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- EMDB-3137: Multiple capsid-stabilizing protein-RNA and protein-protein inter... -

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Basic information

Entry
Database: EMDB / ID: EMD-3137
TitleMultiple capsid-stabilizing protein-RNA and protein-protein interactions revealed in a high-resolution structure of an emerging picornavirus causing neonatal sepsis
Map dataReconstruction of human parechovirus 3
Sample
  • Sample: Human parechovirus 3 capsid structure
  • Virus: Human parechovirus 3
Keywordspicornavirus / parechovirus / human parechovirus 3 / HPeV3 / neonatal sepsis / cryoEM / image processing / single particle anaylsis
Function / homology
Function and homology information


RNA-protein covalent cross-linking / : / host cell cytoplasmic vesicle membrane / cytoplasmic vesicle membrane / : / viral capsid / protein complex oligomerization / monoatomic ion channel activity / RNA helicase activity / symbiont entry into host cell ...RNA-protein covalent cross-linking / : / host cell cytoplasmic vesicle membrane / cytoplasmic vesicle membrane / : / viral capsid / protein complex oligomerization / monoatomic ion channel activity / RNA helicase activity / symbiont entry into host cell / viral RNA genome replication / cysteine-type endopeptidase activity / RNA-dependent RNA polymerase activity / DNA-templated transcription / structural molecule activity / virion attachment to host cell / RNA binding / ATP binding
Similarity search - Function
Viral polyprotein, parechovirus P3B / Parechovirus Genome-linked protein / Viral polyprotein, parechovirus P3A / Picornaviridae P3A protein / Helicase/polymerase/peptidase polyprotein, Calicivirus-type / Picornavirales 3C/3C-like protease domain / Picornavirales 3C/3C-like protease domain profile. / Peptidase C3A/C3B, picornaviral / 3C cysteine protease (picornain 3C) / Picornavirus capsid ...Viral polyprotein, parechovirus P3B / Parechovirus Genome-linked protein / Viral polyprotein, parechovirus P3A / Picornaviridae P3A protein / Helicase/polymerase/peptidase polyprotein, Calicivirus-type / 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. / Peptidase S1, PA clan, chymotrypsin-like fold / DNA/RNA polymerase superfamily / Peptidase S1, PA clan / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Biological speciesHuman parechovirus 3
Methodsingle particle reconstruction / cryo EM / negative staining / Resolution: 4.3 Å
AuthorsShakeel S / Westerhuis BM / Domanska A / Koning RI / Matadeen R / Koster AJ / Bakker AQ / Beaumont T / Wolthers KC / Butcher SJ
CitationJournal: Nat Commun / Year: 2016
Title: Multiple capsid-stabilizing interactions revealed in a high-resolution structure of an emerging picornavirus causing neonatal sepsis.
Authors: Shabih Shakeel / Brenda M Westerhuis / Ausra Domanska / Roman I Koning / Rishi Matadeen / Abraham J Koster / Arjen Q Bakker / Tim Beaumont / Katja C Wolthers / Sarah J Butcher /
Abstract: The poorly studied picornavirus, human parechovirus 3 (HPeV3) causes neonatal sepsis with no therapies available. Our 4.3-Å resolution structure of HPeV3 on its own and at 15 Å resolution in ...The poorly studied picornavirus, human parechovirus 3 (HPeV3) causes neonatal sepsis with no therapies available. Our 4.3-Å resolution structure of HPeV3 on its own and at 15 Å resolution in complex with human monoclonal antibody Fabs demonstrates the expected picornavirus capsid structure with three distinct features. First, 25% of the HPeV3 RNA genome in 60 sites is highly ordered as confirmed by asymmetric reconstruction, and interacts with conserved regions of the capsid proteins VP1 and VP3. Second, the VP0 N terminus stabilizes the capsid inner surface, in contrast to other picornaviruses where on expulsion as VP4, it forms an RNA translocation channel. Last, VP1's hydrophobic pocket, the binding site for the antipicornaviral drug, pleconaril, is blocked and thus inappropriate for antiviral development. Together, these results suggest a direction for development of neutralizing antibodies, antiviral drugs based on targeting the RNA-protein interactions and dissection of virus assembly on the basis of RNA nucleation.
History
DepositionSep 3, 2015-
Header (metadata) releaseJan 13, 2016-
Map releaseJul 27, 2016-
UpdateAug 10, 2016-
Current statusAug 10, 2016Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
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  • Surface view with fitted model
  • Atomic models: PDB-5apm
  • Surface level: 0.05
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-5apm
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Structure viewerEM map:
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Supplemental images

emd_3137.tif

Downloads & links

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Map

FileDownload / File: emd_3137.map.gz / Format: CCP4 / Size: 238.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of human parechovirus 3
Voxel sizeX=Y=Z: 1.14 Å
Density
Contour LevelBy AUTHOR: 0.05 / Movie #1: 0.05
Minimum - Maximum-0.11400025 - 0.20041895
Average (Standard dev.)0.00029651 (±0.01687933)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-200-200-200
Dimensions400400400
Spacing400400400
CellA=B=C: 456.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.141.141.14
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z456.000456.000456.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-200-200-200
NC/NR/NS400400400
D min/max/mean-0.1140.2000.000

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Supplemental data

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Sample components

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Entire : Human parechovirus 3 capsid structure

EntireName: Human parechovirus 3 capsid structure
Components
  • Sample: Human parechovirus 3 capsid structure
  • Virus: Human parechovirus 3

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Supramolecule #1000: Human parechovirus 3 capsid structure

SupramoleculeName: Human parechovirus 3 capsid structure / type: sample / ID: 1000
Details: The purified human parechovirus 3 virions were monodisperse. They were formaldehyde fixed.
Oligomeric state: icosahedrally-symmetric virus with 60 copies each of VP0, VP3 and VP1
Number unique components: 3
Molecular weightExperimental: 5 MDa / Theoretical: 5 MDa
Method: theoretical weight determination from protein sequences, excluding viral genome.

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Supramolecule #1: Human parechovirus 3

SupramoleculeName: Human parechovirus 3 / type: virus / ID: 1 / Name.synonym: HPeV3 / NCBI-ID: 195055 / Sci species name: Human parechovirus 3 / Virus type: VIRION / Virus isolate: OTHER / Virus enveloped: No / Virus empty: No / Syn species name: HPeV3
Host (natural)Organism: Homo sapiens (human) / synonym: VERTEBRATES
Host systemOrganism: Chlorocebus aethiops (grivet) / Recombinant cell: Vero
Molecular weightExperimental: 5 MDa / Theoretical: 5 MDa
Virus shellShell ID: 1 / Diameter: 280 Å / T number (triangulation number): 1

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Experimental details

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Structure determination

Methodnegative staining, cryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1 mg/mL
BufferpH: 7.5 / Details: 10 mM Tris-HCL, 150 mM NaCl, 1 mM MgCl2
StainingType: NEGATIVE / Details: Unstained
GridDetails: Quantifoil holey carbon on copper grids
VitrificationCryogen name: ETHANE / Chamber humidity: 92 % / Chamber temperature: 92 K / Instrument: LEICA EM GP / Method: Blot for 2 seconds on one side before plunging.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 0.01 mm / Nominal defocus max: 2.34 µm / Nominal defocus min: 0.42 µm / Nominal magnification: 59000
Sample stageSpecimen holder: Liquid nitrogen cooled. / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
TemperatureMin: 80 K / Max: 95 K / Average: 87 K
Alignment procedureLegacy - Astigmatism: Done as part of Cs corrector routine.
DetailsCs corrector was used during imaging.
DateJan 21, 2014
Image recordingCategory: CCD / Film or detector model: FEI FALCON II (4k x 4k) / Number real images: 6604 / Average electron dose: 36 e/Å2
Details: Total number of images collected were 6604. Total number of images used in the reconstruction were 1028. Every image is the average of seven aligned frames recorded by the direct electron detector.
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: Each micrograph
Final two d classificationNumber classes: 4
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 4.3 Å / Resolution method: OTHER
Software - Name: Ethan, CTFFIND3, EMAN1, EMAN2, AUTO3DEM, RELION, ResMap
Number images used: 8889
DetailsMovie alignment done using motioncorr software. CTF estimated using CTFFIND3. Particles were selected using Ethan. Bad particles discarded by eye in Eman1 BOXER. Random model generated using Auto3dem. 2d and 3d class averaging done in Relion. Final reconstruction done in Relion.
FSC plot (resolution estimation)

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