Structural mechanism governing the directionality of bridge recombination.
The study reveals that the IS621 recombinase-bRNA complex is less efficient at mediating excision compared to insertion, with structural differences influencing this bias.
Where it sits
this study against the rest of the humanin corpusSummary and findings
The study investigates the mechanism of bridge recombination in IS110 family transposons, specifically IS621, focusing on the role of bridge RNA (bRNA) in mediating excision and insertion reactions. The authors report that the IS621 recombinase-bRNA complex mediates excision less efficiently than insertion. Structural insights are provided through cryo-electron microscopy of the excision complex.
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<sup>1,2</sup>. 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<sup>2</sup>, 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<sup>2</sup>, 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.
Background
Not reported in abstract.
Methods
Not reported in abstract.
Results
Not reported in abstract.
Interpretation
Not reported in abstract.
Limitations
Not reported in abstract.