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- EMDB-4550: Cryo-EM structure of human norovirus GII.4 NSW-2012 VLP with T=4 ... -

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Basic information

Entry
Database: EMDB / ID: EMD-4550
TitleCryo-EM structure of human norovirus GII.4 NSW-2012 VLP with T=4 icosahedral symmetry
Map dataCryo-EM structure of GII.4 NSW virus like particle
Sample
  • Virus: Norovirus
Biological speciesNorovirus
Methodsingle particle reconstruction / cryo EM / Resolution: 7.3 Å
AuthorsDevant J / Hansman G
CitationJournal: Antiviral Res / Year: 2019
Title: Heterologous expression of human norovirus GII.4 VP1 leads to assembly of T=4 virus-like particles.
Authors: Jessica M Devant / Götz Hofhaus / David Bhella / Grant S Hansman /
Abstract: Human noroviruses are a leading cause of acute gastroenteritis, yet there are still no vaccines or antivirals available. Expression of the norovirus capsid protein (VP1) in insect cells typically ...Human noroviruses are a leading cause of acute gastroenteritis, yet there are still no vaccines or antivirals available. Expression of the norovirus capsid protein (VP1) in insect cells typically results in the formation of virus-like particles (VLPs) that are morphologically and antigenically comparable to native virions. Indeed, several different norovirus VLP candidates are currently used in clinical trials. So far, structural analysis of norovirus VLPs showed that the capsid has a T = 3 icosahedral symmetry and is composed of 180 copies of VP1 that are folded into three quasi-equivalent subunits (A, B, and C). In this study, the VLP structures of two norovirus GII.4 genetic variants that were identified in 1974 and 2012 were determined using cryo-EM. Surprisingly, we found that greater than 95% of these GII.4 VLPs were larger than virions and 3D reconstruction showed that these VLPs exhibited T = 4 icosahedral symmetry. We also discovered that the T = 4 VLPs presented several novel structural features. The T = 4 particles assembled from 240 copies of VP1 that adopted four quasi-equivalent conformations (A, B, C, and D) and formed two distinct dimers, A/B and C/D. The protruding domains were elevated ∼21 Å off the capsid shell, which was ∼7 Å more than in the previously studied GII.10 T = 3 VLPs. A small cavity and flap-like structure at the icosahedral two-fold axis disrupted the contiguous T = 4 shell. Overall, our findings indicated that GII.4 VP1 sequences assemble into T = 4 VLPs and these larger particles might have important consequences for VLP-based vaccine development.
History
DepositionJan 17, 2019-
Header (metadata) releaseJun 12, 2019-
Map releaseJun 12, 2019-
UpdateJun 26, 2019-
Current statusJun 26, 2019Processing 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
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_4550.png

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Map

FileDownload / File: emd_4550.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM structure of GII.4 NSW virus like particle
Voxel sizeX=Y=Z: 2.27 Å
Density
Contour LevelBy AUTHOR: 0.05 / Movie #1: 0.05
Minimum - Maximum-0.04992078 - 0.19719508
Average (Standard dev.)0.006455011 (±0.022301873)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 581.12 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.272.272.27
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z581.120581.120581.120
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ304304304
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0500.1970.006

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Supplemental data

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Sample components

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Entire : Norovirus

EntireName: Norovirus
Components
  • Virus: Norovirus

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Supramolecule #1: Norovirus

SupramoleculeName: Norovirus / type: virus / ID: 1 / Parent: 0 / NCBI-ID: 142786 / Sci species name: Norovirus / Virus type: VIRUS-LIKE PARTICLE / Virus isolate: OTHER / Virus enveloped: No / Virus empty: Yes
Host (natural)Organism: Homo sapiens (human)
Host systemOrganism: Trichoplusia ni (cabbage looper) / Recombinant strain: High 5 cells
Molecular weightTheoretical: 13.4 MDa
Virus shellShell ID: 1 / Name: VP1 / Diameter: 500.0 Å / T number (triangulation number): 4

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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

Concentration4 mg/mL
BufferpH: 7.4 / Component - Name: PBS
GridModel: Quantifoil R1.2/1.3 / Material: COPPER/RHODIUM / Mesh: 300
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV

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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: 2.7 mm / Nominal magnification: 64000
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Number grids imaged: 1 / Number real images: 364 / Average electron dose: 20.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 7.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1) / Number images used: 10548

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