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- EMDB-1165: A 9 angstroms single particle reconstruction from CCD captured im... -

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Basic information

Entry
Database: EMDB / ID: EMD-1165
TitleA 9 angstroms single particle reconstruction from CCD captured images on a 200 kV electron cryomicroscope.
Map datathis is a half map of CPV oriented along the 5 fold axis of symmetry. Raw Data Collected on 4k x 4k Gatan US4000 CCD Camera. Reconstruction to 9 A.
Sample
  • Sample: Cytoplasmic Polyhedrosis VirusCypovirus
  • Virus: Bombyx mori cypovirus 1
Biological speciesBombyx mori cypovirus 1
Methodsingle particle reconstruction / cryo EM / negative staining / Resolution: 9.0 Å
AuthorsBooth C / Jiang W / Baker ML / Zhou ZH / Ludtke SJ / Chiu W
CitationJournal: J Struct Biol / Year: 2004
Title: A 9 angstroms single particle reconstruction from CCD captured images on a 200 kV electron cryomicroscope.
Authors: Christopher R Booth / Wen Jiang / Matthew L Baker / Z Hong Zhou / Steven J Ludtke / Wah Chiu /
Abstract: Sub-nanometer resolution structure determination is becoming a common practice in electron cryomicroscopy of macromolecular assemblies. The data for these studies have until now been collected on ...Sub-nanometer resolution structure determination is becoming a common practice in electron cryomicroscopy of macromolecular assemblies. The data for these studies have until now been collected on photographic film. Using cytoplasmic polyhedrosis virus (CPV), a previously determined structure, as a test specimen, we show the feasibility of obtaining a 9 angstroms structure from images acquired from a 4 k x 4 k Gatan CCD on a 200 kV electron cryomicroscope. The match of the alpha-helices in the protein components of the CPV with the previous structure of the same virus validates the suitability of this type of camera as the recording media targeted for single particle reconstructions at sub-nanometer resolution.
History
DepositionSep 7, 2004-
Header (metadata) releaseSep 28, 2005-
Map releaseSep 28, 2006-
UpdateMay 26, 2011-
Current statusMay 26, 2011Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3.5
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 3.5
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

1165.gif

Downloads & links

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Map

FileDownload / File: emd_1165.map.gz / Format: CCP4 / Size: 231.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationthis is a half map of CPV oriented along the 5 fold axis of symmetry. Raw Data Collected on 4k x 4k Gatan US4000 CCD Camera. Reconstruction to 9 A.
Voxel sizeX=Y=Z: 1.8 Å
Density
Contour Level1: 1.71 / Movie #1: 3.5
Minimum - Maximum-9.55039 - 13.1724
Average (Standard dev.)0.0854754 (±1.43191)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-249-249-125
Dimensions499499250
Spacing499499250
CellA: 898.2 Å / B: 898.2 Å / C: 450 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.81.81.8
M x/y/z499499250
origin x/y/z0.0000.0000.000
length x/y/z898.200898.200450.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-64-64-64
NX/NY/NZ128128128
MAP C/R/S123
start NC/NR/NS-249-249-125
NC/NR/NS499499250
D min/max/mean-9.55013.1720.085

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Supplemental data

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Sample components

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Entire : Cytoplasmic Polyhedrosis Virus

EntireName: Cytoplasmic Polyhedrosis VirusCypovirus
Components
  • Sample: Cytoplasmic Polyhedrosis VirusCypovirus
  • Virus: Bombyx mori cypovirus 1

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Supramolecule #1000: Cytoplasmic Polyhedrosis Virus

SupramoleculeName: Cytoplasmic Polyhedrosis Virus / type: sample / ID: 1000 / Number unique components: 1

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Supramolecule #1: Bombyx mori cypovirus 1

SupramoleculeName: Bombyx mori cypovirus 1 / type: virus / ID: 1 / Name.synonym: CPV / NCBI-ID: 110829 / Sci species name: Bombyx mori cypovirus 1 / Virus type: VIRION / Virus isolate: SPECIES / Virus enveloped: No / Virus empty: No / Syn species name: CPV
Host (natural)Organism: Bombyx mori (domestic silkworm) / synonym: INVERTEBRATES
Virus shellShell ID: 1 / Name: capsid / Diameter: 750 Å / T number (triangulation number): 1

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Experimental details

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Structure determination

Methodnegative staining, cryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1 mg/mL
BufferpH: 7.4 / Details: Phosphate Buffered Saline ph 7.4
StainingType: NEGATIVE / Details: flash frozen in liquid ethane
GridDetails: irradiated quantafoil R2-1 holey carbon grids
VitrificationCryogen name: ETHANE / Chamber humidity: 45 % / Chamber temperature: 80 K / Instrument: HOMEMADE PLUNGER
Details: Vitrification instrument: manual plunger. homemade apparatus
Method: blot on 1 side and plunge

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Electron microscopy

MicroscopeJEOL 2010F
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 83062 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 1 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 60000
Sample stageSpecimen holder: Side entry liquid nitrogen-cooled cryo specimen holder
Specimen holder model: GATAN LIQUID NITROGEN
TemperatureMin: 90 K / Max: 90 K / Average: 90 K
DateMar 1, 2003
Image recordingCategory: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 856 / Average electron dose: 15 e/Å2 / Bits/pixel: 12

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Image processing

CTF correctionDetails: SAVR per frame
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 9.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SAVR / Number images used: 6465
Detailsparticles selected using Ethan and manually verified.

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Atomic model buiding 1

SoftwareName: Foldhunter
DetailsProtocol: Rigid Body. Reovirus lambda1 protein was fitted to CSPA. Helixhunter was used to identify alpha helices.
RefinementProtocol: RIGID BODY FIT / Target criteria: Cross correlation

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