Purification of glucose 6-phosphate dehydrogenase from chicken erythrocytes. Investigation of some kinetic properties

Glucose 6-phosphate dehydrogenase (G6PD) was purified from chicken erythrocytes, and some characteristics of the enzyme were investigated. The purification procedure was composed of three steps: hemolysate preparation, ammonium sulfate precipitation, and 2',5'-ADP Sepharose 4B affinity gel chromatography. Thanks to the three consecutive procedures, the enzyme, having the specific activity of 20.862 EU/mg proteins, was purified with a yield of 54.68% and 9,150-fold. Optimal pH, stable pH, optimal temperature, molecular weight, and K-M and V-max values for NADP(+) and glucose 6- phosphate (G6-P) were also determined for the enzyme. In addition, K-i values and the type of inhibition were determined by means of Line-Weaver-Burk graphs obtained for such inhibitors as ATP, ADP, NADH, and NADPH.