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Yorodumi- EMDB-13439: Cryo-EM structure of E. coli TnsB in complex with right end fragm... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-13439 | |||||||||
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Title | Cryo-EM structure of E. coli TnsB in complex with right end fragment of Tn7 transposon | |||||||||
Map data | Primary map used for model generation; sharpened with B-factor of -70 and after Local Filtering | |||||||||
Sample |
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Function / homology | Function and homology information transposase activity / DNA transposition / DNA integration / chromosome / DNA binding Similarity search - Function | |||||||||
Biological species | Escherichia coli (E. coli) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.68 Å | |||||||||
Authors | Kaczmarska Z / Czarnocki-Cieciura M / Rawski M / Nowotny M | |||||||||
Funding support | Poland, European Union, 2 items
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Citation | Journal: Mol Cell / Year: 2022 Title: Structural basis of transposon end recognition explains central features of Tn7 transposition systems. Authors: Zuzanna Kaczmarska / Mariusz Czarnocki-Cieciura / Karolina M Górecka-Minakowska / Robert J Wingo / Justyna Jackiewicz / Weronika Zajko / Jarosław T Poznański / Michał Rawski / Timothy ...Authors: Zuzanna Kaczmarska / Mariusz Czarnocki-Cieciura / Karolina M Górecka-Minakowska / Robert J Wingo / Justyna Jackiewicz / Weronika Zajko / Jarosław T Poznański / Michał Rawski / Timothy Grant / Joseph E Peters / Marcin Nowotny / Abstract: Tn7 is a bacterial transposon with relatives containing element-encoded CRISPR-Cas systems mediating RNA-guided transposon insertion. Here, we present the 2.7 Å cryoelectron microscopy structure of ...Tn7 is a bacterial transposon with relatives containing element-encoded CRISPR-Cas systems mediating RNA-guided transposon insertion. Here, we present the 2.7 Å cryoelectron microscopy structure of prototypic Tn7 transposase TnsB interacting with the transposon end DNA. When TnsB interacts across repeating binding sites, it adopts a beads-on-a-string architecture, where the DNA-binding and catalytic domains are arranged in a tiled and intertwined fashion. The DNA-binding domains form few base-specific contacts leading to a binding preference that requires multiple weakly conserved sites at the appropriate spacing to achieve DNA sequence specificity. TnsB binding imparts differences in the global structure of the protein-bound DNA ends dictated by the spacing or overlap of binding sites explaining functional differences in the left and right ends of the element. We propose a model of the strand-transfer complex in which the terminal TnsB molecule is rearranged so that its catalytic domain is in a position conducive to transposition. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_13439.map.gz | 7.9 MB | EMDB map data format | |
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Header (meta data) | emd-13439-v30.xml emd-13439.xml | 23.4 KB 23.4 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_13439_fsc.xml | 17.8 KB | Display | FSC data file |
Images | emd_13439.png | 121.9 KB | ||
Masks | emd_13439_msk_1.map | 512 MB | Mask map | |
Others | emd_13439_additional_1.map.gz emd_13439_half_map_1.map.gz emd_13439_half_map_2.map.gz | 255.4 MB 475 MB 475 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-13439 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13439 | HTTPS FTP |
-Related structure data
Related structure data | 7pikMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_13439.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Primary map used for model generation; sharpened with B-factor of -70 and after Local Filtering | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.86 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_13439_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Raw map
File | emd_13439_additional_1.map | ||||||||||||
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Annotation | Raw map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map A
File | emd_13439_half_map_1.map | ||||||||||||
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Annotation | Half map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map B
File | emd_13439_half_map_2.map | ||||||||||||
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Annotation | Half map B | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : TnsB-DNA complex
Entire | Name: TnsB-DNA complex |
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Components |
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-Supramolecule #1: TnsB-DNA complex
Supramolecule | Name: TnsB-DNA complex / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: all Details: TnsB from the canonical E. coli Tn7 element in complex with the right transposon end fragment |
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Molecular weight | Theoretical: 43 KDa |
-Supramolecule #2: TnsB
Supramolecule | Name: TnsB / type: complex / Chimera: Yes / ID: 2 / Parent: 1 / Macromolecule list: #1 / Details: TnsB from the canonical E. coli Tn7 element |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Recombinant expression | Organism: Escherichia coli (E. coli) |
-Supramolecule #3: Tn7 transposon right end fragment
Supramolecule | Name: Tn7 transposon right end fragment / type: complex / Chimera: Yes / ID: 3 / Parent: 1 / Macromolecule list: #2 / Details: E. coli Tn7 transposon right end fragment |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Recombinant expression | Organism: synthetic construct (others) |
-Macromolecule #1: Transposon Tn7 transposition protein TnsB
Macromolecule | Name: Transposon Tn7 transposition protein TnsB / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 81.026695 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: SMWQINEVVL FDNDPYRILA IEDGQVVWMQ ISADKGVPQA RAELLLMQYL DEGRLVRTDD PYVHLDLEEP SVDSVSFQKR EEDYRKILP IINSKDRFDP KVRSELVEHV VQEHKVTKAT VYKLLRRYWQ RGQTPNALIP DYKNSGAPGE RRSATGTAKI G RAREYGKG ...String: SMWQINEVVL FDNDPYRILA IEDGQVVWMQ ISADKGVPQA RAELLLMQYL DEGRLVRTDD PYVHLDLEEP SVDSVSFQKR EEDYRKILP IINSKDRFDP KVRSELVEHV VQEHKVTKAT VYKLLRRYWQ RGQTPNALIP DYKNSGAPGE RRSATGTAKI G RAREYGKG EGTKVTPEIE RLFRLTIEKH LLNQKGTKTT VAYRRFVDLF AQYFPRIPQE DYPTLRQFRY FYDREYPKAQ RL KSRVKAG VYKKDVRPLS STATSQALGP GSRYEIDATI ADIYLVDHHD RQKIIGRPTL YIVIDVFSRM ITGFYIGFEN PSY VVAMQA FVNACSDKTA ICAQHDIEIS SSDWPCVGLP DVLLADRGEL MSHQVEALVS SFNVRVESAP PRRGDAKGIV ESTF RTLQA EFKSFAPGIV EGSRIKSHGE TDYRLDASLS VFEFTQIILR TILFRNNHLV MDKYDRDADF PTDLPSIPVQ LWQWG MQHR TGSLRAVEQE QLRVALLPRR KVSISSFGVN LWGLYYSGSE ILREGWLQRS TDIARPQHLE AAYDPVLVDT IYLFPQ VGS RVFWRCNLTE RSRQFKGLSF WEVWDIQAQE KHNKANAKQD ELTKRRELEA FIQQTIQKAN KLTPSTTEPK STRIKQI KT NKKEAVTSER KKRAEHLKPS SSGDEAKVIP FNAVEADDQE DYSLPTYVPE LFQDPPEKDE S |
-Macromolecule #2: Right end fragment of Tn7 transposon
Macromolecule | Name: Right end fragment of Tn7 transposon / type: dna / ID: 2 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 21.471771 KDa |
Sequence | String: (DC)(DT)(DA)(DG)(DT)(DT)(DT)(DA)(DA)(DG) (DA)(DC)(DT)(DT)(DT)(DA)(DT)(DT)(DG)(DT) (DC)(DA)(DT)(DA)(DG)(DT)(DT)(DT)(DA) (DG)(DA)(DT)(DC)(DT)(DA)(DT)(DT)(DT)(DT) (DG) (DT)(DT)(DC)(DA)(DG)(DT) ...String: (DC)(DT)(DA)(DG)(DT)(DT)(DT)(DA)(DA)(DG) (DA)(DC)(DT)(DT)(DT)(DA)(DT)(DT)(DG)(DT) (DC)(DA)(DT)(DA)(DG)(DT)(DT)(DT)(DA) (DG)(DA)(DT)(DC)(DT)(DA)(DT)(DT)(DT)(DT) (DG) (DT)(DT)(DC)(DA)(DG)(DT)(DT)(DT) (DA)(DA)(DG)(DA)(DC)(DT)(DT)(DT)(DA)(DT) (DT)(DG) (DT)(DC)(DC)(DG)(DC)(DC)(DC) (DA)(DC)(DA) |
-Macromolecule #3: Right end fragment of Tn7 transposon
Macromolecule | Name: Right end fragment of Tn7 transposon / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 21.678018 KDa |
Sequence | String: (DT)(DG)(DT)(DG)(DG)(DG)(DC)(DG)(DG)(DA) (DC)(DA)(DA)(DT)(DA)(DA)(DA)(DG)(DT)(DC) (DT)(DT)(DA)(DA)(DA)(DC)(DT)(DG)(DA) (DA)(DC)(DA)(DA)(DA)(DA)(DT)(DA)(DG)(DA) (DT) (DC)(DT)(DA)(DA)(DA)(DC) ...String: (DT)(DG)(DT)(DG)(DG)(DG)(DC)(DG)(DG)(DA) (DC)(DA)(DA)(DT)(DA)(DA)(DA)(DG)(DT)(DC) (DT)(DT)(DA)(DA)(DA)(DC)(DT)(DG)(DA) (DA)(DC)(DA)(DA)(DA)(DA)(DT)(DA)(DG)(DA) (DT) (DC)(DT)(DA)(DA)(DA)(DC)(DT)(DA) (DT)(DG)(DA)(DC)(DA)(DA)(DT)(DA)(DA)(DA) (DG)(DT) (DC)(DT)(DT)(DA)(DA)(DA)(DC) (DT)(DA)(DG) |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | filament |
-Sample preparation
Concentration | 1 mg/mL | |||||||||
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Buffer | pH: 7.5 Component:
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Grid | Model: PELCO Ultrathin Carbon with Lacey Carbon / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE | |||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: 4 s blot time, -5 blot force.. | |||||||||
Details | Sample fixed with 0.05% glutaraldehyde and concentrated prior to vitrification; exact concentration cannot be estimated accurately. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 105000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 40.0 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: AB INITIO MODEL |
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Output model | PDB-7pik: |