BIOSYNTHESIS OF TETRAHYDROBIOPTERIN IN DROSOPHILA MELANOGASTER

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Genetic variations in the synthesis of tetrahydrobiopterin (H4B) have been associated with several human neurological disorders such as atypical phenylketonuria, Parkinson's disease, and Alzheimer's disease. The importance of H4B to these processes is found in its role as an essential cofactor for tyrosine and tryptophan hydroxylase, enzymes involved in the synthesis of catecholamines and serotonin, respectively. BH4 is also required by nitric oxide synthase in the synthesis of nitric oxide, a potent cell-signaling molecule which is involved in the control of vasodilation in mammals. BH4 is structurally conserved among eukaryotes, as is apparently the pathway for its biosynthesis. The biosynthesis of BH4, in organisms as diverse as insects, rats and humans, minimally requires GTP cyclohydrolase, 1, pyruvoyl-H4pterin reductase ina the biosynthesis of BH4 remains unclear. The proposed research includes the purification and biochemical characterization of the Drosophila pyruvoyl- H4pterin reductase. The purified enzyme will be used to raise polyclonal antisera. The alpha-pyruvoyl-H4pterin reductase antibody will be used to isolate a molecular clone encoding for the enzyme and in other biochemical characterizations such as the localization of pyruvoyl-H4pterin reductase in situ. A classical mutant screen for the isolation of a pyruvoyl- H4pterin reductase mutant is proposed as an independent undergraduate student project. Although several human "in born errors" of H4B biosynthesis exist (GTP) cyclohydrolase 1, pyruvoyl-H4pterin synthase, dihydropteridine reductase) there is no known pyruvoyl-H4pterin reductase mutation. Drosophila is the only organism where there is a direct strategy for obtaining such a mutant, due to the involvement of pyruvoyl-H4pterin reductase in pathways leading to eye pigment synthesis. The long term goal of these studies is to contribute to the understanding of the terminal steps of BH4 biosynthesis in Drosophila and other higher organisms.