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- EMDB-49074: FnCas9 scaRNA gRNA 1101 DNA non-productive state -

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ID or keywords:

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Basic information

Entry
Database: EMDB / ID: EMD-49074
TitleFnCas9 scaRNA gRNA 1101 DNA non-productive state
Map dataFnCas9 scaRNA gRNA 1101 DNA non-productive state
Sample
  • Complex: Ternary complex of FnCas9 with scaRNA gRNA and 1101 DNA
    • Protein or peptide: CRISPR-associated endonuclease Cas9
    • DNA: 1101 TS
    • DNA: 1101 NTS
    • RNA: scaRNA gRNA
KeywordsCRISPR / Cas9 / IMMUNE SYSTEM / Hydrolase-DNA-RNA complex
Function / homology
Function and homology information


endonuclease activity / defense response to virus / Hydrolases; Acting on ester bonds / DNA binding / RNA binding / metal ion binding
Similarity search - Function
CRISPR system subtype II-B RNA-guided endonuclease Cas9/Csx12 / : / : / CRISPR-associated endonuclease Cas9 alpha-helical lobe / CRISPR-associated endonuclease Cas9, C-terminal domain / Cas9-type HNH domain / Cas9-type HNH domain profile.
Similarity search - Domain/homology
CRISPR-associated endonuclease Cas9
Similarity search - Component
Biological speciesFrancisella tularensis subsp. novicida (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.77 Å
AuthorsHibshman GN / Taylor DW
Funding support United States, 2 items
OrganizationGrant numberCountry
Welch FoundationF-1938 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM138348 United States
CitationJournal: Nucleic Acids Res. / Year: 2025
Title: Structural basis of a dual-function type II-B CRISPR-Cas9
Authors: Hibshman GN / Taylor DW
History
DepositionFeb 5, 2025-
Header (metadata) releaseOct 1, 2025-
Map releaseOct 1, 2025-
UpdateOct 1, 2025-
Current statusOct 1, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_49074.png

Downloads & links

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Map

FileDownload / File: emd_49074.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFnCas9 scaRNA gRNA 1101 DNA non-productive state
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.93 Å/pix.
x 420 pix.
= 391.86 Å		0.93 Å/pix.
x 420 pix.
= 391.86 Å		0.93 Å/pix.
x 420 pix.
= 391.86 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.933 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.162
Minimum - Maximum-0.3795775 - 1.0539882
Average (Standard dev.)0.00006495739 (±0.013896989)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions420420420
Spacing420420420
CellA=B=C: 391.86002 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half Map B

Fileemd_49074_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_49074_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 : Ternary complex of FnCas9 with scaRNA gRNA and 1101 DNA

EntireName: Ternary complex of FnCas9 with scaRNA gRNA and 1101 DNA
Components
  • Complex: Ternary complex of FnCas9 with scaRNA gRNA and 1101 DNA
    • Protein or peptide: CRISPR-associated endonuclease Cas9
    • DNA: 1101 TS
    • DNA: 1101 NTS
    • RNA: scaRNA gRNA

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Supramolecule #1: Ternary complex of FnCas9 with scaRNA gRNA and 1101 DNA

SupramoleculeName: Ternary complex of FnCas9 with scaRNA gRNA and 1101 DNA
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #4, #1-#3
Source (natural)Organism: Francisella tularensis subsp. novicida (bacteria)
Molecular weightTheoretical: 190.329 KDa

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Macromolecule #1: 1101 TS

MacromoleculeName: 1101 TS / type: dna / ID: 1 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Francisella tularensis subsp. novicida (bacteria)
Molecular weightTheoretical: 7.912127 KDa
SequenceString:
(DG)(DG)(DA)(DT)(DT)(DT)(DC)(DT)(DC)(DC) (DA)(DT)(DT)(DA)(DC)(DA)(DG)(DC)(DT)(DA) (DA)(DT)(DT)(DA)(DC)(DT)

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Macromolecule #2: 1101 NTS

MacromoleculeName: 1101 NTS / type: dna / ID: 2 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Francisella tularensis subsp. novicida (bacteria)
Molecular weightTheoretical: 3.719468 KDa
SequenceString:
(DA)(DA)(DT)(DG)(DG)(DA)(DG)(DA)(DA)(DA) (DT)(DC)

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Macromolecule #3: scaRNA gRNA

MacromoleculeName: scaRNA gRNA / type: rna / ID: 3 / Number of copies: 1
Source (natural)Organism: Francisella tularensis subsp. novicida (bacteria)
Molecular weightTheoretical: 27.886447 KDa
SequenceString:
AAGAUUAGCU GUAAGUUUCA GUUGCGCCGA AAGGCGCUCU GUAAUCAUUU AAAAGUAUUU UGAACGGACC UCUGUUUGAC ACGUCUG

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Macromolecule #4: CRISPR-associated endonuclease Cas9

MacromoleculeName: CRISPR-associated endonuclease Cas9 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO / EC number: Hydrolases; Acting on ester bonds
Source (natural)Organism: Francisella tularensis subsp. novicida (bacteria)
Molecular weightTheoretical: 190.625 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: NFKILPIAID LGVKNTGVFS AFYQKGTSLE RLDNKNGKVY ELSKDSYTLL MNNRTARRHQ RRGIDRKQLV KRLFKLIWTE QLNLEWDKD TQQAISFLFN RRGFSFITDG YSPEYLNIVP EQVKAILMDI FDDYNGEDDL DSYLKLATEQ ESKISEIYNK L MQKILEFK ...String:
NFKILPIAID LGVKNTGVFS AFYQKGTSLE RLDNKNGKVY ELSKDSYTLL MNNRTARRHQ RRGIDRKQLV KRLFKLIWTE QLNLEWDKD TQQAISFLFN RRGFSFITDG YSPEYLNIVP EQVKAILMDI FDDYNGEDDL DSYLKLATEQ ESKISEIYNK L MQKILEFK LMKLCTDIKD DKVSTKTLKE ITSYEFELLA DYLANYSESL KTQKFSYTDK QGNLKELSYY HHDKYNIQEF LK RHATIND RILDTLLTDD LDIWNFNFEK FDFDKNEEKL QNQEDKDHIQ AHLHHFVFAV NKIKSEMASG GRHRSQYFQE ITN VLDENN HQEGYLKNFC ENLHNKKYSN LSVKNLVNLI GNLSNLELKP LRKYFNDKIH AKADHWDEQK FTETYCHWIL GEWR VGVKD QDKKDGAKYS YKDLCNELKQ KVTKAGLVDF LLELDPCRTI PPYLDNNNRK PPKCQSLILN PKFLDNQYPN WQQYL QELK KLQSIQNYLD SFETDLKVLK SSKDQPYFVE YKSSNQQIAS GQRDYKDLDA RILQFIFDRV KASDELLLNE IYFQAK KLK QKASSELEKL ESSKKLDEVI ANSQLSQILK SQHTNGIFEQ GTFLHLVCKY YKQRQRARDS RLYIMPEYRY DKKLHKY NN TGRFDDDNQL LTYCNHKPRQ KRYQLLNDLA GVLQVSPNFL KDKIGSDDDL FISKWLVEHI RGFKKACEDS LKIQKDNR G LLNHKINIAR NTKGKCEKEI FNLICKIEGS EDKKGNYKHG LAYELGVLLF GEPNEASKPE FDRKIKKFNS IYSFAQIQQ IAFAERKGNA NTCAVCSADN AHRMQQIKIT EPVEDNKDKI ILSAKAQRLP AIPTRIVDGA VKKMATILAK NIVDDNWQNI KQVLSAKHQ LHIPIITESN AFEFEPALAD VKGKSLKDRR KKALERISPE NIFKDKNNRI KEFAKGISAY SGANLTDGDF D GAKEELDH IIPRSHKKYG TLNDEANLIC VTRGDNKNKG NRIFCLRDLA DNYKLKQFET TDDLEIEKKI ADTIWDANKK DF KFGNYRS FINLTPQEQK AFRHALFLAD ENPIKQAVIR AINNRNRTFV NGTQRYFAEV LANNIYLRAK KENLNTDKIS FDY FGIPTI GNGRGIAEIR QLYEKVDSDI QAYAKGDKPQ ASYSHLIDAM LAFCIAADEH RNDGSIGLEI DKNYSLYPLD KNTG EVFTK DIFSQIKITD NEFSDKKLVR KKAIEGFNTH RQMTRDGIYA ENYLPILIHK ELNEVRKGYT WKNSEEIKIF KGKKY DIQQ LNNLVYCLKF VDKPISIDIQ ISTLEELRNI LTTNNIAATA EYYYINLKTQ KLHEYYIENY NTALGYKKYS KEMEFL RSL AYRSERVKIK SIDDVKQVLD KDSNFIIGKI TLPFKKEWQR LYREWQNTTI KDDYEFLKSF FNVKSITKLH KKVRKDF SL PISTNEGKFL VKRKTWDNNF IYQILNDSDS RADGTKPFIP AFDISKNEIV EAIIDSFTSK NIFWLPKNIE LQKVDNKN I FAIDTSKWFE VETPSDLRDI GIATIQYKID NNSRPKVRVK LDYVIDDDSK INYFMNHSLL KSRYPDKVLE ILKQSTIIE FESSGFNKTI KEMLGMKLAG IYNETSNN

UniProtKB: CRISPR-associated endonuclease Cas9

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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: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS GLACIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.5 µm

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.77 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 283943
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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