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- EMDB-39167: Type I-FHNH Cascade-dsDNA intermediate complex -

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Basic information

Entry
Database: EMDB / ID: EMD-39167
TitleType I-FHNH Cascade-dsDNA intermediate complex
Map data
Sample
  • Complex: Type I-FHNH Cascade-dsDNA intermediate complex
    • Protein or peptide: Cas5f
    • Protein or peptide: Cas6f
    • RNA: 60-nt crRNA
    • Protein or peptide: Cas7f
    • Protein or peptide: Cas8f fusion with HNH
    • Other: TS
    • Other: NTS
Keywordsprotein / RNA / RNA BINDING PROTEIN/DNA/RNA / RNA BINDING PROTEIN-DNA-RNA complex
Biological speciesSelenomonas sp. (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsLi Z
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Mol.Cell / Year: 2024
Title: Mechanisms for HNH-mediated target DNA cleavage in type I CRISPR-Cas systems
Authors: Li Z
History
DepositionFeb 19, 2024-
Header (metadata) releaseJul 31, 2024-
Map releaseJul 31, 2024-
UpdateJul 31, 2024-
Current statusJul 31, 2024Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_39167.png

Downloads & links

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Map

FileDownload / File: emd_39167.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 320 pix.
= 272. Å		0.85 Å/pix.
x 320 pix.
= 272. Å		0.85 Å/pix.
x 320 pix.
= 272. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.85 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.006
Minimum - Maximum-0.010853409 - 0.028041286
Average (Standard dev.)0.0000465505 (±0.0013339942)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 272.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_39167_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_39167_half_map_2.map
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-FHNH Cascade-dsDNA intermediate complex

EntireName: Type I-FHNH Cascade-dsDNA intermediate complex
Components
  • Complex: Type I-FHNH Cascade-dsDNA intermediate complex
    • Protein or peptide: Cas5f
    • Protein or peptide: Cas6f
    • RNA: 60-nt crRNA
    • Protein or peptide: Cas7f
    • Protein or peptide: Cas8f fusion with HNH
    • Other: TS
    • Other: NTS

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Supramolecule #1: Type I-FHNH Cascade-dsDNA intermediate complex

SupramoleculeName: Type I-FHNH Cascade-dsDNA intermediate complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#7
Source (natural)Organism: Selenomonas sp. (bacteria)

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Macromolecule #1: Cas5f

MacromoleculeName: Cas5f / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Selenomonas sp. (bacteria)
Molecular weightTheoretical: 28.703135 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MMKGYILLEK VNIENANAFN NIIVGIPAIT SFLGFARALE RKLNAKEIAI RINGVGLEFH EYELKGYKNK RGQYVTSCPL PGSIPGQNE KKLDAHIMNQ AYIDLNMSFL LEVEGPHVDM STCKSIKSTM ETLRIAGGII RNYKKIRLID TLADIPYGYF L TLRQDNLN ...String:
MMKGYILLEK VNIENANAFN NIIVGIPAIT SFLGFARALE RKLNAKEIAI RINGVGLEFH EYELKGYKNK RGQYVTSCPL PGSIPGQNE KKLDAHIMNQ AYIDLNMSFL LEVEGPHVDM STCKSIKSTM ETLRIAGGII RNYKKIRLID TLADIPYGYF L TLRQDNLN DAAGDDMLDK MIHALQQEDT LVPIAVGFKA LSEVGHVEGQ RDPEKDHCFV ESIFSLGGFE CSKILEDINS CL WRYKTEE GLYLCTII

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Macromolecule #2: Cas6f

MacromoleculeName: Cas6f / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Selenomonas sp. (bacteria)
Molecular weightTheoretical: 20.735873 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MFSQILIIKP GTGISPNIII SEDIFPVLHS LFVEHDKKFG ITFPAYSFDK KGHLGNIIEV LSEDKEALAS LCLEEHLAEV TDYVKVKKE ITFTDDYVLF KRIREENQYE TTARRMRKRG HTELGRPLEM HIKKKNQQIF CHAYIKVKSA STGQSYNIFL A PTDIKHGS FSAYGLLRGD THA

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Macromolecule #4: Cas7f

MacromoleculeName: Cas7f / type: protein_or_peptide / ID: 4 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Selenomonas sp. (bacteria)
Molecular weightTheoretical: 38.700172 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MAANKKATNV TLKSRPENLS FARCLNTTEA KFWQTDFLKR HTFKLPLLIT DKAVLASKGH EMPPDKLEKE IMDPNPQKSQ SCTLSTECD TLRIDFGIKV LPVKESMYSC SDYNYRTAIY QKIDEYIAED GFLTLAKRYV NNIANARFLW RNRKGAEIIE T IVTIEDKE ...String:
MAANKKATNV TLKSRPENLS FARCLNTTEA KFWQTDFLKR HTFKLPLLIT DKAVLASKGH EMPPDKLEKE IMDPNPQKSQ SCTLSTECD TLRIDFGIKV LPVKESMYSC SDYNYRTAIY QKIDEYIAED GFLTLAKRYV NNIANARFLW RNRKGAEIIE T IVTIEDKE YPSFNSKSFN LDTFVEDNAT INEIAQQIAD TFAGKREYLN IYVTCFVKIG CAMEVYPSQE MTFDDDDKGK KL FKFEGSA GMHSQKINNA LRTIDTWYPD YTTYEFPIPV ENYGAARSIG IPFRPDTKSF YKLIDRMILK NEDLPIEDKH YVM AILIRG GMFSKKQEK

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Macromolecule #5: Cas8f fusion with HNH

MacromoleculeName: Cas8f fusion with HNH / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Selenomonas sp. (bacteria)
Molecular weightTheoretical: 39.339742 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MLRNKILAAI SQKIPEEQKI NKYIEGLFQS IDKNHLATHV AKFTETNSPG NIGAYDILSS DMNCGYLDTA NAGWKEPDIV TNDAKYKRP QGFVAMEMSD GRTVMEHLQE DSAELRHEME ELTDKYDEIR DGILNMPSMQ PYRTNQFIKQ VFFPVGGSYH L LSILPSTV ...String:
MLRNKILAAI SQKIPEEQKI NKYIEGLFQS IDKNHLATHV AKFTETNSPG NIGAYDILSS DMNCGYLDTA NAGWKEPDIV TNDAKYKRP QGFVAMEMSD GRTVMEHLQE DSAELRHEME ELTDKYDEIR DGILNMPSMQ PYRTNQFIKQ VFFPVGGSYH L LSILPSTV LNYEVSDRLY RSKIPKIRLR LLSSNAASTT GSRLVSKNKW PLVFQALPPK FLEKNLAKAL DKEYLLPDIN ID ELEGVDN GCLIDEALLP LIIDEGKRKG EGNYRPRHLR DERKEETVQA FLDKYGYCNI PVGYEVHHIV PLSQGGADSI KNM IMLSIE HHERVTEAHA SYFKWRNT

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Macromolecule #3: 60-nt crRNA

MacromoleculeName: 60-nt crRNA / type: rna / ID: 3 / Number of copies: 1
Source (natural)Organism: Selenomonas sp. (bacteria)
Molecular weightTheoretical: 19.440611 KDa
SequenceString:
UUUAGAAGGA GAAGUCAUUU AAUAAGGCCA CUGUUAAAAA GUGUACCGCC GGAUAGGCGG

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Macromolecule #6: TS

MacromoleculeName: TS / type: other / ID: 6 / Number of copies: 1
Classification: polydeoxyribonucleotide/polyribonucleotide hybrid
Source (natural)Organism: Selenomonas sp. (bacteria)
Molecular weightTheoretical: 10.086293 KDa
SequenceString:
(DG)(DT)GGCC(DT)(DT)A(DT) (DT)AAA(DT)GAC(DT)(DT) C(DT)CCGC(DT)AA(DT) AC
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)

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Macromolecule #7: NTS

MacromoleculeName: NTS / type: other / ID: 7 / Number of copies: 1
Classification: polydeoxyribonucleotide/polyribonucleotide hybrid
Source (natural)Organism: Selenomonas sp. (bacteria)
Molecular weightTheoretical: 3.541243 KDa
SequenceString:
G(DT)A(DT)(DT)AGCGG A
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)

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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

BufferpH: 7
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 54.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.2 µm / Nominal defocus min: 1.2 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 88977
Initial angle assignmentType: RANDOM ASSIGNMENT
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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