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

The hypothesis was tested that the increased IMP dehydrogenase activity in human myelocytic leukemic cells, and along with it guanylate biosynthesis, might be a sensitive target to chemotherapy by tiazofurin. 1. IMP dehydrogenase activity in normal leukocytes was 3.1 +/- 0.5 (means +/- S.E.) nmol/hr/mg protein and in leukemic cells it was elevated 15- to 41-fold. The activity of guanine phosphoribosyltransferase in normal leukocytes was 389 +/- 27 nmol/hr/mg protein and in the leukemic cells it increased 2.8- to 6.8-fold. 2. IMP dehydrogenase was purified 4,900-fold to homogeneity from rat hepatoma 3924A with a yield of 30%. The kinetic properties of the hepatoma enzyme were similar to those of the enzyme in human myelocytic leukemic blast cells because of the similarity of the Km's for IMP (23 microM), NAD (44 and 65 microM); the Ki for TAD was 0.1 microM in both enzymes. 3. There was a selectivity of the in vitro response to tiazofurin in human normal and leukemic leukocytes. When labeled tiazofurin was incubated with leukocytes from normal, healthy volunteers and from leukemic patients, the leukemic leukocytes made 20- to 30-fold more TAD and the GTP content decreased as compared to normal leukocytes. This procedure proved to be a suitable predictive test in a clinical setting because patients with positive tests responded to tiazofurin whereas those with negative ones did not. 4. The National Cancer Institute approved a chemotherapeutic phase I/II trial which concentrates on treatment of refractory acute myelocytic leukemia. Tiazofurin is infused in a 60-minute period with a pump to insure uniform delivery. A novel aspect of the trial was that it was directed primarily by the biochemical impact of tiazofurin on IMP dehydrogenase activity and GTP concentration and the tiazofurin doses were to be adjusted accordingly. Patients received allopurinol as a routine precaution against possible accumulation of uric acid in the kidney. 5. In the first eight patients, there was one complete remission, two entered the chronic phase, two entered into partial remission, one did not respond, and two were not evaluable. In the five patients who responded, there was a rapid, profound decrease in IMP dehydrogenase activity of the blast cells and a gradual decline in GTP concentrations. The blast cell count followed the decrease in the GTP concentration. The white blood cell count was largely preserved. 6. Bone marrow aspirates and peripheral blood samples showed that with tiazofurin treatment there was an induced differentiation of the myelocytes.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Enzyme-pattern-targeted chemotherapy with tiazofurin and allopurinol in human leukemia. 290 68

The pathways of adenine nucleotide catabolism were investigated in cultured beating cardiomyocytes. The activity of the enzymes involved in AMP degradation was assayed in cell extracts. Fluxes of label from ATP to the various purine derivatives were measured in intact cells. Under physiological conditions, cells degraded AMP through deamination to IMP. IMP was rapidly degraded to inosine, hypoxanthine, xanthine and uric acid, which were effluxed from the cells. This is in accord with the fact that the activity of AMP deaminase (EC 3.5.4.6) was 7-fold that of AMP 5'-Nucleotidase (EC 3.1.3.5). Mild ATP-degradation, induced by inhibition of glycolysis by iodoacetate, caused no alterations in the degradation pathways (more than 85% through deamination to IMP). However, fast ATP-degradation (83% of adenine nucleotides/10 min), induced by simultaneous inhibition of glycolysis and electron transport (by antimycin A), caused increased dephosphorylation of AMP to adenosine (50% of total AMP-degradation). The cardiomyocyte extracts were found to contain a significant activity of purine nucleoside phosphorylase (EC 2.4.2.1). Despite the presence of hypoxanthine-guanine phosphoribosyltransferase (EC 2.4.2.8), salvage of hypoxanthine to IMP, both at physiological as well as at conditions associated with ATP degradation, was slow. The salvage of adenosine appeared to be efficient at physiological conditions, but not at fast rates of ATP degradation.
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PMID:Pathways of adenine nucleotide catabolism in primary rat cardiomyocyte cultures. 325 63

We have observed previously that the reactions catalyzed by hypoxanthine/guanine phosphoribosyltransferase (HGPRTase) are activated by Mg(II), Mn(II), and Co(II), and we have defined the mechanism by which these activations proceed [Biochemistry 22, 3419-3424 (1983)]. A more extensive survey of the kinds of metal ions that will activate the HGPRTase catalysis now has been completed through the use of an HPLC assay procedure. Although Fe(II) and Ca(II) are unable to activate this reaction, a significant activation was achieved with the addition of spectroscopically pure Zn(II) to the assay solution. In addition some IMP synthesis resulted from the addition of Ni(II) to the assay mixture. Both the Zn(II) and Ni(II) kinetic effects on HGPRTase over a limited metal ion concentration range have been analyzed through the use of curve-fitting exercises. These results, in addition to the similar pH profiles for the activations by Mg(II), Mn(II), Co(II), and Zn(II), suggest that all of these metal ions activate the HGPRTase-catalyzed synthesis of IMP by way of the same mechanism [model II as defined by London and Steck, Biochemistry 8, 1767-1779 (1969)], during which two divalent ions bind to the HGPRTase active site per molecule of PRibPP.
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PMID:Activation of hypoxanthine/guanine phosphoribosyltransferase from yeast by divalent zinc and nickel ions. 354 95

6-Thio-3-deazaguanine (TDG), a relatively new purine antimetabolite, exhibits significant antitumor activity against a variety of experimental animal tumor models including C3H mammary adenocarcinoma, Lewis lung carcinoma, adenocarcinoma 755, and leukemias L1210 and P388. However, the drug was ineffective against 3-deazaguanine-resistant L1210 (both in vitro and in vivo) and CEM cells (in vitro). The resistant cells appear to lack HGPRTase activity because the extracts from these cell lines failed to convert hypoxanthine to IMP. These data indicate that TDG needs to be activated by hypoxanthine guanine phosphoribosyltransferase prior to its growth inhibitory effects. Cytotoxicity of TDG was completely reversed by hypoxanthine and inosine. TDG inhibited the synthesis of DNA and RNA equally and effectively, whereas the inhibition of protein synthesis required a prolonged drug exposure and appears to be a consequence of the inhibition of DNA and RNA synthesis. Data from these studies suggest that TDG is an effective antitumor agent, and its spectrum of antitumor activity and mechanism of action appears to be different from that of 3-deazaguanine.
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PMID:Antitumor activity and mechanism of action of 6-thio-3-deazaguanine. 381 77

In an attempt to immortalize the gene products of single neurons, somatic cell hybrids were produced by fusion of embryonic rat dorsal root ganglion (DRG) neurons with mouse neuroblastoma cells. Embryonic day 13 rat DRGs were fused with mouse neuroblastoma cells deficient in hypoxanthine phosphoribosyltransferase (HPRT; IMP:pyrophosphate phosphoribosyltransferase, EC 2.4.2.8). The hybrid cells were selected in medium with 100 microM hypoxanthine/1 microM aminopterin/12 microM thymidine to eliminate the neuroblastoma cells and with cis-hydroxyproline to retard fibroblast growth. Of the 17 lines derived, 4 manifested neuronal properties and were cloned. These lines retain both rat and mouse chromosomes and synthesize characteristic rat and mouse isoenzymes. Neuronal gangliosides, action potentials, and extensive neurite-like processes are exhibited by these hybrid cells, properties characteristic of DRG neurons but not of the neuroblastoma parent. Each line manifests a unique combination of action-potential properties and cell-surface markers, suggesting the selective expression of subsets of DRG neuronal genes. All of these neuronal properties are expressed constitutively, without the need for chemical induction or mitotic inhibition, and stably, without diminution after at least 5 months in culture. These lines may prove useful in the identification and isolation of gene products that characterize individual or small subsets of DRG neurons.
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PMID:Neuronal traits of clonal cell lines derived by fusion of dorsal root ganglia neurons with neuroblastoma cells. 385 35

Simple and rapid radiochemical assay procedures for the forward (IMP synthesis) and reverse (IMP pyrophosphorolysis) reactions catalyzed by hypoxanthine phosphoribosyltransferase have been developed. Enzyme activity in the forward direction was assessed by measuring the amount of [8-14C]IMP formed from [8-14C]hypoxanthine following their separation by polyethyleneimine-cellulose TLC in methanol:water (1:1, v/v). [8-14C]IMP has been synthesized from [8-14C]hypoxanthine, using hypoxanthine phosphoribosyltransferase derived from human brain, with subsequent purification by elution from phenyl boronate-agarose. Enzyme activity in the reverse direction was assessed by measuring the amount of [8-14C]uric acid formed from the labeled IMP following their separation by polyethyleneimine-cellulose TLC in 0.2 M LiCl saturated with boric acid (pH 4.5):95% ethanol (1:1, v/v), the transferase reaction being coupled with excess xanthine oxidase and catalase to overcome the unfavorable equilibrium.
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PMID:Hypoxanthine phosphoribosyltransferase: radiochemical assay procedures for the forward and reverse reactions. 400 57

1. Both the acid-soluble fraction and the nucleic acid fraction of wheat embryos were extensively labelled after incubation for 6hr. in the presence of [8-(14)C]adenine. Subsequent incubation in the absence of labelled adenine resulted in no loss of radioactivity to the medium during a 48hr. period. Radioautography indicated that during this period there was a continuous increase in the radioactivity present in the acid-insoluble fractions of the root and leaf tissues relative to that present in the coleorhiza and coleoptile. 2. During incubation at 25 degrees there was a 26-fold increase in the activity of 3'-nucleotidase between 4hr. and 24hr.; the activities of enzymes hydrolysing AMP and IMP increased to a smaller extent. The activities of adenine phosphoribosyltransferase and hypoxanthine phosphoribosyltransferase increased three- to five-fold during incubation at 25 degrees for 24hr. 3. Adenosine kinase, inosine phosphorylase and 5-phosphoribosyl pyrophosphate synthetase activities were high in extracts from dry embryos and did not increase during 48hr. at 25 degrees . 4. The increase in 3'-nucleotidase activity was prevented by cycloheximide, cryptopleurine or incubation at 4 degrees , but not by actinomycin D; these treatments did not depress the activity of the other enzymes measured. 5. The results are discussed in relation to RNA translocation within the wheat embryo during germination.
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PMID:Purine metabolism in germinating wheat embryos. 431 15

IMP:pyrophosphate phosphoribosyltransferase (IPPase) (EC 2.4.2.8) has been purified over 7000-fold from human erythrocytes. The purified enzyme moved as a single band on disc electrophoresis. Antisera prepared in rabbits and rats against the purified enzyme precipitated and neutralized the enzyme, but had no effect on AMP-pyrophosphate phosphoribosyltransferase (EC 2.4.2.7) activity. Evidence was found for isozymes (enzyme variants) of IPPase in normal erythrocytes. Erythrocyte lysates of five patients with Lesch-Nyhan disease reacted with antisera against normal IPPase. Lysates from LN erythrocytes blocked the inactivation of normal enzyme by the antibody. LN erythrocytes had about the same concentration of enzyme protein as normal erythrocytes. The genetic defect in LN results in the production of essentially normal amounts of an immunologically identifiable but catalytically incompetent enzyme. Thus LN is apparently the result of a mutation in a structural gene and is not due to deletion of a structural gene or defect in a regulatory gene.
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PMID:Purification of IMP:pyrophosphate phosphoribosyltransferases, catalytically incompetent enzymes in Lesch-Nyhan disease. 432 3

We here report the establishment of a seemingly permanent hybrid cell line formed by fusion of the cells of two biochemically mutant human lymphocyte lines. One parental line (UM-1-6TGr) was deficient in hypoxanthine-guanine phosphoribosyl transferase (IMP: pyrophosphate phosphoribosyltransferase, EC 2.4.2.8), and had two marker chromosomes. The second parental line (UM-21-5) was a clonal derivative of a citrullinemic lymphocyte line, and was, like the line of origin, dificient in argininosuccinic acid synthetase [(L)-Citrulline: (L)-aspartate ligase (AMP-forming), EC 6.3.4.5]. This line also had a marker chromosome, which was a B5 with a very prominent secondary constriction. After trypsinization of both parental lines, followed by addition to the fusion mixture of beta-propiolactone-inactivated Sendai virus, the cells were placed in a doubly selective medium (hypoxanthine-aminopterin-thymidine-containing medium in which the arginine was replaced with citrulline) to prevent the proliferation of the mutant parents. Under selective conditions, 97-99% of cells were found to be tetraploid, containing the three marker chromosomes; and the specific activities of the hybrid line transferase and synthetase were intermediate between normal and mutant line values. Furthermore, the UM-1-6TGr and UM-21-5 lines were producers of gamma and mu heavy chains of immunoglobulin, and of kappa light chains, as determined by immunodiffusion and immunofluorescence, and the hybrid line continued to synthesize and to secrete detectable levels of these same immunoglobulins. These studies demonstrate the genic and cytogenetic stability of this hybridized lymphocyte cell line, and prove that hybridization per se does not extinguish the activity of either the regulatory of structural genes involved in immunoglobulin synthesis.
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PMID:Establishment of a tetraploid, immunoglobulin-producing cell line from the hybridization of two human lymphocyte lines. 436 96

Tissue culture fibroblasts derived from patients with Lesch-Nyhan disease (congenital hyperuricosuria) have a reduced IMP:pyrophosphate phosphoribosyltransferase (EC 2.4.2.8) activity and therefore incorporate, as detected by radioautography, much smaller amounts of tritiated hypoxanthine or guanine into cell nuclei and cytoplasm than do normal cells. However, Lesch-Nyhan cells grown in close contact with normal fibroblasts incorporate these purines. This phenomenon, which requires cell to cell contact for correction of the mutant phenotype, has been called metabolic cooperation. After separation of Lesch-Nyhan cells from normal cells, there is a prompt reversion to the mutant phenotype although the transferase is stable under these conditions for many hours.These results are most compatible with the transfer from normal to mutant fibroblasts of the product of the normal enzyme, a nucleotide or a nucleotide derivative, rather than the transfer of the transferase or informational macromolecules leading to the synthesis of the enzyme. Metabolic cooperation may provide a mechanism for maintaining normal cell function in the heterozygote in vivo. Evidence has been presented previously that selection of normal cells, presumably during embryogenesis, also provides a means for achieving normal function in the heterozygote.
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PMID:Evidence for transfer of enzyme product as the basis of metabolic cooperation between tissue culture fibroblasts of Lesch-Nyhan disease and normal cells. 527 81


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