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- EMDB-26065: nsEM maps of coxsackievirus A21 viral particle alone and in compl... -

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Basic information

Entry
Database: EMDB / ID: EMD-26065
TitlensEM maps of coxsackievirus A21 viral particle alone and in complex with mouse polyclonal antibodies
Map datansEM map of CV-A21 viral particle in complex with mouse polyclonal antibodies (antigen-specific)
Sample
  • Complex: Immune complexes featuring coxsackievirus A21 (CV-A21) viral particles by themselves or in complex with mouse polyclonal antibodies (reconstructed by nsEM)
Keywordspolyclonal antibodies / immune complex / negative stain EM / VIRUS / VIRUS-IMMUNE SYSTEM complex
Biological speciesCoxsackievirus A21
Methodsingle particle reconstruction / negative staining / Resolution: 15.0 Å
AuthorsAntanasijevic A / Ward AB
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: PNAS Nexus / Year: 2022
Title: High-resolution structural analysis of enterovirus-reactive polyclonal antibodies in complex with whole virions.
Authors: Aleksandar Antanasijevic / Autumn J Schulze / Vijay S Reddy / Andrew B Ward /
Abstract: Non-polio enteroviruses (NPEVs) cause serious illnesses in young children and neonates, including aseptic meningitis, encephalitis, and inflammatory muscle disease, among others. While over 100 ...Non-polio enteroviruses (NPEVs) cause serious illnesses in young children and neonates, including aseptic meningitis, encephalitis, and inflammatory muscle disease, among others. While over 100 serotypes have been described to date, vaccine only exists for EV-A71. Efforts toward rationally designed pan-NPEV vaccines would greatly benefit from structural biology methods for rapid and comprehensive evaluation of vaccine candidates and elicited antibody responses. Toward this goal, we introduced a cryo-electron-microscopy-based approach for structural analysis of virus- or vaccine-elicited polyclonal antibodies (pAbs) in complex with whole NPEV virions. We demonstrated the feasibility using coxsackievirus A21 and reconstructed five structurally distinct pAbs bound to the virus. The pAbs targeted two immunodominant epitopes, one overlapping with the receptor binding site. These results demonstrate that our method can be applied to map broad-spectrum polyclonal immune responses against intact virions and define potentially cross-reactive epitopes.
History
DepositionJan 26, 2022-
Header (metadata) releaseJan 4, 2023-
Map releaseJan 4, 2023-
UpdateJan 17, 2024-
Current statusJan 17, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26065.png

Downloads & links

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Map

FileDownload / File: emd_26065.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationnsEM map of CV-A21 viral particle in complex with mouse polyclonal antibodies (antigen-specific)
Voxel sizeX=Y=Z: 1.77 Å
Density
Contour LevelBy AUTHOR: 0.003
Minimum - Maximum-0.01279412 - 0.027668145
Average (Standard dev.)0.0019828323 (±0.005118759)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 566.4 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: nsEM map of CV-A21 viral particle

Fileemd_26065_additional_1.map
AnnotationnsEM map of CV-A21 viral particle
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: nsEM map of CV-A21 viral particle in complex...

Fileemd_26065_additional_2.map
AnnotationnsEM map of CV-A21 viral particle in complex with mouse polyclonal antibodies (non-specific)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Immune complexes featuring coxsackievirus A21 (CV-A21) viral part...

EntireName: Immune complexes featuring coxsackievirus A21 (CV-A21) viral particles by themselves or in complex with mouse polyclonal antibodies (reconstructed by nsEM)
Components
  • Complex: Immune complexes featuring coxsackievirus A21 (CV-A21) viral particles by themselves or in complex with mouse polyclonal antibodies (reconstructed by nsEM)

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Supramolecule #1: Immune complexes featuring coxsackievirus A21 (CV-A21) viral part...

SupramoleculeName: Immune complexes featuring coxsackievirus A21 (CV-A21) viral particles by themselves or in complex with mouse polyclonal antibodies (reconstructed by nsEM)
type: complex / ID: 1 / Parent: 0
Details: The 3 submitted maps feature nsEM reconstructions of: 1) CV-A21 in complex with mouse polyclonal antibodies (antigen-specific) 2) CV-A21 in complex with mouse polyclonal antibodies (non-specific) 3) CV-A21 alone
Source (natural)Organism: Coxsackievirus A21

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.1 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
10.0 mMTris-HClTrisTris-HClTris
150.0 mMNaClSodium chlorideSodium chloride

Details: TBS, 0.2um filtered
StainingType: NEGATIVE / Material: Uranyl formate / Details: The samples were stained with UF for 60s.
GridModel: Homemade / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 30 sec.
DetailsThe complex was generated by incubation of mouse polyclonal antibodies with recombinantly expressed CV-A21 virions and subsequent SEC purification.

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Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.2 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 62000
Sample stageSpecimen holder model: SIDE ENTRY, EUCENTRIC
Image recordingFilm or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Average electron dose: 25.0 e/Å2
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 6224 / Details: DoG Picker in Appion
Startup modelType of model: OTHER / Details: Low-pass filtered map of coxsackievirus particle
Initial angle assignmentType: OTHER / Software - Name: RELION (ver. 3.0) / Details: Relion, regularized likelihood
Final 3D classificationNumber classes: 4 / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: OTHER / Software - Name: RELION (ver. 3.0) / Details: Relion, regularized likelihood
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 15.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 6224

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