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- EMDB-20817: Cryo-EM structure of HIV-1 neutralizing antibody DH270 UCA3 in co... -

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Basic information

Entry
Database: EMDB / ID: EMD-20817
TitleCryo-EM structure of HIV-1 neutralizing antibody DH270 UCA3 in complex with CH848 10.17DT Env
Map data
Sample
  • Complex: DH270 UCA3 Fab bound to HIV-1 Env CH848 10.17DT
    • Protein or peptide: Envelope glycoprotein gp160
    • Protein or peptide: DH270 UCA3 Fab Heavy Chain
    • Protein or peptide: DH270 UCA3 Fab Light Chain
  • Protein or peptide: CH848 10.17 DT gp120
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
Function / homology
Function and homology information


positive regulation of plasma membrane raft polarization / positive regulation of receptor clustering / positive regulation of establishment of T cell polarity / virus-mediated perturbation of host defense response / host cell endosome membrane / clathrin-dependent endocytosis of virus by host cell / viral protein processing / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope ...positive regulation of plasma membrane raft polarization / positive regulation of receptor clustering / positive regulation of establishment of T cell polarity / virus-mediated perturbation of host defense response / host cell endosome membrane / clathrin-dependent endocytosis of virus by host cell / viral protein processing / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / host cell plasma membrane / virion membrane / structural molecule activity / plasma membrane
Similarity search - Function
Envelope glycoprotein Gp160 / Retroviral envelope protein / Retroviral envelope protein GP41-like / Gp120 core superfamily / Envelope glycoprotein GP120 / Human immunodeficiency virus 1, envelope glycoprotein Gp120
Similarity search - Domain/homology
Envelope glycoprotein gp160 / Envelope glycoprotein gp160
Similarity search - Component
Biological speciesHuman immunodeficiency virus 1 / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsAcharya P / Henderson RC / Saunders KO / Haynes BF
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Human Genome Research Institute (NIH/NHGRI)UM1-AI100645 United States
National Institutes of Health/National Human Genome Research Institute (NIH/NHGRI)UM1-AI144371 United States
CitationJournal: Science / Year: 2019
Title: Targeted selection of HIV-specific antibody mutations by engineering B cell maturation.
Authors: Kevin O Saunders / Kevin Wiehe / Ming Tian / Priyamvada Acharya / Todd Bradley / S Munir Alam / Eden P Go / Richard Scearce / Laura Sutherland / Rory Henderson / Allen L Hsu / Mario J ...Authors: Kevin O Saunders / Kevin Wiehe / Ming Tian / Priyamvada Acharya / Todd Bradley / S Munir Alam / Eden P Go / Richard Scearce / Laura Sutherland / Rory Henderson / Allen L Hsu / Mario J Borgnia / Haiyan Chen / Xiaozhi Lu / Nelson R Wu / Brian Watts / Chuancang Jiang / David Easterhoff / Hwei-Ling Cheng / Kelly McGovern / Peyton Waddicor / Aimee Chapdelaine-Williams / Amanda Eaton / Jinsong Zhang / Wes Rountree / Laurent Verkoczy / Mark Tomai / Mark G Lewis / Heather R Desaire / Robert J Edwards / Derek W Cain / Mattia Bonsignori / David Montefiori / Frederick W Alt / Barton F Haynes /
Abstract: INTRODUCTION: A major goal of HIV-1 vaccine development is the design of immunogens that induce broadly neutralizing antibodies (bnAbs). However, vaccination of humans has not resulted in the ...INTRODUCTION: A major goal of HIV-1 vaccine development is the design of immunogens that induce broadly neutralizing antibodies (bnAbs). However, vaccination of humans has not resulted in the induction of affinity-matured and potent HIV-1 bnAbs. To devise effective vaccine strategies, we previously determined the maturation pathway of select HIV-1 bnAbs from acute infection through neutralizing antibody development. During their evolution, bnAbs acquire an abundance of improbable amino acid substitutions as a result of nucleotide mutations at variable region sequences rarely targeted by activation-induced cytidine deaminase, the enzyme responsible for antibody mutation. A subset of improbable mutations is essential for broad neutralization activity, and their acquisition represents a key roadblock to bnAb development.
RATIONALE: Current bnAb lineage-based vaccine strategies can initiate bnAb lineage development in animal models but have not specifically elicited the improbable mutations required for neutralization ...RATIONALE: Current bnAb lineage-based vaccine strategies can initiate bnAb lineage development in animal models but have not specifically elicited the improbable mutations required for neutralization breadth. Induction of bnAbs requires vaccine strategies that specifically engage bnAb precursors and subsequently select for improbable mutations required for broadly neutralizing activity. We hypothesized that vaccination with immunogens that bind with moderate to high affinity to bnAb B cell precursors, and with higher affinity to precursors that have acquired improbable mutations, could initiate bnAb B cell lineages and select for key improbable mutations required for bnAb development.
RESULTS: We elicited serum neutralizing HIV-1 antibodies in human bnAb precursor knock-in mice and wild-type macaques vaccinated with immunogens designed to select for improbable mutations. We ...RESULTS: We elicited serum neutralizing HIV-1 antibodies in human bnAb precursor knock-in mice and wild-type macaques vaccinated with immunogens designed to select for improbable mutations. We designed two HIV-1 envelope immunogens that bound precursor B cells of either a CD4 binding site or V3-glycan bnAb lineage. In vitro, these immunogens bound more strongly to bnAb precursors once the precursor acquired the desired improbable mutations. Vaccination of macaques with the CD4 binding site–targeting immunogen induced CD4 binding site serum neutralizing antibodies. Antibody sequences elicited in human bnAb precursor knock-in mice encoded functional improbable mutations critical for bnAb development. In bnAb precursor knock-in mice, we isolated a vaccine-elicited monoclonal antibody bearing functional improbable mutations that was capable of neutralizing multiple HIV-1 global isolates. Structures of a bnAb precursor, a bnAb, and the vaccine-elicited antibody revealed the precise roles that acquired improbable mutations played in recognizing the HIV-1 envelope. Thus, our immunogens elicited antibody responses in macaques and knock-in mice that exhibited the mutational patterns, structural characteristics, or neutralization profiles of nascent broadly neutralizing antibodies.
CONCLUSION: Our study represents a proof of concept for targeted selection of improbable mutations to guide antibody affinity maturation. Moreover, this study demonstrates a rational strategy for ...CONCLUSION: Our study represents a proof of concept for targeted selection of improbable mutations to guide antibody affinity maturation. Moreover, this study demonstrates a rational strategy for sequential immunogen design to circumvent the difficult roadblocks in HIV-1 bnAb induction by vaccination. We show that immunogens should exhibit differences in affinity across antibody maturation stages where improbable mutations are necessary for the desired antibody function. This strategy of selection of specific antibody nucleotides by immunogen design can be applied to B cell lineages targeting other pathogens where guided affinity maturation is needed for a protective antibody response.
History
DepositionOct 9, 2019-
Header (metadata) releaseDec 18, 2019-
Map releaseDec 18, 2019-
UpdateJul 29, 2020-
Current statusJul 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.7
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.7
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6um5
  • Surface level: 0.7
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20817.png

Downloads & links

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Map

FileDownload / File: emd_20817.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.08 Å
Density
Contour LevelBy AUTHOR: 0.7 / Movie #1: 0.7
Minimum - Maximum-0.94486827 - 2.0845234
Average (Standard dev.)0.004383294 (±0.09123344)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 345.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.081.081.08
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z345.600345.600345.600
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.9452.0850.004

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Supplemental data

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Sample components

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Entire : DH270 UCA3 Fab bound to HIV-1 Env CH848 10.17DT

EntireName: DH270 UCA3 Fab bound to HIV-1 Env CH848 10.17DT
Components
  • Complex: DH270 UCA3 Fab bound to HIV-1 Env CH848 10.17DT
    • Protein or peptide: Envelope glycoprotein gp160
    • Protein or peptide: DH270 UCA3 Fab Heavy Chain
    • Protein or peptide: DH270 UCA3 Fab Light Chain
  • Protein or peptide: CH848 10.17 DT gp120
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: DH270 UCA3 Fab bound to HIV-1 Env CH848 10.17DT

SupramoleculeName: DH270 UCA3 Fab bound to HIV-1 Env CH848 10.17DT / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #2-#4
Source (natural)Organism: Human immunodeficiency virus 1
Molecular weightExperimental: 360 KDa

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Macromolecule #1: CH848 10.17 DT gp120

MacromoleculeName: CH848 10.17 DT gp120 / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 1
Molecular weightTheoretical: 51.693512 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: ENLWVTVYYG VPVWKEAKTT LFCASDARAY EKEVHNVWAT HACVPTDPSP QELVLGNVTE NFNMWKNDMV DQMHEDIISL WDQSLKPCV KLTPLCVTLI CSDATVKTGT VEEMKNCSFN TTTEIRDKEK KEYALFYKPD IVPLSETNNT SEYRLINCNT S CVTQACPK ...String:
ENLWVTVYYG VPVWKEAKTT LFCASDARAY EKEVHNVWAT HACVPTDPSP QELVLGNVTE NFNMWKNDMV DQMHEDIISL WDQSLKPCV KLTPLCVTLI CSDATVKTGT VEEMKNCSFN TTTEIRDKEK KEYALFYKPD IVPLSETNNT SEYRLINCNT S CVTQACPK VTFEPIPIHY CAPAGYAILK CNDETFNGTG PCSNVSTVQC THGIRPVVST QLLLNGSLAE KEIVIRSENL TN NAKIIIV HLHTPVEIVC TRPNNNTRKS VRIGPGQTFY ATGDIIGDIK QAHCNISEEK WNDTLQKVGI ELQKHFPNKT IKY NQSAGG DMEITTHSFN CGGEFFYCNT SNLFNGTYNG TYISTNSSAN STSTITLQCR IKQIINMWQG VGRCMYAPPI AGNI TCRSN ITGLLLTRDG GTNSNETETF RPAGGDMRDN WRSELYKYKV VKIEPLGVAP TRCKRRV

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Macromolecule #2: Envelope glycoprotein gp160

MacromoleculeName: Envelope glycoprotein gp160 / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 1
Molecular weightTheoretical: 17.162525 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
AVGIGAVFLG FLGAAGSTMG AASMTLTVQA RNLLSGIVQQ QSNLLRAIEA QQHLLKLTVW GIKQLQARVL AVERYLRDQQ LLGIWGCSG KLICCTNVPW NSSWSNRNLS EIWDNMTWLQ WDKEISNYTQ IIYGLLEESQ NQQEKNEQDL LALD

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Macromolecule #3: DH270 UCA3 Fab Heavy Chain

MacromoleculeName: DH270 UCA3 Fab Heavy Chain / type: protein_or_peptide / ID: 3 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 25.779795 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: QVQLVQSGAE VKKPGASVKV SCKASGYTFT GYYMHWVRQA PGQGLEWMGW INPNSGGTNY AQKFQGRVTM TRDTSISTAY MELSRLRSD DTAVYYCARG GWISLYYDSS GYPNFDYWGQ GTLVTVSGAS TKGPSVFPLA PSSKSTSGGT AALGCLVKDY F PEPVTVSW ...String:
QVQLVQSGAE VKKPGASVKV SCKASGYTFT GYYMHWVRQA PGQGLEWMGW INPNSGGTNY AQKFQGRVTM TRDTSISTAY MELSRLRSD DTAVYYCARG GWISLYYDSS GYPNFDYWGQ GTLVTVSGAS TKGPSVFPLA PSSKSTSGGT AALGCLVKDY F PEPVTVSW NSGALTSGVH TFPAVLQSSG LYSLSSVVTV PSSSLGTQTY ICNVNHKPSN TKVDKRVEPK SCDKHHHHHH

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Macromolecule #4: DH270 UCA3 Fab Light Chain

MacromoleculeName: DH270 UCA3 Fab Light Chain / type: protein_or_peptide / ID: 4 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 22.648039 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: QSALTQPASV SGSPGQSITI SCTGTSSDVG SYNLVSWYQQ HPGKAPKLMI YEVSKRPSGV SNRFSGSKSG NTASLTISGL QAEDEADYY CCSYAGSSTV IFGGGTKLTV LGQPKGAPSV TLFPPSSEEL QANKATLVCL ISDFYPGAVT VAWKADSSPV K AGVETTTP ...String:
QSALTQPASV SGSPGQSITI SCTGTSSDVG SYNLVSWYQQ HPGKAPKLMI YEVSKRPSGV SNRFSGSKSG NTASLTISGL QAEDEADYY CCSYAGSSTV IFGGGTKLTV LGQPKGAPSV TLFPPSSEEL QANKATLVCL ISDFYPGAVT VAWKADSSPV K AGVETTTP SKQSNNKYAA SSYLSLTPEQ WKSHRSYSCQ VTHEGSTVEK TVAPTECS

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Macromolecule #9: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 9 / Number of copies: 27 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.2
GridModel: C-flat-1.2/1.3 4C / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 298 K / Instrument: LEICA PLUNGER

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Average electron dose: 42.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 22093

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