+
Open data
-
Basic information
Entry | Database: PDB / ID: 7n6i | ||||||
---|---|---|---|---|---|---|---|
Title | ATP-bound TnsC-TniQ complex from ShCAST system | ||||||
![]() |
| ||||||
![]() | DNA BINDING PROTEIN/DNA / ![]() | ||||||
Function / homology | ADENOSINE-5'-TRIPHOSPHATE / ![]() ![]() | ||||||
Biological species | ![]() synthetic construct (others) | ||||||
Method | ![]() ![]() ![]() | ||||||
![]() | Park, J. / Tsai, A.W.L. / Mehrotra, E. / Kellogg, E.H. | ||||||
Funding support | ![]()
| ||||||
![]() | ![]() Title: Structural basis for target site selection in RNA-guided DNA transposition systems. Authors: Jung-Un Park / Amy Wei-Lun Tsai / Eshan Mehrotra / Michael T Petassi / Shan-Chi Hsieh / Ailong Ke / Joseph E Peters / Elizabeth H Kellogg / ![]() Abstract: CRISPR-associated transposition systems allow guide RNA-directed integration of a single DNA cargo in one orientation at a fixed distance from a programmable target sequence. We used cryo-electron ...CRISPR-associated transposition systems allow guide RNA-directed integration of a single DNA cargo in one orientation at a fixed distance from a programmable target sequence. We used cryo-electron microscopy (cryo-EM) to define the mechanism that underlies this process by characterizing the transposition regulator, TnsC, from a type V-K CRISPR-transposase system. In this scenario, polymerization of adenosine triphosphate-bound TnsC helical filaments could explain how polarity information is passed to the transposase. TniQ caps the TnsC filament, representing a universal mechanism for target information transfer in Tn7/Tn7-like elements. Transposase-driven disassembly establishes delivery of the element only to unused protospacers. Finally, TnsC transitions to define the fixed point of insertion, as revealed by structures with the transition state mimic ADP•AlF These mechanistic findings provide the underpinnings for engineering CRISPR-associated transposition systems for research and therapeutic applications. | ||||||
History |
|
-
Structure visualization
Movie |
![]() |
---|---|
Structure viewer | Molecule: ![]() ![]() |
-
Downloads & links
-
Download
PDBx/mmCIF format | ![]() | 411.5 KB | Display | ![]() |
---|---|---|---|---|
PDB format | ![]() | 322.9 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
---|
-Related structure data
Related structure data | ![]() 23726MC ![]() 7m99C ![]() 7m9aC ![]() 7m9bC ![]() 7m9cC C: citing same article ( M: map data used to model this data |
---|---|
Similar structure data |
-
Links
-
Assembly
Deposited unit | ![]()
|
---|---|
1 |
|
-
Components
-Protein , 2 types, 10 molecules ABCDEFGHIJ
#1: Protein | Mass: 19011.240 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() #2: Protein | Mass: 31444.617 Da / Num. of mol.: 8 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
---|
-DNA chain , 2 types, 2 molecules KL
#3: DNA chain | Mass: 5126.320 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
---|---|
#4: DNA chain | Mass: 5279.559 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
-Non-polymers , 2 types, 14 molecules ![](data/chem/img/ATP.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/MG.gif)
#5: Chemical | ChemComp-ATP / ![]() #6: Chemical | ChemComp-MG / |
---|
-Details
Has ligand of interest | N |
---|
-Experimental details
-Experiment
Experiment | Method: ![]() |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: ![]() |
-
Sample preparation
Component | Name: ATP-bound TnsC-TniQ complex from ShCAST system / Type: COMPLEX / Entity ID: #1-#4 / Source: MULTIPLE SOURCES | |||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Molecular weight | Value: 0.4 MDa / Experimental value: NO | |||||||||||||||||||||||||||||||||||
Source (natural) | Organism: ![]() | |||||||||||||||||||||||||||||||||||
Source (recombinant) | Organism: ![]() ![]() ![]() | |||||||||||||||||||||||||||||||||||
Buffer solution | pH: 7.4 | |||||||||||||||||||||||||||||||||||
Buffer component |
| |||||||||||||||||||||||||||||||||||
Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied![]() ![]() Details: Excess of TniQ was added to ATP-bound TnsC filaments | |||||||||||||||||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 400 divisions/in. / Grid type: UltrAuFoil R2/2 | |||||||||||||||||||||||||||||||||||
Vitrification![]() | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K / Details: Blot for 7 seconds before plunging |
-
Electron microscopy imaging
Experimental equipment | ![]() Model: Talos Arctica / Image courtesy: FEI Company |
---|---|
Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source![]() ![]() |
Electron lens | Mode: BRIGHT FIELD![]() ![]() |
Specimen holder | Cryogen: HELIUM / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 3 sec. / Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4172 |
Image scans | Width: 5760 / Height: 4092 |
-
Processing
EM software |
| ||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
CTF correction![]() | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 648550 / Details: Neural net based particle picking | ||||||||||||||||||||||||||||||||
Symmetry | Point symmetry![]() | ||||||||||||||||||||||||||||||||
3D reconstruction![]() | Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 34546 / Algorithm: BACK PROJECTION / Symmetry type: POINT | ||||||||||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT Details: Homology model was built using the crystal structure of TniQ (PDB:6V9P) by GalaxyTBM server. This model is manually docked into the cryo-EM density using UCSF Chimera. | ||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 6V9P Pdb chain-ID: A / Accession code: 6V9P / Source name: PDB / Type: experimental model |