Purification and properties of 6-phosphogluconate dehydrogenase from rabbit mammary gland.
1. 6-Phosphogluconate dehydrogenase from rabbit mammary gland was purified to homogeneity by the criterion of polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. The molecular weight of the subunit is 52 000. The enzyme was purified 150-fold with a final specific activity of 20 mumol of NADP+ reduced/min per mg of protein and overall yield of 3%. The molecular weight of the native enzyme is estimated to be 104 000 from gel-filtration studies. The final purification step was carried out by affinity chromatography with NADP+-Sepharose. 2. The Km values for 6-phosphogluconate and NADP+ are approx. 54 muM and 23 muM respectively. 3. Citrate and pyrophosphate are competitive inhibitors of the enzyme with respect to both 6-phosphogluconate and NADP+. 4. MgCl2 affects the apparent Km for NADP+ at saturating concentrations of 6-phosphogluconate.
D000645:Ammonium Sulfate; D000818:Animals; D002498:Centrifugation; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008274:Magnesium; D008321:Mammary Glands, Animal / Q000201:enzymology*; D008970:Molecular Weight; D009249:NADP; D010734:Phosphogluconate Dehydrogenase / Q000302:isolation & purification* / Q000378:metabolism; D011506:Proteins / Q000032:analysis; D011817:Rabbits; D012967:Sodium Dodecyl Sulfate
D011506:Proteins; D012967:Sodium Dodecyl Sulfate; D009249:NADP; D010734:Phosphogluconate Dehydrogenase; D008274:Magnesium; D000645:Ammonium Sulfate