Deoxygenation and elevation of intracellular magnesium induce tyrosine phosphorylation of band 3 in human erythrocytes

Deoxygenation increases the level of tyrosine phosphorylation of band 3 by ~25% in human red blood cells (RBCs), as determined by Western blotting. The effect is much more pronounced in osmotically shrunken RBCs or in the presence of vanadate. When the rise in intracellular free Mg²⁺ concentration in deoxygenated RBCs is simulated via clamping of the intracellular magnesium in oxygenated RBCs by ionomycin, band 3 phosphorylation is elevated by up to 10-fold. Phosphorylated band 3 is preferentially retained by RBC skeletons, after mild extraction with Triton X-100. Elevation of intracellular free Mg²⁺ leads to band 3 phosphorylation and is accompanied by rigidification of the membrane skeleton as determined by analysis of RBC membrane mechanical fluctuations. These findings suggest that the visco-elastic properties of human erythrocytes may be regulated by band 3 tyrosine phosphorylation. Copyright (C) 1999 Federation of European Biochemical Societies.