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- EMDB-23824: Open linker DNA nucleosome reconstituted with GUB DNA -

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Basic information

Entry
Database: EMDB / ID: EMD-23824
TitleOpen linker DNA nucleosome reconstituted with GUB DNA
Map data
Sample
  • Complex: Open linker DNA nucleosome reconstituted with GUB DNA
    • Protein or peptide: H3.2
    • Protein or peptide: H4
    • Protein or peptide: H2A
    • Protein or peptide: H2B
    • DNA: GUB_DNA
    • DNA: GUB_DNA_2
Function / homology
Function and homology information


structural constituent of chromatin / nucleosome / heterochromatin formation / nucleosome assembly / protein heterodimerization activity / DNA binding / nucleoplasm / nucleus
Similarity search - Function
: / Histone H2B signature. / Histone H2A conserved site / Histone H2A signature. / Histone H2B / Histone H2B / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone 2A / Histone H2A ...: / Histone H2B signature. / Histone H2A conserved site / Histone H2A signature. / Histone H2B / Histone H2B / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone 2A / Histone H2A / TATA box binding protein associated factor / TATA box binding protein associated factor (TAF), histone-like fold domain / Histone H4, conserved site / Histone H4 signature. / Histone H4 / Histone H4 / CENP-T/Histone H4, histone fold / Centromere kinetochore component CENP-T histone fold / Histone H3 signature 1. / Histone H3 signature 2. / Histone H3 / Histone H3/CENP-A / Histone H2A/H2B/H3 / Core histone H2A/H2B/H3/H4 domain / Histone-fold
Similarity search - Domain/homology
Histone H2B 1.1 / Histone H4 / Histone H3.2 / Histone H2A
Similarity search - Component
Biological speciesXenopus laevis (African clawed frog) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.52 Å
AuthorsArimura Y / Funabiki H
Funding support United States, Japan, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM132111 United States
Japan Society for the Promotion of Science (JSPS)JSPS Overseas Research Fellowships Japan
CitationJournal: Mol Cell / Year: 2021
Title: Structural features of nucleosomes in interphase and metaphase chromosomes.
Authors: Yasuhiro Arimura / Rochelle M Shih / Ruby Froom / Hironori Funabiki /
Abstract: Structural heterogeneity of nucleosomes in functional chromosomes is unknown. Here, we devise the template-, reference- and selection-free (TRSF) cryo-EM pipeline to simultaneously reconstruct cryo- ...Structural heterogeneity of nucleosomes in functional chromosomes is unknown. Here, we devise the template-, reference- and selection-free (TRSF) cryo-EM pipeline to simultaneously reconstruct cryo-EM structures of protein complexes from interphase or metaphase chromosomes. The reconstructed interphase and metaphase nucleosome structures are on average indistinguishable from canonical nucleosome structures, despite DNA sequence heterogeneity, cell-cycle-specific posttranslational modifications, and interacting proteins. Nucleosome structures determined by a decoy-classifying method and structure variability analyses reveal the nucleosome structural variations in linker DNA, histone tails, and nucleosome core particle configurations, suggesting that the opening of linker DNA, which is correlated with H2A C-terminal tail positioning, is suppressed in chromosomes. High-resolution (3.4-3.5 Å) nucleosome structures indicate DNA-sequence-independent stabilization of superhelical locations ±0-1 and ±3.5-4.5. The linker histone H1.8 preferentially binds to metaphase chromatin, from which chromatosome cryo-EM structures with H1.8 at the on-dyad position are reconstituted. This study presents the structural characteristics of nucleosomes in chromosomes.
History
DepositionApr 11, 2021-
Header (metadata) releaseSep 15, 2021-
Map releaseSep 15, 2021-
UpdateNov 17, 2021-
Current statusNov 17, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23824.png

Downloads & links

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Map

FileDownload / File: emd_23824.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.47 Å/pix.
x 200 pix.
= 294. Å		1.47 Å/pix.
x 200 pix.
= 294. Å		1.47 Å/pix.
x 200 pix.
= 294. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.47 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.03 / Movie #1: 0.03
Minimum - Maximum-0.06892017 - 0.122687906
Average (Standard dev.)5.5719378e-05 (±0.0052305125)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 294.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.471.471.47
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z294.000294.000294.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ376376376
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.0690.1230.000

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Supplemental data

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Half map: #1

Fileemd_23824_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_23824_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 : Open linker DNA nucleosome reconstituted with GUB DNA

EntireName: Open linker DNA nucleosome reconstituted with GUB DNA
Components
  • Complex: Open linker DNA nucleosome reconstituted with GUB DNA
    • Protein or peptide: H3.2
    • Protein or peptide: H4
    • Protein or peptide: H2A
    • Protein or peptide: H2B
    • DNA: GUB_DNA
    • DNA: GUB_DNA_2

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Supramolecule #1: Open linker DNA nucleosome reconstituted with GUB DNA

SupramoleculeName: Open linker DNA nucleosome reconstituted with GUB DNA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Xenopus laevis (African clawed frog)

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Macromolecule #1: H3.2

MacromoleculeName: H3.2 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
ARTKQTARKS TGGKAPRKQL ATKAARKSAP ATGGVKKPHR YRPGTVALRE IRRYQKSTEL LIRKLPFQRL VREIAQDFKT DLRFQSSAVM ALQEASEAYL VALFEDTNLC AIHAKRVTIM PKDIQLARRI RGERA

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Macromolecule #2: H4

MacromoleculeName: H4 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
SGRGKGGKGL GKGGAKRHRK VLRDNIQGIT KPAIRRLARR GGVKRISGLI YEETRGVLKV FLENVIRDAV TYTEHAKRKT VTAMDVVYAL KRQGRTLYGF GG

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Macromolecule #3: H2A

MacromoleculeName: H2A / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
SGRGKQGGKT RAKAKTRSSR AGLQFPVGRV HRLLRKGNYA ERVGAGAPVY LAAVLEYLTA EILELAGNAA RDNKKTRIIP RHLQLAVRND EELNKLLGRV TIAQGGVLPN IQSVLLPKKT ESSKSAKSK

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Macromolecule #4: H2B

MacromoleculeName: H2B / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MPEPAKSAPA PKKGSKKAVT KTQKKDGKKR RKTRKESYAI YVYKVLKQVH PDTGISSKAM SIMNSFVNDV FERIAGEASR LAHYNKRSTI TSREIQTAVR LLLPGELAKH AVSEGTKAVT KYTSAK

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Macromolecule #5: GUB_DNA

MacromoleculeName: GUB_DNA / type: dna / ID: 5 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
SequenceString:
ATCCCTCTAG ACGGAGGACA GTCCTCCGGT TACCTTCGAA CCACGTGGCC GTCTAGATGC TGACTCATTG TCGACACGCG TAGATCTGCT AGCATCGATC CATGGACTAG TCTCGAGTTT AAAGATATCC AGCTGCCCGG GAGGCCTTCG CGAAATATTG GTACCCCATG GAAGAT

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Macromolecule #6: GUB_DNA_2

MacromoleculeName: GUB_DNA_2 / type: dna / ID: 6 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
SequenceString:
ATCTTCCATG GGGTACCAAT ATTTCGCGAA GGCCTCCCGG GCAGCTGGAT ATCTTTAAAC TCGAGACTAG TCCATGGATC GATGCTAGCA GATCTACGCG TGTCGACAAT GAGTCAGCAT CTAGACGGCC ACGTGGTTCG AAGGTAACCG GAGGACTGTC CTCCGTCTAG AGGGAT

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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: 7.4
VitrificationCryogen name: ETHANE
DetailsOpen linker DNA nucleosome reconstituted with GUB DNA

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 36.1 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 199125
CTF correctionSoftware: (Name: cryoSPARC (ver. 2), RELION (ver. 3))
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.52 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3) / Number images used: 62823
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 2)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 2)
FSC plot (resolution estimation)

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