The mechanism of phage λ site-specific recombination: Site-specific breakage of DNA by Int topoisomerase

We demonstrate that the topoisomerase activity of bacteriophage λ Int protein introduces single-strand breaks into duplex DNA at specific sites. Strand breakage is accompanied by the covalent linkage of Int to DNA. The linkage connects a residue in Int to the 3′ phosphate of DNA at the site of breakage; the other breakage product has a 5′ OH terminus. Int is the first procaryotic topoisomerase shown to break DNA in this manner. We find that in att sites, Int breaks DNA within the 15 bp homologous core. These sites of Int topoisomerase action result from the interaction of Int with "junction-type" recognition sequences (CAACTTNNT), and Int topoisomerase acts between the 7th and 8th bases of this sequence. The sites of breakage within the cores of attP and attB coincide exactly with positions where breakage and reunion occur during Int-dependent recombination. These results indicate that Int topoisomerase executes strand exchange during recombination.