Fructose 3-phosphate and 5-phosphoribosyl-1-pyrophosphate formation in perfused human erythrocytes: 31P NMR studies.

31P NMR was used to study the formation of fructose 3-phosphate (F3P) and 5-phosphoribosyl-1-pyrophosphate (PRPP) in perfused human erythrocytes, in the presence of 10 different combinations and concentrations of glucose, inosine, pyruvate, fructose, and inorganic phosphate (Pi). (1) The cells were immobilized in alginate-coated agarose threads and perfused with a medium containing fructose, and the level of F3P increased continuously over more than 10 h. The net rate of F3P formation was independent of the concentration of 2,3-bisphosphoglycerate (2,3-DPG) present in the cells. (2) PRPP was formed in high concentrations, relative to normal, in immobilized cells when they were perfused with a medium containing Pi at a low pH (6.6). (3) The 2,3-DPG level decreased simultaneously when the sample was perfused with a medium containing fructose, but without inosine or pyruvate. The measured intracellular pH and free Mg2+ concentration were constant in these experiments. (4) The experiments confirmed the presence of fructose-3-phosphokinase (E.C. 2.7.1.-) and ribose-phosphate pyrophosphokinase (E.C. 2.7.6.1) activity in the human erythrocytes and that the biosynthetic pathways are active in immobilized cells at 37 degrees C. (5) The rates of accumulation of 2,3-DPG and phosphomonoesters (PME) appeared to be strongly correlated.