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- EMDB-6368: Electron Cryo-microscopy of Chikungunya virus-like particles in c... -

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Basic information

Entry
Database: EMDB / ID: EMD-6368
TitleElectron Cryo-microscopy of Chikungunya virus-like particles in complex with neutralizing antibody Fab 8B10
Map dataSingle-particle reconstruction of CHIK-VLP:Fab 8B10
Sample
  • Sample: Fab fragment of human antibody 8B10 bound to Chikungunya virus
  • Virus: Chikungunya virus
  • Protein or peptide: neutralizing antibody Fab 8B10
Keywordsantibody neutralization / Chikungunya / Fab
Biological speciesMus musculus (house mouse) / Chikungunya virus
Methodsingle particle reconstruction / cryo EM / Resolution: 17.6 Å
AuthorsPorta JC / Mangala Prasad V / Wang CI / Akahata W / Ng LFP / Rossmann MG
CitationJournal: J Virol / Year: 2016
Title: Structural Studies of Chikungunya Virus-Like Particles Complexed with Human Antibodies: Neutralization and Cell-to-Cell Transmission.
Authors: Jason Porta / Vidya Mangala Prasad / Cheng-I Wang / Wataru Akahata / Lisa F P Ng / Michael G Rossmann /
Abstract: Chikungunya virus is a positive-stranded RNA alphavirus. Structures of chikungunya virus-like particles in complex with strongly neutralizing antibody Fab fragments (8B10 and 5F10) were determined ...Chikungunya virus is a positive-stranded RNA alphavirus. Structures of chikungunya virus-like particles in complex with strongly neutralizing antibody Fab fragments (8B10 and 5F10) were determined using cryo-electron microscopy and X-ray crystallography. By fitting the crystallographically determined structures of these Fab fragments into the cryo-electron density maps, we show that Fab fragments of antibody 8B10 extend radially from the viral surface and block receptor binding on the E2 glycoprotein. In contrast, Fab fragments of antibody 5F10 bind the tip of the E2 B domain and lie tangentially on the viral surface. Fab 5F10 fixes the B domain rigidly to the surface of the virus, blocking exposure of the fusion loop on glycoprotein E1 and therefore preventing the virus from becoming fusogenic. Although Fab 5F10 can neutralize the wild-type virus, it can also bind to a mutant virus without inhibiting fusion or attachment. Although the mutant virus is no longer able to propagate by extracellular budding, it can, however, enter the next cell by traveling through junctional complexes without being intercepted by a neutralizing antibody to the wild-type virus, thus clarifying how cell-to-cell transmission can occur.
IMPORTANCE: Alphaviral infections are transmitted mainly by mosquitoes. Chikungunya virus (CHIKV), which belongs to the Alphavirus genus, has a wide distribution in the Old World that has expanded in ...IMPORTANCE: Alphaviral infections are transmitted mainly by mosquitoes. Chikungunya virus (CHIKV), which belongs to the Alphavirus genus, has a wide distribution in the Old World that has expanded in recent years into the Americas. There are currently no vaccines or drugs against alphaviral infections. Therefore, a better understanding of CHIKV and its associated neutralizing antibodies will aid in the development of effective treatments.
History
DepositionJun 26, 2015-
Header (metadata) releaseAug 19, 2015-
Map releaseNov 18, 2015-
UpdateJun 22, 2016-
Current statusJun 22, 2016Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1.5
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 1.5
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_6368.png

Downloads & links

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Map

FileDownload / File: emd_6368.map.gz / Format: CCP4 / Size: 62.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSingle-particle reconstruction of CHIK-VLP:Fab 8B10
Voxel sizeX=Y=Z: 4.28 Å
Density
Contour LevelBy AUTHOR: 1.5 / Movie #1: 1.5
Minimum - Maximum-4.6353879 - 4.91174984
Average (Standard dev.)0.00380965 (±0.54962218)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-128-128-128
Dimensions256256256
Spacing256256256
CellA=B=C: 1095.68 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.284.284.28
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z1095.6801095.6801095.680
α/β/γ90.00090.00090.000
start NX/NY/NZ-147-147-146
NX/NY/NZ294294294
MAP C/R/S123
start NC/NR/NS-128-128-128
NC/NR/NS256256256
D min/max/mean-4.6354.9120.004

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Supplemental data

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Sample components

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Entire : Fab fragment of human antibody 8B10 bound to Chikungunya virus

EntireName: Fab fragment of human antibody 8B10 bound to Chikungunya virus
Components
  • Sample: Fab fragment of human antibody 8B10 bound to Chikungunya virus
  • Virus: Chikungunya virus
  • Protein or peptide: neutralizing antibody Fab 8B10

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Supramolecule #1000: Fab fragment of human antibody 8B10 bound to Chikungunya virus

SupramoleculeName: Fab fragment of human antibody 8B10 bound to Chikungunya virus
type: sample / ID: 1000 / Number unique components: 2
Molecular weightExperimental: 5.25 MDa / Method: Sedimentation

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Supramolecule #1: Chikungunya virus

SupramoleculeName: Chikungunya virus / type: virus / ID: 1 / Name.synonym: CHIK-VLP / NCBI-ID: 37124 / Sci species name: Chikungunya virus / Sci species strain: LR2006 OPY-1 / Virus type: VIRUS-LIKE PARTICLE / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: Yes / Syn species name: CHIK-VLP
Host (natural)Organism: Homo sapiens (human) / synonym: VERTEBRATES
Host systemOrganism: Cricetinae (hamsters) / Recombinant strain: LR2006 OPY-1 / Recombinant cell: BHK-15 / Recombinant plasmid: E37997, Eopy-1
Molecular weightExperimental: 5.25 MDa
Virus shellShell ID: 1 / Name: E1E2 / Diameter: 700 Å / T number (triangulation number): 4

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Macromolecule #1: neutralizing antibody Fab 8B10

MacromoleculeName: neutralizing antibody Fab 8B10 / type: protein_or_peptide / ID: 1 / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Mus musculus (house mouse) / synonym: mouse

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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

Concentration2 mg/mL
BufferpH: 7.6 / Details: PBS
GridDetails: 400 mesh Cu holey carbon
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 93 K / Instrument: GATAN CRYOPLUNGE 3 / Method: Blot for 5 seconds before plunging

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 60521 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2 mm / Nominal defocus max: 0.35 µm / Nominal defocus min: 0.185 µm / Nominal magnification: 59000
Specialist opticsEnergy filter - Name: FEI
Sample stageSpecimen holder: Nitrogen-cooled / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
TemperatureMin: 80 K / Max: 100 K / Average: 95 K
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 96,000 times magnification.
DateMay 24, 2014
Image recordingCategory: CCD / Film or detector model: FEI CETA (4k x 4k) / Digitization - Sampling interval: 2.5 µm / Number real images: 96 / Average electron dose: 22 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: Each image
Final two d classificationNumber classes: 10
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 17.6 Å / Resolution method: OTHER / Software - Name: EMAN / Number images used: 1582
DetailsManual selection using e2boxer software

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