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- PDB-7jzz: Cryo-EM structure of CRISPR-Cas surveillance complex with AcrIF14 -

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Basic information

Entry
Database: PDB / ID: 7jzz
TitleCryo-EM structure of CRISPR-Cas surveillance complex with AcrIF14
Components
  • (Type I-F CRISPR-associated ...) x 2
  • AcrF14
  • CRISPR type I-F/YPEST-associated protein Csy3
  • CRISPR-associated protein Csy1
  • RNA (61-MER)
KeywordsHYDROLASE/RNA / CRISPR / HYDROLASE-RNA complex
Function / homology
Function and homology information


maintenance of CRISPR repeat elements / endonuclease activity / defense response to virus / Hydrolases; Acting on ester bonds / RNA binding
Similarity search - Function
CRISPR-associated protein Csy1 / CRISPR-associated protein (Cas_Csy1) / CRISPR-associated endoribonuclease Cas6/Csy4, subtype I-F/YPEST / CRISPR-associated endoribonuclease Cas6/Csy4, subtype I-F/YPEST superfamily / CRISPR-associated protein (Cas_Csy4) / CRISPR-associated protein Csy2 / CRISPR-associated protein (Cas_Csy2) / CRISPR-associated protein Csy3 / CRISPR-associated protein (Cas_Csy3)
Similarity search - Domain/homology
: / RNA / RNA (> 10) / Tail protein / CRISPR type I-F/YPEST-associated protein Csy3 / : / Uncharacterized protein / CRISPR-associated protein Csy1 / CRISPR-associated protein Csy2 / CRISPR-associated protein Csy3 / CRISPR-associated endonuclease Cas6/Csy4
Similarity search - Component
Biological speciesPseudomonas aeruginosa (bacteria)
unidentified (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsChang, L. / Li, Z. / Gabel, C.
CitationJournal: Nucleic Acids Res / Year: 2021
Title: Structural basis for inhibition of the type I-F CRISPR-Cas surveillance complex by AcrIF4, AcrIF7 and AcrIF14.
Authors: Clinton Gabel / Zhuang Li / Heng Zhang / Leifu Chang /
Abstract: CRISPR-Cas systems are adaptive immune systems in bacteria and archaea to defend against mobile genetic elements (MGEs) and have been repurposed as genome editing tools. Anti-CRISPR (Acr) proteins ...CRISPR-Cas systems are adaptive immune systems in bacteria and archaea to defend against mobile genetic elements (MGEs) and have been repurposed as genome editing tools. Anti-CRISPR (Acr) proteins are produced by MGEs to counteract CRISPR-Cas systems and can be used to regulate genome editing by CRISPR techniques. Here, we report the cryo-EM structures of three type I-F Acr proteins, AcrIF4, AcrIF7 and AcrIF14, bound to the type I-F CRISPR-Cas surveillance complex (the Csy complex) from Pseudomonas aeruginosa. AcrIF4 binds to an unprecedented site on the C-terminal helical bundle of Cas8f subunit, precluding conformational changes required for activation of the Csy complex. AcrIF7 mimics the PAM duplex of target DNA and is bound to the N-terminal DNA vise of Cas8f. Two copies of AcrIF14 bind to the thumb domains of Cas7.4f and Cas7.6f, preventing hybridization between target DNA and the crRNA. Our results reveal structural detail of three AcrIF proteins, each binding to a different site on the Csy complex for inhibiting degradation of MGEs.
History
DepositionSep 2, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 30, 2020Provider: repository / Type: Initial release
Revision 1.0Dec 30, 2020Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Dec 30, 2020Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Dec 30, 2020Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.0Dec 30, 2020Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Dec 30, 2020Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.0Dec 30, 2020Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Dec 30, 2020Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.0Dec 30, 2020Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Dec 30, 2020Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.1Jun 2, 2021Group: Structure summary / Category: struct / Item: _struct.title
Revision 1.2Jan 12, 2022Group: Database references / Category: citation / citation_author / database_2
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _database_2.pdbx_DOI / _database_2.pdbx_database_accession
Revision 1.3May 29, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond
Revision 1.4May 21, 2025Group: Data collection / Structure summary / Category: em_admin / em_software / pdbx_entry_details / Item: _em_admin.last_update / _em_software.name
Revision 1.1May 21, 2025Data content type: EM metadata / Data content type: EM metadata / EM metadata / Group: Data processing / Experimental summary / Data content type: EM metadata / EM metadata / Category: em_admin / em_software / Data content type: EM metadata / EM metadata / Item: _em_admin.last_update / _em_software.name

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Structure visualization

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  • Deposited structure unit
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  • Superimposition on EM map
  • EMDB-22585
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Structure viewerMolecule:
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Assembly

Deposited unit
A: CRISPR-associated protein Csy1
B: Type I-F CRISPR-associated protein Csy2
C: Type I-F CRISPR-associated endoribonuclease Cas6/Csy4
D: CRISPR type I-F/YPEST-associated protein Csy3
E: CRISPR type I-F/YPEST-associated protein Csy3
F: CRISPR type I-F/YPEST-associated protein Csy3
G: CRISPR type I-F/YPEST-associated protein Csy3
H: CRISPR type I-F/YPEST-associated protein Csy3
I: CRISPR type I-F/YPEST-associated protein Csy3
J: AcrF14
K: AcrF14
M: RNA (61-MER)


Theoretical massNumber of molelcules
Total (without water)380,72312
Polymers380,72312
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area60240 Å2
ΔGint-187 kcal/mol
Surface area113900 Å2

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Components

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Protein , 3 types, 9 molecules ADEFGHIJK

#1: Protein CRISPR-associated protein Csy1 / Csy1 (Cas8f)


Mass: 49194.168 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa (bacteria) / Gene: csy1, PA14_33330 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q02ML9
#4: Protein
CRISPR type I-F/YPEST-associated protein Csy3 / CRISPR-associated protein Csy3


Mass: 37579.273 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa (bacteria)
Gene: EQH76_13805, F7O93_18150, NCTC13437_01527, NCTC13619_03982
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0A444M080, UniProt: Q02MM1*PLUS
#5: Protein AcrF14


Mass: 14297.373 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) unidentified (others) / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0A0R6PCL0*PLUS

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Type I-F CRISPR-associated ... , 2 types, 2 molecules BC

#2: Protein Type I-F CRISPR-associated protein Csy2 / Csy2 (Cas5f)


Mass: 36244.074 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa (bacteria)
Gene: csy2, ALP65_00953, EQH76_13810, FCG96_17775, PACL_0128
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: B3G161, UniProt: Q02MM0*PLUS
#3: Protein Type I-F CRISPR-associated endoribonuclease Cas6/Csy4


Mass: 21675.781 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa (bacteria) / Gene: cas6f, F7O93_18155 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0A643HZS6, UniProt: Q02MM2*PLUS

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RNA chain , 1 types, 1 molecules M

#6: RNA chain RNA (61-MER)


Mass: 19538.586 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa (bacteria) / Production host: Escherichia coli BL21(DE3) (bacteria) / References: GenBank: 313291946

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Details

Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: CRISPR-Cas complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD
Image recordingElectron dose: 54 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.18.2_3874: / Classification: refinement
EM softwareName: PHENIX / Category: model refinement
CTF correctionType: PHASE FLIPPING ONLY
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 226089 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0125369
ELECTRON MICROSCOPYf_angle_d0.9834733
ELECTRON MICROSCOPYf_dihedral_angle_d21.6279331
ELECTRON MICROSCOPYf_chiral_restr0.0533929
ELECTRON MICROSCOPYf_plane_restr0.0074367

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