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- EMDB-5215: Anthrax toxin PA63 in complex with 1G3 -

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Basic information

Entry
Database: EMDB / ID: EMD-5215
TitleAnthrax toxin PA63 in complex with 1G3
Map dataAnthrax toxin component PA63 in complex with neutralizing antibody 1G3
Sample
  • Sample: Anthrax toxin component PA63 in complex with 1G3
  • Protein or peptide: toxin
Keywordsanthrax toxin protective antigen neutralizing antibody
Biological speciesBacillus anthracis (anthrax bacterium)
Methodsingle particle reconstruction / negative staining / Resolution: 33.0 Å
AuthorsRadjainia M / Hyun J / Leysath CE / Leppla SH / Mitra AK
CitationJournal: Proc Natl Acad Sci U S A / Year: 2010
Title: Anthrax toxin-neutralizing antibody reconfigures the protective antigen heptamer into a supercomplex.
Authors: Mazdak Radjainia / Jae-Kyung Hyun / Clinton E Leysath / Stephen H Leppla / Alok K Mitra /
Abstract: The tripartite protein exotoxin secreted by Bacillus anthracis, a major contributor to its virulence and anthrax pathogenesis, consists of binary complexes of the protective antigen (PA) heptamer ...The tripartite protein exotoxin secreted by Bacillus anthracis, a major contributor to its virulence and anthrax pathogenesis, consists of binary complexes of the protective antigen (PA) heptamer (PA63h), produced by proteolytic cleavage of PA, together with either lethal factor or edema factor. The mouse monoclonal anti-PA antibody 1G3 was previously shown to be a potent antidote that shares F(C) domain dependency with the human monoclonal antibody MDX-1303 currently under clinical development. Here we demonstrate that 1G3 instigates severe perturbation of the PA63h structure and creates a PA supercomplex as visualized by electron microscopy. This phenotype, produced by the unconventional mode of antibody action, highlights the feasibility for optimization of vaccines based on analogous structural modification of PA63h as an additional strategy for future remedies against anthrax.
History
DepositionJul 21, 2010-
Header (metadata) releaseJul 28, 2010-
Map releaseAug 12, 2010-
UpdateNov 2, 2010-
Current statusNov 2, 2010Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0416
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.0416
  • Imaged by UCSF Chimera
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Structure viewerEM map:
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Supplemental images

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Map

FileDownload / File: emd_5215.map.gz / Format: CCP4 / Size: 1.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationAnthrax toxin component PA63 in complex with neutralizing antibody 1G3
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
5 Å/pix.
x 80 pix.
= 400. Å
5 Å/pix.
x 80 pix.
= 400. Å
5 Å/pix.
x 80 pix.
= 400. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 5 Å
Density
Contour LevelBy AUTHOR: 0.0416 / Movie #1: 0.0416
Minimum - Maximum-0.207713 - 0.187278
Average (Standard dev.)-0.0000258488 (±0.014702)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions808080
Spacing808080
CellA=B=C: 400 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z555
M x/y/z808080
origin x/y/z0.0000.0000.000
length x/y/z400.000400.000400.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-99-99-99
NX/NY/NZ200200200
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS808080
D min/max/mean-0.2080.187-0.000

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Supplemental data

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Sample components

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Entire : Anthrax toxin component PA63 in complex with 1G3

EntireName: Anthrax toxin component PA63 in complex with 1G3
Components
  • Sample: Anthrax toxin component PA63 in complex with 1G3
  • Protein or peptide: toxin

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Supramolecule #1000: Anthrax toxin component PA63 in complex with 1G3

SupramoleculeName: Anthrax toxin component PA63 in complex with 1G3 / type: sample / ID: 1000 / Number unique components: 2
Molecular weightExperimental: 1.1 MDa / Method: Size-exclusion chromatography

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Macromolecule #1: toxin

MacromoleculeName: toxin / type: protein_or_peptide / ID: 1 / Name.synonym: protective antigen / Recombinant expression: Yes
Source (natural)Organism: Bacillus anthracis (anthrax bacterium) / synonym: bacillus anthracis
Recombinant expressionOrganism: Bacillus subtilis (bacteria)

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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

Concentration0.5 mg/mL
BufferpH: 9 / Details: 150mM NaCl, 50mM CHES
StainingType: NEGATIVE
Details: Holey grids with samples negatively stained across holes with 1% uranyl acetate
GridDetails: Quantifoil R1.2/1.3
VitrificationCryogen name: NONE / Instrument: OTHER

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Electron microscopy

MicroscopeFEI TECNAI 12
Alignment procedureLegacy - Astigmatism: objective lens astigmatism was corrected at 97000 times magnification
DateSep 1, 2008
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: NIKON SUPER COOLSCAN 9000 / Digitization - Sampling interval: 13 µm / Number real images: 40 / Average electron dose: 10 e/Å2 / Bits/pixel: 8
Tilt angle min0
Electron beamAcceleration voltage: 120 kV / Electron source: LAB6
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 52000
Sample stageSpecimen holder: single-tilt / Specimen holder model: OTHER / Tilt angle max: 45

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Image processing

DetailsThe particles were manually selected.
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 33.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: XMIPP / Number images used: 3459
Final two d classificationNumber classes: 20

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