Specific cation modulation of anion transport across the human erythrocyte membrane

The specific modulation by three cations, Ca²⁺, Mg²⁺, and tetracaine of the equilibrium exchange of SO₄ ^2- across the erythrocyte membrane was investigated. While external calcium had no effect on SO₄ ^2- exchange, internal calcium, and external calcium in the presence of 10 μM A23187 were found to be potent inhibitors of the exchange reaction. The apparent inhibition constants (K₁) for Ca²⁺ were calculated to be 6.1 μM and 5 μM for the above two conditions, respectively. Unlike Ca²⁺, Mg²⁺ was shown to be a weak activator of SO₄ ^2- exchange with an apparent dissociation constant of 3.6 μM. Competition experiments demonstrated that the Ca²⁺ and Mg²⁺ sites associated with anion transport are distinct and noninteracting. Tetracaine, a cation at neutral pH, was also found to be an inhibitor of SO₄ ^2- exchange with an apparent K₁ of 0.8 mM. Although tetracaine was observed to displace calcium from non-specific sites on the erythrocyte membrane, it showed no effect on the apparent inhibition constant of Ca²⁺ for SO₄ ^2- exchange. Thus, the Ca²⁺ and tetracaine sites also appear to be independent. The difficulty of situating three mutually independent sites on a single subunit protein, i.e., band 3, is considered. Using the experimental data obtained from five individuals, the concentration of free calcium in the red cell cytoplasm was calculated to range from 0.2 to 0.7 μM. This concentration was sufficient to reduce SO₄ ^2- exchange only 3-8%. It was concluded that calcium inhibition of anion exchange, and, hence, impairment of CO₂ transport, may be physiologically significant only in senescent cells and in certain types of anemia where calcium concentrations are significantly increased.