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Yorodumi- EMDB-37829: Cryo-EM structure of the IS621 recombinase in complex with bridge... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-37829 | |||||||||
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Title | Cryo-EM structure of the IS621 recombinase in complex with bridge RNA, donor DNA, and target DNA in the post-strand exchange state (Holliday junction intermediate) | |||||||||
Map data | ||||||||||
Sample |
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Keywords | Holliday junction / RNA dependent recombinase / RECOMBINATION-RNA-DNA COMPLEX | |||||||||
Function / homology | Transposase, IS111A/IS1328/IS1533, N-terminal / Transposase, IS116/IS110/IS902 / : / Transposase / Transposase IS116/IS110/IS902 family / transposase activity / DNA transposition / DNA binding / IS621 transposase Function and homology information | |||||||||
Biological species | Escherichia coli (E. coli) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.9 Å | |||||||||
Authors | Hiraizumi M / Yamashita K / Nishimasu H | |||||||||
Funding support | Japan, 2 items
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Citation | Journal: Nature / Year: 2024 Title: Structural mechanism of bridge RNA-guided recombination. Authors: Masahiro Hiraizumi / Nicholas T Perry / Matthew G Durrant / Teppei Soma / Naoto Nagahata / Sae Okazaki / Januka S Athukoralage / Yukari Isayama / James J Pai / April Pawluk / Silvana ...Authors: Masahiro Hiraizumi / Nicholas T Perry / Matthew G Durrant / Teppei Soma / Naoto Nagahata / Sae Okazaki / Januka S Athukoralage / Yukari Isayama / James J Pai / April Pawluk / Silvana Konermann / Keitaro Yamashita / Patrick D Hsu / Hiroshi Nishimasu / Abstract: Insertion sequence (IS) elements are the simplest autonomous transposable elements found in prokaryotic genomes. We recently discovered that IS110 family elements encode a recombinase and a non- ...Insertion sequence (IS) elements are the simplest autonomous transposable elements found in prokaryotic genomes. We recently discovered that IS110 family elements encode a recombinase and a non-coding bridge RNA (bRNA) that confers modular specificity for target DNA and donor DNA through two programmable loops. Here we report the cryo-electron microscopy structures of the IS110 recombinase in complex with its bRNA, target DNA and donor DNA in three different stages of the recombination reaction cycle. The IS110 synaptic complex comprises two recombinase dimers, one of which houses the target-binding loop of the bRNA and binds to target DNA, whereas the other coordinates the bRNA donor-binding loop and donor DNA. We uncovered the formation of a composite RuvC-Tnp active site that spans the two dimers, positioning the catalytic serine residues adjacent to the recombination sites in both target and donor DNA. A comparison of the three structures revealed that (1) the top strands of target and donor DNA are cleaved at the composite active sites to form covalent 5'-phosphoserine intermediates, (2) the cleaved DNA strands are exchanged and religated to create a Holliday junction intermediate, and (3) this intermediate is subsequently resolved by cleavage of the bottom strands. Overall, this study reveals the mechanism by which a bispecific RNA confers target and donor DNA specificity to IS110 recombinases for programmable DNA recombination. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_37829.map.gz | 26.5 MB | EMDB map data format | |
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Header (meta data) | emd-37829-v30.xml emd-37829.xml | 19.7 KB 19.7 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_37829_fsc.xml | 7.9 KB | Display | FSC data file |
Images | emd_37829.png | 131.2 KB | ||
Masks | emd_37829_msk_1.map | 52.7 MB | Mask map | |
Filedesc metadata | emd-37829.cif.gz | 6.4 KB | ||
Others | emd_37829_half_map_1.map.gz emd_37829_half_map_2.map.gz | 49 MB 49 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-37829 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-37829 | HTTPS FTP |
-Validation report
Summary document | emd_37829_validation.pdf.gz | 861.7 KB | Display | EMDB validaton report |
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Full document | emd_37829_full_validation.pdf.gz | 861.3 KB | Display | |
Data in XML | emd_37829_validation.xml.gz | 15.8 KB | Display | |
Data in CIF | emd_37829_validation.cif.gz | 20.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-37829 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-37829 | HTTPS FTP |
-Related structure data
Related structure data | 8wt8MC 8wt6C 8wt7C 8wt9C 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_37829.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.10667 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_37829_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_37829_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_37829_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : The IS621 recombinase in complex with bridge RNA, donor DNA, and ...
Entire | Name: The IS621 recombinase in complex with bridge RNA, donor DNA, and target DNA in the post-strand exchange state (Holliday junction intermediate) |
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Components |
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-Supramolecule #1: The IS621 recombinase in complex with bridge RNA, donor DNA, and ...
Supramolecule | Name: The IS621 recombinase in complex with bridge RNA, donor DNA, and target DNA in the post-strand exchange state (Holliday junction intermediate) type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#6 |
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Source (natural) | Organism: Escherichia coli (E. coli) |
-Macromolecule #1: IS621 transposase
Macromolecule | Name: IS621 transposase / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 36.735355 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: GPMEHELHYI GIDTAKEKLD VDVLRPDGRH RTKKFANTTK GHDELVSWLK GHKIDHAHIC IEATGTYMEP VAECLYDAGY IVSVINPAL GKAFAQSEGL RNKTDTVDAR MLAEFCRQKR PAAWEAPHPL ERALRALVVR HQALTDMHTQ ELNRTETARE V QRPSIDAH ...String: GPMEHELHYI GIDTAKEKLD VDVLRPDGRH RTKKFANTTK GHDELVSWLK GHKIDHAHIC IEATGTYMEP VAECLYDAGY IVSVINPAL GKAFAQSEGL RNKTDTVDAR MLAEFCRQKR PAAWEAPHPL ERALRALVVR HQALTDMHTQ ELNRTETARE V QRPSIDAH LLWLEAELKR LEKQIKDLTD DDPDMKHRRK LLESIPGIGE KTSAVLLAYI GLKDRFAHAR QFAAFAGLTP RR YESGSSV RGASRMSKAG HVSLRRALYM PAMVATSKTE WGRAFRDRLA ANGKKGKVIL GAMMRKLAQV AYGVLKSGVP FDA SRHNPV AA UniProtKB: IS621 transposase |
-Macromolecule #2: bridge RNA
Macromolecule | Name: bridge RNA / type: rna / ID: 2 / Number of copies: 2 |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 57.668918 KDa |
Sequence | String: GGGAGUGCAG AGAAAAUCGG CCAGUUUUCU CUGCCUGCAG UCCGCAUGCC GUAUCGGGCC UUGGGUUCUA ACCUGUUCUG UAGAUUUAU GCAGCGGACU GCCUUUCUCC CAAAGUGAUA AACCGGACAG UAUCAUGGAC CGGUUUUCCC GGUAAUCCGU A UUUACAAG GCUGGUUUCA CU |
-Macromolecule #3: target DNA-donor DNA
Macromolecule | Name: target DNA-donor DNA / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 15.003622 KDa |
Sequence | String: (DG)(DC)(DC)(DG)(DG)(DG)(DT)(DA)(DA)(DT) (DA)(DC)(DC)(DA)(DC)(DC)(DA)(DA)(DG)(DC) (DC)(DG)(DC)(DC)(DT)(DT)(DG)(DT)(DA) (DT)(DT)(DA)(DT)(DC)(DC)(DC)(DT)(DC)(DC) (DA) (DG)(DT)(DG)(DC)(DA)(DG)(DA)(DG) (DA) |
-Macromolecule #4: target DNA
Macromolecule | Name: target DNA / type: dna / ID: 4 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 11.703487 KDa |
Sequence | String: (DC)(DG)(DA)(DG)(DC)(DT)(DC)(DA)(DT)(DC) (DT)(DG)(DT)(DA)(DG)(DG)(DC)(DC)(DC)(DG) (DA)(DT)(DG)(DG)(DT)(DG)(DG)(DT)(DA) (DT)(DT)(DA)(DC)(DC)(DC)(DG)(DG)(DC) |
-Macromolecule #5: donor DNA-target DNA
Macromolecule | Name: donor DNA-target DNA / type: dna / ID: 5 Details: The linkage between 20I and 21I has been cleaved by IS621. Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 10.163561 KDa |
Sequence | String: (DT)(DG)(DC)(DA)(DG)(DG)(DC)(DC)(DA)(DT) (DA)(DA)(DG)(DT)(DC)(DA)(DA)(DT)(DC)(DT) (DA)(DC)(DA)(DG)(DA)(DT)(DG)(DA)(DG) (DC)(DT)(DC)(DG) |
-Macromolecule #6: donor DNA
Macromolecule | Name: donor DNA / type: dna / ID: 6 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 13.587737 KDa |
Sequence | String: (DT)(DC)(DT)(DC)(DT)(DG)(DC)(DA)(DC)(DT) (DG)(DG)(DA)(DG)(DG)(DG)(DA)(DT)(DA)(DA) (DT)(DA)(DC)(DA)(DA)(DG)(DA)(DT)(DA) (DC)(DT)(DG)(DT)(DT)(DA)(DT)(DG)(DG)(DC) (DC) (DT)(DG)(DC)(DA) |
-Macromolecule #7: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 7 / Number of copies: 1 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 62.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.6 µm / Nominal defocus min: 0.8 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |