The heats of binding of protons m b to three globular proteins, lysozyme, chymotrysinogen A, and oxidized cytochrome c, from pH 11-2 or lower a t 25Β°C were determined by flow microcalorimetry. In addition, the acid-base titrations of cbymotrypsinogen A and oxidized cytochrome c were investigated under conditions similar to those used in the calorimetric experiments. The results of the calorimetric experiments and the acid-base titrations for lysozyme and chymotrypsinogen A in the neutral and alkaline pH regions are in accord with the Linderstrom-Lang model with a pH-independent electrostatic factor. However, in order to interpret the results on oxidized cytochrome c in the same pH region, it is necessary to assume the protonation of two unidentified groups; one of these groups, with AHb = -18 kcal/ mol and pK, = 9.4, was detected in previous work [Watt, G . D. & Sturtevant, J. M. (1969) Biochemistry 8,45671. The normal heats of protonation for carboxyl, c-amino, phenolic, a-amino, and imidazole groups on globular proteins as deduced from the study are 0, -10.5, -6.3, -10.0, and -6.3 kcal/mol, respectively. AHb of chymotrypsinogen A between pH 4.5 and 1.3 is +30 kcal/mol. This value, which is undoubtedly too large to be accounted for by the protonation of normal carboxyl groups, leads to the conclusion that this protein undergoes a pH-induced conformational change in this pH region. The same idea can be applied to explain the abnormal AHb observed for oxidized cytochrome c in the alkaline pH region.