Figure 3.

Comparison of the binding isotherms for Ni²⁺-free (panel A) and ZnCl₂-treated PDF_Ec (panel B) titrated with 325 μM actinonin in 50 mM potassium phosphate, pH 7.0 at 25 °C. Total protein concentrations were 31.5 μM in both experiments and the best fit of single binding site model (solid line in panel A) provided n, K_a and ΔH° of 0.41, 2.7 × 10⁶ and 3.6 kcal mol⁻¹, respectively for Ni²⁺-free PDF_Ec. The solid line in panel B corresponds to the best fit of the ZnCl₂-treated PDF_Ec experimental data to an independent two-site model providing n of 0.31 and 0.42, K_a of 3.9 × 10⁸ and 5.7 × 10⁶ M⁻¹ and ΔH° of −2.8 and 2.0 kcal mol⁻¹ for the high and low affinity binding sites, respectively.