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- EMDB-14245: elongated Cascade complex from type I-A CRISPR-Cas system -

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Basic information

Entry
Database: EMDB / ID: EMD-14245
Titleelongated Cascade complex from type I-A CRISPR-Cas system
Map datapartial
Sample
  • Complex: elongated Cascade complex from Pyrococcus furiosus type I-A CRISPR/Cas system
    • Protein or peptide: CRISPR-associated helicase Cas3
    • Protein or peptide: Cas11a
    • DNA: DNA (5'-D(P*TP*AP*CP*AP*AP*GP*GP*GP*A)-3')
    • Protein or peptide: Cas7a
    • Protein or peptide: CRISPR-associated endonuclease Cas3-HD
    • Protein or peptide: Type I-A CRISPR-associated protein Cas5
    • RNA: crRNA (57-MER)
    • DNA: DNA (5'-D(P*TP*CP*CP*C)-3')
    • Protein or peptide: Cas8Roberval (Air Saguenay) Water Aerodrome
  • Ligand: NICKEL (II) ION
Keywordselongated Cascade complex / cascade / type I-A / genome editing / ANTIVIRAL PROTEIN
Function / homology
Function and homology information


exonuclease activity / helicase activity / defense response to virus / endonuclease activity / Hydrolases; Acting on ester bonds / nucleic acid binding / hydrolase activity / ATP binding / metal ion binding
Similarity search - Function
CRISPR-associated protein, MJ0385 / CRISPR-associated protein (Cas_Csa4) / 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 / Helicase Cas3, CRISPR-associated, core / CRISPR-associated Cas3-type HD domain / CRISPR-associated Cas3-type HD domain superfamily / Cas3, HD domain ...CRISPR-associated protein, MJ0385 / CRISPR-associated protein (Cas_Csa4) / 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 / Helicase Cas3, CRISPR-associated, core / CRISPR-associated Cas3-type HD domain / CRISPR-associated Cas3-type HD domain superfamily / Cas3, HD domain / HD Cas3-type domain profile. / CRISPR-associated protein Cas5, N-terminal / DEAD/DEAH box helicase / DEAD/DEAH box helicase domain / Helicase conserved C-terminal domain / helicase superfamily c-terminal domain / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
CRISPR-associated helicase Cas3 / Type I-A CRISPR-associated protein Cas5 / Uncharacterized protein / Type I-A CRISPR-associated protein Cas7/Csa2 / CRISPR-associated endonuclease Cas3-HD / Type I-A CRISPR-associated protein Cas8a2/Csa4
Similarity search - Component
Biological speciesPyrococcus furiosus DSM 3638 (archaea) / Escherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsHu C / Ni D / Nam KH / Terns M / Stahlberg H / Ke A
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_14245.png

Downloads & links

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Map

FileDownload / File: emd_14245.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationpartial
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.33 Å/pix.
x 320 pix.
= 426.4 Å		1.33 Å/pix.
x 320 pix.
= 426.4 Å		1.33 Å/pix.
x 320 pix.
= 426.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.3325 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.16
Minimum - Maximum-0.85244215 - 2.5720727
Average (Standard dev.)-0.0021123928 (±0.07153428)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 426.4 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
    • Protein or peptide: CRISPR-associated helicase Cas3
    • Protein or peptide: Cas11a
    • DNA: DNA (5'-D(P*TP*AP*CP*AP*AP*GP*GP*GP*A)-3')
    • Protein or peptide: Cas7a
    • Protein or peptide: CRISPR-associated endonuclease Cas3-HD
    • Protein or peptide: Type I-A CRISPR-associated protein Cas5
    • RNA: crRNA (57-MER)
    • DNA: DNA (5'-D(P*TP*CP*CP*C)-3')
    • Protein or peptide: Cas8Roberval (Air Saguenay) Water Aerodrome
  • Ligand: NICKEL (II) ION

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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: #1-#9 / Details: Cascade alone
Molecular weightTheoretical: 550 KDa

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Macromolecule #1: CRISPR-associated helicase Cas3

MacromoleculeName: CRISPR-associated helicase Cas3 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pyrococcus furiosus DSM 3638 (archaea) / Strain: ATCC 43587 / DSM 3638 / JCM 8422 / Vc1
Molecular weightTheoretical: 74.475742 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: ELTGFEPYDY QLRAWEKIRE IMNNGGKVII EVPTAGGKTE TAVMPFFAGI YNNNWPVARL VYVLPTRSLV EKQAERLRNL VYKLLQLKG KSKEEAEKLA RELVVVEYGL EKTHAFLGWV VVTTWDAFLY GLAAHRTVGN RFTFPAGAIA QSLVIFDEVQ M YQDESMYM ...String:
ELTGFEPYDY QLRAWEKIRE IMNNGGKVII EVPTAGGKTE TAVMPFFAGI YNNNWPVARL VYVLPTRSLV EKQAERLRNL VYKLLQLKG KSKEEAEKLA RELVVVEYGL EKTHAFLGWV VVTTWDAFLY GLAAHRTVGN RFTFPAGAIA QSLVIFDEVQ M YQDESMYM PRLLSLVVGI LEEANVPLVI MSATIPSKLR EMIAGDTEVI TVDKNDKNKP SKGNVKVRLV EGDITDVLND IK KILKNGK KVLVVRNTVR KAVETYQVLK KKLNDTLANP SDALLIHSRF TIGDRREKER ALDSARLIVA TQVVEAGLDL PNV GLVVTD IAPLDALIQR IGRCARRPGE EGEGIILIPV ENCIEHEKIV RGLSELMEKI GEDTVVFAVV TSTNEYDRVV EIHY GEGKK NFVYVGDIDT ARRVLEKKRS KKLPKDLYII PYSVSPYPDP LVLLTTYDEL SKIGEYLADT TKARKALDRV YKFHY ENNI VPKEFASAYI YFKELKLFSA PPEYEKRSRP ELYVLLYPMN IEKNERVEDK VIDNLETARI IRISYSVKEW KKSDVV IGR LMKEWDKNAE KWVWKVRKSF KIDPYEIYVI DAKYYNSELG FITNLSDTNS HTDSDSKVRT RNSEHSSKKN RSKGKKG QT SLENWGVRV

UniProtKB: CRISPR-associated helicase Cas3

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Macromolecule #2: Cas11a

MacromoleculeName: Cas11a / type: protein_or_peptide / ID: 2 / 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 #4: Cas7a

MacromoleculeName: Cas7a / type: protein_or_peptide / ID: 4 / 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 #5: CRISPR-associated endonuclease Cas3-HD

MacromoleculeName: CRISPR-associated endonuclease Cas3-HD / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO / EC number: Hydrolases; Acting on ester bonds
Source (natural)Organism: Pyrococcus furiosus DSM 3638 (archaea) / Strain: ATCC 43587 / DSM 3638 / JCM 8422 / Vc1
Molecular weightTheoretical: 27.130014 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: SCKAFQGQTL REHIEAMLAA WEIVKNKYIP SIIRVMKTVG VKFTEEDADK FMKTLIILHD VGKCSEVYQK HLSNNEPLRG FRHELVSAY YAYNILKDMF KDETIAFIGA LVVMMHHEPI LMGQIRSLDK EELTPEVVLD KLRTFNGVME GTESFIKSMI K EKLGVIPK ...String:
SCKAFQGQTL REHIEAMLAA WEIVKNKYIP SIIRVMKTVG VKFTEEDADK FMKTLIILHD VGKCSEVYQK HLSNNEPLRG FRHELVSAY YAYNILKDMF KDETIAFIGA LVVMMHHEPI LMGQIRSLDK EELTPEVVLD KLRTFNGVME GTESFIKSMI K EKLGVIPK VPSPTQEDVL REVIRLSVLA RHRPDSGKLR MVVGALLIPL VLCDYKGAKE REGESPKFAE VLRVEMMKAA

UniProtKB: CRISPR-associated endonuclease Cas3-HD

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Macromolecule #6: Type I-A CRISPR-associated protein Cas5

MacromoleculeName: Type I-A CRISPR-associated protein Cas5 / type: protein_or_peptide / ID: 6 / 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 #9: Cas8

MacromoleculeName: Cas8 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pyrococcus furiosus DSM 3638 (archaea) / Strain: ATCC 43587 / DSM 3638 / JCM 8422 / Vc1
Molecular weightTheoretical: 38.800719 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: FNEFKTPQID PIFDLYVAYG YVVSLIRGGA KEATLIPHGA SYLIQTDVSN EEFRHGLVDA LSSMLSLHIA LARHSPREGG KLVSDADFS AGANINNVYW DSVPRNLEKL MKDLEKKRSV KGTATIPITL MPSAGKYMLK HFGVQGGNPI KVDLLNYALA W VGFHYYTP ...String:
FNEFKTPQID PIFDLYVAYG YVVSLIRGGA KEATLIPHGA SYLIQTDVSN EEFRHGLVDA LSSMLSLHIA LARHSPREGG KLVSDADFS AGANINNVYW DSVPRNLEKL MKDLEKKRSV KGTATIPITL MPSAGKYMLK HFGVQGGNPI KVDLLNYALA W VGFHYYTP YIKYAKGDTT WIHIYQIAPV EEVDMISILS LKDLKMHLPH YYESNLDFLI NRRLALLYHL LHSEALELFT EK EFVIHSY TLERSGNNQA IRSFEEEEIG KLMDFLWKLK RRDFYHAIKF IDDLLKKATE GALALIDAIM NERLEGFYTA LKL GKKAGV VSSREIVAAL EDIIC

UniProtKB: Type I-A CRISPR-associated protein Cas8a2/Csa4

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Macromolecule #3: DNA (5'-D(P*TP*AP*CP*AP*AP*GP*GP*GP*A)-3')

MacromoleculeName: DNA (5'-D(P*TP*AP*CP*AP*AP*GP*GP*GP*A)-3') / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 2.788862 KDa
SequenceString:
(DT)(DA)(DC)(DA)(DA)(DG)(DG)(DG)(DA)

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Macromolecule #8: DNA (5'-D(P*TP*CP*CP*C)-3')

MacromoleculeName: DNA (5'-D(P*TP*CP*CP*C)-3') / type: dna / ID: 8 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 1.126781 KDa
SequenceString:
(DT)(DC)(DC)(DC)

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Macromolecule #7: crRNA (57-MER)

MacromoleculeName: crRNA (57-MER) / type: rna / ID: 7 / Number of copies: 1
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 18.389883 KDa
SequenceString:
AUUGAAAGUU GUAGUAUGCG GUCCUUGCGG CUGAGAGCAC UUCAGGAGUU GCCCGCG

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Macromolecule #10: NICKEL (II) ION

MacromoleculeName: NICKEL (II) ION / type: ligand / ID: 10 / Number of copies: 2 / Formula: NI
Molecular weightTheoretical: 58.693 Da
Chemical component information

ChemComp-NI:
NICKEL (II) ION / Nickel

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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 R2/1 / Support film - Material: CARBON / Pretreatment - Type: GLOW DISCHARGE
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: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 49699
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: AB INITIO MODEL
Output model

PDB-7r2k:
elongated Cascade complex from type I-A CRISPR-Cas system

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