Skip to main content
. 2004 Oct 4;32(17):5163–5173. doi: 10.1093/nar/gkh847

Figure 3.

Figure 3

Triplex formation 1 bp 3′ from the Top1 cleavage site interferes with the Top1-mediated religation. (A) The 27mer oligonucleotide labeled on the 3′ end of the scissile strand (shown in Figure 1 and 2A) was reacted with Top1 in the absence of drug (+Top1) or in the presence of 1 μM CPT at 25°C for 30 min. The reactions were reversed in 0.35 M NaCl at 25°C for the indicated times (min). Time 0 refers to the samples taken immediately before the NaCl addition. Similar reactions were carried out using the 27mer triplex oligonucleotide (+TFO) in the absence (DNA) or in the presence of Top1 at 25°C for 30 min. The reactions were stopped before (0) and after reversal at 25°C for the indicated times (min). (B and C) Reactions as in (A) were carried out with salt reversals at 0°C (B), 25°C [(hollow squares and circles; (C)] or 37°C [filled squares and circles; (C)]. The percentage of the Top1-mediated cleavage products remaining after salt reversal for CPT (squares) and TFO (circles) were quantified and represented in semilog plots. Values are normalized to the drug-induced cleavage product remaining at time 0 taken as 100%. Computation of the reversal kinetic constants is summarized in Table 1.