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- EMDB-8837: Sub-tomogram average of I1 dynein Post class -

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Basic information

Entry
Database: EMDB / ID: EMD-8837
TitleSub-tomogram average of I1 dynein Post class
Map dataSub-tomogram average of I1 dynein Post class: the two dynein heavy chains of sea urchin sperm flagellar I1 dynein complex are in the post-powerstroke state.
Sample
  • Cell: Sea urchin sperm
Biological speciesStrongylocentrotus purpuratus (purple sea urchin)
Methodsubtomogram averaging / cryo EM / Resolution: 41.0 Å
AuthorsLin J / Nicastro D
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM083122 United States
March of Dimes Foundation United States
CitationJournal: Science / Year: 2018
Title: Asymmetric distribution and spatial switching of dynein activity generates ciliary motility.
Authors: Jianfeng Lin / Daniela Nicastro /
Abstract: Motile cilia and flagella are essential, highly conserved organelles, and their motility is driven by the coordinated activities of multiple dynein isoforms. The prevailing "switch-point" hypothesis ...Motile cilia and flagella are essential, highly conserved organelles, and their motility is driven by the coordinated activities of multiple dynein isoforms. The prevailing "switch-point" hypothesis posits that dyneins are asymmetrically activated to drive flagellar bending. To test this model, we applied cryo-electron tomography to visualize activity states of individual dyneins relative to their locations along beating flagella of sea urchin sperm cells. As predicted, bending was generated by the asymmetric distribution of dynein activity on opposite sides of the flagellum. However, contrary to predictions, most dyneins were in their active state, and the smaller population of conformationally inactive dyneins switched flagellar sides relative to the bending direction. Thus, our data suggest a "switch-inhibition" mechanism in which force imbalance is generated by inhibiting, rather than activating, dyneins on alternating sides of the flagellum.
History
DepositionJul 19, 2017-
Header (metadata) releaseAug 23, 2017-
Map releaseMay 9, 2018-
UpdateJan 29, 2020-
Current statusJan 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 130
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 130
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_8837.png

Downloads & links

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Map

FileDownload / File: emd_8837.map.gz / Format: CCP4 / Size: 704.1 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSub-tomogram average of I1 dynein Post class: the two dynein heavy chains of sea urchin sperm flagellar I1 dynein complex are in the post-powerstroke state.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
9.86 Å/pix.
x 50 pix.
= 492.8 Å		9.86 Å/pix.
x 45 pix.
= 443.52 Å		9.86 Å/pix.
x 80 pix.
= 788.48 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: 9.856 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 130 / Movie #1: 130
Minimum - Maximum88.75412 - 172.66785
Average (Standard dev.)125.353745 (±9.563898)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions458050
Spacing804550
CellA: 788.48 Å / B: 443.52 Å / C: 492.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z9.8569.8569.856
M x/y/z804550
origin x/y/z0.0000.0000.000
length x/y/z788.480443.520492.800
α/β/γ90.00090.00090.000
start NX/NY/NZ-38-19-20
NX/NY/NZ858082
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS804550
D min/max/mean88.754172.668125.354

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Supplemental data

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Sample components

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Entire : Sea urchin sperm

EntireName: Sea urchin sperm
Components
  • Cell: Sea urchin sperm

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Supramolecule #1: Sea urchin sperm

SupramoleculeName: Sea urchin sperm / type: cell / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Sperm were diluted in demembranation buffer (30 mM HEPES, pH 8.0, 150 mM KCl, 4 mM MgCl2, 0.5 mM EGTA, 0.1% Triton X-100) and incubated for one minute to remove the flagellar membrane. The ...Details: Sperm were diluted in demembranation buffer (30 mM HEPES, pH 8.0, 150 mM KCl, 4 mM MgCl2, 0.5 mM EGTA, 0.1% Triton X-100) and incubated for one minute to remove the flagellar membrane. The sperm were then collected by centrifugation at 1000x g, and resuspended in demembranation buffer (but without Triton X-100).
Source (natural)Organism: Strongylocentrotus purpuratus (purple sea urchin)

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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: 8
Component:
ConcentrationFormulaName
4.0 mMMgCl2magnesium chloride
0.5 mM[-CH2OCH2CH2N(CH2CO2H)2]2EGTA
150.0 mMKClpotassium chloride
30.0 mMC8H18N2O4SHEPES
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER

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Electron microscopy

MicroscopeFEI TECNAI F30
Specialist opticsEnergy filter - Name: GIF
Image recordingFilm or detector model: GATAN MULTISCAN / Average electron dose: 1.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 8.0 µm / Nominal defocus min: 6.0 µm / Nominal magnification: 13500
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Tecnai F30 / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 41.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: PEET (ver. 1.9.0) / Number subtomograms used: 392
ExtractionNumber tomograms: 7 / Number images used: 1134 / Software - Name: PEET (ver. 1.9.0)
Final 3D classificationSoftware - Name: PEET (ver. 1.9.0)
Final angle assignmentType: OTHER

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