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- PDB-7t5o: VFLIP Spike Trimer with GAR03 -

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Basic information

Entry
Database: PDB / ID: 7t5o
TitleVFLIP Spike Trimer with GAR03
Components
  • GAR03 Fab heavy chain
  • GAR03 Fab light chain
  • Spike glycoprotein
KeywordsVIRAL PROTEIN/IMMUNE SYSTEM / spike / Fab / antibody / COVID / VIRAL PROTEIN-IMMUNE SYSTEM complex
Biological speciesSevere acute respiratory syndrome-related coronavirus
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.39 Å
AuthorsSobti, M. / Stewart, A.G. / Rouet, R. / Langley, D.B.
Funding support Australia, 1items
OrganizationGrant numberCountry
Australian Research Council (ARC)DE190100985 Australia
CitationJournal: Nat Commun / Year: 2023
Title: Broadly neutralizing SARS-CoV-2 antibodies through epitope-based selection from convalescent patients.
Authors: Romain Rouet / Jake Y Henry / Matt D Johansen / Meghna Sobti / Harikrishnan Balachandran / David B Langley / Gregory J Walker / Helen Lenthall / Jennifer Jackson / Stephanie Ubiparipovic / ...Authors: Romain Rouet / Jake Y Henry / Matt D Johansen / Meghna Sobti / Harikrishnan Balachandran / David B Langley / Gregory J Walker / Helen Lenthall / Jennifer Jackson / Stephanie Ubiparipovic / Ohan Mazigi / Peter Schofield / Deborah L Burnett / Simon H J Brown / Marianne Martinello / Bernard Hudson / Nicole Gilroy / Jeffrey J Post / Anthony Kelleher / Hans-Martin Jäck / Christopher C Goodnow / Stuart G Turville / William D Rawlinson / Rowena A Bull / Alastair G Stewart / Philip M Hansbro / Daniel Christ /
Abstract: Emerging variants of concern (VOCs) are threatening to limit the effectiveness of SARS-CoV-2 monoclonal antibodies and vaccines currently used in clinical practice; broadly neutralizing antibodies ...Emerging variants of concern (VOCs) are threatening to limit the effectiveness of SARS-CoV-2 monoclonal antibodies and vaccines currently used in clinical practice; broadly neutralizing antibodies and strategies for their identification are therefore urgently required. Here we demonstrate that broadly neutralizing antibodies can be isolated from peripheral blood mononuclear cells of convalescent patients using SARS-CoV-2 receptor binding domains carrying epitope-specific mutations. This is exemplified by two human antibodies, GAR05, binding to epitope class 1, and GAR12, binding to a new epitope class 6 (located between class 3 and 5). Both antibodies broadly neutralize VOCs, exceeding the potency of the clinical monoclonal sotrovimab (S309) by orders of magnitude. They also provide prophylactic and therapeutic in vivo protection of female hACE2 mice against viral challenge. Our results indicate that exposure to SARS-CoV-2 induces antibodies that maintain broad neutralization against emerging VOCs using two unique strategies: either by targeting the divergent class 1 epitope in a manner resistant to VOCs (ACE2 mimicry, as illustrated by GAR05 and mAbs P2C-1F11/S2K14); or alternatively, by targeting rare and highly conserved epitopes, such as the new class 6 epitope identified here (as illustrated by GAR12). Our results provide guidance for next generation monoclonal antibody development and vaccine design.
History
DepositionDec 12, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 25, 2023Provider: repository / Type: Initial release
Revision 1.1Aug 9, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _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: Spike glycoprotein
B: Spike glycoprotein
C: Spike glycoprotein
H: GAR03 Fab heavy chain
L: GAR03 Fab light chain


Theoretical massNumber of molelcules
Total (without water)463,8855
Polymers463,8855
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area29780 Å2
ΔGint-136 kcal/mol
Surface area122160 Å2

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Components

#1: Protein Spike glycoprotein


Mass: 138736.516 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Severe acute respiratory syndrome-related coronavirus
Production host: Homo sapiens (human)
#2: Antibody GAR03 Fab heavy chain


Mass: 24705.656 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 GAR03 Fab light chain


Mass: 22970.273 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Homo sapiens (human)

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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: VFLIP Spike trimer with GAR03 FAB / Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
11Severe acute respiratory syndrome-related coronavirus694009
21Homo sapiens (human)9606
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.5
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: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1800 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K2 QUANTUM (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
CTF correctionType: NONE
3D reconstructionResolution: 3.39 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 130129 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00721748
ELECTRON MICROSCOPYf_angle_d1.37229609
ELECTRON MICROSCOPYf_dihedral_angle_d7.3112956
ELECTRON MICROSCOPYf_chiral_restr0.0773378
ELECTRON MICROSCOPYf_plane_restr0.013853

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