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- EMDB-4580: Structure of inner kinetochore CCAN complex -

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Basic information

Entry
Database: EMDB / ID: EMD-4580
TitleStructure of inner kinetochore CCAN complex
Map data
SampleINNER KINETOCHORE CCAN COMPLEX:
(Central kinetochore subunit ...) x 2 / (Inner kinetochore subunit ...) x 9
Function / homology
Function and homology information


negative regulation of meiotic DNA double-strand break formation involved in reciprocal meiotic recombination / Mis6-Sim4 complex / COMA complex / maintenance of meiotic sister chromatid cohesion / establishment of meiotic sister chromatid cohesion / centromere complex assembly / meiotic sister chromatid segregation / ascospore formation / attachment of spindle microtubules to kinetochore / establishment of mitotic sister chromatid cohesion ...negative regulation of meiotic DNA double-strand break formation involved in reciprocal meiotic recombination / Mis6-Sim4 complex / COMA complex / maintenance of meiotic sister chromatid cohesion / establishment of meiotic sister chromatid cohesion / centromere complex assembly / meiotic sister chromatid segregation / ascospore formation / attachment of spindle microtubules to kinetochore / establishment of mitotic sister chromatid cohesion / condensed nuclear chromosome outer kinetochore / kinetochore assembly / protein localization to chromosome, centromeric region / condensed nuclear chromosome kinetochore / spindle pole body / protein localization to kinetochore / mitotic spindle assembly checkpoint / DNA replication initiation / condensed chromosome kinetochore / kinetochore / meiotic cell cycle / chromosome segregation / cell division / structural molecule activity / nucleus / cytoplasm
Centromere protein O / Kinetochore subunit Nkp2/Cnl2 / Centromere protein Chl4/mis15/CENP-N
Inner kinetochore subunit CTF19 / Inner kinetochore subunit NKP1 / Inner kinetochore subunit MCM16 / Inner kinetochore subunit MCM21 / Inner kinetochore subunit NKP2 / Inner kinetochore subunit CHL4 / Inner kinetochore subunit OKP1 / Inner kinetochore subunit MCM22 / Inner kinetochore subunit AME1 / Inner kinetochore subunit IML3 ...Inner kinetochore subunit CTF19 / Inner kinetochore subunit NKP1 / Inner kinetochore subunit MCM16 / Inner kinetochore subunit MCM21 / Inner kinetochore subunit NKP2 / Inner kinetochore subunit CHL4 / Inner kinetochore subunit OKP1 / Inner kinetochore subunit MCM22 / Inner kinetochore subunit AME1 / Inner kinetochore subunit IML3 / Central kinetochore subunit MCM16 / Inner kinetochore subunit CTF3
Biological speciesSaccharomyces cerevisiae (baker's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.55 Å
AuthorsYan K / Yang J / Zhang Z / McLaughlin SH / Chang L / Fasci D / Heck AJR / Barford D
Funding support United Kingdom, 3 items
OrganizationGrant numberCountry
Medical Research Council (United Kingdom)MC_UP_1201/6 United Kingdom
European CommissionHorizon 2020 program INFRAIA project Epic-XS (Project 823839)
Cancer Research UKC576/A14109 United Kingdom
CitationJournal: Nature / Year: 2019
Title: Structure of the inner kinetochore CCAN complex assembled onto a centromeric nucleosome.
Authors: Kaige Yan / Jing Yang / Ziguo Zhang / Stephen H McLaughlin / Leifu Chang / Domenico Fasci / Ann E Ehrenhofer-Murray / Albert J R Heck / David Barford /
Abstract: In eukaryotes, accurate chromosome segregation in mitosis and meiosis maintains genome stability and prevents aneuploidy. Kinetochores are large protein complexes that, by assembling onto specialized ...In eukaryotes, accurate chromosome segregation in mitosis and meiosis maintains genome stability and prevents aneuploidy. Kinetochores are large protein complexes that, by assembling onto specialized Cenp-A nucleosomes, function to connect centromeric chromatin to microtubules of the mitotic spindle. Whereas the centromeres of vertebrate chromosomes comprise millions of DNA base pairs and attach to multiple microtubules, the simple point centromeres of budding yeast are connected to individual microtubules. All 16 budding yeast chromosomes assemble complete kinetochores using a single Cenp-A nucleosome (Cenp-A), each of which is perfectly centred on its cognate centromere. The inner and outer kinetochore modules are responsible for interacting with centromeric chromatin and microtubules, respectively. Here we describe the cryo-electron microscopy structure of the Saccharomyces cerevisiae inner kinetochore module, the constitutive centromere associated network (CCAN) complex, assembled onto a Cenp-A nucleosome (CCAN-Cenp-A). The structure explains the interdependency of the constituent subcomplexes of CCAN and shows how the Y-shaped opening of CCAN accommodates Cenp-A to enable specific CCAN subunits to contact the nucleosomal DNA and histone subunits. Interactions with the unwrapped DNA duplex at the two termini of Cenp-A are mediated predominantly by a DNA-binding groove in the Cenp-L-Cenp-N subcomplex. Disruption of these interactions impairs assembly of CCAN onto Cenp-A. Our data indicate a mechanism of Cenp-A nucleosome recognition by CCAN and how CCAN acts as a platform for assembly of the outer kinetochore to link centromeres to the mitotic spindle for chromosome segregation.
Validation ReportSummary, Full report, XML, About validation report
History
DepositionJan 31, 2019-
Header (metadata) releaseSep 4, 2019-
Map releaseOct 2, 2019-
UpdateNov 27, 2019-
Current statusNov 27, 2019Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.026
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.026
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6qle
  • Surface level: 0.026
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_4580.png

Downloads & links

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Map

FileDownload / File: emd_4580.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.09 Å/pix.
x 320 pix.
= 348.8 Å		1.09 Å/pix.
x 320 pix.
= 348.8 Å		1.09 Å/pix.
x 320 pix.
= 348.8 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: 0.026 / Movie #1: 0.026
Minimum - Maximum-0.16186026 - 0.22095919
Average (Standard dev.)0.00020103065 (±0.003418988)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 348.80002 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.091.091.09
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z348.800348.800348.800
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ320320320
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.1620.2210.000

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Supplemental data

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Sample components

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Entire INNER KINETOCHORE CCAN COMPLEX

EntireName: INNER KINETOCHORE CCAN COMPLEX / Number of components: 12

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Component #1: protein, INNER KINETOCHORE CCAN COMPLEX

ProteinName: INNER KINETOCHORE CCAN COMPLEX / Recombinant expression: No
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #2: protein, Central kinetochore subunit MCM16,Central kinetochore su...

ProteinName: Central kinetochore subunit MCM16,Central kinetochore subunit MCM16,Inner kinetochore subunit MCM16,Mcm16p
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 20.019242 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #3: protein, Central kinetochore subunit CTF3,Inner kinetochore subun...

ProteinName: Central kinetochore subunit CTF3,Inner kinetochore subunit CTF3,Central kinetochore subunit CTF3,Inner kinetochore subunit CTF3
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 46.149934 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #4: protein, Inner kinetochore subunit MCM22,Inner kinetochore subuni...

ProteinName: Inner kinetochore subunit MCM22,Inner kinetochore subunit MCM22,Inner kinetochore subunit MCM22,Inner kinetochore subunit MCM22
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 21.431949 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #5: protein, Inner kinetochore subunit IML3

ProteinName: Inner kinetochore subunit IML3 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 28.093223 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #6: protein, Inner kinetochore subunit CHL4

ProteinName: Inner kinetochore subunit CHL4 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 52.743723 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #7: protein, Inner kinetochore subunit MCM21

ProteinName: Inner kinetochore subunit MCM21 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 43.028879 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #8: protein, Inner kinetochore subunit CTF19

ProteinName: Inner kinetochore subunit CTF19 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 42.841113 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #9: protein, Inner kinetochore subunit OKP1,Inner kinetochore subunit...

ProteinName: Inner kinetochore subunit OKP1,Inner kinetochore subunit OKP1,Inner kinetochore subunit OKP1,Inner kinetochore subunit OKP1
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 30.460818 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #10: protein, Inner kinetochore subunit AME1,Inner kinetochore subunit...

ProteinName: Inner kinetochore subunit AME1,Inner kinetochore subunit AME1,Inner kinetochore subunit AME1,Inner kinetochore subunit AME1
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 20.940982 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #11: protein, Inner kinetochore subunit NKP1

ProteinName: Inner kinetochore subunit NKP1 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 27.006451 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #12: protein, Inner kinetochore subunit NKP2

ProteinName: Inner kinetochore subunit NKP2 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 17.877033 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 32 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: FEI FALCON III (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 465029
3D reconstruction #1Resolution: 3.55 Å / Resolution method: FSC 0.143 CUT-OFF
3D reconstruction #2Resolution: 3.55 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Output model

6qle

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