Figure 2.

Salt dependence of the non-specific and specific DNA binding affinity determined by fluorescence anisotropy measurements at 20°C. (A) Fraction bound of the non-specific DNA (D^N = 25 nM) as a function of hOGG1 concentration at varying potassium ion concentrations (30 mM – 300 mM). Binding curves for the additional non-specific DNA oligos are provided in Figure S3. The salt concentrations listed match the relative positions of the corresponding data set. (B) Fraction bound of the specific DNA (D^S = 10 nM) as a function of K249Q concentration at various potassium ion concentrations (30 mM – 200 mM). The salt concentrations listed match the relative positions of the corresponding data set. Data presented in both A and B are single trial measurements. The standard deviation in three replicate anisotropy intensity measurements was less than ±10% and that the K_a values and associated errors from independent measurements are reported in Tables 1 and 2. (C) Dependence of non-specific association constant (K_a^N) on the concentration of potassium acetate for WT hOGG1 binding to D^N (black), D^TATA (blue), D^GC (purple), and D^GCGC (red). Error bars are smaller than the size of the data markers. (D) Salt dependence of the non-specific (K_a^N) (black, left y-axis) and specific association constants (K_a^S) (red, right y-axis) for K249Q binding to D^N and D^S. Error bars are smaller than the size of the data markers. For comparison, black and grey dashed lines in C and D show the published salt dependences for non-specific and specific DNA binding by hUNG⁸.