Isolation and characterization of dihydropteridine reductase from Pseudomonas species

Abstract

Dihydropteridine reductase isolated from the bacterium Pseudomonas species (ATCC 11299a) has been purified approximately 450-fold by ammonium sulfate precipitation and diethylaminoethyl-cellulose chromatographic procedures. The preparation is at least 80% pure as judged by polyacrylamide gels. Its molecular weight was determined to be about 44,000. Both dihydropteridine reductase and phenylalanine hydroxylase activities were found to be higher in cells adapted to a medium containing L-phenylalanine or L-tyrosine as the sole carbon source than in those grown in L-asparagine. The substrate of the reductase is quinonoid dihydropteridine, and the product is tentatively identified as a tetrahydropteridine through its ability to serve as a cofactor for phenylalanine hydroxylase. The enzyme shows no marked specificity for the pteridine cofactor that occurs naturally in this organism, L-threo-neopterin. The pH optimum for the reductase is 7.2, and nicotinamide adenine dinucleotide, reduced form, is the preferred cosubstrate. Inhibition of the reduced and untreated enzyme by several sulfhydryl reagents was observed. A metal requirement for the reductase could not be demonstrated. Dihydropteridine reductase was found to be inhibited by aminopterin in a competitive manner with respect to the quinonoid dihydro form of 2-amino-4-hydroxy-6,7-dimethyl-5,6,7,8-tetrahydropteridine.

The NprA nitroreductase required for 2,4‐dinitrophenol reduction in Rhodobacter capsulatus is a dihydropteridine reductase

“…4). Dihydropteridine reductases are present in some bacteria (Williams et al ., 1976; Vasudevan et al ., 1988; 1992; Wilquet et al ., 2004), but no dihydropteridine reductase homologous genes have been described in phototrophic bacteria. Rhodobacter sphaeroides 2.4.1 has a putative dihydrofolate reductase gene (Accession No.…”

Isolation and characterization of dihydropteridine reductase from human liver

“…Involvement ofdihydropteridine reductase in the hydroxylation ofthe aromatic amino acids 0306-3275/81/070031-13$01.50/1 (© 1981 The Biochemical Society Vol. 197 characterized from sheep liver (Craine et al, 1972;Cheema et al, 1973; and brain (Cheema et al, 1973), bovine liver (Hasegawa, 1977;Aksnes et al, 1979), adrenal medulla (Cheema et al, 1973;Aksnes et al, 1979), kidney (Chauvin et al, 1979) and brain (Snady & Musacchio, 1978); rat liver and a Pseudomonas species (Williams et al, 1976). In the main, these studies have indicated a similarity of properties and structure for dihydropteridine reductase from all these sources.…”

Structural insights into the dual substrate specificities of mammalian andDictyosteliumdihydropteridine reductases toward two stereoisomers of quinonoid dihydrobiopterin

“…A 5 min perincubation at 25 °C, 2.8 μg enzyme was added to the cuvette followed by mixing. The reaction mixture was incubated for 1 min at 25 °C, and then the utilization of NADH in the reaction mixture was spectrophotometrically determined at 340 nm based on the oxidation of NADH [8,26].…”