090.000
start NX/NY/NZ-99-99-99
NX/NY/NZ200200200
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS216216216
D min/max/mean-2.6635.1110.080

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Supplemental data

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Sample components

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Entire : Epsilon15

EntireName: Epsilon15 (virus)
Components
  • Sample: Epsilon15Salmonella virus Epsilon15
  • Virus: epsilon15 (virus)

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Supramolecule #1000: Epsilon15

SupramoleculeName: Epsilon15 / type: sample / ID: 1000 / Details: The samples was monodisperse / Number unique components: 6
Molecular weightTheoretical: 500 KDa

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Supramolecule #1: epsilon15

SupramoleculeName: epsilon15 / type: virus / ID: 1 / Name.synonym: bacteriophage / Sci species name: epsilon15 / Database: NCBI / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: No / Virus empty: No / Syn species name: bacteriophage
Host (natural)Organism: Salmonella (bacteria) / synonym: BACTERIA(EUBACTERIA)
Virus shellShell ID: 1 / Diameter: 700 Å / T number (triangulation number): 7

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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: 7.5 / Details: 10 mM Tris pH7.5, 25 mM NaCl and 5 mM MgCl2
GridDetails: Quantifoil R2/2 grid
VitrificationCryogen name: ETHANE / Instrument: OTHER / Details: Vitrification instrument: vitrobot / Method: Blot for 2 seconds before plunging

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Electron microscopy

MicroscopeJEOL 2200FS
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 30000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 4.3 mm / Nominal magnification: 20000
Specialist opticsEnergy filter - Name: JEOL / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV
Sample stageSpecimen holder: Eucentric / Specimen holder model: GATAN LIQUID NITROGEN / Tilt series - Axis1 - Min angle: -70 ° / Tilt series - Axis1 - Max angle: 70 °
TemperatureAverage: 90 K
DetailsWeak beam illumination
Image recordingCategory: CCD / Film or detector model: GENERIC CCD / Digitization - Sampling interval: 15 µm / Number real images: 71 / Average electron dose: 80 e/Å2 / Bits/pixel: 16

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 38.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMOD
Details: Final maps were calculated from fifty averaged datasets
DetailsAverage number of projections used in the 3D reconstructions: 50.

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