Abstract
Ammonium sulfate fractionation and standard column chromatography techniques have been used to purify the enzyme sepiapterin reductase to electrophoretic homogeneity from pupae of Drosophila melanogaster. This purification constitutes a 1000-fold increase in the specific activity of the enzyme. The native molecular weight of the enzyme was determined to be ca 67,000 Da, while the subunit molecular weight is estimated to be 36,000-39,000 Da. The apparent Km for 6-lactoyltetrahydropterin (lactoyl-H4pterin) is 50 microns. The Drosophila enzyme is sensitive to inhibition by the biogenic amine, N-acetyl serotonin, and (to a lesser extent) melatonin, but its activity is not affected by serotonin, epinephrine or norepinephrine. The enzyme was shown to be an integral component of the Drosophila enzyme system which functions in catalyzing the conversion of dihydroneopterin triphosphate (H2NTP) to (6R)-5,6,7,8-tetrahydrobiopterin (H4biopterin). It appears that although purified Drosophila sepiapterin reductase can catalyze low levels of conversion of 6-pyruvoyltetrahydropterin (pyruvoyl-H4pterin) to H4 biopterin in the presence of NADPH, the efficient conversion of pyruvoyl-H4pterin to H4biopterin requires the presence of both sepiapterin reductase and pyruvoyl-H4pterin reductase.
