Partial Purification of D-Xylulokinase in Human Erythrocyes

D-xylulokinase [EC 2. 7. 1. 17] was partialy purified from about 1 liter of acid-citrate-dixtrose (ACD) blood. A large portion of hemoglobin was removed by treatment with DEAE-cellulose, and then enzyme was purified by ammonium phosphate precipitation, calcium phosphate gel treatment, ammonium sulfate fractionation, DEAF-cellulose column chromatography and the sephadex G-200 gel filtration. Four peaks were obtained by eluting with a linear gradient system of KCl concentration with the DEAE-cellulose column chromatography. It seemed that the peak I was hemoglobin and the peak IV was D-xylulokinase. D-xylulokinase was purified 2, 000-fold yielding 9.7mg. Its specific activity was 3 units per mg protein. The molecular weight of the peak IV was about 260, 000 as estimated by sephadex G-200 gel column chromatography.In the phospharylation of D-xylulose by this enzyme, pentose 5-phosphates were produced. Analysis of the reaction product indicate that the main component of the product was xylulose 5-pyosphate.Thus, the present study confirms the postulation that D-xylulose is first phosphorylated by xylulokinase and then metabolized through pentose phosphate pathway to lactic acid.