FIG. 5.

Electrophoretic mobility shift assay of HypR binding to the promoter region of hypR and ahpCF. (A) Effect of HypR concentration. Several half-dilutions of purified HypR were incubated with a ³²P-labeled hypR promoter region obtained with the Phyp5 and Phyp3 primers (Table 1). Lanes 1 to 10 contained 4 μg, 2 μg, 1 μg, 0.5 μg, 0.25 μg, 0.125 μg, 62.5 ng, 32 ng, 16 ng, and 8 ng of HypR, respectively. (B and C) Specificity of the HypR-DNA interaction. Purified HypR was incubated with a ³²P-labeled hypR (B) or ahpCF (C) promoter region obtained with the Pahp5 and Pahp3 primers (Table 1). DNA fragments were incubated without protein (lane 1), with HypR (lane 2), with an unlabeled competitor (lane 3), and with a nonspecific DNA fragment (lane 4).