UNIPROT:P00492 - FACTA Search

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Query: UNIPROT:P00492 (hypoxanthine-guanine phosphoribosyltransferase)
2,385 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The growth inhibitory activity of 3-deazaguanosine toward a mutant line (TGR-3) of Chinese hamster ovary cells deficient in hypoxanthine-guanine phosphoribosyltransferase (EC 2.4.2.8) was substantially reversed by the simultaneous addition of nicotinamide riboside. The activities of most other ribonucleoside analogues tested were unaffected. The formation of cellular 3-deazaGMP and 3-deazaGTP from the ribonucleoside analogue, as measured by high-pressure liquid chromatography, was inhibited by the presence of nicotinamide riboside. The inhibition was dependent on concentration of 3-deazaguanosine and could also be demonstrated by following the metabolism of 3-deazaguanosine, labeled with 14C in the ribose moiety, to [14C]3-deazaGTP. In the presence of 100 microM nicotinamide riboside formation of the labeled triphosphate derivative of 3-deazaguanosine was undetectable. A 3-deazaguanosine phosphorylating activity was separated from other cellular kinases by DEAE-cellulose chromatography. Contaminating purine nucleoside phosphorylase (EC 2.4.2.1) was subsequently removed by sucrose density gradient centrifugation. The resulting enzyme preparation demonstrated the greatest activities with nicotinamide riboside and 3-deazaguanosine and, in addition, could also phosphorylate tiazofurin and guanosine to lesser, but significant, degrees. These and other observations suggest that 3-deazaguanosine, and perhaps other agents such as tiazofurin, may, at least in part, be phosphorylated by a nicotinamide ribonucleoside kinase in these cells. If so, it is possible that the activity of this agent in other types of cells in vivo could be dependent upon the presence of this enzyme and that it could be influenced by cellular concentrations of the natural pyridine nucleoside.
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PMID:Phosphorylation of 3-deazaguanosine by nicotinamide riboside kinase in Chinese hamster ovary cells. 255 47

A series of compounds isolated on the basis of their mutagenicity in the Ames/Salmonella reversion assay were previously identified in fried beef and chemically synthesized for further evaluation. In this study three of these compounds were tested for genotoxic effects in the UV5 line of Chinese hamster ovary (CHO) cells, which is deficient in nucleotide excision repair. Both 2-amino-3,4-dimethyl-imidazo]4,5-f]quinoline (MeIQ) and 2-amino-3,8-dimethyl-imidazo[4,5-f]quinoxaline (MeIQx) gave very weak responses for cell killing, hprt mutation induction and sister chromatid exchange. These effects occurred at doses in the range of 100-800 micrograms/ml (approximately solubility limit), and dose-dependent increases were not observed. Induction of chromosomal aberrations did not occur with either compound. Nor did either of these compounds produce differential cytotoxicity in normal CHO cells versus UV5 cells, indicating that potentially repairable DNA damage was not responsible for the observed cell killing. In contrast to these results, 2-amino-1-methyl-6-phenylimidazo [4,5-b]pyridine (PhIP), which constitutes greater than 90% of the mass of bacterial mutagens in beef, was strongly positive for all endpoints at doses in the range 1-3 micrograms/ml. PhIP also gave marked differential cytotoxicity (ratio of 6) and cell survival curves that were strongly dependent on repair capacity. Because PhIP is 50- to 300-fold less mutagenic than MeIQ and MeIQx in Salmonella TA1538, these results point to major differences between the bacterial and mammalian assays in terms of the relative potency of these food-related compounds.
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PMID:Genotoxicity of compounds from cooked beef in repair-deficient CHO cells versus Salmonella mutagenicity. 332 38

2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), a heterocyclic aromatic amine that is formed in abundance in cooked meats, has been found to be mutagenic in human lymphoblastoid TK6 cells at the thymidine kinase and hypoxanthine-guanine phosphoribosyl transferase (hgprt) loci. The mutations induced at the hgprt locus have been analysed. Of the mutations that have been identified, 60% were found in the coding sequence of the gene. Forty percent were in the introns which resulted in aberrant splicing and consequently, leading to exon losses in the mature hprt mRNA. Mutations resulting in a loss of exonIII appeared most frequently followed by losses of exonVI, exonVIII and partial loss of exonIX. All identified mutations occurred at GC base pairs, consistent with the adducts of PhIP that have been found previously and suggesting that the N-(deoxyguanosin-8-yl)-2-amino-1-methyl-6-phenylimidazo[4,5- b]pyridine, (dG-C8-PhIP) adduct may be the premutagenic lesion. Most of the mutations are GC-->TA transversions except for a cluster of single base pair deletions in a run of guanines. There appears to be strand bias in the induction of mutations with 85% of the mutations on the non-transcribed strand. Although the number of mutations analysed is limited (54 mutants), there are several sites (positions 166 and 207 of the coding sequence, and the splice acceptor site of exonIII) which are overrepresented. There is a preference for a 5' purine but not a strong bias for 3' A as has been found for other mutagens that form a premutagenic lesion at G. Triplet analysis shows that the triplets, 5'GGA3' and 5'AGG3', where the middle base is mutated are preferred.
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PMID:Analysis of mutations induced by 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in human lymphoblastoid cells. 772 48

Phosphoribosyltransferases (PRTases) are enzymes involved in the synthesis of purine, pyrimidine, and pyridine nucleotides. They utilize alpha-D-5-phosphoribosyl-1-pyrophosphate (PRPP) and a nitrogenous base to form a beta-N-riboside monophosphate and pyrophosphate (PPi), and their functional significance in nucleotide homeostasis is evidenced by the devastating effects of inherited diseases associated with the decreased activity and/or stability of these enzymes. The 2.6-A structure of the Salmonella typhimurium orotate phosphoribosyltransferase (OPRTase) complexed with its product orotidine monophosphate (OMP) provides the first detailed image of a member of this group of enzymes. The OPRTase three-dimensional structure was solved using multiple isomorphous replacement methods and reveals two major features: a core five-stranded alpha/beta twisted sheet and an N-terminal region that partially covers the C-terminal portion of the core. PRTases show a very high degree of base specificity. In OPRTase, this is determined by steric constraints and the position of hydrogen bond donors/acceptors of a solvent-inaccessible crevice where the orotate ring of bound OMP resides. Crystalline OPRTase is a dimer, with catalytically important residues from each subunit available to the neighboring subunit, suggesting that oligomerization is necessary for its activity. On the basis of the presence of a common PRPP binding motif among PRTases and the similar chemistry these enzymes perform, we propose that the alpha/beta core found in OPRTase will represent a common feature for PRTases. This generality is demonstrated by construction of a model of the human hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) from secondary structure predictions for HGPRTase and the three-dimensional structure of OPRTase.
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PMID:Crystal structure of orotate phosphoribosyltransferase. 831 45

The mutagenic 'fingerprint' of the cooked food carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) was determined in a Chinese hamster cell line genetically engineered to express human CYP1A2 (XEMh1A2-MZ). The parental Chinese hamster V79 and XEMh1A2-MZ cells were exposed to PhIP at various concentrations for 24h. There was a dose-dependent increase in frequency of mutations at the hypoxanthine-guanine phosphoribosyltransferase (hprt) locus only in the metabolically competent XEMh1A2-MZ cells. The mutant frequency ranged from 25 to 90 X 10(-6) with final concentrations of 2.5 to 100 microM PhIP compared to 8 X 10(-6) in the solvent controls and the V79MZ cells. The molecular nature of the PhIP-induced mutations in XEMh1A2-MZ cells was determined by examining DNA sequence modifications at the hprt locus in forty five 6-thioguanine resistant (6-TGr) mutant clones. Single base substitutions predominantly GC-->TA transversions, were the major class of PhIP-induced mutation. However, a -1 frameshift 'hotspot' in a 5'-GGGA sequence was also observed. With the exception of a compound modification, all of the PhIP-induced mutations involved G.C base pairs. This is consistent with the previously observed PhIP-induced mutations in cultured mammalian cells and 32P-postlabelling experiments that show PhIP adducts to the guanine base and that major adduct is at the C8 position. Furthermore, nearly all of these mutations involved guanine bases on the non-transcribed strand which is possibly indicative of preferential repair of PhIP adducts from the transcribed strand. Nearest neighbor analysis of induced base substitutions indicates a preference for 5' guanine and 3' adenine. These data effectively define a mutation 'fingerprint' for PhIP, which may provide the basis for definitive studies on the role of PhIP in diet associated cancers such as tumours of colon. It is, therefore, intriguing that in their recent report of mutation in tumours of the colon induced by PhIP in male rate Kakiuchi et al. (Proc. Natl Acad. Sci. USA, 92, 910-914) report that four out of eight tumors had identical mutation of the tumour suppressor gene apc which is comprised of a -1 G frameshift in a 5'-GGGA sequence.
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PMID:Mutational spectra of the dietary carcinogen 2-amino-1-methyl-6- phenylimidazo[4,5-b]pyridine(PhIP) at the Chinese hamsters hprt locus. 862 68

The newborn mouse tumorigenicity assay, which involves the treatment of animals during the first two weeks after birth and monitoring tumor induction after a year, has been suggested as a cost- and time-effective alternative to the conventional two-year rodent bioassay. In order to evaluate whether or not lymphocyte hprt mutant induction is an accurate predictor of carcinogenicity in the assay, we determined the frequencies of 6-thioguanine-resistant (TGr) lymphocytes in the spleens of mice neonatally treated with the carcinogenic mutagens N-ethyl-N-nitrosourea (ENU), dimethylnitrosamine (DMN), and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP). Male C57BL/6 pups were injected on postnatal days 8 and 15, and the frequency of TGr T-lymphocytes was measured in groups of three animals, sacrificed periodically up to 31 weeks post-treatment. Compared to background frequencies of 1.1-2.9 x 10(-6), mutant frequencies (MFS) reached 155.1 x 10(-6) following a cumulative dose of 49 mg ENU/kg body weight and 172.3 x 10(-6) following a cumulative dose of 142 mg ENU/kg. These results show that TGr lymphocyte mutations can be induced and measured in mice treated as neonates and that the induced MFs found for mice treated neonatally with ENU are comparable with frequencies reported for the treatment of adult animals with the same chemical. In contrast, treatment with the promutagenic and procarcinogenic compounds DMN (at a maximum concentration of 10.5 mg/kg) and PhIP (26.2 mg/kg) did not result in an increase in lymphocyte MF, suggesting that reactive metabolites of these compounds may not be reaching cells that are sensitive for mutation fixation. The results indicate that the lymphocyte hprt assay may fail to predict the carcinogenicity of some test chemicals in the neonatal mouse bioassay.
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PMID:Mutational response at the splenic T-lymphocyte hprt locus in mice treated as neonates: contrasting effects of the carcinogens N-ethyl-N-nitrosourea, dimethylnitrosamine, and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine. 958 62

Purine and pyridine metabolism were studied in ten Lesch-Nyhan patients, with virtually no hypoxanthine-guanine phosphoribosyltransferase (HPRT) activity in erythrocytes. Increased NAD erythrocyte concentrations were found in all patients. Raised activities of two enzymes catalysing NAD synthesis from nicotinic acid (nicotinic acid phosphoribosyltransferase: NAPRT, and NAD synthetase: NADs) was found in erythrocyte lysates from all patients. The two enzymes had normal apparent Km for their substrates and increased Vmax. The rate of synthesis of pyridine nucleotides from nicotinic acid by intact erythrocytes in vitro was also increased in most patients. These findings suggest that raised NAD concentrations in HPRT- erythrocytes are due to enhanced synthesis as a result of increased enzyme activities.
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PMID:Hypoxanthine-guanine phosphoribosyltransferase deficiency and erythrocyte synthesis of pyridine coenzymes. 1040 7

Mutation frequencies (MnFs) of the lacI transgene and mutation rates (MRs) of the endogenous hprt gene were analyzed in two mammary carcinoma cell lines that we established from mammary carcinomas that had been induced by 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in female lacI-transgenic rats. Using the lacI transgene, corrected MnF, which is the number of independent lacI mutations that occurred while 102 cells expanded into 10(7) cells and which reflect the dynamic increase of point mutations, was measured. The corrected MnFs in the two mammary carcinoma cell lines (59 x 10(-6) and 72 x 10(-6) mutations) were significantly higher than that in the primary culture of normal mammary epithelium (4.7 x 10(-6)). MRs of the hprt gene in the two mammary carcinoma cell lines (8.2 x 10(-7) and 11 x 10(-7) mutations/hprt/cell division) were also higher than the same control (1.4 x 10(-7)). A:T to C:G transversion was observed at significantly higher frequencies in the two cell lines (6 of 24 and 6 of 25 for lacI; 10 of 67 and 19 of 92 for hprt) than in the control (0 of 6 for lacI; 0 of 4 for hprt). Taking advantage of the lacI transgene, high frequencies of A:T to C:G transversion (6 of 38 and 8 of 33, respectively) was also confirmed in the primary carcinomas of the two cell lines, which indicated the presence of a common abnormality in the cell lines and in the primary carcinomas. Both the established cell lines and their primary carcinomas were negative for microsatellite instability, which is known to be caused mainly by mismatch repair insufficiency and to increase point mutations, and for p53 mutations. These findings showed that the two cell lines, and possibly their primary carcinomas, had increases in the MRs of point mutations attributable to a mechanism(s) different from mismatch repair insufficiency, and we would suggest that such a state be designated as single nucleotide instability (SNI).
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PMID:Single nucleotide instability without microsatellite instability in rat mammary carcinomas. 1128 41

Ten heterocyclic amines (HCAs) that are produced by heating amino acids, proteins, or proteinaceous food such as fish and meat were examined for carcinogenicity in rats and mice. Three of them, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), have been shown to induce mammary cancer in female F344 and/or SD rats, but none of the HCAs induced mammary cancer in CDF(1) mice. This report reviews our recent studies on mammary carcinogenesis of PhIP in various strains of mice and on the roles of genomic instability in the rat mammary carcinogenesis of PhIP. We demonstrated that the survival time from mammary adenocarcinomas was shorter in PhIP-treated BALB/c mice than that in the untreated control, and with a significantly higher incidence in the C.B-17 strain of mice compared with that of the control. To clarify mechanisms of mammary carcinogenesis, we examined genomic instability in rat mammary cancer induced by PhIP. Mammary cancers were induced in F344 x SD F(1) rats harboring the lacI transgene, and two cell lines were established from two adenocarcinomas. They showed a greater than 10-fold higher frequency of spontaneous mutations than that of the primary culture of normal mammary epithelial cells, in the lacI transgene and the hprt endogenous gene during cell replication. Nucleotide sequencing revealed that almost all types of mutations were increased, with a remarkable increase of A:T --> C:G mutation. This genomic instability was not attributed either to alterations of mismatch-repair enzymes or to p53. These mutational characteristics were also observed in the original tumors. Single-nucleotide instability (SNI) might be implicated in the mammary cancer induced by PhIP.
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PMID:Studies on mammary carcinogenesis induced by a heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine, in mice and rats. 1192 Nov 84

The relationship between a complete deficiency of the purine enzyme hypoxanthine-guanine phosphoribosyltransferase and the neurobehavioural abnormalities in Lesch-Nyhan disease remains an enigma. In vitro studies using lymphoblasts or fibroblasts have evaluated purine and pyrimidine metabolism with conflicting results. This study focused on pyridine nucleotide metabolism in control and Lesch-Nyhan fibroblasts using radiolabelled salvage precursors to couple the extent of uptake with endocellular nucleotide concentrations. The novel finding, highlighted by specific culture conditions, was a marked NAD depletion in Lesch-Nyhan fibroblasts. ATP and GTP were also 50% of the control, as reported in lymphoblasts. A 6-fold greater incorporation of [(14)C]nicotinic acid into nicotinic acid- adenine dinucleotide by Lesch-Nyhan fibroblasts, with no unmetabolized substrate (20% in controls), supported disturbed pyridine metabolism, NAD depletion being related to utilization by poly(ADP-ribose) polymerase in DNA repair. Although pyrimidine nucleotide concentrations were similar to controls, Lesch-Nyhan cells showed reduced [(14)C]cytidine/uridine salvage into UDP sugars. Incorporation of [(14)C]uridine into CTP by both was minimal, with more than 50% [(14)C]cytidine metabolized to UTP, indicating that fibroblasts, unlike lymphoblasts, lack active CTP synthetase, but possess cytidine deaminase. Restricted culture conditions may be neccesary to mimic the situation in human brain cells at an early developmental stage. Cell type may be equally important. NAD plus ATP depletion in developing brain could restrict DNA repair, leading to neuronal damage/loss by apoptosis, and, with GTP depletion, affect neurotransmitter synthesis and basal ganglia dopaminergic neuronal systems. Thus aberrant pyridine nucleotide metabolism could play a vital role in the pathophysiology of Lesch-Nyhan disease.
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PMID:Severe pyridine nucleotide depletion in fibroblasts from Lesch-Nyhan patients. 1199 69

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