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- PDB-8a5q: Structure of Arp4-Ies4-N-actin-Arp8-Ino80HSA subcomplex (A-module... -

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Basic information

Entry
Database: PDB / ID: 8a5q
TitleStructure of Arp4-Ies4-N-actin-Arp8-Ino80HSA subcomplex (A-module) of Chaetomium thermophilum INO80 on straight DNA
Components
  • (DNA (26-MER)) x 2
  • Actin
  • Actin related protein 4 (Arp4)
  • Actin-related protein 8
  • Ies4
  • Ino80 ATPase
KeywordsDNA BINDING PROTEIN / chromatin remodeler / INO80 / Actin-related protein
Function / homology
Function and homology information


Ino80 complex / SWI/SNF complex / NuA4 histone acetyltransferase complex / double-strand break repair / nervous system development / cytoskeleton / hydrolase activity / chromatin remodeling / DNA repair / chromatin binding ...Ino80 complex / SWI/SNF complex / NuA4 histone acetyltransferase complex / double-strand break repair / nervous system development / cytoskeleton / hydrolase activity / chromatin remodeling / DNA repair / chromatin binding / regulation of DNA-templated transcription / regulation of transcription by RNA polymerase II / ATP binding
Similarity search - Function
INO80 complex subunit Ies4 / INO80 complex, subunit Ies4 / Actins signature 1. / Actin, conserved site / Actins signature 2. / Actin/actin-like conserved site / Actins and actin-related proteins signature. / Actin / Actin family / Actin / ATPase, nucleotide binding domain
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / DNA / DNA (> 10) / DNA (> 100) / Actin-related protein 8 / Actin-related protein 4 / INO80 complex subunit 4 / Actin
Similarity search - Component
Biological speciesThermochaetoides thermophila (fungus)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsKunert, F. / Metzner, F.J. / Eustermann, S. / Jung, J. / Woike, S. / Schall, K. / Kostrewa, D. / Hopfner, K.P.
Funding supportEuropean Union, Germany, 5items
OrganizationGrant numberCountry
European Research Council (ERC)833613 INO3DEuropean Union
German Research Foundation (DFG)CRC1064 Germany
German Research Foundation (DFG)CRC1361 Germany
German Research Foundation (DFG)RTG1721 Germany
German Research Foundation (DFG)Gottfried-Wilhelm-Leibniz Prize Germany
CitationJournal: Sci Adv / Year: 2022
Title: Structural mechanism of extranucleosomal DNA readout by the INO80 complex.
Authors: Franziska Kunert / Felix J Metzner / James Jung / Markus Höpfler / Stephan Woike / Kevin Schall / Dirk Kostrewa / Manuela Moldt / Jia-Xuan Chen / Susanne Bantele / Boris Pfander / Sebastian ...Authors: Franziska Kunert / Felix J Metzner / James Jung / Markus Höpfler / Stephan Woike / Kevin Schall / Dirk Kostrewa / Manuela Moldt / Jia-Xuan Chen / Susanne Bantele / Boris Pfander / Sebastian Eustermann / Karl-Peter Hopfner /
Abstract: The nucleosomal landscape of chromatin depends on the concerted action of chromatin remodelers. The INO80 remodeler specifically places nucleosomes at the boundary of gene regulatory elements, which ...The nucleosomal landscape of chromatin depends on the concerted action of chromatin remodelers. The INO80 remodeler specifically places nucleosomes at the boundary of gene regulatory elements, which is proposed to be the result of an ATP-dependent nucleosome sliding activity that is regulated by extranucleosomal DNA features. Here, we use cryo-electron microscopy and functional assays to reveal how INO80 binds and is regulated by extranucleosomal DNA. Structures of the regulatory A-module bound to DNA clarify the mechanism of linker DNA binding. The A-module is connected to the motor unit via an HSA/post-HSA lever element to chemomechanically couple the motor and linker DNA sensing. Two notable sites of curved DNA recognition by coordinated action of the four actin/actin-related proteins and the motor suggest how sliding by INO80 can be regulated by extranucleosomal DNA features. Last, the structures clarify the recruitment of YY1/Ies4 subunits and reveal deep architectural similarities between the regulatory modules of INO80 and SWI/SNF complexes.
History
DepositionJun 15, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Dec 14, 2022Provider: repository / Type: Initial release
Revision 1.1Dec 28, 2022Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
G: Ino80 ATPase
K: DNA (26-MER)
L: DNA (26-MER)
U: Actin-related protein 8
V: Actin
W: Actin related protein 4 (Arp4)
X: Ies4
hetero molecules


Theoretical massNumber of molelcules
Total (without water)558,87713
Polymers557,2837
Non-polymers1,5946
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area29190 Å2
ΔGint-182 kcal/mol
Surface area70950 Å2
MethodPISA

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Components

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Protein , 5 types, 5 molecules GUVWX

#1: Protein Ino80 ATPase


Mass: 210443.344 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermochaetoides thermophila (fungus) / Production host: Trichoplusia ni (cabbage looper)
#4: Protein Actin-related protein 8


Mass: 84139.266 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermochaetoides thermophila (fungus) / Strain: DSM 1495 / CBS 144.50 / IMI 039719 / Gene: CTHT_0023770 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: G0S519
#5: Protein Actin


Mass: 41735.512 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermochaetoides thermophila (fungus) / Strain: DSM 1495 / CBS 144.50 / IMI 039719 / Gene: CTHT_0062070 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: G0SE15
#6: Protein Actin related protein 4 (Arp4)


Mass: 51761.199 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermochaetoides thermophila (fungus) / Strain: DSM 1495 / CBS 144.50 / IMI 039719 / Gene: CTHT_0055020 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: G0SBW4
#7: Protein Ies4


Mass: 29015.457 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermochaetoides thermophila (fungus) / Strain: DSM 1495 / CBS 144.50 / IMI 039719 / Gene: CTHT_0051540 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: G0SDE9

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DNA chain , 2 types, 2 molecules KL

#2: DNA chain DNA (26-MER)


Mass: 69840.398 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#3: DNA chain DNA (26-MER)


Mass: 70347.750 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)

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Non-polymers , 2 types, 6 molecules

#8: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: ATP, energy-carrying molecule*YM
#9: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY

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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
IDNameTypeEntity IDParent-IDSource
1INO80 A-Module (Arp4-Ies4-N-actin-Arp8-Ino80HSA) on straight DNACOMPLEX#1-#70MULTIPLE SOURCES
2INO80 A-Module (Arp4-Ies4-N-actin-Arp8-Ino80HSA)COMPLEX#1, #4-#71RECOMBINANT
3DNACOMPLEX#2-#31MULTIPLE SOURCES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Thermochaetoides thermophila (fungus)209285
33synthetic construct (others)32630
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
22Trichoplusia ni (cabbage looper)7111
33synthetic construct (others)32630
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2900 nm / Nominal defocus min: 1100 nm
Image recordingElectron dose: 42.4 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: REFMAC / Version: 5.8.0352 / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 97000 / Symmetry type: POINT
RefinementResolution: 3.15→165.2 Å / Cor.coef. Fo:Fc: 0.952 / SU B: 16.512 / SU ML: 0.266 / ESU R: 0.368
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflection
Rwork0.28351 --
obs0.28351 133748 100 %
Solvent computationSolvent model: PARAMETERS FOR MASK CACLULATION
Displacement parametersBiso mean: 127.64 Å2
Refinement stepCycle: 1 / Total: 13537
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
ELECTRON MICROSCOPYr_bond_refined_d0.0080.01213946
ELECTRON MICROSCOPYr_bond_other_d00.01612207
ELECTRON MICROSCOPYr_angle_refined_deg1.721.66119121
ELECTRON MICROSCOPYr_angle_other_deg0.5251.55628511
ELECTRON MICROSCOPYr_dihedral_angle_1_deg7.35251543
ELECTRON MICROSCOPYr_dihedral_angle_2_deg10.65510106
ELECTRON MICROSCOPYr_dihedral_angle_3_deg14.014102212
ELECTRON MICROSCOPYr_dihedral_angle_4_deg
ELECTRON MICROSCOPYr_chiral_restr0.0950.22077
ELECTRON MICROSCOPYr_gen_planes_refined0.0090.0215119
ELECTRON MICROSCOPYr_gen_planes_other0.0010.022722
ELECTRON MICROSCOPYr_nbd_refined
ELECTRON MICROSCOPYr_nbd_other
ELECTRON MICROSCOPYr_nbtor_refined
ELECTRON MICROSCOPYr_nbtor_other
ELECTRON MICROSCOPYr_xyhbond_nbd_refined
ELECTRON MICROSCOPYr_xyhbond_nbd_other
ELECTRON MICROSCOPYr_metal_ion_refined
ELECTRON MICROSCOPYr_metal_ion_other
ELECTRON MICROSCOPYr_symmetry_vdw_refined
ELECTRON MICROSCOPYr_symmetry_vdw_other
ELECTRON MICROSCOPYr_symmetry_hbond_refined
ELECTRON MICROSCOPYr_symmetry_hbond_other
ELECTRON MICROSCOPYr_symmetry_metal_ion_refined
ELECTRON MICROSCOPYr_symmetry_metal_ion_other
ELECTRON MICROSCOPYr_mcbond_it13.96410.2086199
ELECTRON MICROSCOPYr_mcbond_other13.96410.2086199
ELECTRON MICROSCOPYr_mcangle_it20.84915.3167733
ELECTRON MICROSCOPYr_mcangle_other20.84715.3227734
ELECTRON MICROSCOPYr_scbond_it17.49415.9517747
ELECTRON MICROSCOPYr_scbond_other17.49215.9547748
ELECTRON MICROSCOPYr_scangle_it
ELECTRON MICROSCOPYr_scangle_other28.43623.42211389
ELECTRON MICROSCOPYr_long_range_B_refined39.09254086
ELECTRON MICROSCOPYr_long_range_B_other39.09254087
ELECTRON MICROSCOPYr_rigid_bond_restr
ELECTRON MICROSCOPYr_sphericity_free
ELECTRON MICROSCOPYr_sphericity_bonded
LS refinement shellResolution: 3.15→3.232 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0 0 -
Rwork0.766 9875 -
obs--100 %

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