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- EMDB-22188: CryoEM structure of Chimeric Eastern Equine Encephalitis Virus wi... -

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Basic information

Entry
Database: EMDB / ID: EMD-22188
TitleCryoEM structure of Chimeric Eastern Equine Encephalitis Virus with Fab of EEEV-143 Antibody
Map dataEastern Equine Encephalitis Virus (EEEV)_Fab143 complex
Sample
  • Virus: Eastern equine encephalitis virus
Biological speciesEastern equine encephalitis virus
Methodsingle particle reconstruction / cryo EM / Resolution: 8.3 Å
AuthorsWilliamson LE / Gilliland T / Yadav PK / Binshtein E / Bombardi R / Kose N / Nargi RS / Sutton RM / Armstrong E / Carnahan RH ...Williamson LE / Gilliland T / Yadav PK / Binshtein E / Bombardi R / Kose N / Nargi RS / Sutton RM / Armstrong E / Carnahan RH / Walker LM / Kim AS / Fox J / Diamond MS / Ohi M / Klimstra WB / Crowe JE
Funding support United States, United Arab Emirates, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)T32 HL069765 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)U19 AI142790 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)F31 AI145189 United States
National Institutes of Health/National Center for Advancing Translational Sciences (NIH/NCATS)UL1 TR002243United Arab Emirates
Defense Threat Reduction Agency (DTRA)HDTRA1-13-1-0034 United States
CitationJournal: Cell / Year: 2020
Title: Human Antibodies Protect against Aerosolized Eastern Equine Encephalitis Virus Infection.
Authors: Lauren E Williamson / Theron Gilliland / Pramod K Yadav / Elad Binshtein / Robin Bombardi / Nurgun Kose / Rachel S Nargi / Rachel E Sutton / Clarissa L Durie / Erica Armstrong / Robert H ...Authors: Lauren E Williamson / Theron Gilliland / Pramod K Yadav / Elad Binshtein / Robin Bombardi / Nurgun Kose / Rachel S Nargi / Rachel E Sutton / Clarissa L Durie / Erica Armstrong / Robert H Carnahan / Lauren M Walker / Arthur S Kim / Julie M Fox / Michael S Diamond / Melanie D Ohi / William B Klimstra / James E Crowe /
Abstract: Eastern equine encephalitis virus (EEEV) is one of the most virulent viruses endemic to North America. No licensed vaccines or antiviral therapeutics are available to combat this infection, which has ...Eastern equine encephalitis virus (EEEV) is one of the most virulent viruses endemic to North America. No licensed vaccines or antiviral therapeutics are available to combat this infection, which has recently shown an increase in human cases. Here, we characterize human monoclonal antibodies (mAbs) isolated from a survivor of natural EEEV infection with potent (<20 pM) inhibitory activity of EEEV. Cryo-electron microscopy reconstructions of two highly neutralizing mAbs, EEEV-33 and EEEV-143, were solved in complex with chimeric Sindbis/EEEV virions to 7.2 Å and 8.3 Å, respectively. The mAbs recognize two distinct antigenic sites that are critical for inhibiting viral entry into cells. EEEV-33 and EEEV-143 protect against disease following stringent lethal aerosol challenge of mice with highly pathogenic EEEV. These studies provide insight into the molecular basis for the neutralizing human antibody response against EEEV and can facilitate development of vaccines and candidate antibody therapeutics.
History
DepositionJun 19, 2020-
Header (metadata) releaseJan 6, 2021-
Map releaseJan 6, 2021-
UpdateJan 13, 2021-
Current statusJan 13, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.004
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.004
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22188.png

Downloads & links

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Map

FileDownload / File: emd_22188.map.gz / Format: CCP4 / Size: 1.3 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationEastern Equine Encephalitis Virus (EEEV)_Fab143 complex
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2 Å/pix.
x 700 pix.
= 1400. Å		2 Å/pix.
x 700 pix.
= 1400. Å		2 Å/pix.
x 700 pix.
= 1400. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.004 / Movie #1: 0.004
Minimum - Maximum-0.0061711418 - 0.015312157
Average (Standard dev.)0.0002464621 (±0.0011589875)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions700700700
Spacing700700700
CellA=B=C: 1400.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z222
M x/y/z700700700
origin x/y/z0.0000.0000.000
length x/y/z1400.0001400.0001400.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ400400400
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS700700700
D min/max/mean-0.0060.0150.000

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Supplemental data

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Additional map: Eastern Equine Encephalitis Virus (EEEV) Fab143 complex

Fileemd_22188_additional_1.map
AnnotationEastern Equine Encephalitis Virus (EEEV)_Fab143 complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Eastern equine encephalitis virus

EntireName: Eastern equine encephalitis virus
Components
  • Virus: Eastern equine encephalitis virus

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Supramolecule #1: Eastern equine encephalitis virus

SupramoleculeName: Eastern equine encephalitis virus / type: virus / ID: 1 / Parent: 0 / Details: Virus was purified from mammalian cell culture / NCBI-ID: 11021 / Sci species name: Eastern equine encephalitis virus / Sci species strain: Sindbis / Virus type: VIRION / Virus isolate: SUBSPECIES / Virus enveloped: Yes / Virus empty: Yes
Host (natural)Organism: Culiseta melanura (mosquito)
Host systemOrganism: Aedes albopictus (Asian tiger mosquito) / Recombinant cell: C6/36 cells
Molecular weightTheoretical: 6 MDa
Virus shellShell ID: 1 / Name: Eastern equine encephalitis virus / Diameter: 850.0 Å / T number (triangulation number): 4

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation state3D array

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Sample preparation

Concentration0.1 mg/mL
BufferpH: 7.5
Component:
ConcentrationName
50.0 mMTris-HCl
0.1 mMEDTA
100.0 mMSodium Chloride
1.0 xDPBS

Details: Solutions were made fresh
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: blot for 5 seconds before plunging.
DetailsPurified SINV/EEEV particles and recombinant Fab mixture

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Electron microscopy

MicroscopeTFS GLACIOS
TemperatureMin: 70.0 K / Max: 77.0 K
DetailsPreliminary grid screening was performed manually
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-50 / Number grids imaged: 1 / Number real images: 3500 / Average exposure time: 10.0 sec. / Average electron dose: 25.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: OTHER / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 22000
Sample stageSpecimen holder model: OTHER / Cooling holder cryogen: NITROGEN

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Image processing

Particle selectionNumber selected: 10000
CTF correctionSoftware - Name: Gctf (ver. 1.06)
Software - details: Contrast transfer function parameters was estimated using Gctf
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 8.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0)
Software - details: Final masked 3D refinement and postprocessing
Number images used: 7288
Initial angle assignmentType: OTHER
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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