(A–D) Instrumental responses of successive injections of ADP (8.39 mM, 1.92 mM, 5.82 mM and 4.05 mM ADP for ClpB-WT, ClpB-K212A, ClpB-E432A, ClpB-Y503D, respectively) into a solution of ClpB-WT (678 μM), ClpB-K212A (239 μM), ClpB-E432A (467 μM) and ClpB-Y503D (396 μM). The molar ratio of ADP to ClpB hexamer is indicated. Raw isothermal titration calorimetry data are shown. Integrated data after base-line-correction and fitting to the respective binding isotherms to a single-site binding model are given (lower panel). The number of ADP binding sites was calculated.