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- EMDB-8547: Kinesin-1 Binding Microtubules with ADP-AlFx in a One-head-bound ... -

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Basic information

Entry
Database: EMDB / ID: EMD-8547
TitleKinesin-1 Binding Microtubules with ADP-AlFx in a One-head-bound State (low-pass filtered to 8.5 angstrom)
Map data
Sample
  • Complex: Kinesin-1 (1-420) on microtubules with ADP-AlFx in a one-head-bound state
    • Protein or peptide: Kinesin-1Kinesin
    • Protein or peptide: Alpha-tubulin
    • Protein or peptide: Beta-tubulinTubulin
Biological speciesBos taurus (cattle) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.7 Å
AuthorsLiu D / Liu X / Shang Z / Sindelar CV
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01 GM 110530-01 United States
American Cancer SocietyACS- IRG 58-012-55 United States
CitationJournal: Elife / Year: 2017
Title: Structural basis of cooperativity in kinesin revealed by 3D reconstruction of a two-head-bound state on microtubules.
Authors: Daifei Liu / Xueqi Liu / Zhiguo Shang / Charles V Sindelar /
Abstract: The detailed basis of walking by dimeric molecules of kinesin along microtubules has remained unclear, partly because available structural methods have been unable to capture microtubule-bound ...The detailed basis of walking by dimeric molecules of kinesin along microtubules has remained unclear, partly because available structural methods have been unable to capture microtubule-bound intermediates of this process. Utilizing novel electron cryomicroscopy methods, we solved structures of microtubule-attached, dimeric kinesin bound to an ATP analog. We find that under these conditions, the kinesin dimer can attach to the microtubule with either one or two motor domains, and we present sub-nanometer resolution reconstructions of both states. The former structure reveals a novel kinesin conformation that revises the current understanding of how ATP binding is coupled to forward stepping of the motor. The latter structure indicates how tension between the two motor domains keeps their cycles out of phase in order to stimulate directional motility. The methods presented here pave the way for future structural studies of a variety of challenging macromolecules that bind to microtubules and other filaments.
History
DepositionJan 12, 2017-
Header (metadata) releaseJan 18, 2017-
Map releaseJan 25, 2017-
UpdateJan 29, 2020-
Current statusJan 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.046
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.046
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.046
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_8547.png

Downloads & links

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Map

FileDownload / File: emd_8547.map.gz / Format: CCP4 / Size: 163.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.06 Å/pix.
x 350 pix.
= 721. Å		2.06 Å/pix.
x 350 pix.
= 721. Å		2.06 Å/pix.
x 350 pix.
= 721. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.06 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.046 / Movie #1: 0.046
Minimum - Maximum-0.024415813 - 0.1052176
Average (Standard dev.)-0.0010091908 (±0.011948339)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-174-174-174
Dimensions350350350
Spacing350350350
CellA=B=C: 721.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.062.062.06
M x/y/z350350350
origin x/y/z0.0000.0000.000
length x/y/z721.000721.000721.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-38-19-20
NX/NY/NZ858082
MAP C/R/S123
start NC/NR/NS-174-174-174
NC/NR/NS350350350
D min/max/mean-0.0240.105-0.001

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Supplemental data

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Half map: #1

Fileemd_8547_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_8547_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Kinesin-1 (1-420) on microtubules with ADP-AlFx in a one-head-bou...

EntireName: Kinesin-1 (1-420) on microtubules with ADP-AlFx in a one-head-bound state
Components
  • Complex: Kinesin-1 (1-420) on microtubules with ADP-AlFx in a one-head-bound state
    • Protein or peptide: Kinesin-1Kinesin
    • Protein or peptide: Alpha-tubulin
    • Protein or peptide: Beta-tubulinTubulin

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Supramolecule #1: Kinesin-1 (1-420) on microtubules with ADP-AlFx in a one-head-bou...

SupramoleculeName: Kinesin-1 (1-420) on microtubules with ADP-AlFx in a one-head-bound state
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Bos taurus (cattle)

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Macromolecule #1: Kinesin-1

MacromoleculeName: Kinesin-1 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MADLAESNIK VMSRFRPLNE SEVNRGDKYI AKFQGEDTVV IASKPYAFDR VFQSSTSQEQ VYNDSAKKIV KDVLEGYNGT IFAYGQTSSG KTHTMEGKLH DPEGMGIIPR IVQDIFNYIY SMDENLEFHI KVSYFEIYLD KIRDLLDVSK TNLSVHEDKN RVPYVKGSTE ...String:
MADLAESNIK VMSRFRPLNE SEVNRGDKYI AKFQGEDTVV IASKPYAFDR VFQSSTSQEQ VYNDSAKKIV KDVLEGYNGT IFAYGQTSSG KTHTMEGKLH DPEGMGIIPR IVQDIFNYIY SMDENLEFHI KVSYFEIYLD KIRDLLDVSK TNLSVHEDKN RVPYVKGSTE RFVSSPDEVM DTIDEGKSNR HVAVTNMNEH SSRSHSIFLI NVKQENTQTE QKLSGKLYLV DLAGSEKVSK TGAEGAVLDE AKNINKSLSA LGNVISALAE GSTYVPYRDS KMTRILQDSL GGNSRTTIVI SSSPSSYNES ETKSTLLFGQ RAKTIKNTVS VNVELTAEQW KKKYEKEKEK NKILRNTIQW LENELNRWRN GETVPIDEQF DKEKANLEAF TVDKDITLTN DKPATAIGVI GNFTDAERRK

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Macromolecule #2: Alpha-tubulin

MacromoleculeName: Alpha-tubulin / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle)
SequenceString: MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIGGGDDSFN TFFSETGAGK HVPRAVFVDL EPTVIDEVRT GTYRQLFHPE QLITGKEDAA NNYARGHYTI GKEIIDLVLD RIRKLADQCT GLQGFLVFHS FGGGTGSGFT SLLMERLSVD YGKKSKLEFS ...String:
MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIGGGDDSFN TFFSETGAGK HVPRAVFVDL EPTVIDEVRT GTYRQLFHPE QLITGKEDAA NNYARGHYTI GKEIIDLVLD RIRKLADQCT GLQGFLVFHS FGGGTGSGFT SLLMERLSVD YGKKSKLEFS IYPAPQVSTA VVEPYNSILT THTTLEHSDC AFMVDNEAIY DICRRNLDIE RPTYTNLNRL ISQIVSSITA SLRFDGALNV DLTEFQTNLV PYPRIHFPLA TYAPVISAEK AYHEQLSVAE ITNACFEPAN QMVKCDPRHG KYMACCLLYR GDVVPKDVNA AIATIKTKRS IQFVDWCPTG FKVGINYQPP TVVPGGDLAK VQRAVCMLSN TTAIAEAWAR LDHKFDLMYA KRAFVHWYVG EGMEEGEFSE AREDMAALEK DYEEVGVDSV EGEGEEEGEE Y

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Macromolecule #3: Beta-tubulin

MacromoleculeName: Beta-tubulin / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle)
SequenceString: MREIVHLQAG QCGNQIGAKF WEVISDEHGI DPTGTYHGDS DLQLERINVY YNEATGGNYV PRAVLVDLEP GTMDSVRSGP FGQIFRPDNF VFGQSGAGNN WAKGHYTEGA ELVDAVLDVV RKEAESCDCL QGFQLTHSLG GGTGSGMGTL LISKIREEFP DRIMNTFSVV ...String:
MREIVHLQAG QCGNQIGAKF WEVISDEHGI DPTGTYHGDS DLQLERINVY YNEATGGNYV PRAVLVDLEP GTMDSVRSGP FGQIFRPDNF VFGQSGAGNN WAKGHYTEGA ELVDAVLDVV RKEAESCDCL QGFQLTHSLG GGTGSGMGTL LISKIREEFP DRIMNTFSVV PSPKVSDTVV EPYNATLSVH QLVENTDETY CIDNEALYDI CFRTLKLTTP TYGDLNHLVS ATMSGVTTCL RFPGQLNADL RKLAVNMVPF PRLHFFMPGF APLTSRGSQQ YRALTVPELT QQMFDAKNMM AACDPRHGRY LTVAAVFRGR MSMKEVDEQM LSVQSKNSSY FVEWIPNNVK TAVCDIPPRG LKMAATFIGN STAIQELFKR ISEQFTAMFR RKAFLHWYTG EGMDEMEFTE AESNMNDLVS EYQQYQDATA EEGEFEEEAE EEVA

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 6.8
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Average electron dose: 1.65 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionResolution.type: BY AUTHOR / Resolution: 6.7 Å / Resolution method: FSC 0.143 CUT-OFF
Details: a modified version of frealign was used for reconstruction, kinesin dimer was masked before FSC calculation using half maps.
Number images used: 44298

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