MOLECULAR BIOLOGY OF THE PTERIN-DEPENDENT HYDROLASES AND ANCILLARY ENZYMES

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High level (25% of total protein) expression of a phenylalanine hydroxylase sequence lacking the first seven amino acid residues has been achieved in E. coli. A wild type version of this clone is under construction to allow comparisons of catalytic properties and directed mutagenesis. Partial human phenylalanine hydroxylase cDNA clones have been characterized and full length clones are being pursued through the synthesis of optimized cDNA libraries. Expanded experiments with rats have been conducted where chemical induction of diabetes in rats has been performed in conjunction with insulin treatment to clearly define the mechanism of increased phenylalanine hydroxylase activity and mRNA in diabetic rats. A full length coding sequence for human dihydropteridine reductase has been expressed on a bacterial vector and enzyme isolated from large scale cultures has been characterized. The dihydropteridine reductase clone has been used as a probe to identify the location of this gene on chromosome 4 near the Huntington's Disease locus.