FIG 4 .
QStatin affects the interaction between SmcR and target promoter DNAs. (a) For EMSA, a 200-bp PvvpE DNA fragment was radioactively labeled and then used as a DNA probe. Purified SmcR protein was added to the probe (15 ng) along with either QStatin (100 μM) or DMSO (2%). As a control, a random molecule (100 μm) from the library that showed no SmcR-inhibiting activity during initial screening was added instead of QStatin. B, bound DNA; F, free DNA. (b) EMSA was performed as described for panel a, except that SmcR was mixed with DMSO (2%) or increasing amounts of QStatin as indicated. (c to e) QStatin alters the interaction between SmcR and its target promoter DNAs. The promoter DNA of vvpE (c), flhF (d), or VVMO6_03194 (e) was titrated with the apo-SmcR complex (left) or the SmcR-QStatin complex (right). The thermodynamic values calculated from the reactions are shown. Data are representative of two experiments with similar results. (f) The in vivo interaction between SmcR and the promoter DNAs in the presence (20 μM) or absence (0.02% DMSO) of QStatin was analyzed by ChIP. V. vulnificus ΔsmcR was included as a control. A representative image from two independent experiments is shown.