Inhibition of phosphate transport across the human erythrocyte membrane by chemical modification of sulfhydryl groups

Effects of sulfhydryl-reactive reagents on phosphate transport across human erythrocyte membranes were examined using 31P NMR. Phosphate transport was significantly inhibited in erythrocytes treated with sulfhydryl modifiers such as N-ethylmaleimide, diamide, and Cu2+/o-phenanthroline. Quantitation of sulfhydryl groups in band 3 showed that the inhibition is closely associated with the decrease of sulfhydryl groups. Data from erythrocytes treated with diamide or Cu2+/o-phenanthroline demonstrated that intermolecular cross-linking of band 3 by oxidation of a sulfhydryl group, perhaps Cys-201 or Cys-317, decreases the phosphate influx by about 10%. The inhibition was reversed by reduction using dithiothreitol. These results suggest that sulfhydryl groups in the cytoplasmic domain of band 3 may play an important role in the regulation of anion exchange across the membrane.