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- PDB-8glw: CryoEM structure of the TnsC(1-503)-TnsD(1-318)-DNA complex in a ... -
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Open data
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Basic information
Entry | Database: PDB / ID: 8glw | ||||||
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Title | CryoEM structure of the TnsC(1-503)-TnsD(1-318)-DNA complex in a 7:2:1 stoichiometry from E. coli Tn7 | ||||||
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![]() | DNA BINDING PROTEIN/DNA / Transposon / AAA+ ATPase / Oligomer / Complex / DNA BINDING PROTEIN / DNA BINDING PROTEIN-DNA complex | ||||||
Function / homology | ![]() transposition / DNA recombination / ATP hydrolysis activity / DNA binding / ATP binding Similarity search - Function | ||||||
Biological species | ![]() ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.51 Å | ||||||
![]() | Shen, Y. / Guarne, A. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Assembly of the Tn7 targeting complex by a regulated stepwise process. Authors: Yao Shen / Shreya S Krishnan / Michael T Petassi / Mark A Hancock / Joseph E Peters / Alba Guarné / ![]() ![]() Abstract: The Tn7 family of transposons is notable for its highly regulated integration mechanisms, including programmable RNA-guided transposition. The targeting pathways rely on dedicated target selection ...The Tn7 family of transposons is notable for its highly regulated integration mechanisms, including programmable RNA-guided transposition. The targeting pathways rely on dedicated target selection proteins from the TniQ family and the AAA+ adaptor TnsC to recruit and activate the transposase at specific target sites. Here, we report the cryoelectron microscopy (cryo-EM) structures of TnsC bound to the TniQ domain of TnsD from prototypical Tn7 and unveil key regulatory steps stemming from unique behaviors of ATP- versus ADP-bound TnsC. We show that TnsD recruits ADP-bound dimers of TnsC and acts as an exchange factor to release one protomer with exchange to ATP. This loading process explains how TnsC assembles a heptameric ring unidirectionally from the target site. This unique loading process results in functionally distinct TnsC protomers within the ring, providing a checkpoint for target immunity and explaining how insertions at programmed sites precisely occur in a specific orientation across Tn7 elements. | ||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 701.2 KB | Display | ![]() |
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PDB format | ![]() | 574.2 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.7 MB | Display | ![]() |
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Full document | ![]() | 1.7 MB | Display | |
Data in XML | ![]() | 114.8 KB | Display | |
Data in CIF | ![]() | 171 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 40221MC ![]() 8gluC ![]() 8glxC ![]() 8vcjC ![]() 8vctC C: citing same article ( M: map data used to model this data |
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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
-Transposon Tn7 transposition protein ... , 2 types, 9 molecules ABCDEFGYX
#1: Protein | Mass: 59318.926 Da / Num. of mol.: 7 / Mutation: S2G Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() #2: Protein | Mass: 36636.020 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
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-DNA chain , 2 types, 2 molecules HI
#3: DNA chain | Mass: 15438.931 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
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#4: DNA chain | Mass: 15366.818 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
-Non-polymers , 4 types, 16 molecules ![](data/chem/img/ADP.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/ATP.gif)
![](data/chem/img/ZN.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/ATP.gif)
![](data/chem/img/ZN.gif)
#5: Chemical | ChemComp-ADP / #6: Chemical | ChemComp-MG / #7: Chemical | #8: Chemical | |
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-Details
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
Component | Name: Complex of TnsC(1-503) bound to TnsD(1-318) and DNA with cofactors ATP and ADP Type: COMPLEX / Entity ID: #1-#4 / Source: RECOMBINANT |
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Molecular weight | Value: 0.55 MDa / Experimental value: YES |
Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 8 Details: 20 mM Tris pH 8.0, 150 mM NaCl, 1.4 mM beta-mercaptoethanol, 5 mM MgCl2 |
Specimen | Conc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: C-flat-2/1 |
Vitrification | Instrument: 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 |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2750 nm / Nominal defocus min: 1250 nm |
Image recording | Electron dose: 79 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
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Processing
EM software | Name: PHENIX / Version: 1.19.2_4158: / Category: model refinement | ||||||||||||||||||||||||
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CTF correction | Type: NONE | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.51 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 364564 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: OTHER / Space: REAL | ||||||||||||||||||||||||
Atomic model building | PDB-ID: 8GLU Accession code: 8GLU / Details: TnsC and TnsD were modelled using PDB 8GLU. / Source name: PDB / Type: experimental model | ||||||||||||||||||||||||
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