+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-33983 | |||||||||
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Title | Structure of CasPi with guide RNA and target DNA | |||||||||
Map data | ||||||||||
Sample |
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Keywords | CasPi complex / RNA BINDING PROTEIN / RNA BINDING PROTEIN-DNA-RNA complex | |||||||||
Function / homology | Reverse transcriptase domain-containing protein Function and homology information | |||||||||
Biological species | Armatimonadetes bacterium (bacteria) / Armatimonadota bacterium (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.36 Å | |||||||||
Authors | Li CP / Wang J / Liu JJ / Zhang S | |||||||||
Funding support | China, 1 items
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Citation | Journal: Cell Res / Year: 2023 Title: The compact Casπ (Cas12l) 'bracelet' provides a unique structural platform for DNA manipulation. Authors: Ao Sun / Cheng-Ping Li / Zhihang Chen / Shouyue Zhang / Dan-Yuan Li / Yun Yang / Long-Qi Li / Yuqian Zhao / Kaichen Wang / Zhaofu Li / Jinxia Liu / Sitong Liu / Jia Wang / Jun-Jie Gogo Liu / Abstract: CRISPR-Cas modules serve as the adaptive nucleic acid immune systems for prokaryotes, and provide versatile tools for nucleic acid manipulation in various organisms. Here, we discovered a new ...CRISPR-Cas modules serve as the adaptive nucleic acid immune systems for prokaryotes, and provide versatile tools for nucleic acid manipulation in various organisms. Here, we discovered a new miniature type V system, CRISPR-Casπ (Cas12l) (~860 aa), from the environmental metagenome. Complexed with a large guide RNA (~170 nt) comprising the tracrRNA and crRNA, Casπ (Cas12l) recognizes a unique 5' C-rich PAM for DNA cleavage under a broad range of biochemical conditions, and generates gene editing in mammalian cells. Cryo-EM study reveals a 'bracelet' architecture of Casπ effector encircling the DNA target at 3.4 Å resolution, substantially different from the canonical 'two-lobe' architectures of Cas12 and Cas9 nucleases. The large guide RNA serves as a 'two-arm' scaffold for effector assembly. Our study expands the knowledge of DNA targeting mechanisms by CRISPR effectors, and offers an efficient but compact platform for DNA manipulation. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_33983.map.gz | 49.7 MB | EMDB map data format | |
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Header (meta data) | emd-33983-v30.xml emd-33983.xml | 19.7 KB 19.7 KB | Display Display | EMDB header |
Images | emd_33983.png | 106.5 KB | ||
Filedesc metadata | emd-33983.cif.gz | 6.9 KB | ||
Others | emd_33983_half_map_1.map.gz emd_33983_half_map_2.map.gz | 49 MB 49 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-33983 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-33983 | HTTPS FTP |
-Validation report
Summary document | emd_33983_validation.pdf.gz | 1 MB | Display | EMDB validaton report |
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Full document | emd_33983_full_validation.pdf.gz | 1 MB | Display | |
Data in XML | emd_33983_validation.xml.gz | 11.9 KB | Display | |
Data in CIF | emd_33983_validation.cif.gz | 13.8 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-33983 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-33983 | HTTPS FTP |
-Related structure data
Related structure data | 7yojMC 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_33983.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Voxel size | X=Y=Z: 0.856 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #2
File | emd_33983_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_33983_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : CasPi
Entire | Name: CasPi |
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Components |
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-Supramolecule #1: CasPi
Supramolecule | Name: CasPi / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Armatimonadetes bacterium (bacteria) |
Molecular weight | Theoretical: 177.28 KDa |
-Macromolecule #1: CasPi
Macromolecule | Name: CasPi / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Armatimonadota bacterium (bacteria) |
Molecular weight | Theoretical: 98.731234 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MAKATKEVKS KRVEALRQVA YQRLERLERK AQKIGAHLRK PGKAADLQSL HYLLHKVEVE YHDIARNLEK DPTWTPKPKM RREKRAIVP ESGPAAPLPT TAKGEPGRPA NRHIPPPVPL DSARIPEDQQ SMGQGSGGRS WCSAPFVEVK LPPTQWSNVR E KLLKFRIE ...String: MAKATKEVKS KRVEALRQVA YQRLERLERK AQKIGAHLRK PGKAADLQSL HYLLHKVEVE YHDIARNLEK DPTWTPKPKM RREKRAIVP ESGPAAPLPT TAKGEPGRPA NRHIPPPVPL DSARIPEDQQ SMGQGSGGRS WCSAPFVEVK LPPTQWSNVR E KLLKFRIE DDADIVRRWA EAKFGSIETA RDGLRASAEI GTSPDVWRSF ISRAISNGKK DFEPLLSLDD DELTADATAE RV VRRWHQI DWVGRMLDSI LETVPSGVSK DTFRSRVESR LKTFHSSVNS FELKKRKDGT VERKRKHTNP QFPYLSPSAV SID PDVVTM EAVELLQMQP EERFAKDPND ANGRMRLRVL QAELGKARRE ALGRRGEKAP PWSGRKVFRG TTTRKREACL VWDK EAQAD GLYFALVMSG GPKIDDKRFV YMDGQPLQSD WQLHNGVAGK AKSCRAMPLI LKHDFLRWYH RHIKNHDVNA PLEKR CVHT TTQFVFVEPD EKKGLQPRLF IRPVFKFYDP VYEVPDSHSI DKKPDCRYLI GIARGVNYPY RAAVYDCETN SIIADK FVD GRKADWERIR NELAYHQRRR DLLRNSRASS AAIQREIRAI ARIRKRERGL NKVETVESIA RLVDWAEENL GKCNYCF VL ADLSSNLNLG RNNRVKHIAA IKEALINQMR KRGYRFKKSG KVDGVREESA WYTSAVAPSG WWAKKEEVDG AWKADKTR P LARKIGSYYC CEEIDGLHLR GVLKGLGRAK RLVLQSDDPS APTRRRGFGS ELFWDPYCTE LCGHAFPQGV VLDADFIGA FNIALRPLVR EELGKKAKAV DLADRHQTLN PTVALRCGVT AYEFVEVGGD PRGGLRKILL NPAEAVI UniProtKB: Reverse transcriptase domain-containing protein |
-Macromolecule #2: DNA (5'-D(P*CP*GP*GP*GP*AP*TP*GP*CP*CP*CP*AP*G)-3')
Macromolecule | Name: DNA (5'-D(P*CP*GP*GP*GP*AP*TP*GP*CP*CP*CP*AP*G)-3') / type: dna / ID: 2 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Armatimonadetes bacterium (bacteria) |
Molecular weight | Theoretical: 3.688405 KDa |
Sequence | String: (DC)(DG)(DG)(DG)(DA)(DT)(DG)(DC)(DC)(DC) (DA)(DG) |
-Macromolecule #3: DNA (30-MER)
Macromolecule | Name: DNA (30-MER) / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Armatimonadetes bacterium (bacteria) |
Molecular weight | Theoretical: 9.192878 KDa |
Sequence | String: (DG)(DT)(DT)(DC)(DA)(DC)(DC)(DA)(DG)(DG) (DG)(DT)(DG)(DT)(DC)(DG)(DC)(DC)(DC)(DT) (DG)(DG)(DG)(DC)(DA)(DT)(DC)(DC)(DC) (DG) |
-Macromolecule #4: RNA (174-MER)
Macromolecule | Name: RNA (174-MER) / type: rna / ID: 4 / Number of copies: 1 |
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Source (natural) | Organism: Armatimonadetes bacterium (bacteria) |
Molecular weight | Theoretical: 56.411703 KDa |
Sequence | String: GUCUGCCGAA GACGCCGCAC GGAGCCUGGG CCGGAAUCGU AGAUCGAACG CGGCAUCGAA GCCCUGCAGC CCUUCGGGGC CAAGGCGGC GCAGCAAGCC UCUUUCAGGC GGCAGAGUCC UUUAGAGUGU GAGAGACACU CUAAAGGAAU GAAAGAGGGC G ACACCCUG GUGAAC |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 8 |
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Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GRAPHENE OXIDE / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 15 sec. / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.01 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 81000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing #1
+Image processing #2
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: BACKBONE TRACE |
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Output model | PDB-7yoj: |