Figure 3. The effects of CFS-1686 or CPT on Top1-mediated DNA relaxation and cleavage.

(A) CFS-1686 demonstrated 4-fold higher inhibition of Top1-mediated DNA relaxation than CPT. Recombinant Top1 enzymes were incubated with plasmid pUC19 DNA with/without CFS-1686 or CPT. Agarose electrophoresis was run to resolve the supercoiling or relaxed form of DNA. (B) CFS-1686 could not induce Top1-DNAcc, but it reduced CPT-induced DNA cleavage of Top1. The recombinant Top1 enzymes were incubated with γP³²-labelled pUC19 DNA with/without CFS-1686 or CPT. Then natural PAGE was run to resolve the intact and cleavage products. Experiments were repeated twice and representative results are shown. Arrows “a” and “b” indicate the compacted form of DNA and smear cleavage DNA, respectively.