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- EMDB-8938: Cryo-EM structure of nucleosome in complex with a single chain an... -

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Basic information

Entry
Database: EMDB / ID: EMD-8938
TitleCryo-EM structure of nucleosome in complex with a single chain antibody fragment
Map data
SampleNucleosome-Antibody complex:
Histone H3 / Histone H4 / Histone H2A / Histone H2B / (nucleic-acidNucleic acid) x 2 / scFvSingle-chain variable fragment
Function / homology
Function and homology information


SIRT1 negatively regulates rRNA expression / Interleukin-7 signaling / PRC2 methylates histones and DNA / Condensation of Prophase Chromosomes / Oxidative Stress Induced Senescence / Senescence-Associated Secretory Phenotype (SASP) / PKMTs methylate histone lysines / HDMs demethylate histones / HATs acetylate histones / Chromatin modifying enzymes ...SIRT1 negatively regulates rRNA expression / Interleukin-7 signaling / PRC2 methylates histones and DNA / Condensation of Prophase Chromosomes / Oxidative Stress Induced Senescence / Senescence-Associated Secretory Phenotype (SASP) / PKMTs methylate histone lysines / HDMs demethylate histones / HATs acetylate histones / Chromatin modifying enzymes / Factors involved in megakaryocyte development and platelet production / Estrogen-dependent gene expression / RUNX1 regulates genes involved in megakaryocyte differentiation and platelet function / E3 ubiquitin ligases ubiquitinate target proteins / RNA Polymerase I Promoter Escape / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / Ub-specific processing proteases / Transcriptional regulation by small RNAs / Activated PKN1 stimulates transcription of AR (androgen receptor) regulated genes KLK2 and KLK3 / SUMOylation of chromatin organization proteins / RMTs methylate histone arginines / RCAF complex / larval somatic muscle development / polytene chromosome band / polytene chromosome / chromatin assembly or disassembly / female meiosis chromosome segregation / nuclear nucleosome / nucleosomal DNA binding / nuclear chromosome / DNA-templated transcription, initiation / nucleosome assembly / nucleosome / chromosome / chromatin organization / histone binding / protein-containing complex binding / nuclear chromatin / protein heterodimerization activity / DNA binding / nucleus
Centromere kinetochore component CENP-T histone fold / Histone H3 signature 1. / Histone H4, conserved site / Histone H2A, C-terminal domain / Histone H2A conserved site / CENP-T/Histone H4, histone fold / Core histone H2A/H2B/H3/H4 / C-terminus of histone H2A / Histone H2A signature. / Histone H4 signature. ...Centromere kinetochore component CENP-T histone fold / Histone H3 signature 1. / Histone H4, conserved site / Histone H2A, C-terminal domain / Histone H2A conserved site / CENP-T/Histone H4, histone fold / Core histone H2A/H2B/H3/H4 / C-terminus of histone H2A / Histone H2A signature. / Histone H4 signature. / Histone H2B signature. / Histone H2A/H2B/H3 / Histone H3 signature 2. / Histone-fold / TATA box binding protein associated factor (TAF) / Histone H2A / Histone H3/CENP-A / Histone H2B / Histone H4
Histone H2B / Histone H3 / Histone H4 / Histone H2A
Biological speciesDrosophila melanogaster (fruit fly) / Escherichia coli (E. coli) / Mus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.99 Å
AuthorsYadav KNS / Zhou B-R
Funding support3 items
OrganizationGrant numberCountry
CitationJournal: Nat Commun / Year: 2019
Title: Atomic resolution cryo-EM structure of a native-like CENP-A nucleosome aided by an antibody fragment.
Authors: Bing-Rui Zhou / K N Sathish Yadav / Mario Borgnia / Jingjun Hong / Baohua Cao / Ada L Olins / Donald E Olins / Yawen Bai / Ping Zhang /
Abstract: Genomic DNA in eukaryotes is organized into chromatin through association with core histones to form nucleosomes, each distinguished by their DNA sequences and histone variants. Here, we used a ...Genomic DNA in eukaryotes is organized into chromatin through association with core histones to form nucleosomes, each distinguished by their DNA sequences and histone variants. Here, we used a single-chain antibody fragment (scFv) derived from the anti-nucleosome antibody mAb PL2-6 to stabilize human CENP-A nucleosome containing a native α-satellite DNA and solved its structure by the cryo-electron microscopy (cryo-EM) to 2.6 Å resolution. In comparison, the corresponding cryo-EM structure of the free CENP-A nucleosome could only reach 3.4 Å resolution. We find that scFv binds to a conserved acidic patch on the histone H2A-H2B dimer without perturbing the nucleosome structure. Our results provide an atomic resolution cryo-EM structure of a nucleosome and insight into the structure and function of the CENP-A nucleosome. The scFv approach is applicable to the structural determination of other native-like nucleosomes with distinct DNA sequences.
Validation ReportPDB-ID: 6dzt

SummaryFull reportAbout validation report
History
DepositionJul 5, 2018-
Header (metadata) releaseOct 31, 2018-
Map releaseMay 22, 2019-
UpdateJun 19, 2019-
Current statusJun 19, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6dzt
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_8938.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.99 Å/pix.
x 256 pix.
= 254.106 Å
0.99 Å/pix.
x 256 pix.
= 254.106 Å
0.99 Å/pix.
x 256 pix.
= 254.106 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.9926 Å
Density
Contour LevelBy AUTHOR: 0.025 / Movie #1: 0.025
Minimum - Maximum-0.099544995 - 0.17028357
Average (Standard dev.)0.0000022051981 (±0.0054613072)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 254.1056 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.99260156250.99260156250.9926015625
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z254.106254.106254.106
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ513513513
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.1000.1700.000

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Supplemental data

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Mask #1

Fileemd_8938_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire Nucleosome-Antibody complex

EntireName: Nucleosome-Antibody complex / Number of components: 8

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Component #1: protein, Nucleosome-Antibody complex

ProteinName: Nucleosome-Antibody complex / Recombinant expression: No
SourceSpecies: Drosophila melanogaster (fruit fly)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #2: protein, Histone H3

ProteinName: Histone H3 / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 15.421101 kDa
SourceSpecies: Drosophila melanogaster (fruit fly)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #3: protein, Histone H4

ProteinName: Histone H4 / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 11.521611 kDa
SourceSpecies: Drosophila melanogaster (fruit fly)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #4: protein, Histone H2A

ProteinName: Histone H2A / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 13.388727 kDa
SourceSpecies: Drosophila melanogaster (fruit fly)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #5: protein, Histone H2B

ProteinName: Histone H2B / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 13.840224 kDa
SourceSpecies: Drosophila melanogaster (fruit fly)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #6: nucleic-acid, DNA (147-MER)

nucleic acidName: DNA (147-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DA)(DT)(DC)(DG)(DG)(DA)(DT)(DG)(DT)(DA) (DT)(DA)(DT)(DA)(DT)(DC)(DT)(DG)(DA)(DC) (DA)(DC)(DG)(DT)(DG)(DC)(DC)(DT)(DG)(DG) (DA)(DG)(DA)(DC)(DT)(DA)(DG)(DG)(DG)(DA) (DG)(DT)(DA)(DA)(DT)(DC)(DC)(DC)(DC)(DT) (DT)(DG)(DG)(DC)(DG)(DG)(DT)(DT)(DA)(DA) (DA)(DA)(DC)(DG)(DC)(DG)(DG)(DG)(DG)(DG) (DA)(DC)(DA)(DG)(DC)(DG)(DC)(DG)(DT)(DA) (DC)(DG)(DT)(DG)(DC)(DG)(DT)(DT)(DT)(DA) (DA)(DG)(DC)(DG)(DG)(DT)(DG)(DC)(DT)(DA) (DG)(DA)(DG)(DC)(DT)(DG)(DT)(DC)(DT)(DA) (DC)(DG)(DA)(DC)(DC)(DA)(DA)(DT)(DT)(DG) (DA)(DG)(DC)(DG)(DG)(DC)(DC)(DT)(DC)(DG) (DG)(DC)(DA)(DC)(DC)(DG)(DG)(DG)(DA)(DT) (DT)(DC)(DT)(DC)(DG)(DA)(DT)
MassTheoretical: 45.610043 kDa
SourceSpecies: Escherichia coli (E. coli)

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Component #7: nucleic-acid, DNA (147-MER)

nucleic acidName: DNA (147-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DA)(DT)(DC)(DG)(DA)(DG)(DA)(DA)(DT)(DC) (DC)(DC)(DG)(DG)(DT)(DG)(DC)(DC)(DG)(DA) (DG)(DG)(DC)(DC)(DG)(DC)(DT)(DC)(DA)(DA) (DT)(DT)(DG)(DG)(DT)(DC)(DG)(DT)(DA)(DG) (DA)(DC)(DA)(DG)(DC)(DT)(DC)(DT)(DA)(DG) (DC)(DA)(DC)(DC)(DG)(DC)(DT)(DT)(DA)(DA) (DA)(DC)(DG)(DC)(DA)(DC)(DG)(DT)(DA)(DC) (DG)(DC)(DG)(DC)(DT)(DG)(DT)(DC)(DC)(DC) (DC)(DC)(DG)(DC)(DG)(DT)(DT)(DT)(DT)(DA) (DA)(DC)(DC)(DG)(DC)(DC)(DA)(DA)(DG)(DG) (DG)(DG)(DA)(DT)(DT)(DA)(DC)(DT)(DC)(DC) (DC)(DT)(DA)(DG)(DT)(DC)(DT)(DC)(DC)(DA) (DG)(DG)(DC)(DA)(DC)(DG)(DT)(DG)(DT)(DC) (DA)(DG)(DA)(DT)(DA)(DT)(DA)(DT)(DA)(DC) (DA)(DT)(DC)(DC)(DG)(DA)(DT)
MassTheoretical: 45.13877 kDa
SourceSpecies: Escherichia coli (E. coli)

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Component #8: protein, scFv

ProteinName: scFvSingle-chain variable fragment / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 29.030146 kDa
SourceSpecies: Mus musculus (house mouse)
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 solutionpH: 7.4
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: 40 e/Å2 / Illumination mode: OTHER
LensImaging mode: OTHER
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 238790
3D reconstructionSoftware: RELION / Resolution: 2.99 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Output model

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