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Yorodumi- EMDB-37656: Structure of CbCas9-PcrIIC1 complex bound to 28-bp DNA substrate ... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-37656 | |||||||||
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Title | Structure of CbCas9-PcrIIC1 complex bound to 28-bp DNA substrate (20-nt complementary) | |||||||||
Map data | ||||||||||
Sample |
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Keywords | Cas9 complex / DNA BINDING PROTEIN / DNA BINDING PROTEIN-DNA-RNA complex | |||||||||
Biological species | Chryseobacterium (bacteria) | |||||||||
Method | single particle reconstruction / Resolution: 2.98 Å | |||||||||
Authors | Zhang S / Lin S / Liu JJG | |||||||||
Funding support | China, 2 items
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Citation | Journal: Nature / Year: 2024 Title: Pro-CRISPR PcrIIC1-associated Cas9 system for enhanced bacterial immunity. Authors: Shouyue Zhang / Ao Sun / Jing-Mei Qian / Shuo Lin / Wenjing Xing / Yun Yang / Han-Zhou Zhu / Xin-Yi Zhou / Yan-Shuo Guo / Yun Liu / Yu Meng / Shu-Lin Jin / Wenhao Song / Cheng-Ping Li / ...Authors: Shouyue Zhang / Ao Sun / Jing-Mei Qian / Shuo Lin / Wenjing Xing / Yun Yang / Han-Zhou Zhu / Xin-Yi Zhou / Yan-Shuo Guo / Yun Liu / Yu Meng / Shu-Lin Jin / Wenhao Song / Cheng-Ping Li / Zhaofu Li / Shuai Jin / Jian-Hua Wang / Meng-Qiu Dong / Caixia Gao / Chunlai Chen / Yang Bai / Jun-Jie Gogo Liu / Abstract: The CRISPR system is an adaptive immune system found in prokaryotes that defends host cells against the invasion of foreign DNA. As part of the ongoing struggle between phages and the bacterial ...The CRISPR system is an adaptive immune system found in prokaryotes that defends host cells against the invasion of foreign DNA. As part of the ongoing struggle between phages and the bacterial immune system, the CRISPR system has evolved into various types, each with distinct functionalities. Type II Cas9 is the most extensively studied of these systems and has diverse subtypes. It remains uncertain whether members of this family can evolve additional mechanisms to counter viral invasions. Here we identify 2,062 complete Cas9 loci, predict the structures of their associated proteins and reveal three structural growth trajectories for type II-C Cas9. We found that novel associated genes (NAGs) tended to be present within the loci of larger II-C Cas9s. Further investigation revealed that CbCas9 from Chryseobacterium species contains a novel β-REC2 domain, and forms a heterotetrameric complex with an NAG-encoded CRISPR-Cas-system-promoting (pro-CRISPR) protein of II-C Cas9 (PcrIIC1). The CbCas9-PcrIIC1 complex exhibits enhanced DNA binding and cleavage activity, broader compatibility for protospacer adjacent motif sequences, increased tolerance for mismatches and improved anti-phage immunity, compared with stand-alone CbCas9. Overall, our work sheds light on the diversity and 'growth evolutionary' trajectories of II-C Cas9 proteins at the structural level, and identifies many NAGs-such as PcrIIC1, which serves as a pro-CRISPR factor to enhance CRISPR-mediated immunity. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_37656.map.gz | 145.1 MB | EMDB map data format | |
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Header (meta data) | emd-37656-v30.xml emd-37656.xml | 20 KB 20 KB | Display Display | EMDB header |
Images | emd_37656.png | 38.9 KB | ||
Filedesc metadata | emd-37656.cif.gz | 6.9 KB | ||
Others | emd_37656_half_map_1.map.gz emd_37656_half_map_2.map.gz | 151.4 MB 151.4 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-37656 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-37656 | HTTPS FTP |
-Validation report
Summary document | emd_37656_validation.pdf.gz | 654 KB | Display | EMDB validaton report |
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Full document | emd_37656_full_validation.pdf.gz | 653.5 KB | Display | |
Data in XML | emd_37656_validation.xml.gz | 14.5 KB | Display | |
Data in CIF | emd_37656_validation.cif.gz | 17.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-37656 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-37656 | HTTPS FTP |
-Related structure data
Related structure data | 8wmmMC 8iyqC 8wmhC 8wmnC 8wr4C M: atomic model generated by this map C: citing same article (ref.) |
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-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_37656.map.gz / Format: CCP4 / Size: 163.6 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: 1.0825 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #2
File | emd_37656_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_37656_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Structure of CbCas9-PcrIIC1 complex bound to 28-bp DNA substrate ...
Entire | Name: Structure of CbCas9-PcrIIC1 complex bound to 28-bp DNA substrate (20-nt complementary) |
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Components |
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-Supramolecule #1: Structure of CbCas9-PcrIIC1 complex bound to 28-bp DNA substrate ...
Supramolecule | Name: Structure of CbCas9-PcrIIC1 complex bound to 28-bp DNA substrate (20-nt complementary) type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#5 |
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Source (natural) | Organism: Chryseobacterium (bacteria) |
Molecular weight | Theoretical: 469.6 KDa |
-Macromolecule #1: deadCbCas9
Macromolecule | Name: deadCbCas9 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Chryseobacterium (bacteria) |
Molecular weight | Theoretical: 170.029109 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MIKNILGLAL GTNSIGWALV KQDFENKQGE ILGMGSRIIP MSQDILGDFG KGNSVSQTAE RTKYRSVRRL RERFLLRRER LHRVLYILN FLPEHYASQI DFEKRLGKFK VETEPKLVWK NTDGQFSFLF QNSFNEMLED FKAAGQELKI PYDWTIYHLR K KAISQKIE ...String: MIKNILGLAL GTNSIGWALV KQDFENKQGE ILGMGSRIIP MSQDILGDFG KGNSVSQTAE RTKYRSVRRL RERFLLRRER LHRVLYILN FLPEHYASQI DFEKRLGKFK VETEPKLVWK NTDGQFSFLF QNSFNEMLED FKAAGQELKI PYDWTIYHLR K KAISQKIE KEELAWILLN FNHKRGYYQL RGEDFEEEKD KTFVRLKVDR IVDSGENVKG KILYDVYFEN GWKYDKQVVK TE DWVDRTK EFIVSESILK NGETKRTFKA VDSEKDWIAI KTKTEQEIEH SHKTVGTYIY ETLLQNPKQK IKGKLVRTIE RKF YKEELR QILEKQKEFH QELQSDDLYN DCIRELYRNN EVHQLTLRKK DFVHLFMEDI IFYQRPLRSQ KSSVSNCTLE FRKY KGENG AEHTQYLKAI PKSNPYYQEF RLWQWIFNLN LYTKDNDENV TKVFLNTTQD FENLFEFLNT RKEVDQKALL KHFKL NEKT HRWNFVEDKK YPCNETKTMI SSRLDKVENI SDDFLTRDIE QKIWHIIYSV NDKVEYEKAL KSFARKHHLD ESSFFE AFR KFPPFKSEYG SFSEKAIKKL LPLMRLGKYW NYAEIDKYSR ERIQKIITGE YDENIKDKVR EKSVHLTIEN DFQGLQL WL AQYIVYGRHS EASMIGKWNS ANDLEVFLKD FKQHSLRNPI VEQVITETLR VVKDIWLKYG NGTKDFFNEI HIELGREM K LPADDRKKLT NQITENENTN LRIKALLAEM MNDHSVENVR PFSPMQQEIL KIYEDGVLKS DIEIEDDILK ISKTAQPSS SDLKRYKLWL EQKYKSPYTG QIIPLNKLFT PEYEIEAIIP QSRYFDDSFS NKIICESAVN KLKDNYIGLG FIKQFGGTII ELGFGKSVK VFDTEEYEDF VKKHYANNRG KRNKLLMEDI PEKMIERQLN DTRYISKYIS GILSNIVRVE DGSDEGVNSK N IVPGNGKI TTQLKQDWGL NDVWNDLILP RFERMNQLTN SKDFTAWNEN HQKFLPTVPI EFSKGFSKKR IDHRHHALDA LV IACATTD HVNLLNNQSA KSDTKRYDLK KKLMKFEKVV YHHTQTGEKI EREIPKQFLK PWEKFTVDAK HNLESIIVSF KQN LRVINK ATNYYEKYVE KDGTKNKERV EQAGTNWAIR KPMHKDTVSG KVDLPWVKVP KGKILTATRK SLDSSFDLKS IGSI TDTGI QKILKNYLAF KDGNPELAFS PEGIDDLNKN IEKYNDGKPH QPINKVRVFE LGSKFQVGQT GNKKGKYVEA AKGTN LFFA VYEDEKGKRS YETIPLNEVI ERQKQGLTSV PLENEKGSRL LFDLSPNDLV YVPEIDENID SNFVFSNLNK EKISRI YKV EKTSGTECYF VRQDIAYLIK QYDAKTKIGE LESQNKLQVT MTDDRIRITD TCVKINCDRL GNINFITKEK IKQIFNE FR |
-Macromolecule #4: PcrIIC1
Macromolecule | Name: PcrIIC1 / type: protein_or_peptide / ID: 4 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Chryseobacterium (bacteria) |
Molecular weight | Theoretical: 15.991991 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MSLDKIAIDT NILLYAYDNR DLDKQDRAVE ILLKKPFVTQ LVVFEFIKVL ERRFKMDKKE ITKLTIKLLK EVIIPLSLHR DIYNYSQFL LQRYNFGLSD ILVLSDSILN NCTILLSEDM CNGMIVDKKL KIVNPFL |
-Macromolecule #2: TS
Macromolecule | Name: TS / type: dna / ID: 2 / Number of copies: 2 / Classification: DNA |
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Source (natural) | Organism: Chryseobacterium (bacteria) |
Molecular weight | Theoretical: 8.450413 KDa |
Sequence | String: (DC)(DG)(DT)(DT)(DT)(DT)(DG)(DT)(DC)(DT) (DC)(DG)(DG)(DC)(DT)(DC)(DC)(DC)(DC)(DG) (DA)(DC)(DA)(DT)(DT)(DC)(DT)(DC) |
-Macromolecule #5: NTS
Macromolecule | Name: NTS / type: dna / ID: 5 / Number of copies: 2 / Classification: DNA |
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Source (natural) | Organism: Chryseobacterium (bacteria) |
Molecular weight | Theoretical: 8.76267 KDa |
Sequence | String: (DG)(DA)(DG)(DA)(DA)(DT)(DG)(DT)(DC)(DG) (DG)(DG)(DG)(DA)(DG)(DC)(DC)(DG)(DA)(DG) (DA)(DC)(DA)(DA)(DA)(DA)(DC)(DG) |
-Macromolecule #3: sgRNA
Macromolecule | Name: sgRNA / type: rna / ID: 3 / Number of copies: 2 |
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Source (natural) | Organism: Chryseobacterium (bacteria) |
Molecular weight | Theoretical: 40.936227 KDa |
Sequence | String: GAGAAUGUCG GGGAGCCGAG GUUGUGAAUU GCUUUCAAAA AUUAUUGAGA AAUAAUUUUG AAAAGCAAUU CACAAUAAGG AUUAUUCCG UUGUGAAAAC AUUCAAGGCG GGGCAACUCG CCUUUUUU |
-Macromolecule #6: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 6 / Number of copies: 2 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Experimental details
-Structure determination
Processing | single particle reconstruction |
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Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 / Component - Concentration: 1.0 uM / Component - Formula: NaCl / Component - Name: sodium chloride Details: 150mM NaCl, 20mM Hepes (pH=7.5), 5mM MgCl2, 1mM Tcep, 0.1% Glycerol |
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-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: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 1.3 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
Startup model | Type of model: NONE / Details: Ab-initial in cryosparc |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 2.98 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 355919 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |