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- EMDB-43770: Structure of Acidothermus cellulolyticus Cas9 bound with methylat... -

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Basic information

Entry
Database: EMDB / ID: EMD-43770
TitleStructure of Acidothermus cellulolyticus Cas9 bound with methylated DNA
Map data
Sample
  • Complex: CRISPR-Cas9
    • Protein or peptide: CRISPR-associated endonuclease, Csn1 family
    • RNA: RNA (92-MER)
    • DNA: DNA (25-MER)
    • DNA: DNA/RNA (5'-D(P*TP*AP*CP*A)-R(P*(5MC))-D(P*CP*AP*AP*GP*CP*T)-3')
KeywordsCRISPR-Cas9 / Methylated DNA / AceCas9 / HNH rotation / DNA BINDING PROTEIN-DNA-RNA complex
Function / homology
Function and homology information


endonuclease activity / defense response to virus / DNA binding / RNA binding / zinc ion binding
Similarity search - Function
CRISPR-associated endonuclease Cas9, beta-hairpin domain / Cas9 C-terminal domain / Cas9, topo homolgy domain / CRISPR-associated endonuclease Cas9 beta-hairpin domain / Topo homolgy domain in CRISPR-associated endonuclease Cas9 / Cas9 C-terminal domain / : / Cas9, alpha-helical lobe domain / Cas9 alpha-helical lobe domain / HNH endonuclease ...CRISPR-associated endonuclease Cas9, beta-hairpin domain / Cas9 C-terminal domain / Cas9, topo homolgy domain / CRISPR-associated endonuclease Cas9 beta-hairpin domain / Topo homolgy domain in CRISPR-associated endonuclease Cas9 / Cas9 C-terminal domain / : / Cas9, alpha-helical lobe domain / Cas9 alpha-helical lobe domain / HNH endonuclease / HNH endonuclease / RuvC endonuclease subdomain 3 / RuvC endonuclease subdomain 3 / HNH nucleases / CRISPR-associated endonuclease Cas9 / Cas9-type HNH domain / Cas9-type HNH domain profile. / HNH nuclease / Ribonuclease H superfamily
Similarity search - Domain/homology
CRISPR-associated endonuclease, Csn1 family
Similarity search - Component
Biological speciesAcidothermus cellulolyticus (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.88 Å
AuthorsDas A / Rai J / Wang B / Li H
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM099604 United States
CitationJournal: To be Published
Title: Molecular Basis for Epigenetic-sensitive Editing by Cas9
Authors: Roth MO / Shu Y / Zhao Y / Trasanidou D / Hoffman RD / Trasanidis N / Gelasco MK / Medina ML / Das D / Rai J / Wang B / Li H
History
DepositionFeb 22, 2024-
Header (metadata) releaseFeb 26, 2025-
Map releaseFeb 26, 2025-
UpdateFeb 26, 2025-
Current statusFeb 26, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43770.png

Downloads & links

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Map

FileDownload / File: emd_43770.map.gz / Format: CCP4 / Size: 103 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 300 pix.
= 246. Å		0.82 Å/pix.
x 300 pix.
= 246. Å		0.82 Å/pix.
x 300 pix.
= 246. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.82 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.28
Minimum - Maximum-2.9603522 - 4.827367
Average (Standard dev.)0.004212022 (±0.09618883)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 246.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_43770_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_43770_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 : CRISPR-Cas9

EntireName: CRISPR-Cas9
Components
  • Complex: CRISPR-Cas9
    • Protein or peptide: CRISPR-associated endonuclease, Csn1 family
    • RNA: RNA (92-MER)
    • DNA: DNA (25-MER)
    • DNA: DNA/RNA (5'-D(P*TP*AP*CP*A)-R(P*(5MC))-D(P*CP*AP*AP*GP*CP*T)-3')

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Supramolecule #1: CRISPR-Cas9

SupramoleculeName: CRISPR-Cas9 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Acidothermus cellulolyticus (bacteria)

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Macromolecule #1: CRISPR-associated endonuclease, Csn1 family

MacromoleculeName: CRISPR-associated endonuclease, Csn1 family / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Acidothermus cellulolyticus (bacteria)
Molecular weightTheoretical: 127.498094 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MGGSEVGTVP VTWRLGVDVG ERSIGLAAVS YEEDKPKEIL AAVSWIHDGG VGDERSGASR LALRGMARRA RRLRRFRRAR LRDLDMLLS ELGWTPLPDK NVSPVDAWLA RKRLAEEYVV DETERRRLLG YAVSHMARHR GWRNPWTTIK DLKNLPQPSD S WERTRESL ...String:
MGGSEVGTVP VTWRLGVDVG ERSIGLAAVS YEEDKPKEIL AAVSWIHDGG VGDERSGASR LALRGMARRA RRLRRFRRAR LRDLDMLLS ELGWTPLPDK NVSPVDAWLA RKRLAEEYVV DETERRRLLG YAVSHMARHR GWRNPWTTIK DLKNLPQPSD S WERTRESL EARYSVSLEP GTVGQWAGYL LQRAPGIRLN PTQQSAGRRA ELSNATAFET RLRQEDVLWE LRCIADVQGL PE DVVSNVI DAVFCQKRPS VPAERIGRDP LDPSQLRASR ACLEFQEYRI VAAVANLRIR DGSGSRPLSL EERNAVIEAL LAQ TERSLT WSDIALEILK LPNESDLTSV PEEDGPSSLA YSQFAPFDET SARIAEFIAK NRRKIPTFAQ WWQEQDRTSR SDLV AALAD NSIAGEEEQE LLVHLPDAEL EALEGLALPS GRVAYSRLTL SGLTRVMRDD GVDVHNARKT CFGVDDNWRP PLPAL HEAT GHPVVDRNLA ILRKFLSSAT MRWGPPQSIV VELARGASES RERQAEEEAA RRAHRKANDR IRAELRASGL SDPSPA DLV RARLLELYDC HCMYCGAPIS WENSELDHIV PRTDGGSNRH ENLAITCGAC NKEKGRRPFA SWAETSNRVQ LRDVIDR VQ KLKYSGNMYW TRDEFSRYKK SVVARLKRRT SDPEVIQSIE STGYAAVALR DRLLSYGEKN GVAQVAVFRG GVTAEARR W LDISIERLFS RVAIFAQSTS TKRLDRRHHA VDAVVLTTLT PGVAKTLADA RSRRVSAEFW RRPSDVNRHS TEEPQSPAY RQWKESCSGL GDLLISTAAR DSIAVAAPLR LRPTGALHEE TLRAFSEHTV GAAWKGAELR RIVEPEVYAA FLALTDPGGR FLKVSPSED VLPADENRHI VLSDRVLGPR DRVKLFPDDR GSIRVRGGAA YIASFHHARV FRWGSSHSPS FALLRVSLAD L AVAGLLRD GVDVFTAELP PWTPAWRYAS IALVKAVESG DAKQVGWLVP GDELDFGPEG VTTAAGDLSM FLKYFPERHW VV TGFEDDK RINLKPAFLS AEQAEVLRTE RSDRPDTLTE AGEILAQFFP RCWRATVAKV LCHPGLTVIR RTALGQPRWR RGH LPYSWR PWSADPWSGG TP

UniProtKB: CRISPR-associated endonuclease, Csn1 family

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Macromolecule #2: RNA (92-MER)

MacromoleculeName: RNA (92-MER) / type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: Acidothermus cellulolyticus (bacteria)
Molecular weightTheoretical: 34.230328 KDa
SequenceString:
GGUAGGAUGG CAAGAUCCUG GUAUGCUGGG GAGCCUGAAA AGGCUACCUA GCAAGACCCC UUCGUGGGGU CGCAUUCUUC ACCCCCUCG CAGCAGCGAG GGGGUUC

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Macromolecule #3: DNA (25-MER)

MacromoleculeName: DNA (25-MER) / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Acidothermus cellulolyticus (bacteria)
Molecular weightTheoretical: 7.704969 KDa
SequenceString:
(DA)(DG)(DC)(DT)(DT)(DG)(DG)(DT)(DG)(DT) (DA)(DT)(DA)(DC)(DC)(DA)(DG)(DG)(DA)(DT) (DC)(DT)(DT)(DG)(DC)

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Macromolecule #4: DNA/RNA (5'-D(P*TP*AP*CP*A)-R(P*(5MC))-D(P*CP*AP*AP*GP*CP*T)-3')

MacromoleculeName: DNA/RNA (5'-D(P*TP*AP*CP*A)-R(P*(5MC))-D(P*CP*AP*AP*GP*CP*T)-3')
type: dna / ID: 4 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Acidothermus cellulolyticus (bacteria)
Molecular weightTheoretical: 3.332214 KDa
SequenceString:
(DT)(DA)(DC)(DA)(5MC)(DC)(DA)(DA)(DG)(DC) (DT)

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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

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

8d2n

Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.88 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 275004
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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