Structural mechanism governing the directionality of bridge recombination

Area of Science:

• Molecular Biology
• Genetics
• Structural Biology

Background:

• IS110 family transposons, like IS621, utilize bridge recombinases and bridge RNA (bRNA) for DNA recombination.
• While DNA insertion mechanisms are understood, the process of IS621 element excision and circular DNA intermediate formation remains unclear.

Purpose of the Study:

• To elucidate the mechanism of IS621 element excision from host genomes.
• To determine the structural basis for the observed bias favoring DNA insertion over excision.

Main Methods:

• Cryo-electron microscopy to determine the structure of the IS621 recombinase-bRNA complex bound to DNA excision substrates.
• Biochemical assays to compare the efficiency of DNA insertion and excision reactions.

Main Results:

• Bridge RNA (bRNA) is expressed at low levels from IS621 loci in Escherichia coli.
• The IS621 recombinase-bRNA complex mediates DNA excision less efficiently than insertion.
• Cryo-EM structures reveal distinct linear DNA conformations in the excision complex compared to the bent U-shape in the insertion complex, forming an X-shaped structure.
• Handshake guides in bRNA modulate both insertion and excision efficiencies.

Conclusions:

• The study provides mechanistic insights into the complete IS110 transposition cycle, including the less efficient excision pathway.
• The structural differences explain the inherent bias towards DNA insertion over excision.
• Findings facilitate the design of programmable bridge-editing applications.