Abstract
Quinolone antimicrobial drugs target both DNA gyrase and topoisomerase IV (Topo IV) and convert these essential enzymes into cellular poisons. Topoisomerase poisoning results in the inhibition of DNA replication and the generation of double-strand breaks. Double-strand breaks are repaired by homologous recombination. Here, we have investigated the interaction between the RuvAB branch migration complex and the Topo IV.quinolone.DNA ternary complex. A strand-displacement assay is employed to assess the helicase activity of the RuvAB complex in vitro. RuvAB-catalyzed strand displacement requires both RuvA and RuvB proteins, and it is stimulated by a 3'-non-hybridized tail. Interestingly, Topo IV.quinolone.DNA ternary complexes do not inhibit the translocation of the RuvAB complex. In fact, Topo IV.quinolone.DNA ternary complexes are reversed and displaced from the DNA upon their collisions with the RuvAB complex. These results suggest that the RuvAB branch migration complex can actively remove quinolone-induced covalent topoisomerase.DNA complexes from DNA and complete the homologous recombination process in vivo.
Highlights
The generation of genetic diversity is a result of homologous recombination but it is not the main function of recombination [1,2,3]
Topo IV1⁄7quinolone1⁄7DNA ternary complexes are reversed and displaced from the DNA upon their collisions with the RuvAB complex. These results suggest that the RuvAB branch migration complex can actively remove quinolone-induced covalent topoisomerase1⁄7DNA complexes from DNA and complete the homologous recombination process in vivo
We assumed that the topoisomerase IV (Topo IV)-catalyzed cleavage in the presence of various concentrations of norfloxacin shown in Fig. 2 correlated with the extent of Topo IV1⁄7norfloxacin1⁄7DNA ternary complexes formed on the partial duplex DNA
Summary
The generation of genetic diversity is a result of homologous recombination but it is not the main function of recombination [1,2,3]. These results suggest that the RuvAB branch migration complex can actively remove quinolone-induced covalent topoisomerase1⁄7DNA complexes from DNA and complete the homologous recombination process in vivo. The homologous recombination process, including the RuvABcatalyzed branch migration, must proceed in the presence of topoisomerase1⁄7quinolone1⁄7DNA ternary complexes to repair DSBs generated by the quinolone treatment and/or restart the replication fork stalled by a topoisomerase1⁄7quinolone1⁄7DNA ternary complex.
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