FIG. 2.
Determination of Ni-HpNikR K50 on PfecA3 wild type. (A) Electrophoretic mobility shifts of the wild-type promoter in the presence of increasing amounts of holo-HpNikR and 200 ng of nonspecific competitor DNA. Lane 1, no protein added; lane 2, 0.34 nM; lane 3, 3.4 nM; lane 4, 9.8 nM, lane 5, 34 nM; lane 6, 68.6 nM; lane 7, 137.2 nM. Mobility shifts were quantified by ImageQuant analysis (Molecular Dynamics). (B) The K50 was extrapolated graphically by the binding curves. A K50 equal to 3.5 nM was derived from the binding curves obtained by plotting the percentage of free probe against increasing HpNikR (inset). The error bars represent the standard deviations between the calculated K50s of at least two independent experiments.