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Open data
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Basic information
Entry | Database: PDB / ID: 8q3w | |||||||||
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Title | ATP-bound IstB in complex to duplex DNA | |||||||||
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![]() | DNA BINDING PROTEIN / DNA Transposition / transposon / transpososome / AAA+ ATPases / target DNA / IS21 / IstB / Insertion sequence / cryo-electron microscopy. | |||||||||
Function / homology | ![]() | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.18 Å | |||||||||
![]() | de la Gandara, A. / Spinola-Amilibia, M. / Araujo-Bazan, L. / Nunez-Ramirez, R. / Berger, J.M. / Arias-Palomo, E. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Molecular basis for transposase activation by a dedicated AAA+ ATPase. Authors: Álvaro de la Gándara / Mercedes Spínola-Amilibia / Lidia Araújo-Bazán / Rafael Núñez-Ramírez / James M Berger / Ernesto Arias-Palomo / ![]() ![]() Abstract: Transposases drive chromosomal rearrangements and the dissemination of drug-resistance genes and toxins. Although some transposases act alone, many rely on dedicated AAA+ ATPase subunits that ...Transposases drive chromosomal rearrangements and the dissemination of drug-resistance genes and toxins. Although some transposases act alone, many rely on dedicated AAA+ ATPase subunits that regulate site selectivity and catalytic function through poorly understood mechanisms. Using IS21 as a model transposase system, we show how an ATPase regulator uses nucleotide-controlled assembly and DNA deformation to enable structure-based site selectivity, transposase recruitment, and activation and integration. Solution and cryogenic electron microscopy studies show that the IstB ATPase self-assembles into an autoinhibited pentamer of dimers that tightly curves target DNA into a half-coil. Two of these decamers dimerize, which stabilizes the target nucleic acid into a kinked S-shaped configuration that engages the IstA transposase at the interface between the two IstB oligomers to form an approximately 1 MDa transpososome complex. Specific interactions stimulate regulator ATPase activity and trigger a large conformational change on the transposase that positions the catalytic site to perform DNA strand transfer. These studies help explain how AAA+ ATPase regulators-which are used by classical transposition systems such as Tn7, Mu and CRISPR-associated elements-can remodel their substrate DNA and cognate transposases to promote function. | |||||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 903.5 KB | Display | ![]() |
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PDB format | ![]() | 756.5 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.8 MB | Display | ![]() |
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Full document | ![]() | 1.8 MB | Display | |
Data in XML | ![]() | 78.4 KB | Display | |
Data in CIF | ![]() | 113.2 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 18136MC ![]() 8q4dC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 29602.275 Da / Num. of mol.: 10 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Plasmid: pHMKS Details (production host): TEV cleavable MBP-His6 tag at the N-terminus Production host: ![]() ![]() #2: DNA chain | | Mass: 18401.746 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() #3: DNA chain | | Mass: 18584.918 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() #4: Chemical | ChemComp-MG / #5: Chemical | ChemComp-ATP / Has ligand of interest | Y | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
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Molecular weight | Value: 0.337 MDa / Experimental value: NO | |||||||||||||||||||||||||||||||||||
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Buffer solution | pH: 7.5 | |||||||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1 | |||||||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 298 K |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2700 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 5 sec. / Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
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Processing
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Image processing | Details: Super-resolution counting mode | ||||||||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 2439865 | ||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.18 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 306745 / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 95.21 / Protocol: AB INITIO MODEL / Space: REAL | ||||||||||||||||||||||||||||||||||||
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