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Yorodumi- EMDB-19690: DNA-bound Type IV-A3 CRISPR effector in complex with DinG helicas... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-19690 | |||||||||||||||
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Title | DNA-bound Type IV-A3 CRISPR effector in complex with DinG helicase from K. pneumoniae (state III) | |||||||||||||||
Map data | ||||||||||||||||
Sample |
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Keywords | CRISPR / crRNA / DNA binding / type IV CRISPR-Cas / CRISPRi / nuclease deficient / GENE REGULATION / ANTIVIRAL PROTEIN | |||||||||||||||
Function / homology | Function and homology information hydrolase activity, acting on acid anhydrides, in phosphorus-containing anhydrides / DNA 5'-3' helicase / nucleobase-containing compound metabolic process / DNA helicase activity / nucleic acid binding / ATP binding Similarity search - Function | |||||||||||||||
Biological species | Klebsiella pneumoniae (bacteria) | |||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.9 Å | |||||||||||||||
Authors | Skorupskaite A / Ragozius V / Cepaite R / Klein N / Randau L / Malinauskaite L / Pausch P | |||||||||||||||
Funding support | European Union, 4 items
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Citation | Journal: Nat Commun / Year: 2024 Title: Structural variation of types IV-A1- and IV-A3-mediated CRISPR interference. Authors: R Čepaitė / N Klein / A Mikšys / S Camara-Wilpert / V Ragožius / F Benz / A Skorupskaitė / H Becker / G Žvejytė / N Steube / G K A Hochberg / L Randau / R Pinilla-Redondo / L ...Authors: R Čepaitė / N Klein / A Mikšys / S Camara-Wilpert / V Ragožius / F Benz / A Skorupskaitė / H Becker / G Žvejytė / N Steube / G K A Hochberg / L Randau / R Pinilla-Redondo / L Malinauskaitė / P Pausch / Abstract: CRISPR-Cas mediated DNA-interference typically relies on sequence-specific binding and nucleolytic degradation of foreign genetic material. Type IV-A CRISPR-Cas systems diverge from this general ...CRISPR-Cas mediated DNA-interference typically relies on sequence-specific binding and nucleolytic degradation of foreign genetic material. Type IV-A CRISPR-Cas systems diverge from this general mechanism, using a nuclease-independent interference pathway to suppress gene expression for gene regulation and plasmid competition. To understand how the type IV-A system associated effector complex achieves this interference, we determine cryo-EM structures of two evolutionarily distinct type IV-A complexes (types IV-A1 and IV-A3) bound to cognate DNA-targets in the presence and absence of the type IV-A signature DinG effector helicase. The structures reveal how the effector complexes recognize the protospacer adjacent motif and target-strand DNA to form an R-loop structure. Additionally, we reveal differences between types IV-A1 and IV-A3 in DNA interactions and structural motifs that allow for in trans recruitment of DinG. Our study provides a detailed view of type IV-A mediated DNA-interference and presents a structural foundation for engineering type IV-A-based genome editing tools. | |||||||||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_19690.map.gz | 90.7 MB | EMDB map data format | |
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Header (meta data) | emd-19690-v30.xml emd-19690.xml | 30.9 KB 30.9 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_19690_fsc.xml | 11.8 KB | Display | FSC data file |
Images | emd_19690.png | 60.6 KB | ||
Filedesc metadata | emd-19690.cif.gz | 8.4 KB | ||
Others | emd_19690_additional_1.map.gz emd_19690_half_map_1.map.gz emd_19690_half_map_2.map.gz | 168.1 MB 165.2 MB 165.2 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-19690 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-19690 | HTTPS FTP |
-Validation report
Summary document | emd_19690_validation.pdf.gz | 995.9 KB | Display | EMDB validaton report |
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Full document | emd_19690_full_validation.pdf.gz | 995.5 KB | Display | |
Data in XML | emd_19690_validation.xml.gz | 20.5 KB | Display | |
Data in CIF | emd_19690_validation.cif.gz | 26.5 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-19690 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-19690 | HTTPS FTP |
-Related structure data
Related structure data | 8s37MC 8rc2C 8rc3C 8rfjC 8s35C 8s36C C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_19690.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.1 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Additional map: Volume map
File | emd_19690_additional_1.map | ||||||||||||
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Annotation | Volume map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map A
File | emd_19690_half_map_1.map | ||||||||||||
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Annotation | Half map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map B
File | emd_19690_half_map_2.map | ||||||||||||
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Annotation | Half map B | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : Type IV-A3 CRISPR-Cas effector complex with DinG from Klebsiella ...
+Supramolecule #1: Type IV-A3 CRISPR-Cas effector complex with DinG from Klebsiella ...
+Supramolecule #2: CRISPR effector in complex with a DinG helicase and crRNA
+Supramolecule #3: Double-stranded DNA
+Macromolecule #1: CRISPR type AFERR-associated protein Csf2
+Macromolecule #2: CRISPR type AFERR-associated protein Csf3
+Macromolecule #3: CRISPR type AFERR-associated protein Csf1
+Macromolecule #7: DEAD/DEAH box helicase
+Macromolecule #4: crRNA
+Macromolecule #5: TS-DNA
+Macromolecule #6: NTS-DNA
+Macromolecule #8: ZINC ION
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.5 mg/mL | ||||||
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Buffer | pH: 7.5 / Component:
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Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 7 sec. / Pretreatment - Atmosphere: AIR / Details: 20 mA current | ||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | TFS GLACIOS |
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Software | Name: EPU (ver. 3.5.1) |
Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 4350 / Average exposure time: 46.33 sec. / Average electron dose: 30.31 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated defocus max: 2.0 µm / Calibrated defocus min: 1.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 92000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |