FIGURE 7.

ITC analysis of nucleotide binding to AtAPSK D136A. A, titration of AtAPSK D136A with ADP (solid squares), ATPγS (open squares), ATP (closed circles), and AMP-PNP (open circles). B, titration of AtAPSK D136A with APS (solid squares), APS + 5 mm Mg²⁺ (open squares), and AtAPSK with APS (open circles). C, titration of AtAPSK D136A·ADP with APS (open squares), APS + 5 mm Mg²⁺ (solid squares), and AtAPSK D136A·AMP-PNP (open circles) with APS + 5 mm Mg²⁺. Representative experimental data for the ADP (A) and APS (B and C) titrations are plotted as heat signal (μcal s⁻¹) versus time (min) in each upper panel. Each experiment consisted of 20 to 30 injections of 10 μl each of nucleotide into a solution containing AtAPSK dimer. Each lower panel shows the integrated heat responses per injection. The solid line is the linear regression fit using a two-site binding model (see Table 6).