Fig. 4. Decatenation of pseudo-replicative catenanes. (A) Schematic of catenanes with parallel dif sites formed by Cre recombination at directly repeated loxP sites, and their subsequent decatenation by FtsK50C–XerCD-mediated recombination at dif. The initial plasmid was constructed in a similar way to that described in Figure 1A, except that the dif sites were in inverted repeat with respect to each other. Therefore, the catenanes generated by Cre–loxP recombination have parallel dif sites and mimic replicative catenanes. (B) FtsK50C–XerCD recombination on catenanes containing parallel dif sites. The abbreviations are as in Figure 1A. (C) Comparison of decatenation by FtsK50C–XerCD and TopoIV at 20°C. Note that only a single FtsK50C–XerCD-dif decatenating unit can be active on each molecule, whereas multiple TopoIV molecules could be active.