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- EMDB-10421: Human kinesin-5 motor domain in the GSK-1 state bound to microtubules -

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Entry
Database: EMDB / ID: EMD-10421
TitleHuman kinesin-5 motor domain in the GSK-1 state bound to microtubules
Map dataHuman kinesin-5 motor domain bound to microtubules and the drug GSK-1
Sample
  • Complex: Ternary complex of Human Kinesin 5 and microtubules
    • Complex: Tubulin alpha-1B chain and Tubulin beta chain
      • Protein or peptide: Tubulin beta chain
      • Protein or peptide: Tubulin alpha-1B chain
    • Complex: Kinesin-like protein KIF11
      • Protein or peptide: Kinesin-like protein KIF11
  • Ligand: PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER
  • Ligand: MAGNESIUM ION
  • Ligand: 6-[4-(trifluoromethyl)phenyl]-3,4-dihydro-1~{H}-quinolin-2-one
Keywordskinesin / microtubule / mitosis / inhibition / motor / CELL CYCLE
Function / homology
Function and homology information


spindle elongation / regulation of mitotic centrosome separation / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Resolution of Sister Chromatid Cohesion / Hedgehog 'off' state / Cilium Assembly / Intraflagellar transport / COPI-dependent Golgi-to-ER retrograde traffic / Mitotic Prometaphase / Carboxyterminal post-translational modifications of tubulin ...spindle elongation / regulation of mitotic centrosome separation / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Resolution of Sister Chromatid Cohesion / Hedgehog 'off' state / Cilium Assembly / Intraflagellar transport / COPI-dependent Golgi-to-ER retrograde traffic / Mitotic Prometaphase / Carboxyterminal post-translational modifications of tubulin / RHOH GTPase cycle / EML4 and NUDC in mitotic spindle formation / Sealing of the nuclear envelope (NE) by ESCRT-III / Kinesins / PKR-mediated signaling / Separation of Sister Chromatids / The role of GTSE1 in G2/M progression after G2 checkpoint / Aggrephagy / plus-end-directed microtubule motor activity / mitotic centrosome separation / Kinesins / RHO GTPases activate IQGAPs / RHO GTPases Activate Formins / kinesin complex / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / MHC class II antigen presentation / Recruitment of NuMA to mitotic centrosomes / microtubule motor activity / COPI-mediated anterograde transport / spindle organization / COPI-dependent Golgi-to-ER retrograde traffic / microtubule-based movement / mitotic spindle assembly / MHC class II antigen presentation / mitotic spindle organization / structural constituent of cytoskeleton / microtubule cytoskeleton organization / spindle / neuron migration / spindle pole / mitotic spindle / mitotic cell cycle / microtubule cytoskeleton / microtubule binding / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / microtubule / ciliary basal body / cell division / GTPase activity / intracellular membrane-bounded organelle / protein kinase binding / GTP binding / protein-containing complex / ATP binding / metal ion binding / nucleus / membrane / cytosol / cytoplasm
Similarity search - Function
Kinesin-associated microtubule-binding domain / Kinesin-associated microtubule-binding / : / : / Kinesin motor domain signature. / Kinesin motor domain, conserved site / Kinesin motor domain / Kinesin motor domain profile. / Kinesin motor, catalytic domain. ATPase. / Kinesin motor domain ...Kinesin-associated microtubule-binding domain / Kinesin-associated microtubule-binding / : / : / Kinesin motor domain signature. / Kinesin motor domain, conserved site / Kinesin motor domain / Kinesin motor domain profile. / Kinesin motor, catalytic domain. ATPase. / Kinesin motor domain / Kinesin motor domain superfamily / Tubulin-beta mRNA autoregulation signal. / Alpha tubulin / Beta tubulin, autoregulation binding site / Beta tubulin / Tubulin / Tubulin, C-terminal / Tubulin C-terminal domain / Tubulin, conserved site / Tubulin subunits alpha, beta, and gamma signature. / Tubulin/FtsZ family, C-terminal domain / Tubulin/FtsZ-like, C-terminal domain / Tubulin/FtsZ, C-terminal / Tubulin/FtsZ, 2-layer sandwich domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ, GTPase domain / Tubulin/FtsZ, GTPase domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Tubulin beta chain / Kinesin-like protein KIF11 / Tubulin alpha-1B chain
Similarity search - Component
Biological speciesSus scrofa (pig) / Homo sapiens (human)
Methodhelical reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsPena A / Sweeney A
Funding support United Kingdom, 3 items
OrganizationGrant numberCountry
Wellcome Trust202679/Z/16/Z and 206166/Z/17/Z United Kingdom
Medical Research Council (United Kingdom)MR/J003867/1 United Kingdom
Medical Research Council (United Kingdom)MR/N013867/1 United Kingdom
Citation
Journal: Structure / Year: 2020
Title: Structure of Microtubule-Trapped Human Kinesin-5 and Its Mechanism of Inhibition Revealed Using Cryoelectron Microscopy.
Authors: Alejandro Peña / Aaron Sweeney / Alexander D Cook / Julia Locke / Maya Topf / Carolyn A Moores /
Abstract: Kinesin-5 motors are vital mitotic spindle components, and disruption of their function perturbs cell division. We investigated the molecular mechanism of the human kinesin-5 inhibitor GSK-1, which ...Kinesin-5 motors are vital mitotic spindle components, and disruption of their function perturbs cell division. We investigated the molecular mechanism of the human kinesin-5 inhibitor GSK-1, which allosterically promotes tight microtubule binding. GSK-1 inhibits monomeric human kinesin-5 ATPase and microtubule gliding activities, and promotes the motor's microtubule stabilization activity. Using cryoelectron microscopy, we determined the 3D structure of the microtubule-bound motor-GSK-1 at 3.8 Å overall resolution. The structure reveals that GSK-1 stabilizes the microtubule binding surface of the motor in an ATP-like conformation, while destabilizing regions of the motor around the empty nucleotide binding pocket. Density corresponding to GSK-1 is located between helix-α4 and helix-α6 in the motor domain at its interface with the microtubule. Using a combination of difference mapping and protein-ligand docking, we characterized the kinesin-5-GSK-1 interaction and further validated this binding site using mutagenesis. This work opens up new avenues of investigation of kinesin inhibition and spindle perturbation.
#1: Journal: To Be Published
Title: Mechanism of microtubule-trapped human kinesin-5 inhibition revealed using cryo-EM
Authors: Pena AP / Sweeney A
History
DepositionOct 29, 2019-
Header (metadata) releaseDec 4, 2019-
Map releaseApr 22, 2020-
UpdateApr 9, 2025-
Current statusApr 9, 2025Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.005
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  • Surface view colored by height
  • Surface level: 0.005
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  • Surface view with fitted model
  • Atomic models: PDB-6ta3
  • Surface level: 0.005
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  • Surface view with fitted model
  • Atomic models: PDB-6tiw
  • Surface level: 0.005
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6ta3
  • Imaged by Jmol
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6tiw
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10421.png

Downloads & links

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Map

FileDownload / File: emd_10421.map.gz / Format: CCP4 / Size: 506 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationHuman kinesin-5 motor domain bound to microtubules and the drug GSK-1
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.09 Å/pix.
x 510 pix.
= 555.9 Å		1.09 Å/pix.
x 510 pix.
= 555.9 Å		1.09 Å/pix.
x 510 pix.
= 555.9 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.09 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.005 / Movie #1: 0.005
Minimum - Maximum0.0 - 0.021588022
Average (Standard dev.)0.00001147205 (±0.00027406844)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions510510510
Spacing510510510
CellA=B=C: 555.9 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.091.091.09
M x/y/z510510510
origin x/y/z0.0000.0000.000
length x/y/z555.900555.900555.900
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS510510510
D min/max/mean0.0000.0220.000

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Supplemental data

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Sample components

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Entire : Ternary complex of Human Kinesin 5 and microtubules

EntireName: Ternary complex of Human Kinesin 5 and microtubules
Components
  • Complex: Ternary complex of Human Kinesin 5 and microtubules
    • Complex: Tubulin alpha-1B chain and Tubulin beta chain
      • Protein or peptide: Tubulin beta chain
      • Protein or peptide: Tubulin alpha-1B chain
    • Complex: Kinesin-like protein KIF11
      • Protein or peptide: Kinesin-like protein KIF11
  • Ligand: PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER
  • Ligand: MAGNESIUM ION
  • Ligand: 6-[4-(trifluoromethyl)phenyl]-3,4-dihydro-1~{H}-quinolin-2-one

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Supramolecule #1: Ternary complex of Human Kinesin 5 and microtubules

SupramoleculeName: Ternary complex of Human Kinesin 5 and microtubules / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3

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Supramolecule #2: Tubulin alpha-1B chain and Tubulin beta chain

SupramoleculeName: Tubulin alpha-1B chain and Tubulin beta chain / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#2
Source (natural)Organism: Sus scrofa (pig)

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Supramolecule #3: Kinesin-like protein KIF11

SupramoleculeName: Kinesin-like protein KIF11 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #3
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Tubulin beta chain

MacromoleculeName: Tubulin beta chain / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Sus scrofa (pig)
Molecular weightTheoretical: 48.113129 KDa
SequenceString: MREIVHIQAG QCGNQIGAKF WEVISDEHGI DPTGSYHGDS DLQLERINVY YNEAAGNKYV PRAILVDLEP GTMDSVRSGP FGQIFRPDN FVFGQSGAGN NWAKGHYTEG AELVDSVLDV VRKESESCDC LQGFQLTHSL GGGTGSGMGT LLISKIREEY P DRIMNTFS ...String:
MREIVHIQAG QCGNQIGAKF WEVISDEHGI DPTGSYHGDS DLQLERINVY YNEAAGNKYV PRAILVDLEP GTMDSVRSGP FGQIFRPDN FVFGQSGAGN NWAKGHYTEG AELVDSVLDV VRKESESCDC LQGFQLTHSL GGGTGSGMGT LLISKIREEY P DRIMNTFS VVPSPKVSDT VVEPYNATLS VHQLVENTDE TYCIDNEALY DICFRTLKLT TPTYGDLNHL VSATMSGVTT CL RFPGQLN ADLRKLAVNM VPFPRLHFFM PGFAPLTSRG SQQYRALTVP ELTQQMFDAK NMMAACDPRH GRYLTVAAVF RGR MSMKEV DEQMLNVQNK NSSYFVEWIP NNVKTAVCDI PPRGLKMSAT FIGNSTAIQE LFKRISEQFT AMFRRKAFLH WYTG EGMDE MEFTEAESNM NDLVSEYQQY QDAT

UniProtKB: Tubulin beta chain

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Macromolecule #2: Tubulin alpha-1B chain

MacromoleculeName: Tubulin alpha-1B chain / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Sus scrofa (pig)
Molecular weightTheoretical: 48.780117 KDa
SequenceString: MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIGGGDDSFN TFFSETGAGK HVPRAVFVDL EPTVIDEVRT GTYRQLFHP EQLITGKEDA ANNYARGHYT IGKEIIDLVL DRIRKLADQC TGLQGFLVFH SFGGGTGSGF TSLLMERLSV D YGKKSKLE ...String:
MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIGGGDDSFN TFFSETGAGK HVPRAVFVDL EPTVIDEVRT GTYRQLFHP EQLITGKEDA ANNYARGHYT IGKEIIDLVL DRIRKLADQC TGLQGFLVFH SFGGGTGSGF TSLLMERLSV D YGKKSKLE FSIYPAPQVS TAVVEPYNSI LTTHTTLEHS DCAFMVDNEA IYDICRRNLD IERPTYTNLN RLISQIVSSI TA SLRFDGA LNVDLTEFQT NLVPYPRIHF PLATYAPVIS AEKAYHEQLS VAEITNACFE PANQMVKCDP RHGKYMACCL LYR GDVVPK DVNAAIATIK TKRSIQFVDW CPTGFKVGIN YQPPTVVPGG DLAKVQRAVC MLSNTTAIAE AWARLDHKFD LMYA KRAFV HWYVGEGMEE GEFSEAREDM AALEKDYEEV GVD

UniProtKB: Tubulin alpha-1B chain

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Macromolecule #3: Kinesin-like protein KIF11

MacromoleculeName: Kinesin-like protein KIF11 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 43.727461 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHSSG VDLGTENLYF QSMASQPNSS AKKKEEKGKN IQVVVRCRPF NLAERKASAH SIVECDPVRK EVSVRTGGLA DKSSRKTYT FDMVFGASTK QIDVYRSVVC PILDEVIMGY NCTIFAYGQT GTGKTFTMEG ERSPNEEYTW EEDPLAGIIP R TLHQIFEK ...String:
MHHHHHHSSG VDLGTENLYF QSMASQPNSS AKKKEEKGKN IQVVVRCRPF NLAERKASAH SIVECDPVRK EVSVRTGGLA DKSSRKTYT FDMVFGASTK QIDVYRSVVC PILDEVIMGY NCTIFAYGQT GTGKTFTMEG ERSPNEEYTW EEDPLAGIIP R TLHQIFEK LTDNGTEFSV KVSLLEIYNE ELFDLLNPSS DVSERLQMFD DPRNKRGVII KGLEEITVHN KDEVYQILEK GA AKRTTAA TLMNAYSSRS HSVFSVTIHM KETTIDGEEL VKIGKLNLVD LAGSENIGRS GAVDKRAREA GNINQSLLTL GRV ITALVE RTPHVPYRES KLTRILQDSL GGRTRTSIIA TISPASLNLE ETLSTLEYAH RAKNILNKPE VNQKL

UniProtKB: Kinesin-like protein KIF11

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Macromolecule #4: PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER

MacromoleculeName: PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER / type: ligand / ID: 4 / Number of copies: 2 / Formula: G2P
Molecular weightTheoretical: 521.208 Da
Chemical component information

ChemComp-G2P:
PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER / GMP-CPP, energy-carrying molecule analogue*YM

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Macromolecule #5: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 5 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #6: 6-[4-(trifluoromethyl)phenyl]-3,4-dihydro-1~{H}-quinolin-2-one

MacromoleculeName: 6-[4-(trifluoromethyl)phenyl]-3,4-dihydro-1~{H}-quinolin-2-one
type: ligand / ID: 6 / Number of copies: 1 / Formula: MZK
Molecular weightTheoretical: 291.268 Da
Chemical component information

ChemComp-MZK:
6-[4-(trifluoromethyl)phenyl]-3,4-dihydro-1~{H}-quinolin-2-one

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7.6
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 45.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 8.9 Å
Applied symmetry - Helical parameters - Δ&Phi: -25.7 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.5 CUT-OFF / Number images used: 507219
Startup modelType of model: INSILICO MODEL
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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