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- EMDB-7469: Structure of the DASH/Dam1 complex shows its role at the yeast ki... -

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Basic information

Entry
Database: EMDB / ID: EMD-7469
TitleStructure of the DASH/Dam1 complex shows its role at the yeast kinetochore-microtubule interface (asymmetric reconstruction)
Map dataDASH/Dam1 complex (asymmetric reconstruction)
Sample
  • Complex: DASH/Dam1 complex
    • Protein or peptide: Ask1
    • Protein or peptide: Dad3
    • Protein or peptide: Dad2
    • Protein or peptide: Duo1
    • Protein or peptide: Dad4
    • Protein or peptide: Dad1,Dad1
    • Protein or peptide: Hsk3
    • Protein or peptide: Dam1
    • Protein or peptide: Spc19
    • Protein or peptide: Spc34
KeywordsDASH / Dam1 complex / Ask1 / Dad1 / Dad2 / Dad3 / Dad4 / Dam1 / Duo1 / Hsk3 / Spc19 / Spc34 / kinetochore / kinetochore-microtubule interface / chromosome segregation / cell-division / point-centromere yeast / NUCLEAR PROTEIN
Function / homology
Function and homology information


DASH complex / protein transport along microtubule to mitotic spindle pole body / mitotic sister chromatid biorientation / attachment of spindle microtubules to kinetochore / Ino80 complex / attachment of mitotic spindle microtubules to kinetochore / mitotic spindle organization / chromosome segregation / spindle microtubule / mitotic spindle ...DASH complex / protein transport along microtubule to mitotic spindle pole body / mitotic sister chromatid biorientation / attachment of spindle microtubules to kinetochore / Ino80 complex / attachment of mitotic spindle microtubules to kinetochore / mitotic spindle organization / chromosome segregation / spindle microtubule / mitotic spindle / kinetochore / mitotic cell cycle / microtubule / cell division / regulation of DNA-templated transcription / cytoplasm
Similarity search - Function
DASH complex subunit Duo1 / DASH complex subunit Spc34 / DASH complex subunit Duo1 / DASH complex subunit Spc34 / DASH complex subunit Spc19 / DASH complex subunit Dad1 / DASH complex subunit Dad2 / DASH complex subunit Ask1 / DASH complex subunit Dad3 / Spc19 ...DASH complex subunit Duo1 / DASH complex subunit Spc34 / DASH complex subunit Duo1 / DASH complex subunit Spc34 / DASH complex subunit Spc19 / DASH complex subunit Dad1 / DASH complex subunit Dad2 / DASH complex subunit Ask1 / DASH complex subunit Dad3 / Spc19 / DASH complex subunit Dad1 / DASH complex subunit Dad2 / DASH complex subunit Ask1 / DASH complex subunit Dad3 / DASH complex subunit Dam1 / DASH complex subunit Dam1 / DASH complex subunit Dad4 / DASH complex subunit Dad4 / Actin / Actin family / Actin / ATPase, nucleotide binding domain
Similarity search - Domain/homology
Uncharacterized protein / DASH complex subunit DAD1 / DASH complex subunit DAD2 / DASH complex subunit SPC19 / Uncharacterized protein / DASH complex subunit DAM1 / Uncharacterized protein / Associated with spindles and kinetochores protein 1 / DASH complex subunit duo1-like protein
Similarity search - Component
Biological speciesChaetomium thermophilum (fungus)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.5 Å
AuthorsJenni S / Harrison SC
Funding support United States, 1 items
OrganizationGrant numberCountry
Damon Runyon Cancer Research FoundationDRG-2004-09 United States
CitationJournal: Science / Year: 2018
Title: Structure of the DASH/Dam1 complex shows its role at the yeast kinetochore-microtubule interface.
Authors: Simon Jenni / Stephen C Harrison /
Abstract: Kinetochores connect mitotic-spindle microtubules with chromosomes, allowing microtubule depolymerization to pull chromosomes apart during anaphase while resisting detachment as the microtubule ...Kinetochores connect mitotic-spindle microtubules with chromosomes, allowing microtubule depolymerization to pull chromosomes apart during anaphase while resisting detachment as the microtubule shortens. The heterodecameric DASH/Dam1 complex (DASH/Dam1c), an essential component of yeast kinetochores, assembles into a microtubule-encircling ring. The ring associates with rodlike Ndc80 complexes to organize the kinetochore-microtubule interface. We report the cryo-electron microscopy structure (at ~4.5-angstrom resolution) of a DASH/Dam1c ring and a molecular model of its ordered components, validated by evolutionary direct-coupling analysis. Integrating this structure with that of the Ndc80 complex and with published interaction data yields a molecular picture of kinetochore-microtubule attachment, including how flexible, C-terminal extensions of DASH/Dam1c subunits project and contact widely separated sites on the Ndc80 complex rod and how phosphorylation at previously identified sites might regulate kinetochore assembly.
History
DepositionFeb 19, 2018-
Header (metadata) releaseMay 2, 2018-
Map releaseMay 2, 2018-
UpdateMar 13, 2024-
Current statusMar 13, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 9.2
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 9.2
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6cfz
  • Surface level: 9.2
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6cfz
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_7469.png

Downloads & links

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Map

FileDownload / File: emd_7469.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationDASH/Dam1 complex (asymmetric reconstruction)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.64 Å/pix.
x 512 pix.
= 839.68 Å		1.64 Å/pix.
x 512 pix.
= 839.68 Å		1.64 Å/pix.
x 512 pix.
= 839.68 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.64 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 9.199999999999999 / Movie #1: 9.2
Minimum - Maximum-28.424824000000001 - 64.93665
Average (Standard dev.)0.01115092 (±0.47197706)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 839.68 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.641.641.64
M x/y/z512512512
origin x/y/z0.0000.0000.000
length x/y/z839.680839.680839.680
α/β/γ90.00090.00090.000
start NX/NY/NZ-64-64-64
NX/NY/NZ128128128
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS512512512
D min/max/mean-28.42564.9370.011

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Supplemental data

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Mask #1

Fileemd_7469_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: DASH/Dam1 complex (asymmetric reconstruction, boxed volume)

Fileemd_7469_additional.map
AnnotationDASH/Dam1 complex (asymmetric reconstruction, boxed volume)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half-map 1 (not masked, not filtered)

Fileemd_7469_half_map_1.map
AnnotationHalf-map 1 (not masked, not filtered)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half-map 2 (not masked, not filtered)

Fileemd_7469_half_map_2.map
AnnotationHalf-map 2 (not masked, not filtered)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : DASH/Dam1 complex

EntireName: DASH/Dam1 complex
Components
  • Complex: DASH/Dam1 complex
    • Protein or peptide: Ask1
    • Protein or peptide: Dad3
    • Protein or peptide: Dad2
    • Protein or peptide: Duo1
    • Protein or peptide: Dad4
    • Protein or peptide: Dad1,Dad1
    • Protein or peptide: Hsk3
    • Protein or peptide: Dam1
    • Protein or peptide: Spc19
    • Protein or peptide: Spc34

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Supramolecule #1: DASH/Dam1 complex

SupramoleculeName: DASH/Dam1 complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Chaetomium thermophilum (fungus) / Location in cell: Nucleus
Molecular weightTheoretical: 115 KDa

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Macromolecule #1: Ask1

MacromoleculeName: Ask1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 8.897934 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MTLTEELEKL EQQITLTLQE IDSNFAKAHR IVTTSILPLV EQYGEHSRAV WEATKFWKQF FEASANVSLS GYEELVD

UniProtKB: Associated with spindles and kinetochores protein 1

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Macromolecule #2: Dad3

MacromoleculeName: Dad3 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 7.599788 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MLSPLEQEVL DEYERLSENM KKLAVLLDEL ASAPATEILD GLRELERKTS LVFTLLKASV YSIVLQQ

UniProtKB: Uncharacterized protein

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Macromolecule #3: Dad2

MacromoleculeName: Dad2 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 10.229733 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MSPALLARVN EKKAELENLK ELRDLSAAVA AQMEALEQKL STLSSGTEAI ATVLANWHNV LRAISMASAK IPEPKEETEE NTVPLPQTL VRIPT

UniProtKB: DASH complex subunit DAD2

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Macromolecule #4: Duo1

MacromoleculeName: Duo1 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 12.907559 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MEAREAALRK ELEGVRKINE VIEGMIGTLE RAKGNMGTVS QTVTNATTLL NTWTRMLSQT EHNQRLILNP EWKGATQDLL ELEAEERRR QEEVERRAAE AERRREEARR KA

UniProtKB: DASH complex subunit duo1-like protein

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Macromolecule #5: Dad4

MacromoleculeName: Dad4 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 8.400615 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MESPHEHQQN LLLSRIITNV EKLNEAIMVM NKTLQEINIQ NMNIELVAQM FKNYQSNVLF HLEATDNLKD PA

UniProtKB: Uncharacterized protein

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Macromolecule #6: Dad1,Dad1

MacromoleculeName: Dad1,Dad1 / type: protein_or_peptide / ID: 6 / Details: DASH/Dam1 complex subunit Dad1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 10.684892 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MSYFEQQRQA LIEEIAMNFE HVLANINKLN RSLEAVIAVG NEFSSVEALW SQFENVMSWS HPQFEKGAMT GWSHPQFEKR SAGSWSHPQ FEK

UniProtKB: DASH complex subunit DAD1

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Macromolecule #7: Hsk3

MacromoleculeName: Hsk3 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 6.641681 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MAVKARQLAH LHSQLTQLSH NLATTENLMR MTAVQAEAMR GLGSWHAGLF MAASKVLGEE S

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Macromolecule #8: Dam1

MacromoleculeName: Dam1 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 6.314139 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MNAFEPAFAE LADAVADLEA NMMHFQLMHE SLARFSESFA SFLYGLNMNA FCVDFP

UniProtKB: DASH complex subunit DAM1

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Macromolecule #9: Spc19

MacromoleculeName: Spc19 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 17.028398 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MSYADCVCSL RTSLAFLESS VATLDNGVQD FPRLCHVLRT VRHYELIPQT TLAAAEASLR DEIGPFIQLL LDRAEKHLDR QARRIETLK ARAELNAGRL SQYSGDGHNN GKFSGQGMDF RKSRPLNGEA ALRAKVVRQR KEALKYSVER LE

UniProtKB: DASH complex subunit SPC19

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Macromolecule #10: Spc34

MacromoleculeName: Spc34 / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Chaetomium thermophilum (fungus)
Molecular weightTheoretical: 27.443814 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MSLLSAHLEQ ISISCQGIDS LPFPPPKIFT NALLSNPDIT SLIRDTEAHE RALFSVPPPP PRQTTLTAEQ QQQQKPSNRR QTVFNVTGG EIRTGGVGSA STARRNTAVA AVLGGDLHAQ IMRGTRARPG QQPGSGDIDM EVLLRGVEKL CAVYPLPGAL E RVPVIRQK ...String:
MSLLSAHLEQ ISISCQGIDS LPFPPPKIFT NALLSNPDIT SLIRDTEAHE RALFSVPPPP PRQTTLTAEQ QQQQKPSNRR QTVFNVTGG EIRTGGVGSA STARRNTAVA AVLGGDLHAQ IMRGTRARPG QQPGSGDIDM EVLLRGVEKL CAVYPLPGAL E RVPVIRQK WQAQSNTLAY YEAKIAEQQE MLDRIAQERM MNDGDGDVEM EDVEEVGMTE EDLRREEEEV RELDKRKRDL QH HHHHH

UniProtKB: Uncharacterized protein

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statehelical array

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Sample preparation

Concentration2.6 mg/mL
BufferpH: 7.5
GridModel: C-flat-1.2/1.3 4C / Material: COPPER / Mesh: 400 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 35 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 89 % / Chamber temperature: 293 K / Instrument: GATAN CRYOPLUNGE 3

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Number real images: 2723 / Average electron dose: 1.115 e/Å2
Details: Movies collected: 50 frames with 1.115 e/A2 per frame
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 30488 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.2 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 34997
Startup modelType of model: OTHER / Details: Hollow cylinder with random noise
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 4.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cisTEM / Software - details: FrealignX / Number images used: 34997
Initial angle assignmentType: PROJECTION MATCHING / Software: (Name: SPARX, EMAN2)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: cisTEM / Software - details: FrealignX
FSC plot (resolution estimation)

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Atomic model buiding 1

Detailsphenix.real_space_refine
RefinementSpace: REAL / Protocol: AB INITIO MODEL / Overall B value: 240 / Target criteria: CC
Output model

PDB-6cfz:
Structure of the DASH/Dam1 complex shows its role at the yeast kinetochore-microtubule interface

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