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- PDB-8f9m: HIV Env germline targeting BG505_MD64_N332-GT5 SOSIP in complex w... -

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Basic information

Entry
Database: PDB / ID: 8f9m
TitleHIV Env germline targeting BG505_MD64_N332-GT5 SOSIP in complex with V3-glycan polyclonal Fab isolated from immunized wild type mice, and NHP monoclonal Fab RM20A3
Components
  • (BG505_MD64_N332-GT5 ...) x 2
  • (V3-glycan epitope polyclonal Fab ...) x 2
  • RM20A3 Fab Heavy Chain
  • RM20A3 Fab Light Chain
KeywordsVIRAL PROTEIN/IMMUNE SYSTEM / mouse antibody / germline targeting / HIV-1 / vaccine design / polyclonal / VIRAL PROTEIN-IMMUNE SYSTEM complex
Biological speciessynthetic construct (others)
Macaca mulatta (Rhesus monkey)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsOzorowski, G. / Ward, A.B.
Funding support United States, 2items
OrganizationGrant numberCountry
Bill & Melinda Gates FoundationOPP1115782/INV-002916 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)A1144462 United States
CitationJournal: Science / Year: 2024
Title: mRNA-LNP HIV-1 trimer boosters elicit precursors to broad neutralizing antibodies.
Authors: Zhenfei Xie / Ying-Cing Lin / Jon M Steichen / Gabriel Ozorowski / Sven Kratochvil / Rashmi Ray / Jonathan L Torres / Alessia Liguori / Oleksandr Kalyuzhniy / Xuesong Wang / John E Warner / ...Authors: Zhenfei Xie / Ying-Cing Lin / Jon M Steichen / Gabriel Ozorowski / Sven Kratochvil / Rashmi Ray / Jonathan L Torres / Alessia Liguori / Oleksandr Kalyuzhniy / Xuesong Wang / John E Warner / Stephanie R Weldon / Gordon A Dale / Kathrin H Kirsch / Usha Nair / Sabyasachi Baboo / Erik Georgeson / Yumiko Adachi / Michael Kubitz / Abigail M Jackson / Sara T Richey / Reid M Volk / Jeong Hyun Lee / Jolene K Diedrich / Thavaleak Prum / Samantha Falcone / Sunny Himansu / Andrea Carfi / John R Yates / James C Paulson / Devin Sok / Andrew B Ward / William R Schief / Facundo D Batista /
Abstract: Germline-targeting (GT) HIV vaccine strategies are predicated on deriving broadly neutralizing antibodies (bnAbs) through multiple boost immunogens. However, as the recruitment of memory B cells ...Germline-targeting (GT) HIV vaccine strategies are predicated on deriving broadly neutralizing antibodies (bnAbs) through multiple boost immunogens. However, as the recruitment of memory B cells (MBCs) to germinal centers (GCs) is inefficient and may be derailed by serum antibody-induced epitope masking, driving further B cell receptor (BCR) modification in GC-experienced B cells after boosting poses a challenge. Using humanized immunoglobulin knockin mice, we found that GT protein trimer immunogen N332-GT5 could prime inferred-germline precursors to the V3-glycan-targeted bnAb BG18 and that B cells primed by N332-GT5 were effectively boosted by either of two novel protein immunogens designed to have minimum cross-reactivity with the off-target V1-binding responses. The delivery of the prime and boost immunogens as messenger RNA lipid nanoparticles (mRNA-LNPs) generated long-lasting GCs, somatic hypermutation, and affinity maturation and may be an effective tool in HIV vaccine development.
History
DepositionNov 23, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 15, 2024Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Mask / Data content type: Mask / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Mask / Data content type: Mask / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Mask / Data content type: Mask / Provider: repository / Type: Initial release
Revision 1.0May 15, 2024Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.1May 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.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.2Nov 20, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification
Revision 1.3May 21, 2025Group: Data collection / Category: em_admin / em_software / Item: _em_admin.last_update / _em_software.name
Revision 1.1May 21, 2025Data content type: EM metadata / Data content type: EM metadata / EM metadata / Group: Data processing / Experimental summary / Data content type: EM metadata / EM metadata / Category: em_admin / em_software / Data content type: EM metadata / EM metadata / Item: _em_admin.last_update / _em_software.name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: BG505_MD64_N332-GT5 gp120
C: RM20A3 Fab Heavy Chain
D: RM20A3 Fab Light Chain
B: BG505_MD64_N332-GT5 gp41
E: BG505_MD64_N332-GT5 gp120
J: RM20A3 Fab Heavy Chain
M: RM20A3 Fab Light Chain
G: BG505_MD64_N332-GT5 gp41
F: BG505_MD64_N332-GT5 gp120
K: RM20A3 Fab Heavy Chain
N: RM20A3 Fab Light Chain
I: BG505_MD64_N332-GT5 gp41
H: V3-glycan epitope polyclonal Fab heavy chain
L: V3-glycan epitope polyclonal Fab light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)331,29565
Polymers317,53614
Non-polymers13,75951
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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BG505 MD64 N332-GT5 ... , 2 types, 6 molecules AEFBGI

#1: Protein BG505_MD64_N332-GT5 gp120


Mass: 54236.801 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Homo sapiens (human)
#4: Protein BG505_MD64_N332-GT5 gp41


Mass: 18250.541 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Homo sapiens (human)

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Antibody , 4 types, 8 molecules CJKDMNHL

#2: Antibody RM20A3 Fab Heavy Chain


Mass: 13511.111 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Macaca mulatta (Rhesus monkey) / Production host: Homo sapiens (human)
#3: Antibody RM20A3 Fab Light Chain


Mass: 13508.800 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Macaca mulatta (Rhesus monkey) / Production host: Homo sapiens (human)
#5: Antibody V3-glycan epitope polyclonal Fab heavy chain


Mass: 9634.868 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#6: Antibody V3-glycan epitope polyclonal Fab light chain


Mass: 9379.553 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)

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Sugars , 3 types, 51 molecules

#7: 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.: 7
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
#8: Polysaccharide alpha-D-mannopyranose-(1-3)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1- ...alpha-D-mannopyranose-(1-3)-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
DManpa1-3DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-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_c3-d1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][a-D-Manp]{}}}}LINUCSPDB-CARE
#9: 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.: 42 / Source method: obtained synthetically / 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
Has protein modificationY

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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: HIV Env germline targeting BG505_MD64_N332-GT5 SOSIP in complex with V3-glycan polyclonal Fab isolated from immunized wild type mice, and NHP monoclonal Fab RM20A3
Type: COMPLEX / Entity ID: #1-#6 / Source: MULTIPLE SOURCES
Molecular weightExperimental value: NO
Source (natural)Organism: Mus musculus (house mouse)
Buffer solutionpH: 7.4
SpecimenConc.: 7 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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 magnification: 190000 X / Nominal defocus max: 1500 nm / Nominal defocus min: 700 nm / Alignment procedure: COMA FREE
Specimen holderSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 40 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.20.1_4487: / Classification: refinement
EM software
IDNameVersionCategoryDetails
2EPUimage acquisition
4cryoSPARC3.2CTF correctionPatch CTF
8PHENIXmodel refinement
12RELION3.1classification
13cryoSPARC3.23D reconstructionLocal Refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 11058 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00520707
ELECTRON MICROSCOPYf_angle_d0.86128122
ELECTRON MICROSCOPYf_dihedral_angle_d10.3237392
ELECTRON MICROSCOPYf_chiral_restr0.0533355
ELECTRON MICROSCOPYf_plane_restr0.0053520

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