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- EMDB-8500: Subtomogram average of the non-piliated toxin-coregulated pilus m... -

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Basic information

Entry
Database: EMDB / ID: EMD-8500
TitleSubtomogram average of the non-piliated toxin-coregulated pilus machine in Vibrio cholerae cells with tcpD knockout (aligning OM-parts)
Map dataSubtomogram average of the non-piliated toxin-coregulated pilus machine in Vibrio cholerae cells with tcpD knockout (aligning OM-parts)
Sample
  • Complex: Toxin-coregulated pilus machine
Biological speciesVibrio cholerae (bacteria)
Methodsubtomogram averaging / cryo EM / Resolution: 50.0 Å
AuthorsChang Y-W / Kjaer A / Jensen GJ
CitationJournal: Nat Microbiol / Year: 2017
Title: Architecture of the Vibrio cholerae toxin-coregulated pilus machine revealed by electron cryotomography.
Authors: Yi-Wei Chang / Andreas Kjær / Davi R Ortega / Gabriela Kovacikova / John A Sutherland / Lee A Rettberg / Ronald K Taylor / Grant J Jensen /
Abstract: Type IV pili (T4P) are filamentous appendages found on many Bacteria and Archaea. They are helical fibres of pilin proteins assembled by a multi-component macromolecular machine we call the basal ...Type IV pili (T4P) are filamentous appendages found on many Bacteria and Archaea. They are helical fibres of pilin proteins assembled by a multi-component macromolecular machine we call the basal body. Based on pilin features, T4P are classified into type IVa pili (T4aP) and type IVb pili (T4bP). T4aP are more widespread and are involved in cell motility, DNA transfer, host predation and electron transfer. T4bP are less prevalent and are mainly found in enteropathogenic bacteria, where they play key roles in host colonization. Following similar work on T4aP machines, here we use electron cryotomography to reveal the three-dimensional in situ structure of a T4bP machine in its piliated and non-piliated states. The specific machine we analyse is the Vibrio cholerae toxin-coregulated pilus machine (TCPM). Although only about half of the components of the TCPM show sequence homology to components of the previously analysed Myxococcus xanthus T4aP machine (T4aPM), we find that their structures are nevertheless remarkably similar. Based on homologies with components of the M. xanthus T4aPM and additional reconstructions of TCPM mutants in which the non-homologous proteins are individually deleted, we propose locations for all eight TCPM components within the complex. Non-homologous proteins in the T4aPM and TCPM are found to form similar structures, suggesting new hypotheses for their functions and evolutionary histories.
History
DepositionDec 1, 2016-
Header (metadata) releaseDec 14, 2016-
Map releaseFeb 15, 2017-
UpdateFeb 15, 2017-
Current statusFeb 15, 2017Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 140
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 140
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

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Map

FileDownload / File: emd_8500.map.gz / Format: CCP4 / Size: 1.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSubtomogram average of the non-piliated toxin-coregulated pilus machine in Vibrio cholerae cells with tcpD knockout (aligning OM-parts)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
7.8 Å/pix.
x 50 pix.
= 390. Å
7.8 Å/pix.
x 150 pix.
= 1170. Å
7.8 Å/pix.
x 50 pix.
= 390. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 7.8 Å
Density
Contour LevelBy AUTHOR: 140. / Movie #1: 140
Minimum - Maximum0. - 255.
Average (Standard dev.)78.476685000000003 (±55.272390000000001)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions1505050
Spacing5015050
CellA: 390.0 Å / B: 1170.0 Å / C: 390.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z7.87.87.8
M x/y/z5015050
origin x/y/z0.0000.0000.000
length x/y/z390.0001170.000390.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-190-190-190
NX/NY/NZ380380380
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS5015050
D min/max/mean0.000255.00078.477

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Supplemental data

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Sample components

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Entire : Toxin-coregulated pilus machine

EntireName: Toxin-coregulated pilus machine
Components
  • Complex: Toxin-coregulated pilus machine

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Supramolecule #1: Toxin-coregulated pilus machine

SupramoleculeName: Toxin-coregulated pilus machine / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Vibrio cholerae (bacteria)

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferpH: 7
VitrificationCryogen name: ETHANE-PROPANE

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Electron microscopy

MicroscopeFEI POLARA 300
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 1.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 50.0 Å / Resolution method: OTHER / Software - Name: Tomo3d / Number subtomograms used: 325
ExtractionNumber tomograms: 42 / Number images used: 325 / Software - Name: PEET
CTF correctionSoftware - Name: IMOD
Final 3D classificationSoftware - Name: PEET
Final angle assignmentType: OTHER / Software - Name: PEET

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