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1.
Peroxisomes were isolated from pea ( Pisum sativum L.) leaves and the peroxisomal membranes were purified by treatment with Na 2CO 3. The production of superoxide radicals (O 2-) induced by NADH was investigated in peroxisomal membranes from intact organelles incubated with proteases (pronase E and proteinase K). Under isoosmotic conditions, in the presence of pronase E, the production of O 2- radicals was inhibited by 80%. SDS-PAGE of peroxisomal membranes after protease treatment demonstrated a decrease in the 18-kDa PMP. This suggests that this polypeptide has a small fragment exposed to the cytosolic side of the peroxisomal membrane which is essential for O 2-production. The 18-kDa PMP was purified by preparative SDS-PAGE and in the reconstituted protein the NADH-driven production of O 2- radicals was investigated. The isolated polypeptide showed a high generation rate of superoxide (about 300 nmol O 2- × mg- 1 protein × min -1) which was completely inhibited by 50 mM pyridine. The 18-kDa PMP was recognized by a polyclonal antibody against Cyt b5 from human ery-throcytes. The presence of b-type cytochrome in peroxisomal membranes was demonstrated by difference spectroscopy. Results obtained show that in the NADH-dependent O 2- radical generating system of peroxisomal membranes, the 18-kDa integral membrane polypeptide, which appears to be Cyt b 5, is clearly involved in superoxide radical production. 相似文献
2.
The in vivo production of HO- requires iron ions, H 2O 2 and O 2- or other oxidants but probably does not occur through the Haber-Weiss reaction. Instead oxidants, such as O 2-, increase free iron by releasing Fe(II) from the iron-sulfur clusters of dehydratases and by interfering with the iron-sulfur clusters reassembly. Fe(II) then reduces H 2O 2, and in turn Fe(III) and the oxidized cluster are re-reduced by cellular reductants such as NADPH and glutathione. In this way, SOD cooperates with cellular reductants in keeping the iron-sulfur clusters intact and the rate of HO- production to a minimum.
O 2- and other oxidants can release iron from Fe(II)-containing enzymes as well as copper from thionein. The released Fe(III) and Cu(II) are then reduced to Fe(II) and Cu(I) and can then participate in the Fenton reaction.
In mammalian cells oxidants are able to convert cytosolic aconitase into active IRE-BP, which increases the “free” iron concentration intracellularly both by decreasing the biosynthesis of ferritin and increasing biosynthesis of transferrin receptors.
The biological role of the soxRS regulon of Escherichia coli, which is involved in the adaptation toward oxidative stress, is presumably to counteract the oxidative inactivation of the iron clusters and the subsequent release of iron with consequent increased rate of production of HO. 相似文献
3.
Of production by homogenates and isolated membranes of E. coli has been examined. Approximately one-fourth of the O 2-generated by extracts in the prescence of NAD (P) H is attributable to the membranes. The autoxidizable membrane component is a member of the respiratory chain, since O 2-production is NADH-specific, amplified by cyanide, and absent from membranes lacking the respiratory NADH dehyd-rogenase. Other respiratory substrates (succinate, I -phosphoglycerol, D-lactate. and L-lactate) supported Or production at efficiencies between 3 and 30 O 2-released per 10.000 electrons transferred, under conditions of substrate saturation.
Membranes from quinoneless mutants quantitatively retain the ability to evolve O 2-. indicating that the dehydrogenases are the sites of O 2-production. Relative O 2-production was greater at low substrate concentrations, probably reflecting the facilitation of unpairing of electrons that may occur when enzymes with multiple redox centers are only partially reduced.
Respiration rate, cell volume, rates of membraneous and cytosolic O 2-production, and SOD levels were used to calculate a steady-state concentration of O 2-between 10 -- 10 and 10 -- 9 M in well-fed, aerobic, SOD-proficient cells. 相似文献
4.
在CO2浓度分别为当今CO2浓度(360 μL/L)和加富浓度(5 000 μL/L)条件下,研究了UV-B胁迫对亚心形扁藻(Platymonas subcordiformis(Wille)Hazen)的光合作用、膜脂过氧化和抗氧化酶活性的影响.实验结果表明:(1)UV-B单独作用下,亚心形扁藻的干重、光合速率、叶绿素a(Chl a)和类胡萝卜素(Car.)含量显著降低,CO2加富单独作用下,亚心形扁藻的干重和光合速率显著升高,叶绿素a和类胡萝卜素含量与对照相比没有显著变化,而UV-B与CO2共同作用则使亚心形扁藻的干重和光合速率与对照相比没有显著变化,叶绿素a和类胡萝卜素含量显著降低.(2)UV-B单独作用和CO2加富单独作用都使可溶性蛋白含量显著降低,UV-B与CO2共同作用下的可溶性蛋白含量比UV-B单独作用的要高.高CO2对藻的可溶性蛋白含量的变化在很大程度上归因于Rubisco蛋白的降低.(3)UV-B单独作用下,O-.2产生速率、H2O2含量和MDA含量显著升高,而CO2加富单独作用下,O-.2产生速率、H2O2含量和MDA含量显著降低,与UV-B单独作用相比,UV-B与CO2共同作用使O-.2产生速率、H2O2含量和MDA含量显著降低.说明CO2加富可以减少活性氧对亚心形扁藻的氧化胁迫,同时减少UV-B对亚心形扁藻的膜脂过氧化伤害.(4)UV-B单独作用下,SOD、POD、CAT、GR和GPx活性显著升高,高CO2单独作用使SOD、POD和GR活性显著降低,而CAT和GPx活性与对照相比稍有所降低,但降低不明显,而UV-B与CO2共同作用则使SOD、POD、CAT、GR和GPx活性比UV-B单独作用少得多.结果表明,高CO2对UV-B胁迫所造成的氧化胁迫具有一定的改善作用,因此CO2浓度升高可能对增强海洋微藻的抗逆能力有利. 相似文献
5.
It is shown that the copper zinc superoxide dismutase is unique in its ability to catalyze O 2- dismutation in vivo in contrast to other copper compounds which have this feature only in vitro. The reasons for this difference are discussed in terms of kinetic and thermodynamic parameters. 相似文献
6.
The polymorphonuclear leukocyte secretes both O 2-and H 2O 2 when stimulated by various soluble or particulate stimuli. Since a rcaction involving iron, O 2-, and H 2O 2 could generate the hydroxyl radical (HO.) there has been speculation that the HO-may participate in the bactericidal activity of the neutroph-il. A variety of water-soluble HO. scavengers have been used to test for the participation of HO. and the results imply that HO. might participate. However, other workers have not been able to detect the formation of significant amounts of HO-by the activated neutrophil. We have examined the effect of several commonly used HO. radical scavengers on the ability of the neutrophil to secrete O 2-and H 2O 2. Several of these compounds actively inhibit secretion without affecting the viability of the neutrophil. After considering the various complications inherent in using water soluble radical scavengers, we suggest that they only be used with well defined experimental systems. 相似文献
8.
Human neutrophils (PMN) activated by N-formyl-methionyl-leucyl-phenylalanine (fMLP) simultaneously release nitric oxide (.NO), superoxide anion (O 2-) and its dismutation product, hydrogen peroxide (H 2O 2). To assess whether NO production shares common steps with the activation of the NADPH oxidase, PMN were treated with inhibitors and antagonists of intracellular signaling pathways and subsequently stimulated either with fMLP or with a phorbol ester (PMA). The G-protein inhibitor, pertussis toxin (1-10 μg/ml) decreased H 2O 2 yield without significantly changing. NO production in fMLP-stimulated neutrophils; no effects were observed in PMA-activated cells. The inhibition of tyrosine kinases by genistein (1-25 μg/ml) completely abolished H 2O 2 release by fMLP-activated neutrophils; conversely, NO production increased about 1.5- and 3-fold with fMLP and PMA, respectively. Accordingly, orthovanadate, an inhibitor of phosphotyrosine phosphatase, markedly decreased -NO production and increased O 2;- release. On the other hand, inhibition of protein kinase C with staurosporine and the use of burst antagonists like adenosine, cholera toxin or dibutyryl-cAMP diminished both H 2O 2 and NO production. The results suggest that the activation of the tyrosine kinase pathway in stimulated human neutrophils controls positively O 2- and H 2O 2 generation and simultaneously maintains -NO production in low levels. In contrast, activation of protein kinase C is a positive modulator for O 2;-and *NO production. 相似文献
9.
Isolated hepatocytes incubated with selenite (30–100 μM) exhibited changes in the glutathione redox system as shown by an increase in O 2 consumption, oxidation of glutathione and loss of NADPH. Selenite (50 μM) raised O 2 consumption within the 1 h and induced an partial depletion of thiols with a concomitant increase in oxidized glutathione, as well as a decrease in NADPH levels within 2 h. With 100 μM selenite more pronounced effects were obtained such as a total depletion of thiols. This concentration of selenite also lysed cells within 3 h. Arsenite, HgCl 2 and KCN prevented the increase in O 2 uptake, counteracted loss of thiols and delayed selenite induced lysis. p-Tert-butylbenzoic acid, an inhibitor of gluconeogenesis, decreased selenite dependent O 2 consumption and potentiated the effect on NADPH levels as well as the toxic effect. Finally, methionine further enhanced O 2 consumption by selenite and also delayed loss of thiols and potentiated selenite toxicity. These results indicated that selenite catalyzed a reduction of O 2 in glutathione dependent redox cycles with NADPH as an electron donor. With subtoxic concentrations of selenite (50 μM) there were indications that O 2 reduction was terminated by selenite biotransformation to methylated metabolites. With toxic concentrations of selenite (100 μM) it appeared that O 2 reduction was eventually limited by the capacity of the cell to regenerate NADPH. It is suggested that a depletion of NADPH mediated the observed cytotoxicity of selenite. 相似文献
10.
The relationship between O 2 and an active oxygen scavenging system in Chlorella vulgaris var. vulgaris (IAM C-534) was investigated. When Chlorella vulgaris was exposed to 2% O 2, only traces of active oxygen scavenging enzymes were found. When the Chlorella vulgaris was treated with 20% or 50% O 2, it was shown that the level of enzyme activity increased as the O 2 concentration increased. An increase in enzyme activity was not found in any specific enzyme but in all of the enzymes, but the level of glutathione and ascorbate remained the same in all the cases. In addition, the photosynthetic efficiency also decreased as the concentration of O 2 was increased. These results suggest that an O 2 enriched environment can lead to an increase in the production of active oxygen species such as O bullet2 and H 2O 2 and to a decrease in the photosynthetic efficiency in Chlorella vulgaris. The hydroxyl radical ( bulletOH) was detected directly in the Chlorella vulgaris suspension with a spin trapping reagent. It was also clear that the increase in the bulletOH intensity as the visible light intensity increased was unrelated to the O 2 concentration. It was suggested that the conditions for producing bulletOH and the other active oxygen species were different, and that two types of oxygen stress should exist in the Chlorella vulgaris. 相似文献
11.
1. 1. The mechanism of the photooxidation of ascorbate and of Mn2+ by isolated chloroplasts was reinvestigated. 2. 2. Our results suggest that ascorbate or Mn2+ oxidation is the result of the Photosystem I-mediated production of the radical superoxide, and that neither ascorbate nor Mn2+ compete with water as electron donors to Photosystem II nor affect the rate of electron transport through the two photosystems: The radical superoxide is formed as a result of the autooxidation of the reduced forms of low potential electron acceptors, such as methylviologen, diquat, napthaquinone, or ferredoxin. 3. 3. In the absence of ascorbate or Mn2+ the superoxide formed dismutases either spontaneously or enzymatically producing O2 and H2O2. In the presence of ascorbate or Mn2+, however, the superoxide is reduced to H2O2 with no formation of O2. Consequently, in the absence of reducing compounds, in the reaction H2O to low potential acceptor one O2 (net) is taken up per four electrons transported where as in the presence of ascorbate, Mn2+ or other suitable reductants up to three molecules O2 can be taken up per four electrons transported. 4. 4. This interpretation is supported by the following observations: (a) in a chloroplast-free model system containing NADPH and ferredoxin-NADP reductase, methylviologen can be reduced to a free radical which is autooxidizable in the presence of O2; the addition of ascorbate or Mn2+ to this system results in a two fold stimulation of O2 uptake, with no stimulation of NADPH oxidation. The stimulation of O2 uptake is inhibited by the enzyme superoxide dismutase; (b) the stimulation of light-dependent O2 uptake in the system H2O → methylviologen in chloroplasts is likewise inhibited by the enzyme superoxide dismutase. 5. 5. In Class II chloroplasts in the system H2O → NADP upon the addition of ascorbate or Mn2+ an apparent inhibition of O2 evolution is observed. This is explained by the interaction of these reductants with the superoxide formed by the autooxidation of ferredoxin, a reaction which proceeds simultaneously with the photoreduction of NADP. Such an effect usually does not occur in Class I chloroplasts in which the enzyme superoxide dismutase is presumably more active than in Class II chloroplasts. 6. 6. It is proposed that since in the Photosystem I-mediated reaction from reduced 2,4-dichlorophenolindophenol to such low potential electron acceptor as methylviologen, superoxide is formed and results in the oxidation of the ascorbate present in the system, the ratio ATP/2e in this system (when the rate of electron flow is based on the rate of O2 uptake) should be revised in the upward direction.
Abbreviations: DCMU, 3-(3′,4′-dichlorophenyl)-1,1-dimethylurea; HEPES, hydroxyethyl-piperazineethanesulfonic acid; MES, (N-morpholino)ethanesulfonic acid; DCIP, 2,4-dichlorophenol-indophenol 相似文献
12.
The reduction of ferricytochrome c by O 2− and CO 2− was studied in the pH range 6.6–9.2 and Arrhenius as well as Eyring parameters were derived from the rate constants and their temperature dependence. Ionic effects on the rate indicate that the redox process proceeds through a multiply-positively charged interaction site on cytochrome c. It is shown that the reaction with O 2− and correspondingly with O 2 of ferrocytochrome c) is by a factor of approx. 10 3 slower than warranted by factors such as redox potential. Evidence is adduced to support the view that this slowness is connected with the role of water in the interaction between O 2−/O 2 and ferri-ferrocytochrome c in the positively charged interaction site on cytochrome c in which water molecules are specifically involved in maintaining the local structure of cytochrome c and participate in the process of electron equivalent transfer. 相似文献
13.
The protein IF 1 is a natural inhibitor of the mitochondrial F oF 1-ATPase. Many investigators have been prompted to identify the shortest segment of IF 1, retaining its native activity, for use in biomedical applications. Here, the activity of the synthetic peptides IF 1-(42–58) and IF 1-(22–46) is correlated to their structure and conformational plasticity determined by CD and [ 1H]-NMR spectroscopy. Among all the IF 1 segments tested, IF 1-(42–58) exerts the most potent, pH and temperature dependent activity on the F oF 1 complex. The results suggest that, due to its flexible structure, it can fold in helical and/or β-spiral arrangements that favor the binding to the F oF 1 complex, where the native IF 1 binds. IF 1-(22–46), instead, as it adopts a rigid -helical conformation, it inhibits ATP hydrolysis only in the soluble F 1 moiety. 相似文献
14.
In dimethylformamide superoxide ion forms a l:l adduct with tctrakis (2.6-dichlorophenyl) porphinatoiron, (Cl 8 TPP)FeOO -, as well as with its manganese analogue, (Cl 8 TPP)MnOO -. On the basis of their electrochemical, spectroscopic, and magnetic properties these adducts have a metal-oxygen covalent bond (PorM-OO -), oxygen-centered redox chemistry. and reactivities that are similar to the hydroperoxide ion (HOO -). Addition of -OH to a solution of PorFe and O 2 results in the formation of PorFe(OH)(OO -), which can be electrochemically oxidized to PorFeOH plus O 2 (-0.2 V vs SCE). Addition of protons to the PorM-OO - adducts promotes their rapid decomposition to PorM, HOOH. and O 2. This chemistry provides insight to the reactions of biological superoxide and superoxide dismutases. 相似文献
15.
Phagocytic cells such as neutrophils generate superoxide anions (O 2−) within phagocytic vacuoles for killing and digesting microorganisms. Here we report the simultaneous observation of morphological changes and O 2− generation in single phagocytic cells during phagocytosis. Point stimulation of a cell by contact with an opsonized microelectrode at the cell surface induced significant deformation to engulf the electrode, and also induced the O 2− generation which was measured by the electrode. Periodic fluctuations in the magnitude of the O 2− generation were observed in the time course. These oscillations may be caused by metabolic regulation of the formation of NADPH, which is the substrate for the O 2− generation. 相似文献
16.
Vanadate stimulates the oxidation of NAD(P)H by biological membranes because such membranes contain NAD(P)H oxidases which are capable of reducing dioxygen to O 2− and because vanadate catalyzes the oxidation of NAD(P)H by O 2−, by a free radical chain mechanism. Dihydropyridines, such as reduced nicotinamide mononucleotide (NMNH), which are not substrates for membrane-associated NAD(P)H oxidases, are not oxidized by membranes plus vanadate unless NAD(P)H is present to serve as a source of O 2−. When [NMNH] greatly exceeds [NAD(P)H], in such reaction mixtures, one can observe the oxidation of many molecules of NMNH per NAD(P)H consumed. This reflects the chain length of the free radical chain mechanism. We have discussed the mechanism and significance of this process and have tried to clarify the pertinent but confusing literature. 相似文献
17.
A study of the blood oxygenation in pulmonary capillaries is made by considering the transport mechanisms of molecular diffusion, convection and the facilitated diffusion due to the presence of haemoglobin. The resistance offered by the pulmonary membrane on the transport of gases has been incorporated. The resulting system of coupled, non-linear partial differential equations is solved numerically. It is found that, in the immediate neighbourhood of the entry, the amount of dissolved O2 decreases. This decreases further as the resistance offered by the pulmonary membrane increases. The rate of oxygenation of blood increases as the permeability coefficient for O2(Po) increases. It is shown that the ideally permeable case for both O2 and CO2 can be approximated by taking Po ˜ 10 cm/s. Further, it is shown that the oxygen takes longest and CO2 is the fastest to attain equilibration. The equilibration length increases as the resistance offered by the membrane increases. Finally, some of the pulmonary diseases such as pulmonary oedema and fibrosis have been analyzed. 相似文献
19.
Influence of oxygen on the electron transfers of photosynthesis. I. Influence of some oxygen concentrations on some Hill reactionsThe influence of O2 concentrations on the Hill reactions in the presence of p-benzoquinone, ferricyanide, NADP+, NADP+ plus ferredoxin has been studied with isolated spinach chloroplasts. Because of the partial reoxidation of the hydroquinone, which is depending upon the O2 concentration, it does not seem possible to localize a site of action for O2. With ferricyanide the influence of O2 is weak. However, the rate of ferricyanide reduction is increased in the presence of O2. The observed stimulation is greater for 21% O2 than for 70% O2. Bicarbonate stimulates the ferricyanide reduction and decreases the stimulating effect of 21% O2. O2 decreases the rate of NADP+ reduction. Ferredoxin as well as bicarbonate stimulate the NADP+ reduction and reduce the O2 inhibition. These results seem to indicate that O2 may enter the electron transport chain at a site situated near Photosystem I and before the ferredoxin's site. The inhibitory effect of O2 on the Hill reactions with p-benzoquinone and NADP+ is depending upon the plants' growth conditions. It is greater with plants grown under weak light. 相似文献
20.
通过开顶箱(OTCs)模拟,以环境臭氧(O 3)浓度约40 nmol·mol -1为对照,研究大气O 3浓度升高(80和160 nmol·mol -1O 3)对冷季型草坪草高羊茅生长、亚细胞结构及其活性氧代谢的影响.结果表明: 14 d的80 nmol·mol -1O 3熏蒸使高羊茅株高和叶宽降低,总生物量降低43.7%,老叶变黄,而160 nmol·mol -1O 3处理高羊茅叶出现大量枯死褐斑,叶尖坏死,新叶卷曲,总生物量降低46.2%,叶肉细胞膜卷曲,叶绿体和线粒体受损严重.与对照相比,80和160 nmol·mol -1O 3熏蒸下高羊茅叶片超氧阴离子(O 2-·)产生速率、过氧化氢(H 2O 2)和丙二醛(MDA)含量显著增加,抗氧化酶活性显著升高,但叶片总酚含量和抗氧化能力随O 3浓度升高而先升高后降低.在明显O 3伤害症状出现之前,O 3已对高羊茅的生长和抗氧化代谢产生不利影响;高羊茅抗氧化系统虽对O 3浓度的升高存在一定的适应性反应,但其不能抵御过高浓度的长期胁迫和伤害. 相似文献
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