Developmental and biochemical studies on the phenylalanine hydroxylation system in Drosophila melanogaster

Abstract

The enzyme phenylalanine hydroxylase, the substrate phenylalanine, the product of the reaction tyrosine, and the probable in vivo cofactors (6R)- l-erytro-5,6,7,8-tetrahydrobiopterin (H 4Bip) and 5,6,7,8-tetrahydropterin (H 4Ptr), have been measured during development in Drosophila. The developmental profile of phenylalanine hydroxylase activity shows two peaks. The larger occurs at the time of pupation, coiciding with an important accumulation of tyrosine in the insect. The minor peak appears at the time of adult emergence. The developmental profile of H 4Bip shows also two peaks, coinciding with those of maximal phenylalanine hydroxylase activity. However, H 4Ptr is only detectable after late pupa, with a small peak at adult emergence. The results of the analyses of the phenylalanine hydroxylase activity in vitro showed that H 4Bip is a better cofactor than H 4Ptr, based on the difference in V max (4–5 times) and in K m for phenylalanine (1.6 times). Moreover, the difference in activity of the enzyme with H 4Bip related to that with H 4Ptr, increases even more when larvae are reared on phenylalanine-supplemented media (40-fold difference in V max). Also, the inhibition of H 4Bip synthesis with N-acetylserotonin resulted in an increase of phenylalanine. As in most other organisms studied, our results indicate that H 4Bip is by far the preferred natural cofactor for the Drosophila phenylalanine hydroxylase.