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- EMDB-40308: Sub-tomogram average of the RSV F pairs from the surface of nativ... -

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Basic information

Entry
Database: EMDB / ID: EMD-40308
TitleSub-tomogram average of the RSV F pairs from the surface of native virions released from RSV-infected BEAS-2B cells cultured on EM grids collected via montage parallel array cryo-tomography (MPACT)
Map dataSTA of RSV glycoprotein F pair on the surface of native RSV virion collected via parallel array montage cryo-electron tomography
Sample
  • Virus: Respiratory syncytial virus A2
Keywordsnatively folded / class I viral fusion protein / pre-fusion native state / whole virion / VIRUS
Biological speciesRespiratory syncytial virus A2
Methodsubtomogram averaging / cryo EM / Resolution: 19.9 Å
AuthorsYang J / Wright ER
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01 GM114561 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)U24 GM139168 United States
CitationJournal: Nat Methods / Year: 2023
Title: Correlative montage parallel array cryo-tomography for in situ structural cell biology.
Authors: Jie E Yang / Matthew R Larson / Bryan S Sibert / Joseph Y Kim / Daniel Parrell / Juan C Sanchez / Victoria Pappas / Anil Kumar / Kai Cai / Keith Thompson / Elizabeth R Wright /
Abstract: Imaging large fields of view while preserving high-resolution structural information remains a challenge in low-dose cryo-electron tomography. Here we present robust tools for montage parallel array ...Imaging large fields of view while preserving high-resolution structural information remains a challenge in low-dose cryo-electron tomography. Here we present robust tools for montage parallel array cryo-tomography (MPACT) tailored for vitrified specimens. The combination of correlative cryo-fluorescence microscopy, focused-ion-beam milling, substrate micropatterning, and MPACT supports studies that contextually define the three-dimensional architecture of cells. To further extend the flexibility of MPACT, tilt series may be processed in their entirety or as individual tiles suitable for sub-tomogram averaging, enabling efficient data processing and analysis.
History
DepositionApr 3, 2023-
Header (metadata) releaseSep 27, 2023-
Map releaseSep 27, 2023-
UpdateOct 18, 2023-
Current statusOct 18, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_40308.png

Downloads & links

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Map

FileDownload / File: emd_40308.map.gz / Format: CCP4 / Size: 8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSTA of RSV glycoprotein F pair on the surface of native RSV virion collected via parallel array montage cryo-electron tomography
Voxel sizeX=Y=Z: 2.503 Å
Density
Contour LevelBy AUTHOR: 0.0658
Minimum - Maximum-0.18480107 - 0.31608817
Average (Standard dev.)0.00023386958 (±0.030943437)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions128128128
Spacing128128128
CellA=B=C: 320.384 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: STA of RSV glycoprotein F pair on the...

Fileemd_40308_half_map_1.map
AnnotationSTA of RSV glycoprotein F pair on the surface of native RSV virion collected via parallel array montage cryo-electron tomography
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: STA of RSV glycoprotein F pair on the...

Fileemd_40308_half_map_2.map
AnnotationSTA of RSV glycoprotein F pair on the surface of native RSV virion collected via parallel array montage cryo-electron tomography
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Respiratory syncytial virus A2

EntireName: Respiratory syncytial virus A2
Components
  • Virus: Respiratory syncytial virus A2

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Supramolecule #1: Respiratory syncytial virus A2

SupramoleculeName: Respiratory syncytial virus A2 / type: virus / ID: 1 / Parent: 0
Details: The strain was kindly provided by Dr. Martin L. Moore. The reference is PMCID: PMC3492879/PMID:23062737
NCBI-ID: 1972429 / Sci species name: Respiratory syncytial virus A2 / Sci species strain: rA2-mK+ / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: No
Host (natural)Organism: Homo sapiens (human)

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferpH: 7
GridModel: Quantifoil R2/1 / Material: GOLD / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 17
VitrificationCryogen name: ETHANE / Chamber humidity: 80 % / Chamber temperature: 303 K / Instrument: GATAN CRYOPLUNGE 3
DetailsRespiratory syncytial virus released from infected whole BEAS-2B mammalian cells (human non-tumorigenic lung epithelial cell)

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Average exposure time: 0.5 sec. / Average electron dose: 1.8 e/Å2
Details: Tilt series of defocus collection ranging from -3 to -6 micrometer, with a step of -0.5 micrometer
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated defocus max: 6.8500000000000005 µm / Calibrated defocus min: 3.3200000000000003 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 6.0 µm / Nominal defocus min: 3.0 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Details5760 x 4092 (counting mode, bin 1x)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: EXACT BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 19.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: IMOD (ver. 4.11.11) / Number subtomograms used: 13021
ExtractionNumber tomograms: 20 / Number images used: 15000 / Method: Semi-automatic and manual picking / Software - Name: IMOD (ver. 4.11.11)
Details: The initial reference model was generated from the manually picked particles. The first round of reference was used for template-matching picking in some tomograms. For those tomograms ...Details: The initial reference model was generated from the manually picked particles. The first round of reference was used for template-matching picking in some tomograms. For those tomograms without good template matching picking, manual picking was done.
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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