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- EMDB-2506: Cryo-electron tomography average of an C1-IgG complex -

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Basic information

Entry
Database: EMDB / ID: EMD-2506
TitleCryo-electron tomography average of an C1-IgG complex
Map dataC1-IgG
Sample
  • Sample: DNP-haptenated liposomes, polyclonal goat anti-DNP Antibody, human C1 (originating from C4 depleted serum)
  • Protein or peptide: polyclonal IgG
  • Protein or peptide: C1
KeywordsC1-IgG complex / C1-immune complex / C1 / C1qr2S2 / IgG / classical complement activation
Biological speciesCapra hircus (goat) / Homo sapiens (human)
Methodsubtomogram averaging / cryo EM / Resolution: 66.0 Å
AuthorsDiebolder CA / Beurskens FJ / de Jong RN / Koning RI / Strumane K / Lindorfer MA / Voorhorst M / Ugurlar D / Rosati S / Heck AJR ...Diebolder CA / Beurskens FJ / de Jong RN / Koning RI / Strumane K / Lindorfer MA / Voorhorst M / Ugurlar D / Rosati S / Heck AJR / van de Winkel JGJ / Wilson IA / Koster AJ / Taylor RP / Ollmann-Saphire E / Burton DR / Schuurman J / Gros P / Parren PWHI
CitationJournal: Science / Year: 2014
Title: Complement is activated by IgG hexamers assembled at the cell surface.
Authors: Christoph A Diebolder / Frank J Beurskens / Rob N de Jong / Roman I Koning / Kristin Strumane / Margaret A Lindorfer / Marleen Voorhorst / Deniz Ugurlar / Sara Rosati / Albert J R Heck / Jan ...Authors: Christoph A Diebolder / Frank J Beurskens / Rob N de Jong / Roman I Koning / Kristin Strumane / Margaret A Lindorfer / Marleen Voorhorst / Deniz Ugurlar / Sara Rosati / Albert J R Heck / Jan G J van de Winkel / Ian A Wilson / Abraham J Koster / Ronald P Taylor / Erica Ollmann Saphire / Dennis R Burton / Janine Schuurman / Piet Gros / Paul W H I Parren /
Abstract: Complement activation by antibodies bound to pathogens, tumors, and self antigens is a critical feature of natural immune defense, a number of disease processes, and immunotherapies. How antibodies ...Complement activation by antibodies bound to pathogens, tumors, and self antigens is a critical feature of natural immune defense, a number of disease processes, and immunotherapies. How antibodies activate the complement cascade, however, is poorly understood. We found that specific noncovalent interactions between Fc segments of immunoglobulin G (IgG) antibodies resulted in the formation of ordered antibody hexamers after antigen binding on cells. These hexamers recruited and activated C1, the first component of complement, thereby triggering the complement cascade. The interactions between neighboring Fc segments could be manipulated to block, reconstitute, and enhance complement activation and killing of target cells, using all four human IgG subclasses. We offer a general model for understanding antibody-mediated complement activation and the design of antibody therapeutics with enhanced efficacy.
History
DepositionNov 4, 2013-
Header (metadata) releaseDec 11, 2013-
Map releaseMar 26, 2014-
UpdateMar 26, 2014-
Current statusMar 26, 2014Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.5
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.5
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_2506.png

Downloads & links

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Map

FileDownload / File: emd_2506.map.gz / Format: CCP4 / Size: 19.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationC1-IgG
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
4.3 Å/pix.
x 174 pix.
= 748.2 Å		4.3 Å/pix.
x 174 pix.
= 748.2 Å		4.3 Å/pix.
x 174 pix.
= 748.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 4.3 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.5 / Movie #1: 0.5
Minimum - Maximum-0.00031829 - 1.00010979
Average (Standard dev.)0.3666459 (±0.09346347)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions174174174
Spacing174174174
CellA=B=C: 748.2 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.34.34.3
M x/y/z174174174
origin x/y/z0.0000.0000.000
length x/y/z748.200748.200748.200
α/β/γ90.00090.00090.000
start NX/NY/NZ00-40
NX/NY/NZ555581
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS174174174
D min/max/mean-0.0001.0000.367

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Supplemental data

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Sample components

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Entire : DNP-haptenated liposomes, polyclonal goat anti-DNP Antibody, huma...

EntireName: DNP-haptenated liposomes, polyclonal goat anti-DNP Antibody, human C1 (originating from C4 depleted serum)
Components
  • Sample: DNP-haptenated liposomes, polyclonal goat anti-DNP Antibody, human C1 (originating from C4 depleted serum)
  • Protein or peptide: polyclonal IgG
  • Protein or peptide: C1

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Supramolecule #1000: DNP-haptenated liposomes, polyclonal goat anti-DNP Antibody, huma...

SupramoleculeName: DNP-haptenated liposomes, polyclonal goat anti-DNP Antibody, human C1 (originating from C4 depleted serum)
type: sample / ID: 1000
Oligomeric state: one C1 complex binds to one hexamer of membrane bound IgG
Number unique components: 2
Molecular weightTheoretical: 1.65 MDa

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Macromolecule #1: polyclonal IgG

MacromoleculeName: polyclonal IgG / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Oligomeric state: Hexamer / Recombinant expression: No
Source (natural)Organism: Capra hircus (goat) / synonym: Goat / Tissue: blood
Molecular weightTheoretical: 150 KDa

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Macromolecule #2: C1

MacromoleculeName: C1 / type: protein_or_peptide / ID: 2 / Name.synonym: C1r2s2 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No
Source (natural)Organism: Homo sapiens (human) / synonym: Human / Tissue: Blood
Molecular weightTheoretical: 766 KDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

VitrificationCryogen name: ETHANE-PROPANE MIXTURE / Chamber humidity: 95 % / Instrument: LEICA EM GP / Method: Blot 3 seconds before plunging

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: Gatan Quantum SE/963
DateJun 6, 2012
Image recordingCategory: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 51 / Average electron dose: 100 e/Å2 / Bits/pixel: 16
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 7.5 µm / Nominal defocus min: 5.0 µm / Nominal magnification: 50000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Tilt series - Axis1 - Min angle: -60 ° / Tilt series - Axis1 - Max angle: 60 °
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Detailsparticle selected using IMOD stalkInit. C6 for IgG hexamer (lower platform) C2 for C1r2s2 hetero tetramer (upper platform)
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 66.0 Å / Resolution method: OTHER / Software - Name: IMOD, PEET, Spider
Details: Final map is an average of 107 particles, representing the first class after pca classification and k-means clustering of 200 particles. The 200 particles originate from ~50 single and dual axis tomograms
Number subtomograms used: 107
CTF correctionDetails: Phase flipping in IMOD after defocus esimation in TOMOCTF

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