+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-5758 | |||||||||
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Title | Structural mechanism of the dynein powerstroke | |||||||||
Map data | Reconstruction of axonemal dyneins in pre-powerstroke states. The dyneins show two distinct pre-powerstroke conformations: pre-I (detached, bottom dynein) and pre-II (microtubule-bound, top dynein). | |||||||||
Sample |
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Keywords | dynein movement / flagellar motility / axoneme | |||||||||
Function / homology | Function and homology information karyogamy / establishment of mitotic spindle localization / astral microtubule / nuclear migration along microtubule / minus-end-directed microtubule motor activity / dynein light intermediate chain binding / cytoplasmic dynein complex / nuclear migration / spindle pole body / dynein intermediate chain binding ...karyogamy / establishment of mitotic spindle localization / astral microtubule / nuclear migration along microtubule / minus-end-directed microtubule motor activity / dynein light intermediate chain binding / cytoplasmic dynein complex / nuclear migration / spindle pole body / dynein intermediate chain binding / cytoplasmic microtubule / establishment of mitotic spindle orientation / mitotic sister chromatid segregation / cytoplasmic microtubule organization / Neutrophil degranulation / mitotic spindle organization / cell cortex / ATP hydrolysis activity / ATP binding / cytoplasm Similarity search - Function | |||||||||
Biological species | Strongylocentrotus purpuratus (purple sea urchin) | |||||||||
Method | subtomogram averaging / cryo EM / Resolution: 30.0 Å | |||||||||
Authors | Lin J / Okada K / Raytchev M / Smith MC / Nicastro D | |||||||||
Citation | Journal: Nat Cell Biol / Year: 2014 Title: Structural mechanism of the dynein power stroke. Authors: Jianfeng Lin / Kyoko Okada / Milen Raytchev / Maria C Smith / Daniela Nicastro / Abstract: Dyneins are large microtubule motor proteins required for mitosis, intracellular transport and ciliary and flagellar motility. They generate force through a power-stroke mechanism, which is an ATP- ...Dyneins are large microtubule motor proteins required for mitosis, intracellular transport and ciliary and flagellar motility. They generate force through a power-stroke mechanism, which is an ATP-consuming cycle of pre- and post-power-stroke conformational changes that cause relative motion between different dynein domains. However, key structural details of dynein's force generation remain elusive. Here, using cryo-electron tomography of intact, active (that is, beating), rapidly frozen sea urchin sperm flagella, we determined the in situ three-dimensional structures of all domains of both pre- and post-power-stroke dynein, including the previously unresolved linker and stalk of pre-power-stroke dynein. Our results reveal that the rotation of the head relative to the linker is the key action in dynein movement, and that there are at least two distinct pre-power-stroke conformations: pre-I (microtubule-detached) and pre-II (microtubule-bound). We provide three-dimensional reconstructions of native dyneins in three conformational states, in situ, allowing us to propose a molecular model of the structural cycle underlying dynein movement. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_5758.map.gz | 293.1 KB | EMDB map data format | |
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Header (meta data) | emd-5758-v30.xml emd-5758.xml | 10.3 KB 10.3 KB | Display Display | EMDB header |
Images | 400_5758.gif 80_5758.gif | 66.2 KB 5 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-5758 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-5758 | HTTPS FTP |
-Validation report
Summary document | emd_5758_validation.pdf.gz | 308.1 KB | Display | EMDB validaton report |
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Full document | emd_5758_full_validation.pdf.gz | 307.7 KB | Display | |
Data in XML | emd_5758_validation.xml.gz | 4.6 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5758 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5758 | HTTPS FTP |
-Related structure data
Related structure data | 3j68MC 5757C 3j67C M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_5758.map.gz / Format: CCP4 / Size: 344.7 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Reconstruction of axonemal dyneins in pre-powerstroke states. The dyneins show two distinct pre-powerstroke conformations: pre-I (detached, bottom dynein) and pre-II (microtubule-bound, top dynein). | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 9.856 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Cryo-electron tomography and subtomographic average (2800 axonema...
Entire | Name: Cryo-electron tomography and subtomographic average (2800 axonemal repeats) of active sea urchin sperm flagella reveal two distinct pre-powerstroke conformations: pre-I (detached) and pre-II (microtubule-bound). |
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Components |
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-Supramolecule #1000: Cryo-electron tomography and subtomographic average (2800 axonema...
Supramolecule | Name: Cryo-electron tomography and subtomographic average (2800 axonemal repeats) of active sea urchin sperm flagella reveal two distinct pre-powerstroke conformations: pre-I (detached) and pre-II (microtubule-bound). type: sample / ID: 1000 / Number unique components: 1 |
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-Supramolecule #1: dynein
Supramolecule | Name: dynein / type: organelle_or_cellular_component / ID: 1 / Details: Sperm were frozen while actively beating. / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Strongylocentrotus purpuratus (purple sea urchin) / synonym: sea urchin / Cell: sperm / Organelle: flagella |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | subtomogram averaging |
Aggregation state | cell |
-Sample preparation
Buffer | pH: 8 Details: 360 mM NaCl, 50 mM MgCl2, 10 mM CaCl2, 10 mM KCl, 30 mM HEPES, pH 8.0 |
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Grid | Details: Quantifoil holey carbon grids Cu 200 mesh R2/2 |
Vitrification | Cryogen name: ETHANE / Chamber temperature: 100 K / Instrument: HOMEMADE PLUNGER / Method: Blot for 1.5-2.5 seconds before plunging |
-Electron microscopy
Microscope | FEI TECNAI F30 |
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Temperature | Average: 80 K |
Specialist optics | Energy filter - Name: GATAN postcolumn filter GIF / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV |
Date | Apr 7, 2012 |
Image recording | Category: CCD / Film or detector model: GENERIC GATAN (2k x 2k) / Average electron dose: 100 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 8.0 µm / Nominal defocus min: 6.0 µm / Nominal magnification: 13500 |
Sample stage | Specimen holder: Liquid nitrogen cooled / Specimen holder model: GATAN LIQUID NITROGEN / Tilt series - Axis1 - Min angle: -65 ° / Tilt series - Axis1 - Max angle: 65 ° |
Experimental equipment | Model: Tecnai F30 / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 30.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMOD, PEET Details: Final maps were calculated by averaging 2800 particles from 41 tomograms. Axonemal repeats (96 nm long) from 41 tomograms (reconstructed using fiducial alignment and weighted backprojection, ...Details: Final maps were calculated by averaging 2800 particles from 41 tomograms. Axonemal repeats (96 nm long) from 41 tomograms (reconstructed using fiducial alignment and weighted backprojection, IMOD software, Kremer et al. 1996) were aligned and averaged using the PEET software (bio3d.colorado.edu, Nicastro et al. 2006). To obtain structures with consistent conformations, classification of the different conformational states of dynein was performed using a clustering approach implemented in PEET (Heumann et al. 2011). Number subtomograms used: 2800 |
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Final 3D classification | Number classes: 12 |