+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-9015 | |||||||||
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Title | Apo-EsCas13d | |||||||||
Map data | Apo-EsCas13d | |||||||||
Sample |
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Biological species | unidentified bacterium (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 6.5 Å | |||||||||
Authors | Zhang C / Lyumkis D | |||||||||
Funding support | United States, 2 items
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Citation | Journal: Cell / Year: 2018 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 |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_9015.map.gz | 56.3 MB | EMDB map data format | |
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Header (meta data) | emd-9015-v30.xml emd-9015.xml | 20.9 KB 20.9 KB | Display Display | EMDB header |
Images | emd_9015.png | 77.1 KB | ||
Others | emd_9015_additional.map.gz emd_9015_half_map_1.map.gz emd_9015_half_map_2.map.gz | 1 MB 11.1 MB 11.2 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-9015 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-9015 | HTTPS FTP |
-Validation report
Summary document | emd_9015_validation.pdf.gz | 78.4 KB | Display | EMDB validaton report |
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Full document | emd_9015_full_validation.pdf.gz | 77.5 KB | Display | |
Data in XML | emd_9015_validation.xml.gz | 492 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-9015 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-9015 | HTTPS FTP |
-Related structure data
Related structure data | 9013C 9014C 6e9eC 6e9fC C: citing same article (ref.) |
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Similar structure data | |
EM raw data | EMPIAR-10229 (Title: Cryo-EM Reconstruction of apo EsCas13d / Data size: 66.9 Data #1: stack of ~150k particles used for ab initio orientation assignment and reconstruction [picked particles - single frame - unprocessed] Data #2: stack of ~16k best particles from above [picked particles - single frame - processed]) |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_9015.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Apo-EsCas13d | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.79 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Additional map: Apo-EsCas13d, additional map
File | emd_9015_additional.map | ||||||||||||
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Annotation | Apo-EsCas13d, additional map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Apo-EsCas13d, half map #1
File | emd_9015_half_map_1.map | ||||||||||||
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Annotation | Apo-EsCas13d, half map #1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Apo-EsCas13d, half map #2
File | emd_9015_half_map_2.map | ||||||||||||
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Annotation | Apo-EsCas13d, half map #2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Apo-EsCas13d
Entire | Name: Apo-EsCas13d |
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Components |
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-Supramolecule #1: Apo-EsCas13d
Supramolecule | Name: Apo-EsCas13d / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: unidentified bacterium (bacteria) |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 110 KDa |
-Macromolecule #1: Apo-esCas13d
Macromolecule | Name: Apo-esCas13d / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: unidentified bacterium (bacteria) |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MGKKIHARDL REQRKTDRTE KFADQNKKRE AERAVPKKDA AVSVKSVSSV SSKKDNVTKS MAKAAGVKSV FAVGNTVYMT SFGRGNDAVL EQKIVDTSHE PLNIDDPAYQ LNVVTMNGYS VTGHRGETVS AVTDNPLRRF NGRKKDEPEQ SVPTDMLCLK PTLEKKFFGK ...String: MGKKIHARDL REQRKTDRTE KFADQNKKRE AERAVPKKDA AVSVKSVSSV SSKKDNVTKS MAKAAGVKSV FAVGNTVYMT SFGRGNDAVL EQKIVDTSHE PLNIDDPAYQ LNVVTMNGYS VTGHRGETVS AVTDNPLRRF NGRKKDEPEQ SVPTDMLCLK PTLEKKFFGK EFDDNIHIQL IYNILDIEKI LAVYSTNAIY ALNNMSADEN IENSDFFMKR TTDETFDDFE KKKESTNSRE KADFDAFEKF IGNYRLAYFA DAFYVNKKNP KGKAKNVLRE DKELYSVLTL IGKLRHWCVH SEEGRAEFWL YKLDELKDDF KNVLDVVYNR PVEEINNRFI ENNKVNIQIL GSVYKNTDIA ELVRSYYEFL ITKKYKNMGF SIKKLRESML EGKGYADKEY DSVRNKLYQM TDFILYTGYI NEDSDRADDL VNTLRSSLKE DDKTTVYCKE ADYLWKKYRE SIREVADALD GDNIKKLSKS NIEIQEDKLR KCFISYADSV SEFTKLIYLL TRFLSGKEIN DLVTTLINKF DNIRSFLEIM DELGLDRTFT AEYSFFEGST KYLAELVELN SFVKSCSFDI NAKRTMYRDA LDILGIESDK TEEDIEKMID NILQIDANGD KKLKKNNGLR NFIASNVIDS NRFKYLVRYG NPKKIRETAK CKPAVRFVLN EIPDAQIERY YEACCPKNTA LCSANKRREK LADMIAEIKF ENFSDAGNYQ KANVTSRTSE AEIKRKNQAI IRLYLTVMYI MLKNLVNVNA RYVIAFHCVE RDTKLYAESG LEVGNIEKNK TNLTMAVMGV KLENGIIKTE FDKSFAENAA NRYLRNARWY KLILDNLKKS ERAVVNEFRN TVCHLNAIRN ININIKEIKE VENYFALYHY LIQKHLENRF ADKKVERDTG DFISKLEEHK TYCKDFVKAY CTPFGYNLVR YKNLTIDGLF DKNYPGKDDS DEQK |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1 mg/mL | |||||||||||||||||||||
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Buffer | pH: 7.5 Component:
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Grid | Pretreatment - Type: PLASMA CLEANING / Pretreatment - Atmosphere: OTHER / Details: unspecified | |||||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 80 % / Instrument: HOMEMADE PLUNGER |
-Electron microscopy
Microscope | FEI TALOS ARCTICA |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Digitization - Frames/image: 1-120 / Number real images: 1158 / Average electron dose: 56.8 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 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: 57000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: AB INITIO MODEL / Target criteria: Correlation coefficient |
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