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Yorodumi- EMDB-6188: High-resolution structures of kinesin on microtubules provide a b... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-6188 | |||||||||
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Title | High-resolution structures of kinesin on microtubules provide a basis for nucleotide-gated force generation | |||||||||
Map data | Microtubule decorated with monomeric human kinesin (K349 construct) having ADP aluminum fluoride complex bound in the nucleotide pocket. | |||||||||
Sample |
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Function / homology | Function and homology information cytoplasm organization / cytolytic granule membrane / plus-end-directed vesicle transport along microtubule / mitocytosis / anterograde dendritic transport of neurotransmitter receptor complex / anterograde neuronal dense core vesicle transport / anterograde axonal protein transport / retrograde neuronal dense core vesicle transport / vesicle transport along microtubule / lysosome localization ...cytoplasm organization / cytolytic granule membrane / plus-end-directed vesicle transport along microtubule / mitocytosis / anterograde dendritic transport of neurotransmitter receptor complex / anterograde neuronal dense core vesicle transport / anterograde axonal protein transport / retrograde neuronal dense core vesicle transport / vesicle transport along microtubule / lysosome localization / positive regulation of potassium ion transport / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Hedgehog 'off' state / Cilium Assembly / Intraflagellar transport / COPI-dependent Golgi-to-ER retrograde traffic / Carboxyterminal post-translational modifications of tubulin / RHOH GTPase cycle / Sealing of the nuclear envelope (NE) by ESCRT-III / Kinesins / PKR-mediated signaling / The role of GTSE1 in G2/M progression after G2 checkpoint / Aggrephagy / Resolution of Sister Chromatid Cohesion / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / Separation of Sister Chromatids / Kinesins / plus-end-directed microtubule motor activity / RHO GTPases activate KTN1 / RHO GTPases activate IQGAPs / RHO GTPases Activate Formins / stress granule disassembly / Recruitment of NuMA to mitotic centrosomes / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / MHC class II antigen presentation / mitochondrion transport along microtubule / COPI-mediated anterograde transport / centrosome localization / COPI-dependent Golgi-to-ER retrograde traffic / microtubule motor activity / ciliary rootlet / natural killer cell mediated cytotoxicity / kinesin complex / synaptic vesicle transport / Insulin processing / microtubule-based movement / microtubule-based process / centriolar satellite / axon cytoplasm / phagocytic vesicle / MHC class II antigen presentation / dendrite cytoplasm / positive regulation of protein localization to plasma membrane / regulation of membrane potential / axon guidance / positive regulation of synaptic transmission, GABAergic / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / structural constituent of cytoskeleton / cellular response to type II interferon / microtubule cytoskeleton organization / microtubule cytoskeleton / Signaling by ALK fusions and activated point mutants / mitotic cell cycle / microtubule binding / vesicle / microtubule / cadherin binding / GTPase activity / protein-containing complex binding / GTP binding / perinuclear region of cytoplasm / ATP hydrolysis activity / mitochondrion / ATP binding / identical protein binding / membrane / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) / Sus scrofa (pig) | |||||||||
Method | helical reconstruction / cryo EM / Resolution: 5.0 Å | |||||||||
Authors | Shang ZG / Zhou KF / Xu C / Csencsits R / Cochran JC / Sindelar CV | |||||||||
Citation | Journal: Elife / Year: 2014 Title: High-resolution structures of kinesin on microtubules provide a basis for nucleotide-gated force-generation. Authors: Zhiguo Shang / Kaifeng Zhou / Chen Xu / Roseann Csencsits / Jared C Cochran / Charles V Sindelar / Abstract: Microtubule-based transport by the kinesin motors, powered by ATP hydrolysis, is essential for a wide range of vital processes in eukaryotes. We obtained insight into this process by developing ...Microtubule-based transport by the kinesin motors, powered by ATP hydrolysis, is essential for a wide range of vital processes in eukaryotes. We obtained insight into this process by developing atomic models for no-nucleotide and ATP states of the monomeric kinesin motor domain on microtubules from cryo-EM reconstructions at 5-6 Å resolution. By comparing these models with existing X-ray structures of ADP-bound kinesin, we infer a mechanistic scheme in which microtubule attachment, mediated by a universally conserved 'linchpin' residue in kinesin (N255), triggers a clamshell opening of the nucleotide cleft and accompanying release of ADP. Binding of ATP re-closes the cleft in a manner that tightly couples to translocation of cargo, via kinesin's 'neck linker' element. These structural transitions are reminiscent of the analogous nucleotide-exchange steps in the myosin and F1-ATPase motors and inform how the two heads of a kinesin dimer 'gate' each other to promote coordinated stepping along microtubules. | |||||||||
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_6188.map.gz | 12.1 MB | EMDB map data format | |
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Header (meta data) | emd-6188-v30.xml emd-6188.xml | 12.5 KB 12.5 KB | Display Display | EMDB header |
Images | 400_6188.gif 80_6188.gif | 86.1 KB 5.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-6188 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-6188 | HTTPS FTP |
-Validation report
Summary document | emd_6188_validation.pdf.gz | 330.1 KB | Display | EMDB validaton report |
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Full document | emd_6188_full_validation.pdf.gz | 329.7 KB | Display | |
Data in XML | emd_6188_validation.xml.gz | 4.2 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6188 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6188 | HTTPS FTP |
-Related structure data
Related structure data | 3j8yMC 6187C 3j8xC 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_6188.map.gz / Format: CCP4 / Size: 34.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Microtubule decorated with monomeric human kinesin (K349 construct) having ADP aluminum fluoride complex bound in the nucleotide pocket. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.097 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 : Microtubule decorated with monomeric human kinesin (K349 construc...
Entire | Name: Microtubule decorated with monomeric human kinesin (K349 construct) having ADP aluminum fluoride complex bound in the nucleotide pocket. |
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Components |
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-Supramolecule #1000: Microtubule decorated with monomeric human kinesin (K349 construc...
Supramolecule | Name: Microtubule decorated with monomeric human kinesin (K349 construct) having ADP aluminum fluoride complex bound in the nucleotide pocket. type: sample / ID: 1000 / Details: Microtubule decorated with monomeric human kinesin Oligomeric state: One monomer of kinesin binds to one heterodimer of tubulin Number unique components: 2 |
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Molecular weight | Theoretical: 135 KDa |
-Macromolecule #1: monomeric kinesin-1A
Macromolecule | Name: monomeric kinesin-1A / type: protein_or_peptide / ID: 1 / Details: K349 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: human |
Molecular weight | Theoretical: 38 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant strain: BL21(DE3) / Recombinant plasmid: pHB40p |
Sequence | UniProtKB: Kinesin-1 heavy chain |
-Macromolecule #2: tubulin
Macromolecule | Name: tubulin / type: protein_or_peptide / ID: 2 / Oligomeric state: heterodimer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Sus scrofa (pig) / synonym: pig |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | helical reconstruction |
Aggregation state | filament |
-Sample preparation
Buffer | pH: 6.8 / Details: 25 mM PIPES, 25 mM NaCl, 2 mM MgCl2, 1 mM EGTA |
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Grid | Details: 300 mesh copper grid with homemade holey carbon |
Vitrification | Cryogen name: ETHANE / Instrument: HOMEMADE PLUNGER Method: No glow discharged applied; after sample application to grid, liquid was mostly 'wicked' away by edgewise application of filter paper. Subsequently, blotting and plunge freezing were ...Method: No glow discharged applied; after sample application to grid, liquid was mostly 'wicked' away by edgewise application of filter paper. Subsequently, blotting and plunge freezing were performed with a ~0.5 second delay after blotting but prior to plunging. |
-Electron microscopy
Microscope | FEI TITAN |
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Details | 4K x 4K counting mode was used; 24 frames total were collected. |
Date | Jun 2, 2013 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 51 / Average electron dose: 15 e/Å2 Details: Each image was collected as a stack of 24 movie frames. |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 23859.4 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
-Image processing
Details | Initial alignment was done using customized SPIDER scripts. Reconstruction and subsequent refinement were done by FREALIGN. |
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Final reconstruction | Applied symmetry - Helical parameters - Δz: 9.455 Å Applied symmetry - Helical parameters - Δ&Phi: 25.71 ° Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric) Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 5.0 Å / Resolution method: OTHER / Software - Name: SPIDER, FREALIGN Details: Approximately 33,600 asymmetric units were averaged in the final reconstruction. |
CTF correction | Details: done within FREALIGN |
-Atomic model buiding 1
Initial model | PDB ID: Chain - #0 - Chain ID: K / Chain - #1 - Chain ID: A / Chain - #2 - Chain ID: B |
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Software | Name: MDFF |
Details | MDFF was performed using explicit solvation. Side chains were removed from the MDFF target potential. Following several equilibration steps, the relative strength of the EM map potential (GSCALE term) was slowly increased from 0 to 1 over the course of 10 nanoseconds. The t = 1.2 ns time point was selected to represent the final fitted model, based on the approximate convergence of the RMSD from the starting structure. |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT Target criteria: RMSD from the starting structure was monitored for convergence |
Output model | PDB-3j8y: |