RGD Reference Report - Comparative activity of rat liver dihydrofolate reductase with 7,8-dihydrofolate and other 7,8-dihydropteridines. - Rat Genome Database

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Comparative activity of rat liver dihydrofolate reductase with 7,8-dihydrofolate and other 7,8-dihydropteridines.

Authors: Webber, S  Whiteley, JM 
Citation: Webber S and Whiteley JM, Arch Biochem Biophys. 1985 Feb 1;236(2):681-90.
RGD ID: 2300196
Pubmed: (View Article at PubMed) PMID:3970530

The various interactions of rat liver dihydrofolate reductase with two unconjugated 7,8-dihydropteridines, 7,8-dihydrobiopterin and 6-methyl-7,8-dihydropteridine, have been compared with those of 7,8-dihydrofolate and folate. Of particular interest was the reactivity demonstrated by 7,8-dihydrobiopterin because of the potential physiological significance of this reaction both in the regeneration of tetrahydrobiopterin, a cofactor for various biological hydroxylations, and as a step in the biosynthesis of this compound from GTP. Kinetic experiments gave Km values of 0.17, 6.42, and 10.2 microM for 7,8-dihydrofolate, 7,8-dihydrobiopterin, and 6-methyl-7,8-dihydropteridine, respectively, with Vmax = 6.22, 2.39, and 1.54 mumol min-1 mg-1. With folate the enzyme showed high affinity (Km = 0.88 microM) but low Vmax (0.20 mumol min-1 mg-1). The natural cofactor was NADPH and a Km of approximately 0.7 microM was measured with each substrate. The enzyme was activated by both p-hydroxymercuribenzoate and urea when assayed with 7,8-dihydrofolate but was inhibited when 7,8-dihydrobiopterin was the substrate. The pH optimum for dihydrofolate reduction was 4 with enhancement at pH greater than or equal to 5.5 in the presence of 1 M NaCl. Peak activity with 7,8-dihydrobiopterin occurred at pH 4.8; this was shifted to pH 5.3 but was not enhanced by 1 M NaCl. Inhibition with methotrexate was similar whether the enzyme was assayed with either the conjugated or unconjugated 7,8-dihydro derivatives. The rat liver enzyme, highly unstable after purification, was stabilized in the presence of the nonionic detergent, Tween-20 (0.1%); however, the comparative properties toward the conjugated and unconjugated substrates were not altered by this treatment.

Annotation

Gene Ontology Annotations    

Biological Process

Molecular Function

Objects Annotated

Genes (Rattus norvegicus)
Dhfr  (dihydrofolate reductase)


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