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- PDB-8com: Structure of the Nucleosome Core Particle from Trypanosoma brucei -

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Basic information

Entry
Database: PDB / ID: 8com
TitleStructure of the Nucleosome Core Particle from Trypanosoma brucei
Components
  • (Widom 601 145 bp DNA (127-mer ordered and ...) x 2
  • Histone H2A
  • Histone H2B
  • Histone H3, putative
  • Histone H4
KeywordsDNA BINDING PROTEIN / Nucleosome Chromatin Parasite Trypanosome Kinetoplast
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
DNA / DNA (> 10) / DNA (> 100) / Histone H2B / Histone H3, putative / Histone H2A / Histone H4
Similarity search - Component
Biological speciesTrypanosoma brucei brucei TREU927 (eukaryote)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsBurdett, H. / Deak, G. / Wilson, M.D.
Funding support United Kingdom, 2items
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, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jul 12, 2023Provider: repository / Type: Initial release
Revision 1.1Jul 19, 2023Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name
Revision 1.2Sep 6, 2023Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Histone H3, putative
B: Histone H4
C: Histone H2A
D: Histone H2B
E: Histone H3, putative
F: Histone H4
G: Histone H2A
H: Histone H2B
I: Widom 601 145 bp DNA (127-mer ordered and built)
J: Widom 601 145 bp DNA (127-mer ordered and built)


Theoretical massNumber of molelcules
Total (without water)194,03010
Polymers194,03010
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: SAXS, native gels, SAXS, negative stain all support monodispersity of sample
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area51590 Å2
ΔGint-348 kcal/mol
Surface area63340 Å2
MethodPISA

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Components

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Protein , 4 types, 8 molecules AEBFCGDH

#1: Protein Histone H3, putative


Mass: 14647.858 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Trypanosoma brucei brucei TREU927 (eukaryote)
Strain: TREU927
Gene: TB927.1.2430, TB927.1.2450, TB927.1.2470, TB927.1.2490, TB927.1.2510, TB927.1.2530, TB927.1.2550
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q4GYX7
#2: Protein Histone H4


Mass: 11037.914 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Trypanosoma brucei brucei TREU927 (eukaryote)
Strain: TREU927
Gene: Tb05.45E22.530, Tb05.45E22.470, Tb05.45E22.480, Tb05.45E22.490, Tb05.45E22.500, Tb05.45E22.510, Tb05.45E22.520, Tb05.45E22.540, Tb05.45E22.550, Tb05.45E22.560, Tb927.5.4170, Tb927.5.4180, Tb927. ...Gene: Tb05.45E22.530, Tb05.45E22.470, Tb05.45E22.480, Tb05.45E22.490, Tb05.45E22.500, Tb05.45E22.510, Tb05.45E22.520, Tb05.45E22.540, Tb05.45E22.550, Tb05.45E22.560, Tb927.5.4170, Tb927.5.4180, Tb927.5.4190, Tb927.5.4200, Tb927.5.4210, Tb927.5.4220, Tb927.5.4230, Tb927.5.4240, Tb927.5.4250, Tb927.5.4260
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q57Z31
#3: Protein Histone H2A


Mass: 14108.614 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Trypanosoma brucei brucei TREU927 (eukaryote)
Strain: TREU927
Gene: Tb07.13M20.520, Tb07.13M20.510, Tb07.13M20.530, Tb07.13M20.540, Tb07.13M20.550, Tb07.13M20.560, Tb07.13M20.570, Tb07.13M20.580, Tb07.13M20.590, Tb07.13M20.600, Tb07.13M20.610, Tb07.13M20.620, ...Gene: Tb07.13M20.520, Tb07.13M20.510, Tb07.13M20.530, Tb07.13M20.540, Tb07.13M20.550, Tb07.13M20.560, Tb07.13M20.570, Tb07.13M20.580, Tb07.13M20.590, Tb07.13M20.600, Tb07.13M20.610, Tb07.13M20.620, Tb07.13M20.630, Tb927.7.2820, Tb927.7.2830, Tb927.7.2840, Tb927.7.2850, Tb927.7.2860, Tb927.7.2870, Tb927.7.2880, Tb927.7.2890, Tb927.7.2900, Tb927.7.2910, Tb927.7.2920, Tb927.7.2930, Tb927.7.2940
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q57YA3
#4: Protein Histone H2B


Mass: 12464.503 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Trypanosoma brucei brucei TREU927 (eukaryote)
Strain: TREU927
Gene: Tb10.406.0330, Tb10.406.0350, Tb10.406.0360, Tb10.406.0370, Tb10.406.0380, Tb10.406.0400, Tb10.406.0410, Tb10.406.0420, Tb10.406.0430, Tb10.406.0440, Tb10.406.0450, Tb10.406.0460
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q389T1

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Widom 601 145 bp DNA (127-mer ordered and ... , 2 types, 2 molecules IJ

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


Mass: 44991.660 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli K2 (bacteria)
#6: DNA chain Widom 601 145 bp DNA (127-mer ordered and built)


Mass: 44520.383 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli K2 (bacteria)

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeDetailsEntity IDParent-IDSource
1Nucleosome core particleCOMPLEXNucleosome core particle with histones from Trypanosoma brucei and Widom 601 145 bp DNAall0RECOMBINANT
2Histone octamerCOMPLEXTrypanosoma brucei histone octamer composed of an H4-H3-H3-H4 tetramer and two H2A-H2B dimers#1-#41RECOMBINANT
3Widom 601 145 bp DNACOMPLEX#5-#61RECOMBINANT
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
110.1938 MDaNO
210.1042 MDaNO
310.0896 MDaNO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Trypanosoma brucei brucei TREU927 (eukaryote)185431
32Trypanosoma brucei brucei TREU927 (eukaryote)185431
43synthetic construct (others)32630
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDStrain
21Escherichia coli (E. coli)562BL21 (DE3)
32Escherichia coli (E. coli)562BL21 (DE3)
43Escherichia coli (E. coli)562
Buffer solutionpH: 7.5 / Details: 20 mM HEPES pH 7.5, 150 mM NaCl, 1 mM DTT
SpecimenConc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Glutaraldehyde fixation, S200 purification
Specimen supportDetails: PELCO easiglow / Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 105000 X / Nominal defocus max: 3200 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm / C2 aperture diameter: 75 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 7 sec. / Electron dose: 45.7 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4913
Details: correlated double sampling used, super-resolution, binning at motion correction stage

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Processing

EM software
IDNameVersionCategory
2SerialEMimage acquisition
4Gctf1.08CTF correction
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
12cryoSPARCclassification
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 306475 / Algorithm: FOURIER SPACE / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00411391
ELECTRON MICROSCOPYf_angle_d0.55716465
ELECTRON MICROSCOPYf_dihedral_angle_d31.7083343
ELECTRON MICROSCOPYf_chiral_restr0.0321900
ELECTRON MICROSCOPYf_plane_restr0.0031198

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