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- EMDB-0916: Neutralization mechanism of a monoclonal antibody targeting a por... -

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Basic information

Entry
Database: EMDB / ID: EMD-0916
TitleNeutralization mechanism of a monoclonal antibody targeting a porcine circovirus type 2 Cap protein conformational epitope
Map data3D cry-EM map of PCV2 vision
Sample
  • Complex: PCV2 virion in complex with Fab fragments of the mAb 3A5
    • Protein or peptide: Capsid protein
KeywordsPorcine circovirus type 2 / Cap protein / VIRUS
Function / homology
Function and homology information


viral capsid assembly / T=1 icosahedral viral capsid / viral penetration into host nucleus / host cell / endocytosis involved in viral entry into host cell / virion attachment to host cell / host cell nucleus
Similarity search - Function
Circovirus capsid protein / Circovirus capsid superfamily / Circovirus capsid protein
Similarity search - Domain/homology
Biological speciesPorcine circovirus 2
Methodsingle particle reconstruction / cryo EM / Resolution: 6.7 Å
AuthorsSun Z / Huang L
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31873012 China
CitationJournal: J Virol / Year: 2020
Title: Neutralization Mechanism of a Monoclonal Antibody Targeting a Porcine Circovirus Type 2 Cap Protein Conformational Epitope.
Authors: Liping Huang / Zhenzhao Sun / Deli Xia / Yanwu Wei / Encheng Sun / Chunguo Liu / Hongzhen Zhu / Haiqiao Bian / Hongli Wu / Li Feng / Jingfei Wang / Changming Liu /
Abstract: Porcine circovirus type 2 (PCV2) is an important pathogen in swine herds, and its infection of pigs has caused severe economic losses to the pig industry worldwide. The capsid protein of PCV2 is the ...Porcine circovirus type 2 (PCV2) is an important pathogen in swine herds, and its infection of pigs has caused severe economic losses to the pig industry worldwide. The capsid protein of PCV2 is the only structural protein that is associated with PCV2 infection and immunity. Here, we report a neutralizing monoclonal antibody (MAb), MAb 3A5, that binds to intact PCV2 virions of the PCV2a, PCV2b, and PCV2d genotypes. MAb 3A5 neutralized PCV2 by blocking viral attachment to PK15 cells. To further explore the neutralization mechanism, we resolved the structure of the PCV2 virion in complex with MAb 3A5 Fab fragments by using cryo-electron microscopy single-particle analysis. The binding sites were located at the topmost edges around 5-fold icosahedral symmetry axes, with each footprint covering amino acids from two adjacent capsid proteins. Most of the epitope residues (15/18 residues) were conserved among 2,273 PCV2 strains. Mutations of some amino acids within the epitope had significant effects on the neutralizing activity of MAb 3A5. This study reveals the molecular and structural bases of this PCV2-neutralizing antibody and provides new and important information for vaccine design and therapeutic antibody development against PCV2 infections. PCV2 is associated with several clinical manifestations collectively known as PCV2-associated diseases (PCVADs). Neutralizing antibodies play a crucial role in the prevention of PCVADs. We demonstrated previously that a MAb, MAb 3A5, neutralizes the PCV2a, PCV2b, and PCV2d genotypes with different degrees of efficiency, but the underlying mechanism remains elusive. Here, we report the neutralization mechanism of this MAb and the structure of the PCV2 virion in complex with MAb 3A5 Fabs, showing a binding mode in which one Fab interacted with more than two loops from two adjacent capsid proteins. This binding mode has not been observed previously for PCV2-neutralizing antibodies. Our work provides new and important information for vaccine design and therapeutic antibody development against PCV2 infections.
History
DepositionDec 24, 2019-
Header (metadata) releaseFeb 12, 2020-
Map releaseFeb 12, 2020-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6lm3
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6lm3
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_0916.png

Downloads & links

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Map

FileDownload / File: emd_0916.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation3D cry-EM map of PCV2 vision
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.8 Å/pix.
x 300 pix.
= 240. Å		0.8 Å/pix.
x 300 pix.
= 240. Å		0.8 Å/pix.
x 300 pix.
= 240. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.8 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.02 / Movie #1: 0.02
Minimum - Maximum-0.07332169 - 0.09048094
Average (Standard dev.)0.000999484 (±0.006954318)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-150-150-150
Dimensions300300300
Spacing300300300
CellA=B=C: 240.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.80.80.8
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z240.000240.000240.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-150-150-150
NC/NR/NS300300300
D min/max/mean-0.0730.0900.001

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Supplemental data

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Sample components

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Entire : PCV2 virion in complex with Fab fragments of the mAb 3A5

EntireName: PCV2 virion in complex with Fab fragments of the mAb 3A5
Components
  • Complex: PCV2 virion in complex with Fab fragments of the mAb 3A5
    • Protein or peptide: Capsid protein

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Supramolecule #1: PCV2 virion in complex with Fab fragments of the mAb 3A5

SupramoleculeName: PCV2 virion in complex with Fab fragments of the mAb 3A5
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Porcine circovirus 2

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Macromolecule #1: Capsid protein

MacromoleculeName: Capsid protein / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Porcine circovirus 2
Molecular weightTheoretical: 27.55952 KDa
Recombinant expressionOrganism: Porcine circovirus 2
SequenceString: MTYPRRRFRR RRHRPRSHLG LILRRRPWLV HPRHRYRWRR KNGIFNTRLS CTFGYTVKAT TVRTPSWAVD MMRFNINDFV PPGGGTNKI SIPFEYYRIR KVKVEFWPCS PITQGDRGVG STAVILDDNF VTKATALTYD PYVNYSSRHT IPQPFSYHSR Y FTPKPVLD ...String:
MTYPRRRFRR RRHRPRSHLG LILRRRPWLV HPRHRYRWRR KNGIFNTRLS CTFGYTVKAT TVRTPSWAVD MMRFNINDFV PPGGGTNKI SIPFEYYRIR KVKVEFWPCS PITQGDRGVG STAVILDDNF VTKATALTYD PYVNYSSRHT IPQPFSYHSR Y FTPKPVLD STIDYFQPNN KRNQLWLRLQ TSANVDHVGL GIAFENSTYD QDYNIRVTMY VQFREFNLKD PPL

UniProtKB: Capsid protein

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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

BufferpH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: FEI CETA (4k x 4k) / Average electron dose: 35.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 6.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 2011
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE

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