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- PDB-8a1e: Rabies virus glycoprotein in complex with Fab fragments of 17C7 a... -

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Basic information

Entry
Database: PDB / ID: 8a1e
TitleRabies virus glycoprotein in complex with Fab fragments of 17C7 and 1112-1 neutralizing antibodies
Components
  • Fab 1112-1 heavy chain variable domain
  • Fab 1112-1 light chain variable domain
  • Fab 17C7 heavy chain variable domain
  • Fab 17C7 light chain variable domain
  • Glycoprotein
KeywordsVIRAL PROTEIN / Viral Glycoprotein / Antibody / Complex
Function / homologyRhabdovirus glycoprotein / Rhabdovirus spike glycoprotein / membrane => GO:0016020 / viral envelope / virion membrane / Glycoprotein
Function and homology information
Biological speciesRabies virus strain Pasteur vaccin
Homo sapiens (human)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.83 Å
AuthorsNg, W.M. / Fedosyuk, S. / English, S. / Augusto, G. / Berg, A. / Thorley, L. / Haselon, A.S. / Segireddy, R.R. / Bowden, T.A. / Douglas, A.D.
Funding support United Kingdom, European Union, 7items
OrganizationGrant numberCountry
Wellcome Trust220679/Z/20/Z United Kingdom
Medical Research Council (MRC, United Kingdom)MR/P017339/1 United Kingdom
Medical Research Council (MRC, United Kingdom)MR/S007555/1 United Kingdom
Marie Sklodowska-Curie Actions, FragNET ITN840866European Union
Wellcome Trust060208/Z/00/Z United Kingdom
Wellcome Trust093305/Z/10/Z United Kingdom
Wellcome Trust203141/Z/16/Z United Kingdom
CitationJournal: Cell Host Microbe / Year: 2022
Title: Structure of trimeric pre-fusion rabies virus glycoprotein in complex with two protective antibodies.
Authors: Weng M Ng / Sofiya Fedosyuk / Solomon English / Gilles Augusto / Adam Berg / Luke Thorley / Anna-Sophie Haselon / Rameswara R Segireddy / Thomas A Bowden / Alexander D Douglas /
Abstract: Rabies virus (RABV) causes lethal encephalitis and is responsible for approximately 60,000 deaths per year. As the sole virion-surface protein, the rabies virus glycoprotein (RABV-G) mediates host- ...Rabies virus (RABV) causes lethal encephalitis and is responsible for approximately 60,000 deaths per year. As the sole virion-surface protein, the rabies virus glycoprotein (RABV-G) mediates host-cell entry. RABV-G's pre-fusion trimeric conformation displays epitopes bound by protective neutralizing antibodies that can be induced by vaccination or passively administered for post-exposure prophylaxis. We report a 2.8-Å structure of a RABV-G trimer in the pre-fusion conformation, in complex with two neutralizing and protective monoclonal antibodies, 17C7 and 1112-1, that recognize distinct epitopes. One of these antibodies is a licensed prophylactic (17C7, Rabishield), which we show locks the protein in pre-fusion conformation. Targeted mutations can similarly stabilize RABV-G in the pre-fusion conformation, a key step toward structure-guided vaccine design. These data reveal the higher-order architecture of a key therapeutic target and the structural basis of neutralization by antibodies binding two key antigenic sites, and this will facilitate the development of improved vaccines and prophylactic antibodies.
History
DepositionJun 1, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Aug 17, 2022Provider: repository / Type: Initial release
Revision 1.1Aug 31, 2022Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Sep 28, 2022Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Glycoprotein
D: Fab 17C7 heavy chain variable domain
E: Fab 17C7 light chain variable domain
C: Fab 1112-1 light chain variable domain
B: Fab 1112-1 heavy chain variable domain


Theoretical massNumber of molelcules
Total (without water)106,5715
Polymers106,5715
Non-polymers00
Water0
1
A: Glycoprotein
D: Fab 17C7 heavy chain variable domain
E: Fab 17C7 light chain variable domain
C: Fab 1112-1 light chain variable domain
B: Fab 1112-1 heavy chain variable domain

A: Glycoprotein
D: Fab 17C7 heavy chain variable domain
E: Fab 17C7 light chain variable domain
C: Fab 1112-1 light chain variable domain
B: Fab 1112-1 heavy chain variable domain

A: Glycoprotein
D: Fab 17C7 heavy chain variable domain
E: Fab 17C7 light chain variable domain
C: Fab 1112-1 light chain variable domain
B: Fab 1112-1 heavy chain variable domain


Theoretical massNumber of molelcules
Total (without water)319,71415
Polymers319,71415
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation2

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Components

#1: Protein Glycoprotein /


Mass: 56494.551 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rabies virus strain Pasteur vaccin / Strain: Pasteur vaccins / PV / Production host: Homo sapiens (human) / References: UniProt: P08667
#2: Antibody Fab 17C7 heavy chain variable domain


Mass: 13227.727 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Homo sapiens (human)
#3: Antibody Fab 17C7 light chain variable domain


Mass: 11754.044 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Homo sapiens (human)
#4: Antibody Fab 1112-1 light chain variable domain


Mass: 11668.073 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Production host: Mus musculus (house mouse)
#5: Antibody Fab 1112-1 heavy chain variable domain


Mass: 13426.977 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Production host: Mus musculus (house mouse)

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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: Rabies virus glycoprotein (chain A) in complex with Fab 1112-1 (chains B and C) and Fab 17C7 (chains D and E)
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.33 MDa / Experimental value: NO
Source (natural)Organism: Rabies lyssavirus
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
1150 mMsodium chlorideNaClSodium chloride1
250 mM4-(2-hydroxyethyl)-1-piperazineethanesulfonic acidHEPES1
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Purified sample was concentrated and mixed with 0.07% n-octyl-beta-d-glucoside to final concentrations of 1 mg/mL.
Specimen supportGrid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K
Details: Blot for 3.5 seconds at -15 N blot force before plunging.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 81000 X / Nominal defocus max: 2600 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 3.7 sec. / Electron dose: 44.4 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 12884
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
EM software
IDNameVersionCategory
1cryoSPARC3.1particle selection
2SerialEMimage acquisition
4Gctf1.06CTF correction
7UCSF Chimeramodel fitting
9cryoSPARC3.1initial Euler assignment
10cryoSPARC3.1final Euler assignment
12cryoSPARC3.13D reconstruction
19PHENIX1.19.2_4158model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 3112037
Details: Particles were automatically picked using circular blobs with a diameter of 80-150 Angstrom on cryoSPARC.
SymmetryPoint symmetry: C3 (3 fold cyclic)
3D reconstructionResolution: 2.83 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 458014 / Details: cryoSPARC non-uniform refinement was used. / Symmetry type: POINT
Atomic model buildingB value: 108 / Protocol: RIGID BODY FIT / Space: REAL
Details: Model building was performed with Coot and refined with Phenix.
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 59.69 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00316287
ELECTRON MICROSCOPYf_angle_d0.5238522
ELECTRON MICROSCOPYf_chiral_restr0.0417928
ELECTRON MICROSCOPYf_plane_restr0.00391088
ELECTRON MICROSCOPYf_dihedral_angle_d4.7288855

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