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Oxidative metabolism of cholesterol precursors: sensitivity to ketoconazole, an inhibitor of cytochrome P-450 总被引:2,自引:0,他引:2
The effects of ketoconazole, an inhibitor of cytochrome P-450, on the metabolism of the cholesterol precursors lanosterol, dihydrolanosterol, lanost-8-en-3 beta,32-diol, and 3 beta-hydroxylanost-8-en-32-al were investigated in subcellular fractions of rat liver and in rat hepatocytes in culture. At low (1-2 microM) concentrations of the drug, the oxidative demethylation of lanosterol was inhibited by about 70% in the subcellular fractions but there was no effect on the metabolism of the 3 beta, 32-diol or the 32-aldehyde. Higher drug concentrations (10-20 microM) were required to inhibit the oxidative metabolism of these cholesterol precursors. Similar results were obtained during longer-term incubations using hepatocytes in culture medium, but higher concentrations of ketoconazole were required to effect the same degree of inhibition of each precursor. In the subcellular fractions, dihydrolanosterol, the 3 beta,32-diol and the 32-aldehyde were each metabolized to more polar sterols, in addition to cholesterol. Ketoconazole also inhibited the formation of these polar substances. 相似文献
4.
Regulatory and Structural Properties of the Cyanobacterial ADPglucose Pyrophosphorylases 总被引:7,自引:5,他引:2 下载免费PDF全文
ADPglucose pyrophosphorylase (EC 2.7.7.27) has been purified from two cyanobacteria: the filamentous, heterocystic, Anabaena PCC 7120 and the unicellular Synechocystis PCC 6803. The purification procedure gave highly purified enzymes from both cynobacteria with specific activities of 134 (Synechocystis) and 111 (Anabaena) units per milligram protein. The purified enzymes migrated as a single protein band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with molecular mass corresponding to 53 (Synechocystis) and 50 (Anabaena) kilodaltons. Tetrameric structures were determined for the native enzymes by analysis of gel filtrations. Kinetic and regulatory properties were characterized for the cyanobacterial ADPglucose pyrophosphorylases. Inorganic phosphate and 3-phosphoglycerate were the most potent inhibitor and activator, respectively. The Synechocystis enzyme was activated 126-fold by 3-phosphoglycerate, with saturation curves exhibiting sigmoidicity (A0.5 = 0.81 millimolar; nH = 2.0). Activation by 3-phosphoglycerate of the enzyme from Anabaena demonstrated hyperbolic kinetics (A0.5 = 0.12 millimolar; nH = 1.0), having a maximal stimulation of 17-fold. I0.5 values of 95 and 44 micromolar were calculated for the inhibition by inorganic phosphate of the Synechocystis and Anabaena enzyme, respectively. Pyridoxal-phosphate behaved as an activator of the cyanobacterial enzyme. It activated the enzyme from Synechocystis nearly 10-fold with high apparent affinity (A0.5 = 10 micromolar; nH = 1.8). Phenylglyoxal modified the cyanobacterial enzyme by inactivating the activity in the presence of 3-phosphoglycerate. Antibody neutralization experiments showed that anti-spinach leaf (but not anti-Escherichia coli) ADPglucose pyrophosphorylase serum inactivated the enzyme from cyanobacteria. When the cyanobacterial enzymes were resolved on sodium dodecyl sulfate- and two-dimensional polyacrylamide gel electrophoresis and probed with Western blots, only one protein band was recognized by the anti-spinach leaf serum. The same polypeptide strongly reacted with antiserum prepared against the smaller spinach leaf 51 kilodalton subunit, whereas the anti-54 kilodalton antibody raised against the spinach subunit reacted weakly to the cyanobacterial subunit. Regulatory and immunological properties of the cyanobacterial enzyme are more related to the higher plant than the bacterial enzyme. Despite this, results suggest that the ADPglucose pyrophosphorylase from cyanobacteria is homotetrameric in structure, in contrast to the reported heterotetrameric structures of the higher plant ADPglucose pyrophosphorylase. 相似文献
5.
R Mu?oz R Iglesias J M Ferreras F J Arias M A Rojo T Girbés 《Cellular and molecular biology, including cyto-enzymology》1991,37(7):671-677
Long-term ingestion of sublethal n-butanol doses by rats led to a noteworthy increase in the resistance of in vitro brain ribosomal function to the acute inhibitory action of ethanol and isopropanol. Withdrawal of n-butanol did not change this adaptation process immediately. The step affected seems to be the elongation of polypeptide chains. The dependence of in vitro translation on incubation temperature was affected by the adaptation process, the translation system of chronic animals being less stimulatable than that of control animals at low temperature. 相似文献
6.
Evidence for a differential physiological modulation of brown fat iodothyronine 5'-deiodinase activity in the perinatal period 总被引:1,自引:0,他引:1
M Giralt I Martin T Mampel F Villarroya R Iglesias O Vi?as 《Biochemical and biophysical research communications》1988,156(1):493-499
Brown adipose tissue iodothyronine 5'-deiodinase increases progressively in fetuses from the day 17 of pregnancy on, it reaches peak values on the 20th day of gestation and declines in the last days of fetal life as well as during the first day of life. Birth of premature fetuses causes a sudden drop in the enzyme activity. Postmaturity is associated to a decrease in brown fat 5'-deiodinase similar to that found after birth in fetuses born at term. In the first hours of life brown fat iodothyronine 5'-deiodinase is essentially insensitive to the cold-stimulus. Present data indicates that, differently from adult rats, brown fat iodothyronine 5'-deiodinase activity during the perinatal period is dissociated from the thermogenic activity of the tissue. It is suggested that factors different from the action of the sympathetic nervous system may play a main role in brown fat iodothyronine 5'-deiodinase activity modulation in the fetal and neonatal life. 相似文献
7.
Protein kinase C from small intestine epithelial cells 总被引:1,自引:0,他引:1
G Velasco C F Iglesias P Domínguez F Barros S Gascón P S Lazo 《Biochemical and biophysical research communications》1986,139(3):875-882
Protein kinase C activity has been identified in cytosolic and membrane fractions from rat and rabbit small intestine epithelial cells. The cytosolic fraction comprised about the 75% of total activity. Protein kinase C activity was resolved from other protein kinase activities by ion exchange chromatography. Phosphatidylserine or phosphatidylinositol were required for protein kinase C to be active. In addition, the activity was enhanced by the presence of a diacylglycerol. Diolein and dimyristin were the most effective (13-14 fold activation). In the presence of phosphatidylserine and diolein, the Ka for activation by Ca2+ was 10(-7)M. The phorbol ester TPA substituted for diacylglycerol in activating protein kinase C. Brush border and basolateral membranes contained protein kinase C activity, although the specific activity of the basal lateral membranes was four-fold higher than the specific activity of the brush border membranes. The presence of PKC in small intestine epithelial cells might have important implications in the Ca2+ mediated control of ionic transport in this tissue. 相似文献
8.
Dielectrophoretic behavior of yeast cells: effect of growth sources and cell wall and a comparison with fungal spores. 下载免费PDF全文
Saccharomyces cerevisiae showed different dielectrophoretic behavior depending on the source of carbon for growth. Growth on fermentable carbon sources produced a dielectrophoretic response that decreased according to the amount of sugar present in the culture medium. Growth on nonfermentable carbon sources produced a constant dielectrophoretic yield, independent of the amount and source of carbon present in the medium. The dielectrophoretic yield, however, was independent of the nitrogen source. The yield spectrum for S. cerevisiae protoplasts was similar to that for the cells, although a decrease in the absolute value was observed. This decrease could be explained by the reduction in cell size and by assuming that the cell wall contributes a negative net charge to the yield. Fungal spores responded to the nonuniform electric field in the same range of frequencies as assayed for yeast cells. 相似文献
9.
Homogeneous groups of cells were excised at regular intervals from opposing (extensor and flexor) motor tissue of Samanea saman (Jacq) Merrill maintained in white light for 34 hours. H+ fluxes between the tissue and bathing solution were then monitored during 30 minutes of darkness. Flux rates in both cell types vary with circadian rhythms. Flexor cells secrete H+ to the medium during two-thirds of the circadian cycle and take up H+ during the remainder of the cycle, while extensor cells take up H+ from the medium during the entire cycle. 相似文献
10.
H fluxes in excised samanea motor tissue : I. Promotion by light 总被引:11,自引:10,他引:1
Previous investigators revealed that white light-promoted leaflet opening in Samanea saman (Jacq) Merrill depends upon K+ uptake by extensor cells and efflux from flexor cells of the pulvinus, while dark-promoted closure depends upon K+ fluxes in the opposite directions. We now monitored H+ fluxes during pulvinar movement to test a model proposing coupled H+/K+ fluxes. H+ fluxes were monitored by measuring changes in the pH of a weakly buffered solution (initial pH = 5.5) bathing excised strips of extensor or flexor tissue. White light at hour 3 of the usual dark period promoted pulvinar opening, H+ efflux from extensor cells and uptake by flexor cells, while darkness at hours 2 to 4 of the usual light period promoted pulvinar closure, H+ uptake by extensor cells and efflux from flexor cells. The following conditions altered H+ fluxes during dark-promoted closure. (a) Light reversed the directions of the fluxes in both extensor and flexor cells. (b) Anoxia increased the rate of H+ uptake by extensor cells and promoted H+ uptake (rather than efflux) by flexor cells, consistent with an outwardly directed H+ pump. KCN showed similar effects initially, but they were transient. (c) Increase in external pH from 5.5 to 6.7 promoted H+ efflux (rather than uptake) by extensor cells and increased the rate of H+ efflux from flexor cells, presumably by decreasing the rate of inward diffusion. (d) Change in external K+ did not alter H+ fluxes by extensor cells, but removal of external K+ decreased the rate of H+ efflux from flexor cells by 70%. These observations support a model for coupled H+/K+ fluxes in pulvinar cells during light-and dark-promoted leaflet movements. 相似文献