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. 2005 Jun;187(11):3762–3778. doi: 10.1128/JB.187.11.3762-3778.2005

FIG. 2.

FIG. 2.

Gel mobility shift assay for FhuR binding to different regulated gene promoter regions. The promoter regions of FhuR target genes were PCR amplified and labeled. Radiolabeled DNA probes (10 × 10−15 mole) for metB2 (A), plpA (B), yjgC (C), cysM (D), cysD (E), and metA (F) were incubated with purified FhuR protein at various concentrations and analyzed on nondenaturing polyacrylamide gel electrophoresis (see Materials and Methods for details). (A and B) Lane 1, no protein; lanes 2 to 5, 0.6, 1.2, 2.4, and 3.6 μM final concentrations of the FhuR protein, respectively; lane 6, 3.6 μM final concentration of the FhuR protein with an excess of unlabeled DNA probe (10 × 10−13 mole). As a negative control, lanes 7 and 8 contain no protein and 3.6 μM (final) of the FhuR protein, respectively, with a purH gene 32P-labeled internal fragment as DNA probe. (C to F) Lanes 1 and 2, no protein and 3.6 μM (final) of the FhuR protein, respectively; lane 3, 3.6 μM (final) of the FhuR protein with an excess of unlabeled DNA probe (10 × 10−13 mole). In all panels, the bands for the probe (free) and the FhuR-probe complex (bound) are indicated by an arrow and an arrow with an asterisk, respectively.