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- EMDB-45796: Post-targeting aCascade Type I-A CRISPR-Cas Surveillance Complexes -

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Basic information

Entry
Database: EMDB / ID: EMD-45796
TitlePost-targeting aCascade Type I-A CRISPR-Cas Surveillance Complexes
Map dataPost-targeting aCascade Type I-A CRISPR-Cas Surveillance Complexes
Sample
  • Complex: Type I-A CRISPR-Cas post-targeting surveillance subcomplex from Saccharolobus solfataricus
    • Protein or peptide: CRISPR-associated aCascade subunit Cas7/Csa2 2
    • Protein or peptide: CRISPR system aCascade subunit Cas5 1
    • RNA: RNA (38-MER)
    • DNA: DNA (5'-D(P*CP*GP*GP*GP*TP*TP*TP*TP*CP*CP*T)-3')
  • Ligand: 2-AMINO-2-HYDROXYMETHYL-PROPANE-1,3-DIOL
KeywordsCascade CRISPR CRISPR-Cas Cas Cas5 Cas7 crRNA TypeI-A / DNA-RNA HYBRID / RNA BINDING PROTEIN-RNA-DNA complex
Function / homology
Function and homology information


defense response to virus / RNA binding
Similarity search - Function
CRISPR-associated protein, Cas5a type / : / CRISPR-associated protein Cas7/Cst2/DevR, subtype I-a/Apern / CRISPR-associated protein Cas7/Cst2/DevR / : / CRISPR-associated negative auto-regulator DevR/Csa2 / CRISPR-associated protein Cas5, N-terminal
Similarity search - Domain/homology
CRISPR-associated aCascade subunit Cas7/Csa2 2 / CRISPR system aCascade subunit Cas5 1
Similarity search - Component
Biological speciesSaccharolobus solfataricus (archaea) / Saccharolobus solfataricus P2 (archaea)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.97 Å
AuthorsGentry JK / Findlay JL / Lawrence CM
Funding support United States, 2 items
OrganizationGrant numberCountry
National Science Foundation (NSF, United States)DBI-1828765 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P30GM140963 United States
CitationJournal: To Be Published
Title: Structures and Disassembly of Post-Targeting Type I-A CRISPR-Cas Surveillance Complexes
Authors: Gentry JK / Findlay JL / Lawrence CM
History
DepositionJul 17, 2024-
Header (metadata) releaseJul 23, 2025-
Map releaseJul 23, 2025-
UpdateJul 23, 2025-
Current statusJul 23, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45796.png

Downloads & links

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Map

FileDownload / File: emd_45796.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationPost-targeting aCascade Type I-A CRISPR-Cas Surveillance Complexes
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.42 Å/pix.
x 256 pix.
= 363.52 Å		1.42 Å/pix.
x 256 pix.
= 363.52 Å		1.42 Å/pix.
x 256 pix.
= 363.52 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.42 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.114
Minimum - Maximum-0.55791265 - 1.3773557
Average (Standard dev.)0.00007065394 (±0.026831154)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 363.52 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half Map B

Fileemd_45796_half_map_1.map
AnnotationHalf Map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map A

Fileemd_45796_half_map_2.map
AnnotationHalf Map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Type I-A CRISPR-Cas post-targeting surveillance subcomplex from S...

EntireName: Type I-A CRISPR-Cas post-targeting surveillance subcomplex from Saccharolobus solfataricus
Components
  • Complex: Type I-A CRISPR-Cas post-targeting surveillance subcomplex from Saccharolobus solfataricus
    • Protein or peptide: CRISPR-associated aCascade subunit Cas7/Csa2 2
    • Protein or peptide: CRISPR system aCascade subunit Cas5 1
    • RNA: RNA (38-MER)
    • DNA: DNA (5'-D(P*CP*GP*GP*GP*TP*TP*TP*TP*CP*CP*T)-3')
  • Ligand: 2-AMINO-2-HYDROXYMETHYL-PROPANE-1,3-DIOL

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Supramolecule #1: Type I-A CRISPR-Cas post-targeting surveillance subcomplex from S...

SupramoleculeName: Type I-A CRISPR-Cas post-targeting surveillance subcomplex from Saccharolobus solfataricus
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4 / Details: Composed of Cas5, 6Cas7, crRNA and DNA
Source (natural)Organism: Saccharolobus solfataricus (archaea) / Strain: P2
Molecular weightTheoretical: 250 KDa

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Macromolecule #1: CRISPR-associated aCascade subunit Cas7/Csa2 2

MacromoleculeName: CRISPR-associated aCascade subunit Cas7/Csa2 2 / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Saccharolobus solfataricus P2 (archaea)
Molecular weightTheoretical: 35.308508 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MISGSVRFLV NLESLNGVES IGNLTKHRTA PVVLKTSTGY LVRYVPVISG EALAHAYQAS LVDIAKKEGL PVGSLSSQYE FIKFSTDEA LKIEGIKEPK DYNDARRFEV EVMLKDVIAD VGGFMYAGGA PVRRTSRIKL GYMIPALRGD EIPAQLEAQF H VRFSNKPV ...String:
MISGSVRFLV NLESLNGVES IGNLTKHRTA PVVLKTSTGY LVRYVPVISG EALAHAYQAS LVDIAKKEGL PVGSLSSQYE FIKFSTDEA LKIEGIKEPK DYNDARRFEV EVMLKDVIAD VGGFMYAGGA PVRRTSRIKL GYMIPALRGD EIPAQLEAQF H VRFSNKPV SGSQAIFNVE VSSALYTFSF ELDEDLIAVP STFGEKVKGE EELERQKAKR VKSAIKALYS LLSGNFGGKR SR FLPSMKL MSLVVTKTDF PFMPEPAHDD DYIKTTIMRL GKAKGVLNGN LAKAYVINNE GIEVGEGVTV LSTVEDLVVK LEE E

UniProtKB: CRISPR-associated aCascade subunit Cas7/Csa2 2

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Macromolecule #2: CRISPR system aCascade subunit Cas5 1

MacromoleculeName: CRISPR system aCascade subunit Cas5 1 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharolobus solfataricus P2 (archaea)
Molecular weightTheoretical: 27.431582 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MIYSKVFLKL HWGFSVVKPL AAKAKPGFYL PPPTTLIGAL SYGKFRGVDN INLGNVYGSP AYNFRNIMAT ARLESEGVYT EDIIRNVIS YFQRKERREN PRYIYGVIPT GKVYIPNGRL VVVYVTDSIS KEELEKLCWS ITRIGCKECL ASVENVEVGE A KKVSGRVK ...String:
MIYSKVFLKL HWGFSVVKPL AAKAKPGFYL PPPTTLIGAL SYGKFRGVDN INLGNVYGSP AYNFRNIMAT ARLESEGVYT EDIIRNVIS YFQRKERREN PRYIYGVIPT GKVYIPNGRL VVVYVTDSIS KEELEKLCWS ITRIGCKECL ASVENVEVGE A KKVSGRVK TRYYFRDTVK VVGRKEFLEY VTFWEENGYI WGKEGSPVRY ILPITTYPLA SKEVEVEAKE AYEVGGEYVV FS

UniProtKB: CRISPR system aCascade subunit Cas5 1

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Macromolecule #3: RNA (38-MER)

MacromoleculeName: RNA (38-MER) / type: rna / ID: 3 / Number of copies: 1
Source (natural)Organism: Saccharolobus solfataricus (archaea) / Strain: P1-16
Molecular weightTheoretical: 20.161945 KDa
SequenceString:
AUUGAAAGUU CUGUUUCGAA GAAAACCCGC CUCAGAUUCA UUAUGGGGAU AAUCUCUUAU AGA

GENBANK: GENBANK: CP050869.1

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Macromolecule #4: DNA (5'-D(P*CP*GP*GP*GP*TP*TP*TP*TP*CP*CP*T)-3')

MacromoleculeName: DNA (5'-D(P*CP*GP*GP*GP*TP*TP*TP*TP*CP*CP*T)-3') / type: dna / ID: 4
Details: Target DNA from the plasmid carrying the CRISPR array that copurified with the complex.
Number of copies: 1 / Classification: DNA
Source (natural)Organism: Saccharolobus solfataricus (archaea) / Strain: P1-16
Molecular weightTheoretical: 3.331169 KDa
SequenceString:
(DC)(DG)(DG)(DG)(DT)(DT)(DT)(DT)(DC)(DC) (DT)

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Macromolecule #5: 2-AMINO-2-HYDROXYMETHYL-PROPANE-1,3-DIOL

MacromoleculeName: 2-AMINO-2-HYDROXYMETHYL-PROPANE-1,3-DIOL / type: ligand / ID: 5 / Number of copies: 6 / Formula: TRS
Molecular weightTheoretical: 122.143 Da
Chemical component information

ChemComp-TRS:
2-AMINO-2-HYDROXYMETHYL-PROPANE-1,3-DIOL / pH buffer*YM

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1.1 mg/mL
BufferpH: 8
Component:
ConcentrationNameFormula
10.0 mMTris
50.0 mMsodium chlorideNaCl
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 11 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: OTHER / Pretreatment - Pressure: 39.0 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 85 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: Blot time: 4 seconds Blot force: 5.
DetailsThe sample was a mixture of different subcomplex assemblies of aCASCDE purified by size-exclusion chromatography.

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Electron microscopy

MicroscopeTFS TALOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number real images: 3901 / Average exposure time: 3.7 sec. / Average electron dose: 55.3 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 36000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN

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Image processing

Particle selectionNumber selected: 5400000
Details: Following blob picking and washing with 2D classification
CTF correctionSoftware - Name: cryoSPARC / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.97 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 389106
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationSoftware - Name: cryoSPARC / Details: See manuscript
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
DetailsAlphafold 3 homology model including RNA was rigid body fit into the density.DNA modeling was ab initio. After rigid body fitting iterative real-space refinement and model building with PHENIX and Coot was performed.
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 510 / Target criteria: Correlation Coefficient
Output model

PDB-9cp1:
Post-targeting aCascade Type I-A CRISPR-Cas Surveillance Complexes

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