Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.5.1.3 (dihydrofolate reductase)
5,819 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The natural killer-like cell line YT constitutively expresses GTP-cyclohydrolase activity whereas 6-pyruvoyltetrahydropterin synthase and sepiapterin reductase are absent. The product, dihydroneopterin triphosphate, is dephosphorylated and oxidized causing neopterin to accumulate in the cells. The activities of the H4biopterin synthesizing enzymes are not controlled by IFN-gamma or the synergistic action of both IFN-gamma and IL-2 as has been shown for monocytes/macrophages (Huber C. et al. (1984) J. Exp. Med. 160, 310) and CD4+ T cells, respectively (Ziegler I. et al. (1990) J. Biol. Chem. 265, 17026). Sepiapterin reductase specifically is induced by incubation of the cells with sepiapterin, leaving GTP-cyclohydrolase, 6-pyruvoyltetrahydropterin synthase and other enzymes related to pteridine metabolism (dihydropteridine reductase, dihydrofolate reductase) unaffected. The data indicate that H4biopterin synthesis is individually regulated in the diverse cellular components of the immune system.
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PMID:Control of pteridine biosynthesis in the natural killer-like cell line YT. 203 22

The novel tetrahydrofolate, 5,10-dideazatetrahydrofolic acid (DDATHF), was designed as an inhibitor of folate metabolism at a site other than dihydrofolate reductase. DDATHF has been shown to inhibit glycinamide ribonucleotide transformylase, a folate-requiring enzyme that catalyzes the first of two one-carbon transfer reactions in the de novo purine nucleotide biosynthetic pathway. Incubation of HL-60 promyelocytic leukemia cells with 5 x 10(-8) to 10(-5) M DDATHF resulted in a marked inhibition of growth after 48 h, with a complete cessation of cellular replication by day 4. Cell cycle analyses of DDATHF-treated HL-60 cells demonstrated an initial block in early S phase by day 3 followed by an accumulation of cells in the G1 and G2 + M phases of the cell cycle. Inhibition of growth was accompanied by a concentration-dependent increase in the percentage of mature myeloid cells that expressed nitroblue tetrazolium positivity, and a small increase in nonspecific esterase activity. Induction of differentiation and inhibition of growth by DDATHF were completely prevented by hypoxanthine and 5(4)-amino-4(5)-imidazole carboxamide, suggesting that depletion of intracellular purine nucleotide pools has an important role in the biological effects of this inhibitor. This possibility was confirmed by the finding that DDATHF caused a pronounced reduction in intracellular GTP and ATP levels within 2 h, with maximum decreases being observed by 24 h, a time interval which preceded the inhibition of cellular proliferation by this agent. Pyrimidine nucleoside triphosphate levels were markedly increased under these conditions. The findings indicate the importance of purine nucleotides to both the inhibition of growth and the induction of differentiation of HL-60 leukemia cells by DDATHF.
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PMID:Induction of HL-60 leukemia cell differentiation by the novel antifolate 5,10-dideazatetrahydrofolic acid. 275 15

5,10-Dideazatetrahydrofolate (DDATHF) is a new antimetabolite designed as an inhibitor of folate metabolism at sites other than dihydrofolate reductase. DDATHF was found to inhibit the growth of L1210 and CCRF-CEM cells in culture at concentrations in the range of 10-30 nM. The inhibitory effect of DDATHF on the growth of L1210 and CCRF-CEM cells was reversed by either hypoxanthine or aminoimidazole carboxamide. Growth inhibition by DDATHF was prevented by addition of both thymidine and hypoxanthine, but not by thymidine alone. 5-Formyltetrahydrofolate reversed the effects of DDATHF in a dose-dependent manner. DDATHF had no appreciable inhibitory activity against either dihydrofolate reductase or thymidylate synthase in vitro, but was found to be an excellent substrate for folylpolyglutamate synthetase. DDATHF had little or no effect on incorporation of either deoxyuridine or thymidine into DNA, in distinct contrast to the effects of the classical dihydrofolate reductase inhibitor, methotrexate. DDATHF was found to deplete cellular ATP and GTP over the same concentrations as those inhibitory to leukemic cell growth, suggesting that the locus of DDATHF action was in the de novo purine biosynthesis pathway. The synthesis of formylglycinamide ribonucleotide in intact L1210 cells was inhibited by DDATHF with the same concentration dependence as inhibition of growth. This suggested that DDATHF inhibited glycinamide ribonucleotide transformylase, the first folate-dependent enzyme of de novo purine synthesis. DDATHF is a potent folate analog which suppresses purine synthesis through direct or indirect inhibition of glycinamide ribonucleotide transformylase.
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PMID:A new folate antimetabolite, 5,10-dideaza-5,6,7,8-tetrahydrofolate is a potent inhibitor of de novo purine synthesis. 290 24

We have investigated the energy requirement of mitochondrial protein import with a simplified system containing only isolated yeast mitochondria, energy sources and a purified precursor protein. This precursor was a fusion protein composed of 22 residues of the cytochrome oxidase subunit IV pre-sequence fused to mouse dihydrofolate reductase. Import of this protein required not only an energized inner membrane, but also ATP. ATP could be replaced by GTP, but not by CTP, TTP or non-hydrolyzable ATP analogs. Added ATP did not increase the membrane potential of respiring mitochondria; it supported import even if the proton-translocating mitochondrial ATPase and the entry of ATP into the matrix were blocked. We conclude that ATP exerts its effect on mitochondrial protein import outside the inner membrane.
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PMID:Both ATP and an energized inner membrane are required to import a purified precursor protein into mitochondria. 303 90

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.
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PMID:Comparative activity of rat liver dihydrofolate reductase with 7,8-dihydrofolate and other 7,8-dihydropteridines. 397 May 30

The involvement of the nucleoside triphosphates in the initiation of the synthesis of the messenger ribonucleic acid of five T4 specific enzymes has been studied. Only one of these, the messenger RNA for deoxynucleosidemonophosphate kinase, can be initiated in the presence of one nucleoside triphosphate, namely ATP. All of the remaining four require the presence of at least two nucleoside triphosphates during the initiation period. The combination of ATP and UTP was best for the initiation of messenger RNA for dihydrofolate reductase, ATP and CTP for deoxycytidylate hydroxymethyltransferase and beta-glucosyltransferase, and ATP and GTP for alpha-glucosyltransferase. We have concluded that there is a great variation in the nucleotide composition and sequence of the initiation sites in T4 DNA. No correlation in the requirements of nucleoside triphosphates during the initiation period could be observed among the five systems studied according to their classification as one type or another of "early" T4 messenger RNA.
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PMID:Initiation characteristics for the synthesis of five T4 phage-specific messenger RNAs in vitro. 436 Sep 43

The regulation of GTP-cyclohydrolase (GTP-CH) activity and tetrahydrobiopterin (BH4) levels in the adrenal cortex were studied in intact and hypophysectomized rats. Treatment with a single dose of reserpine (5 mg/kg) or insulin-induced hypoglycemia (4 h) elevated adrenocortical BH4 3-fold by 10 h; BH4 levels remained elevated after 24 h and returned to control levels by 48-72 h. GTP-CH was significantly increased 24 h after hypoglycemic shock, and the increased enzyme activity preceded the changes in BH4 levels after reserpine treatment. Two and a half hours of stress by immobilization also increased GTP-CH activity and BH4 levels in the adrenal cortex. The activities of sepiapterin reductase and dihydrofolate reductase, putative enzymes in the biosynthetic pathway from GTP to BH4, were not increased by reserpine. Both reserpine and insulin increased the apparent maximum velocity for GTP, with no increase in the affinity of the enzyme for its substrate, further suggesting that the experimental treatments induce the synthesis of GTP-CH. Hypophysectomy completely blocked the reserpine-dependent increase in both cortical GTP-CH activity and BH4 content. The administration of purified porcine ACTH to intact and hypophysectomized rats elevated adrenocortical GTP-CH activity and cofactor levels. Synthetic ACTH-(1-24) also enhanced the enzyme activity and BH4 levels in the adrenal cortex. Thus, pituitary control of adrenal cortical GTP-CH synthesis and biopterin levels appears to be mediated through the secretion of ACTH. The changes in enzyme activity and cofactor levels after activation of the hypothalamo-hypophyseal axis or ACTH administration suggest that BH4, a cofactor for certain monooxygenases, has some function, as yet unknown, in the adaptive changes of the adrenal cortex in response to stress.
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PMID:Hormonal regulation of guanosine triphosphate cyclohydrolase activity and biopterin levels in the rat adrenal cortex. 613 42

Mammalian cells and tissues were found to have two pathways for the biosynthesis of tetrahydrobiopterin (BH4): (i) the conversion of GTP to BH4 by a methotrexate-insensitive de novo pathway, and (ii) the conversion of sepiapterin to BH4 by a pterin salvage pathway dependent on dihydrofolate reductase (5,6,7,8-tetrahydrofolate: NADP+ oxidoreductase, EC 1.5.1.3) activity. In a Chinese hamster ovary cell mutant lacking dihydrofolate reductase (DUKX-B11), endogenous formation of BH4 proceeds normally but, unlike the parent cells, these cells or extracts of them do not convert sepiapterin or 7,8-dihydrobiopterin to BH4. KB cells, which do not contain detectable levels of GTP cyclohydrolase or BH4 but do contain dihydrofolate reductase, readily convert sepiapterin to BH4 and this conversion is completely prevented by methotrexate. In supernatant fractions of bovine adrenal medulla, the conversion of sepiapterin to BH4 is completely inhibited by methotrexate. Similarly, this conversion in rat brain in vivo is methotrexate-sensitive. Sepiapterin and 7,8-dihydrobiopterin apparently do not enter the de novo pathway of BH4 biosynthesis and may be derived from labile intermediates which have not yet been characterized.
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PMID:Biosynthesis of tetrahydrobiopterin by de novo and salvage pathways in adrenal medulla extracts, mammalian cell cultures, and rat brain in vivo. 657 16

Culture of rat pineal glands in methotrexate (0.5, 5, or 10 microM) for 6 or 24 h did not alter pineal tetrahydrobiopterin (85-90% of total biopterin in cultured glands), except for a decrease of 30% after 24 h culture in 10 microM methotrexate. However, pineal dihydrobiopterin and/or biopterin (10-15% of total biopterin) was increased by methotrexate up to 2.5-fold. Biopterin detected in the culture medium following pineal culture was also increased to a similar extent after methotrexate treatment and appeared to represent leakage of pineal dihydrobiopterin and/or biopterin. Culture of glands in 5 microM methotrexate did not alter the conversion of [U-14C]-guanosine to [14C]biopterin, suggesting that pineal tetrahydrobiopterin synthesis was not altered by methotrexate. Complete inhibition of dihydrofolate reductase activity measured in pineal homogenates was obtained following culture of glands in all concentrations of methotrexate studied. Therefore, dihydrofolate reductase and dihydrobiopterin do not appear to be involved in a major biosynthetic pathway for pineal tetrahydrobiopterin from GTP, although they may have a minor role in tetrahydrobiopterin synthesis.
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PMID:Effects of methotrexate on biopterin levels and synthesis in rat cultured pineal glands. 672 34

L1210 cells resistant to 6MP and 6TG exhibit increased sensitivity to MTX compared to the parent line. The differential response of parent and purine analog-resistant cell lines to MTX is not due to host influences, for both L1210/6MP and L1210/6TG cell lines are cross-resistant to 6-MeMPR, an inhibitor of de novo synthesis, and cultured L1210/6MP cells are more sensitive to MTX than the parent cell line. Following treatment of tumor-bearing mice with MTX, the drug concentration in L1210/6TG cells was about 50% greater than in L1210/0 cells for 24 hr and may account, wholly or in part, for the increased sensitivity of the L1210/6TG cell line to MTX. L1210/6MP cells, however, accumulated less MTX than L1210/0 cells, indicating that an equivalent mechanism is not operative in these cells. DHFR activity in L1210/6TG cells was the same as that in L1210/0 cells, but activity in L1210/6MP cells was lower by 60%. Cultured L1210/6MP cells also exhibited a deficiency in DHFR activity as compared to the parent cell line. The sensitivity of the enzyme to MTX was the same for all three cell lines propagated in vivo. Therefore, the increased sensitivity of the L1210/6MP cell line to MTX may be due, in part, to decreased DHFR activity. Significantly lower levels of GTP + GDP and CTP in 6TG-resistant cells than in parent cells 4 hr after the administration of MTX to tumor-bearing mice may be related to the increased MTX sensitivity of these cells. Our results indicate that the observed alterations in drug sensitivity are associated with more than one biochemical change and that these changes are different in the two purine analog-resistant cell lines.
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PMID:Some biochemical characteristics of L1210 cell lines resistant to 6-mercaptopurine and 6-thioguanine and with increased sensitivity to methotrexate. 689 Aug 14


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