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- PDB-6vzg: Cryo-EM structure of Sth1-Arp7-Arp9-Rtt102 -

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Basic information

Entry
Database: PDB / ID: 6vzg
TitleCryo-EM structure of Sth1-Arp7-Arp9-Rtt102
Components
  • Actin-like protein ARP9
  • Actin-related protein 7
  • Nuclear protein STH1/NPS1
  • Regulator of Ty1 transposition protein 102
KeywordsMOTOR PROTEIN / Chromatin remodeling / Nucleosome / Gene Regulation
Function / homology
Function and homology information


chromatin remodeling at centromere / RHO GTPases activate IQGAPs / RHO GTPases Activate WASPs and WAVEs / Regulation of actin dynamics for phagocytic cup formation / : / Platelet degranulation / DNA translocase activity / nucleosome disassembly / RSC-type complex / ATP-dependent chromatin remodeler activity ...chromatin remodeling at centromere / RHO GTPases activate IQGAPs / RHO GTPases Activate WASPs and WAVEs / Regulation of actin dynamics for phagocytic cup formation / : / Platelet degranulation / DNA translocase activity / nucleosome disassembly / RSC-type complex / ATP-dependent chromatin remodeler activity / SWI/SNF complex / NuA4 histone acetyltransferase complex / chromosome, centromeric region / ATP-dependent activity, acting on DNA / cytoskeleton organization / helicase activity / meiotic cell cycle / chromosome segregation / transcription elongation by RNA polymerase II / lysine-acetylated histone binding / base-excision repair / double-strand break repair / chromatin organization / DNA helicase / chromatin remodeling / chromatin binding / chromatin / regulation of DNA-templated transcription / regulation of transcription by RNA polymerase II / structural molecule activity / ATP hydrolysis activity / positive regulation of transcription by RNA polymerase II / DNA binding / ATP binding / nucleus
Similarity search - Function
Transcription regulator protein Rtt102 / Rtt102p-like transcription regulator protein / Snf2, ATP coupling domain / Snf2-ATP coupling, chromatin remodelling complex / Snf2-ATP coupling, chromatin remodelling complex / HSA domain / Helicase/SANT-associated domain / HSA domain profile. / : / SNF2-like, N-terminal domain superfamily ...Transcription regulator protein Rtt102 / Rtt102p-like transcription regulator protein / Snf2, ATP coupling domain / Snf2-ATP coupling, chromatin remodelling complex / Snf2-ATP coupling, chromatin remodelling complex / HSA domain / Helicase/SANT-associated domain / HSA domain profile. / : / SNF2-like, N-terminal domain superfamily / SNF2, N-terminal / SNF2-related domain / Actin / Actin family / Actin / Helicase conserved C-terminal domain / ATPase, nucleotide binding domain / Bromodomain, conserved site / Bromodomain signature. / Bromodomain / Bromodomain profile. / bromo domain / Bromodomain / Bromodomain-like superfamily / helicase superfamily c-terminal domain / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / Nuclear protein STH1/NPS1 / Regulator of Ty1 transposition protein 102 / Actin-like protein ARP9 / Actin-related protein 7
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsLeschziner, A.E. / Baker, R.W.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)5R01GM092895-08 United States
CitationJournal: Nat Struct Mol Biol / Year: 2021
Title: Structural insights into assembly and function of the RSC chromatin remodeling complex.
Authors: Richard W Baker / Janice M Reimer / Peter J Carman / Bengi Turegun / Tsutomu Arakawa / Roberto Dominguez / Andres E Leschziner /
Abstract: SWI/SNF chromatin remodelers modify the position and spacing of nucleosomes and, in humans, are linked to cancer. To provide insights into the assembly and regulation of this protein family, we ...SWI/SNF chromatin remodelers modify the position and spacing of nucleosomes and, in humans, are linked to cancer. To provide insights into the assembly and regulation of this protein family, we focused on a subcomplex of the Saccharomyces cerevisiae RSC comprising its ATPase (Sth1), the essential actin-related proteins (ARPs) Arp7 and Arp9 and the ARP-binding protein Rtt102. Cryo-EM and biochemical analyses of this subcomplex shows that ARP binding induces a helical conformation in the helicase-SANT-associated (HSA) domain of Sth1. Surprisingly, the ARP module is rotated 120° relative to the full RSC about a pivot point previously identified as a regulatory hub in Sth1, suggesting that large conformational changes are part of Sth1 regulation and RSC assembly. We also show that a conserved interaction between Sth1 and the nucleosome acidic patch enhances remodeling. As some cancer-associated mutations dysregulate rather than inactivate SWI/SNF remodelers, our insights into RSC complex regulation advance a mechanistic understanding of chromatin remodeling in disease states.
History
DepositionFeb 28, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 2, 2020Provider: repository / Type: Initial release
Revision 1.1Dec 23, 2020Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Jan 27, 2021Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.year
Revision 1.3Mar 6, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

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  • Deposited structure unit
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  • Imaged by UCSF Chimera
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Assembly

Deposited unit
L: Actin-related protein 7
M: Actin-like protein ARP9
K: Nuclear protein STH1/NPS1
N: Regulator of Ty1 transposition protein 102
hetero molecules


Theoretical massNumber of molelcules
Total (without water)220,3975
Polymers219,8904
Non-polymers5071
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area11580 Å2
ΔGint-55 kcal/mol
Surface area42150 Å2
Number of models10

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Components

#1: Protein Actin-related protein 7 / Actin-like protein ARP7 / Chromatin structure-remodeling complex protein ARP7 / SWI/SNF complex component ARP7


Mass: 53863.016 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c / Gene: ARP7, SWP61, YPR034W, YP9367.14 / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 Rosetta / References: UniProt: Q12406
#2: Protein Actin-like protein ARP9 / Chromatin structure-remodeling complex protein ARP9 / SWI/SNF complex component ARP9


Mass: 53131.930 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: ARP9 / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 Rosetta / References: UniProt: Q05123
#3: Protein Nuclear protein STH1/NPS1 / ATP-dependent helicase STH1 / Chromatin structure-remodeling complex protein STH1 / SNF2 homolog


Mass: 95077.289 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c / Gene: STH1, NPS1, YIL126W / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 Rosetta / References: UniProt: P32597, DNA helicase
#4: Protein Regulator of Ty1 transposition protein 102


Mass: 17817.615 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c / Gene: RTT102, YGR275W, G9378 / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 Rosetta / References: UniProt: P53330
#5: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE / Adenosine triphosphate


Mass: 507.181 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Comment: ATP, energy-carrying molecule*YM
Has ligand of interestN

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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

ComponentName: cryo-EM structure of Sth1-Arp7-Arp9-Rtt102 / Type: COMPLEX / Entity ID: #1-#4 / Source: RECOMBINANT
Molecular weightValue: 0.2 MDa / Experimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Source (recombinant)Organism: Escherichia coli (E. coli) / Strain: BL21 Rosetta
Buffer solutionpH: 7.5
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: Glow discharge for 20 seconds, 20 mAmp
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K / Details: 4 second blot time, blot force 20

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 36000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN
Image recordingAverage exposure time: 7 sec. / Electron dose: 53 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
EM imaging opticsPhase plate: VOLTA PHASE PLATE
Image scansMovie frames/image: 35

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Processing

EM software
IDNameVersionCategory
1Appion3.3particle selection
2Leginon3.3image acquisition
4CTFFIND4CTF correction
7UCSF Chimeramodel fitting
12RELION33D reconstruction
13Rosettamodel refinement
14PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1986341
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 415957 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Details: Initial model docking was done in Chimera. Sth1-Arp7-Arp9-Rtt102 were refined in Rosetta and the top ten models were deposited.
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
14I6M

4i6m

14I6M1PDBexperimental model
25TGC

5tgc

15TGC2PDBexperimental model

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