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- EMDB-6200: Cryo-electron microscopy of Enterovirus 71 (EV71) in complex with... -

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Basic information

Entry
Database: EMDB / ID: EMD-6200
TitleCryo-electron microscopy of Enterovirus 71 (EV71) in complex with Fab fragments of neutralizing antibody 22A12
Map dataReconstruction of EV71 procapsid in complex with Fab22A12
Sample
  • Sample: Fab fragment of 22A12 monoclonal antibody bound to Enterovirus 71 procapsid at the VP1 GH Loop
  • Virus: Human enterovirus 71
  • Protein or peptide: neutralizing antibody 22A12 Fab
KeywordsEV71 / picornavirus / Mab22A12 / antibody / Fab / neutralization / canyon / enterovirus / enterovirus 71
Function / homology
Function and homology information


symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of MDA-5 activity / picornain 2A / symbiont-mediated suppression of host mRNA export from nucleus / symbiont genome entry into host cell via pore formation in plasma membrane / picornain 3C / T=pseudo3 icosahedral viral capsid / host cell cytoplasmic vesicle membrane / endocytosis involved in viral entry into host cell / cytoplasmic vesicle membrane / : ...symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of MDA-5 activity / picornain 2A / symbiont-mediated suppression of host mRNA export from nucleus / symbiont genome entry into host cell via pore formation in plasma membrane / picornain 3C / T=pseudo3 icosahedral viral capsid / host cell cytoplasmic vesicle membrane / endocytosis involved in viral entry into host cell / cytoplasmic vesicle membrane / : / nucleoside-triphosphate phosphatase / protein complex oligomerization / monoatomic ion channel activity / RNA helicase activity / induction by virus of host autophagy / RNA-directed RNA polymerase / symbiont-mediated suppression of host gene expression / viral RNA genome replication / cysteine-type endopeptidase activity / RNA-dependent RNA polymerase activity / DNA-templated transcription / host cell nucleus / structural molecule activity / virion attachment to host cell / ATP hydrolysis activity / proteolysis / RNA binding / ATP binding / metal ion binding
Similarity search - Function
Poliovirus 3A protein-like / Poliovirus 3A protein like / Picornavirus 2B protein / Poliovirus core protein 3a, soluble domain / Picornavirus 2B protein / Peptidase C3, picornavirus core protein 2A / Picornavirus core protein 2A / Picornavirus coat protein VP4 / Picornavirus coat protein (VP4) / Picornavirales 3C/3C-like protease domain ...Poliovirus 3A protein-like / Poliovirus 3A protein like / Picornavirus 2B protein / Poliovirus core protein 3a, soluble domain / Picornavirus 2B protein / Peptidase C3, picornavirus core protein 2A / Picornavirus core protein 2A / Picornavirus coat protein VP4 / Picornavirus coat protein (VP4) / Picornavirales 3C/3C-like protease domain / Picornavirales 3C/3C-like protease domain profile. / Peptidase C3A/C3B, picornaviral / 3C cysteine protease (picornain 3C) / Picornavirus capsid / picornavirus capsid protein / Helicase, superfamily 3, single-stranded RNA virus / Superfamily 3 helicase of positive ssRNA viruses domain profile. / Helicase, superfamily 3, single-stranded DNA/RNA virus / RNA helicase / Picornavirus/Calicivirus coat protein / Viral coat protein subunit / RNA-directed RNA polymerase, C-terminal domain / Viral RNA-dependent RNA polymerase / Reverse transcriptase/Diguanylate cyclase domain / RNA-directed RNA polymerase, catalytic domain / RdRp of positive ssRNA viruses catalytic domain profile. / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / Peptidase S1, PA clan, chymotrypsin-like fold / Peptidase S1, PA clan / DNA/RNA polymerase superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Biological speciesunidentified (others) / Human enterovirus 71
Methodsingle particle reconstruction / cryo EM / Resolution: 8.8 Å
AuthorsShingler KL / Cifuente JO / Ashley RE / Makhov AM / Conway JF / Hafenstein S
CitationJournal: J Virol / Year: 2015
Title: The enterovirus 71 procapsid binds neutralizing antibodies and rescues virus infection in vitro.
Authors: Kristin L Shingler / Javier O Cifuente / Robert E Ashley / Alexander M Makhov / James F Conway / Susan Hafenstein /
Abstract: Enterovirus 71 (EV71) is responsible for seasonal outbreaks of hand, foot, and mouth disease in the Asia-Pacific region. The virus has the capability to cause severe disease and death, especially in ...Enterovirus 71 (EV71) is responsible for seasonal outbreaks of hand, foot, and mouth disease in the Asia-Pacific region. The virus has the capability to cause severe disease and death, especially in young children. Although several vaccines are currently in clinical trials, no vaccines or therapeutics have been approved for use. Previous structural studies have revealed that two antigenically distinct capsid forms are produced in EV71-infected cells: an expanded empty capsid, sometimes called a procapsid, and the infectious virus. Specifically, an immunodominant epitope of EV71 that maps to the virus canyon is structurally different in the procapsid and virus. This structure-function study shows that the procapsid can sequester antibodies, thus enhancing EV71 infection in vitro. The results presented here suggest that, due to conformational differences between the EV71 procapsid and virus, the presence of the procapsid in natural virus infections should be considered in the future design of vaccines or therapeutics.
IMPORTANCE: In a picornavirus infection, both an infectious and a noninfectious empty capsid, sometimes referred to as a procapsid, are produced. It was novel to discover that the procapsid form of ...IMPORTANCE: In a picornavirus infection, both an infectious and a noninfectious empty capsid, sometimes referred to as a procapsid, are produced. It was novel to discover that the procapsid form of EV71 was expanded and antigenically distinct from the infectious virus. Previously, it had been supposed that this empty capsid was an off-pathway dead end or at best served for storage of pentameric subunits, which was later shown to be unlikely. It remains unexplained why picornaviruses evolutionarily conserve the wasteful production of so much noninfectious capsid. Here, we demonstrate that the EV71 procapsid has different antigenic properties than the infectious virus. Thus, the procapsid has the capacity to sequester neutralizing antibody and protect the virus, promoting or restoring a successful infection in vitro. This important observation should be considered in the future design and development of vaccines and therapeutics.
History
DepositionNov 21, 2014-
Header (metadata) releaseDec 10, 2014-
Map releaseDec 10, 2014-
UpdateJan 28, 2015-
Current statusJan 28, 2015Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 1
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-3j91, PDB-3j93
  • Surface level: 1
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-3j91
  • Imaged by Jmol
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-3j93
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_6200.tif

Downloads & links

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Map

FileDownload / File: emd_6200.map.gz / Format: CCP4 / Size: 365 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of EV71 procapsid in complex with Fab22A12
Voxel sizeX=Y=Z: 1.27 Å
Density
Contour LevelBy AUTHOR: 1.0 / Movie #1: 1
Minimum - Maximum-4.29300165 - 8.482726100000001
Average (Standard dev.)0.0 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-230-230-230
Dimensions461461461
Spacing461461461
CellA=B=C: 585.47 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.271.271.27
M x/y/z461461461
origin x/y/z0.0000.0000.000
length x/y/z585.470585.470585.470
α/β/γ90.00090.00090.000
start NX/NY/NZ-72-72-72
NX/NY/NZ145145145
MAP C/R/S123
start NC/NR/NS-230-230-230
NC/NR/NS461461461
D min/max/mean-4.2938.483-0.000

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Supplemental data

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Sample components

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Entire : Fab fragment of 22A12 monoclonal antibody bound to Enterovirus 71...

EntireName: Fab fragment of 22A12 monoclonal antibody bound to Enterovirus 71 procapsid at the VP1 GH Loop
Components
  • Sample: Fab fragment of 22A12 monoclonal antibody bound to Enterovirus 71 procapsid at the VP1 GH Loop
  • Virus: Human enterovirus 71
  • Protein or peptide: neutralizing antibody 22A12 Fab

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Supramolecule #1000: Fab fragment of 22A12 monoclonal antibody bound to Enterovirus 71...

SupramoleculeName: Fab fragment of 22A12 monoclonal antibody bound to Enterovirus 71 procapsid at the VP1 GH Loop
type: sample / ID: 1000 / Number unique components: 2

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Supramolecule #1: Human enterovirus 71

SupramoleculeName: Human enterovirus 71 / type: virus / ID: 1 / Name.synonym: EV71
Details: Virus procapsid was complexed with Fab fragments of neutralizing antibody 22A12.
NCBI-ID: 39054 / Sci species name: Human enterovirus 71 / Sci species strain: 1095/Shiga / Database: NCBI / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: No / Virus empty: Yes / Syn species name: EV71
Host (natural)Organism: Homo sapiens (human) / synonym: VERTEBRATES
Virus shellShell ID: 1 / Diameter: 300 Å / T number (triangulation number): 3

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Macromolecule #1: neutralizing antibody 22A12 Fab

MacromoleculeName: neutralizing antibody 22A12 Fab / type: protein_or_peptide / ID: 1 / Recombinant expression: No / Database: NCBI
Source (natural)Organism: unidentified (others)

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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

Concentration1.0 mg/mL
BufferpH: 7.5 / Details: 10 mM Tris, 200 mM NaCl, 50 mM MgCl2
GridDetails: holey carbon Quantifoil EM grids
VitrificationCryogen name: ETHANE-PROPANE MIXTURE / Instrument: FEI VITROBOT MARK III

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Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal magnification: 50000
Sample stageSpecimen holder model: GATAN LIQUID NITROGEN
DateJul 15, 2011
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: NIKON SUPER COOLSCAN 9000 / Digitization - Sampling interval: 6.35 µm
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: CTFFind3
Final reconstructionResolution.type: BY AUTHOR / Resolution: 8.8 Å / Resolution method: OTHER / Software - Name: EMAN, EMAN2, AUTO3DEM, Chimera / Number images used: 15226
DetailsProcessing was completed with EMAN2 and AUTO3DEM.

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Atomic model buiding 1

Initial modelPDB ID:

4gmp

SoftwareName: Situs
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-3j91:
Cryo-electron microscopy of Enterovirus 71 (EV71) procapsid in complex with Fab fragments of neutralizing antibody 22A12

PDB-3j93:
Fitting of Fab into the cryoEM density map of EV71 procapsid in complex with Fab22A12

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