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- PDB-9eap: MNV Allosteric escape mutant V339I + GCDCA -

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Basic information

Entry
Database: PDB / ID: 9eap
TitleMNV Allosteric escape mutant V339I + GCDCA
ComponentsCapsid protein VP1
KeywordsVIRUS / norovirus / escape / allosteric / mutant
Function / homologyCalicivirus coat protein C-terminal / Calicivirus coat protein C-terminal / Calicivirus coat protein / Calicivirus coat protein / virion component / Picornavirus/Calicivirus coat protein / Viral coat protein subunit / host cell cytoplasm / Capsid protein VP1
Function and homology information
Biological speciesMurine norovirus 1
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsSmith, T.J. / Sherman, M.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)NIH 1R01-AI141465 United States
CitationJournal: J Virol / Year: 2025
Title: Murine norovirus allosteric escape mutants mimic gut activation.
Authors: Michael B Sherman / Hong Q Smith / Faith Cox / Christiane E Wobus / Gillian C Lynch / B Montgomery Pettitt / Thomas J Smith /
Abstract: Murine norovirus (MNV) undergoes large conformational changes in response to the environment. The T=3 icosahedral capsid is composed of 180 copies of ~58 kDa VP1 that has N-terminal (N), shell (S), ...Murine norovirus (MNV) undergoes large conformational changes in response to the environment. The T=3 icosahedral capsid is composed of 180 copies of ~58 kDa VP1 that has N-terminal (N), shell (S), and C-terminal protruding (P) domains. In phosphate-buffered saline, the P domains are loosely tethered to the shell and float ~15 Å above the surface. At conditions found in the gut (i.e., low pH with high metal ion and bile salt concentrations), the P domain rotates and drops onto the shell with intra P domain changes that enhance receptor interactions while blocking antibody binding. Two of our monoclonal antibodies (2D3 and 4F9) have broad strain recognition, and the only escape mutants, V339I and D348E, are located on the C'D' loop and ~20 Å from the epitope. Here, we determined the cryo-EM structures of V339I and D348E at neutral pH +/-metal ions and bile salts. These allosteric escape mutants have the activated conformation in the absence of gut triggers. Since this conformation is not recognized by antibodies, it explains how these mutants evade antibody recognition. Dynamic simulations of the P domain further suggest that movement of the C'D' loop may be the rate-limiting step in the conformational change and that V339I increases the motion of the A'B'/E'F' loops compared to the wild-type (WT), facilitating the transition to the activated state. These findings have important implications for norovirus vaccine design since they uncover a form of the viral capsid that should lend superior immune protection against subsequent challenge by wild-type virus.IMPORTANCEImmune protection from norovirus infection is notoriously transient in both humans and mice. Our results strongly suggest that this is likely because the "activated" form of the virus found in gut conditions is not recognized by antibodies created in the circulation. By reversibly presenting one structure in the gut and a completely different antigenic structure in circulation, the gut tissue can be infected in subsequent challenges, while extraintestinal organs are protected. We find here that allosteric escape mutants to the most broadly neutralizing antibodies thwart recognition by transitioning to the activated state without the need for gut triggers (i.e., bile, low pH, or metal ions). These findings are significant because it is now feasible to present the activated form of the virus to the immune system (for example, as a vaccine) to better protect the gut tissue for longer periods of time.
History
DepositionNov 11, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 7, 2025Provider: repository / Type: Initial release
Revision 1.0May 7, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Capsid protein VP1
B: Capsid protein VP1
C: Capsid protein VP1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)177,0996
Polymers175,7503
Non-polymers1,3493
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein Capsid protein VP1 / CP / Coat protein


Mass: 58583.434 Da / Num. of mol.: 3 / Mutation: V339I / Source method: isolated from a natural source / Source: (natural) Murine norovirus 1 / References: UniProt: Q80J94
#2: Chemical ChemComp-CHO / GLYCOCHENODEOXYCHOLIC ACID


Mass: 449.623 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C26H43NO5 / Feature type: SUBJECT OF INVESTIGATION / Comment: detergent*YM
Has ligand of interestY
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Murine norovirus 1 / Type: VIRUS / Entity ID: #1 / Source: NATURAL
Source (natural)Organism: Murine norovirus 1
Details of virusEmpty: NO / Enveloped: NO / Isolate: STRAIN / Type: VIRION
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1500 nm
Image recordingElectron dose: 40 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.21rc1_5134: / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 209918 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00412219
ELECTRON MICROSCOPYf_angle_d0.69916772
ELECTRON MICROSCOPYf_dihedral_angle_d6.0531677
ELECTRON MICROSCOPYf_chiral_restr0.0451920
ELECTRON MICROSCOPYf_plane_restr0.0052183

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