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- EMDB-7052: Cryo-EM structure of Type I-F CRISPR crRNA-guided Csy surveillanc... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-7052 | |||||||||
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Title | Cryo-EM structure of Type I-F CRISPR crRNA-guided Csy surveillance complex with bound anti-CRISPR protein AcrF10 | |||||||||
![]() | Final sharpened map of type I-F CRISPR crRNA-guided Csy surveillance complex with bound anti-CRISPR protein AcrF10 | |||||||||
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Function / homology | ![]() maintenance of CRISPR repeat elements / endonuclease activity / defense response to virus / Hydrolases; Acting on ester bonds / RNA binding Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.6 Å | |||||||||
![]() | Guo TW / Bartesaghi A / Yang H / Falconieri V / Rao P / Merk A / Fox T / Earl L / Patel DJ / Subramaniam S | |||||||||
![]() | ![]() Title: Cryo-EM Structures Reveal Mechanism and Inhibition of DNA Targeting by a CRISPR-Cas Surveillance Complex. Authors: Tai Wei Guo / Alberto Bartesaghi / Hui Yang / Veronica Falconieri / Prashant Rao / Alan Merk / Edward T Eng / Ashleigh M Raczkowski / Tara Fox / Lesley A Earl / Dinshaw J Patel / Sriram Subramaniam / ![]() Abstract: Prokaryotic cells possess CRISPR-mediated adaptive immune systems that protect them from foreign genetic elements, such as invading viruses. A central element of this immune system is an RNA-guided ...Prokaryotic cells possess CRISPR-mediated adaptive immune systems that protect them from foreign genetic elements, such as invading viruses. A central element of this immune system is an RNA-guided surveillance complex capable of targeting non-self DNA or RNA for degradation in a sequence- and site-specific manner analogous to RNA interference. Although the complexes display considerable diversity in their composition and architecture, many basic mechanisms underlying target recognition and cleavage are highly conserved. Using cryoelectron microscopy (cryo-EM), we show that the binding of target double-stranded DNA (dsDNA) to a type I-F CRISPR system yersinia (Csy) surveillance complex leads to large quaternary and tertiary structural changes in the complex that are likely necessary in the pathway leading to target dsDNA degradation by a trans-acting helicase-nuclease. Comparison of the structure of the surveillance complex before and after dsDNA binding, or in complex with three virally encoded anti-CRISPR suppressors that inhibit dsDNA binding, reveals mechanistic details underlying target recognition and inhibition. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 47.8 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 21.4 KB 21.4 KB | Display Display | ![]() |
Images | ![]() | 175.7 KB | ||
Others | ![]() | 47.7 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6b48MC ![]() 7048C ![]() 7049C ![]() 7050C ![]() 7051C ![]() 6anvC ![]() 6anwC ![]() 6b44C ![]() 6b45C ![]() 6b46C ![]() 6b47C C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Map
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Annotation | Final sharpened map of type I-F CRISPR crRNA-guided Csy surveillance complex with bound anti-CRISPR protein AcrF10 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.84 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Additional map: Unsharpened map of type I-F CRISPR crRNA-guided Csy...
File | emd_7052_additional.map | ||||||||||||
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Annotation | Unsharpened map of type I-F CRISPR crRNA-guided Csy surveillance complex with bound anti-CRISPR protein AcrF10 | ||||||||||||
Projections & Slices |
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Density Histograms |
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Sample components
-Entire : Type I-F CRISPR crRNA-guided Csy surveillance complex with bound ...
Entire | Name: Type I-F CRISPR crRNA-guided Csy surveillance complex with bound anti-CRISPR protein AcrF10 |
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Components |
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-Supramolecule #1: Type I-F CRISPR crRNA-guided Csy surveillance complex with bound ...
Supramolecule | Name: Type I-F CRISPR crRNA-guided Csy surveillance complex with bound anti-CRISPR protein AcrF10 type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: ![]() ![]() |
Recombinant expression | Organism: ![]() ![]() |
Molecular weight | Theoretical: 350 KDa |
-Macromolecule #1: CRISPR-associated protein Csy1
Macromolecule | Name: CRISPR-associated protein Csy1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() Strain: UCBPP-PA14 |
Molecular weight | Theoretical: 49.338297 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: GSMTSPLPTP TWQELRQFIE SFIQERLQGK LDKLQPDEDD KRQTLLATHR REAWLADAAR RVGQLQLVTH TLKPIHPDAR GSNLHSLPQ APGQPGLAGS HELGDRLVSD VVGNAAALDV FKFLSLQYQG KNLLNWLTED SAEALQALSD NAEQAREWRQ A FIGITTVK ...String: GSMTSPLPTP TWQELRQFIE SFIQERLQGK LDKLQPDEDD KRQTLLATHR REAWLADAAR RVGQLQLVTH TLKPIHPDAR GSNLHSLPQ APGQPGLAGS HELGDRLVSD VVGNAAALDV FKFLSLQYQG KNLLNWLTED SAEALQALSD NAEQAREWRQ A FIGITTVK GAPASHSLAK QLYFPLPGSG YHLLAPLFPT SLVHHVHALL REARFGDAAK AAREARSRQE SWPHGFSEYP NL AIQKFGG TKPQNISQLN NERRGENWLL PSLPPNWQRQ NVNAPMRHSS VFEHDFGRTP EVSRLTRTLQ RFLAKTVHNN LAI RQRRAQ LVAQICDEAL QYAARLRELE PGWSATPGCQ LHDAEQLWLD PLRAQTDETF LQRRLRGDWP AEVGNRFANW LNRA VSSDS QILGSPEAAQ WSQELSKELT MFKEILEDER D |
-Macromolecule #2: CRISPR-associated protein Csy2
Macromolecule | Name: CRISPR-associated protein Csy2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() Strain: UCBPP-PA14 |
Molecular weight | Theoretical: 36.446352 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MAMSVTDPEA LLLLPRLSIQ NANAISSPLT WGFPSPGAFT GFVHALQRRV GISLDIELDG VGIVCHRFEA QISQPAGKRT KVFNLTRNP LNRDGSTAAI VEEGRAHLEV SLLLGVHGDG LDDHPAQEIA RQVQEQAGAM RLAGGSILPW CNERFPAPNA E LLMLGGSD ...String: MAMSVTDPEA LLLLPRLSIQ NANAISSPLT WGFPSPGAFT GFVHALQRRV GISLDIELDG VGIVCHRFEA QISQPAGKRT KVFNLTRNP LNRDGSTAAI VEEGRAHLEV SLLLGVHGDG LDDHPAQEIA RQVQEQAGAM RLAGGSILPW CNERFPAPNA E LLMLGGSD EQRRKNQRRL TRRLLPGFAL VSREALLQQH LETLRTTLPE ATTLDALLDL CRINFEPPAT SSEEEASPPD AA WQVRDKP GWLVPIPAGY NALSPLYLPG EVRNARDRET PLRFVENLFG LGEWLSPHRV AALSDLLWYH HAEPDKGLYR WST PRFVEH AIA |
-Macromolecule #3: CRISPR-associated protein Csy3
Macromolecule | Name: CRISPR-associated protein Csy3 / type: protein_or_peptide / ID: 3 / Number of copies: 6 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() Strain: UCBPP-PA14 |
Molecular weight | Theoretical: 37.781547 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MAMSKPILST ASVLAFERKL DPSDALMSAG AWAQRDASQE WPAVTVREKS VRGTISNRLK TKDRDPAKLD ASIQSPNLQT VDVANLPSD ADTLKVRFTL RVLGGAGTPS ACNDAAYRDK LLQTVATYVN DQGFAELARR YAHNLANARF LWRNRVGAEA V EVRINHIR ...String: MAMSKPILST ASVLAFERKL DPSDALMSAG AWAQRDASQE WPAVTVREKS VRGTISNRLK TKDRDPAKLD ASIQSPNLQT VDVANLPSD ADTLKVRFTL RVLGGAGTPS ACNDAAYRDK LLQTVATYVN DQGFAELARR YAHNLANARF LWRNRVGAEA V EVRINHIR QGEVARAWRF DALAIGLRDF KADAELDALA ELIASGLSGS GHVLLEVVAF ARIGDGQEVF PSQELILDKG DK KGQKSKT LYSVRDAAAI HSQKIGNALR TIDTWYPDED GLGPIAVEPY GSVTSQGKAY RQPKQKLDFY TLLDNWVLRD EAP AVEQQH YVIANLIRGG VFGEAEEK |
-Macromolecule #4: Anti-CRISPR protein AcrF10
Macromolecule | Name: Anti-CRISPR protein AcrF10 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 11.614141 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: GS(MSE)TTFRIEN VRIETINDFD (MSE)VKFDLVTDL GRVELAEHVN YDSEGDFKSV EYTDSNIRYN (MSE)VDELCSV F DLTDKPSL(MSE)P AIDYVTFAEI IEAVEE(MSE)LEA |
-Macromolecule #5: CRISPR-associated endonuclease Cas6/Csy4
Macromolecule | Name: CRISPR-associated endonuclease Cas6/Csy4 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO / EC number: Hydrolases; Acting on ester bonds |
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Source (natural) | Organism: ![]() Strain: UCBPP-PA14 |
Molecular weight | Theoretical: 21.629777 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MAMDHYLDIR LRPDPEFPPA QLMSVLFGKL HQALVAQGGD RIGVSFPDLD ESRSRLGERL RIHASADDLR ALLARPWLEG LRDHLQFGE PAVVPHPTPY RQVSRVQAKS NPERLRRRLM RRHDLSEEEA RKRIPDTVAR ALDLPFVTLR SQSTGQHFRL F IRHGPLQV TAEEGGFTCY GLSKGGFVPW F |
-Macromolecule #6: Pseudomonas aeruginosa strain SMC4485 CRISPR repeat sequence
Macromolecule | Name: Pseudomonas aeruginosa strain SMC4485 CRISPR repeat sequence type: rna / ID: 6 / Number of copies: 1 |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 19.265404 KDa |
Sequence | String: CUAAGAAAUU CACGGCGGGC UUGAUGUCCG CGUCUACCUG GUUCACUGCC GUGUAGGCAG |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 1 mg/mL |
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Buffer | pH: 7.2 Details: 10 mM HEPES, pH 7.2, 150 mM NaCl, 2nM MgCl2, 1 mM DTT |
Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Pretreatment - Type: PLASMA CLEANING |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 98 % / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number real images: 900 / Average exposure time: 15.2 sec. / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
CTF correction | Software - Name: CTFFIND (ver. 4) |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: FREALIGN (ver. 9.11) / Number images used: 42498 |
Initial angle assignment | Type: PROJECTION MATCHING / Software - Name: FREALIGN (ver. 9.11) |
Final angle assignment | Type: PROJECTION MATCHING / Software - Name: FREALIGN (ver. 9.11) |