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Yorodumi- PDB-8glw: CryoEM structure of the TnsC(1-503)-TnsD(1-318)-DNA complex in a ... -
+Open data
-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 | ||||||
Components |
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Keywords | DNA BINDING PROTEIN/DNA / Transposon / AAA+ ATPase / Oligomer / Complex / DNA BINDING PROTEIN / DNA BINDING PROTEIN-DNA complex | ||||||
Function / homology | Function and homology information transposition / DNA recombination / ATP hydrolysis activity / DNA binding / ATP binding Similarity search - Function | ||||||
Biological species | Escherichia coli (E. coli) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.51 Å | ||||||
Authors | Shen, Y. / Guarne, A. | ||||||
Funding support | Canada, 1items
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Citation | Journal: Mol Cell / Year: 2024 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: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8glw.cif.gz | 701.2 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8glw.ent.gz | 574.2 KB | Display | PDB format |
PDBx/mmJSON format | 8glw.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8glw_validation.pdf.gz | 1.7 MB | Display | wwPDB validaton report |
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Full document | 8glw_full_validation.pdf.gz | 1.7 MB | Display | |
Data in XML | 8glw_validation.xml.gz | 114.8 KB | Display | |
Data in CIF | 8glw_validation.cif.gz | 171 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/gl/8glw ftp://data.pdbj.org/pub/pdb/validation_reports/gl/8glw | HTTPS FTP |
-Related structure data
Related structure data | 40221MC 8gluC 8glxC 8vcjC 8vctC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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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.) Escherichia coli (E. coli) / Gene: tnsC / Production host: Escherichia coli (E. coli) / References: UniProt: P05846 #2: Protein | Mass: 36636.020 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: tnsD / Production host: Escherichia coli (E. coli) / References: UniProt: P13991 |
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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.) Escherichia coli (E. coli) |
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#4: DNA chain | Mass: 15366.818 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
-Non-polymers , 4 types, 16 molecules
#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 |
-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: Escherichia coli (E. coli) |
Source (recombinant) | Organism: Escherichia coli (E. coli) |
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 |
-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: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
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) |
-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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