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- EMDB-3623: Microtubule-bound MKLP2 motor domain in the presence of AMPPNP -

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Basic information

Entry
Database: EMDB / ID: EMD-3623
TitleMicrotubule-bound MKLP2 motor domain in the presence of AMPPNP
Map data'Microtubule-bound MKLP2 motor domain with AMPPNP'
Sample
  • Complex: Complex of 13pf microtubule with bound MKLP2 motor domain in the presence of AMPPNP
    • Complex: Kinesin-like protein KIF20A
      • Protein or peptide: Kinesin-like protein KIF20A
    • Complex: Tubulin alpha and beta chains
      • Protein or peptide: Tubulin alpha chain
      • Protein or peptide: Tubulin beta-2B chain
  • Ligand: MAGNESIUM ION
  • Ligand: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER
  • Ligand: GUANOSINE-5'-TRIPHOSPHATE
  • Ligand: GUANOSINE-5'-DIPHOSPHATE
  • Ligand: TAXOL
Function / homology
Function and homology information


Mitotic Telophase/Cytokinesis / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly / Intraflagellar transport / Carboxyterminal post-translational modifications of tubulin / Sealing of the nuclear envelope (NE) by ESCRT-III / Kinesins / Resolution of Sister Chromatid Cohesion / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation ...Mitotic Telophase/Cytokinesis / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly / Intraflagellar transport / Carboxyterminal post-translational modifications of tubulin / Sealing of the nuclear envelope (NE) by ESCRT-III / Kinesins / Resolution of Sister Chromatid Cohesion / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / COPI-dependent Golgi-to-ER retrograde traffic / COPI-independent Golgi-to-ER retrograde traffic / COPI-mediated anterograde transport / RHO GTPases activate IQGAPs / RHO GTPases Activate Formins / Kinesins / MHC class II antigen presentation / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / Aggrephagy / COPI-dependent Golgi-to-ER retrograde traffic / The role of GTSE1 in G2/M progression after G2 checkpoint / Separation of Sister Chromatids / Loss of Nlp from mitotic centrosomes / Recruitment of mitotic centrosome proteins and complexes / Loss of proteins required for interphase microtubule organization from the centrosome / Anchoring of the basal body to the plasma membrane / AURKA Activation by TPX2 / Recruitment of NuMA to mitotic centrosomes / midbody abscission / Regulation of PLK1 Activity at G2/M Transition / Hedgehog 'off' state / MHC class II antigen presentation / microtubule bundle formation / positive regulation of axon guidance / microtubule motor activity / kinesin complex / intercellular bridge / microtubule-based movement / microtubule-based process / mitotic cytokinesis / regulation of cytokinesis / structural constituent of cytoskeleton / microtubule cytoskeleton organization / spindle / microtubule cytoskeleton / protein transport / mitotic cell cycle / nervous system development / midbody / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / microtubule binding / microtubule / hydrolase activity / protein heterodimerization activity / GTPase activity / GTP binding / protein kinase binding / Golgi apparatus / ATP hydrolysis activity / nucleoplasm / ATP binding / nucleus / metal ion binding / cytosol / cytoplasm
Similarity search - Function
Kinesin-like protein / 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 / Alpha tubulin / Tubulin-beta mRNA autoregulation signal. ...Kinesin-like protein / 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 / Alpha tubulin / Tubulin-beta mRNA autoregulation signal. / 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 alpha chain / Tubulin alpha-1B chain / Kinesin-like protein KIF20A / Tubulin beta-2B chain
Similarity search - Component
Biological speciesMus musculus (house mouse) / Bos taurus (cattle) / Bovine (cattle)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.9 Å
AuthorsAtherton J / Yu I-M / Cook A / Muretta JM / Joseph AP / Major J / Sourigues Y / Clause J / Topf M / Rosenfeld SS ...Atherton J / Yu I-M / Cook A / Muretta JM / Joseph AP / Major J / Sourigues Y / Clause J / Topf M / Rosenfeld SS / Houdusse A / Moores CA
CitationJournal: Elife / Year: 2017
Title: The divergent mitotic kinesin MKLP2 exhibits atypical structure and mechanochemistry.
Authors: Joseph Atherton / I-Mei Yu / Alexander Cook / Joseph M Muretta / Agnel Joseph / Jennifer Major / Yannick Sourigues / Jeffrey Clause / Maya Topf / Steven S Rosenfeld / Anne Houdusse / Carolyn A Moores /
Abstract: MKLP2, a kinesin-6, has critical roles during the metaphase-anaphase transition and cytokinesis. Its motor domain contains conserved nucleotide binding motifs, but is divergent in sequence (~35% ...MKLP2, a kinesin-6, has critical roles during the metaphase-anaphase transition and cytokinesis. Its motor domain contains conserved nucleotide binding motifs, but is divergent in sequence (~35% identity) and size (~40% larger) compared to other kinesins. Using cryo-electron microscopy and biophysical assays, we have undertaken a mechanochemical dissection of the microtubule-bound MKLP2 motor domain during its ATPase cycle, and show that many facets of its mechanism are distinct from other kinesins. While the MKLP2 neck-linker is directed towards the microtubule plus-end in an ATP-like state, it does not fully dock along the motor domain. Furthermore, the footprint of the MKLP2 motor domain on the MT surface is altered compared to motile kinesins, and enhanced by kinesin-6-specific sequences. The conformation of the highly extended loop6 insertion characteristic of kinesin-6s is nucleotide-independent and does not contact the MT surface. Our results emphasize the role of family-specific insertions in modulating kinesin motor function.
History
DepositionMar 7, 2017-
Header (metadata) releaseAug 16, 2017-
Map releaseOct 4, 2017-
UpdateOct 4, 2017-
Current statusOct 4, 2017Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0322
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.0322
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5nd7
  • Surface level: 0.0322
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-5nd7
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3623.png

Downloads & links

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Map

FileDownload / File: emd_3623.map.gz / Format: CCP4 / Size: 13.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation'Microtubule-bound MKLP2 motor domain with AMPPNP'
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.54 Å/pix.
x 161 pix.
= 247.94 Å		1.54 Å/pix.
x 139 pix.
= 214.06 Å		1.54 Å/pix.
x 160 pix.
= 246.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 1.54 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0322 / Movie #1: 0.0322
Minimum - Maximum-0.030743273 - 0.08448133
Average (Standard dev.)0.0028416982 (±0.010940165)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin7058123
Dimensions139160161
Spacing160139161
CellA: 246.4 Å / B: 214.06 Å / C: 247.93999 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.541.541.54
M x/y/z160139161
origin x/y/z0.0000.0000.000
length x/y/z246.400214.060247.940
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS5870123
NC/NR/NS160139161
D min/max/mean-0.0310.0840.003

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Supplemental data

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Sample components

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Entire : Complex of 13pf microtubule with bound MKLP2 motor domain in the ...

EntireName: Complex of 13pf microtubule with bound MKLP2 motor domain in the presence of AMPPNP
Components
  • Complex: Complex of 13pf microtubule with bound MKLP2 motor domain in the presence of AMPPNP
    • Complex: Kinesin-like protein KIF20A
      • Protein or peptide: Kinesin-like protein KIF20A
    • Complex: Tubulin alpha and beta chains
      • Protein or peptide: Tubulin alpha chain
      • Protein or peptide: Tubulin beta-2B chain
  • Ligand: MAGNESIUM ION
  • Ligand: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER
  • Ligand: GUANOSINE-5'-TRIPHOSPHATE
  • Ligand: GUANOSINE-5'-DIPHOSPHATE
  • Ligand: TAXOL

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Supramolecule #1: Complex of 13pf microtubule with bound MKLP2 motor domain in the ...

SupramoleculeName: Complex of 13pf microtubule with bound MKLP2 motor domain in the presence of AMPPNP
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 / Details: Microtubule bound to taxol
Source (natural)Organism: Mus musculus (house mouse)

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Supramolecule #2: Kinesin-like protein KIF20A

SupramoleculeName: Kinesin-like protein KIF20A / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1 / Details: Microtubule bound to taxol
Source (natural)Organism: Bos taurus (cattle)

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Supramolecule #3: Tubulin alpha and beta chains

SupramoleculeName: Tubulin alpha and beta chains / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2-#3 / Details: Microtubule bound to taxol

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Macromolecule #1: Kinesin-like protein KIF20A

MacromoleculeName: Kinesin-like protein KIF20A / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 56.021332 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: SPILESTAAD LRSVVRKDLL SDCSVISASL EDKQALLEDT SEKVKVYLRI RPFLTSELDR QEDQGCVCIE NTETLVLQAP KDSFALKSN ERGVGQATHK FTFSQIFGPE VGQVAFFNLT MKEMVKDVLK GQNWLIYTYG VTNSGKTYTI QGTSKDAGIL P QSLALIFN ...String:
SPILESTAAD LRSVVRKDLL SDCSVISASL EDKQALLEDT SEKVKVYLRI RPFLTSELDR QEDQGCVCIE NTETLVLQAP KDSFALKSN ERGVGQATHK FTFSQIFGPE VGQVAFFNLT MKEMVKDVLK GQNWLIYTYG VTNSGKTYTI QGTSKDAGIL P QSLALIFN SLQGQLHPTP DLKPLLSNEV IWLDSKQIRQ EEMKKLSLLI GGLQEEELST SVKKRVHTES RIGASNSFDS GV AGLSSTS QFTSSSQLDE TSQLWAQPDT VPVSVPADIR FSVWISFFEI YNELLYDLLE PPSHQHKRQT LRLCEDQNGN PYV KDLNWI HVRDVEEAWK LLKVGRKNQS FASTHMNQQS SRSHSIFSIR ILHLQGEGDI VPKISELSLC DLAGSERCKH QKSG ERLKE AGNINTSLHT LGRCIAALRQ NQQNRSKQNL IPFRDSKLTR VFQGFFTGRG RSCMIVNVNP CASTYDETLH AAKFS ALAS QLVHAPPVHL GIPSLHS

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Macromolecule #2: Tubulin alpha chain

MacromoleculeName: Tubulin alpha chain / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bovine (cattle)
Molecular weightTheoretical: 50.107238 KDa
SequenceString: MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIGGGDDSFN TFFSETGAGK HVPRAVFVDL EPTVIDEVRT GTYRQLFHP EQLITGKEDA ANNYARGHYT IGKEIIDLVL DRIRKLADQC TGLQGFSVFH SFGGGTGSGF TSLLMERLSV D YGKKSKLE ...String:
MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIGGGDDSFN TFFSETGAGK HVPRAVFVDL EPTVIDEVRT GTYRQLFHP EQLITGKEDA ANNYARGHYT IGKEIIDLVL DRIRKLADQC TGLQGFSVFH SFGGGTGSGF TSLLMERLSV D YGKKSKLE FSIYPAPQVS TAVVEPYNSI LTTHTTLEHS DCAFMVDNEA IYDICRRNLD IERPTYTNLN RLIGQIVSSI TA SLRFDGA LNVDLTEFQT NLVPYPRGHF PLATYAPVIS AEKAYHEQLS VAEITNACFE PANQMVKCDP RHGKYMACCL LYR GDVVPK DVNAAIATIK TKRTIQFVDW CPTGFKVGIN YEPPTVVPGG DLAKVQRAVC MLSNTTAIAE AWARLDHKFD LMYA KRAFV HWYVGEGMEE GEFSEAREDM AALEKDYEEV GVDSVEGEGE EEGEEY

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Macromolecule #3: Tubulin beta-2B chain

MacromoleculeName: Tubulin beta-2B chain / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bovine (cattle)
Molecular weightTheoretical: 49.90777 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 QDATADEQGE FEEEEGEDEA

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #5: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER

MacromoleculeName: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / type: ligand / ID: 5 / Number of copies: 1 / Formula: ANP
Molecular weightTheoretical: 506.196 Da
Chemical component information

ChemComp-ANP:
PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / AMP-PNP, energy-carrying molecule analogue*YM

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Macromolecule #6: GUANOSINE-5'-TRIPHOSPHATE

MacromoleculeName: GUANOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 6 / Number of copies: 1 / Formula: GTP
Molecular weightTheoretical: 523.18 Da
Chemical component information

ChemComp-GTP:
GUANOSINE-5'-TRIPHOSPHATE / GTP, energy-carrying molecule*YM

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Macromolecule #7: GUANOSINE-5'-DIPHOSPHATE

MacromoleculeName: GUANOSINE-5'-DIPHOSPHATE / type: ligand / ID: 7 / Number of copies: 1 / Formula: GDP
Molecular weightTheoretical: 443.201 Da
Chemical component information

ChemComp-GDP:
GUANOSINE-5'-DIPHOSPHATE / GDP, energy-carrying molecule*YM

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Macromolecule #8: TAXOL

MacromoleculeName: TAXOL / type: ligand / ID: 8 / Number of copies: 1 / Formula: TA1
Molecular weightTheoretical: 853.906 Da
Chemical component information

ChemComp-TA1:
TAXOL / medication, chemotherapy*YM

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 6.8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TECNAI F20
Image recordingFilm or detector model: DIRECT ELECTRON DE-20 (5k x 3k) / Detector mode: INTEGRATING / Average electron dose: 20.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 7.9 Å / Resolution method: FSC 0.143 CUT-OFF
Details: Gold-standard FSCtrue using noise substitution test (Chen et al., 2014)
Number images used: 10858
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING

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