A New Assay For 6-Pyruvoyl Tetrahydropterin Synthase From A Rat Kidney Model

Abstract

The enzyme 6-pyruvoyl tetrahydropterin synthase (PTP synthase) is the second enzyme in a four - step pathway which leads to the production of tetrahydrobiopterin, the cofactor for aromatic amino acid hydroxylations. In attempts to develop an assay, PTP synthase was characterized in terms of what effect its concentration, substrate (dihydroneopterin triphosphate-h^NTP) concentration, incubation time, and cofactor concentration had on product (sepiapterin) formation. In a reaction mixture of 100 pi, sepiapterin production is proportional to enzyme amount only up to 10 pi of enzyme. Sepiapterin production is linear to substrate concentration up to 60 pM after which, sepiapterin production levels off. Sepiapterin production increases linearly for 60 min and then falls off this course and levels off. The cofactors, Mg++ and NADPH had no limiting or inhibiting effects of sepiapterin production at the concentrations tested.