|Entry||Database: EMDB / ID: EMD-4549|
|Title||Cryo-EM structure of the GII.4 CHDC-1974 VLP with T=4 icosahedral symmetry|
|Method||single particle reconstruction / cryo EM / Resolution: 6.1 Å|
|Authors||Devant J / Hansman G|
|Citation||Journal: 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 1976 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) that 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.
|Date||Deposition: Jan 17, 2019 / Header (metadata) release: Jun 12, 2019 / Map release: Jun 12, 2019 / Update: Jun 12, 2019|
|Structure viewer||EM map: |
Downloads & links
|File||Download / File: emd_4549.map.gz / Format: CCP4 / Size: 476.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 1.375 Å|
|Symmetry||Space group: 1|
CCP4 map header:
|Entire||Name: Norovirus / Number of components: 1|
-Component #1: virus, Norovirus
|Virus||Name: Norovirus / Class: VIRUS-LIKE PARTICLE / Empty: Yes / Enveloped: No / Isolate: OTHER|
|Mass||Theoretical: 13.4 MDa|
|Source (engineered)||Expression System: Trichoplusia ni (cabbage looper) / Cell of expression system: High5 cells|
|Source (natural)||Host Species: Homo sapiens (human)|
|Shell #1||Name of element: VP1 / Diameter: 500.0 Å / T number (triangulation number): 4|
|Specimen||Specimen state: Particle / Method: cryo EM|
|Sample solution||Specimen conc.: 4 mg/mL / pH: 7.4|
|Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Imaging||Microscope: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 20 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Magnification: 64000.0 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD|
|Specimen Holder||Model: OTHER|
|Image acquisition||Number of digital images: 591|
|Processing||Method: single particle reconstruction / Applied symmetry: I (icosahedral) / Number of projections: 42485|
|3D reconstruction||Software: cryoSPARC / Resolution: 6.1 Å / Resolution method: FSC 0.143 CUT-OFF|
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