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- EMDB-65979: Cryo-EM structure of the IS621 recombinase in complex with bridge... -

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Basic information

Entry
Database: EMDB / ID: EMD-65979
TitleCryo-EM structure of the IS621 recombinase in complex with bridge RNA, left-half DNA, and right-half DNA in the post-strand exchange state
Map data
Sample
  • Complex: Cryo-EM structure of the IS621 recombinase in complex with bridge RNA, left-half DNA, and right-half DNA in the post-strand exchange state
    • Protein or peptide: IS621 transposase
    • RNA: bridge RNA
    • DNA: LH/RH, top strand
    • DNA: LH, bottom strand
    • DNA: RH/LH, top strand
    • DNA: RH, bottom strand
  • Ligand: MAGNESIUM ION
  • Ligand: water
KeywordsHolliday junction / RNA dependent recombinase / RECOMBINATION-DNA-RNA complex / DNA BINDING PROTEIN
Function / homologyTransposase, IS111A/IS1328/IS1533, N-terminal / : / Transposase / Transposase, IS116/IS110/IS902 / Transposase IS116/IS110/IS902 family / transposase activity / DNA transposition / DNA binding / IS621 transposase
Function and homology information
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.6 Å
AuthorsHiraizumi M / Tsujimoto E / Shiojiri N / Nagahata N / Yamashita K / Nishimasu H
Funding support Japan, 2 items
OrganizationGrant numberCountry
Japan Science and TechnologyJPMJCR23B6 Japan
Other privateInamori Foundation
CitationJournal: Nature / Year: 2026
Title: Structural mechanism governing the directionality of bridge recombination.
Authors: Masahiro Hiraizumi / Januka S Athukoralage / Nicholas T Perry / Eisuke Tsujimoto / Nami Shiojiri / Naoto Nagahata / Lauren Lee / Gwanggyu Sun / Matthew G Durrant / Sita S Chandrasekaran / ...Authors: Masahiro Hiraizumi / Januka S Athukoralage / Nicholas T Perry / Eisuke Tsujimoto / Nami Shiojiri / Naoto Nagahata / Lauren Lee / Gwanggyu Sun / Matthew G Durrant / Sita S Chandrasekaran / Silvana Konermann / Keitaro Yamashita / Patrick D Hsu / Hiroshi Nishimasu /
Abstract: Bridge recombinases from the IS110 family of transposons, such as IS621, associate with a bridge RNA (bRNA) to mediate programmable recombination between donor DNA and target DNA. Although insertion ...Bridge recombinases from the IS110 family of transposons, such as IS621, associate with a bridge RNA (bRNA) to mediate programmable recombination between donor DNA and target DNA. Although insertion is mediated by the recombinase-bRNA complex, it remains unknown how IS621 elements are excised from host genomes to form the circular DNA intermediates required for transposition. Here we show that bRNA is weakly expressed from IS621 loci in the Escherichia coli genome and that the IS621 recombinase-bRNA complex mediates excision less efficiently than insertion. Furthermore, we present the cryo-electron microscopy structures of the IS621 recombinase-bRNA complex bound to excision DNA substrates, providing mechanistic insights into the excision reaction. Similar to the previously reported donor- and target-bound insertion complex, the excision complex comprises two recombinase dimers, each accommodating the target- and donor-binding loops of the bRNA. However, DNA recognition differs notably between the two complexes. Although the donor and target DNAs form a bent U-shape during insertion, the excision substrates adopt linear conformations and bind across both bRNA loops, forming an X-shaped structure. This geometry reduces the efficiency of top-strand exchange and contributes to the naturally observed bias favouring insertion over excision. Despite these differences, the efficiencies of both reactions are similarly modulated by base pairing between specific dinucleotides in the bRNA, termed handshake guides, and the top strands of the DNA substrates. Overall, this study provides mechanistic insights into the complete IS110 transposition cycle and facilitates the optimal design of programmable bridge-editing applications.
History
DepositionAug 27, 2025-
Header (metadata) releaseAug 12, 2026-
Map releaseAug 12, 2026-
UpdateAug 26, 2026-
Current statusAug 26, 2026Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_65979.png

Downloads & links

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Map

FileDownload / File: emd_65979.map.gz / Format: CCP4 / Size: 36.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 212 pix.
= 175.96 Å		0.83 Å/pix.
x 212 pix.
= 175.96 Å		0.83 Å/pix.
x 212 pix.
= 175.96 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.02
Minimum - Maximum-0.07221764 - 0.24651182
Average (Standard dev.)0.00040890195 (±0.012199536)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin545454
Dimensions212212212
Spacing212212212
CellA=B=C: 175.95999 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_65979_msk_1.map
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AxesZYX

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Half map: #2

Fileemd_65979_half_map_1.map
Projections & Slices
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Half map: #1

Fileemd_65979_half_map_2.map
Projections & Slices
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Sample components

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Entire : Cryo-EM structure of the IS621 recombinase in complex with bridge...

EntireName: Cryo-EM structure of the IS621 recombinase in complex with bridge RNA, left-half DNA, and right-half DNA in the post-strand exchange state
Components
  • Complex: Cryo-EM structure of the IS621 recombinase in complex with bridge RNA, left-half DNA, and right-half DNA in the post-strand exchange state
    • Protein or peptide: IS621 transposase
    • RNA: bridge RNA
    • DNA: LH/RH, top strand
    • DNA: LH, bottom strand
    • DNA: RH/LH, top strand
    • DNA: RH, bottom strand
  • Ligand: MAGNESIUM ION
  • Ligand: water

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Supramolecule #1: Cryo-EM structure of the IS621 recombinase in complex with bridge...

SupramoleculeName: Cryo-EM structure of the IS621 recombinase in complex with bridge RNA, left-half DNA, and right-half DNA in the post-strand exchange state
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#6
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: IS621 transposase

MacromoleculeName: IS621 transposase / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 36.735355 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: 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

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Macromolecule #2: bridge RNA

MacromoleculeName: bridge RNA / type: rna / ID: 2 / Details: cell-free synthesis / Number of copies: 2
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 57.668918 KDa
SequenceString:
GGGAGUGCAG AGAAAAUCGG CCAGUUUUCU CUGCCUGCAG UCCGCAUGCC GUAUCGGGCC UUGGGUUCUA ACCUGUUCUG UAGGCUUAU GCAGCGGACU GCCUUUCUCC CAAAGUGAUA AACCGGACAG UAUCAUGGAC CGGUUUUCCC GGUAAUCCGU A UUUACAAG AUUGGUUUCA CU

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Macromolecule #3: LH/RH, top strand

MacromoleculeName: LH/RH, top strand / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 15.03264 KDa
SequenceString:
(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)(DA)(DC)(DA)(DG) (DA)(DT)(DG)(DA)(DG)(DC)(DT)(DC)(DG)(DC) (DC) (DC)(DC)(DA)(DA)(DC)(DG)(DA)(DG) (DG)

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Macromolecule #4: LH, bottom strand

MacromoleculeName: LH, bottom strand / type: dna / ID: 4 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 11.809589 KDa
SequenceString:
(DG)(DA)(DG)(DG)(DG)(DA)(DT)(DA)(DA)(DT) (DA)(DC)(DA)(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)

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Macromolecule #5: RH/LH, top strand

MacromoleculeName: RH/LH, top strand / type: dna / ID: 5 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 10.055483 KDa
SequenceString:
(DT)(DG)(DC)(DA)(DG)(DG)(DC)(DC)(DA)(DT) (DA)(DA)(DG)(DT)(DC)(DA)(DA)(DT)(DC)(DT) (DT)(DG)(DT)(DA)(DT)(DT)(DA)(DT)(DC) (DC)(DC)(DT)(DC)

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Macromolecule #6: RH, bottom strand

MacromoleculeName: RH, bottom strand / type: dna / ID: 6 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 13.562671 KDa
SequenceString:
(DC)(DC)(DT)(DC)(DG)(DT)(DT)(DG)(DG)(DG) (DG)(DC)(DG)(DA)(DG)(DC)(DT)(DC)(DA)(DT) (DC)(DT)(DG)(DT)(DA)(DG)(DA)(DT)(DA) (DC)(DT)(DG)(DT)(DT)(DA)(DT)(DG)(DG)(DC) (DC) (DT)(DG)(DC)(DA)

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Macromolecule #7: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 7 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #8: water

MacromoleculeName: water / type: ligand / ID: 8 / Number of copies: 4 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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 KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: cryoSPARC / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 515470
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
FSC plot (resolution estimation)

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