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

---

Query: EC:2.4.2.8 (hypoxanthine-guanine phosphoribosyltransferase)
2,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We constructed an expression plasmid (pMAMCRR51) that carried the entire protein-coding sequence of the rabbit cardiac ryanodine receptor cDNA, linked to the dexamethasone-inducible mouse mammary tumor virus promoter and Escherichia coli xanthine-guanine phosphoribosyltransferase (gpt). Chinese hamster ovary (CHO) cells were transfected with pMAMCRR51 and mycophenolic acid-resistant cells showing caffeine-induced intracellular Ca2+ transients were selected. Immunoprecipitation with a monoclonal antibody against the canine cardiac ryanodine receptor revealed that the cell clones thus selected exhibited Ca(2+)-dependent [3H]ryanodine binding activity, which was stimulated by 5 mM ATP or 1 M KCl. The apparent dissociation constant (Kd) for [3H]ryanodine was 6.6 nM in 1 M KCl, which was similar to the Kd obtained with cardiac microsomes. Immunoprecipitation also demonstrated that these cell clones expressed a protein indistinguishable in M(r) from the ryanodine receptor in canine cardiac microsomes. The ryanodine binding activity expressed in CHO cells increased significantly after dexamethasone induction. In saponin-skinned CHO cells transfected with pMAMCRR51, micromolar Ca2+ or millimolar caffeine evoked rapid Ca2+ release from the intracellular Ca2+ stores. In skinned control CHO cells, we did not observe such Ca2+ release activity. These results clearly demonstrate that the cardiac ryanodine receptor is stably expressed in internal membranes of CHO cells and functions as Ca(2+)-induced Ca2+ release channels.
...
PMID:Expression of Ca(2+)-induced Ca2+ release channel activity from cardiac ryanodine receptor cDNA in Chinese hamster ovary cells. 133 83

We describe a highly efficient stable gene transfection procedure for Chinese hamster ovary (CHO) cells using a modification of the calcium phosphate-DNA coprecipitation method. We have found that treatment of CHO cells with chloroquine increases the efficiency of gene transfer by up to 20-fold (from approx. 0.01% to approx. 0.2%) when transfection is done using the pSV2-neo plasmid. The optimized transfection procedure requires that CHO cells to be incubated with calcium phosphate-DNA coprecipitate and chloroquine (100 microM) for a total of 16 h. By using high-molecular-weight human genomic DNA as a DNA source for transfection, we show that this procedure is equally efficient for stably transferring a much larger gene, such as the 49-kb human hypoxanthine phosphoribosyltransferase gene.
...
PMID:High-efficiency stable gene transfection using chloroquine-treated Chinese hamster ovary cells. 176 89

Adenine phosphoribosyltransferase (APRTase) and hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) have been purified from Artemia cysts and nauplii to apparent homogeneity, as determined by SDS-PAGE. The purification includes affinity chromatography on AMP-Sepharose, which binds both enzymes, and they are eluted at different 5-phospho-alpha-D-ribosyl diphosphate (PP-Rib-P) concentrations. The purified enzymes from Artemia cysts were similar to nauplii enzymes with respect to Mr in denaturing gel electrophoresis and gel filtration, pH and cation dependence and kinetic constants for substrates and inhibitors. By Sephadex G-100 filtration, the native Mr of the adenine and hypoxanthine-guanine enzymes was estimated to be Mr 28,000 and 66,000, respectively. Analysis by SDS-PAGE revealed that the APRTase was a dimer of Mr 15,000 sub-units and the HGPRTase, a tetramer of four identical Mr 19,000 sub-units. The pH profile of the HGPRTase shows two apparent buffer-independent pH optima, at 7.0 and 9.5, while the APRTase has just one, at about pH 8-9. The purine phosphoribosyltransferase activity with adenine was highest, about tenfold the HGPRTase activity with hypoxanthine and fivefold that with guanine. Both enzymes exhibited similar requirements for divalent cations, either Mg2+, Mn2+ or Zn2+, while Ca2+ is highly inhibitory. The Km values of APRTase for adenine and PP-Rib-P are 2 and 30 microM, respectively, and the Km values of HGPRTase for hypoxanthine, guanine and PP-Rib-P are less than 1, less than 1 and 15 microM, respectively. Plots of the reciprocal enzyme activities versus reciprocal concentrations of one substrate at several fixed levels of the second one yield a pattern of inhibition by guanine and hypoxanthine. Product-inhibition studies indicated that AMP is a competitive inhibitor with respect to PP-Rib-P in the APRTase reaction, while the HGPRTase shows a mixed inhibition by GMP.
...
PMID:Artemia purine phosphoribosyltransferases. Purification and characterization. 185 Sep 82

The data concerning the mutagenic, clastogenic and carcinogenic properties of inorganic lead compounds have been conflicting. To investigate whether the genotoxicity of lead is due to indirect effects such as interference with DNA-repair processes, the induction of mutations, sister-chromatid exchanges and strand breaks by lead ions alone as well as in combination with UV light as a standard mutagen were determined. Lead acetate alone does not induce DNA-strand breaks in HeLa cells or mutations at the HPRT locus and sister-chromatid exchanges in V79 Chinese hamster cells. However, at all endpoints tested, lead ions interfere with the processing of UV-induced DNA damage. They inhibit the closing of DNA-strand breaks after UV irradiation and enhance the number of UV-induced mutations and sister-chromatid exchanges, indicating an inhibition of DNA repair. These data point out the necessity to consider such indirect effects when assessing the genotoxicity of metal compounds. As possible mechanisms of repair inhibition we suggest either the interaction with repair enzymes such as polymerase or ligase or else the interaction with calcium-regulated processes, for example with calmodulin.
...
PMID:Indirect mechanism of lead-induced genotoxicity in cultured mammalian cells. 233 87

An aryl hydrocarbon hydroxylase (AHH)-deficient gene A- mutant of the mouse line Hepa-1 was treated with calcium phosphate precipitates of DNA from Hepa-1, the rat line H4IIEC3, or an A- -human hybrid in which the A- mutation is complemented by the corresponding human gene. AHH+ transfectants were isolated by selection with benzo[ghi]perylene plus near UV. In addition, a gene A- mutant which also carries a mutation for hypoxanthine phosphoribosyltransferase deficiency was treated with the above genomic DNAs together with pSV2-gpt DNA, and cotransfectants were isolated after treatment with both benzo[ghi]pereylene and HAT. All transfectants and cotransfectants were inducible for AHH by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Both transfectants and cotransfectants were unstable during culture, rapidly losing AHH activity. Rat DNA-derived transfectants were probed in Southern blots with a cDNA probe to mouse cytochrome P1-450 that cross-hybridizes to the corresponding rat gene. All rat DNA-derived transfectants contained the rat P1-450 gene. In half of the transfectants, the rat gene was amplified four- to sevenfold. In one transfectant, the rat gene was truncated at the 3' end. The proportion of rat DNA in different transfectants, as determined by hybridization to a rat repetitive sequence, ranged from less than 1% to 5%. AHH activity and the rat P1-450 gene segregated together in subclones of one of the transfectants. These results demonstrate that the A gene is either the structural gene for cytochrome P1-450, or another very closely linked gene. Previous results (O. Hankinson et al., J. Biol. Chem. 260:1790-1795, 1985) favor the former alternative.
...
PMID:Transfection by genomic DNA of cytochrome P1-450 enzymatic activity and inducibility. 399 Jun 91

PCC3 mouse teratocarcinoma (TCC) stem cells were cotransfected with either the plasmid p delta C-1A or p delta C-1B carrying the chicken delta-crystallin gene, and with the plasmid pSV2gpt containing the selectable bacterial xanthine-guanine phosphoribosyltransferase (XGPRT) gene, using the calcium phosphate technique. Nine transformed PCC3 stem cell lines, each of which was clonally derived from respective colonies surviving after the selection process, were isolated. Southern blot analysis revealed that all of them stably maintained delta-crystallin sequences associated with high mol. wt. cellular DNA after propagation in non-selective medium in vitro, and after the production of solid tumors in the syngenic host mice. Six cell lines contain the intact delta-crystallin gene sequence and eight contain the gpt sequence. The number of delta-crystallin DNA copies was highly variable among transformed lines, 1-500 delta-crystallin genes per diploid mouse genome. No expression of the exogenous genes was detected in the transformed cells as long as they were in the undifferentiated state. However, the synthesis of delta-crystallin in certain types of cells was detected immunohistologically in three lines after the differentiation. The positive cell types were unique to each line, skeletal muscle in Y delta-9, certain columnar epithelia in Y delta-2, and unidentified spindle-shaped cells in Y delta-3. Authentic delta-crystallin polypeptides with a mol. wt. of 48 000 are synthesized upon differentiation of line Y delta-3 in solid tumors in syngenic mice.
...
PMID:Differentiation-dependent expression of the chicken delta-crystallin gene introduced into mouse teratocarcinoma stem cells. 609 52

Chromosome-mediated transfer of genes between human cell lines was accomplished using HeLa cells as chromosome donors and HT1080 fibrosarcoma lines as recipients. This report describes the intraspecific transfer of two genetic markers, hypoxanthine-guanine-phosphoribosyltransferase (HPRT+) and adenine phosphoribosyltransferase (APRT+). The isolation and characterization of the necessary enzyme-deficient (HPRT- and APRT-) recipient HT1080 cell lines are also described. The chromosome-mediated gene transfer was carried out using a modification of the procedure of Miller and Ruddle, including treatment of the donor chromosomes with calcium phosphate and subsequent exposure of the recipient cells of dimethyl sulfoxide. In experiments to optimize this procedure for HT1080 cell recipients, we found that a brief (2-min) exposure to high DMSO concentration (20%) was effective for enhancing transfer efficiencies in this system. Transfer frequencies (transferents per recipient cells assayed) averaged approximately 1 x 10(-6) for HPRT+ and were greater than 2 x 10(-6) for APRT+.
...
PMID:Chromosome-mediated gene transfer of HPRT and APRT in an intraspecific human cell system. 683 54

Human DNA purified from HeLa cells and from three strains of skin fibroblasts was precipitated with calcium phosphate and added to mouse cells that were deficient in adenine phosphoribosyltransferase (APRT) and hypoxanthine phosphoribosyltransferase (HPRT). Selection for cells possessing either of the phosphoribosyltransferases was imposed by blocking de novo synthesis of purine nucleotides with azaserine in a medium supplemented with adenine and hypoxanthine. The frequency of colony formation after selection was 1.7 x 10(-7)-3.3 x 10(-6). Excepting some azaserine-resistant colonies that appeared only in the first experiment and infrequent revertants expressing moust APRT, all characterized clones expressed the human forms of APRT or HPRT according to the criteria of specific immunoprecipitation and electrophoretic mobility. The frequency of transfer of the human APRT gene was much greater than that of HPRT. Transfer efficiency was not significantly reduced when HeLa DNA was sheared to 6.5-13.5 kb size or when the donor DNA was isolated from a transferent that expressed human APRT.
...
PMID:Expression of human genes for adenine phosphoribosyltransferase and hypoxanthine-guanine phosphoribosyltransferase after genetic transformation of mouse cells with purified human DNA. 699 64

Thioguanine resistant CHO cells (HPRT-) were stably cotransfected with pSV2-gpt and pi H3-CD2 vectors using the calcium phosphate coprecipitation technique. The effects of single low doses of ionizing radiation were studied in a CD2+ CHO clone. The CD2+ phenotype responsible for binding sheep erythrocytes and rosette formation, was not affected by X-rays doses in the range 2-6 cGy. However, after 10 cGy of X-irradiation, 50% of the cells lost the CD2+ phenotype. These results suggest that this CD2+ clone might be a very sensitive indicator of very low X-ray doses. The implications of the phenotypic changes, observed after very low doses of irradiation, are discussed.
...
PMID:Effects of low-doses of X-rays on the expression of human cell surface CD2 antigen in CD2+ CHO cells. 790 2

Inhibition of poly(ADP-ribosylation) reduces random genomic integration of transfected DNA and mildly stimulates intrachromosomal homologous recombination in mammalian cells. We investigated the effect of inhibition of poly(ADP-ribosylation) on the efficiency of gene targeting in Chinese hamster ovary (CHO) cell line ATS-49tg. This cell line is hemizygous for a defective adenine phosphoribosyltransferase (aprt) gene and is hypoxanthine phosphoribosyltransferase (hprt) deficient. Plasmid pAG100 contains a portion of the CHO aprt gene sufficient to correct the defect in ATS-49tg cells via gene targeting; pAG100 also contains an Escherichia coli guanine phosphoribosyltransferase (gpt) gene. Following transfection of ATS-49tg cells with pAG100, selection for gpt-positive transfectants allowed recovery of cells that had randomly integrated pAG100 while selection for aprt-positive cells allowed recovery of cells that had undergone gene targeting at the endogenous aprt locus. Treatment of cells with 3 mM 3-methoxybenzamide (3-MB), an inhibitor of poly(ADP-ribose) polymerase, decreased random integration and gene targeting of electroporated pAG100 about 5-fold. In contrast, treatment with 3 mM 3-MB during calcium phosphate transfection could reduce random integration more than 150-fold while reducing gene targeting less than two-fold. Therefore, as much as a 100-fold enrichment for gene targeting was achieved with calcium phosphate transfection.
...
PMID:Enrichment for gene targeting in mammalian cells by inhibition of poly(ADP-ribosylation). 880 16


1 2 Next >>

---