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- PDB-8tye: Lassa GPC (strain Josiah) bound to rabbit polyclonal fusion-pepti... -

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Basic information

Entry
Database: PDB / ID: 8tye
TitleLassa GPC (strain Josiah) bound to rabbit polyclonal fusion-peptide-targeting antibody FP-1
Components
  • (Glycoprotein ...) x 2
  • (Polyclonal Fv ...) x 2
KeywordsVIRAL PROTEIN/IMMUNE SYSTEM / Lassa virus glycoprotein complex / GPC / immune complex / antibody / polyclonal antibody / base antibody / VIRAL PROTEIN / VIRAL PROTEIN-IMMUNE SYSTEM complex
Function / homology
Function and homology information


host cell Golgi membrane / receptor-mediated endocytosis of virus by host cell / host cell endoplasmic reticulum membrane / lyase activity / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / host cell plasma membrane / virion membrane / membrane / metal ion binding
Similarity search - Function
KDPG/KHG aldolase / KDPG and KHG aldolase / Arenavirus glycoprotein, zinc binding domain / Arenavirus glycoprotein / Arenavirus glycoprotein / Aldolase-type TIM barrel
Similarity search - Domain/homology
Pre-glycoprotein polyprotein GP complex / 2-dehydro-3-deoxyphosphogluconate aldolase/4-hydroxy-2-oxoglutarate aldolase
Similarity search - Component
Biological speciesLassa virus
Thermotoga maritima (bacteria)
Oryctolagus cuniculus (rabbit)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsPerrett, H.R. / Brouwer, P.J.M. / Ward, A.B.
Funding support United States, Netherlands, 4items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)1F31Al172358 United States
Netherlands Organisation for Scientific Research (NWO)Rubicon Fellowship Netherlands
Bill & Melinda Gates FoundationINV-004923 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI71438 United States
CitationJournal: Cell Rep / Year: 2024
Title: Defining bottlenecks and opportunities for Lassa virus neutralization by structural profiling of vaccine-induced polyclonal antibody responses.
Authors: Philip J M Brouwer / Hailee R Perrett / Tim Beaumont / Haye Nijhuis / Sabine Kruijer / Judith A Burger / Ilja Bontjer / Wen-Hsin Lee / James A Ferguson / Martin Schauflinger / Helena Müller- ...Authors: Philip J M Brouwer / Hailee R Perrett / Tim Beaumont / Haye Nijhuis / Sabine Kruijer / Judith A Burger / Ilja Bontjer / Wen-Hsin Lee / James A Ferguson / Martin Schauflinger / Helena Müller-Kräuter / Rogier W Sanders / Thomas Strecker / Marit J van Gils / Andrew B Ward /
Abstract: Lassa fever continues to be a major public health burden in West Africa, yet effective therapies or vaccines are lacking. The isolation of protective neutralizing antibodies against the Lassa virus ...Lassa fever continues to be a major public health burden in West Africa, yet effective therapies or vaccines are lacking. The isolation of protective neutralizing antibodies against the Lassa virus glycoprotein complex (GPC) justifies the development of vaccines that can elicit strong neutralizing antibody responses. However, Lassa vaccine candidates have generally been unsuccessful at doing so, and the associated antibody responses to these vaccines remain poorly characterized. Here, we establish an electron microscopy-based epitope mapping workflow that enables high-resolution structural characterization of polyclonal antibodies to the GPC. By applying this method to rabbits vaccinated with a recombinant GPC vaccine and a GPC-derived virus-like particle, we reveal determinants of neutralization that involve epitopes of the GPC-A competition cluster. Furthermore, by identifying undescribed immunogenic off-target epitopes, we expose the challenges that recombinant GPC vaccines face. By enabling detailed polyclonal antibody characterization, our work ushers in a next generation of more rational Lassa vaccine design.
History
DepositionAug 25, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 11, 2024Provider: repository / Type: Initial release
Revision 1.1Sep 25, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
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 / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
H: Polyclonal Fv heavy chain
L: Polyclonal Fv light chain
C: Glycoprotein GP1
A: Glycoprotein GP1
B: Glycoprotein GP1
c: Glycoprotein GP2
a: Glycoprotein GP2
b: Glycoprotein GP2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)252,44641
Polymers239,9158
Non-polymers12,53133
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

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Glycoprotein ... , 2 types, 6 molecules CABcab

#3: Protein Glycoprotein GP1


Mass: 29010.352 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lassa virus (strain Mouse/Sierra Leone/Josiah/1976)
Strain: Mouse/Sierra Leone/Josiah/1976 / Gene: GPC, GP-C / Production host: Homo sapiens (human) / References: UniProt: P08669
#4: Protein Glycoprotein GP2 / GP2


Mass: 44765.047 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lassa virus (strain Mouse/Sierra Leone/Josiah/1976), (gene. exp.) Thermotoga maritima (strain ATCC 43589 / DSM 3109 / JCM 10099 / NBRC 100826 / MSB8) (bacteria)
Strain: Mouse/Sierra Leone/Josiah/1976, ATCC 43589 / DSM 3109 / JCM 10099 / NBRC 100826 / MSB8
Gene: GPC, GP-C, TM_0066 / Production host: Homo sapiens (human) / References: UniProt: P08669, UniProt: Q9WXS1

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Antibody , 2 types, 2 molecules HL

#1: Antibody Polyclonal Fv heavy chain


Mass: 9890.183 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit)
#2: Antibody Polyclonal Fv light chain


Mass: 8698.714 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit)

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Sugars , 6 types, 33 molecules

#5: Polysaccharide alpha-L-fucopyranose-(1-6)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 367.349 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
LFucpa1-6DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,2,1/[a2122h-1b_1-5_2*NCC/3=O][a1221m-1a_1-5]/1-2/a6-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(6+1)][a-L-Fucp]{}}LINUCSPDB-CARE
#6: Polysaccharide alpha-D-mannopyranose-(1-6)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1- ...alpha-D-mannopyranose-(1-6)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 748.682 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpa1-6DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/3,4,3/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3/a4-b1_b4-c1_c6-d1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(6+1)][a-D-Manp]{}}}}LINUCSPDB-CARE
#7: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta- ...2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 570.542 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4[LFucpa1-6]DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1221m-1a_1-5]/1-1-2/a4-b1_a6-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}[(6+1)][a-L-Fucp]{}}LINUCSPDB-CARE
#8: Polysaccharide
2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 11
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#9: Polysaccharide beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 586.542 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5]/1-1-2/a4-b1_b4-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{}}}LINUCSPDB-CARE
#10: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 14
Source method: isolated from a genetically manipulated source
Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Details

Has ligand of interestN
Sequence detailsThe authors state that the sequences of the antibody fragments are unknown.

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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

Component
IDNameTypeEntity IDParent-IDSource
1Heterotrimeric Lassa GPC bound to heterodimeric rabbit polyclonal fusion-peptide-targeting antibody FP-1COMPLEX#1-#40MULTIPLE SOURCES
2Glycoprotein G1, Glycoprotein G2, 2-dehydro-3-deoxyphosphogluconate aldolase/4-hydroxy-2-oxoglutarate aldolase fusion proteinCOMPLEX#3-#41RECOMBINANT
3Polyclonal antibody FP-1 heavy chain, Polyclonal antibody FP-1 light chainCOMPLEX#1-#21NATURAL
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Lassa virus Josiah11622
23Oryctolagus cuniculus (rabbit)9986
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.4
SpecimenConc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

MicroscopyModel: TFS GLACIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: OTHER
Image recordingElectron dose: 43.3 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

EM software
IDNameCategory
2Leginonimage acquisition
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
13cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 59236 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00610092
ELECTRON MICROSCOPYf_angle_d0.97613699
ELECTRON MICROSCOPYf_dihedral_angle_d16.2064405
ELECTRON MICROSCOPYf_chiral_restr0.0651746
ELECTRON MICROSCOPYf_plane_restr0.0251665

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