The Reaction of Oxygen with Hemoglobin and the Kinetic Basis of the Effect of Salt on Binding of Oxygen

The reaction of hemoglobin with oxygen has been studied by stopped flow methods, and, under suitably restricted conditions, it can be adequately represented by a system of four consecutive reversible reactions. The numerical values given to the eight rate constants permit a satisfactory fit to combination and dissociation velocity data, and yield an equilibrium curve of the appropriate form. The distribution of rates among the various steps in the reaction requires that cooperativity in oxygen binding be attributed primarily to deviations of the successive dissociation velocity constants from their statistical values, and is consistent with the idea that the major change in reactivity occurs after 1 ligand molecule has dissociated from saturated hemoglobin. The difference in affinity between hemoglobin in phosphate buffers and hemoglobin freed from salts is due to reduction in the rate of dissociation of the 2nd, 3rd, and 4th molecules leaving oxyhemoglobin. The rate of dissociation of the 1st molecule from saturated hemoglobin is not changed.