The interaction between erythrocyte organic phosphates, magnesium ion, and hemoglobin.

Oxygen equilibria have been used to study the interaction between red cell organic phosphates, magnesium ion, and hemoglobin. We measured the oxygen affinity of hemoglobin solutions containing either 2,3-diphosphoglycerate (2,3-DPG) or ATP and varying concentrations of magnesium ion at lo”, ZO”, and 30”. Deoxyhemoglobin and Mg2f were competitors, binding reciprocally with the organic phosphates. Stability constants derived from these data were in the order: HbDPG > HbATP > MgATP > MgDPG. The binding of deoxyhemoglobin to 2,3-DPG was more temperature dependent than its binding to ATP (AI? = 11 kcal per mole and -6.1 kcal per mole respectively). Unlike MgATP, the binding of magnesium ion to 2,3-DPG was independent of temperature. From these equilibria and a knowledge of the total red cell concentration of magnesium, ATP, 2,3-DPG, and hemoglobin, we have estimated the distribution of bound and unbound molecules within oxygenated and deoxygenated erythrocytes under physiological conditions. The bulk of red cell ATP is bound to magnesium and therefore is ineffective in mediating hemoglobin function. Upon deoxygenation there is a considerable increase in the concentration of free magnesium ion.