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)

The specific activity of HMG-CoA reductase, the major rate-limiting enzyme in the sterol biosynthetic pathway, declined linearly with increasing cell density in four different lines of mammalian cell cultures. As expected, this caused the rates of sterol synthesis from [14C]acetate to decline in a parallel manner. The decrease in reductase activity in the dense cultures was also correlated with decreased incorporation of [14C]acetate into fatty acids and [3H]thymidine into DNA. In contrast, the activities of two enzymes, NADH dehydrogenase and 5'-nucleotidase, which are not involved in lipid synthesis, were independent of changes in cell density. The simplest explanation for these data is tht HMG-CoA reductase and the synthesis of sterol and fatty acids are regulated in concordance with the rate of cell growth and proliferation.
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PMID:The activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase and the rate of sterol synthesis diminish in cultures with high cell density. 626 81

The experiments reported herein compare growth kinetics and biochemical properties of cultured skin fibroblasts from patients with Duchenne muscular dystrophy (DMD) and matched normal controls. On day 7 after plating (6000 cells/cm2) cell number and DNA per dish are significantly reduced (P less than 0.0001) in the cultures from DMD patients (n = 14), compared to those from controls (n = 10). Moreover DMD cells contain less lipids and proteins per dish but more per cell than normal fibroblasts (not significant). Variations of media (McCoy's medium instead of Eagle's minimum essential medium) resulted in the same differences between DMD and control cells. Cell kinetic experiments (plating density: 2000 cells/cm2) show increased doubling times of DMD fibroblasts (P less than 0.001; nDMD = 5; ncontrols = 4) whereas plating efficiency is equal for both DMD and controls. On day 7 activity of the membrane bound enzyme 5'-nucleotidase either per mg protein or per microgram DNA is significantly elevated in cells from DMD patients (P less than 0.0005; nDMD = 8; ncontrols = 9) independent of cell density. Thus all findings in cultured DMD fibroblasts: increased doubling time, tendency to more voluminous cells, and elevated 5'-nucleotidase activity per cell suggest, that the DMD cells behave similar to prematurely aging cells. Until now we were not able to check whether any alterations of the plasma membrane are inducing early senescence or, reversely, premature aging is the cause of the postulated membrane alterations. If these findings were to be confirmed in cultured amniotic cells from DMD fetuses, thay could serve as a potential prenatal diagnosis of the disease.
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PMID:Abnormal growth kinetics and 5'-nucleotidase activities in cultured skin fibroblasts from patients with Duchenne muscular dystrophy. 627 84

A zymogen granule fraction has been isolated from rat pancreas, and its purity has been assessed by biochemical and morphological criteria. Specific activities of two marker enzymes, amylase and chymotrypsin, are increased by 4.6 and 5.4-fold, respectively, as compared to the homogenate. The purified fraction is devoid of detectable RNA, DNA and 5'-nucleotidase, glucose-6-phosphatase, and cytochrome c oxidase activities. Electron micrographs confirm the absence of mitochondria, lysosomes, and rough endoplasmic reticulum fragments. Zymogen granule membranes were isolated from this fraction on a sucrose gradient following lysis in alkaline buffer. Secretory contaminants were efficiently removed from the membranes as indicated by experiments in which labeled secretory proteins were added during the isolation procedure and secondly by measuring residual levels of amylase and chymotrypsin. Three enzyme activities were found in the membranes: thiamine pyrophosphatase, ATP-diphosphohydrolase, and low levels of acid phosphatase. Membrane proteins were solubilized by urea-Triton X-100 and separated in double-dimension (isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis). Isoelectric point and molecular weight of each protein band were determined.
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PMID:Isolation of zymogen granules from rat pancreas and characterization of their membrane proteins. 629 Feb 20

A method using sucrose gradient centrifugation is described for the purification of plasma membranes of guinea pig peritoneal macrophages. The subcellular fractions obtained have been submitted to a biochemical and ultrastructural analysis. Two plasma membrane markers, 5'-nucleotidase and alkaline phosphodiesterase I, have been assayed at the same time as markers for other subcellular organelles, DNA (nuclei), succinic dehydrogenase (mitochondria), inosine diphosphatase (endoplasmic reticulum), and acid phosphatase (lysosomes). The exposure of the plasma membranes to a low concentration of digitonin allowed us to obtain their high purification. They are only contaminated by 2-3% of other cell components present in the macrophages homogenate. The representative ultrastructural technique used has confirmed the purity of the plasma membranes isolated.
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PMID:Analytical subcellular fractionation of guinea pig peritoneal macrophages: preparation of purified plasma membranes. 629 13

The persistence of normal thymidine nucleotidase (ThyNase) activity in subjects with pyrimidine nucleotidase (PyrNase) deficiency suggested the possible existence of separate isozymes in normal human erythrocytes. This hypothesis was confirmed by studies of PyrNase-deficient individuals from five unrelated families. Erythrocytes deficient in PyrNase retained normal activity of an enzyme system preferentially active at pH 6.2 with a variety of 2'-deoxyribonucleoside 5'-monophosphate substrates, including those of uridine, thymidine, and cytidine. Lesser activities were observed with the corresponding ribonucleotides. Normal control hemolysates were also found capable of effectively dephosphorylating purine nucleotides (dAMP greater than AMP) when pH was lowered sufficiently from the pH 7.4-8.0 region commonly used in conventional assays. Variations in substrate specificity, pH optima, kinetics, and sensitivity to inactivation by Pb2+ indicated the existence of multiple 5'-nucleotidase isozymes in normal erythrocytes: PyrNase and deoxyribonucleotidase(s) that might function physiologically in the conversion of DNA-derived nucleotides to diffusible nucleosides. Evolution of such a unique 5'-nucleotidase suggests that normal erythroblast maturation and nuclear extrusion is accompanied by a degree of karyolysis sufficient to require dephosphorylation and clearance of DNA degradation products.
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PMID:Identification of thymidine nucleotidase and deoxyribonucleotidase activities among normal isozymes of 5'-nucleotidase in human erythrocytes. 632 Jan 96

Glucagon receptor levels, glucagon-stimulated and other forms of adenylyl cyclase activity, and regulatory component activity of adenylyl cyclase were determined in hepatic plasma membranes of rats administered streptozotocin without and with insulin to produce varying degrees of hyperglycemia. Receptor levels were assayed by direct binding of the specific probe [125I-Tyr10]-iodoglucagon; regulatory component activity was assayed by the capacity to reconstitute stimulatory regulation in deficient membranes from cyc- S49 murine lymphoma cells. In rats given 150 mg streptozotocin, glucagon stimulation of adenylyl cyclase as well as basal, sodium fluoride, 5' guanylylimidodiphosphate [GMP-P(NH)P] and Mn-dependent activities were reduced 50%, glucagon receptor levels but not affinity were reduced 67%, and regulatory component activity was decreased 50%. In addition, alpha 1-adrenergic receptors and 5'-nucleotidase were similarly reduced in diabetes. However, specific ouabain-inhibitable Na+, K+, ATPase activity was not altered by streptozotocin treatment. The streptozotocin-induced changes were noted within 24 h and became maximal by 120 h after its administration. All of these decreases were partially reversed by in vivo insulin treatment. DNA, cytochrome c oxidase, glucose-6-phosphatase, and N-acetyl-beta-glucosaminidase content in hepatic plasma membrane preparations were not substantially different in diabetic as compared with control animals. The data demonstrate that glucagon-mediated regulation of cyclic AMP formation is deranged in insulin deficiency owing to a combined decrease in receptors, derangement of the coupling mechanism intervening between receptor and adenylyl cyclase, and possibly, an altered basal effector system. Some of these changes appear to reflect a "desensitization-like" phenomenon which may or may not be attributable to the hyperglucagonemia of diabetes mellitus. There also appears to be a concurrent generalized decrease in several but not all plasma membrane receptor and enzymatic proteins. This may be the result of a number of processes among which is the accelerated proteolysis of uncontrolled diabetes.
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PMID:Glucagon-stimulable adenylyl cyclase in rat liver. The impact of streptozotocin-induced diabetes mellitus. 632 32

The activities of adenosine deaminase, purine nucleoside phosphorylase, membrane 5'-nucleotidase and DNA content in thymus and spleen lymphocytes as well as the immune function of T and B lymphocytes of the spleen of C3HA mice with o-AAT-induced hepatomas were studied 1, 3 weeks and 3, 8, 12 months after o-AAT treatment was instituted. In the early stages of the hepatocarcinogenesis (up to 3 months), the elevation of the activity of all the enzymes and DNA content in thymocytes and T and B lymphocytes was observed. These changes coincided with the enhancement of the immune responses that manifested in the increased content of PFC, EA-RFC and high response to PHA and Con A. In the late stages, the decreased activities of purine nucleosides and nucleotide metabolizing enzymes correlated with disturbances of lymphocyte differentiation and lowering of the host immune response.
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PMID:[Changes in purine metabolism and in the immune response in the thymic and splenic lymphocytes of C3HA mice during chemical hepatocarcinogenesis]. 633 63

To define reproducible conditions for the homogenization of small-intestinal biopsy samples, tissue homogenization has been studied by the use of three different homogenizers. Tissue samples of increasing wet weights (0.5-10.8 mg) were homogenized in a fixed volume (1 ml) before DNA and protein were determined. The DNA to protein ratio was calculated for all wet weights and used as a measure for reproducible homogenization. The minimum tissue wet weight needed for analysis (2 mg) was determined from the values obtained for the DNA to protein ratio. Highly sensitive techniques are described in detail for the assay of brush border (maltase, lactase, sucrase, neutral alpha-glucosidase, alkaline phosphatase, gamma-glutamyl transferase, leucyl-beta-naphthylamidase), basolateral membrane (5'-nucleotidase), and mitochondrial (succinate dehydrogenase) marker enzymes and for four acid hydrolases (acid phosphatase, acid beta-D-galactosidase, N-acetyl-beta-D-glucosaminidase, acid diesterase) in human and rat jejunal mucosa. Linear kinetics have been established for all enzyme assays. The optimal dilution of tissue homogenate for the assay of the various enzymes has been determined to enable the determination of a maximum number of enzymes in each homogenate. The range of enzyme activities in samples of human and rat jejunal mucosa has been determined.
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PMID:Enzyme activities in human and rat jejunal mucosa. 667 54

The purpose of this study was to compare the adenylate cyclase of a tumour (rat osteosarcoma) growing in vivo with that of fast-growing embryonic bone. In the tumour the enzyme activity per total protein or DNA (under the same assay conditions) was 6--10-fold lower than in embryonic bone. To characterize this difference, we examined the kinetic properties of the enzyme in partially purified plasma membranes from the two tissues. A purification procedure based on differential centrifugation and discontinuous-sucrose-gradient centrifugation yielded a 10-fold increase in the specific activities of adenylate cyclase and 5'-nucleotidase in bone. The same procedure yielded an enriched membrane preparation from the tumour, but, relative to 5'-nucleotidase, a loss of 30% in adenylate cyclase occurred, which could not be recovered from another fraction. Kinetic analysis revealed that the lower adenylate cyclase activity in the tumour was due to a decrease in Vmax.. There was no significant difference in Ks (approx. 0.15 mM), and in the Km for GTP and p[NH]ppG. There were marked differences, however, in the extent of stimulation by p[NH]ppG, GTP and hormone, which was greater in tumour, and in the K1 for adenosine inhibition, which was 140 microM in bone and 500 microM in tumour. Under maximum stimulatory conditions, the enzyme activity in the tumour approached that in bone. The kinetic differences between bone and tumour enzyme were decreased by detergent solubilization, suggesting that the membrane environment plays a role in the generation of the observed differences.
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PMID:Comparison of bone and osteosarcoma adenylate cyclase. Partial purification of membranes and kinetic properties of enzyme. 693 Feb 65

Different human T cell populations were assayed for susceptibility of DNA synthesis to inhibition by deoxyguanosine. T lymphocytes from the thymus were most sensitive to inhibition of proliferation by deoxyguanosine (90% inhibition at 10 microM deoxyguanosine). This exquisite sensitivity of thymocytes appeared related to an enhanced ability of these cells for uptake and phosphorylation of deoxyguanosine to deoxyGTP and by their reduced ability to degrade accumulated deoxyGTP. Compared to more mature T lymphocytes and B cells, thymocytes contained the highest level of the salvage enzyme deoxynucleoside kinase and the lowest level of the nucleotide degrading enzyme, 5'-nucleotidase. The present study suggests that the levels of these 2 enzymes can serve as differentiation markers, identifying T cells at various stages of maturation, and that the loss of sensitivity to deoxyguanosine toxicity may be a stepwise process. Further, a deficiency in purine nucleoside phosphorylase may preferentially interfere with T cell maturation at an intrathymic stage of T cell differentiation.
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PMID:The expression of deoxyguanosine toxicity in T lymphocytes at different stages of maturation. 696 9


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