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- PDB-6orn: Modified BG505 SOSIP-based immunogen RC1 in complex with the elic... -

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Entry
Database: PDB / ID: 6orn
TitleModified BG505 SOSIP-based immunogen RC1 in complex with the elicited V3-glycan patch bNAb 10-1074
Components
  • (10-1074 antibody Fab ...) x 2
  • (RC1 variant of HIV-1 Env glycoprotein ...) x 2
KeywordsVIRAL PROTEIN/Immune System / HIV-1 broadly-neutralizing antibody / Env trimer structure / V3-glycan patch / cryo-EM / RC1 / immunogen design / ANTIVIRAL PROTEIN-IMMUNE SYSTEM complex / ANTIVIRAL PROTEIN / VIRAL PROTEIN-Immune System complex
Biological speciesHuman immunodeficiency virus 1
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.05 Å
AuthorsAbernathy, M.E. / Gristick, H.B. / Bjorkman, P.J.
Funding supportUnited States , 2件
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious DiseasesP01 AI100148United States
National Institutes of Health/National Institute of General Medical SciencesP50 GM082545-06United States
CitationJournal: Nature / Year: 2019
Title: Immunization expands B cells specific to HIV-1 V3 glycan in mice and macaques.
Authors: Amelia Escolano / Harry B Gristick / Morgan E Abernathy / Julia Merkenschlager / Rajeev Gautam / Thiago Y Oliveira / Joy Pai / Anthony P West / Christopher O Barnes / Alexander A Cohen / Haoqing Wang / Jovana Golijanin / Daniel Yost / Jennifer R Keeffe / Zijun Wang / Peng Zhao / Kai-Hui Yao / Jens Bauer / Lilian Nogueira / Han Gao / Alisa V Voll / David C Montefiori / Michael S Seaman / Anna Gazumyan / Murillo Silva / Andrew T McGuire / Leonidas Stamatatos / Darrell J Irvine / Lance Wells / Malcolm A Martin / Pamela J Bjorkman / Michel C Nussenzweig /
Abstract: Broadly neutralizing monoclonal antibodies protect against infection with HIV-1 in animal models, suggesting that a vaccine that elicits these antibodies would be protective in humans. However, it ...Broadly neutralizing monoclonal antibodies protect against infection with HIV-1 in animal models, suggesting that a vaccine that elicits these antibodies would be protective in humans. However, it has not yet been possible to induce adequate serological responses by vaccination. Here, to activate B cells that express precursors of broadly neutralizing antibodies within polyclonal repertoires, we developed an immunogen, RC1, that facilitates the recognition of the variable loop 3 (V3)-glycan patch on the envelope protein of HIV-1. RC1 conceals non-conserved immunodominant regions by the addition of glycans and/or multimerization on virus-like particles. Immunization of mice, rabbits and rhesus macaques with RC1 elicited serological responses that targeted the V3-glycan patch. Antibody cloning and cryo-electron microscopy structures of antibody-envelope complexes confirmed that immunization with RC1 expands clones of B cells that carry the anti-V3-glycan patch antibodies, which resemble precursors of human broadly neutralizing antibodies. Thus, RC1 may be a suitable priming immunogen for sequential vaccination strategies in the context of polyclonal repertoires.
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Apr 30, 2019 / Release: Jun 12, 2019
RevisionDateData content typeProviderType
1.0Jun 12, 2019Structure modelrepositoryInitial release

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Structure visualization

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Structure viewerMolecule:
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Assembly

Deposited unit
C: RC1 variant of HIV-1 Env glycoprotein gp41
G: RC1 variant of HIV-1 Env glycoprotein gp120
J: RC1 variant of HIV-1 Env glycoprotein gp41
K: RC1 variant of HIV-1 Env glycoprotein gp120
Q: RC1 variant of HIV-1 Env glycoprotein gp41
R: RC1 variant of HIV-1 Env glycoprotein gp120
A: 10-1074 antibody Fab heavy chain
F: 10-1074 antibody Fab heavy chain
O: 10-1074 antibody Fab heavy chain
B: 10-1074 antibody Fab light chain
I: 10-1074 antibody Fab light chain
P: 10-1074 antibody Fab light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)381,531113
Polymers360,37912
Non-polymers21,152101
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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RC1 variant of HIV-1 Env glycoprotein ... , 2 types, 6 molecules CJQGKR

#1: Protein/peptide RC1 variant of HIV-1 Env glycoprotein gp41


Mass: 17134.324 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human immunodeficiency virus 1 / Cell (production host): 293 6E / Production host: Homo sapiens (human)
#2: Protein/peptide RC1 variant of HIV-1 Env glycoprotein gp120


Mass: 54149.566 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human immunodeficiency virus 1 / Cell (production host): 293 6E / Production host: Homo sapiens (human)

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10-1074 antibody Fab ... , 2 types, 6 molecules AFOBIP

#3: Protein/peptide 10-1074 antibody Fab heavy chain


Mass: 25661.688 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell (production host): Expi293 / Production host: Homo sapiens (human)
#4: Protein/peptide 10-1074 antibody Fab light chain


Mass: 23180.760 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell (production host): Expi293 / Production host: Homo sapiens (human)

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Non-polymers , 3 types, 101 molecules

#5: Chemical...
ChemComp-NAG / N-ACETYL-D-GLUCOSAMINE


Mass: 221.208 Da / Num. of mol.: 72
Source method: isolated from a genetically manipulated source
Formula: C8H15NO6 / N-Acetylglucosamine
#6: Chemical
ChemComp-BMA / BETA-D-MANNOSE


Mass: 180.156 Da / Num. of mol.: 9
Source method: isolated from a genetically manipulated source
Formula: C6H12O6 / Mannose
#7: Chemical
ChemComp-MAN / ALPHA-D-MANNOSE


Mass: 180.156 Da / Num. of mol.: 20
Source method: isolated from a genetically manipulated source
Formula: C6H12O6 / Mannose

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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

Type: COMPLEX

IDNameDetailsEntity IDParent-IDSource
1Complex of RC1 variant of BG505 SOSIP.664 trimer with three 10-1074 Fabs and three CD4bs FabsFab fragments generated by recombinant expression and complexed with the RC1 variant of BG505 SOSIP.664 trimer1,2,3,40MULTIPLE SOURCES
2RC1 variant of BG505 SOSIP.664 trimer1,21RECOMBINANT
310-1074 Fab3,41RECOMBINANT
Molecular weightValue: 0.6 MDa / Experimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
11Human immunodeficiency virus 111676
21Homo sapiens (human)9606
32Human immunodeficiency virus 111676
43Homo sapiens (human)9606
Source (recombinant)

Ncbi tax-ID: 9606 / Organism: Homo sapiens (human)

IDEntity assembly-IDCell
11293 6E
22Expi293
33Expi293
Buffer solutionpH: 8
Buffer component

Buffer-ID: 1

IDConc.NameFormula
120 mMTris-HClTris
2120 mMsodium chlorideNaClSodium chloride
SpecimenConc.: 0.75 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K
Details: 0 blot force, 3.5 second blot time, 3 uL sample added to freshly glow-discharged grids

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 73000 X / Nominal defocus max: 3400 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 684
Image scansMovie frames/image: 40 / Used frames/image: 1-40

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Processing

SoftwareName: PHENIX / Version: 1.14_3219: / Classification: refinement
EM software
IDNameVersionCategory
1RELION3particle selection
2EPUimage acquisition
4Gctf1.06CTF correction
7UCSF Chimeramodel fitting
9PHENIX1.14model refinement
10cryoSPARC2.1initial Euler assignment
11cryoSPARC2.1final Euler assignment
12cryoSPARC2.1classification
13cryoSPARC2.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C3 (3 fold cyclic)
3D reconstructionResolution: 4.05 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 122013 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL / Target criteria: Correlation coefficient
Atomic model building

3D fitting-ID: 1 / PDB-ID: 5T3Z

IDPdb chain-ID
1G
2B
3H
4L
Refine LS restraints

Refinement-ID: ELECTRON MICROSCOPY

TypeDev idealNumber
f_bond_d0.00720889
f_angle_d0.83328362
f_dihedral_angle_d10.29912300
f_chiral_restr0.1143426
f_plane_restr0.0053507

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