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- PDB-6t8b: FtsK motor domain with dsDNA, translocating state -

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Basic information

Entry
Database: PDB / ID: 6t8b
TitleFtsK motor domain with dsDNA, translocating state
Components
  • DNA translocase FtsK
  • dsDNA substrate
KeywordsDNA BINDING PROTEIN / DNA translocation / DNA motor / RecA fold / Divisome
Function / homology
Function and homology information


cellular response to antibiotic / chromosome segregation / cell division / DNA binding / ATP binding / plasma membrane
Similarity search - Function
Threonyl-tRNA Synthetase; Chain A, domain 2 - #40 / FtsK gamma domain / DNA translocase FtsK, 4TM region / FtsK alpha domain / Ftsk gamma domain / 4TM region of DNA translocase FtsK/SpoIIIE / FtsK alpha domain / Ftsk_gamma / Threonyl-tRNA Synthetase; Chain A, domain 2 / FtsK domain ...Threonyl-tRNA Synthetase; Chain A, domain 2 - #40 / FtsK gamma domain / DNA translocase FtsK, 4TM region / FtsK alpha domain / Ftsk gamma domain / 4TM region of DNA translocase FtsK/SpoIIIE / FtsK alpha domain / Ftsk_gamma / Threonyl-tRNA Synthetase; Chain A, domain 2 / FtsK domain / FtsK/SpoIIIE family / FtsK domain profile. / Winged helix DNA-binding domain superfamily / P-loop containing nucleotide triphosphate hydrolases / Winged helix-like DNA-binding domain superfamily / P-loop containing nucleoside triphosphate hydrolase / Rossmann fold / 2-Layer Sandwich / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / DNA / DNA (> 10) / DNA translocase FtsK
Similarity search - Component
Biological speciesPseudomonas aeruginosa PAO1 (bacteria)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.65 Å
AuthorsJean, N.L. / Lowe, J.
Funding support United Kingdom, 2items
OrganizationGrant numberCountry
Wellcome Trust202754/Z/16/Z United Kingdom
European Molecular Biology OrganizationALTF-128-2016
Citation
Journal: Proc Natl Acad Sci U S A / Year: 2020
Title: FtsK in motion reveals its mechanism for double-stranded DNA translocation.
Authors: Nicolas L Jean / Trevor J Rutherford / Jan Löwe /
Abstract: FtsK protein contains a fast DNA motor that is involved in bacterial chromosome dimer resolution. During cell division, FtsK translocates double-stranded DNA until both recombination sites are ...FtsK protein contains a fast DNA motor that is involved in bacterial chromosome dimer resolution. During cell division, FtsK translocates double-stranded DNA until both recombination sites are placed at mid cell for subsequent dimer resolution. Here, we solved the 3.6-Å resolution electron cryo-microscopy structure of the motor domain of FtsK while translocating on its DNA substrate. Each subunit of the homo-hexameric ring adopts a unique conformation and one of three nucleotide states. Two DNA-binding loops within four subunits form a pair of spiral staircases within the ring, interacting with the two DNA strands. This suggests that simultaneous conformational changes in all ATPase domains at each catalytic step generate movement through a mechanism related to filament treadmilling. While the ring is only rotating around the DNA slowly, it is instead the conformational states that rotate around the ring as the DNA substrate is pushed through.
#1: Journal: Biorxiv
Title: FtsK in motion reveals its mechanism for double-stranded DNA translocation
Authors: Jean, N.L. / Rutherford, T.J. / Lowe, J.
History
DepositionOct 24, 2019Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 20, 2019Provider: repository / Type: Initial release
Revision 1.1Jun 24, 2020Group: Database references / Category: citation / citation_author
Revision 1.2Jul 1, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID

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Structure visualization

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  • Deposited structure unit
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Structure viewerMolecule:
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Assembly

Deposited unit
A: DNA translocase FtsK
B: DNA translocase FtsK
C: DNA translocase FtsK
D: DNA translocase FtsK
E: DNA translocase FtsK
F: DNA translocase FtsK
G: dsDNA substrate
H: dsDNA substrate
hetero molecules


Theoretical massNumber of molelcules
Total (without water)339,21518
Polymers336,2678
Non-polymers2,94910
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area26600 Å2
ΔGint-158 kcal/mol
Surface area95900 Å2
MethodPISA

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Components

#1: Protein
DNA translocase FtsK


Mass: 54001.441 Da / Num. of mol.: 6 / Fragment: Motor domain, residues 247-728
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa PAO1 (bacteria) / Gene: ftsK, PA2615 / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): C41 / References: UniProt: Q9I0M3
#2: DNA chain dsDNA substrate


Mass: 6129.039 Da / Num. of mol.: 2 / Source method: obtained synthetically
Details: Sequence not determined due to sequence heterogeneity of the protein-bound DNA
Source: (synth.) synthetic construct (others)
#3: Chemical ChemComp-AGS / PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / ATP-GAMMA-S / ADENOSINE 5'-(3-THIOTRIPHOSPHATE) / ADENOSINE 5'-(GAMMA-THIOTRIPHOSPHATE) / ADENOSINE-5'-DIPHOSPHATE MONOTHIOPHOSPHATE


Mass: 523.247 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C10H16N5O12P3S / Comment: ATP-gamma-S, energy-carrying molecule analogue*YM
#4: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Mg
#5: Chemical ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE / Adenosine diphosphate


Mass: 427.201 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Comment: ADP, 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

Component
IDNameTypeEntity IDParent-IDSource
1FtsK motor domain with dsDNA, translocating stateCOMPLEX#1-#20RECOMBINANT
2FtsK motor domainCOMPLEX#11RECOMBINANT
3DNACOMPLEX#21RECOMBINANT
Molecular weightUnits: MEGADALTONS / Experimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)208964
23synthetic construct (others)32630
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
12Escherichia coli BL21(DE3) (bacteria)469008
23synthetic construct (others)32630
Buffer solutionpH: 7.5
Buffer component
IDConc.NameBuffer-ID
125 mMTris1
22 mMATPgammaS1
34 mMMgCl21
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: 0.7 m/mL FtsK 1.5 uM 45 bp DNA
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
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: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 42.95 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 3300
EM imaging opticsEnergyfilter slit width: 20 eV

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Processing

EM software
IDNameVersionCategory
1RELION3particle selection
2EPUimage acquisition
4CTFFIND4.1CTF correction
9PHENIX1.14rc2-3139model refinement
10RELION3initial Euler assignment
11RELION3final Euler assignment
12RELION3classification
13RELION33D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.65 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 56485 / Symmetry type: POINT
Atomic model buildingPDB-ID: 2IUU

2iuu


Pdb chain-ID: A

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