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Open data
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Basic information
Entry | Database: PDB / ID: 6e9f | |||||||||
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Title | EsCas13d-crRNA-target RNA ternary complex | |||||||||
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![]() | RNA BINDING PROTEIN/RNA / CRISPR-Cas / RNase / Complex / RNA BINDING PROTEIN-RNA complex | |||||||||
Function / homology | RNA / RNA (> 10) / Uncharacterized protein![]() | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å | |||||||||
![]() | Zhang, C. / Lyumkis, D. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural Basis for the RNA-Guided Ribonuclease Activity of CRISPR-Cas13d. Authors: Cheng Zhang / Silvana Konermann / Nicholas J Brideau / Peter Lotfy / Xuebing Wu / Scott J Novick / Timothy Strutzenberg / Patrick R Griffin / Patrick D Hsu / Dmitry Lyumkis / ![]() Abstract: CRISPR-Cas endonucleases directed against foreign nucleic acids mediate prokaryotic adaptive immunity and have been tailored for broad genetic engineering applications. Type VI-D CRISPR systems ...CRISPR-Cas endonucleases directed against foreign nucleic acids mediate prokaryotic adaptive immunity and have been tailored for broad genetic engineering applications. Type VI-D CRISPR systems contain the smallest known family of single effector Cas enzymes, and their signature Cas13d ribonuclease employs guide RNAs to cleave matching target RNAs. To understand the molecular basis for Cas13d function and explain its compact molecular architecture, we resolved cryoelectron microscopy structures of Cas13d-guide RNA binary complex and Cas13d-guide-target RNA ternary complex to 3.4 and 3.3 Å resolution, respectively. Furthermore, a 6.5 Å reconstruction of apo Cas13d combined with hydrogen-deuterium exchange revealed conformational dynamics that have implications for RNA scanning. These structures, together with biochemical and cellular characterization, provide insights into its RNA-guided, RNA-targeting mechanism and delineate a blueprint for the rational design of improved transcriptome engineering technologies. | |||||||||
History |
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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 | ![]() | 207 KB | Display | ![]() |
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PDB format | ![]() | 157.6 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 9014MC ![]() 9013C ![]() 9015C ![]() 6e9eC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 110522.570 Da / Num. of mol.: 1 / Mutation: R295A, H300A, R849A, H854A Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: EUBSIR_02687 / Production host: ![]() ![]() |
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#2: RNA chain | Mass: 16715.051 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() |
#3: RNA chain | Mass: 8574.064 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() |
#4: Chemical | ChemComp-MG / |
-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: Ternary complex of EsCas13d(R295A/H300A/R849A/H854A) with crRNA, target RNA and Mg2+ Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT | |||||||||||||||||||||||||||||||||||
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Molecular weight | Value: 0.135 MDa / Experimental value: NO | |||||||||||||||||||||||||||||||||||
Source (natural) | Organism: ![]() | |||||||||||||||||||||||||||||||||||
Source (recombinant) | Organism: ![]() ![]() | |||||||||||||||||||||||||||||||||||
Buffer solution | pH: 7.5 | |||||||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||||||||||||||||||||||
Specimen support | Details: unspecified | |||||||||||||||||||||||||||||||||||
Vitrification | Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 80 % |
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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 magnification: 39000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 57.3 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 2205 |
Image scans | Width: 3838 / Height: 3710 / Movie frames/image: 60 / Used frames/image: 1-60 |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 51885 / Symmetry type: POINT | ||||||||||||||||||||||||||||||
Atomic model building | Protocol: AB INITIO MODEL / Space: REAL / Target criteria: Correlation coefficient |