title: Purification and characterization of 6-pyruvoyl-tetrahydropterin synthase from Drosophila melanogaster.
abstract: The enzyme 6-pyruvoyl-tetrahydropterin synthase (PTP synthase), which catalyzes the conversion of 7,8-dihydroneopterin triphosphate to 6-pyruvoyl tetrahydropterin, has been purified approx. 230-fold to apparent homogeneity from head extracts of Drosophila melanogaster. A partially purified 6-pyruvoyl-tetrahydropterin reductase (PTP reductase) was also prepared and in its presence, along with Mg2+ and NADPH, the purified PTP synthase converted 7,8-dihydroneopterin triphosphate to metastable 6-lactoyltetrahydropterin, which was autoxidized to sepiapterin under aerobic conditions. Purified PTP synthase had a specific activity of 3792 units per mg protein and migrated as a single protein band on both nondenaturing polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The purified active enzyme consisted of at least two identical subunits which had a molecular mass of 37.5 kDa on SDS-PAGE and NH2-Asx-Pro- as N-terminal amino acids. The native enzyme in crude extract was shown to be more complex, existing as higher multimeric forms.
authors.affiliation: Department of Microbiology, Seoul National University, Korea.
authors.forename: Y S
authors.initials: YS
authors.lastname: Park
authors.forename: J H
authors.initials: JH
authors.lastname: Kim
authors.forename: K B
authors.initials: KB
authors.lastname: Jacobson
authors.forename: J J
authors.initials: JJ
authors.lastname: Yim
mesh_terms: D000429:Alcohol Oxidoreductases / Q000302:isolation & purification* / Q000378:metabolism; D000645:Ammonium Sulfate; D000818:Animals; D000906:Antibodies; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004331:Drosophila melanogaster / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D007658:Ketone Oxidoreductases / Q000302:isolation & purification; D019761:Phosphorus-Oxygen Lyases
chemical_list: D000906:Antibodies; D000429:Alcohol Oxidoreductases; D007658:Ketone Oxidoreductases; C059712:6-pyruvoyl tetrahydropterin (2'-oxo)reductase; D019761:Phosphorus-Oxygen Lyases; C043497:6-pyruvoyltetrahydropterin synthase; D000645:Ammonium Sulfate