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Open data
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Basic information
Entry | Database: PDB / ID: 7kft | ||||||
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Title | Partial Cas6-RT-Cas1--Cas2 complex | ||||||
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![]() | HYDROLASE / Complex / CRISPR Cas Protein / Reverse Transcriptase | ||||||
Biological species | ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||
![]() | Hoel, C.M. / Wang, J.Y. / Doudna, J.A. / Brohawn, S.G. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Structural coordination between active sites of a CRISPR reverse transcriptase-integrase complex. Authors: Joy Y Wang / Christopher M Hoel / Basem Al-Shayeb / Jillian F Banfield / Stephen G Brohawn / Jennifer A Doudna / ![]() Abstract: CRISPR-Cas systems provide adaptive immunity in bacteria and archaea, beginning with integration of foreign sequences into the host CRISPR genomic locus and followed by transcription and maturation ...CRISPR-Cas systems provide adaptive immunity in bacteria and archaea, beginning with integration of foreign sequences into the host CRISPR genomic locus and followed by transcription and maturation of CRISPR RNAs (crRNAs). In some CRISPR systems, a reverse transcriptase (RT) fusion to the Cas1 integrase and Cas6 maturase creates a single protein that enables concerted sequence integration and crRNA production. To elucidate how the RT-integrase organizes distinct enzymatic activities, we present the cryo-EM structure of a Cas6-RT-Cas1-Cas2 CRISPR integrase complex. The structure reveals a heterohexamer in which the RT directly contacts the integrase and maturase domains, suggesting functional coordination between all three active sites. Together with biochemical experiments, our data support a model of sequential enzymatic activities that enable CRISPR sequence acquisition from RNA and DNA substrates. These findings highlight an expanded capacity of some CRISPR systems to acquire diverse sequences that direct CRISPR-mediated interference. | ||||||
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Structure visualization
Movie |
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 297.2 KB | Display | ![]() |
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PDB format | ![]() | 218 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 915.2 KB | Display | ![]() |
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Full document | ![]() | 926 KB | Display | |
Data in XML | ![]() | 42.3 KB | Display | |
Data in CIF | ![]() | 64.1 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 22855MC ![]() 7kfuC M: map data used to model this data C: citing same article ( |
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Similar structure data | |
EM raw data | ![]() Data size: 2.2 TB Data #1: Unaligned multi-frame movies of a Cas6-reverse transcriptase-Cas1—Cas2 CRISPR integrase complex [micrographs - multiframe]) |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 11583.263 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() #2: Protein | Mass: 112869.414 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
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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: Cas6-RT-Cas1--Cas2 complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Value: 0.47 MDa / Experimental value: NO |
Source (natural) | Organism: ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 7.5 |
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 R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K / Details: Blot force 1, 3 second blot |
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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 |
Image recording | Electron dose: 49.9 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
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Processing
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 129175 / Symmetry type: POINT | ||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||
Displacement parameters | Biso mean: 57.06 Å2 | ||||||||||||||||||||||||
Refine LS restraints |
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