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- EMDB-16777: Structure of the Nucleosome Core Particle from Trypanosoma brucei -

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Basic information

Entry
Database: EMDB / ID: EMD-16777
TitleStructure of the Nucleosome Core Particle from Trypanosoma brucei
Map dataMasked sharpened map of Nucleosome core particle from Trypanosoma brucei
Sample
  • Complex: Nucleosome core particle
    • Complex: Histone octamer
      • Protein or peptide: Histone H3, putative
      • Protein or peptide: Histone H4
      • Protein or peptide: Histone H2A
      • Protein or peptide: Histone H2B
    • Complex: Widom 601 145 bp DNA
      • DNA: Widom 601 145 bp DNA (127-mer ordered and built)
      • DNA: Widom 601 145 bp DNA (127-mer ordered and built)
KeywordsNucleosome Chromatin Parasite Trypanosome Kinetoplast / DNA BINDING PROTEIN
Function / homology
Function and homology information


ciliary transition zone / nuclear lumen / ciliary plasm / phosphate ion binding / chromosome organization / structural constituent of chromatin / nucleosome / nucleosome assembly / protein heterodimerization activity / DNA binding ...ciliary transition zone / nuclear lumen / ciliary plasm / phosphate ion binding / chromosome organization / structural constituent of chromatin / nucleosome / nucleosome assembly / protein heterodimerization activity / DNA binding / nucleoplasm / nucleus / cytoplasm
Similarity search - Function
Histone H2B signature. / Histone H2B / Histone H2B / Histone H2A conserved site / Histone H2A signature. / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone H2A / Histone 2A / Histone H4 ...Histone H2B signature. / Histone H2B / Histone H2B / Histone H2A conserved site / Histone H2A signature. / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone H2A / Histone 2A / Histone H4 / Histone H4 / Histone H3 signature 2. / Histone H3 / Histone H3/CENP-A / Histone H2A/H2B/H3 / Core histone H2A/H2B/H3/H4 / Histone-fold
Similarity search - Domain/homology
Histone H2B / Histone H3, putative / Histone H2A / Histone H4
Similarity search - Component
Biological speciesTrypanosoma brucei brucei TREU927 (eukaryote) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsSandoval G / Deak G / Tuijtel MW / Wilson MD
Funding support United Kingdom, 2 items
OrganizationGrant numberCountry
Wellcome Trust210493 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/M010996/1 United Kingdom
CitationJournal: Nucleic Acids Res / Year: 2023
Title: Histone divergence in trypanosomes results in unique alterations to nucleosome structure.
Authors: Gauri Deák / Hannah Wapenaar / Gorka Sandoval / Ruofan Chen / Mark R D Taylor / Hayden Burdett / James A Watson / Maarten W Tuijtel / Shaun Webb / Marcus D Wilson /
Abstract: Eukaryotes have a multitude of diverse mechanisms for organising and using their genomes, but the histones that make up chromatin are highly conserved. Unusually, histones from kinetoplastids are ...Eukaryotes have a multitude of diverse mechanisms for organising and using their genomes, but the histones that make up chromatin are highly conserved. Unusually, histones from kinetoplastids are highly divergent. The structural and functional consequences of this variation are unknown. Here, we have biochemically and structurally characterised nucleosome core particles (NCPs) from the kinetoplastid parasite Trypanosoma brucei. A structure of the T. brucei NCP reveals that global histone architecture is conserved, but specific sequence alterations lead to distinct DNA and protein interaction interfaces. The T. brucei NCP is unstable and has weakened overall DNA binding. However, dramatic changes at the H2A-H2B interface introduce local reinforcement of DNA contacts. The T. brucei acidic patch has altered topology and is refractory to known binders, indicating that the nature of chromatin interactions in T. brucei may be unique. Overall, our results provide a detailed molecular basis for understanding evolutionary divergence in chromatin structure.
History
DepositionFeb 28, 2023-
Header (metadata) releaseJul 12, 2023-
Map releaseJul 12, 2023-
UpdateSep 6, 2023-
Current statusSep 6, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_16777.map.gz / Format: CCP4 / Size: 120.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMasked sharpened map of Nucleosome core particle from Trypanosoma brucei
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 316 pix.
= 261.964 Å
0.83 Å/pix.
x 316 pix.
= 261.964 Å
0.83 Å/pix.
x 316 pix.
= 261.964 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.829 Å
Density
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-1.5084459 - 2.5465097
Average (Standard dev.)0.00081927225 (±0.076253876)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions316316316
Spacing316316316
CellA=B=C: 261.964 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_16777_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: unsharpened map of Nucleosome core particle from Trypanosoma brucei

Fileemd_16777_additional_1.map
Annotationunsharpened map of Nucleosome core particle from Trypanosoma brucei
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: half map 2 of Nucleosome core particle from Trypanosoma brucei

Fileemd_16777_half_map_1.map
Annotationhalf map 2 of Nucleosome core particle from Trypanosoma brucei
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: half map 1 of Nucleosome core particle from Trypanosoma brucei

Fileemd_16777_half_map_2.map
Annotationhalf map 1 of Nucleosome core particle from Trypanosoma brucei
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : Nucleosome core particle

EntireName: Nucleosome core particle
Components
  • Complex: Nucleosome core particle
    • Complex: Histone octamer
      • Protein or peptide: Histone H3, putative
      • Protein or peptide: Histone H4
      • Protein or peptide: Histone H2A
      • Protein or peptide: Histone H2B
    • Complex: Widom 601 145 bp DNA
      • DNA: Widom 601 145 bp DNA (127-mer ordered and built)
      • DNA: Widom 601 145 bp DNA (127-mer ordered and built)

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Supramolecule #1: Nucleosome core particle

SupramoleculeName: Nucleosome core particle / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Nucleosome core particle with histones from Trypanosoma brucei and Widom 601 145 bp DNA
Source (natural)Organism: Trypanosoma brucei brucei TREU927 (eukaryote)
Molecular weightTheoretical: 89.6 KDa

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Supramolecule #2: Histone octamer

SupramoleculeName: Histone octamer / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#4
Details: Trypanosoma brucei histone octamer composed of an H4-H3-H3-H4 tetramer and two H2A-H2B dimers
Source (natural)Organism: Trypanosoma brucei brucei TREU927 (eukaryote)

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Supramolecule #3: Widom 601 145 bp DNA

SupramoleculeName: Widom 601 145 bp DNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #5-#6
Source (natural)Organism: synthetic construct (others)

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Macromolecule #1: Histone H3, putative

MacromoleculeName: Histone H3, putative / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Trypanosoma brucei brucei TREU927 (eukaryote) / Strain: TREU927
Molecular weightTheoretical: 14.647858 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
SRTKETARTK KTITSKKSKK ASKGSDAASG VKTAQRRWRP GTVALREIRQ FQRSTDLLLQ KAPFQRLVRE VSGAQKEGLR FQSSAILAA QEATESYIVS LLADTNRACI HSGRVTIQPK DIHLALCLRG ERA

UniProtKB: Histone H3, putative

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Macromolecule #2: Histone H4

MacromoleculeName: Histone H4 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Trypanosoma brucei brucei TREU927 (eukaryote) / Strain: TREU927
Molecular weightTheoretical: 11.037914 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
AKGKKSGEAK GSQKRQKKVL RENVRGITRG SIRRLARRGG VKRISGVIYD EVRGVLKSFV EGVVRDATAY TEYSRKKTVT AVDVVNALR KRGKILYGYA

UniProtKB: Histone H4

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Macromolecule #3: Histone H2A

MacromoleculeName: Histone H2A / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Trypanosoma brucei brucei TREU927 (eukaryote) / Strain: TREU927
Molecular weightTheoretical: 14.108614 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
ATPKQAVKKA SKGGSSRSVK AGLIFPVGRV GTLLRRGQYA RRIGASGAVY MAAVLEYLTA ELLELSVKAA AQQTKKTKRL TPRTVTLAV RHDDDLGALL RNVTMSRGGV MPSLNKALAK KQKSGKHAKA TPSV

UniProtKB: Histone H2A

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Macromolecule #4: Histone H2B

MacromoleculeName: Histone H2B / type: protein_or_peptide / ID: 4 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Trypanosoma brucei brucei TREU927 (eukaryote) / Strain: TREU927
Molecular weightTheoretical: 12.464503 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
ATPKSTPAKT RKEAKKTRRQ RKRTWNVYVS RSLRSINSQM SMTSRTMKIV NSFVNDLFER IAAEAATIVR VNRKRTLGAR ELQTAVRLV LPADLAKHAM AEGTKAVSHA SS

UniProtKB: Histone H2B

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Macromolecule #5: Widom 601 145 bp DNA (127-mer ordered and built)

MacromoleculeName: Widom 601 145 bp DNA (127-mer ordered and built) / type: dna / ID: 5 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 44.99166 KDa
SequenceString: (DA)(DT)(DC)(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) ...String:
(DA)(DT)(DC)(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)(DG) (DA)(DT)

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Macromolecule #6: Widom 601 145 bp DNA (127-mer ordered and built)

MacromoleculeName: Widom 601 145 bp DNA (127-mer ordered and built) / type: dna / ID: 6 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 44.520383 KDa
SequenceString: (DA)(DT)(DC)(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) ...String:
(DA)(DT)(DC)(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)(DG) (DA)(DT)

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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

Concentration0.5 mg/mL
BufferpH: 7.5 / Details: 20 mM HEPES pH 7.5, 150 mM NaCl, 1 mM DTT
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 120 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 3e-05 kPa / Details: PELCO easiglow
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
DetailsGlutaraldehyde fixation, S200 purification

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 4913 / Average exposure time: 7.0 sec. / Average electron dose: 45.7 e/Å2
Details: correlated double sampling used, super-resolution, binning at motion correction stage
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 75.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.2 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL / In silico model: cryoSPARC ab initio
Final reconstructionAlgorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 306475
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationNumber classes: 2 / Software - Name: cryoSPARC

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