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1.
Three phosphorylated dinucleosides designated HS1, HS2, and HS3, isolated from the water-mould Achlya, were shown to significantly inhibit ribonucleotide reductase activity from Achlya. All three compounds decreased CDP reduction in fungal extracts by 50% at concentrations of 0.1mM. At the same concentration HS3 also inhibited partially purified CDP reductase from Chinese hamster ovary cells by at least 80% but showed only 10% inhibition with enzyme from E.coli. ADP reductase activity from Achlya was inhibited 50% by both HS1 and HS3 at 0.1mM. HS2 however, showed no inhibitory effect on purine reduction. The levels of ribonucleotide reductase during the asexual growth cycle of Achlya correlated with thymidine uptake into DNA and with the synthesis of HS compounds.  相似文献   

2.
6-phosphogluconate, potentiated activation of ribulose bisphosphate carboxylase from Pseudomonasoxalaticus whereas fructose-1,6-bisphosphate inhibited activation and fructose-6-phosphate had no effect. The presence of 1 mM 6-phosphogluconate during activation reduced the Kact for Mg2+ from 1.4 mM to approximately 0.2 mM. In the absence of 6-phosphogluconate, the enzyme responded sigmoidally to increasing CO2 (Hill coefficient, h, of 1.8), with a concentration causing half maximal activation, Act0.5, of 15 mM NaHCO3. In the presence of 1 mM 6-phosphogluconate h was reduced to 1.1 and an Act0.5 value of 5 mM NaHCO3 was obtained. 6-phosphogluconate appeared to saturate at or below 20 μmM.  相似文献   

3.
A single subcutaneous injection of folate, homofolate or MTX resulted in the inhibition of the activity of dihydrofolate reductase in homogenates prepared from the kidneys of normal mice. Stimulation of 3H-thymidine uptake occurred in the kidneys of treated animals approximately 30 hr after administration of either folate or homofolate, and reached a peak 72 hr after administration. The effects of folate and MTX on dihydrofolate reductase activity invivo were also determined. One hr after administration of 15 mg/kg methotrexate (MTX) or 300 mg/kg folate, enzyme activity invivo was inhibited by 90%.3H-deoxyuridine uptake was neither stimulated nor depressed after treatment with MTX. After administration of folate, uptake of 3H-deoxyuridine was stimulated at approximately 30 hr after drug-treatment and reached a peak at 72 hr after folate administration. Treatment with xanthopterin had no effect on the activity of dihydrofolate reductase invitro. Xanthopterin stimulated uptake of both deoxyuridine and thymidine in an identical manner.The increased DNA synthesis that occurs in animals after treatment with agents that cause renal damage is distinct from the effect these agents have upon dihydrofolate reductase. Nucleoside incorporation after treatment with folate, homofolate, MTX or xanthopterin cannot be predicted on the basis of enzyme inhibition. Treatment with MTX, folate or homofolate results in enzyme inhibition which is not correlated with the uptake of deoxyuridine into DNA.  相似文献   

4.
The enzyme related to the synthesis of 4,16- androstadien-3-one from progesterone was investigated using boar testis. To elucidate the activity, in the soluble form, the lyophilized powder of 12,000 g supernatant from homogenate was first treated with n-butanol after which the enzyme could be extracted with 1 mM ethylenediaminete-traacetic acid(EDTA), 1 mM dithiothreitol(DTT) and 20 % glycerol. The enzyme was stable in this medium.The enzyme filtered through a column of Sephacryl S-200 super fine gel exhibited requirements of NADPH-cytochrome C reductase and phosphatidylcholine for maximum enzymatic activity. The requirements of reductase and phosphatidylcholine were not observed in the crude extract fraction. The enzyme separated by column chromatography with DEAE-cellulose required phosphatidylcholine for the synthesis but the reductase had no effect. These lines of evidence suggest that the activity of the enzyme, as related to synthesis of 4,16-androstadien-3-one from progesterone might be regulated by phosphatidylcholine and reductase, in situ.  相似文献   

5.
Both the cytochrome b5 level and NADH cytochrome b5 reductase activity in rat liver microsomes were increased 2-fold by repeated i.p. administration of 1.5 mmol/kg propylthiouracil (PTU) for 2 weeks, but neither the cytochrome P-450 level nor NADPH cytochrome P-450 reductase activity were affected by the treatment. Liver microsomes from PTU-treated rats showed a significant decrease in aminopyrine N-demethylation, but not in benzphetamine N-demethylation, aniline hydroxylation or 7-ethoxycoumarin O-deethylation. A single administration of the compound had no effect on any components of the system. In vitro, drug hydroxylation activities were not affected by PTU up to 1.0 mM. From the above evidence, repeated administration of PTU selectively induced cytochrome b5 and NADH cytochrome b5 reductase in rat liver microsomes.  相似文献   

6.
F H Faas  W J Carter  J O Wynn 《Life sciences》1974,15(12):2059-2068
Rat liver microsomal NADH-cytochrome c reductase activity is stimulated by 20 μM thyroxine invitro. Thyroxine does not influence microsomal NADH-dichlorophenolindophenol reductase, NADPH-cytochrome c reductase, or NADPH-dichlorophenolindophenol reductase activity. Stimulation of NADH-cytochrome c reductase activity is not mediated by super-oxide and is likely due to enhanced reduction or oxidation of cytochrome b5.  相似文献   

7.
The transport of 3-O-methylglucose in white fat cells was measured under equilibrium exchange conditions at 3-O-methylglucose concentrations up to 50 mM with a previously described method (Vinten, J., Gliemann, J. and Østerlind, K. (1976) J. Biol. Chem. 251, 794–800). Under these conditions the main part of the transport was inhibitable by cytochalasin B. The inhibition was found to be of competitive type with an inhibition constant of about 2.5 · 10?7 M, both in the absence and in the presence of insulin (1μM). The cytochalasin B-insensitive part of the 3-O-methylglucose permeability was about 2 · 10?9 cm · s?1, and was not affected by insulin. As calculated from the maximum transport capacity, the half saturation constant and the volume/ surface ratio, the maximum permeability of the fat cell membrane to 3-O-methylglucose at 37°C and in the presence of insulin was 4.3 · 10?6 cm · s?1. From the temperature dependence of the maximum transport capacity in the interval 18–37°C and in the presence of insulin, an Arrhenius activation energy of 14.8 ± 0.44 kcal/mol was found. The corresponding value was 13.9 ± 0.89 in the absence of insulin. The half saturating concentration of 3-O-methylglucose was about 6 mM in the temperature interval used, and it was not affected by insulin, although this hormone increased the maximum transport capacity about ten-fold to 1.7 mmol · s?1 per 1 intracellular water at 37°C.  相似文献   

8.
By using fluorogenic peptidyl-3-amino-9-ethyl-carbazole a highly selective endopeptidase for the Val-Leu-Gly-Arg sequence was demonstrated in endoerythrocytic stages of Plasmodium berghei. Val-Leu-Gly-Arg-endopeptidase showed a maximum activity in pH range 7.0–8.0; it was completely inhibited by 1 mM leupeptin and 1 mM antipain. A complete inhibition was also obtained by 15 mM chloroquine. This trypsin-like activity was negligible in uninfected red blood cells. The high sensitive fluorogenic procedure could be performed on cell fractions, cell lysates as well as supernatants.  相似文献   

9.
The effect of pretreatment with phenobarbitone, rifampicin, β-naphthoflavone, antipyrine and spironolactone on the irreversible binding of ethynyloestradiol to guinea pig liver microsomes has been examined and the corresponding changes in microsomal P-450 content and cytochrome c reductase activity measured. Rifampicin produced the greatest increase (220%) in irreversible binding while phenobarbitone produced the greatest increase in both microsomal P-450 content (172%) and cytochrome c reductase activity (210%). There was no correlation of irreversible binding with either microsomal P-450 content or with cytochrome c reductase activity.  相似文献   

10.
The effect of Cl? and other anions on (3H)-noradrenaline line (NA) transport by bisected rat heart atrial appendages invitro has been studied. It was found that NA active transport, at the plasma membrane level, shows an absolute dependency on Cl?, with a half-maximal activation of transport occurring at 6 mM Cl? and complete saturation at 50 mM. Cl? effects are due to changes in transport Km, while Vmax is not changed. Only one class of sites for Cl? seem to be present in the transport system. Br? can substitute for Cl? with 90% effectiveness, nitrate and iodide are less effective, while larger anions are very poor substitutes. In addition, heart atrial hemi-appendages have been characterized as a suitable preparation for studies of this type.  相似文献   

11.
The activity of pure Escherichia coli murein (peptidoglycan) amidase (N-acetylmuramoyl-l-alanine amidase, EC 3.5.1.28) was measured after preincubation with E. coli phosphatidylglycerol microdispersions in final concentration ranging over micro- and millimolarities. The enzyme activity was increased up to 160% of the control for phosphatidylglycerol concentrations increasing from 2 to 50 μM. After a plateau extending from 0.05 to 0.3 mM, higher phosphatidylglycerol concentrations inactivated the enzyme down to 15% of initial activity for concentrations of 2 mM. Positive kinetic cooperativity was observed for the activation as well as for the inactivation processes. Cardiolipin (or diphosphatidylglycerol) from the same origin and under same conditions had no significant effect. Molecular sieving experiments have shown that, when inactivated, the enzyme remained firmly bound to the phosphatidylglycerol vesicles, whereas the activated phosphatidylglycerol-enzyme complex was totally dissociable by dilution. Activated phosphatidylglycerol complexes were recovered by gel exclusion chromatography at equilibrium in 40 μM phosphatidylglycerol. Possible physiological meaning of the results is briefly discussed in the context of our work and that done previously by others.  相似文献   

12.
Rabbit pulmonary alveolar macrophages produce a collagenase which lyses labeled collagen gels, specifically cleaves collagen types I, II and III, is inhibited by ethylenediaminetetraacetate, cysteine, dithiothreitol and serum but is not inhibited by a serine protease inhibitor. Alveolar macrophage collagenase activity can be enhanced by in vivo BCG activation, in vitro latex, silica or mycobacterium activation and by in vitro uncovering of latent enzymatic activity with trypsin treatment. The production of collagenase by unactivated alveolar macrophages and the presence of “latent” collagenase in culture media of alveolar macrophages are examples of significant differences between alveolar and peritoneal macrophages.  相似文献   

13.
Rainbow trout were treated with β-naphthoflavone and the hepatic microsomal cytochrome P-450 solubilized with 3-[(3-cholaminopropyl)dimethylammonio]-1-propanesulfonate. Chromatography on tryptamine-Sepharose 4B gave a single cytochrome P-450 peak which was further resolved into three components by elution from DEAE-Sepharose. The two main peaks were then chromatographed on hydroxyapatite and a total of four fractions obtained. Two of these fractions had similar properties and significantly metabolized [14C]benzo[a]pyrene in a reconstituted system containing rat cytochrome P-450 reductase. This activity was inhibited by α-naphthoflavone but not by metyrapone or SKF-525A. Purified cytochromes P-448 from 3-methylcholanthrene-treated rat had similar spectral properties and activity towards [14C]benzo[a]pyrene suggesting similarities between these forms.  相似文献   

14.
Benzoate 1,2-dioxygenase system which catalyzed double hydroxylation of benzoate was obtained from Pseudomonas arvilla and was shown to consist of two protein components (component A and B). Component A which was purified and was shown to be homogeneous upon sodium dodecyl sulfate disc gel electrophoresis retained high activity of NADH-cytochrome c reductase. Both of benzoate 1,2-dioxygenase activity and NADH-cytochrome c reductase activity were simultaneously induced by benzoate. Dichlorophenolindophenol which could serve as an electron acceptor of the NADH-cytochrome c reductase inhibited the activity of benzoate 1,2-dioxygenase. These results suggest the possibility that NADH-cytochrome c reductase activity is required for benzoate 1,2-dioxygenase.  相似文献   

15.
Antimycin, when added to resolved succinate-cytochrome c reductase complex in amounts sufficient to partially inhibit succinate-cytochrome c reductase activity, causes a decrease in inhibition of the residual succinate-cytochrome c reductase activity by 2-thenoyltrifluoroacetone. Antimycin has no effect on the inhibition of succinate-ubiquinone reductase activity by 2-thenoyltrifluoroacetone. We propose that antimycin increases the steady state concentration of ubisemiquinone in the reductase complex, and that 2-thenoyltrifluoracetone is competitive with ubisemiquinone.  相似文献   

16.
The activity of hydroxymethylglutaryl CoA reductase (NADPH) (EC 1.1.1.34) was studied in the latex of regularly tapped mature trees of Hevea brasiliensis. The reductase activity was found mainly (95% of the total activity) in the pellet fraction (40 000 g) of the centrifuged latex. The enzyme in this fraction had a specific requirement for NADPH as the cofactor and, while not obligatory for activity, was activated by dithiothreitol at the optimum concentration of 2 mM. The pH optimum was found to be 6.6–6.9 in 0.1 M phosphate buffer. Mevalonate and CoA (at 2 mM each) did not affect enzyme activity, while hydroxymethylglutarate (2 mM) was slightly inhibitory. p-Chloromercuribenzoate (1 mM) completely inhibited this enzyme. The reductase activity in the 40 000 g pellet was not easily solubilized either using Triton X-100 or by sonication. The apparent Km for the washed, membrane-bound enzyme (103 000 g pellet) was 56 μ M (RS-HMG-CoA). Magnesium-ATP (4 mM) inactivated the reductase but this effect was greatly diminished or was absent upon washing the 40 000 g pellet.  相似文献   

17.
Peter Nicholls 《BBA》1976,430(1):13-29
1. Formate inhibits cytochrome c oxidase activity both in intact mitochondria and submitochondrial particles, and in isolated cytochrome aa3. The inhibition increases with decreasing pH, indicating that HCOOH may be the inhibitory species.2. Formate induces a blue shift in the absorption spectrum of oxidized cytochrome aa3 (a3+a33+) and in the half-reduced species (a2+a33+). Comparison with cyanide-induced spectral shifts, towards the red, indicates that formate and cyanide have opposite effects on the aa3 spectrum, both in the fully oxidized and the half-reduced states. The formate spectra provide a new method of obtaining the difference spectrum of a32+ minus a33+, free of the difficulties with cyanide (which induces marked high → low spin spectral shifts in cytochrome a33+) and azide (which induces peak shifts of cytochrome a2+ towards the blue in both α- and Soret regions).3. The rate of formate dissociation from cytochrome a2+a33+-HCOOH is faster than its rate of dissociation from a3+a33+-HCOOH, especially in the presence of cytochrome c. The Ki for formate inhibition of respiration is a function of the reduction state of the system, varying from 30 mM (100% reduction) to 1 mM (100% oxidation) at pH 7.4, 30 °C.4. Succinate-cytochrome c reductase activity is also inhibited by formate, in a reaction competitive with succinate and dependent on [formate]2.5. Formate inhibition of ascorbate plus N,N,N′,N′-tetramethyl-p-phenyl-enediamine oxidation by intact rat liver mitochondria is partially released by uncoupler addition. Formate is permeable through the inner mitochondrial membrane and no differences in ‘on’ or ‘off’ inhibition rates were observed when intact mitochondria were compared with submitochondrial particles.6. NADH-cytochrome c reductase activity is unaffected by formate in submitochondrial particles, but mitochondrial oxidation of glutamate plus malate is subject both to terminal inhibition at the cytochrome aa3 level and to a slow extra inhibition by formate following uncoupler addition, indicating a third site of formate action in the intact mitochondrion.  相似文献   

18.
An anaerobic incubation period of varying duration is required to induce hydrogenase activity in C., reinhardtii. Inclusion of sodium acetate, a metabolizable carbonaceous substrate, in the medium during anaerobic incubation accelerates the activation process. Thus, in the presence of sodium acetate, hydrogen photoproduction is detected within 7 to 15 minutes after the onset of anaerobiosis. On the contrary, if an uncoupler of phosphorylation, such as CCCP or sodium arsenate, is present during anaerobic incubation, little activation of the hydrogenase is observed even after hours of anaerobic adaptation. Since the uncouplers had no inhibitory effect on hydrogen photoproduction by the alga when added to previously activated cells, they are not inhibitors of activated hydrogenase. The uncouplers interfere, most likely, with the activation of hydrogenase. Similar effects of uncouplers on the hydrogenase activation process were obtained using a cell-free assay of hydrogenase activity. These observations provide strong evidence that anaerobic activation of the hydrogenase is an energy requiring process.  相似文献   

19.
邓治  刘实忠  校现周 《广西植物》2010,30(6):876-880
通过丙酮沉淀、DEAE-纤维素离子交换柱层析和Sephadex G-100凝胶过滤柱层析等分离纯化技术,对巴西橡胶树胶乳C-乳清磷脂酶A2进行分离纯化。用SDS-PAGE测定其亚基的相对分子量。测定该酶最适温度和pH,动力学常数Km和Vmax。并测定Ca2+和La3+对酶活性的影响。结果显示:该酶被纯化了49.47倍,产率为5.12%。SDS-PAGE检测为单一条带,其亚基相对分子量约43kDa。最适反应温度为37℃,最适反应pH为8.0,Km为0.44mmol·L-1,Vmax为7.22μmol.(mL.min)-1。最适Ca2+浓度为50μmol·L-1,稀土元素La3+离子对磷脂酶A2活性有抑制作用,但加入Ca2+后可缓解La3+对磷脂酶A2活性的抑制作用。胶乳C-乳清磷脂酶A2与其他植物磷脂酶A2在Ca2+的依赖性上存在差异。研究结果为今后探索橡胶树胶乳磷脂酶A2的催化机理、调节机理及生理功能等奠定了基础。  相似文献   

20.
Pent-4-enoate at 0.1 to 1.0 mM strongly inhibited urea synthesis in isolated rat hepatocytes. Pent-4-enoate at the same concentrations markedly decreased concentrations of N-acetyl-L-glutamate, an essential activator of carbamoyl-phosphate synthase-I (EC 2.7.2.5), and the decrease was well parallel with the inhibition of urea synthesis by pent-4-enoate. This compound also lowered cellular concentrations of acetyl-CoA, a substrate of acetylglutamate synthase (EC 2.3.1.1). Pent-4-enoate in a dose of 1 mM did not significantly affect cellular concentrations of ATP, and had no direct effect on acetylglutamate synthase activity. These results suggest that the inhibition of urea synthesis by pent-4-enoate is due to decrease in N-acetyl-L-glutamate concentration and that the decrease is probably brought about by decreased rate of its synthesis due to the lowered concentration of cellular acetyl-CoA.  相似文献   

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