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Yorodumi- EMDB-2769: Conserved mechanisms of microtubule-stimulated ADP release, ATP b... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-2769 | |||||||||
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Title | Conserved mechanisms of microtubule-stimulated ADP release, ATP binding, and force generation in transport kinesins | |||||||||
Map data | 13-protofilament microtubule-bound human kinesin-1 motor domain in absence of nucleotides | |||||||||
Sample |
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Keywords | kinesin / microtubule / cryo-EM / cryo-electron microscopy | |||||||||
Function / homology | Function and homology information anterograde dendritic transport of neurotransmitter receptor complex / anterograde axonal protein transport / retrograde neuronal dense core vesicle transport / Kinesins / plus-end-directed microtubule motor activity / RHO GTPases activate KTN1 / positive regulation of axon guidance / COPI-dependent Golgi-to-ER retrograde traffic / ciliary rootlet / kinesin complex ...anterograde dendritic transport of neurotransmitter receptor complex / anterograde axonal protein transport / retrograde neuronal dense core vesicle transport / Kinesins / plus-end-directed microtubule motor activity / RHO GTPases activate KTN1 / positive regulation of axon guidance / COPI-dependent Golgi-to-ER retrograde traffic / ciliary rootlet / kinesin complex / synaptic vesicle transport / Insulin processing / microtubule-based movement / kinesin binding / cytoskeletal motor activity / microtubule-based process / vesicle-mediated transport / axon cytoplasm / MHC class II antigen presentation / dendrite cytoplasm / axon guidance / structural constituent of cytoskeleton / microtubule cytoskeleton organization / microtubule cytoskeleton / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / mitotic cell cycle / nervous system development / microtubule binding / chemical synaptic transmission / perikaryon / microtubule / protein heterodimerization activity / GTPase activity / synapse / GTP binding / perinuclear region of cytoplasm / ATP hydrolysis activity / ATP binding / membrane / metal ion binding / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Bos taurus (cattle) / Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 7.4 Å | |||||||||
Authors | Atherton J / Farabella I / Yu IM / Rosenfeld SS / Houdusse A / Topf M / Moores C | |||||||||
Citation | Journal: Elife / Year: 2014 Title: Conserved mechanisms of microtubule-stimulated ADP release, ATP binding, and force generation in transport kinesins. Authors: Joseph Atherton / Irene Farabella / I-Mei Yu / Steven S Rosenfeld / Anne Houdusse / Maya Topf / Carolyn A Moores / Abstract: Kinesins are a superfamily of microtubule-based ATP-powered motors, important for multiple, essential cellular functions. How microtubule binding stimulates their ATPase and controls force generation ...Kinesins are a superfamily of microtubule-based ATP-powered motors, important for multiple, essential cellular functions. How microtubule binding stimulates their ATPase and controls force generation is not understood. To address this fundamental question, we visualized microtubule-bound kinesin-1 and kinesin-3 motor domains at multiple steps in their ATPase cycles--including their nucleotide-free states--at ∼ 7 Å resolution using cryo-electron microscopy. In both motors, microtubule binding promotes ordered conformations of conserved loops that stimulate ADP release, enhance microtubule affinity and prime the catalytic site for ATP binding. ATP binding causes only small shifts of these nucleotide-coordinating loops but induces large conformational changes elsewhere that allow force generation and neck linker docking towards the microtubule plus end. Family-specific differences across the kinesin-microtubule interface account for the distinctive properties of each motor. Our data thus provide evidence for a conserved ATP-driven mechanism for kinesins and reveal the critical mechanistic contribution of the microtubule interface. | |||||||||
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_2769.map.gz | 204.8 MB | EMDB map data format | |
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Header (meta data) | emd-2769-v30.xml emd-2769.xml | 13.1 KB 13.1 KB | Display Display | EMDB header |
Images | emd_2769.jpg | 140.6 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-2769 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-2769 | HTTPS FTP |
-Validation report
Summary document | emd_2769_validation.pdf.gz | 303 KB | Display | EMDB validaton report |
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Full document | emd_2769_full_validation.pdf.gz | 302.1 KB | Display | |
Data in XML | emd_2769_validation.xml.gz | 6.9 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2769 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2769 | HTTPS FTP |
-Related structure data
Related structure data | 4uxtMC 2765C 2766C 2767C 2768C 2770C 2771C 4uxoC 4uxpC 4uxrC 4uxsC 4uxyC 4uy0C 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_2769.map.gz / Format: CCP4 / Size: 238.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | 13-protofilament microtubule-bound human kinesin-1 motor domain in absence of nucleotides | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.5 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 : 13-protofilament microtubule-bound human kinesin-1 motor domain i...
Entire | Name: 13-protofilament microtubule-bound human kinesin-1 motor domain in absence of nucleotides |
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Components |
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-Supramolecule #1000: 13-protofilament microtubule-bound human kinesin-1 motor domain i...
Supramolecule | Name: 13-protofilament microtubule-bound human kinesin-1 motor domain in absence of nucleotides type: sample / ID: 1000 Oligomeric state: A kinesin motor domain binds to each alpha-beta tubulin heterodimer Number unique components: 3 |
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-Macromolecule #1: alpha tubulin
Macromolecule | Name: alpha tubulin / type: protein_or_peptide / ID: 1 / Oligomeric state: heterodimer / Recombinant expression: No |
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Source (natural) | Organism: Bos taurus (cattle) / synonym: Bovine |
Sequence | InterPro: Alpha tubulin |
-Macromolecule #2: beta tubulin
Macromolecule | Name: beta tubulin / type: protein_or_peptide / ID: 2 / Oligomeric state: heterodimer / Recombinant expression: No |
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Source (natural) | Organism: Bos taurus (cattle) / synonym: Bovine |
Sequence | InterPro: Beta tubulin, autoregulation binding site |
-Macromolecule #3: Kinesin-1 motor domain
Macromolecule | Name: Kinesin-1 motor domain / type: protein_or_peptide / ID: 3 / Name.synonym: Kif5A, Kin1 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant plasmid: pET151-D-TOPO |
Sequence | InterPro: Kinesin motor domain, conserved site |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 6.8 Details: 20mM PIPES, 2mM MgCl2, 1mM EGTA, 2mM DTT, 10 U/mL apyrase |
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Grid | Details: 400 mesh holey carbon grids, air glow discharged |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK I Method: Chamber at 24 degrees C, add microtubule droplet, blot 0.5 sec, add kinesin motor domain droplet, blot 3.5s. |
-Electron microscopy
Microscope | FEI POLARA 300 |
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Temperature | Average: 90 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 150000 times magnification |
Date | Dec 10, 2012 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Digitization - Sampling interval: 15 µm / Number real images: 497 / Average electron dose: 20 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 100000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.3 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 0.4 µm |
Sample stage | Specimen holder model: OTHER |
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |
-Image processing
Details | The particles were selected interactively at the computer terminal |
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CTF correction | Details: Frealign |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 7.4 Å / Resolution method: OTHER / Software - Name: Spider, Frealign Details: Pseudo-symmetry was utilised to generate the asymmetric unit at improved resolution Number images used: 168974 |
-Atomic model buiding 1
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: Chimera, Flex-EM |
Details | Initial model based on multiple PDBs rigid fitted in Chimera, then flexible fitting performed in Flex-EM (Topf et al., 2008) |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT / Target criteria: cross correlation |
Output model | PDB-4uxt: |
-Atomic model buiding 2
Initial model | PDB ID: Chain - Chain ID: K |
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Software | Name: Chimera, Flex-EM |
Details | Initial model based on multiple PDBs rigid fitted in Chimera, then flexible fitting performed in Flex-EM (Topf et al., 2008) |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT / Target criteria: cross correlation |
Output model | PDB-4uxt: |
-Atomic model buiding 3
Initial model | PDB ID: Chain - Chain ID: C |
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Software | Name: Chimera, Flex-EM |
Details | Initial model based on multiple PDBs rigid fitted in Chimera, then flexible fitting performed in Flex-EM (Topf et al., 2008) |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT / Target criteria: cross correlation |
Output model | PDB-4uxt: |