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- EMDB-9385: The central pair complex of sea urchin (Strongylocentrotus purpur... -

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Entry
Database: EMDB / ID: EMD-9385
TitleThe central pair complex of sea urchin (Strongylocentrotus purpuratus) sperm resolved by cryo-electron tomography and sub-tomogram averaging
Map dataThe central pair complex of sea urchin (Strongylocentrotus purpuratus) sperm resolved by cryo-electron tomography and sub-tomogram averaging
Sample
  • Organelle or cellular component: The central pair complex/apparatus (2167 repeats) averaged from 41 Strongylocentrotus sea urchin sperms
Biological speciesStrongylocentrotus purpuratus (purple sea urchin)
Methodsubtomogram averaging / cryo EM / Resolution: 35.0 Å
AuthorsCarbajal-Gonzalez BI / Heuser T / Fu X / Lin J / Smith BW / Mitchell DR / Fu G / Nicastro D
CitationJournal: Cytoskeleton (Hoboken) / Year: 2013
Title: Conserved structural motifs in the central pair complex of eukaryotic flagella.
Authors: Blanca I Carbajal-González / Thomas Heuser / Xiaofeng Fu / Jianfeng Lin / Brandon W Smith / David R Mitchell / Daniela Nicastro /
Abstract: Cilia and flagella are conserved hair-like appendages of eukaryotic cells that function as sensing and motility generating organelles. Motility is driven by thousands of axonemal dyneins that require ...Cilia and flagella are conserved hair-like appendages of eukaryotic cells that function as sensing and motility generating organelles. Motility is driven by thousands of axonemal dyneins that require precise regulation. One essential motility regulator is the central pair complex (CPC) and many CPC defects cause paralysis of cilia/flagella. Several human diseases, such as immotile cilia syndrome, show CPC abnormalities, but little is known about the detailed three-dimensional (3D) structure and function of the CPC. The CPC is located in the center of typical [9+2] cilia/flagella and is composed of two singlet microtubules (MTs), each with a set of associated projections that extend toward the surrounding nine doublet MTs. Using cryo-electron tomography coupled with subtomogram averaging, we visualized and compared the 3D structures of the CPC in both the green alga Chlamydomonas and the sea urchin Strongylocentrotus at the highest resolution published to date. Despite the evolutionary distance between these species, their CPCs exhibit remarkable structural conservation. We identified several new projections, including those that form the elusive sheath, and show that the bridge has a more complex architecture than previously thought. Organism-specific differences include the presence of MT inner proteins in Chlamydomonas, but not Strongylocentrotus, and different overall outlines of the highly connected projection network, which forms a round-shaped cylinder in algae, but is more oval in sea urchin. These differences could be adaptations to the mechanical requirements of the rotating CPC in Chlamydomonas, compared to the Strongylocentrotus CPC which has a fixed orientation.
History
DepositionJan 2, 2019-
Header (metadata) releaseJan 16, 2019-
Map releaseJan 16, 2019-
UpdateJan 16, 2019-
Current statusJan 16, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 126
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 126
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_9385.png

Downloads & links

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Map

FileDownload / File: emd_9385.map.gz / Format: CCP4 / Size: 3.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThe central pair complex of sea urchin (Strongylocentrotus purpuratus) sperm resolved by cryo-electron tomography and sub-tomogram averaging
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
9.86 Å/pix.
x 100 pix.
= 985.6 Å		9.86 Å/pix.
x 100 pix.
= 985.6 Å		9.86 Å/pix.
x 100 pix.
= 985.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 9.856 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 126.0 / Movie #1: 126
Minimum - Maximum107.892334000000005 - 139.692520000000002
Average (Standard dev.)118.929749999999999 (±4.037084)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions100100100
Spacing100100100
CellA=B=C: 985.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z9.8569.8569.856
M x/y/z100100100
origin x/y/z0.0000.0000.000
length x/y/z985.600985.600985.600
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS100100100
D min/max/mean107.892139.693118.930

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Supplemental data

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Sample components

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Entire : The central pair complex/apparatus (2167 repeats) averaged from 4...

EntireName: The central pair complex/apparatus (2167 repeats) averaged from 41 Strongylocentrotus sea urchin sperms
Components
  • Organelle or cellular component: The central pair complex/apparatus (2167 repeats) averaged from 41 Strongylocentrotus sea urchin sperms

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Supramolecule #1: The central pair complex/apparatus (2167 repeats) averaged from 4...

SupramoleculeName: The central pair complex/apparatus (2167 repeats) averaged from 41 Strongylocentrotus sea urchin sperms
type: organelle_or_cellular_component / ID: 1 / Parent: 0
Source (natural)Organism: Strongylocentrotus purpuratus (purple sea urchin) / Organelle: sperm / Location in cell: tail of sperm

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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: 7.4 / Details: artificial sea water
GridModel: Quantifoil R2/2 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 101.325 kPa
VitrificationCryogen name: ETHANE / Chamber temperature: 298 K / Instrument: HOMEMADE PLUNGER
Details: back-side blotting for 1.5-2.5 seconds before plunging.
DetailsFreshly spawned sea urchin sperms in artificial sea water were used.

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Electron microscopy

MicroscopeFEI TECNAI F30
Specialist opticsEnergy filter - Name: GIF 2000 / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN ULTRASCAN 1000 (2k x 2k) / Average electron dose: 1.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated defocus max: 8.0 µm / Calibrated defocus min: 6.0 µm / Calibrated magnification: 13500 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Sample stageSpecimen holder model: GATAN LIQUID NITROGEN / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Tecnai F30 / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 35.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMOD / Number subtomograms used: 2167
ExtractionNumber tomograms: 41 / Number images used: 2167
Final angle assignmentType: OTHER

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