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- EMDB-49174: Cryo-EM structure of the dCas12f-gRNA-dsDNA complex (full R-Loop) -

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Basic information

Entry
Database: EMDB / ID: EMD-49174
TitleCryo-EM structure of the dCas12f-gRNA-dsDNA complex (full R-Loop)
Map data
Sample
  • Complex: dCas12f-gRNA-target DNA complex
    • Protein or peptide: Nuclease-deactivated Cas12f
    • DNA: DNA non-template strand
    • DNA: DNA template strand
    • RNA: gRNA (85-MER)
KeywordsdCas12f / Full R-Loop / RNA BINDING PROTEIN / RNA BINDING PROTEIN-DNA-RNA complex
Biological speciesFlagellimonas taeanensis (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.42 Å
AuthorsChang L / Sternberg SH / Xiao R / Hoffmann FT / Wiegand T / Xie D
Funding support United States, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM138675 United States
National Science Foundation (NSF, United States)2339799 United States
National Science Foundation (NSF, United States)2239685 United States
The Pew Charitable Trusts United States
CitationJournal: To Be Published
Title: Structural basis of RNA-guided transcription via Cas12f-Sigma factor association
Authors: Renjian X / Leifu C
History
DepositionFeb 11, 2025-
Header (metadata) releaseDec 24, 2025-
Map releaseDec 24, 2025-
UpdateDec 24, 2025-
Current statusDec 24, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_49174.png

Downloads & links

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Map

FileDownload / File: emd_49174.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.82 Å/pix.
x 256 pix.
= 210.432 Å		0.82 Å/pix.
x 256 pix.
= 210.432 Å		0.82 Å/pix.
x 256 pix.
= 210.432 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.822 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.03
Minimum - Maximum-0.0017551545 - 2.5478501
Average (Standard dev.)0.0016605813 (±0.02961542)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 210.432 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_49174_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_49174_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 : dCas12f-gRNA-target DNA complex

EntireName: dCas12f-gRNA-target DNA complex
Components
  • Complex: dCas12f-gRNA-target DNA complex
    • Protein or peptide: Nuclease-deactivated Cas12f
    • DNA: DNA non-template strand
    • DNA: DNA template strand
    • RNA: gRNA (85-MER)

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Supramolecule #1: dCas12f-gRNA-target DNA complex

SupramoleculeName: dCas12f-gRNA-target DNA complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Flagellimonas taeanensis (bacteria)

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Macromolecule #1: Nuclease-deactivated Cas12f

MacromoleculeName: Nuclease-deactivated Cas12f / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Flagellimonas taeanensis (bacteria)
Molecular weightTheoretical: 42.913262 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGKSTLKHTR KIQILIDLPT KDEKKEVMDM MYQWRDRCFR AANIIVTHLY VQEMIKDFFY LSEGIKYKLA DEKKDEKGIL QRSRMNTTY RVVSDRFKGE MPTNILSTLN HGLISSFNKN RVQYWKGERS LPNFKKDMAF PFGLQGISRL VYDEEKKAFC F RLYRVPFK ...String:
MGKSTLKHTR KIQILIDLPT KDEKKEVMDM MYQWRDRCFR AANIIVTHLY VQEMIKDFFY LSEGIKYKLA DEKKDEKGIL QRSRMNTTY RVVSDRFKGE MPTNILSTLN HGLISSFNKN RVQYWKGERS LPNFKKDMAF PFGLQGISRL VYDEEKKAFC F RLYRVPFK TYLGKDFTDK RMLLERLVKG DVKLCASNIQ LNGGKIFWLA VFEIEKEKHS LKPEVIAEAS LSLEYPIVVK TG KNRLTIG TKEEFLYRRL AIQAARRRTQ VGATYSRSGK GKKRKLKAVD KYHKTESNYV AHRIHVYSRK LIDFCIKHQA GTL ILMNQE DKVGIAKEEE FVLRNWSYYE LMTKIKYKAE KAGIELIIG

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Macromolecule #2: DNA non-template strand

MacromoleculeName: DNA non-template strand / type: dna / ID: 2 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Flagellimonas taeanensis (bacteria)
Molecular weightTheoretical: 18.349744 KDa
SequenceString: (DC)(DT)(DA)(DG)(DC)(DG)(DT)(DT)(DG)(DT) (DG)(DG)(DG)(DA)(DT)(DA)(DC)(DC)(DC)(DG) (DT)(DT)(DA)(DC)(DA)(DC)(DT)(DA)(DA) (DT)(DT)(DT)(DT)(DT)(DC)(DG)(DG)(DT)(DT) (DC) (DA)(DC)(DG)(DA)(DT)(DA) ...String:
(DC)(DT)(DA)(DG)(DC)(DG)(DT)(DT)(DG)(DT) (DG)(DG)(DG)(DA)(DT)(DA)(DC)(DC)(DC)(DG) (DT)(DT)(DA)(DC)(DA)(DC)(DT)(DA)(DA) (DT)(DT)(DT)(DT)(DT)(DC)(DG)(DG)(DT)(DT) (DC) (DA)(DC)(DG)(DA)(DT)(DA)(DT)(DC) (DC)(DT)(DT)(DT)(DA)(DG)(DT)(DC)(DC)(DC) (DA)(DC)

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Macromolecule #3: DNA template strand

MacromoleculeName: DNA template strand / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Flagellimonas taeanensis (bacteria)
Molecular weightTheoretical: 18.63099 KDa
SequenceString: (DG)(DT)(DG)(DG)(DG)(DA)(DC)(DT)(DA)(DA) (DA)(DG)(DG)(DA)(DT)(DA)(DT)(DC)(DG)(DT) (DG)(DA)(DA)(DC)(DC)(DG)(DA)(DA)(DA) (DA)(DA)(DT)(DT)(DA)(DG)(DT)(DG)(DT)(DA) (DA) (DC)(DG)(DG)(DG)(DT)(DA) ...String:
(DG)(DT)(DG)(DG)(DG)(DA)(DC)(DT)(DA)(DA) (DA)(DG)(DG)(DA)(DT)(DA)(DT)(DC)(DG)(DT) (DG)(DA)(DA)(DC)(DC)(DG)(DA)(DA)(DA) (DA)(DA)(DT)(DT)(DA)(DG)(DT)(DG)(DT)(DA) (DA) (DC)(DG)(DG)(DG)(DT)(DA)(DT)(DC) (DC)(DC)(DA)(DC)(DA)(DA)(DC)(DG)(DC)(DT) (DA)(DG)

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Macromolecule #4: gRNA (85-MER)

MacromoleculeName: gRNA (85-MER) / type: rna / ID: 4 / Number of copies: 1
Source (natural)Organism: Flagellimonas taeanensis (bacteria)
Molecular weightTheoretical: 29.92277 KDa
SequenceString:
AUUUUGAGGG UGCUUGUACA CCCAUAGGGU GAGGUUAAGA AUUACACUCA CUAAGUGUGA ACAACACAUA CAACUUGUGG GAUAUACGC UAAC

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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: 7.6
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 59.2 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: DARK FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING ONLY
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.42 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 16447
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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