Thermodynamical Studies of Oxygen Equilibrium of Hemoglobin NONUNIFORM HEATS AND ENTROPY CHANGES FOR THE INDIVIDUAL OXYGENATION STEPS

Precise oxygen equilibrium curves of human adult hemoglobin were determined by the automatic recording method at several temperatures in the presence and absence of 2,3diphosphoglycerate (DPG) or inositol hexaphosphate (IHP) with 0.05 M 2,2-bis(hydroxymethyl)-2,2’,2”nitrolotriethanol (bis-tris) buffers (pH 7.4) containing 0.1 M Cl-. The equilibrium data were analyzed according to the Adair scheme, and the heats, AHi (i = 1,2,3,4) and the entropy changes, A& (i = 1,2,3,4), for the individual oxygenation steps were obtained. The shape of the equilibrium curve varies on temperature changes whether DPG or IHP is present or absent. In consequence, the AHi value depends on i and on the presence of DPG and IHP. Behavior of ASi is similar to that of AHi. The similar behavior of AHi and A& resulted in a compensation phenomenon. The contribution of T,ASi to the free energy change is compensated by the contribution of AHi at the first three oxygenation steps but not at the fourth step, and for i = 1,2, and 3 changes of T,ASi value upon the addition of DPG and IHP are compensated by accompanied changes of AHi value, where T c ( = 260 K) is the compensation temperature. A major part of both the enthalpy-entropy compensation and nonuniformity of AHi and ASi appears to be attributable to contributions of the oxygen-linked binding of Cl-, DPG and IHP, by hemoglobin. The present results do not necessarily support the earlier idea of Wyman that the cooperative oxygen binding is essentially an entropy effect.