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- EMDB-0772: H3-H3-H3 tri-nucleosome with the 30 base-pair linker DNA -

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Basic information

Entry
Database: EMDB / ID: EMD-0772
TitleH3-H3-H3 tri-nucleosome with the 30 base-pair linker DNA
Map data
SampleH3-H3-H3 tri-nucleosome with the 30 base-pair linker DNA
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 15.7 Å
AuthorsTakizawa Y / Ho C-H / Tachiwana H / Ohi M / Wolf M / Kurumizaka H
Funding support Japan, 6 items
OrganizationGrant numberCountry
Japan Society for the Promotion of ScienceJP18H05534 Japan
Japan Society for the Promotion of ScienceJP17H05013 Japan
Japan Society for the Promotion of ScienceJP17H01408 Japan
Japan Society for the Promotion of Science15H05972 Japan
Japan Society for the Promotion of Science17H06167 Japan
Japan Society for the Promotion of ScienceJP19K06522 Japan
CitationJournal: Structure / Year: 2019
Title: Cryo-EM Structures of Centromeric Tri-nucleosomes Containing a Central CENP-A Nucleosome.
Authors: Yoshimasa Takizawa / Cheng-Han Ho / Hiroaki Tachiwana / Hideyuki Matsunami / Wataru Kobayashi / Midori Suzuki / Yasuhiro Arimura / Tetsuya Hori / Tatsuo Fukagawa / Melanie D Ohi / Matthias Wolf / Hitoshi Kurumizaka /
Abstract: The histone H3 variant CENP-A is a crucial epigenetic marker for centromere specification. CENP-A forms a characteristic nucleosome and dictates the higher-order configuration of centromeric ...The histone H3 variant CENP-A is a crucial epigenetic marker for centromere specification. CENP-A forms a characteristic nucleosome and dictates the higher-order configuration of centromeric chromatin. However, little is known about how the CENP-A nucleosome affects the architecture of centromeric chromatin. In this study, we reconstituted tri-nucleosomes mimicking a centromeric nucleosome arrangement containing the CENP-A nucleosome, and determined their 3D structures by cryoelectron microscopy. The H3-CENP-A-H3 tri-nucleosomes adopt an untwisted architecture, with an outward-facing linker DNA path between nucleosomes. This is distinct from the H3-H3-H3 tri-nucleosome architecture, with an inward-facing DNA path. Intriguingly, the untwisted architecture may allow the CENP-A nucleosome to be exposed to the solvent in the condensed chromatin model. These results provide a structural basis for understanding the 3D configuration of CENP-A-containing chromatin, and may explain how centromeric proteins can specifically target the CENP-A nucleosomes buried in robust amounts of H3 nucleosomes in centromeres.
History
DepositionAug 24, 2019-
Header (metadata) releaseDec 4, 2019-
Map releaseDec 4, 2019-
UpdateDec 4, 2019-
Current statusDec 4, 2019Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3.6
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 3.6
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_0772.png

Downloads & links

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Map

FileDownload / File: emd_0772.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.4 Å/pix.
x 280 pix.
= 392. Å		1.4 Å/pix.
x 280 pix.
= 392. Å		1.4 Å/pix.
x 280 pix.
= 392. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.4 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 3.6 / Movie #1: 3.6
Minimum - Maximum-4.573675 - 16.096634
Average (Standard dev.)0.00000000659 (±1)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 392.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.41.41.4
M x/y/z280280280
origin x/y/z0.0000.0000.000
length x/y/z392.000392.000392.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS280280280
D min/max/mean-4.57416.0970.000

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Supplemental data

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Sample components

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Entire H3-H3-H3 tri-nucleosome with the 30 base-pair linker DNA

EntireName: H3-H3-H3 tri-nucleosome with the 30 base-pair linker DNA
Number of components: 1

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Component #1: protein, H3-H3-H3 tri-nucleosome with the 30 base-pair linker DNA

ProteinName: H3-H3-H3 tri-nucleosome with the 30 base-pair linker DNA
Recombinant expression: No
MassTheoretical: 600 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.03 mg/mL / pH: 7.8
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
ImagingMicroscope: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 25 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 2.7 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: FEI FALCON III (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 8035

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 4534
3D reconstructionSoftware: RELION / Resolution: 15.7 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Modeling #1Refinement protocol: rigid body

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