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. 2014 Aug 12;53(34):5558–5567. doi: 10.1021/bi500682e

Figure 6.

Figure 6

Competition between ciprofloxacin and 8-methyl-3′-(AM)P-dione and the effects of ciprofloxacin on DNA relaxation. The ability of 0–200 μM ciprofloxacin to compete with 10 μM 8-methyl-3′-(AM)P-dione for ParCS80L (S80L, blue), ParCS80I (S80I, red), ParCS80F (S80F, green), and ParCE84K (E84K, yellow) E. coli topoisomerase IV was determined using DNA cleavage assays (left). Both drugs were added to reaction mixtures simultaneously. For ParCS80L, the level of cleavage seen in the presence of ciprofloxacin alone was used as a baseline and subtracted from the total observed in the presence of both drugs. The level of DNA cleavage observed in the presence of only the quinazolinedione was set to 1.0 to facilitate direct comparisons. The effects of ciprofloxacin on the relaxation activities of wild-type (WT, black), ParCS80L (S80L, blue), ParCS80I (S80I, red), ParCS80F (S80F, green), and ParCE84K (E84K, yellow) topoisomerase IV are shown at the right. Wild-type, ParCS80L, and ParCS80F relaxation reactions were stopped at 10 min, ParCS80I reactions at 20 min, and ParCE84K reactions at 30 min to account for the different baseline DNA relaxation rates of the different enzymes. The amount of relaxed DNA observed in the absence of drug (“0” point) was set to 100% to facilitate direct comparisons. Error bars represent the standard deviation of three or more independent experiments.