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- EMDB-5694: Type III secretion system (Injectisome) of Yersinia enterocolitic... -

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Basic information

Entry
Database: EMDB / ID: EMD-5694
TitleType III secretion system (Injectisome) of Yersinia enterocolitica in situ
Map dataIn situ structure of the Yersinia enterocolitica Injectisome
Sample
  • Sample: Yersinia enterocolitica intact bacterial cells containing type III secretion injectisomes
  • Protein or peptide: Bacterial Type III Secretion System
KeywordsT3SS / YscC / YscD / YscJ / YscV / cryo electron tomography / sub-volume averaging.
Biological speciesYersinia enterocolitica (bacteria)
Methodsubtomogram averaging / cryo EM / Resolution: 40.0 Å
AuthorsKudryashev M / Stenta M / Schmelz S / Amstutz M / Wiesand U / Castano-Diez D / Degiacomi M / Muennich S / Bleck CKE / Kowal J ...Kudryashev M / Stenta M / Schmelz S / Amstutz M / Wiesand U / Castano-Diez D / Degiacomi M / Muennich S / Bleck CKE / Kowal J / Diepold A / Heinz DW / Dal Peraro M / Cornelis GR / Stahlberg H
CitationJournal: Elife / Year: 2013
Title: In situ structural analysis of the Yersinia enterocolitica injectisome.
Authors: Mikhail Kudryashev / Marco Stenta / Stefan Schmelz / Marlise Amstutz / Ulrich Wiesand / Daniel Castaño-Díez / Matteo T Degiacomi / Stefan Münnich / Christopher Ke Bleck / Julia Kowal / ...Authors: Mikhail Kudryashev / Marco Stenta / Stefan Schmelz / Marlise Amstutz / Ulrich Wiesand / Daniel Castaño-Díez / Matteo T Degiacomi / Stefan Münnich / Christopher Ke Bleck / Julia Kowal / Andreas Diepold / Dirk W Heinz / Matteo Dal Peraro / Guy R Cornelis / Henning Stahlberg /
Abstract: Injectisomes are multi-protein transmembrane machines allowing pathogenic bacteria to inject effector proteins into eukaryotic host cells, a process called type III secretion. Here we present the ...Injectisomes are multi-protein transmembrane machines allowing pathogenic bacteria to inject effector proteins into eukaryotic host cells, a process called type III secretion. Here we present the first three-dimensional structure of Yersinia enterocolitica and Shigella flexneri injectisomes in situ and the first structural analysis of the Yersinia injectisome. Unexpectedly, basal bodies of injectisomes inside the bacterial cells showed length variations of 20%. The in situ structures of the Y. enterocolitica and S. flexneri injectisomes had similar dimensions and were significantly longer than the isolated structures of related injectisomes. The crystal structure of the inner membrane injectisome component YscD appeared elongated compared to a homologous protein, and molecular dynamics simulations documented its elongation elasticity. The ring-shaped secretin YscC at the outer membrane was stretched by 30-40% in situ, compared to its isolated liposome-embedded conformation. We suggest that elasticity is critical for some two-membrane spanning protein complexes to cope with variations in the intermembrane distance. DOI:http://dx.doi.org/10.7554/eLife.00792.001.
History
DepositionJun 18, 2013-
Header (metadata) releaseJul 17, 2013-
Map releaseJul 17, 2013-
UpdateAug 14, 2013-
Current statusAug 14, 2013Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.6
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.6
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_5694_1.jpg

Downloads & links

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Map

FileDownload / File: emd_5694.map.gz / Format: CCP4 / Size: 7.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationIn situ structure of the Yersinia enterocolitica Injectisome
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.74 Å/pix.
x 128 pix.
= 94.72 Å		0.74 Å/pix.
x 128 pix.
= 94.72 Å		0.74 Å/pix.
x 128 pix.
= 94.72 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel size
XYZ
EMDB info.0.740.740.74
CCP4 map header0.740.740.74
EM Navigator Movie #17.47.47.4
Density

Histogram

Histogram (log scale)
Contour LevelBy EMDB: 1.1 / Movie #1: 0.6
Minimum - Maximum-3.88122559 - 5.41991997
Average (Standard dev.)0.0 (±0.99111927)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-64-64-64
Dimensions128128128
Spacing128128128
CellA=B=C: 94.72 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.740.740.74
M x/y/z128128128
origin x/y/z0.0000.0000.000
length x/y/z94.72094.72094.720
α/β/γ90.00090.00090.000
start NX/NY/NZ-132-122-147
NX/NY/NZ250274261
MAP C/R/S123
start NC/NR/NS-64-64-64
NC/NR/NS128128128
D min/max/mean-3.8815.420-0.000

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Supplemental data

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Sample components

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Entire : Yersinia enterocolitica intact bacterial cells containing type II...

EntireName: Yersinia enterocolitica intact bacterial cells containing type III secretion injectisomes
Components
  • Sample: Yersinia enterocolitica intact bacterial cells containing type III secretion injectisomes
  • Protein or peptide: Bacterial Type III Secretion System

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Supramolecule #1000: Yersinia enterocolitica intact bacterial cells containing type II...

SupramoleculeName: Yersinia enterocolitica intact bacterial cells containing type III secretion injectisomes
type: sample / ID: 1000
Details: Bacteria were taken from cell culture and were rapidly prepared for cryo imaging
Oligomeric state: various / Number unique components: 9

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Macromolecule #1: Bacterial Type III Secretion System

MacromoleculeName: Bacterial Type III Secretion System / type: protein_or_peptide / ID: 1 / Name.synonym: Injectisome, T3SS / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Yersinia enterocolitica (bacteria)

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferDetails: PBS
GridDetails: Quantifoil holey carbon grids with various hole sizes
VitrificationCryogen name: ETHANE / Chamber temperature: 77 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Tridem / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV
DateNov 1, 2009
Image recordingCategory: CCD / Film or detector model: GATAN ULTRASCAN 1000 (2k x 2k) / Number real images: 61 / Average electron dose: 100 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 19500 / Illumination mode: FLOOD BEAM / Imaging mode: DARK FIELD / Cs: 2.2 mm / Nominal defocus max: -15.0 µm / Nominal defocus min: -3.0 µm
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

Final reconstructionResolution.type: BY AUTHOR / Resolution: 40.0 Å / Resolution method: OTHER / Number subtomograms used: 941
CTF correctionDetails: Each micrograph was phase flipped assuming the value of nominal defocus. Focal pair tomography was employed.
Final 3D classificationNumber classes: 1

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