Figure 2. In vitro analysis of CNBD–cAMP... | Rockefeller University Press

Figure 2.

In vitro analysis of CNBD–cAMP interaction. Examples of binding of cAMP to His6-MBP-tagged human HCN2 CNBD WT (A) and mutants (B–D) measured by ITC. Top: Heat changes (μcal/s) during successive injections of 8 µl of the corresponding cAMP (200 µM) into the chamber containing CNBD (20 µM). Bottom: Binding curve obtained from data displayed in the upper panel. The peaks were integrated, normalized to cAMP concentration, and plotted against the molar ratio (cAMP/CNBD). Solid red line represents a nonlinear least-squares fit to a single-site binding model (see Materials and methods).

In vitro analysis of CNBD–cAMP interaction. Examples of binding of cAMP to His₆-MBP-tagged human HCN2 CNBD WT (A) and mutants (B–D) measured by ITC. Top: Heat changes (μcal/s) during successive injections of 8 µl of the corresponding cAMP (200 µM) into the chamber containing CNBD (20 µM). Bottom: Binding curve obtained from data displayed in the upper panel. The peaks were integrated, normalized to cAMP concentration, and plotted against the molar ratio (cAMP/CNBD). Solid red line represents a nonlinear least-squares fit to a single-site binding model (see Materials and methods).