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:3.1.3.5 (5'-nucleotidase)
3,167 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

2-Chlorodeoxyadenosine (CdA) is active in chronic lymphocytic leukemia, hairy-cell leukemia, and low-grade lymphomas. In part, this spectrum of activity may be attributable to the selective toxicity of CdA to nondividing lymphocytes and monocytes. However, CdA is unstable at acidic pH and is degraded by bacterial nucleoside phosphorylases. The present experiments demonstrate that the 2'-arabino-fluoro derivative of CdA, designated CAFdA, is also directly toxic to quiescent lymphocytes and macrophages. Unlike CdA, CAFdA was stable at pH 2 and resisted degradation by Escherichia coli nucleoside phosphorylase. Cell killing was preceded by the formation of DNA strand breaks and could be prevented by supplementation of the medium with deoxycytidine. The initial DNA damage initiated the pattern of oligonucleosomal DNA fragmentation characteristic of apoptosis. Mutant lymphoblasts, deficient in deoxycytidine kinase, with elevated cytoplasmic 5'-nucleotidase, or with expanded deoxynucleotide pools secondary to increased ribonucleotide reductase activity, were cross-resistant to both CAFdA and CdA toxicity. One-week oral treatment with CAFdA (1 mg/ml in drinking water) achieved an average plasma concentration of 0.56 microM and eliminated 90% of chronic lymphocytic leukemia cells transplanted into severe combined immunodeficiency (scid) mice. Under the same conditions, CdA was much less active. Collectively, these results suggest that CAFdA could be effective as an oral agent in indolent lymphoproliferative diseases and in autoimmune diseases where lymphocyte and monocyte depletion is desirable.
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PMID:Oral antilymphocyte activity and induction of apoptosis by 2-chloro-2'-arabino-fluoro-2'-deoxyadenosine. 134 62

Cytosolic 5'-nucleotidase has been implicated in the phosphorylation of certain nucleosides of therapeutic interest. In vitro, IMP and GMP serve as the optimal phosphate donors for this nucleoside phosphotransferase reaction. Existing assays for nucleoside phosphorylation effected by 5'-nucleotidase require a radiolabeled nucleoside as the phosphate acceptor and separation of the substrate-nucleoside from product-nucleotide has been accomplished either by a filter binding method or HPLC. However, detection of the phosphorylation of unlabeled nucleoside by HPLC is difficult since the ultraviolet absorbance of the phosphate donor, IMP, frequently obscures the absorbance of newly formed nucleotide. The use of ribavirin 5'-phosphate (RMP, 1,2,4-triazole-3-carboxamide riboside 5-monophosphate) as the phosphate donor obviates this difficulty since this triazole heterocycle does not significantly absorb at the wavelengths used to detect most nucleoside analogs. Using this procedure, a 5'-nucleotidase activity from the 100,000 x g supernatant fraction of human T-lymphoblasts deficient in adenosine kinase, hypoxanthine-guanine phosphoribosyltransferase, and deoxycytidine kinase, was characterized with regard to structure-activity relationships for certain inosine and guanosine analogs.
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PMID:A novel non-radioactive method for detection of nucleoside analog phosphorylation by 5'-nucleotidase. 143 Jul 86

Carbovir (CBV) is a highly selective carbocyclic nucleoside inhibitor of HIV replication in human lymphocytes and is potentially useful in the treatment of AIDS [Vince et al. (1988) Biochem. Biophys. Res. Commun. 156, 1046-1053]. Using human lymphoid cells severely deficient in nucleoside kinases, we were able to identify the route of activation of CBV metabolism. The present studies have demonstrated that CBV is anabolized to the mono-, di-, and triphosphates and to guanosine 5'-triphosphate in CCRF-CEM cells. Conversion to GTP amounted to 15-20% of the total analogue nucleotides formed in the cells and may arise from CBV through depurination and salvage via HGPRT. Evidence was obtained that neither deoxycytidine kinase, adenosine kinase, or mitochondrial deoxyguanosine kinase is primarily involved in the initial step of phosphorylation of CBV in CCRF-CEM cells. In contrast, earlier studies [Johnson & Fridland (1989) Mol. Pharmacol. 36, 291-295] showed that a cytosolic 5'-nucleotidase catalyzes the activation of CBV to the monosphosphate. Other biochemical effects examined showed that the nucleobases hypoxanthine and adenine, but not guanine, their respective nucleosides, and the dideoxynucleosides 2',3'-dideoxyinosine, 2',3'-dideoxyguanosine, and 3'-azido-3'-deoxythymidine produced significant increased accumulation of CBV nucleotides in CEM cells. The exact mechanism for this potentiation of CBV phosphorylation has not been elucidated but may be due to a modulating effect of intracellular nucleotides on 5'-nucleotidase activity.
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PMID:Metabolism of the carbocyclic nucleoside analogue carbovir, an inhibitor of human immunodeficiency virus, in human lymphoid cells. 227 22

2',3'-Dideoxyinosine (ddlno) is a potent and selective inhibitor of human immunodeficiency virus in human lymphoid cells and monocytes/macrophages. Earlier studies [J. Biol. Chem. 263:15354 (1988)] showed that anabolism of ddlno in human lymphoid cells is mediated via an initial step of phosphorylation and subsequent amination to dideoxy-AMP via adenylosuccinate synthetase/lyase. Evidence was obtained that neither adenosine kinase nor deoxycytidine kinase is involved in the phosphorylation of this compound in human lymphoid cells. We now find that, in the presence of MgCl2, KCl, and inosine-5'-monophosphate as phosphate donor, purified cytosolic 5'-nucleotidase catalyzed the phosphorylation of ddlno. Although not phosphate donors, ATP, diadenosine tetraphosphate, and glycerate-2,3-bisphosphate stimulate this phosphorylation by the nucleotidase 4-5-fold. In addition to ddlno, the antiviral nucleoside analogs 2',3'-dideoxyguanosine and carbovir were substrates for this enzyme. The relative phosphorylation of these compounds varied with the concentration of the phosphate donor IMP. Approximate Km values of the nucleotidase for inosine, ddlno, dideoxyguanosine, and carbovir were, respectively, 3.4, 0.5, 0.9, and 1.7 mM. Although the substrate activity of dideoxynucleosides is inefficient, it appears likely that this nucleotidase is responsible for the metabolism of these compounds to their active nucleotides, yielding antiviral activity in human lymphoid cells.
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PMID:Phosphorylation of 2',3'-dideoxyinosine by cytosolic 5'-nucleotidase of human lymphoid cells. 254 85

A microassay requiring as few as 2 X 10(5) cells per assay was developed for systematic analysis of 9 purine enzymes in lymphocytes from equine peripheral blood, spleen, lymph node, thymus and bone marrow. The activities of adenosine deaminase (ADA), purine nucleoside phosphorylase (PNP), adenosine kinase (AK), deoxyadenosine kinase (dAK), deoxycytidine kinase (dCK), 5'-nucleotidase (5'-N), AMP deaminase, hypoxanthine-guanine phosphoribosyl transferase (HGPRT or HPRT), and adenine phosphoribosyl transferase (APRT) were measured by this microassay in lymphocytes from peripheral blood from four different breeds of horses (Arabian, Quarter Horse, Thoroughbred and Shetland Pony). There were no significant differences in the enzyme activities among the various breeds. Peripheral blood lymphocytes (PBL) from foals exhibited enzyme activities similar to those observed for adult animals. All lymphoid tissue contained similar levels of activity for each kinase (AK, dAK and dCK). Spleen had the highest activity for ADA, PNP, 5'-N, and HGPRT. The lowest activity for ADA, APRT, PNP and AMP deaminase was found in thymus. Enzymatic activities that varied the most among the tissue were 5'-N, ADA, APRT, HGPRT and AMP deaminase.
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PMID:Distribution of enzymes of purine metabolism in lymphocytes of horse, Equus caballus. 299 Aug 11

In an attempt to determine the metabolic defect causing severe combined immunodeficiency (SCID) in horses in which altered purine metabolism has been observed, various parameters of purine and pyrimidine metabolism were evaluated. The activities of nine purine enzymes (adenosine kinase, purine nucleoside phosphorylase, deoxyadenosine kinase, deoxycytidine kinase, 5'-nucleotidase, AMP deaminase, hypoxanthine-guanine phosphoribosyl transferase, and adenine phosphoribosyl transferase were measured in fibroblasts. All activities determined for SCID horses were normal. Uptake of 10 microM adenosine or 2'-deoxyadenosine (a growth inhibitory concentration for SCID fibroblasts) by SCID fibroblasts was identical to that found for normal fibroblasts in the presence of both 1 and 50 microM phosphate. The Km determined for the transport of both adenosine and 2'-deoxyadenosine was 35 microM. In the presence of p-nitrobenzylthioguanosine (a nucleoside transport inhibitor), 2'-deoxyadenosine uptake was inhibited to the same extent in all fibroblast lines tested. To determine if the last step in pyrimidine biosynthesis might be altered in SCID fibroblasts, UMP synthase activities were evaluated but found to be normal (0.5 nmol UMP formed/min/mg protein).
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PMID:Metabolic investigations of fibroblasts from horses, Equus caballus, with hereditary severe combined immunodeficiency. 299 78

WI-L2 cells (a B-lymphoblastoid cell line) were more resistant than CEM cells (a T-lymphoblastoid cell line) to deoxyadenosine, ara-A (9-beta-D-arabinofuranosyladenine), or ara-C (1-beta-D-arabinofuranosylcytosine) inhibition. This was caused by a difference in the composition of cytosol 5'-nucleotidases between WI-L2 and CEM cells. In intact cells, the endogenous production of deoxyadenosine from WI-L2 cells deficient in adenosine kinase (EC 2.7.1.20) and deoxycytidine kinase (EC 2.7.1.74) was consistently high, despite changes in endogenous adenosine production. Endogenous production of deoxyadenosine from CEM cells deficient in adenosine kinase and deoxycytidine kinase was, however, coordinated with endogenous adenosine production. In broken cells, cytosol dAMPase (2'-deoxyadenosine 5'-monophosphate 5'-nucleotidase) activity of WI-L2 cells was 3-5-fold higher than that of CEM cells. dAMPase activity could be separated from ATP-activated IMPase (inosine 5'-monophosphate 5'-nucleotidase) by gel filtration (molecular weight: dAMPase; 39,000-46,000; ATP-activated IMPase, greater than 150,000). Cytosol ATP-activated IMPase and dAMPase were isolated by phosphocellulose or DEAE-Bio-Gel A chromatography from non-specific phosphatases. The ATP-activated IMPase showed only marginal activity towards dAMP (2'-deoxyadenosine 5'-monophosphate), ara-AMP (9-beta-D-arabinofuranosyladenine 5'-monophosphate), or ara-CMP (cytosine-beta-D-arabinofuranoside 5'-monophosphate), even in the presence of ATP. The activity of ATP-activated IMPase was similar in WI-L2 and CEM cells. dAMPase was separated into two peaks by DEAE-Bio-Gel A chromatography; one of these peaks degraded ara-AMP and ara-CMP. The activities of both peaks from WI-L2 cells were higher than those from CEM cells. These results show that the degradation of dAMP, ara-AMP or ara-CMP was more specific and rapid in WI-L2 than in CEM cells.
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PMID:Differential composition of cytosol 5'-nucleotidases between T and B lymphoblasts. 301 Nov 14

Long-term bovine lymphocyte cultures were initiated by stimulation with alloantigens and maintained in continuous culture using medium containing recombinant human interleukin-2 (rh IL-2). The development of specific and lectin-dependent killing was monitored following primary alloantigen challenge. Cytolytic activity was barely detectable after 7 days of culture, but gradually increased with peak activity occurring after 21 days of culture. A panel of monoclonal antibodies (MoAb) was used to determine whether a shift in the antigen phenotype of the cell population occurred during culture. The primary cell type that grew in culture was of the T-cell lineage with minimal or no expression of class II antigens. The activities of adenosine deaminase (ADA), purine nucleotide phosphorylase (PNP), adenosine kinase (AK), deoxyadenosine kinase (dAK), deoxycytidine kinase (dCK), 5'-nucleotidase (5'-N), AMP deaminase, hypoxanthine-guanine phosphoribosyl transferase (HGPRT or HPRT), and adenine phosphoribosyl transferase (APRT) were measured by microassay in resting peripheral blood lymphocytes (PBL) and in cells from long-term cultures. Large increases in the activities of PNP and HPRT with a decrease in the activity of ADA were observed. The data show that long-term cultures of lymphocytes can be readily generated, and that sequential changes in antigenic phenotype and function can be monitored and correlated with quantitative changes in enzyme activity.
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PMID:Development and maintenance of bovine cytotoxic lymphocytes with recombinant human interleukin-2. 348 20

Purine and pyrimidine enzyme profiles of human cell lines have been investigated. A novel observation was the finding that most of the cell lines showed very low or undetectable levels of cytidine (deoxycytidine) deaminase, while they possessed pyrimidine 5'-nucleotidase, cytidine and deoxycytidine kinase activities. Most cell lines showed high levels of adenosine deaminase and purine nucleoside phosphorylase activities and low levels of purine 5'-nucleotidase. We propose that high adenosine deaminase and purine nucleoside phosphorylase activities and low cytidine deaminase activity may be of importance for immature hematopoietic cells in order to ensure a balanced synthesis of the DNA precursors.
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PMID:Low cytidine deaminase levels in human hematopoietic cell lines. 362 11

The mammalian deoxyribonucleoside kinases are deoxycytidine kinase, thymidine kinase 1 and 2 and deoxyguanosine kinase. These enzymes phosphorylate deoxyribonucleosides and thereby provide an alternative to de novo synthesis of DNA precursors. Their activities are essential for the activation of several chemotherapeutically important nucleoside analogues. In recent years, these enzymes have been thoroughly characterised with regard to structure, substrate specificity and patterns of expression. In this review, these results are reviewed and furthermore, the physiologic metabolic role of the anabolic enzymes is discussed in relation to catabolic pathways. The significance of this information for the development of therapeutic protocols and choice of animal model systems is discussed. Finally, alternative pathways for nucleoside analogue phosphorylation are surveyed, such as the phosphotransfer capacity of 5'-nucleotidase.
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PMID:Mammalian deoxyribonucleoside kinases. 749 63


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