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- PDB-8i5l: Crystal structure of P domain from norovirus GI.4 capsid protein ... -

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Basic information

Entry
Database: PDB / ID: 8i5l
TitleCrystal structure of P domain from norovirus GI.4 capsid protein in complex with broad specificity antibody single-chain Fv fragment CV-2F5.
Components
  • Capsid protein
  • scFv fragment
KeywordsVIRAL PROTEIN / norovirus / p-domain / capsid / single-chain Fv fragment / antibody-protein complex
Function / homologyCalicivirus coat protein C-terminal / Calicivirus coat protein C-terminal / Calicivirus coat protein / Calicivirus coat protein / Picornavirus/Calicivirus coat protein / Viral coat protein subunit / Capsid protein
Function and homology information
Biological speciesChiba virus
Homo sapiens (human)
MethodX-RAY DIFFRACTION / MOLECULAR REPLACEMENT / Resolution: 2.7 Å
AuthorsKato-Murayama, M. / Murayama, K. / Shirouzu, M.
Funding support Japan, 1items
OrganizationGrant numberCountry
Japan Agency for Medical Research and Development (AMED) 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
DepositionJan 26, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 31, 2024Provider: repository / Type: Initial release
Revision 1.1May 8, 2024Group: Database references / Source and taxonomy / Category: citation / citation_author / entity_src_gen
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _entity_src_gen.pdbx_gene_src_gene / _entity_src_gen.pdbx_gene_src_ncbi_taxonomy_id / _entity_src_gen.pdbx_gene_src_scientific_name
Revision 1.2May 29, 2024Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.3Jun 5, 2024Group: Source and taxonomy / Category: entity_src_gen
Item: _entity_src_gen.gene_src_strain / _entity_src_gen.pdbx_gene_src_scientific_name
Revision 1.4Jun 19, 2024Group: Database references / Category: citation / citation_author
Item: _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Capsid protein
B: Capsid protein
C: Capsid protein
D: scFv fragment
E: scFv fragment


Theoretical massNumber of molelcules
Total (without water)156,3585
Polymers156,3585
Non-polymers00
Water3,189177
1
A: Capsid protein
B: Capsid protein
D: scFv fragment
E: scFv fragment


Theoretical massNumber of molelcules
Total (without water)122,4624
Polymers122,4624
Non-polymers00
Water724
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
C: Capsid protein

C: Capsid protein


Theoretical massNumber of molelcules
Total (without water)67,7922
Polymers67,7922
Non-polymers00
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_656-x+1,y,-z+11
Unit cell
Length a, b, c (Å)176.669, 102.248, 90.793
Angle α, β, γ (deg.)90.000, 109.933, 90.000
Int Tables number5
Space group name H-MC121
Space group name HallC2y
Symmetry operation#1: x,y,z
#2: -x,y,-z
#3: x+1/2,y+1/2,z
#4: -x+1/2,y+1/2,-z
Components on special symmetry positions
IDModelComponents
11C-609-

HOH

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Components

#1: Protein Capsid protein


Mass: 33895.895 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Chiba virus / Strain: GI/Human/Japan/Chiba 407/1987 / Production host: Escherichia coli (E. coli) / References: UniProt: Q9DU46
#2: Antibody scFv fragment


Mass: 27334.945 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 177 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.6 Å3/Da / Density % sol: 52.71 %
Crystal growTemperature: 298 K / Method: vapor diffusion, sitting drop / pH: 7 / Details: Tacsimate

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: ROTATING ANODE / Type: RIGAKU FR-E SUPERBRIGHT / Wavelength: 1.5418 Å
DetectorType: RIGAKU RAXIS IV++ / Detector: IMAGE PLATE / Date: Jul 18, 2018
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.5418 Å / Relative weight: 1
ReflectionResolution: 2.7→41.35 Å / Num. obs: 41894 / % possible obs: 99.9 % / Redundancy: 3.7 % / Biso Wilson estimate: 45.37 Å2 / Rrim(I) all: 0.111 / Net I/σ(I): 14
Reflection shellResolution: 2.7→2.8 Å / Num. unique obs: 4144 / Rrim(I) all: 0.664

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Processing

Software
NameVersionClassification
PHENIX1.13_2998refinement
HKL-2000data reduction
HKL-2000data scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.7→41.35 Å / SU ML: 0.4187 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 27.8119 / Stereochemistry target values: GeoStd + Monomer Library
RfactorNum. reflection% reflection
Rfree0.259 1999 4.77 %
Rwork0.1843 39879 -
obs0.1879 41878 99.8 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 41.66 Å2
Refinement stepCycle: LAST / Resolution: 2.7→41.35 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms10438 0 0 177 10615
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.008510729
X-RAY DIFFRACTIONf_angle_d1.053414661
X-RAY DIFFRACTIONf_chiral_restr0.05751592
X-RAY DIFFRACTIONf_plane_restr0.00771943
X-RAY DIFFRACTIONf_dihedral_angle_d18.47743884
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.7-2.770.36031400.29042801X-RAY DIFFRACTION97.94
2.77-2.840.3511430.26552854X-RAY DIFFRACTION99.87
2.84-2.920.35231420.24782815X-RAY DIFFRACTION99.97
2.92-3.020.35921420.24812838X-RAY DIFFRACTION100
3.02-3.130.34271420.25212832X-RAY DIFFRACTION100
3.13-3.250.32421430.22842846X-RAY DIFFRACTION100
3.25-3.40.30891430.22222857X-RAY DIFFRACTION100
3.4-3.580.32221410.21242812X-RAY DIFFRACTION99.93
3.58-3.80.29331430.21022874X-RAY DIFFRACTION99.77
3.8-4.090.21411440.16692857X-RAY DIFFRACTION100
4.09-4.510.21911410.13182829X-RAY DIFFRACTION99.97
4.51-5.160.17341430.12872852X-RAY DIFFRACTION100
5.16-6.490.22041460.14822889X-RAY DIFFRACTION100
6.49-41.350.18811460.14282923X-RAY DIFFRACTION99.84

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