title: Purification of 6-phosphogluconate dehydrogenase from chicken liver and investigation of some kinetic properties.
abstract: 6-phosphogluconate (6PG) dehydrogenase (EC 1.1.1.44; 6PGD) was purified from chicken liver; some kinetic and characteristic properties of the enzyme were investigated. The purification procedure consisted of four steps: preparation of the hemolysate, ammonium sulfate precipitation, 2',5'-ADP Sepharose 4B affinity chromatography, and Sephadex G-200 gel filtration chromatography. Thanks to the four consecutive procedures, product having a specific activity of 61 U (mg proteins)(-1), was purified 344-fold with a yield of 5.57%. Optimum pH, stable pH, optimum temperature, and KM and Vmax values for NADP+ and 6PG substrates were determined for the enzyme. Molecular weight of the enzyme was also determined by Sephadex G-200 gel filtration chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). In addition, Ki values and inhibition types were estimated by means of Lineweaver-Burk graphs obtained for NADPH and CO2 products.
authors.affiliation: Biotechnology Application and Research Center, Atatürk University, Erzurum, Turkey. eratm@atauni.edu.tr
authors.forename: Mustafa
authors.initials: M
authors.lastname: Erat
mesh_terms: D000818:Animals; D005591:Chemical Fractionation / Q000379:methods*; D002645:Chickens; D002846:Chromatography, Affinity / Q000379:methods*; D003911:Dextrans; D004789:Enzyme Activation; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D010734:Phosphogluconate Dehydrogenase / Q000737:chemistry* / Q000302:isolation & purification*; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D003911:Dextrans; C025614:sephadex; D010734:Phosphogluconate Dehydrogenase