+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-27252 | |||||||||
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Title | gRAMP non-match PFS target RNA | |||||||||
Map data | ||||||||||
Sample |
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Keywords | CRISPR / GRAMP / RNA BINDING PROTEIN-RNA complex | |||||||||
Function / homology | : / CRISPR type III-associated protein / RAMP superfamily / defense response to virus / RNA binding / RAMP superfamily protein Function and homology information | |||||||||
Biological species | Candidatus Scalindua brodae (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.6 Å | |||||||||
Authors | Hu C / Nam KH / Schuler G / Ke A | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Science / Year: 2022 Title: Craspase is a CRISPR RNA-guided, RNA-activated protease. Authors: Chunyi Hu / Sam P B van Beljouw / Ki Hyun Nam / Gabriel Schuler / Fran Ding / Yanru Cui / Alicia Rodríguez-Molina / Anna C Haagsma / Menno Valk / Martin Pabst / Stan J J Brouns / Ailong Ke / Abstract: The CRISPR-Cas type III-E RNA-targeting effector complex gRAMP/Cas7-11 is associated with a caspase-like protein (TPR-CHAT/Csx29) to form Craspase (CRISPR-guided caspase). Here, we use cryo-electron ...The CRISPR-Cas type III-E RNA-targeting effector complex gRAMP/Cas7-11 is associated with a caspase-like protein (TPR-CHAT/Csx29) to form Craspase (CRISPR-guided caspase). Here, we use cryo-electron microscopy snapshots of Craspase to explain its target RNA cleavage and protease activation mechanisms. Target-guide pairing extending into the 5' region of the guide RNA displaces a gating loop in gRAMP, which triggers an extensive conformational relay that allosterically aligns the protease catalytic dyad and opens an amino acid side-chain-binding pocket. We further define Csx30 as the endogenous protein substrate that is site-specifically proteolyzed by RNA-activated Craspase. This protease activity is switched off by target RNA cleavage by gRAMP and is not activated by RNA targets containing a matching protospacer flanking sequence. We thus conclude that Craspase is a target RNA-activated protease with self-regulatory capacity. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_27252.map.gz | 43.4 MB | EMDB map data format | |
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Header (meta data) | emd-27252-v30.xml emd-27252.xml | 18.3 KB 18.3 KB | Display Display | EMDB header |
Images | emd_27252.png | 85.5 KB | ||
Filedesc metadata | emd-27252.cif.gz | 6.7 KB | ||
Others | emd_27252_additional_1.map.gz emd_27252_half_map_1.map.gz emd_27252_half_map_2.map.gz | 43.6 MB 55.3 MB 55.3 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-27252 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-27252 | HTTPS FTP |
-Validation report
Summary document | emd_27252_validation.pdf.gz | 1 MB | Display | EMDB validaton report |
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Full document | emd_27252_full_validation.pdf.gz | 1 MB | Display | |
Data in XML | emd_27252_validation.xml.gz | 12.2 KB | Display | |
Data in CIF | emd_27252_validation.cif.gz | 14.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-27252 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-27252 | HTTPS FTP |
-Related structure data
Related structure data | 8d8nMC 8d97C 8d9eC 8d9fC 8d9gC 8d9hC 8d9iC M: atomic model generated by this map C: citing same article (ref.) |
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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_27252.map.gz / Format: CCP4 / Size: 59.6 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: 1.284 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Additional map: #1
File | emd_27252_additional_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_27252_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_27252_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Gramp-Non matching PFS target
Entire | Name: Gramp-Non matching PFS target |
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Components |
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-Supramolecule #1: Gramp-Non matching PFS target
Supramolecule | Name: Gramp-Non matching PFS target / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #2-#3 |
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Source (natural) | Organism: Candidatus Scalindua brodae (bacteria) |
-Macromolecule #1: RAMP superfamily protein
Macromolecule | Name: RAMP superfamily protein / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Candidatus Scalindua brodae (bacteria) |
Molecular weight | Theoretical: 197.219141 KDa |
Recombinant expression | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Sequence | String: MNITVELTFF EPYRLVEWFD WDARKKSHSA MRGQAFAQWT WKGKGRTAGK SFITGTLVRS AVIKAVEELL SLNNGKWEGV PCCNGSFQT DESKGKKPSF LRKRHTLQWQ ANNKNICDKE EACPFCILLG RFDNAGKVHE RNKDYDIHFS NFDLDHKQEK N DLRLVDIA ...String: MNITVELTFF EPYRLVEWFD WDARKKSHSA MRGQAFAQWT WKGKGRTAGK SFITGTLVRS AVIKAVEELL SLNNGKWEGV PCCNGSFQT DESKGKKPSF LRKRHTLQWQ ANNKNICDKE EACPFCILLG RFDNAGKVHE RNKDYDIHFS NFDLDHKQEK N DLRLVDIA SGRILNRVDF DTGKAKDYFR TWEADYETYG TYTGRITLRN EHAKKLLLAS LGFVDKLCGA LCRIEVIKKS ES PLPSDTK EQSYTKDDTV EVLSEDHNDE LRKQAEVIVE AFKQNDKLEK IRILADAIRT LRLHGEGVIE KDELPDGKEE RDK GHHLWD IKVQGTALRT KLKELWQSNK DIGWRKFTEM LGSNLYLIYK KETGGVSTRF RILGDTEYYS KAHDSEGSDL FIPV TPPEG IETKEWIIVG RLKAATPFYF GVQQPSDSIP GKEKKSEDSL VINEHTSFNI LLDKENRYRI PRSALRGALR RDLRT AFGS GCNVSLGGQI LCNCKVCIEM RRITLKDSVS DFSEPPEIRY RIAKNPGTAT VEDGSLFDIE VGPEGLTFPF VLRYRG HKF PEQLSSVIRY WEENDGKNGM AWLGGLDSTG KGRFALKDIK IFEWDLNQKI NEYIKERGMR GKEKELLEMG ESSLPDG LI PYKFFEEREC LFPYKENLKP QWSEVQYTIE VGSPLLTADT ISALTEPGNR DAIAYKKRVY NDGNNAIEPE PRFAVKSE T HRGIFRTAVG RRTGDLGKED HEDCTCDMCI IFGNEHESSK IRFEDLELIN GNEFEKLEKH IDHVAIDRFT GGALDKAKF DTYPLAGSPK KPLKLKGRFW IKKGFSGDHK LLITTALSDI RDGLYPLGSK GGVGYGWVAG ISIDDNVPDD FKEMINKTEM PLPEEVEES NNGPINNDYV HPGHQSPKQD HKNKNIYYPH YFLDSGSKVY REKDIITHEE FTEELLSGKI NCKLETLTPL I IPDTSDEN GLKLQGNKPG HKNYKFFNIN GELMIPGSEL RGMLRTHFEA LTKSCFAIFG EDSTLSWRMN ADEKDYKIDS NS IRKMESQ RNPKYRIPDE LQKELRNSGN GLFNRLYTSE RRFWSDVSNK FENSIDYKRE ILRCAGRPKN YKGGIIRQRK DSL MAEELK VHRLPLYDNF DIPDSAYKAN DHCRKSATCS TSRGCRERFT CGIKVRDKNR VFLNAANNNR QYLNNIKKSN HDLY LQYLK GEKKIRFNSK VITGSERSPI DVIAELNERG RQTGFIKLSG LNNSNKSQGN TGTTFNSGWD RFELNILLDD LETRP SKSD YPRPRLLFTK DQYEYNITKR CERVFEIDKG NKTGYPVDDQ IKKNYEDILD SYDGIKDQEV AERFDTFTRG SKLKVG DLV YFHIDGDNKI DSLIPVRISR KCASKTLGGK LDKALHPCTG LSDGLCPGCH LFGTTDYKGR VKFGFAKYEN GPEWLIT RG NNPERSLTLG VLESPRPAFS IPDDESEIPG RKFYLHHNGW RIIRQKQLEI RETVQPERNV TTEVMDKGNV FSFDVKFE N LREWELGLLL QSLDPGKNIA HKLGKGKPYG FGSVKIKIDS LHTFKINSNN DKIKRVPQSD IREYINKGYQ KLIEWSGNN SIQKGNVLPQ WHVIPHIDKL YKLLWVPFLN DSKLEPDVRY PVLNEESKGY IEGSDYTYKK LGDKDNLPYK TRVKGLTTPW SPWNPFQVI AEHEEQEVNV TGSRPSVTDK IERDGKMV UniProtKB: RAMP superfamily protein |
-Macromolecule #2: RNA (35-MER)
Macromolecule | Name: RNA (35-MER) / type: rna / ID: 2 / Number of copies: 1 |
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Source (natural) | Organism: Candidatus Scalindua brodae (bacteria) |
Molecular weight | Theoretical: 11.078582 KDa |
Sequence | String: ACUUAAUGUC ACGGUACCCA AUUUUCUGCC CCGGA |
-Macromolecule #3: RNA (5'-R(P*UP*CP*CP*GP*GP*GP*GP*CP*AP*GP*AP*AP*AP*AP*UP*UP*GP*GP...
Macromolecule | Name: RNA (5'-R(P*UP*CP*CP*GP*GP*GP*GP*CP*AP*GP*AP*AP*AP*AP*UP*UP*GP*GP*AP*CP*A)-3') type: rna / ID: 3 / Number of copies: 1 |
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Source (natural) | Organism: Candidatus Scalindua brodae (bacteria) |
Molecular weight | Theoretical: 6.815149 KDa |
Sequence | String: UCCGGGGCAG AAAAUUGGAC A |
-Macromolecule #4: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 4 / Number of copies: 4 / Formula: ZN |
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Molecular weight | Theoretical: 65.409 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TALOS ARCTICA |
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Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 59.0 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm |
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: NONE |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 89342 |
Initial angle assignment | Type: NOT APPLICABLE |
Final angle assignment | Type: NOT APPLICABLE |