SEARCH · PubMed Health

Results for “PYRIMIDINES”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 721 records · Page 40Linked to original sources

PubMed2005

Pyrimidine pathways in health and disease.

Genetic defects involving enzymes essential for pyrimidine nucleotide metabolism have provided new insights into the vital physiological functions of these molecules in addition to nucleic acid synthesis. Such aberrations disrupt the haematological, nervous or mitochondrial systems and can cause adverse reactions to analogue therapy. Regulation of pyrimidine pathways is also known to be disrupted in malignancies. Nine genetic defects have now been identified but only one is currently treatable. Diagnosis is aided by the accumulation of specific metabolites. Recently, progress has been made in understanding the molecular mechanisms underlying inborn errors of pyrimidine metabolism, together with the key clinical issues and the implications for the future development of novel drugs and therapeutic strategies.

Health↗

PubMed1985

Assay of human erythrocyte pyrimidine and deoxypyrimidine 5'-nucleotidase by isocratic reversed-phase high-performance liquid chromatography.

We report a rapid and reproducible assay for activity of human erythrocyte pyrimidine 5'-nucleotidase and deoxypyrimidine 5'-nucleotidase. The nucleotides CMP, UMP, dUMP, dCMP or dTMP are individually incubated 30 min at 37 degrees C with erythrocyte hemolysate and 4 mM magnesium chloride in Tris, pH 7.5. Data are provided for standardization of the reaction with each substrate. Individual nucleoside products are assayed in less than 10 min by reversed-phase high-performance liquid chromatography at 280 nm with 0-14% methanol in 0.01 M potassium dihydrogen phosphate. This is the first report of a high-performance liquid chromatographic assay system which allows quantitation of the activity of pyrimidine 5'-nucleotidase isozymes using five individual pyrimidine and deoxypyrimidine nucleotides as the substrates.

Chromatography, High Pressure Liquid↗

PubMed1988

Site-directed mutagenesis of the T4 endonuclease V gene: role of tyrosine-129 and -131 in pyrimidine dimer-specific binding.

T4 endonuclease V incises DNA at the sites of pyrimidine dimers through a two-step mechanism. These breakage reactions are preceded by the scanning of nontarget DNA and binding to pyrimidine dimers. In analogy to the synthetic tripeptides Lys-Trp-Lys and Lys-Tyr-Lys, which have been shown to be capable of producing single-strand scissions in DNA containing apurinic sites, endonuclease V has the amino acid sequence Trp-Tyr-Lys-Tyr-Tyr (128-132). Site-directed mutagenesis of the endonuclease V gene, denV, was performed at the Tyr-129 and at the Tyr-129 and Tyr-131 positions in order to convert the Tyr residues to nonaromatic amino acids to test their role in dimer-specific binding. The UV survival of repair-deficient (uvrA recA) Escherichia coli cells harboring the denV N-129 construction was dramatically reduced relative to wild-type denV+ cells. The survival of denV N-129,131 cells was indistinguishable from that of the parental strain lacking the denV gene. The mutant endonuclease V proteins were then characterized with regard to (1) dimer-specific nicking activity, (2) apurinic nicking activity, and (3) binding affinity to UV-irradiated DNA. Dimer-specific nicking activity and dimer-specific binding for both denV N-129 and N-129,131 were abolished, while apurinic-specific nicking was substantially retained in denV N-129,131 but was abolished in denV N-129. These results indicate that Tyr-129 and Tyr-131 positions of endonuclease V are at least important in pyrimidine dimer-specific binding and possibly nicking activity.

DNA, Superhelical↗

PubMed1996

Synthesis of non-nucleoside analogs of toyocamycin, sangivamycin, and thiosangivamycin: the effect of certain 4- and 4,6-substituents on the antiviral activity of pyrrolo[2,3-d]pyrimidines.

A number of 4-substituted 7-(ethoxymethyl)- and 7-[(2-methoxyethoxy)methyl]pyrrolo[2,3-d]-pyrimidine-5-carbonitrile and -5-thiocarboxamide derivatives and several 7-substituted 4,6-diaminopyrrolo[2,3-d]pyrimidine-5-carbonitrile, -5-carboxamide, and -5-thiocarboxamide analogs related to the nucleoside antibiotics toyocamycin and sangivamycin were prepared and tested for activity against human cytomegalovirus (HCMV) and herpes simplex virus type 1 (HSV-1). Biologically, modifications at the 4-position were not well tolerated in cell culture, and in almost all cases no activity against HCMV or HSV-1 was observed. Furthermore, none of the compounds inhibited the growth of L1210 murine leukemic cells in vitro. In sharp contrast to the 4-substituted compounds, all of the 4,6-diamino 5-nitrile and the 5-thioamide analogs were active against HCMV, whereas the 5-carboxamides were inactive. The corresponding 4-amino 6-methylamino and 6-dimethylamino 5-nitrile analogs were inactive against HCMV, establishing that an amino group at both C-4 and C-6 is a likely requirement for antiviral activity. Overall, our results demonstrate that an amino group at C-4 and a thioamide moiety at C-5 of a 7-substituted pyrrolo[2,3-d]pyrimidine are essential for activity against HCMV, whereas a 4,6-diamino analog does not necessarily require a thioamide group at C-5 for activity against HCMV.

Antiviral Agents↗

PubMed2025

De novo pyrimidine synthesis is a collateral metabolic vulnerability in NF2-deficient mesothelioma.

Pleural mesothelioma (PM) is one of the deadliest cancers, with limited therapeutic options due to its therapeutically intractable genome, which is characterized by the functional inactivation of tumor suppressor genes (TSGs) and high tumor heterogeneity, including diverse metabolic adaptations. However, the molecular mechanisms underlying these metabolic alterations remain poorly understood, particularly how TSG inactivation rewires tumor metabolism to drive tumorigenesis and create metabolic dependencies. Through integrated multi-omics analysis, we identify for the first time that NF2 loss of function defines a distinct PM subtype characterized by enhanced de novo pyrimidine synthesis, which NF2-deficient PM cells are critically dependent on for sustained proliferation in vitro and in vivo. Mechanistically, NF2 loss activates YAP, a downstream proto-oncogenic transcriptional coactivator in the Hippo signalling pathway, which in turn upregulates CAD and DHODH, key enzymes in the de novo pyrimidine biosynthesis pathway. Our findings provide novel insights into metabolic reprogramming in PM, revealing de novo pyrimidine synthesis as a synthetic lethal vulnerability in NF2-deficient tumors. This work highlights a potential therapeutic strategy for targeting NF2-deficient mesothelioma through metabolic intervention.

Pyrimidines↗

PubMed1992

Role of the basic amino acid cluster and Glu-23 in pyrimidine dimer glycosylase activity of T4 endonuclease V.

T4 endonuclease V [endodeoxyribonuclease (pyrimidine dimer); deoxyribonuclease (pyrimidine dimer), EC 3.1.25.1] initiates repair of damaged DNA by hydrolysis of the N-glycosyl bond at the 5' side of a pyrimidine photodimer in double-stranded DNA. To study one of the active sites of T4 endonuclease V, systematic site-directed mutagenesis was performed on the synthetic T4 endonuclease V gene, in parallel with three-dimensional structure analysis by x-ray crystallography. The mutant proteins were evaluated for DNA glycosylase activity using an oligonucleotide duplex (14-mer) containing a single thymidine dimer as a substrate. Replacement of either Glu-23 with glutamine or asparatic acid or Arg-3 with glutamine completely abolished DNA glycosylase activity. Mutation of Arg-3 to lysine or of Arg-26 to glutamine or lysine in a basic amino acid cluster caused serious defects in DNA glycosylase activity, which are reflected in the increases in Km and decreases in kcat of DNA glycosylase activity. On the other hand, substitutions of lysine for Arg-22 or of glutamine for Arg-117 or Lys-121 resulted in increases in the Km value. The completely inactive mutant proteins, E23Q and R3Q, in which glutamine was substituted for Glu-23 and Arg-3, respectively, were further investigated by CD spectroscopy for their ability to bind the oligonucleotide substrate. It was found that the E23Q protein retained specific substrate-binding ability, whereas the R3Q protein did not. These results indicate that Glu-23 plays an important role in catalysis of the DNA glycosylase reaction, and that Arg-3 is a crucial residue for substrate binding. In addition, Arg-22, Arg-26, Arg-117, and Lys-121 in the basic amino acid cluster also participate in substrate binding. We conclude that the basic amino acid cluster in T4 endonuclease V is an essential structure for DNA glycosylase activity.

Amino Acid Sequence↗

PubMed1997

UV endonuclease of Micrococcus luteus, a cyclobutane pyrimidine dimer-DNA glycosylase/abasic lyase: cloning and characterization of the gene.

The gene of Micrococcus luteus UV endonuclease (cyclobutane pyrimidine dimer-DNA glycosylase/ abasic lyase) was cloned and characterized. The cloned gene, whose product had a predicted molecular mass of 17,120 Da, was found to be capable of complementing the Escherichia coli uvrA6 mutation in vivo with respect to resistance to acetonemediated molecular photosensitization, a treatment producing exclusively cyclobutane pyrimidine dimers in DNA. It also generated a nicking activity specific for photosensitization-treated DNA by in vitro transcription/translation. When expressed in E. coli cells, the gene produced a protein structurally identical with UV endonuclease and possessing an activity consistent with cyclobutane pyrimidine dimer-DNA glycosylase/abasic lyase with respect to the effect of inhibitors and the site of the DNA backbone scission. Furthermore, the UV endonuclease-deficient mutant DB7 was shown to regain the enzyme through transformation with the cloned gene. The deduced amino acid sequence of the gene product was at best 27% identical with that of endonuclease V of phage T4, an enzyme strikingly similar to UV endonuclease in molecular and catalytic properties. Despite this marginal overall similarity in amino acid sequence, four of the seven amino acid residues reported to be functionally important in the T4 enzyme were found to be conserved in the M. luteus enzyme. We propose that the gene be called uveA.

Adenosine Triphosphatases↗

PubMed2003

Compromised incision of oxidized pyrimidines in liver mitochondria of mice deficient in NTH1 and OGG1 glycosylases.

Mitochondrial DNA is constantly exposed to high levels of endogenously produced reactive oxygen species, resulting in elevated levels of oxidative damaged DNA bases. A large spectrum of DNA base alterations can be detected after oxidative stress, and many of these are highly mutagenic. Thus, an efficient repair of these is necessary for survival. Some of the DNA repair pathways involved have been characterized, but others are not yet determined. A DNA repair activity for thymine glycol and other oxidized pyrimidines has been described in mammalian mitochondria, but the nature of the glycosylases involved in this pathway remains unclear. The generation of mouse strains lacking murine thymine glycol-DNA glycosylase (mNTH1) and/or murine 8-oxoguanine-DNA glycosylase (mOGG1), the two major DNA N-glycosylase/apurinic/apyrimidinic (AP) lyases involved in the repair of oxidative base damage in the nucleus, has provided very useful biological model systems for the study of the function of these and other glycosylases in mitochondrial DNA repair. In this study, mouse liver mitochondrial extracts were generated from mNTH1-, mOGG1-, and [mNTH1, mOGG1]-deficient mice to ascertain the role of each of these glycosylases in the repair of oxidized pyrimidine base damage. We also characterized for the first time the incision of various modified bases in mitochondrial extracts from a double-knock-out [mNTH1, mOGG1]-deficient mouse. We show that mNTH1 is responsible for the repair of thymine glycols in mitochondrial DNA, whereas other glycosylase/AP lyases also participate in removing other oxidized pyrimidines, such as 5-hydroxycytosine and 5-hydroxyuracil. We did not detect a backup glycosylase or glycosylase/AP lyase activity for thymine glycol in the mitochondrial mouse extracts.

Animals↗

PubMed1982

Purine and pyrimidine metabolism.

The pathways of purine biosynthesis and degradation have been elucidated during the last 30 years; the regulation of the mechanisms involved is not yet fully understood, particularly with respect to quantitative aspects. Research into inborn errors of purine metabolism has provided valuable insights into purine synthesis and salvage pathways. Nutrition experiments using purine-free formula diets and supplements with defined purine sources permit precise descriptions of the influence of various dietary purines on uric acid formation. Supplements of dietary purines produce dose-proportional increases in plasma uric acid concentrations, uric acid pool size and renal uric acid excretion. The magnitude of these increases depends on the type of purine compound administered, which may limit the value of food tables for human dietetics. Purine content of food must be related not only to weight but also to energy and to protein, particularly if new foodstuffs or a vegetarian diet are ingested. Dietary purines appear to influence the biosynthesis of pyrimidines. In contrast to dietary purines, pyrimidines in the diet, if administered as nucleosides or nucleotides, are utilized in animals for the synthesis of nucleic acids. Much further work is necessary for a better understanding of the inter-relationships of purine and pyrimidine metabolism.

Animals↗

PubMed1988

Variations in excision repair of UVB-induced pyrimidine dimers in DNA of human skin in situ.

The excision repair kinetics of UVB (280-320 nm)*-induced pyrimidine dimers in DNA of human skin in situ was determined for seventeen volunteers using a dimer-specific endonuclease from Micrococcus luteus in conjunction with agarose gel electrophoresis. Removal of pyrimidine dimers from human skin could be detected within 6 h after irradiation and the average half-life for removal of pyrimidine dimers was 11.0 h (+/- 4.3 h). However, there was significant inter-individual variability of repair as indicated by a half-life coefficient of variation of 38%.

Adult↗

PubMed1996

Mutations in Bacillus subtilis PyrR, the pyr regulatory protein, with defects in regulation by pyrimidines.

The pyrimidine nucleotide biosynthetic (pyr) operon in Bacillus subtilis is regulated by a transcriptional attenuation mechanism in which PyrR, a bifunctional pyr RNA-binding attenuation protein/uracil phosphoribosyltransferase, plays a crucial role. A convenient procedure for isolation of pyrR mutants with defects in the regulation of pyr operon expression is described. The selection is based on the selection of spontaneous mutations that convert the pyrimidine-sensitive growth of cpa strain (lacking arginine-repressible carbamyl phosphate synthetase) to pyrimidine resistance. Twelve such mutants were isolated and sequenced. All resulted from point mutations in the pyrR gene.

Amino Acid Sequence↗

PubMed1990

Role of the purine repressor in the regulation of pyrimidine gene expression in Escherichia coli K-12.

The pyrC and pyrD genes of Escherichia coli K-12 encode the pyrimidine biosynthetic enzymes dihydroorotase and dihydroorotate dehydrogenase, respectively. A highly conserved sequence in the promoter regions of these two genes is similar to the pur operator, which is the binding site for the purine repressor (PurR). In this study, we examined the role of PurR in the regulation of pyrC and pyrD expression. Our results show that pyrC and pyrD expression was repressed approximately twofold in cells grown in the presence of adenine [corrected] through a mechanism requiring PurR. A mutation, designated pyrCp926, which alters a 6-base-pair region within the conserved sequence in the pyrC promoter eliminated PurR-mediated repression of pyrC expression. This result indicates that PurR binds to the pyrC (and presumably to the pyrD) conserved sequence and inhibits transcriptional initiation. We also demonstrated that the pyrCp926 mutation had no effect on pyrimidine-mediated regulation of pyrC expression, indicating that pyrimidine and purine effectors act through independent mechanisms to control the expression of the pyrC and pyrD genes.

Amidohydrolases↗

PubMed2000

Rapid screening of high-risk patients for disorders of purine and pyrimidine metabolism using HPLC-electrospray tandem mass spectrometry of liquid urine or urine-soaked filter paper strips.

BACKGROUND: A rapid and specific screening method for patients at risk of inherited disorders of purine and pyrimidine metabolism is desirable because symptoms are varied and nonspecific. The aim of this study was to develop a rapid and specific method for screening with use of liquid urine samples or urine-soaked filter paper strips. METHODS: Reverse-phase HPLC was combined with electrospray ionization (ESI), tandem mass spectrometry (MS/MS), and detection performed by multiple reaction monitoring. Transitions and instrument settings were established for 17 purines or pyrimidines. Stable-isotope-labeled reference compounds were used as internal standards when available. RESULTS: Total analysis time of this method was 15 min, approximately one-third that of conventional HPLC with ultraviolet detection. Recoveries were 96-107% in urine with added analyte, with two exceptions (hypoxanthine, 64%; xanthine, 79%), and 89-110% in urine-soaked filter paper strips, with three exceptions (hypoxanthine, 65%; xanthine, 77%; 5-hydroxymethyluracil, 80%). The expected abnormalities were easily found in samples from patients with purine nucleoside phosphorylase deficiency, ornithine transcarbamylase deficiency, molybdenum cofactor deficiency, adenylosuccinase deficiency, or dihydropyrimidine dehydrogenase deficiency. CONCLUSIONS: HPLC-ESI MS/MS of urine allows rapid screening for disorders of purine and pyrimidine metabolism. The filter paper strips offer the advantage of easy collection, transport, and storage of the urine samples.

Chromatography, High Pressure Liquid↗

PubMed1992

Pyrimidine dimer removal enhanced by DNA repair liposomes reduces the incidence of UV skin cancer in mice.

UV exposure has been linked to skin cancer in humans by epidemiology and the rare genetic disease xeroderma pigmentosum. However, UV produces multiple photoproducts in DNA, and their relative contribution is uncertain. An enzyme which specifically repairs cyclobutane pyrimidine dimers in DNA, T4 endonuclease V, was encapsulated in liposomes for topical delivery into mouse and human skin. In both species, liposomes applied after UV exposure localized in the epidermis and stimulated the removal of cyclobutane pyrimidine dimers. UV-irradiated mice treated with these liposomes had a dose-dependent decrease in the incidence of squamous cell carcinoma compared to controls. The results demonstrate that unrepaired cyclobutane pyrimidine dimers in DNA are a direct cause of cancer in mammalian skin.

Animals↗

PubMed1985

Reaction of l-halogenoses with nitrogen nucleophiles in the presence of tert-amine. Synthesis of 2'-deoxyribofuranosyl and arabinofuranosyl pyrimidines.

Selective introduction of 2'-deoxyribofuranosyl and arabinofuranosyl moieties onto one of two nitrogen atoms in the pyrimidine ring was demonstrated by the reaction of 1-glycosyl chlorides with pyrimidines protected by the (octylthio)carbonyl group in the presence of tert-amine. Replacing pyrimidines by imidazole derivatives such as benzimidazole and theophylline as nucleophiles gave no nucleosides but orthoamide derivatives.

Amines↗

PubMed1981

Salvage of circulating pyrimidine nucleosides in the rat.

A new procedure was developed to measure uridine and cytidine in plasma. These nucleosides are present in micromolar concentrations in the plasma of rats, mice, and humans. Inhibitors of pyrimidine synthesis de novo (pyrazofurin or N-phosphonacetyl-L-aspartate) produce only modest decreases in the concentration of circulating uridine or cytidine in the rat. Since both uridine and cytidine are rapidly cleared from the circulation of the rat, constant infusions of radiolabeled uridine and cytidine were used to establish a steady-state specific activity of circulating nucleoside without altering the normal endogenous concentration. These studies permitted an estimation of the contribution of circulating pyrimidine nucleoside to the nucleotide pools of various rat tissues. Most of the uridine entering the circulation (greater than 70%) is catabolized rather than salvaged by formation of nucleotides. Cytidine in the circulation is much more efficiently utilized and is predominantly salvaged. The implication of these results for chemotherapy based on inhibition of pyrimidine synthesis de novo is discussed.

Amides↗

PubMed1993

Inhibition of 5-phosphoribosyl-1-pyrophosphate synthetase by the monophosphate metabolite of 4-amino-8-(beta-D-ribofuranosylamino)pyrimido[5,4-d]pyrimidine: a novel mechanism for antitumor activity.

The aminopyrimidopyrimidine nucleoside 4-amino-8-(beta-D-ribofuranosylamino)pyrimido[5,4-d]pyrimidine (APP), which was previously shown to possess experimental antitumor and antiviral activity, was metabolized within WI-L2 human lymphoblastoid cells to a derivative identified as the beta-D-ribonucleotide (APP-MP). In a subline of WI-L2 cells deficient in adenosine kinase, this metabolite was not formed and APP was not cytotoxic, suggesting that APP is converted by adenosine kinase to its 5'-monophosphate. Because no evidence of di- or triphosphates was seen, the monophosphate appeared to be the active species. Treatment of WI-L2 or L1210 cells with APP (10 microM) for 30 min caused extensive depletion of both purine and pyrimidine ribonucleotides. Purine and pyrimidine deoxyribonucleotides were also depleted. Cells were not protected from the cytotoxicity of APP by hypoxanthine plus uridine, but uridine plus adenosine plus 2-deoxycoformycin gave considerable protection. This result was consistent with APP-MP acting as an inhibitor of 5-phosphoribosyl-1-pyrophosphate (PRPP) synthetase, a hypothesis that was confirmed by preparing PRPP synthetase from Novikoff hepatoma cells; APP-MP was a noncompetitive inhibitor, with a Ki of 0.43 mM. APP-MP was found to accumulate in APP-treated cells to a concentration of almost 3 mM. The relevance of PRPP synthetase inhibition to the cytotoxic mechanism of APP is indicated by the fact that depletion of the PRPP pool was seen as early as 15 min after treatment, before any change was apparent in cellular levels of ATP or UTP. DNA synthesis was markedly suppressed within 30 min of APP treatment of WI-L2 cells, and a lesser degree of inhibition of RNA synthesis was apparent after 45 min.

Animals↗

PubMed1998

Sunlight and carcinogenesis: expression of p53 and pyrimidine dimers in human skin following UVA I, UVA I + II and solar simulating radiations.

DNA damage by UV radiation plays an essential role in skin cancer induction. We report that even sub-erythemal doses of solar simulating radiation, are capable of inducing substantial nuclear damage, namely pyrimidine dimers and p53 induction in human skin in situ. The quantity and distribution of p53 induced in human skin by UV radiation depended highly on the waveband and dose of UV used. Solar simulating radiation induced very high levels of p53 throughout all layers in epidermal keratinocytes 24 hr following an erythemal dose (230+/-15.9/1000 cells), and the induction followed a dose response. Following UVA I + II and UVA I radiations, p53 expression was approximately half of that seen with equivalent biological doses of solar simulating radiation (63.5+/-28.5 and 103+/-15.9, respectively). Expression of p53 was seen in basal cell keratinocytes at lower doses of UVA, but all layers of the epidermis were affected at higher doses. Pyrimidine dimer induction, however, was seen to be the same for equivalent biological doses of UVA I, UVA I + II and solar simulating radiations, which coincides with previous findings that pyrimidine dimers initiate the erythemal response and are implicated in skin carcinogenesis. When equivalent biological doses of pure UVA are used with no UVB contamination, significant nuclear alterations occur in human skin in situ, which can approach those seen with UVB radiation. Our results suggest that DNA damage assessed in vivo by immunohistochemistry could provide a very sensitive endpoint for determining the efficacy of protective measures, such as sunscreens or protective clothing, against both UVB- and UVA-induced damage in human skin.

Adult↗