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- PDB-8jg5: Cryo-EM structure of the GI.4 Chiba VLP complexed with the CV-1A1... -

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Basic information

Entry
Database: PDB / ID: 8jg5
TitleCryo-EM structure of the GI.4 Chiba VLP complexed with the CV-1A1 Fv-clasp
Components
  • VH,SARAH
  • VL,SARAH
  • VP1
KeywordsVIRUS / NOROVIRUS / GI.4 / VIRUS LIKE PARTICLE / human monoclonal antibody / Fv-clasp
Biological speciesNorovirus
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.04 Å
AuthorsHosaka, T. / Katsura, K. / Kimura-Someya, T. / Someya, Y. / Shirouzu, M.
Funding support Japan, 2items
OrganizationGrant numberCountry
Japan Agency for Medical Research and Development (AMED)JP21fk0108121 Japan
Japan Agency for Medical Research and Development (AMED)JP22fk0108121 Japan
CitationJournal: J Virol / Year: 2024
Title: Structural analyses of the GI.4 norovirus by cryo-electron microscopy and X-ray crystallography revealing binding sites for human monoclonal antibodies.
Authors: Tomomi Kimura-Someya / Kazushige Katsura / Miyuki Kato-Murayama / Toshiaki Hosaka / Tomomi Uchikubo-Kamo / Kentaro Ihara / Kazuharu Hanada / Shin Sato / Kazutaka Murayama / Michiyo Kataoka / ...Authors: Tomomi Kimura-Someya / Kazushige Katsura / Miyuki Kato-Murayama / Toshiaki Hosaka / Tomomi Uchikubo-Kamo / Kentaro Ihara / Kazuharu Hanada / Shin Sato / Kazutaka Murayama / Michiyo Kataoka / Mikako Shirouzu / Yuichi Someya /
Abstract: Noroviruses are major causative agents of acute nonbacterial gastroenteritis in humans. There are neither antiviral therapeutic agents nor vaccines for noroviruses at this time. To evaluate the ...Noroviruses are major causative agents of acute nonbacterial gastroenteritis in humans. There are neither antiviral therapeutic agents nor vaccines for noroviruses at this time. To evaluate the potential usefulness of two previously isolated human monoclonal antibody fragments, CV-1A1 and CV-2F5, we first conducted a single-particle analysis to determine the cryo-electron microscopy structure of virus-like particles (VLPs) from the genogroup I genotype 4 (GI.4) Chiba strain uniformly coated with CV-1A1 fragments. The results revealed that the GI.4-specific CV-1A1 antibody bound to the P2 subdomain, in which amino acids are less conserved and variable. Interestingly, a part of the CV-1A1 intrudes into the histo-blood group antigen-binding site, suggesting that this antibody might exert neutralizing activity. Next, we determined the crystal structure of the protruding (P) domain of the capsid protein in the complex form with the CV-2F5 antibody fragment. Consistent with the cross-reactivity, the CV-2F5 bound to the P1 subdomain, which is rich in amino acids conserved among the GI strains, and moreover induced a disruption of Chiba VLPs. These results suggest that the broadly reactive CV-2F5 antibody can be used as both a universal detection reagent and an antiviral drug for GI noroviruses.
IMPORTANCE: We conducted the structural analyses of the VP1 protein from the GI.4 Chiba norovirus to identify the binding sites of the previously isolated human monoclonal antibodies CV-1A1 and CV- ...IMPORTANCE: We conducted the structural analyses of the VP1 protein from the GI.4 Chiba norovirus to identify the binding sites of the previously isolated human monoclonal antibodies CV-1A1 and CV-2F5. The cryo-electron microscopy of the Chiba virus-like particles (VLPs) complexed with the Fv-clasp forms of GI.4-specific CV-1A1 revealed that this antibody binds to the highly variable P2 subdomain, suggesting that this antibody may have neutralizing ability against the GI.4 strains. X-ray crystallography revealed that the CV-2F5 antibody bound to the P1 subdomain, which is rich in conserved amino acids. This result is consistent with the ability of the CV-2F5 antibody to react with a wide variety of GI norovirus strains. It is also found that the CV-2F5 antibody caused a disruption of VLPs. Our findings, together with previous reports on the structures of VP1 proteins and VLPs, are expected to open a path for the structure-based development of antivirals and vaccines against norovirus disease.
History
DepositionMay 19, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Apr 17, 2024Provider: repository / Type: Initial release
Revision 1.1May 29, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: VP1
B: VP1
C: VH,SARAH
D: VL,SARAH
E: VH,SARAH
F: VL,SARAH
G: VP1


Theoretical massNumber of molelcules
Total (without water)250,4067
Polymers250,4067
Non-polymers00
Water0
1
A: VP1
B: VP1
C: VH,SARAH
D: VL,SARAH
E: VH,SARAH
F: VL,SARAH
G: VP1
x 60


Theoretical massNumber of molelcules
Total (without water)15,024,348420
Polymers15,024,348420
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation59
2


  • Idetical with deposited unit
  • icosahedral asymmetric unit
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
A: VP1
B: VP1
C: VH,SARAH
D: VL,SARAH
E: VH,SARAH
F: VL,SARAH
G: VP1
x 5


  • icosahedral pentamer
  • 1.25 MDa, 35 polymers
Theoretical massNumber of molelcules
Total (without water)1,252,02935
Polymers1,252,02935
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation4
4
A: VP1
B: VP1
C: VH,SARAH
D: VL,SARAH
E: VH,SARAH
F: VL,SARAH
G: VP1
x 6


  • icosahedral 23 hexamer
  • 1.5 MDa, 42 polymers
Theoretical massNumber of molelcules
Total (without water)1,502,43542
Polymers1,502,43542
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation5
5


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit, std point frame
TypeNameSymmetry operationNumber
transform to point frame1
SymmetryPoint symmetry: (Schoenflies symbol: I (icosahedral))

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Components

#1: Protein VP1


Mass: 58108.332 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Details: GenBank ID AB042808 / Source: (gene. exp.) Norovirus / Gene: ORF2 / Production host: Spodoptera frugiperda (fall armyworm)
#2: Antibody VH,SARAH


Mass: 19847.254 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: VH-SARAH chimera of linker (residues (-6)-0), VH (residues 1-118), and SARAH (residues 119-171)
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)
#3: Antibody VL,SARAH


Mass: 18193.150 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: VL-SARAH chimera of linker (residues (-6)-0), VL (residues 1-112), and SARAH (residues 113-162)
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)

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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: Viral protein 1 with its antibody fragments / Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Source (natural)Organism: Norovirus
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 8
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: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TECNAI ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1600 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.20.1_4487: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: I (icosahedral)
3D reconstructionResolution: 3.04 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 47125 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00416478
ELECTRON MICROSCOPYf_angle_d0.64122486
ELECTRON MICROSCOPYf_dihedral_angle_d4.2422222
ELECTRON MICROSCOPYf_chiral_restr0.0442489
ELECTRON MICROSCOPYf_plane_restr0.0052955

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