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- EMDB-2625: Negative stain reconstruction of 8ANC195 Fab in complex with BG50... -

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Basic information

Entry
Database: EMDB / ID: EMD-2625
TitleNegative stain reconstruction of 8ANC195 Fab in complex with BG505 SOSIP.664 trimer
Map dataReconstruction of 8ANC195 Fab bound to soluble Env trimer BG505.SOSIP
Sample
  • Sample: 8ANC195 Fab in complex with BG505 SOSIP.664 gp140 trimer
  • Protein or peptide: Soluble HIV-1 Envelope glycoprotein
  • Protein or peptide: Fab 8ANC195
KeywordsHIV-1 / broadly neutralizing antibodies / bnAb / 8ANC195 / SOSIP
Biological speciesHuman Immunodeficiency Virus-1 / Homo sapiens (human)
Methodsingle particle reconstruction / negative staining / Resolution: 18.7 Å
AuthorsLee JH / Scharf L / Bjorkman PJ / Ward AB
CitationJournal: Cell Rep / Year: 2014
Title: Antibody 8ANC195 reveals a site of broad vulnerability on the HIV-1 envelope spike.
Authors: Louise Scharf / Johannes F Scheid / Jeong Hyun Lee / Anthony P West / Courtney Chen / Han Gao / Priyanthi N P Gnanapragasam / René Mares / Michael S Seaman / Andrew B Ward / Michel C ...Authors: Louise Scharf / Johannes F Scheid / Jeong Hyun Lee / Anthony P West / Courtney Chen / Han Gao / Priyanthi N P Gnanapragasam / René Mares / Michael S Seaman / Andrew B Ward / Michel C Nussenzweig / Pamela J Bjorkman /
Abstract: Broadly neutralizing antibodies (bNAbs) to HIV-1 envelope glycoprotein (Env) can prevent infection in animal models. Characterized bNAb targets, although key to vaccine and therapeutic strategies, ...Broadly neutralizing antibodies (bNAbs) to HIV-1 envelope glycoprotein (Env) can prevent infection in animal models. Characterized bNAb targets, although key to vaccine and therapeutic strategies, are currently limited. We defined a new site of vulnerability by solving structures of bNAb 8ANC195 complexed with monomeric gp120 by X-ray crystallography and trimeric Env by electron microscopy. The site includes portions of gp41 and N-linked glycans adjacent to the CD4-binding site on gp120, making 8ANC195 the first donor-derived anti-HIV-1 bNAb with an epitope spanning both Env subunits. Rather than penetrating the glycan shield by using a single variable-region CDR loop, 8ANC195 inserted its entire heavy-chain variable domain into a gap to form a large interface with gp120 glycans and regions of the gp120 inner domain not contacted by other bNAbs. By isolating additional 8ANC195 clonal variants, we identified a more potent variant, which may be valuable for therapeutic approaches using bNAb combinations with nonoverlapping epitopes.
History
DepositionApr 5, 2014-
Header (metadata) releaseMay 14, 2014-
Map releaseMay 14, 2014-
UpdateJul 9, 2014-
Current statusJul 9, 2014Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0123
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0123
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_2625.png

Downloads & links

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Map

FileDownload / File: emd_2625.map.gz / Format: CCP4 / Size: 15.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of 8ANC195 Fab bound to soluble Env trimer BG505.SOSIP
Voxel sizeX=Y=Z: 2.05 Å
Density
Contour LevelBy AUTHOR: 0.0123 / Movie #1: 0.0123
Minimum - Maximum-0.01134914 - 0.05044185
Average (Standard dev.)0.00076809 (±0.00487608)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions160160160
Spacing160160160
CellA=B=C: 328.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.052.052.05
M x/y/z160160160
origin x/y/z0.0000.0000.000
length x/y/z328.000328.000328.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-184-184-183
NX/NY/NZ368368368
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS160160160
D min/max/mean-0.0110.0500.001

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Supplemental data

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Sample components

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Entire : 8ANC195 Fab in complex with BG505 SOSIP.664 gp140 trimer

EntireName: 8ANC195 Fab in complex with BG505 SOSIP.664 gp140 trimer
Components
  • Sample: 8ANC195 Fab in complex with BG505 SOSIP.664 gp140 trimer
  • Protein or peptide: Soluble HIV-1 Envelope glycoprotein
  • Protein or peptide: Fab 8ANC195

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Supramolecule #1000: 8ANC195 Fab in complex with BG505 SOSIP.664 gp140 trimer

SupramoleculeName: 8ANC195 Fab in complex with BG505 SOSIP.664 gp140 trimer
type: sample / ID: 1000 / Oligomeric state: One Fab binds per gp140 monomer / Number unique components: 2
Molecular weightTheoretical: 500 KDa

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Macromolecule #1: Soluble HIV-1 Envelope glycoprotein

MacromoleculeName: Soluble HIV-1 Envelope glycoprotein / type: protein_or_peptide / ID: 1 / Name.synonym: BG505 SOSIP.664 / Number of copies: 1 / Oligomeric state: Trimer / Recombinant expression: Yes
Source (natural)Organism: Human Immunodeficiency Virus-1 / Strain: BG505 / synonym: HIV-1
Molecular weightTheoretical: 350 KDa
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293T

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Macromolecule #2: Fab 8ANC195

MacromoleculeName: Fab 8ANC195 / type: protein_or_peptide / ID: 2 / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 50 KDa
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293T

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.03 mg/mL
BufferpH: 7.4 / Details: 150 mM NaCl, 50 mM Tris
StainingType: NEGATIVE
Details: Grids with adsorbed sample stained with 2% UF for 30 seconds
GridDetails: 400 mesh Cu with thin carbon support, glow discharged
VitrificationCryogen name: NONE / Instrument: OTHER

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Electron microscopy

MicroscopeFEI TECNAI 12
Electron beamAcceleration voltage: 120 kV / Electron source: LAB6
Electron opticsCalibrated magnification: 52000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 0.8 µm / Nominal defocus min: 0.7 µm / Nominal magnification: 52000
Sample stageSpecimen holder model: SIDE ENTRY, EUCENTRIC
TemperatureMax: 293 K
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism corrected at 52,000x mag
DetailsData collected from 0 to -40 degrees in 10 degree increments
DateMay 13, 2013
Image recordingCategory: CCD / Film or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Number real images: 195 / Average electron dose: 25 e/Å2 / Details: Data collected on CCD
Tilt angle min0

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Image processing

CTF correctionDetails: not corrected
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 18.7 Å / Resolution method: OTHER / Software - Name: EMAN2, Sparx / Number images used: 11637

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Atomic model buiding 1

Initial modelPDB ID:

4nco


Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B / Chain - #2 - Chain ID: E / Chain - #3 - Chain ID: F / Chain - #4 - Chain ID: I / Chain - #5 - Chain ID: J
SoftwareName: Chimera
DetailsThe models were fit using the Fit function in Chimera.
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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Atomic model buiding 2

Initial modelPDB ID:

4p9h


Chain - #0 - Chain ID: G / Chain - #1 - Chain ID: H / Chain - #2 - Chain ID: L
SoftwareName: Chimera
DetailsThis structure was fit by either superimposing the gp120 onto gp120 of the SOSIP trimer, or by using the Fit function in Chimera.
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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