Fig. 3. Comparison of in vivo and in vitro activities of DmpR-Flag derivatives. (A) Western analysis of expression levels of DmpR-Flag (lane 1), DmpR-scramble-Flag (lane 2), DmpR-L215,219,222E-Flag (lane 3) and DmpR-L215,219,222A-Flag (lane 4) in 30 µg of total protein derived from cultures used to determine in vivo transcriptional responses. (B–E) Histograms show the in vivo luciferase transcriptional response of P.putida KT2440::Po-luxAB harbouring plasmids expressing DmpR-Flag (B), DmpR-scramble-Flag (C), DmpR-L215,219,222E-Flag (D) or DmpR-L215,219,222A-Flag (E) measured in the absence or presence of 2 mM 2-methylphenol (2-mp) or 4-methylphenol (4-mp). Values are the averages of triplicate determinations from each of two independent experiments. Line graphs show the level of in vitro ATP hydrolysis mediated by 1 µl of bead-bound Flag-tagged protein, in the absence (open symbols) and presence (closed symbols) of 1 mM 2-methylphenol. Data are the average of duplicate experiments and expressed as the percentage of total ATP (180 nM) hydrolysed.