Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.4.2.7 (adenine phosphoribosyltransferase)
 692 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have studied the mechanism of targeted recombination in mammalian cells using a hemizygous adenine phosphoribosyltransferase-deficient (APRT-) Chinese hamster ovary (CHO) cell mutant as a recipient. Three structurally different targeting vectors with a 5' or a 3', or both, end-deleted aprt sequence, in either a closed-circular or linear form, were transfected to the cells with a mutated aprt gene by electroporation. APRT-positive (APRT+) recombinant clones were selected and analyzed to study the gene correction events of the deletion mutation. Some half of 58 recombinant clones obtained resulted from corrections of the deleted chromosomal aprt gene by either gene replacement or gene insertion, a mechanism which is currently accepted for homologous recombination in mammalian cells. However, the chromosomal sequence in the remaining half of the recombinants remained uncorrected but their truncated end of the aprt gene in the incoming vectors was corrected by extending the end beyond the region of homology to the target locus; the corrected vector was then randomly integrated into the genome. This extension, termed end extension repair, was observed with all three vectors used and was as far as 4.6-kilobase (kb) or more long. It is evident that the novel repair reaction mediated by homologous recombination, in addition to gene replacement and gene insertion, is also involved in gene correction events in mammalian cells. We discuss the model which may account for this phenomenon.
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PMID:End extension repair of introduced targeting vectors mediated by homologous recombination in mammalian cells. 140 93

The measurement of the activity of the X-linked enzyme HPRT has been widely used as an indicator of X-chromosome activity during preimplantation development in the mouse. More recently, the concomitant measurement of the activity of the autosomally-encoded enzyme APRT has been used in an attempt to decrease the variability inherent in the measurement of enzyme activity from minute samples such as preimplantation embryos. In this study the use of the HPRT-deficient mouse mutant, Hprtb-m3, allowed the unequivocal identification of the parental origin of HPRT activity measured in embryos derived from crosses between wild-type mice, and mice which were homozygous or hemizygous for the Hprtb-m3 allele. Results were similar to those of a previous study, where oocyte-encoded HPRT activity accounted for about 10% of total HPRT activity at 76 hours post human chorionic gonadotrophin injection and the paternally-derived Hprt allele was shown to be transcriptionally active by the late 2-cell stage. In contrast to other studies, differential expression of the two Hprt alleles was detected during the preimplantation period, in embryos derived from crosses between wild-type and HPRT-deficient mice. Evidence was also found for the existence of an X-linked locus which influences the amount of APRT activity in the unfertilized oocyte. We propose that the expression pattern of this locus may be influenced by its parental origin.
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PMID:Imprinting of phosphoribosyltransferases during preimplantation development of the mouse mutant, Hprtb-m3. 145 55

Two APRT- clones (V79-E3 and V79-E1A) were isolated from V79 hamster fibroblasts treated with ethyl methanesulfonate. Selection involved sequential exposure of the mutagenized cells to the adenine analogues 8-azaadenine and 2,6-diaminopurine. To examine the influence of APRT deficiency on cell metabolism we determined the size and turnover of adenine ribonucleotide pools, the deoxyribonucleoside triphosphate pools, the rate of DNA synthesis, and the length of the cell cycle. Clone V79-E3 was hemizygous for aprt and carried a new chromosome, 3p-. Clone V79-E1A was quasi-tetraploid with a cell volume more than twice that of the WT cells. When the difference in size was taken into account, both clones behaved similarly. While WT V79 cells released no adenine into the medium, they excreted adenine at a rate of 6 pmol/min. This did not affect the size of the ATP pool. The main change in the deoxynucleotide pools was a marked decrease of the concentration of dCTP. The rate of DNA synthesis was the same in WT cells and in the diploid V79-E3 clone. APRT is known to recycle adenine produced during polyamine synthesis, but the enzyme apparently contributes little to the maintenance of adenine ribonucleotide pools of V79 fibroblasts.
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PMID:Metabolic consequences of adenine-phosphoribosyl transferase deficiency in V79 hamster fibroblasts. 145 99

Five purine auxotrophic mutants of Lactococcus lactis were isolated. L. lactis was capable of converting adenine, guanine and hypoxanthine to AMP, GMP and IMP, respectively, indicating the existence of adenine phosphoribosyltransferase (APRT) and hypoxanthine guanine phosphoribosyltransferase (HGPRT) activities. A 1.3 kb DNA fragment from L. lactis was cloned by complementation of the hpt mutation in Escherichia coli. Introduction of this fragment into L. lactis resulted in an increase in HGPRT activity. In vitro transcription and translation analysis showed that the fragment coded for a polypeptide with M(r) of 22,000. The nucleotide sequence of this hpt gene was determined.
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PMID:Isolation of purine auxotrophic mutants of Lactococcus lactis and characterization of the gene hpt encoding hypoxanthine guanine phosphoribosyltransferase. 146 8

A series of clones displaying high frequency "switching" phenotypes for expression of the adenine phosphoribosyltransferase (aprt) gene were previously isolated from the P19 mouse embryonal carcinoma stem cell line. Most clones contained only one aprt allele. We report here the characterization of each of these clones with regards to enzymatic activity, mRNA steady state levels, DNA methylation, and chromatin conformation. When clones were selected for resistance to the purine analog 2,6-diaminopurine, which requires markedly reduced levels of APRT enzymatic activity, two distinct classes were observed. The first class was associated with reduced or undetectable levels of aprt mRNA, hypermethylation of the 5' CpG island, and a closed chromatin conformation within this region. When clones of this class were selected for reacquisition of APRT enzymatic activity they were found to have increased mRNA levels, a hypomethylated CpG island, and an open chromatin conformation. In contrast, the second class of clones displayed wild-type levels of mRNA, CpG island hypomethylation, and an open chromatin conformation regardless of whether they were selected for the presence or absence of APRT enzymatic activity. The implications of these results for general mechanisms of epigenetic change in somatic cells and the possibility that expression of the mouse aprt gene may be developmentally regulated are discussed.
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PMID:At least two distinct epigenetic mechanisms are correlated with high-frequency "switching" for APRT phenotypic expression in mouse embryonal carcinoma stem cells. 149 18

Growth factors induce the sequential expression of cellular genes whose products are thought to mediate long-term responses to the growth factors. In mouse 3T3 fibroblastic cells, the first genes to be expressed (immediate-early genes) are activated within minutes after the addition of platelet-derived growth factor, fibroblast growth factor, or serum. By cDNA cloning, we have identified genes that are activated after a delay of a few hours and several hours prior to serum-induced DNA replication. Activation of these delayed early response genes requires new protein synthesis, presumably the synthesis of immediate-early transcription factors described previously. Partial or complete sequencing of 13 different delayed early cDNAs, representing about 40% of the 650 primary cDNA isolates, revealed that 8 were related to known gene sequences and 5 were not. Among the former are cDNAs encoding nonhistone chromosomal proteins [HMGI(Y) and HMGI-C], adenine phosphoribosyltransferase (APRT), a protein related to human macrophage migration inhibitory factor (MIF), a protein of the major intrinsic protein (MIP) family homologous to the integral membrane protein of human erythrocytes, and cyclin CYL1. In 3T3 cells, the delayed early gene response to growth factors appears to be at least as complex as the immediate-early gene response previously described.
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PMID:Growth factor-induced delayed early response genes. 150 93

A 48-year-old man with a history of recurrent urolithiasis and chronic renal failure underwent a nephrectomy for a renal mass. At surgery the mass proved to be a calculus impacted in a dilated calyx. Gross examination of the kidney revealed chalky white deposits in the deep medulla and papillary tips. Histologic examination revealed chronic interstitial nephritis with brown spicules within some tubular epithelial cells and larger deposits of brown crystals within tubular lumina, the interstitium of the medulla, and papillary tips. Polarization microscopy revealed individual crystals scattered throughout the renal parenchyma. Although the arrangement of the crystals was reminiscent of uric acid, and, in fact, a clinical diagnosis of gouty nephropathy was made, x-ray diffraction analysis demonstrated crystals of 2,8-dihydroxyadenine. Enzymatic studies confirmed the complete absence of adenine phosphoribosyltransferase activity in erythrocyte lysates.
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PMID:Renal insufficiency secondary to 2,8-dihydroxyadenine urolithiasis. 151 30

We have examined the clonal variation in rates of amino acid transport, protein synthesis, protein degradation, growth and proliferation for CHO cells with mutations in the purine and pyrimidine salvage pathways. First we compared three clonal cell lines, each with a different mutation, with the heterozygous parental line AT3-2. Overall, the correlation between rates of protein turnover, growth and proliferation was excellent. The slower growth and proliferation of one mutant, AB3 (TK-, APRT-), is explained by a low intrinsic rate of protein synthesis coupled with a smaller response in rates of amino acid transport, protein synthesis and protein degradation to insulin, serum and dexamethasone. Secondly, we compared seven aza-adenine-resistant and 14 thioguanine-resistant mutants of AT3-2 and found significant differences in control and insulin-stimulated rates of protein turnover both within and between mutant populations. A significant difference between the populations was unexpected because each individual cell line was cloned from a spontaneous pre-existing mutation in AT3-2, and each population should have the same average rate. Remarkably, all 24 mutants had lower rates of protein synthesis than AT3-2. We cannot explain the data solely in terms of mutations in the salvage pathways. Rather, we propose that the mutant survivors have randomly down-regulated the intrinsically fixed growth factor-regulated pathways of protein turnover, resulting in a broad spectrum of lower metabolic rates.
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PMID:Protein turnover, growth and proliferation in CHO cells. Variation within and between mutant classes for salvage pathway enzymes. 154 Jan 46

An intact cDNA from Arabidopsis thaliana for adenine phosphoribosyltransferase (APRT) was isolated and sequenced. The cDNA is 729 nucleotides in length and predicts a protein of Mr 27,140. The deduced amino acid sequence has been compared with those of other APRTs and shown to be most similar to the Escherichia coli protein. Construction of a molecular tree of the known APRT amino acid sequences indicates the A. thaliana and E. coli APRT sequences form one cluster and the currently available vertebrate and invertebrate sequences form a separate grouping. Since it is possible to select either for or against the expression of APRT, the isolation of this APRT cDNA clone will allow these selection schemes to be used in plant genetic experiments.
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PMID:A complete cDNA for adenine phosphoribosyltransferase from Arabidopsis thaliana. 155 43

Compared with other purine salvage and nitrogen catabolism enzymatic activities, adenine deaminase (adenine aminohydrolase [AAH]; EC 3.5.4.2) activity in Saccharomyces cerevisiae is uniquely regulated. AAH specific activity is not induced by adenine and is reduced sevenfold when cells are cultivated in medium containing proline in place of ammonium as the sole nitrogen source. Exogenous adenine enters metabolic pathways primarily via the function of either AAH or adenine phosphoribosyltransferase (APRT; EC 2.4.2.7). Exogenous adenosine cannot normally be utilized as a purine source. Strains efficiently utilized adenosine or inosine when grown in pH 4.5 medium containing Triton X-100. A recessive mutation permitting utilization of adenosine or inosine in standard media was isolated. In both situations, growth of purine auxotrophs required either AAH or APRT activity. With medium containing either ammonium or proline as a nitrogen source, minimum doubling times of purine auxotrophs deficient in either APRT or AAH were measured. In proline-based medium, AAH and APRT permitted equal utilization of exogenous adenine. In ammonium-based medium, the absence of APRT increased the minimum doubling time by 50%. Similar experiments using sufficient exogenous histidine to feedback inhibit histidine biosynthesis failed to affect the growth rates of adenine auxotrophs blocked in AAH or APRT, indicating that the histidine-biosynthetic pathway does not play a significant role in adenine utilization. The gene that encodes AAH in S. cerevisiae was isolated by complementation using yeast strain XD1-1, which is deficient in AAH, APRT, and purine synthesis. A 1.36-kb EcoRI-SphI fragment was demonstrated to contain the structural gene for AAH by expressing this DNA in Escherichia coli under control of the trp promoter-operator. Northern (RNA) studies using the AAH-, APRT-, and CDC3-coding regions indicated that AAH regulation was not mediated at the level of transcription or mRNA degradation.
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PMID:Adenine deaminase and adenine utilization in Saccharomyces cerevisiae. 157 82


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