Chloride-bicarbonate exchange in red blood cells: physiology of transport and chemical modification of binding sites

Abstract

About 80% of the CO 2 formed by metabolism is transported from tissues to lungs as bicarbonate ions dissolved in the water phases of red cells and plasma. The catalysed hydration of CO 2 to bicarbonate takes place in the erythrocytes but most of the bicarbonate thus formed must be exchanged with extracellular chloride to make full use of the carbon dioxide transporting capacity of the blood. The anion transport capacity of the red cell membrane is among the largest ionic transport capacities of any biological membrane. Exchange diffusion of chloride and bicarbonate is nevertheless a rate-limiting step for the transfer of CO 2 from tissues to lungs. Measurements of chloride and bicarbonate self-exchange form the basis for calculations that demonstrate that the ionic exchange processes cannot run to complete equilibration at capillary transit times less than about 0.5 s. The anion exchange diffusion is mediated by a large transmembrane protein, constituting almost 30% of the total membrane protein. The kinetics of exchange diffusion must depend on conformational changes of the protein molecule, associated with the binding and subsequent translocation of the transported anion. We have characterized the nature of anion-binding sites facing the extracellular medium by acid-base titration of the transport function and modification of the transport protein in situ with group-specific amino acid reagents. Anion binding and translocation depend on the integrity and the degree of protonation of two sets of exofacial groups with apparent pA values of 12 and 5, respectively. From the chemical reactivities towards amino acid reagents it appears that the groups whose p K = 1 2 are guanidino groups of arginyl residues, while the groups whose p K = 5 are likely to be carboxylates of glutamic or aspartic acid. Our studies suggest that the characteristics of anion recognition sites in watersoluble proteins and in the integral transport proteins are closely related.