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. 2010 Dec 17;286(8):6402–6413. doi: 10.1074/jbc.M110.148148

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FIGURE 4. — TopA is inhibited by phenanthrene alkaloids. A, inhibition of TopA by boldine, compound 2 (secoboldine) and compound 5 (N-methyl-secoboldine). Supercoiled pBR322 (0.5 μg) was treated with 1 unit of purified TopA in 15-μl reactions containing the alkaloids at the concentrations (μm) indicated. Mw, molecular mass standard; S, covalently closed supercoiled pBR322 used as substrate. B, inhibition of TopA by compounds 16 (N-metilseconeolitsine), 17 (seconeolitsine), and 14 (neolitsine). The reactions were carried out as in A. C, the inhibition of TopA by seconeolitsine is enhanced by preincubation of the enzyme with the alkaloid. pBR322 was treated with 1 unit of purified TopA in the absence of the inhibitor (lane 0), or in the presence of 30 μm of compound 17 with (lane +) or without (lane −) preincubation during 10 min at 4 °C. D, effect of EtBr on TopA activity. The reactions were carried out as in A containing the compound at the indicated concentrations (μm).