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- EMDB-0097: Cryo-EM structure of the CBF3-msk complex of the budding yeast ki... -

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Basic information

Entry
Database: EMDB / ID: EMD-0097
TitleCryo-EM structure of the CBF3-msk complex of the budding yeast kinetochore
Map data
SampleCBF3-msk complex
  • (Centromere DNA-binding protein complex CBF3 subunit ...) x 4
  • Suppressor of kinetochore protein 1
  • ligand
Function / homology
Function and homology information


RAVE complex / CBF3 complex / Neddylation / septin ring assembly / negative regulation of cytoplasmic translation / mitotic spindle elongation / centromeric DNA binding / vacuolar acidification / regulation of exit from mitosis / kinetochore assembly ...RAVE complex / CBF3 complex / Neddylation / septin ring assembly / negative regulation of cytoplasmic translation / mitotic spindle elongation / centromeric DNA binding / vacuolar acidification / regulation of exit from mitosis / kinetochore assembly / exit from mitosis / condensed chromosome, centromeric region / protein neddylation / spindle pole body / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / SCF ubiquitin ligase complex / DNA binding, bending / cullin family protein binding / mitotic spindle assembly checkpoint signaling / protein-containing complex assembly => GO:0065003 / DNA replication origin binding / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / : / Orc1 removal from chromatin / Antigen processing: Ubiquitination & Proteasome degradation / spindle midzone / regulation of protein-containing complex assembly / kinetochore / chromosome segregation / G1/S transition of mitotic cell cycle / spindle / G2/M transition of mitotic cell cycle / ubiquitin-dependent protein catabolic process / mitotic cell cycle / protein ubiquitination / DNA-binding transcription factor activity, RNA polymerase II-specific / zinc ion binding / identical protein binding / nucleus / cytoplasm
Similarity search - Function
Transcriptional activator of glycolytic enzymes / Centromere DNA-binding protein complex CBF3 subunit, domain 2 / Transcription activator GCR1-like domain / Ndc10, domain 2 / Ndc10, domain 2 superfamily / Centromere DNA-binding protein complex CBF3 subunit B, C-terminal / Centromere DNA-binding protein complex CBF3 subunit B / Zn(2)-C6 fungal-type DNA-binding domain signature. / Fungal Zn(2)-Cys(6) binuclear cluster domain / Zn(2)-C6 fungal-type DNA-binding domain profile. ...Transcriptional activator of glycolytic enzymes / Centromere DNA-binding protein complex CBF3 subunit, domain 2 / Transcription activator GCR1-like domain / Ndc10, domain 2 / Ndc10, domain 2 superfamily / Centromere DNA-binding protein complex CBF3 subunit B, C-terminal / Centromere DNA-binding protein complex CBF3 subunit B / Zn(2)-C6 fungal-type DNA-binding domain signature. / Fungal Zn(2)-Cys(6) binuclear cluster domain / Zn(2)-C6 fungal-type DNA-binding domain profile. / Zn(2)-C6 fungal-type DNA-binding domain superfamily / GAL4-like Zn(II)2Cys6 (or C6 zinc) binuclear cluster DNA-binding domain / Zn(2)-C6 fungal-type DNA-binding domain / Integrase/recombinase, N-terminal / Skp1 family, dimerisation domain / S-phase kinase-associated protein 1 / SKP1-like, dimerisation domain superfamily / SKP1 component, dimerisation / Skp1 family, tetramerisation domain / Found in Skp1 protein family / SKP1 component, POZ domain / S-phase kinase-associated protein 1-like / SKP1/BTB/POZ domain superfamily / Leucine-rich repeat domain superfamily
Similarity search - Domain/homology
Centromere DNA-binding protein complex CBF3 subunit A / Centromere DNA-binding protein complex CBF3 subunit C / Centromere DNA-binding protein complex CBF3 subunit B / Suppressor of kinetochore protein 1
Similarity search - Component
Biological speciesSaccharomyces cerevisiae S288C (yeast) / Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3 Å
AuthorsYan K / Zhang Z / Yang J / McLaughlin SH / Barford D
Funding support United Kingdom, 2 items
OrganizationGrant numberCountry
Medical Research Council (United Kingdom)MRC_UP_1201/6 United Kingdom
Cancer Research UKC576/A14109 United Kingdom
CitationJournal: Nat Struct Mol Biol / Year: 2018
Title: Architecture of the CBF3-centromere complex of the budding yeast kinetochore.
Authors: Kaige Yan / Ziguo Zhang / Jing Yang / Stephen H McLaughlin / David Barford /
Abstract: Kinetochores are multicomponent complexes responsible for coordinating the attachment of centromeric DNA to mitotic-spindle microtubules. The point centromeres of budding yeast are organized into ...Kinetochores are multicomponent complexes responsible for coordinating the attachment of centromeric DNA to mitotic-spindle microtubules. The point centromeres of budding yeast are organized into three centromeric determining elements (CDEs), and are associated with the centromere-specific nucleosome Cse4. Deposition of Cse4 at CEN loci is dependent on the CBF3 complex that engages CDEIII to direct Cse4 nucleosomes to CDEII. To understand how CBF3 recognizes CDEIII and positions Cse4, we determined a cryo-EM structure of a CBF3-CEN complex. CBF3 interacts with CEN DNA as a head-to-head dimer that includes the whole of CDEIII and immediate 3' regions. Specific CEN-binding of CBF3 is mediated by a Cep3 subunit of one of the CBF3 protomers that forms major groove interactions with the conserved and essential CCG and TGT motifs of CDEIII. We propose a model for a CBF3-Cse4-CEN complex with implications for understanding CBF3-directed deposition of the Cse4 nucleosome at CEN loci.
History
DepositionJul 2, 2018-
Header (metadata) releaseAug 22, 2018-
Map releaseDec 5, 2018-
UpdateDec 18, 2019-
Current statusDec 18, 2019Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.064
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.064
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-6gyu
  • Surface level: 0.064
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_0097.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 240 pix.
= 254.4 Å
1.06 Å/pix.
x 240 pix.
= 254.4 Å
1.06 Å/pix.
x 240 pix.
= 254.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 0.064 / Movie #1: 0.064
Minimum - Maximum-0.2236886 - 0.44195563
Average (Standard dev.)0.0010160384 (±0.013003029)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 254.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.061.061.06
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z254.400254.400254.400
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ450450450
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.2240.4420.001

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Supplemental data

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Sample components

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Entire CBF3-msk complex

EntireName: CBF3-msk complex / Number of Components: 7

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Component #1: protein, CBF3-msk complex

ProteinName: CBF3-msk complex / Recombinant expression: No
SourceSpecies: Saccharomyces cerevisiae S288C (yeast)
Source (engineered)Expression System: unidentified baculovirus

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Component #2: protein, Centromere DNA-binding protein complex CBF3 subunit B

ProteinName: Centromere DNA-binding protein complex CBF3 subunit B / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 71.439891 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c
Source (engineered)Expression System: unidentified baculovirus

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Component #3: protein, Centromere DNA-binding protein complex CBF3 subunit C

ProteinName: Centromere DNA-binding protein complex CBF3 subunit C / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 56.416863 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c
Source (engineered)Expression System: unidentified baculovirus

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Component #4: protein, Centromere DNA-binding protein complex CBF3 subunit B

ProteinName: Centromere DNA-binding protein complex CBF3 subunit B / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 66.401859 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c
Source (engineered)Expression System: unidentified baculovirus

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Component #5: protein, Suppressor of kinetochore protein 1

ProteinName: Suppressor of kinetochore protein 1 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 22.35727 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c
Source (engineered)Expression System: unidentified baculovirus

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Component #6: protein, Centromere DNA-binding protein complex CBF3 subunit A

ProteinName: Centromere DNA-binding protein complex CBF3 subunit A / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 112.066031 kDa
SourceSpecies: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c
Source (engineered)Expression System: unidentified baculovirus

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Component #7: ligand, ZINC ION

LigandName: ZINC ION / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 6.540905 MDa

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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: 27 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: 198010
3D reconstructionResolution: 3 Å / Resolution Method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement protocol: flexible / Refinement space: REAL
Input PDB model: 6GYP
Output model

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