6-phosphogluconate dehydrogenase of Aspergillus oryzae (Ahlburg).II.Separation and study of isoenzymes

Abstract

Through ammonium sulphate fractionation followed by ion-exchange chromatography, two forms of 6-phosphogluconate dehydrogenase (6j-phosphogluconate: NADP+ oxidoreductase E.C.1.1.1.44) were isolated from mycelium of Aspergillus oryzae.The Km values for 6PG were the same, but the Km for NADP+ was smaller for isoenzyme I.The Vmx values were also different, with greater activity for isoenzyme II.The optimum pH for isoenzyme I was 7.5.whereas the isoenzyme II had two maximum peaks, at pH 7.5 and 9.Isoenzyme II was the more resistant to thermal inactivation.Inhibition by NADPH was competitive with respect to NADP+, with a Ki value similar for both isoenzymes.The GSSG inhibits isoenzyme I and activates isoenzyme II.The apparent molecular weights determined by gel filtration were 79,000 +/- 2,000 daltons for isoenzyme I and 112,000 +/- 2,000 daltons for isoenzyme II.