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- PDB-7qfw: S.c. Condensin peripheral Ycg1 subcomplex bound to DNA -

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Basic information

Entry
Database: PDB / ID: 7qfw
TitleS.c. Condensin peripheral Ycg1 subcomplex bound to DNA
Components
  • Condensin complex subunit 2
  • Condensin complex subunit 3
  • Synthetic DNA ligand
  • synthetic DNA ligand
KeywordsDNA BINDING PROTEIN / SMC-motor protein
Function / homology
Function and homology information


negative regulation of meiotic DNA double-strand break formation / meiotic chromosome condensation / Condensation of Prometaphase Chromosomes / tRNA gene clustering / meiotic chromosome separation / condensin complex / rDNA chromatin condensation / synaptonemal complex assembly / mitotic chromosome condensation / chromosome condensation ...negative regulation of meiotic DNA double-strand break formation / meiotic chromosome condensation / Condensation of Prometaphase Chromosomes / tRNA gene clustering / meiotic chromosome separation / condensin complex / rDNA chromatin condensation / synaptonemal complex assembly / mitotic chromosome condensation / chromosome condensation / mitotic sister chromatid segregation / condensed chromosome / cell division / chromatin binding / nucleus / cytoplasm
Similarity search - Function
Nuclear condensin complex subunit 3, C-terminal domain / Condensin complex subunit 3 / Nuclear condensing complex subunits, C-term domain / Condensin complex subunit 2/barren / Condensin complex subunit 2 / Armadillo-like helical / Armadillo-type fold
Similarity search - Domain/homology
DNA / DNA (> 10) / Condensin complex subunit 2 / Condensin complex subunit 3
Similarity search - Component
Biological speciesSaccharomyces cerevisiae S288C (yeast)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.86 Å
AuthorsLecomte, L. / Hassler, M. / Haering, C. / Eustermann, S.
Funding supportEuropean Union, 1items
OrganizationGrant numberCountry
European Research Council (ERC)681365European Union
CitationJournal: Science / Year: 2022
Title: A hold-and-feed mechanism drives directional DNA loop extrusion by condensin.
Authors: Indra A Shaltiel / Sumanjit Datta / Léa Lecomte / Markus Hassler / Marc Kschonsak / Sol Bravo / Catherine Stober / Jenny Ormanns / Sebastian Eustermann / Christian H Haering /
Abstract: Structural maintenance of chromosomes (SMC) protein complexes structure genomes by extruding DNA loops, but the molecular mechanism that underlies their activity has remained unknown. We show that ...Structural maintenance of chromosomes (SMC) protein complexes structure genomes by extruding DNA loops, but the molecular mechanism that underlies their activity has remained unknown. We show that the active condensin complex entraps the bases of a DNA loop transiently in two separate chambers. Single-molecule imaging and cryo-electron microscopy suggest a putative power-stroke movement at the first chamber that feeds DNA into the SMC-kleisin ring upon adenosine triphosphate binding, whereas the second chamber holds on upstream of the same DNA double helix. Unlocking the strict separation of "motor" and "anchor" chambers turns condensin from a one-sided into a bidirectional DNA loop extruder. We conclude that the orientation of two topologically bound DNA segments during the SMC reaction cycle determines the directionality of DNA loop extrusion.
History
DepositionDec 6, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 15, 2022Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Condensin complex subunit 3
B: Condensin complex subunit 2
C: Synthetic DNA ligand
D: synthetic DNA ligand


Theoretical massNumber of molelcules
Total (without water)241,4824
Polymers241,4824
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area13360 Å2
ΔGint-84 kcal/mol
Surface area51890 Å2

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Components

#1: Protein Condensin complex subunit 3 / CAPG homolog


Mass: 117981.000 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c / Gene: YCG1, YCS5, YDR325W / Production host: Saccharomyces cerevisiae W303 (yeast) / References: UniProt: Q06680
#2: Protein Condensin complex subunit 2 / Barren homolog / CAPH homolog


Mass: 92721.219 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c / Gene: BRN1, YBL097W, YBL0830 / Production host: Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P38170
#3: DNA chain Synthetic DNA ligand


Mass: 15615.376 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: 50bp synthetic DNA ligand with the sequence: 5'-GTTGACAGTG TCGCAACCTG CACAGGCAAG CTGCTGAGTC TGGTGTAGAC-3' The DNA ligand was modelled as poly(dA) as the register could not be determined by cryoEM density.
Source: (gene. exp.) synthetic construct (others) / Production host: synthetic construct (others)
#4: DNA chain synthetic DNA ligand


Mass: 15164.683 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: 50bp synthetic DNA ligand with the sequence: 5'-GTCTACACCAGACTCAGCAGCTTGCCTGTGCAGGTTGCGACACTGTCAAC-3' The DNA ligand was modelled as poly(dT) as the register could not be determined by cryoEM density.
Source: (gene. exp.) synthetic construct (others) / Production host: synthetic construct (others)

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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: S.c. Condensin peripheral Ycg1 complex bound to DNA / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.707 MDa / Experimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Source (recombinant)Organism: Saccharomyces cerevisiae (brewer's yeast)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMSodium ChlorideNaCl1
220 mMHepesHepes1
31 mMDithiothreitolDTT1
45 mMMagnesium ChlorideMgCl21
SpecimenConc.: 0.707 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 281 K

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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 magnification: 130000 X / Calibrated magnification: 130000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Calibrated defocus min: 1000 nm / Calibrated defocus max: 2000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 8 sec. / Electron dose: 1 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 6544
EM imaging opticsEnergyfilter slit width: 20 eV
Image scansSampling size: 5 µm / Width: 3838 / Height: 3710 / Movie frames/image: 40 / Used frames/image: 1-40

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Processing

SoftwareName: PHENIX / Version: 1.14_3260: / Classification: refinement
EM software
IDNameVersionCategory
2SerialEMimage acquisition
4CTFFIND4.1CTF correction
7UCSF ChimeraXmodel fitting
8Cootmodel fitting
10RELION3.1.3initial Euler assignment
11RELION3.1.3final Euler assignment
12RELION3.1.3classification
13RELION3.1.33D reconstruction
14PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 91522
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.86 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 27701 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
Atomic model buildingPDB-ID: 5OQN

5oqn

RefinementStereochemistry target values: CDL v1.2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.01047586
ELECTRON MICROSCOPYf_angle_d1.023812498
ELECTRON MICROSCOPYf_chiral_restr0.06311438
ELECTRON MICROSCOPYf_plane_restr0.00811375
ELECTRON MICROSCOPYf_dihedral_angle_d16.87195284

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