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- EMDB-14244: elongated Cascade complex from type I-A CRISPR-Cas system -

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Basic information

Entry
Database: EMDB / ID: EMD-14244
Titleelongated Cascade complex from type I-A CRISPR-Cas system
Map datapao-elongate
Sample
  • Complex: elongated Cascade complex from Pyrococcus furiosus type I-A CRISPR/Cas system
    • Complex: Cas11a, Cas7a and Type I-A CRISPR-associated protein Cas5
      • Protein or peptide: Cas11a
      • Protein or peptide: Cas7a
      • Protein or peptide: Type I-A CRISPR-associated protein Cas5
    • Complex: CrRNA (62-MER)
      • RNA: CrRNA (62-MER)
Keywordselongated Cascade complex / cascade / type I-A / genome editing / ANTIVIRAL PROTEIN
Function / homologyCRISPR-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 / defense response to virus / Type I-A CRISPR-associated protein Cas5 / Uncharacterized protein / Type I-A CRISPR-associated protein Cas7/Csa2
Function and homology information
Biological speciesPyrococcus furiosus DSM 3638 (archaea) / Escherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsHu C / Ni D
Funding support United States, Switzerland, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM118117 United States
Swiss National Science Foundation(NCCR) TransCure Switzerland
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM118160 United States
CitationJournal: To Be Published
Title: Structural snapshots for an atypic type I CRISPR-Cas system
Authors: Ni D / Hu C / Nam KH / Terns M / Stahlberg H / Ke A
History
DepositionFeb 4, 2022-
Header (metadata) releaseAug 16, 2023-
Map releaseAug 16, 2023-
UpdateAug 16, 2023-
Current statusAug 16, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_14244.png

Downloads & links

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Map

FileDownload / File: emd_14244.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationpao-elongate
Voxel sizeX=Y=Z: 1.3667 Å
Density
Contour LevelBy AUTHOR: 0.574
Minimum - Maximum-3.8518164 - 7.654628
Average (Standard dev.)-0.0024215458 (±0.18068118)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 410.01 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : elongated Cascade complex from Pyrococcus furiosus type I-A CRISP...

EntireName: elongated Cascade complex from Pyrococcus furiosus type I-A CRISPR/Cas system
Components
  • Complex: elongated Cascade complex from Pyrococcus furiosus type I-A CRISPR/Cas system
    • Complex: Cas11a, Cas7a and Type I-A CRISPR-associated protein Cas5
      • Protein or peptide: Cas11a
      • Protein or peptide: Cas7a
      • Protein or peptide: Type I-A CRISPR-associated protein Cas5
    • Complex: CrRNA (62-MER)
      • RNA: CrRNA (62-MER)

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Supramolecule #1: elongated Cascade complex from Pyrococcus furiosus type I-A CRISP...

SupramoleculeName: elongated Cascade complex from Pyrococcus furiosus type I-A CRISPR/Cas system
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all / Details: Cascade alone
Molecular weightTheoretical: 550 KDa

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Supramolecule #2: Cas11a, Cas7a and Type I-A CRISPR-associated protein Cas5

SupramoleculeName: Cas11a, Cas7a and Type I-A CRISPR-associated protein Cas5
type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#3
Source (natural)Organism: Pyrococcus furiosus DSM 3638 (archaea)

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Supramolecule #3: CrRNA (62-MER)

SupramoleculeName: CrRNA (62-MER) / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #4
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: Cas11a

MacromoleculeName: Cas11a / type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO
Source (natural)Organism: Pyrococcus furiosus DSM 3638 (archaea) / Strain: ATCC 43587 / DSM 3638 / JCM 8422 / Vc1
Molecular weightTheoretical: 12.208933 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
GGWIRNIGRY LSYLVDDTFE EYAYDVVDGI AKARTQEELL EGVYKALRLA PKLKKKAESK GCPPPRIPSP EDIEALEEKV EQLSNPKDL RKLAVSLALW AFASWNNCP

UniProtKB: Uncharacterized protein

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Macromolecule #2: Cas7a

MacromoleculeName: Cas7a / type: protein_or_peptide / ID: 2 / Number of copies: 10 / Enantiomer: LEVO
Source (natural)Organism: Pyrococcus furiosus DSM 3638 (archaea) / Strain: ATCC 43587 / DSM 3638 / JCM 8422 / Vc1
Molecular weightTheoretical: 36.989148 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MYVRISGRIR LNAHSLNAQG GGGTNYIEIT KTKVTVRTEN GWTVVEVPAI TGNMLKHWHF VGFVDYFKTT PYGVNLTERA LRYNGTRFG QGETTATKAN GATVQLNDEA TIIKELADAD VHGFLAPKTG RRRVSLVKAS FILPTEDFIK EVEGERLITA I KHNRVDVD ...String:
MYVRISGRIR LNAHSLNAQG GGGTNYIEIT KTKVTVRTEN GWTVVEVPAI TGNMLKHWHF VGFVDYFKTT PYGVNLTERA LRYNGTRFG QGETTATKAN GATVQLNDEA TIIKELADAD VHGFLAPKTG RRRVSLVKAS FILPTEDFIK EVEGERLITA I KHNRVDVD EKGAIGSSKE GTAQMLFSRE YATGLYGFSI VLDLGLVGIP QGLPVKFEEN QPRPNIVIDP NERKARIESA LK ALIPMLS GYIGANLARS FPVFKVEELV AIASEGPIPA LVHGFYEDYI EANRSIIKNA RALGFNIEVF TYNVDLGEDI EAT KVSSVE ELVANLVKMV

UniProtKB: Type I-A CRISPR-associated protein Cas7/Csa2

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Macromolecule #3: Type I-A CRISPR-associated protein Cas5

MacromoleculeName: Type I-A CRISPR-associated protein Cas5 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pyrococcus furiosus DSM 3638 (archaea) / Strain: ATCC 43587 / DSM 3638 / JCM 8422 / Vc1
Molecular weightTheoretical: 29.147219 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MDILLVCLRF PFFSVAKRSY QVRTSFLLPP PSALKGALAK GLILLKPEKY ASSSLDEAAL KAIKEIESKL VDIKAVSVAP LSPLIRNAF LLKRLRNLES GSNAEKSDAM RREYTFTREL LVAYIFKNLT QEEKNLYLKA AMLIDVIGDT ESLATPVWAS F VKPEDKKA ...String:
MDILLVCLRF PFFSVAKRSY QVRTSFLLPP PSALKGALAK GLILLKPEKY ASSSLDEAAL KAIKEIESKL VDIKAVSVAP LSPLIRNAF LLKRLRNLES GSNAEKSDAM RREYTFTREL LVAYIFKNLT QEEKNLYLKA AMLIDVIGDT ESLATPVWAS F VKPEDKKA PLAFSAPYTE IYSLLSSKIQ AKGKIRMYIE KMRVSPEYSK TKGPQEEIFY LPIEERRYKR IVYYARIYPP EV EKALTVD GEVLGIWIP

UniProtKB: Type I-A CRISPR-associated protein Cas5

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Macromolecule #4: CrRNA (62-MER)

MacromoleculeName: CrRNA (62-MER) / type: rna / ID: 4 / Number of copies: 1
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 19.97984 KDa
SequenceString:
AUUGAAAGUU GUAGUAUGCG GUCCUUGCGG CUGAGAGCAC UUCAGGAGUU GCCCGCGCCA GC

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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

Concentration0.8 mg/mL
BufferpH: 7.5 / Component - Concentration: 150.0 mM / Component - Formula: NaClSodium chloride / Component - Name: sodium chloride
GridModel: Quantifoil / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 20 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV / Details: 4s force 0.

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated defocus min: 0.8 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number real images: 8500 / Average electron dose: 70.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC
Final 3D classificationNumber classes: 4 / Software - Name: cryoSPARC
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 240851
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: AB INITIO MODEL
Output model

PDB-7r21:
elongated Cascade complex from type I-A CRISPR-Cas system

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