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- EMDB-2520: Injectisome in the tomogram of Salmonella mini-cell -

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Basic information

Entry
Database: EMDB / ID: EMD-2520
TitleInjectisome in the tomogram of Salmonella mini-cellType three secretion system
Map dataSubtomogram averaged structure of Injectisome from Salmonella typhimurium
Sample
  • Sample: Injectisome from Salmonella typhimurium mini-cell in situType three secretion system
  • Organelle or cellular component: type III secretion systemType three secretion system
KeywordsInjectisome / Type III secretion system / Salmonella typhimurium / Mini-cell / Electron cryotomography
Biological speciesSalmonella enterica subsp. enterica serovar Typhimurium (bacteria)
Methodsubtomogram averaging / cryo EM / Resolution: 44.0 Å
AuthorsKawamoto A / Morimoto VY / Miyata T / Minamino T / Hughes TK / Kato T / Namba K
CitationJournal: Sci Rep / Year: 2013
Title: Common and distinct structural features of Salmonella injectisome and flagellar basal body.
Authors: Akihiro Kawamoto / Yusuke V Morimoto / Tomoko Miyata / Tohru Minamino / Kelly T Hughes / Takayuki Kato / Keiichi Namba /
Abstract: Bacterial pathogens use an injectisome to deliver virulence proteins into eukaryotic host cells. The bacterial flagellum and injectisome export their component proteins for self-assembly. These two ...Bacterial pathogens use an injectisome to deliver virulence proteins into eukaryotic host cells. The bacterial flagellum and injectisome export their component proteins for self-assembly. These two systems show high structural similarities and are classified as the type III secretion system, but it remains elusive how similar they are in situ because the structures of these complexes isolated from cells and visualized by electron cryomicroscopy have shown only the export channel and housing for the export apparatus. Here we report in situ structures of Salmonella injectisome and flagellum by electron cryotomography. The injectisome lacks the flagellar basal body C-ring, but a wing-like disc and a globular density corresponding to the export gate platform and ATPase hexamer ring, respectively, are stably attached through thin connectors, revealing yet unidentified common architectures of the two systems. The ATPase ring is far from the disc, suggesting that both apparatuses are observed in an export-off state.
History
DepositionNov 26, 2013-
Header (metadata) releaseDec 18, 2013-
Map releaseDec 25, 2013-
UpdateDec 25, 2013-
Current statusDec 25, 2013Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.237
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.237
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

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Map

FileDownload / File: emd_2520.map.gz / Format: CCP4 / Size: 2.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSubtomogram averaged structure of Injectisome from Salmonella typhimurium
Voxel sizeX=Y=Z: 11.4 Å
Density
Contour LevelBy AUTHOR: 0.237 / Movie #1: 0.237
Minimum - Maximum-2.02735949 - 5.20517445
Average (Standard dev.)0.01956169 (±0.89927512)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-767155240
Dimensions828282
Spacing828282
CellA=B=C: 934.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z11.411.411.4
M x/y/z828282
origin x/y/z0.0000.0000.000
length x/y/z934.800934.800934.800
α/β/γ90.00090.00090.000
start NX/NY/NZ-184-184-183
NX/NY/NZ368368368
MAP C/R/S123
start NC/NR/NS155-767240
NC/NR/NS828282
D min/max/mean-2.0275.2050.020

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Supplemental data

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Sample components

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Entire : Injectisome from Salmonella typhimurium mini-cell in situ

EntireName: Injectisome from Salmonella typhimurium mini-cell in situType three secretion system
Components
  • Sample: Injectisome from Salmonella typhimurium mini-cell in situType three secretion system
  • Organelle or cellular component: type III secretion systemType three secretion system

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Supramolecule #1000: Injectisome from Salmonella typhimurium mini-cell in situ

SupramoleculeName: Injectisome from Salmonella typhimurium mini-cell in situ
type: sample / ID: 1000 / Number unique components: 1
Molecular weightTheoretical: 3.5 MDa

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Supramolecule #1: type III secretion system

SupramoleculeName: type III secretion system / type: organelle_or_cellular_component / ID: 1 / Name.synonym: injectisome / Recombinant expression: No
Source (natural)Organism: Salmonella enterica subsp. enterica serovar Typhimurium (bacteria)
Location in cell: Plasma membrane
Molecular weightTheoretical: 3.5 MDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferDetails: M9 medium (17.1g Na2HPO4-12H2O, 3g KH2PO4, 0.5g NaCl, 1g NH4Cl, 0.2%glycerol, 1% typtone, 1mM MgSO4 per litre)
GridDetails: Quantifoil molybdenum 200 mesh R0.6/1.0 grid with thin carbon support
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK II

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 49030 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 7.0 µm / Nominal defocus min: 4.0 µm / Nominal magnification: 29000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Tilt series - Axis1 - Min angle: -70 ° / Tilt series - Axis1 - Max angle: 70 °
TemperatureMin: 80 K
DateDec 17, 2012
Image recordingCategory: CCD / Film or detector model: FEI FALCON I (4k x 4k) / Average electron dose: 200 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C24 (24 fold cyclic) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 44.0 Å / Resolution method: OTHER / Software - Name: EMAN / Number subtomograms used: 75

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