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- EMDB-8532: Cryo-EM structure of 16nm repeat ciliary microtubule doublet -

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Basic information

Entry
Database: EMDB / ID: EMD-8532
TitleCryo-EM structure of 16nm repeat ciliary microtubule doublet
Map data16nm repeat map of the doublet from Tetrahymena thermophila
Sample
  • Organelle or cellular component: 16nm-repeat of microtubule doublet
Function / homology
Function and homology information


structural constituent of cytoskeleton / microtubule cytoskeleton organization / mitotic cell cycle / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / microtubule / hydrolase activity / GTPase activity / GTP binding / metal ion binding / cytoplasm
Similarity search - Function
Alpha tubulin / Tubulin-beta mRNA autoregulation signal. / Beta tubulin, autoregulation binding site / Beta tubulin / Tubulin / Tubulin, C-terminal / Tubulin C-terminal domain / Tubulin, conserved site / Tubulin subunits alpha, beta, and gamma signature. / Tubulin/FtsZ family, C-terminal domain ...Alpha tubulin / Tubulin-beta mRNA autoregulation signal. / Beta tubulin, autoregulation binding site / Beta tubulin / Tubulin / Tubulin, C-terminal / Tubulin C-terminal domain / Tubulin, conserved site / Tubulin subunits alpha, beta, and gamma signature. / Tubulin/FtsZ family, C-terminal domain / Tubulin/FtsZ-like, C-terminal domain / Tubulin/FtsZ, C-terminal / Tubulin/FtsZ, 2-layer sandwich domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ, GTPase domain / Tubulin/FtsZ, GTPase domain superfamily
Similarity search - Domain/homology
Tubulin alpha chain / Tubulin beta chain
Similarity search - Component
Biological speciesTetrahymena thermophila (eukaryote)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.2 Å
AuthorsIchikawa M / Liu D / Kastritis PL / Basu K / Bui KH
CitationJournal: Nat Commun / Year: 2017
Title: Subnanometre-resolution structure of the doublet microtubule reveals new classes of microtubule-associated proteins.
Authors: Muneyoshi Ichikawa / Dinan Liu / Panagiotis L Kastritis / Kaustuv Basu / Tzu Chin Hsu / Shunkai Yang / Khanh Huy Bui /
Abstract: Cilia are ubiquitous, hair-like appendages found in eukaryotic cells that carry out functions of cell motility and sensory reception. Cilia contain an intriguing cytoskeletal structure, termed the ...Cilia are ubiquitous, hair-like appendages found in eukaryotic cells that carry out functions of cell motility and sensory reception. Cilia contain an intriguing cytoskeletal structure, termed the axoneme that consists of nine doublet microtubules radially interlinked and longitudinally organized in multiple specific repeat units. Little is known, however, about how the axoneme allows cilia to be both actively bendable and sturdy or how it is assembled. To answer these questions, we used cryo-electron microscopy to structurally analyse several of the repeating units of the doublet at sub-nanometre resolution. This structural detail enables us to unambiguously assign α- and β-tubulins in the doublet microtubule lattice. Our study demonstrates the existence of an inner sheath composed of different kinds of microtubule inner proteins inside the doublet that likely stabilizes the structure and facilitates the specific building of the B-tubule.
History
DepositionDec 21, 2016-
Header (metadata) releaseJan 18, 2017-
Map releaseMay 10, 2017-
UpdateFeb 14, 2018-
Current statusFeb 14, 2018Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_8532.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation16nm repeat map of the doublet from Tetrahymena thermophila
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.75 Å/pix.
x 256 pix.
= 704. Å
2.75 Å/pix.
x 256 pix.
= 704. Å
2.75 Å/pix.
x 256 pix.
= 704. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.75 Å
Density
Contour LevelBy AUTHOR: 0.03 / Movie #1: 0.03
Minimum - Maximum-0.074407496 - 0.15960655
Average (Standard dev.)0.0019532945 (±0.014449678)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 704.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.752.752.75
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z704.000704.000704.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ281156
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0740.1600.002

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Supplemental data

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Sample components

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Entire : 16nm-repeat of microtubule doublet

EntireName: 16nm-repeat of microtubule doublet
Components
  • Organelle or cellular component: 16nm-repeat of microtubule doublet

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Supramolecule #1: 16nm-repeat of microtubule doublet

SupramoleculeName: 16nm-repeat of microtubule doublet / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Tetrahymena thermophila (eukaryote) / Strain: SB255 / Organelle: cilia / Location in cell: cilia

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statefilament

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Sample preparation

Concentration2 mg/mL
BufferpH: 7.4
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV / Details: Blot force 3 for 5 seconds.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Digitization - Frames/image: 1-7 / Number real images: 5983 / Average exposure time: 1.4 sec. / Average electron dose: 45.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.8 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 59000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 64116
CTF correctionSoftware - Name: CTFFIND4
Startup modelType of model: EMDB MAP
EMDB ID:
Final reconstructionNumber classes used: 1 / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 6.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.4,2.0) / Number images used: 64116
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: SPIDER (ver. 19.02)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 1.4,2.0)

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