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- EMDB-2397: Cryo-EM reconstruction of enterovirus 71 in complex with a neutra... -

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Basic information

Entry
Database: EMDB / ID: EMD-2397
TitleCryo-EM reconstruction of enterovirus 71 in complex with a neutralizing antibody E18
Map dataReconstruction of EV71 E18 Fab complex
Sample
  • Sample: Reconstruction of human enterovirus 71 in complex with Fab fragment of antibody E18
  • Virus: Human enterovirus 71
  • Protein or peptide: Fab EV18/4/D6-1/F1/G9
KeywordsEV71 / human / enterovirus 71 / virus / antibody / Fab / E18 / complex / neutralizing / pathogen
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 / 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 speciesMus musculus (house mouse) / Human enterovirus 71
Methodsingle particle reconstruction / cryo EM / Resolution: 10.0 Å
AuthorsPlevka P / Perera R / Cardosa J / Suksatu A / Kuhn RJ / Rossmann MG
CitationJournal: Proc Natl Acad Sci U S A / Year: 2014
Title: Neutralizing antibodies can initiate genome release from human enterovirus 71.
Authors: Pavel Plevka / Pei-Yin Lim / Rushika Perera / Jane Cardosa / Ampa Suksatu / Richard J Kuhn / Michael G Rossmann /
Abstract: Antibodies were prepared by immunizing mice with empty, immature particles of human enterovirus 71 (EV71), a picornavirus that causes severe neurological disease in young children. The capsid ...Antibodies were prepared by immunizing mice with empty, immature particles of human enterovirus 71 (EV71), a picornavirus that causes severe neurological disease in young children. The capsid structure of these empty particles is different from that of the mature virus and is similar to "A" particles encountered when picornaviruses recognize a potential host cell before genome release. The monoclonal antibody E18, generated by this immunization, induced a conformational change when incubated at temperatures between 4 °C and 37 °C with mature virus, transforming infectious virions into A particles. The resultant loss of genome that was observed by cryo-EM and a fluorescent SYBR Green dye assay inactivated the virus, establishing the mechanism by which the virus is inactivated and demonstrating that the E18 antibody has potential as an anti-EV71 therapy. The antibody-mediated virus neutralization by the induction of genome release has not been previously demonstrated. Furthermore, the present results indicate that antibodies with genome-release activity could also be produced for other picornaviruses by immunization with immature particles.
History
DepositionJun 11, 2013-
Header (metadata) releaseJul 17, 2013-
Map releaseFeb 5, 2014-
UpdateFeb 19, 2014-
Current statusFeb 19, 2014Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1
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  • Surface view colored by radius
  • Surface level: 1
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  • Surface view with fitted model
  • Atomic models: PDB-4c0u
  • Surface level: 1
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-4c0u
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_2397.map.gz / Format: CCP4 / Size: 29.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of EV71 E18 Fab complex
Voxel sizeX=Y=Z: 2.48 Å
Density
Contour LevelBy AUTHOR: 1.0 / Movie #1: 1
Minimum - Maximum-4.85787678 - 4.86891079
Average (Standard dev.)0.02293283 (±0.91755581)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-100-100-100
Dimensions200200200
Spacing200200200
CellA=B=C: 496.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.482.482.48
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z496.000496.000496.000
α/β/γ90.00090.00090.000
start NX/NY/NZ00-40
NX/NY/NZ555581
MAP C/R/S123
start NC/NR/NS-100-100-100
NC/NR/NS200200200
D min/max/mean-4.8584.8690.023

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Supplemental data

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Sample components

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Entire : Reconstruction of human enterovirus 71 in complex with Fab fragme...

EntireName: Reconstruction of human enterovirus 71 in complex with Fab fragment of antibody E18
Components
  • Sample: Reconstruction of human enterovirus 71 in complex with Fab fragment of antibody E18
  • Virus: Human enterovirus 71
  • Protein or peptide: Fab EV18/4/D6-1/F1/G9

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Supramolecule #1000: Reconstruction of human enterovirus 71 in complex with Fab fragme...

SupramoleculeName: Reconstruction of human enterovirus 71 in complex with Fab fragment of antibody E18
type: sample / ID: 1000
Oligomeric state: Icosahedral virus and 60 copies of bound Fab
Number unique components: 2
Molecular weightTheoretical: 6.8 MDa

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Supramolecule #1: Human enterovirus 71

SupramoleculeName: Human enterovirus 71 / type: virus / ID: 1 / Name.synonym: EV71 / NCBI-ID: 39054 / Sci species name: Human enterovirus 71 / Virus type: VIRION / Virus isolate: SEROTYPE / Virus enveloped: No / Virus empty: No / Syn species name: EV71 / Sci species serotype: EV-174
Host (natural)Organism: Homo sapiens (human) / synonym: VERTEBRATES
Molecular weightTheoretical: 3.2 MDa
Virus shellShell ID: 1 / Name: capsid / Diameter: 280 Å / T number (triangulation number): 1

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Macromolecule #1: Fab EV18/4/D6-1/F1/G9

MacromoleculeName: Fab EV18/4/D6-1/F1/G9 / type: protein_or_peptide / ID: 1 / Name.synonym: E18 / Number of copies: 60 / Oligomeric state: monomer / Recombinant expression: No
Source (natural)Organism: Mus musculus (house mouse) / synonym: house mouse
Molecular weightTheoretical: 4.6 KDa

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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: 8 / Details: 10 mM Tris-HCl, pH 8.0, 0.5 M NaCl, and 1 mM EDTA
GridDetails: Quantifoin R1.2/1.3 Holey carbon 200Mesh
VitrificationCryogen name: ETHANE / Chamber humidity: 70 % / Chamber temperature: 100 K / Instrument: HOMEMADE PLUNGER / Method: Blot for 2 second before plunging

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Electron microscopy

MicroscopeFEI/PHILIPS CM200FEG
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 4950 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 0.0045 µm / Nominal defocus min: 0.00075 µm / Nominal magnification: 50000
Sample stageSpecimen holder model: OTHER
TemperatureMin: 95 K / Max: 120 K / Average: 119 K
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification
DateSep 10, 2013
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: NIKON SUPER COOLSCAN 9000 / Number real images: 112 / Average electron dose: 18 e/Å2 / Bits/pixel: 16

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Image processing

CTF correctionDetails: CTF determined for all particles from one film, phase flip
Final two d classificationNumber classes: 122
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 10.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: eman, eman2 / Number images used: 8335
DetailsParticles were boxed with e2boxer.py

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Atomic model buiding 1

Initial modelPDB ID:

Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B / Chain - #2 - Chain ID: C
SoftwareName: EMFit
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Rcrit
Output model

PDB-4c0u:
Cryo-EM reconstruction of enterovirus 71 in complex with a neutralizing antibody E18

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