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- EMDB-23219: BG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOS... -

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Basic information

Entry
Database: EMDB / ID: EMD-23219
TitleBG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B)
Map dataBG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B) - Main map
Sample
  • Complex: BG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B)
    • Protein or peptide: BG505 SOSIP.v5.2(7S) - gp120
    • Protein or peptide: BG505 SOSIP.v5.2(7S) - gp41
  • 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 / identical protein binding / 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
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsAntanasijevic A / Froes Rocha R / Sewall LM / Ward AB
Funding support United States, 1 items
OrganizationGrant numberCountry
International AIDS Vaccine InitiativeOPP1196345 United States
CitationJournal: Nat Commun / Year: 2021
Title: Polyclonal antibody responses to HIV Env immunogens resolved using cryoEM.
Authors: Aleksandar Antanasijevic / Leigh M Sewall / Christopher A Cottrell / Diane G Carnathan / Luis E Jimenez / Julia T Ngo / Jennifer B Silverman / Bettina Groschel / Erik Georgeson / Jinal ...Authors: Aleksandar Antanasijevic / Leigh M Sewall / Christopher A Cottrell / Diane G Carnathan / Luis E Jimenez / Julia T Ngo / Jennifer B Silverman / Bettina Groschel / Erik Georgeson / Jinal Bhiman / Raiza Bastidas / Celia LaBranche / Joel D Allen / Jeffrey Copps / Hailee R Perrett / Kimmo Rantalainen / Fabien Cannac / Yuhe R Yang / Alba Torrents de la Peña / Rebeca Froes Rocha / Zachary T Berndsen / David Baker / Neil P King / Rogier W Sanders / John P Moore / Shane Crotty / Max Crispin / David C Montefiori / Dennis R Burton / William R Schief / Guido Silvestri / Andrew B Ward /
Abstract: Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate ...Engineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal antibody responses without the need for monoclonal antibody isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic.
History
DepositionDec 30, 2020-
Header (metadata) releaseAug 4, 2021-
Map releaseAug 4, 2021-
UpdateAug 25, 2021-
Current statusAug 25, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-7l7u
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23219.png

Downloads & links

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Map

FileDownload / File: emd_23219.map.gz / Format: CCP4 / Size: 22.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationBG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B) - Main map
Voxel sizeX=Y=Z: 1.15 Å
Density
Contour LevelBy AUTHOR: 0.03 / Movie #1: 0.03
Minimum - Maximum-0.1494741 - 0.23227064
Average (Standard dev.)0.0022507317 (±0.010273769)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions180180180
Spacing180180180
CellA=B=C: 207.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.151.151.15
M x/y/z180180180
origin x/y/z0.0000.0000.000
length x/y/z207.000207.000207.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS180180180
D min/max/mean-0.1490.2320.002

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Supplemental data

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Mask #1

Fileemd_23219_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: BG505 SOSIP reconstructed from a designed nanoparticle, BG505...

Fileemd_23219_half_map_1.map
AnnotationBG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B) - Half-map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: BG505 SOSIP reconstructed from a designed nanoparticle, BG505...

Fileemd_23219_half_map_2.map
AnnotationBG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B) - Half-map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : BG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOS...

EntireName: BG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B)
Components
  • Complex: BG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B)
    • Protein or peptide: BG505 SOSIP.v5.2(7S) - gp120
    • Protein or peptide: BG505 SOSIP.v5.2(7S) - gp41
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: BG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOS...

SupramoleculeName: BG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Details: The map was generated by subparticle extraction from the BG505 SOSIP-T33-31 nanoparticle using the localized reconstruction approach.
Source (natural)Organism: Human immunodeficiency virus 1
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293 / Recombinant plasmid: pPPI4

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Macromolecule #1: BG505 SOSIP.v5.2(7S) - gp120

MacromoleculeName: BG505 SOSIP.v5.2(7S) - gp120 / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 1
Molecular weightTheoretical: 56.70748 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MKRGLCCVLL LCGAVFVSPS QEIHARFRRG ARAENLWVTV YYGVPVWKDA ETTLFCASDA KAYETKKHNV WATHCCVPTD PNPQEIHLE NVTEEFNMWK NNMVEQMHTD IISLWDQSLK PCVKLTPLCV TLQCTNVTNN ITDDMRGELK NCSFNMTTEL R DKKQKVYS ...String:
MKRGLCCVLL LCGAVFVSPS QEIHARFRRG ARAENLWVTV YYGVPVWKDA ETTLFCASDA KAYETKKHNV WATHCCVPTD PNPQEIHLE NVTEEFNMWK NNMVEQMHTD IISLWDQSLK PCVKLTPLCV TLQCTNVTNN ITDDMRGELK NCSFNMTTEL R DKKQKVYS LFYRLDVVQI NENQGNRSNN SNKEYRLINC NTSAITQACP KVSFEPIPIH YCAPAGFAIL KCKDKKFNGT GP CTNVSTV QCTHGIKPVV STQLLLNGSL AEEEVIIRSE NITNNAKNIL VQLNESVQIN CTRPNNNTVK SIRIGPGQWF YYT GDIIGD IRQAHCNVSK ATWNETLGKV VKQLRKHFGN NTIIRFANSS GGDLEVTTHS FNCGGEFFYC NTSGLFNSTW ISNT SVQGS NSTGSNDSIT LPCRIKQIIN MWQRIGQAMY APPIQGVIRC VSNITGLILT RDGGSTNSTT ETFRPGGGDM RDNWR SELY KYKVVKIEPL GVAPTRCKRR V

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Macromolecule #2: BG505 SOSIP.v5.2(7S) - gp41

MacromoleculeName: BG505 SOSIP.v5.2(7S) - gp41 / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Human immunodeficiency virus 1
Molecular weightTheoretical: 16.543627 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
LGFLGAAGST MGAASMTLTV QARNLLSGIV QQQSNLLRAP ECQQHLLKDT HWGIKQLQAR VLAVEHYLRD QQLLGIWGCS GKLICCTNV PWNSSWSNRN LSEIWDNMTW LQWDKEISNY TQIIYGLLEE SQNQQEKNEQ DLLELD

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Macromolecule #5: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 5 / Number of copies: 39 / 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

Concentration4.1 mg/mL
BufferpH: 7.4
Component:
ConcentrationNameFormula
20.0 mMTris-HClTris
150.0 mMSodium chlorideNaClSodium chloride

Details: TBS buffer prepared from a 10X stock
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV / Details: Blotting time varied between 3 and 7 seconds..
DetailsBG505 SOSIP-T33-31 nanoparticle was prepared by combining equimolar amounts of BG505 SOSIP-T33-31A and BG505 SOSIP-T33-31B components that were expressed separately.

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 36000
Sample stageSpecimen holder model: OTHER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-41 / Number grids imaged: 2 / Number real images: 1751 / Average exposure time: 10.25 sec. / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 441476
Details: Subparticles were generated by extraction from 110,369 BG505 SOSIP-T33-31 nanoparticles (4 subparticles per 1 nanoparticle).
CTF correctionSoftware - Name: RELION (ver. 3.0)
Startup modelType of model: OTHER
Details: Map obtained from Ab initio reconstruction in cryoSPARC
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationSoftware - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 64726
DetailsFrames were aligned using MotionCorr and GCTF was applied for estimation of CTF parameters.
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6vl5

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-7l7u:
BG505 SOSIP reconstructed from a designed nanoparticle, BG505 SOSIP-T33-31 (Component B)

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