Osmotic pressure measurements of dissociating bovine hemoglobin

Abstract

While the dissociation of hemoglobin, in general, has been studied extensively using various techniques under both alkaline and acidic conditions, the dissociation under alkaline conditions using membrane osmometry has not been previously reported. An equilibrium mathematical model describing the two-stage dissociation is presented in this paper and agrees with the experimental osmotic pressure measurements of bovine hemoglobin at pH's above 7.4. Results of this study show that the pH dependency under alkaline conditions at 37°C can be regressed to give approximate values of the dissociation constants for both the dissociation of the tetramer to the dimer and the dimer to the monomer as a function of pH. The data also indicate that, in the presence of the borate buffer used in these experiments, the liberation of approximately one proton occurs for the dissociation of the tetramer into the dimers; approximately two protons are liberated for the dissociation of the dimer into the monomer. The dissociation constants reported here for bovine hemoglobin are in reasonable agreement with those obtained under similar conditions using other techniques.