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- EMDB-34958: Cryo-EM structure of the CnCas12f1-sgRNA-DNA complex -

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ID or keywords:

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Basic information

Entry
Database: EMDB / ID: EMD-34958
TitleCryo-EM structure of the CnCas12f1-sgRNA-DNA complex
Map data
Sample
  • Complex: Ternary complex of CnCas12f1-sgRNA-DNA
    • Protein or peptide: Transposase
    • RNA: sgRNASubgenomic mRNA
    • DNA: DNA (28-MER)
    • DNA: DNA (5'-D(*TP*AP*AP*CP*CP*TP*AP*AP*TP*AP*GP*AP*TP*GP*TP*GP*AP*A)-3')
KeywordsComplex / CRISPR-Cas / RNA BINDING PROTEIN/RNA/DNA / RNA BINDING PROTEIN-RNA-DNA complex
Function / homologyTransposase IS605, OrfB, C-terminal / Putative transposase DNA-binding domain / transposition / DNA recombination / DNA binding / Transposase
Function and homology information
Biological speciesClostridium novyi (bacteria) / synthetic construct (others)
Methodsingle particle reconstruction / Resolution: 3.73 Å
AuthorsLi F / Ji Q
Funding support China, 3 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)21922705 China
National Natural Science Foundation of China (NSFC)2207783 China
National Natural Science Foundation of China (NSFC)91753127 China
CitationJournal: To Be Published
Title: Structure and engineering of miniature Clostridium novyi CRISPR-Cas12f1 with rare C-rich PAM specificity
Authors: Li F / Ji Q
History
DepositionDec 14, 2022-
Header (metadata) releaseSep 6, 2023-
Map releaseSep 6, 2023-
UpdateSep 6, 2023-
Current statusSep 6, 2023Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_34958.png

Downloads & links

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Map

FileDownload / File: emd_34958.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.82 Å
Density
Contour LevelBy AUTHOR: 0.1
Minimum - Maximum-1.6911359 - 2.6486044
Average (Standard dev.)-0.0004276748 (±0.037480395)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 314.88 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_34958_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_34958_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 : Ternary complex of CnCas12f1-sgRNA-DNA

EntireName: Ternary complex of CnCas12f1-sgRNA-DNA
Components
  • Complex: Ternary complex of CnCas12f1-sgRNA-DNA
    • Protein or peptide: Transposase
    • RNA: sgRNASubgenomic mRNA
    • DNA: DNA (28-MER)
    • DNA: DNA (5'-D(*TP*AP*AP*CP*CP*TP*AP*AP*TP*AP*GP*AP*TP*GP*TP*GP*AP*A)-3')

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Supramolecule #1: Ternary complex of CnCas12f1-sgRNA-DNA

SupramoleculeName: Ternary complex of CnCas12f1-sgRNA-DNA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Clostridium novyi (bacteria)

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Macromolecule #1: Transposase

MacromoleculeName: Transposase / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Clostridium novyi (bacteria)
Molecular weightTheoretical: 58.60968 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MITVRKIKLT IMGDKDTRNS QYKWIRDEQY NQYRALNMGM TYLAVNDILY MNESGLEIRT IKDLKDCEKD IDKNKKEIEK LTARLEKEQ NKKNSSSEKL DEIKYKISLV ENKIEDYKLK IVELNKILEE TQKERMDIQK EFKEKYVDDL YQVLDKIPFK H LDNKSLVT ...String:
MITVRKIKLT IMGDKDTRNS QYKWIRDEQY NQYRALNMGM TYLAVNDILY MNESGLEIRT IKDLKDCEKD IDKNKKEIEK LTARLEKEQ NKKNSSSEKL DEIKYKISLV ENKIEDYKLK IVELNKILEE TQKERMDIQK EFKEKYVDDL YQVLDKIPFK H LDNKSLVT QRIKADIKSD KSNGLLKGER SIRNYKRNFP LMTRGRDLKF KYDDNDDIEI KWMEGIKFKV ILGNRIKNSL EL RHTLHKV IEGKYKICDS SLQFDKNNNL ILNLTLDIPI DIVNKKVSGR VVGVDLGLKI PAYCALNDVE YIKKSIGRID DFL KVRTQM QSRRRRLQIA IQSAKGGKGR VNKLQALERF AEKEKNFAKT YNHFLSSNIV KFAVSNQAEQ INMELLSLKE TQNK SILRN WSYYQLQTMI EYKAQREGIK VKYIDPYHTS QTCSKCGNYE EGQRESQADF ICKKCGYKVN ADYNAARNIA MSNKY ITKK EESKYYKIKE SMV

UniProtKB: Transposase

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Macromolecule #2: sgRNA

MacromoleculeName: sgRNA / type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 75.498406 KDa
SequenceString: AACAUAGUUA AACUAAUAAA AACAGGGCGA UUUAACGUCC UAAGGCUGAG AGAAGUUUUU UCUACUCGGC AAGGGUUAAU CUCGAUUGU UGUGUUACCG AUCGAGCGUU UCACAAAAUG CGAGAGAAAU CUCGCAUUUU UAAUUUUGCA GUAAGGCUAG U UUUUAUAU ...String:
AACAUAGUUA AACUAAUAAA AACAGGGCGA UUUAACGUCC UAAGGCUGAG AGAAGUUUUU UCUACUCGGC AAGGGUUAAU CUCGAUUGU UGUGUUACCG AUCGAGCGUU UCACAAAAUG CGAGAGAAAU CUCGCAUUUU UAAUUUUGCA GUAAGGCUAG U UUUUAUAU AAAUAUGCUA UAACUAGGGU UUUAGUUUAA CUAUGUGAAA UGUAAAUAAU AGAUGUGAAG UCGCUUU

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Macromolecule #3: DNA (28-MER)

MacromoleculeName: DNA (28-MER) / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 8.612591 KDa
SequenceString:
(DG)(DA)(DA)(DA)(DA)(DG)(DC)(DG)(DA)(DC) (DT)(DT)(DC)(DA)(DC)(DA)(DT)(DC)(DT)(DA) (DT)(DT)(DA)(DG)(DG)(DT)(DT)(DA)

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Macromolecule #4: DNA (5'-D(*TP*AP*AP*CP*CP*TP*AP*AP*TP*AP*GP*AP*TP*GP*TP*GP*AP*A)-3')

MacromoleculeName: DNA (5'-D(*TP*AP*AP*CP*CP*TP*AP*AP*TP*AP*GP*AP*TP*GP*TP*GP*AP*A)-3')
type: dna / ID: 4 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 5.547643 KDa
SequenceString:
(DT)(DA)(DA)(DC)(DC)(DT)(DA)(DA)(DT)(DA) (DG)(DA)(DT)(DG)(DT)(DG)(DA)(DA)

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Experimental details

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Structure determination

Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1.8 µm / Nominal defocus min: 1.2 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 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: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.73 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 100719
FSC plot (resolution estimation)

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