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Yorodumi- EMDB-8408: Cryo-EM reconstruction of the yeast kinesin-5, Cin8, bound to GDP... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-8408 | |||||||||
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Title | Cryo-EM reconstruction of the yeast kinesin-5, Cin8, bound to GDP-taxol microtubules in ADP-AlFx state | |||||||||
Map data | 3D helical reconstruction of Cin8 motor domain in ADP-AlFx state bound to microtubule using cryo-EM | |||||||||
Sample |
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Biological species | Saccharomyces cerevisiae (brewer's yeast) | |||||||||
Method | helical reconstruction / cryo EM / Resolution: 7.3 Å | |||||||||
Authors | Cha HK | |||||||||
Citation | Journal: J Biol Chem / Year: 2017 Title: The yeast kinesin-5 Cin8 interacts with the microtubule in a noncanonical manner. Authors: Kayla M Bell / Hyo Keun Cha / Charles V Sindelar / Jared C Cochran / Abstract: Kinesin motors play central roles in establishing and maintaining the mitotic spindle during cell division. Unlike most other kinesins, Cin8, a kinesin-5 motor in can move bidirectionally along ...Kinesin motors play central roles in establishing and maintaining the mitotic spindle during cell division. Unlike most other kinesins, Cin8, a kinesin-5 motor in can move bidirectionally along microtubules, switching directionality according to biochemical conditions, a behavior that remains largely unexplained. To this end, we used biochemical rate and equilibrium constant measurements as well as cryo-electron microscopy methodologies to investigate the microtubule interactions of the Cin8 motor domain. These experiments unexpectedly revealed that, whereas Cin8 ATPase kinetics fell within measured ranges for kinesins (especially kinesin-5 proteins), approximately four motors can bind each αβ-tubulin dimer within the microtubule lattice. This result contrasted with those observations on other known kinesins, which can bind only a single "canonical" site per tubulin dimer. Competition assays with human kinesin-5 (Eg5) only partially abrogated this behavior, indicating that Cin8 binds microtubules not only at the canonical site, but also one or more separate ("noncanonical") sites. Moreover, we found that deleting the large, class-specific insert in the microtubule-binding loop 8 reverts Cin8 to one motor per αβ-tubulin in the microtubule. The novel microtubule-binding mode of Cin8 identified here provides a potential explanation for Cin8 clustering along microtubules and potentially may contribute to the mechanism for direction reversal. | |||||||||
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_8408.map.gz | 9.4 MB | EMDB map data format | |
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Header (meta data) | emd-8408-v30.xml emd-8408.xml | 9.5 KB 9.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_8408_fsc.xml | 11.6 KB | Display | FSC data file |
Images | emd_8408.png | 210.4 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-8408 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-8408 | HTTPS FTP |
-Validation report
Summary document | emd_8408_validation.pdf.gz | 78.9 KB | Display | EMDB validaton report |
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Full document | emd_8408_full_validation.pdf.gz | 78 KB | Display | |
Data in XML | emd_8408_validation.xml.gz | 493 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8408 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8408 | HTTPS FTP |
-Related structure data
Similar structure data |
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-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_8408.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | 3D helical reconstruction of Cin8 motor domain in ADP-AlFx state bound to microtubule using cryo-EM | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.49402 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-bound kinesin-5 Cin8 in ADP-AlFx state
Entire | Name: Microtubule-bound kinesin-5 Cin8 in ADP-AlFx state |
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Components |
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-Supramolecule #1: Microtubule-bound kinesin-5 Cin8 in ADP-AlFx state
Supramolecule | Name: Microtubule-bound kinesin-5 Cin8 in ADP-AlFx state / type: complex / ID: 1 / Parent: 0 Details: Cin8-his monomer (residues 70-535) Taxol-stabilized microtubules |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant cell: B834(DE3) / Recombinant plasmid: pET16b |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | helical reconstruction |
Aggregation state | filament |
-Sample preparation
Buffer | pH: 6.8 Component:
Details: adjusted to pH 6.8 with KOH | |||||||||||||||
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Vitrification | Cryogen name: ETHANE / Instrument: HOMEMADE PLUNGER |
-Electron microscopy
Microscope | FEI TECNAI F20 |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 1.71 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | Model: Tecnai F20 / Image courtesy: FEI Company |