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- EMDB-5563: Electron microscopy of Everglades virus -

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Basic information

Entry
Database: EMDB / ID: EMD-5563
TitleElectron microscopy of Everglades virus
Map dataReconstruction of wild type everglades virus
Sample
  • Sample: Everglades virus
  • Virus: Everglades virus
Keywordselectron microscopy / cryo-electron microscopy / alphavirus / everglades virus
Biological speciesEverglades virus
Methodsingle particle reconstruction / cryo EM / Resolution: 10.0 Å
AuthorsSherman MB / Trujillo J / Leahy I / Razmus D / DeHate R / Lorcheim P / Czarneski MA / Zimmerman D / Newton JPM / Haddow AM / Weaver SC
CitationJournal: J Struct Biol / Year: 2013
Title: Construction and organization of a BSL-3 cryo-electron microscopy laboratory at UTMB.
Authors: Michael B Sherman / Juan Trujillo / Ian Leahy / Dennis Razmus / Robert Dehate / Paul Lorcheim / Mark A Czarneski / Domenica Zimmerman / Je T'aime M Newton / Andrew D Haddow / Scott C Weaver /
Abstract: A unique cryo-electron microscopy facility has been designed and constructed at the University of Texas Medical Branch (UTMB) to study the three-dimensional organization of viruses and bacteria ...A unique cryo-electron microscopy facility has been designed and constructed at the University of Texas Medical Branch (UTMB) to study the three-dimensional organization of viruses and bacteria classified as select agents at biological safety level (BSL)-3, and their interactions with host cells. A 200keV high-end cryo-electron microscope was installed inside a BSL-3 containment laboratory and standard operating procedures were developed and implemented to ensure its safe and efficient operation. We also developed a new microscope decontamination protocol based on chlorine dioxide gas with a continuous flow system, which allowed us to expand the facility capabilities to study bacterial agents including spore-forming species. The new unified protocol does not require agent-specific treatment in contrast to the previously used heat decontamination. To optimize the use of the cryo-electron microscope and to improve safety conditions, it can be remotely controlled from a room outside of containment, or through a computer network world-wide. Automated data collection is provided by using JADAS (single particle imaging) and SerialEM (tomography). The facility has successfully operated for more than a year without an incident and was certified as a select agent facility by the Centers for Disease Control.
History
DepositionJan 6, 2013-
Header (metadata) releaseFeb 13, 2013-
Map releaseFeb 13, 2013-
UpdateFeb 27, 2013-
Current statusFeb 27, 2013Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 15
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 15
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_5563_1.jpg

Downloads & links

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Map

FileDownload / File: emd_5563.map.gz / Format: CCP4 / Size: 29.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of wild type everglades virus
Voxel sizeX=Y=Z: 4 Å
Density
Contour LevelBy EMDB: 13.0 / Movie #1: 15
Minimum - Maximum-72.682418819999995 - 68.751846310000005
Average (Standard dev.)0.0 (±9.223706249999999)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-100-100-100
Dimensions200200200
Spacing200200200
CellA=B=C: 800.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z444
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z800.000800.000800.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-5029166
NX/NY/NZ106122134
MAP C/R/S123
start NC/NR/NS-100-100-100
NC/NR/NS200200200
D min/max/mean-72.68268.7520.000

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Supplemental data

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Sample components

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Entire : Everglades virus

EntireName: Everglades virus
Components
  • Sample: Everglades virus
  • Virus: Everglades virus

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Supramolecule #1000: Everglades virus

SupramoleculeName: Everglades virus / type: sample / ID: 1000 / Details: monodisperse sample / Number unique components: 4

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Supramolecule #1: Everglades virus

SupramoleculeName: Everglades virus / type: virus / ID: 1 / Sci species name: Everglades virus / Sci species strain: Fe5-47et / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: No
Host (natural)Organism: unidentified (others) / synonym: VERTEBRATES
Host systemOrganism: unidentified (others)
Virus shellShell ID: 1 / Name: glycoprotein E1/E2 / Diameter: 700 Å / T number (triangulation number): 4

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1 mg/mL
BufferpH: 7.4 / Details: 0.05 M Tris HCl, pH 7.4, 0.1 M NaCl, 0.001 M EDTA
GridDetails: C-flat 1.2/1.3 holey film
VitrificationCryogen name: ETHANE / Chamber humidity: 60 % / Chamber temperature: 90 K / Instrument: HOMEMADE PLUNGER / Method: blot for 4 seconds before plunging

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Electron microscopy

MicroscopeJEOL 2200FS
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 74800 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 2.3 µm / Nominal defocus min: 0.3 µm / Nominal magnification: 60000
Specialist opticsEnergy filter - Name: omega
Sample stageSpecimen holder: Gatan 626 70 degree holder / Specimen holder model: GATAN LIQUID NITROGEN
TemperatureMin: 87 K / Max: 90 K / Average: 88 K
Alignment procedureLegacy - Astigmatism: manual correction at 150,000x
DateFeb 9, 2012
Image recordingCategory: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Digitization - Sampling interval: 15 µm / Number real images: 80 / Average electron dose: 20 e/Å2 / Bits/pixel: 16

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Image processing

CTF correctionDetails: micrograph
Final angle assignmentDetails: icosahedral asymmetric unit
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 10.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMAGIC-5
Details: final maps were calculated from individual particles
Number images used: 1000
Detailsparticles picked with EMAN2 boxer, reconstructed using IMAGIC-5

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