Figure 5.

DB270 binds PU.1 in the absence of DNA. Representative direct titrations of 10 nM of DB270 with PU.1 ETS domain at 150 mM (A), 300 mM (B) and 600 mM NaCl (C). Curves represent an empirical fit of the data to the Hill equation (dashed) or a mechanistic fit (solid) to a model in which DB270 binds n independent sites on PU.1 with a microscopic dissociation constant k₁₀₁, as defined by Scheme 2. Parametric values are given in Table 2. (D) Emission spectra of 10 nM of DB270 in the free state (dashed) and when saturated with 1 μM of PU.1Δ167 (solid). The sample was excited at 358 nm. (E) Electrophoretic resoloution of the PU.1/DB270 complex. Mixtures of 1 μg of PU.1 (∼70 pmol) were titrated with graded amounts of DB270 before separation on a 12% non-denaturing polyacrylamide gel. Note the anode at the bottom of the gel.