Fig. 4. Calorimetric titration of pepsin with DRV and APV.

The titrations were performed at 28 °C in 50 mM sodium formate buffer, pH 3. Pepsin concentration in the cell was 14.4 μM for the titration with DRV and 21 μM for the titration with APV. Panels A and B are experimental traces of the thermal response on titration with successive aliquots of the inhibitor solutions; panels C and D are integrated data after processing by the instrument's Origin software. The data in panel D were corrected for a baseline of 1.1 kcal/mol per injection of APV. Curve fitting of the integrated data gave a binding constant (K_L) of (6.5 ± 0.6) × 10⁵ M and a stoichiometry of 0.90 ± 0.01 for DRV; and a binding constant of (5.5 ± 0.5) × 10⁶ and a stoichiometry of 0.97 ± 0.004 for APV.