Human erythrocyte ammonium transport is mediated by functional interaction of ammonium (RhAG) and anion (AE1) transporters

The ammonia/ammonium (NH₃/NH₄⁺) influx into red blood cells (RBCs) is mediated by surface glycoprotein RhAG that forms a structural complex with anion exchanger 1 (AE1, band 3). Owing to the activity of this complex, RBCs exposed to the isosmotic ammonium buffer swell and finally lyse. Isoosmotic NH₄⁺-containing media alters the pH gradient in RBCs (intracellular alkalosis in response to NH₃/NH₄⁺ influx) and triggers the AE1 activity resulting in redundant chloride and water influx and finally in cell swelling. Here we demonstrate that the ammonia/ammonium transport in human RBCs depends on the pH (pH optimum 7.4 ± 0.1), temperature (Q₁₀ 2.6 ± 0.3), HCO₃⁻ concentration (EC₅₀ 4.7 ± 0.3 mM), and AE1 function. The data confirm functional interactions between AE1 and RhAG. The initial velocity of cell swelling increased almost 50-fold in the isosmotic ammonium buffer containing 25 mM HCO₃⁻ (37°C) in comparison to the reaction in the same buffer without HCO₃⁻. This indicates that the reaction is facilitated mostly by the carrier proteins, not just owing to the simple diffusion of NH₃ across the erythrocyte membrane. We demonstrate that pH_i reaches its maximum value much faster than the volume increase does. These data suggest that there is no direct correlation between pH_i changes and the influx of NH₃/NH₄⁺. Taken together, our data show that the RhAG and AE1 complex activity enables erythrocytes to be ammonia/ammonium storage sites in order to maintain the physiological blood ammonia/ammonium equilibrium.