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- EMDB-6588: Negative stain EM of Ebola virus GP bound to human survivor antibodies -

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Basic information

Entry
Database: EMDB / ID: EMD-6588
TitleNegative stain EM of Ebola virus GP bound to human survivor antibodies
Map dataReconstruction of human survivor antibody Fabs bound to EBOV GPdTM
Sample
  • Sample: Reconstruction of Ebola virus GP ectodomain containing mucin-like domains in complex with human survivor antibody Fab ADI-15758
  • Protein or peptide: Ebola virus glycoprotein
  • Protein or peptide: Immunoglobulin 1 fragment
KeywordsEbola virus / antibodies / negative stain EM
Biological speciesEbola virus sp. / Homo sapiens (human)
Methodsingle particle reconstruction / negative staining / Resolution: 24.0 Å
AuthorsBorndoldt ZA / Turner HL / Murin CD / Li W / Sok D / Sounders C / Piper AE / Fusco ML / Pommert K / Klempner M ...Borndoldt ZA / Turner HL / Murin CD / Li W / Sok D / Sounders C / Piper AE / Fusco ML / Pommert K / Klempner M / Reimann KA / Golf A / Wittrup DK / Gerngross TU / Krauland E / Glass PJ / Burton DR / Saphire EO / Ward AB / Walker LM
CitationJournal: Science / Year: 2016
Title: Isolation of potent neutralizing antibodies from a survivor of the 2014 Ebola virus outbreak.
Authors: Zachary A Bornholdt / Hannah L Turner / Charles D Murin / Wen Li / Devin Sok / Colby A Souders / Ashley E Piper / Arthur Goff / Joshua D Shamblin / Suzanne E Wollen / Thomas R Sprague / ...Authors: Zachary A Bornholdt / Hannah L Turner / Charles D Murin / Wen Li / Devin Sok / Colby A Souders / Ashley E Piper / Arthur Goff / Joshua D Shamblin / Suzanne E Wollen / Thomas R Sprague / Marnie L Fusco / Kathleen B J Pommert / Lisa A Cavacini / Heidi L Smith / Mark Klempner / Keith A Reimann / Eric Krauland / Tillman U Gerngross / Karl D Wittrup / Erica Ollmann Saphire / Dennis R Burton / Pamela J Glass / Andrew B Ward / Laura M Walker /
Abstract: Antibodies targeting the Ebola virus surface glycoprotein (EBOV GP) are implicated in protection against lethal disease, but the characteristics of the human antibody response to EBOV GP remain ...Antibodies targeting the Ebola virus surface glycoprotein (EBOV GP) are implicated in protection against lethal disease, but the characteristics of the human antibody response to EBOV GP remain poorly understood. We isolated and characterized 349 GP-specific monoclonal antibodies (mAbs) from the peripheral B cells of a convalescent donor who survived the 2014 EBOV Zaire outbreak. Remarkably, 77% of the mAbs neutralize live EBOV, and several mAbs exhibit unprecedented potency. Structures of selected mAbs in complex with GP reveal a site of vulnerability located in the GP stalk region proximal to the viral membrane. Neutralizing antibodies targeting this site show potent therapeutic efficacy against lethal EBOV challenge in mice. The results provide a framework for the design of new EBOV vaccine candidates and immunotherapies.
History
DepositionFeb 3, 2016-
Header (metadata) releaseMar 9, 2016-
Map releaseApr 6, 2016-
UpdateApr 6, 2016-
Current statusApr 6, 2016Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3.26
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 3.26
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_6588.jpg

Downloads & links

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Map

FileDownload / File: emd_6588.map.gz / Format: CCP4 / Size: 1001 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of human survivor antibody Fabs bound to EBOV GPdTM
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
4.1 Å/pix.
x 64 pix.
= 262.4 Å		4.1 Å/pix.
x 64 pix.
= 262.4 Å		4.1 Å/pix.
x 64 pix.
= 262.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 4.1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 3.26 / Movie #1: 3.26
Minimum - Maximum-4.76629543 - 10.37817669
Average (Standard dev.)0.0 (±0.99999809)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions646464
Spacing646464
CellA=B=C: 262.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.14.14.1
M x/y/z646464
origin x/y/z0.0000.0000.000
length x/y/z262.400262.400262.400
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS646464
D min/max/mean-4.76610.378-0.000

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Supplemental data

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Sample components

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Entire : Reconstruction of Ebola virus GP ectodomain containing mucin-like...

EntireName: Reconstruction of Ebola virus GP ectodomain containing mucin-like domains in complex with human survivor antibody Fab ADI-15758
Components
  • Sample: Reconstruction of Ebola virus GP ectodomain containing mucin-like domains in complex with human survivor antibody Fab ADI-15758
  • Protein or peptide: Ebola virus glycoprotein
  • Protein or peptide: Immunoglobulin 1 fragment

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Supramolecule #1000: Reconstruction of Ebola virus GP ectodomain containing mucin-like...

SupramoleculeName: Reconstruction of Ebola virus GP ectodomain containing mucin-like domains in complex with human survivor antibody Fab ADI-15758
type: sample / ID: 1000 / Oligomeric state: One GP trimer bound to three Fab monomers / Number unique components: 2
Molecular weightTheoretical: 360 KDa

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Macromolecule #1: Ebola virus glycoprotein

MacromoleculeName: Ebola virus glycoprotein / type: protein_or_peptide / ID: 1 / Name.synonym: EBOV GPdTM / Number of copies: 1 / Oligomeric state: Trimer / Recombinant expression: Yes
Source (natural)Organism: Ebola virus sp. / synonym: Ebola virus
Molecular weightTheoretical: 210 KDa
Recombinant expressionOrganism: Drosophila melanogaster (fruit fly) / Recombinant cell: Schneider S2 / Recombinant plasmid: pMT-puro

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Macromolecule #2: Immunoglobulin 1 fragment

MacromoleculeName: Immunoglobulin 1 fragment / type: protein_or_peptide / ID: 2 / Name.synonym: IgG1 Fab / Number of copies: 3 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human) / synonym: Human
Molecular weightTheoretical: 50 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)

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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, 20 mM Tris
StainingType: NEGATIVE / Details: 2% uranyl formate
GridDetails: Cu mesh grid
VitrificationCryogen name: NONE / Instrument: OTHER

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Electron microscopy

MicroscopeFEI TECNAI SPIRIT
DateSep 10, 2015
Image recordingCategory: CCD / Film or detector model: TVIPS TEMCAM-F415 (4k x 4k) / Number real images: 98 / Average electron dose: 25 e/Å2
Electron beamAcceleration voltage: 120 kV / Electron source: LAB6
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 52000
Sample stageSpecimen holder model: SIDE ENTRY, EUCENTRIC
Experimental equipment
Model: Tecnai Spirit / Image courtesy: FEI Company

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Image processing

DetailsParticles were picked using DoG Picker.
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 24.0 Å / Resolution method: OTHER / Software - Name: EMAN2, XMipp, IMAGIC / Number images used: 17771

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