Measurement of the Bohr Effect: Dependence on pH of Oxygen Affinity of Hemoglobin

This chapter explains the measurement of the Bohr effect. The Bohr effect is the pH dependence of oxygen affinity in hemoglobin and is the expression of the linkage between the binding of oxygen and the binding of protons by the protein. The oxygen-linked ionizable groups change pK upon addition or removal of oxygen, thereby exchanging protons with the solvent. The Bohr effect can be measured either as the variation of oxygen affinity with pH or as the number of protons that dissociate from the hemoglobin molecule upon addition of oxygen. The chapter discusses the determination of the Bohr effect by the measurement of changes in proton binding. The potentiometric measurements of the amount of protons liberated or absorbed by hemoglobin upon addition of oxygen allow a very accurate determination of the Bohr effect measured by proton binding (BEh). The accuracy stems from the high precision with which voltages originating in the glass-calomel cell can be measured.