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- EMDB-5619: Outer dynein arm - microtubule complex, labeled with biotin-ADP a... -

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Basic information

Entry
Database: EMDB / ID: EMD-5619
TitleOuter dynein arm - microtubule complex, labeled with biotin-ADP and streptavidin
Map dataReconstruction of outer dynein arm-microtubule complex labeled with biotin-ADP and streptavidin
Sample
  • Sample: Outer dynein arms from Chlamydomonas reinhardtii bound to porcine microtubules
  • Organelle or cellular component: Outer dynen arm-microbutule complex
Keywordsflagella / Chlamydomonas / outer dynein arm / microtubule
Biological speciesChlamydomonas reinhardtii (plant)
Methodsingle particle reconstruction / cryo EM / Resolution: 34.0 Å
AuthorsOda T / Yagi T / Yanagisawa HA / Kikkawa M
CitationJournal: Curr Biol / Year: 2013
Title: Identification of the outer-inner dynein linker as a hub controller for axonemal dynein activities.
Authors: Toshiyuki Oda / Toshiki Yagi / Haruaki Yanagisawa / Masahide Kikkawa /
Abstract: BACKGROUND: In flagella, the outer dynein arm (ODA) and inner dynein arm (IDA) play distinct roles in generating beating motion. However, functional communications between the two dyneins have not been investigated.
RESULTS: Here, we demonstrated by cryo-electron microscopy and chemical crosslinking that intermediate chain 2 (IC2) of ODAs interacts with the dynein regulatory complex in the axoneme and ...RESULTS: Here, we demonstrated by cryo-electron microscopy and chemical crosslinking that intermediate chain 2 (IC2) of ODAs interacts with the dynein regulatory complex in the axoneme and constitutes part of the outer-inner dynein (OID) linker. Furthermore, we identified IC2 as a functional hub between ODAs and IDAs based on the phenotypes of Chlamydomonas mutants expressing biotinylation-tagged IC2. The flagella of the IC2 mutant showed activated microtubule sliding and enhanced ATPase activities of ODAs, as well as an altered waveform, indicating attenuated IDA activity.
CONCLUSIONS: We concluded that the OID linker controls both ODAs and IDAs and regulates flagellar beating.
History
DepositionMar 22, 2013-
Header (metadata) releaseApr 24, 2013-
Map releaseApr 24, 2013-
UpdateApr 23, 2014-
Current statusApr 23, 2014Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 82.1
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 82.1
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

400_5619.gif

Downloads & links

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Map

FileDownload / File: emd_5619.map.gz / Format: CCP4 / Size: 8.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of outer dynein arm-microtubule complex labeled with biotin-ADP and streptavidin
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
4.87 Å/pix.
x 100 pix.
= 487. Å		4.87 Å/pix.
x 186 pix.
= 905.82 Å		4.87 Å/pix.
x 126 pix.
= 613.62 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: 4.87 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 82.099999999999994 / Movie #1: 82.1
Minimum - Maximum-82.260139469999999 - 214.341033939999988
Average (Standard dev.)10.847085 (±53.627422330000002)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions186126100
Spacing186126100
CellA: 613.62 Å / B: 905.82 Å / C: 487.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.874.874.87
M x/y/z126186100
origin x/y/z0.0000.0000.000
length x/y/z613.620905.820487.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-132-122-147
NX/NY/NZ250274261
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS126186100
D min/max/mean-82.260214.34110.847

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Supplemental data

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Sample components

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Entire : Outer dynein arms from Chlamydomonas reinhardtii bound to porcine...

EntireName: Outer dynein arms from Chlamydomonas reinhardtii bound to porcine microtubules
Components
  • Sample: Outer dynein arms from Chlamydomonas reinhardtii bound to porcine microtubules
  • Organelle or cellular component: Outer dynen arm-microbutule complex

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Supramolecule #1000: Outer dynein arms from Chlamydomonas reinhardtii bound to porcine...

SupramoleculeName: Outer dynein arms from Chlamydomonas reinhardtii bound to porcine microtubules
type: sample / ID: 1000 / Number unique components: 2

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Supramolecule #1: Outer dynen arm-microbutule complex

SupramoleculeName: Outer dynen arm-microbutule complex / type: organelle_or_cellular_component / ID: 1 / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Chlamydomonas reinhardtii (plant) / Strain: 137c / Organelle: flagella / Location in cell: axoneme

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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

Concentration0.03 mg/mL
BufferpH: 7.4
Details: 10 mM Hepes-NaOH pH 7.4, 1 mM DTT, 5 mM MgCl2, 1 mM EGTA, 10% trehalose, 0.1% Nonidet P-40, 5 microM paclitaxel
GridDetails: 400 mesh copper grid with home-made holey carbon support, glow discharged in amylamine atmosphere
VitrificationCryogen name: ETHANE / Chamber temperature: 90 K / Instrument: LEICA KF80
Method: Two-side blot by hand for 10 seconds before plunging

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Electron microscopy

MicroscopeJEOL 2010F
TemperatureMin: 80 K / Max: 97 K / Average: 95 K
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 200,000 times magnification
DateMay 29, 2011
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: OTHER / Digitization - Sampling interval: 10 µm / Number real images: 48 / Average electron dose: 7 e/Å2 / Bits/pixel: 16
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 40000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 3.3 mm / Nominal defocus max: 3.3 µm / Nominal defocus min: 1.6 µm / Nominal magnification: 40000
Sample stageSpecimen holder: Gatan 626 Single Tilt Liquid Nitrogen Cryo Transfer Holder
Specimen holder model: GATAN LIQUID NITROGEN

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Image processing

CTF correctionDetails: Wiener filter
Final reconstructionApplied symmetry - Helical parameters - Δz: 122.3 Å
Applied symmetry - Helical parameters - Δ&Phi: 179.2 °
Resolution.type: BY AUTHOR / Resolution: 34.0 Å / Resolution method: FSC 0.33 CUT-OFF / Software - Name: FREALIGN / Number images used: 3792

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