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1.
Glutathione reductase (GR) is an enzyme that recycles a key cellular antioxidant molecule glutathione (GSH) from its oxidized form (GSSG) thus maintaining cellular redox homeostasis. A recombinant plasmid to overexpress a GR of Brassica rapa subsp. pekinensis (BrGR) in E. coli BL21 (DE3) was constructed using an expression vector pKM260. Expression of the introduced gene was confirmed by semiquantitative RT-PCR, immunoblotting and enzyme assays. Purification of the BrGR protein was performed by IMAC method and indicated that the BrGR was a dimmer. The BrGR required NADPH as a cofactor and specific activity was approximately 458 U. The BrGR-expressing E. coli cells showed increased GR activity and tolerance to H2O2, menadione, and heavy metal (CdCl2, ZnCl2 and AlCl2)-mediated growth inhibition. The ectopic expression of BrGR provoked the co-regulation of a variety of antioxidant enzymes including catalase, superoxide dismutase, glutathione peroxidase, and glucose-6-phosphate dehydrogenase. Consequently, the transformed cells showed decreased hydroperoxide levels when exposed to stressful conditions. A proteomic analysis demonstrated the higher level of induction of proteins involved in glycolysis, detoxification/oxidative stress response, protein folding, transport/binding proteins, cell envelope/porins, and protein translation and modification when exposed to H2O2 stress. Taken together, these results indicate that the plant GR protein is functional in a cooperative way in the E. coli system to protect cells against oxidative stress.  相似文献   

2.
Oxidative stress causes damage to proteins, lipids and nucleic acids, and thereby compromises cell viability. Some of the oxidative stress markers in an eukaryotic model organism, fission yeast Schizosaccharomyces pombe, were evaluated in this study. Intracellular oxidation, protein carbonyls, lipid peroxidation and reduced glutathione (GSH) levels were investigated in H2O2-treated and non-treated control cells. It was observed that increased H2O2 concentration proportionally lowered the cell number and increased the intracellular oxidation, lipid peroxidation and protein carbonyl levels in S. pombe. A dose-dependent decrease in GSH level was also detected. The fission yeast S. pombe is best known for its contribution to understanding of eukaryotic cell cycle control. S. pombe displays a different physiology from Saccharomyces cerevisiae in several ways and is thus probably more closely related to higher eukaryotes. The purpose of this study was to provide some data about the effects of hydrogen peroxide on the proteins and lipids in the fission yeast. The data obtained here is expected to constitute a basis for the further studies on redox balance and related processes in yeast and mammalian cells.  相似文献   

3.
Peroxiredoxins (Prxs) are ubiquitous thiol-specific antioxidant enzymes that are critically involved in cell defense and protect cells from oxidative damage. In this study, a putative Type II Prx (ThPrx1) was identified and characterized from Tamarix hispida. The expression of ThPrx1 is highly induced in response to hydrogen peroxide (H2O2) and methyl viologen (MV) stresses. When expressed ectopically, ThPrx1 showed enhanced tolerance against oxidative stress in yeast and Arabidopsis. In addition, transgenic Arabidopsis plants overexpressing ThPrx1 displayed improved seedling survival rates and increased root growth and fresh weight gain under H2O2 and MV treatments. Moreover, transgenic Arabidopsis plants showed decreased accumulation of H2O2, superoxide (O2??) and malondialdehyde (MDA), increased superoxide dismutase (SOD) activity compared to wild-type (WT) plants under oxidative stress. Moreover, transgenic plants maintained higher photosynthesis efficiency and lower electrolyte leakage rates than that of WT plants under stress conditions. These results clearly indicated that ThPrx1 plays an important role in cellular redox homeostasis under stress conditions, leading to the maintenance of membrane integrity and increased tolerance to oxidative stress.  相似文献   

4.
Lee SC  Hwang IS  Hwang BK 《Planta》2011,234(6):1111-1125
Proteomics facilitates our understanding of cellular processes and network functions in the plant defense response during abiotic and biotic stresses. Here, we demonstrate that the ectopic expression of the Capsicum annuum antimicrobial protein CaAMP1 gene in Arabidopsis thaliana confers enhanced tolerance to methyl viologen (MV)-induced oxidative stress, which is accompanied by lower levels of lipid peroxidation. Quantitative comparative proteome analyses using two-dimensional gel electrophoresis coupled with mass spectrometry identified some of the oxidative stress- and disease-related proteins that are differentially regulated by CaAMP1 overexpression in Arabidopsis leaves. Antioxidant- and defense-related proteins, such as 2-cys peroxiredoxin, l-ascorbate peroxidase, peroxiredoxin, glutathione S-transferase and copper homeostasis factor, were up-regulated in the CaAMP1 transgenic leaf tissues. In contrast, GSH-dependent dehydroascorbate reductase and WD-40 repeat family protein were down-regulated by CaAMP1 overexpression. In addition, CaAMP1 overexpression enhanced resistance to Pseudomonas syringae pv. tomato (Pst) DC3000 infection and also H2O2 accumulation in Arabidopsis. The identified antioxidant- and defense-related genes were differentially expressed during MV-induced oxidative stress and Pst DC3000 infection. Taken together, we conclude that CaAMP1 overexpression can regulate the differential expression of defense-related proteins in response to environmental stresses to maintain reactive oxygen species (ROS) homeostasis.  相似文献   

5.
N-Acetyltransferase Mpr1 of Saccharomyces cerevisiae can reduce intracellular oxidation levels and protect yeast cells under oxidative stress, including H2O2, heat-shock, or freeze-thaw treatment. Unlike many antioxidant enzyme genes induced in response to oxidative stress, the MPR1 gene seems to be constitutively expressed in yeast cells. Based on a recent report that ethanol toxicity is correlated with the production of reactive oxygen species (ROS), we examined here the role of Mpr1 under ethanol stress conditions. The null mutant of the MPR1 and MPR2 genes showed hypersensitivity to ethanol stress, and the expression of the MPR1 gene conferred stress tolerance. We also found that yeast cells exhibited increased ROS levels during exposure to ethanol stress, and that Mpr1 protects yeast cells from ethanol stress by reducing intracellular ROS levels. When the MPR1 gene was overexpressed in antioxidant enzyme-deficient mutants, increased resistance to H2O2 or heat shock was observed in cells lacking the CTA1, CTT1, or GPX1 gene encoding catalase A, catalase T, or glutathione peroxidase, respectively. These results suggest that Mpr1 might compensate the function of enzymes that detoxify H2O2. Hence, Mpr1 has promising potential for the breeding of novel ethanol-tolerant yeast strains.  相似文献   

6.
The objective of the present study was to determine the influence of potassium deprivation on the halophyte species Hordeum maritimum grown in hydroponics for 2 weeks. Treatments were with potassium (+K) or without potassium (−K). Growth, water status, mineral nutrition, parameters of oxidative stress [malondialdehyde (MDA), carbonyl groups (C=O), and hydrogen peroxide concentration (H2O2) contents], antioxidant enzyme activities [superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6), guaiacol peroxidase (GPX, EC 1.11.1.7), ascorbate peroxidase (APX, EC 1.11.1.11), monodehydroascorbate peroxidase (MDHAR, EC 1.6.5.4), dehydroascorbate peroxidase (DHAR, EC 1.8.5.1), and glutathione reductase (GR, EC 1.6.4.2)], and antioxidant molecules [ascorbate (ASC), and glutathione (GSH)] were determined. Results showed that the growth of vegetative organs decreased owing to potassium deficiency with roots (−36%) more affected than shoots (−12%). Water status was only diminished in roots (reduction of 24%). Potassium deprivation decreased potassium concentration in both organs, this decrease was more pronounced in roots (−81%) than in shoots (−55%). In contrast to carbonyl groups, MDA content increased owing to potassium deprivation. Except for CAT activity that remained unaffected; SOD, GPX, APX, GR, MDHAR, and DHAR activities were significantly increased. H2O2 concentration was negatively correlated with the activities of enzymes and the accumulation of non-enzymatic antioxidants implicated in its detoxification. In conclusion, a cooperative process between the antioxidant systems is important for the tolerance of H. maritimum to potassium deficiency.  相似文献   

7.

Objectives

To improve the oxidative stress tolerance, biomass yield, and ascorbate/dehydroascorbate (AsA/DHA) ratio of Synechococcus elongatus PCC 7942 in the presence of H2O2, by heterologous expression of the dehydroascorbate reductase (DHAR) gene from Brassica juncea (BrDHAR).

Results

Under H2O2 stress, overexpression of BrDHAR in the transgenic strain (BrD) of S. elongatus greatly increased the AsA/DHA ratio. As part of the AsA recycling system, the oxidative stress response induced by reactive oxygen species was enhanced, and intracellular H2O2 level decreased. In addition, under H2O2 stress conditions, the BrD strain displayed increased growth rate and biomass, as well as higher chlorophyll content and deeper pigmentation than did wild-type and control strains.

Conclusion

By maintaining the AsA pool and redox homeostasis, the heterologous expression of BrDHAR increased S. elongatus tolerance to H2O2 stress, improving the biomass yield under these conditions. The results suggest that the BrD strain of S. elongatus, with its ability to attenuate the deleterious effects of ROS caused by environmental stressors, could be a promising platform for the generation of biofuels and other valuable bioproducts.
  相似文献   

8.
In order to elucidate the response of the ascorbate-glutathione (ASC-GSH) cycle to drought stress, the activities of antioxidant enzymes and the levels of molecules involved in the ASC-GSH metabolism were studied in Trifolium repens L. seedlings subjected to PEG-induced water deficit. Compared to the control, the contents of H2O2, thiobarbituric acid reactive substances (TBARS), ascorbate (ASC), dehydroascorbate (DHA), and glutathione disulfide (GSSG) increased in PEG-treated seedlings, whereas the glutathione (GSH) content kept constant during the drought period. Further more, the ASC/DHA and GSH/GSSG ratios decreased in the presence of PEG. Except for that of monodehydroascorbate reductase (MDHAR), the activities of ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR), and glutathione reductase (GR) were up-regulated during water deficit, and the increases in APX and DHAR activities were much higher than those in GR activity. These data indicate that fluctuations in the ASC-GSH metabolism resulted from PEG treatment may have a positive effect on drought stress mitigation in T. repens.  相似文献   

9.
We studied how plant cell modulated redox homeostasis in cytoplasmic male-sterility (CMS) Brassica juncea. The CMS Brassica juncea was identified to be mutated in several mitochondrial genes that suggested the changes of cell redox homeostasis. We observed that it was not associated with increased oxidative stress as shown by decreased H2O2 and OH contents in this type of CMS. The expressions of several anti-oxidative genes were up-regulated in 5-day-old seedlings of CMS than MF lines under light and dark conditions. The mitochondrial alternative oxidase pathway was not activated, as indicated by no increased expression of AOX1a gene in CMS. Interestingly, the expression of Ferritin1 gene was markedly activated in 5-day-old seedlings of CMS than MF line under light and dark conditions. Consequently, we detected increased content of total iron in 30-day-old leaves in CMS than MF line. We isolated Ferritin1 orthologous gene from Brassica juncea, which was targeted to the chloroplast and induced by Fe-citrate and H2O2, not ABA. Taken together, we proposed that increased expressions of BjFer1 and several antioxidant genes protected cell from oxidative stress in CMS Brassica juncea.  相似文献   

10.
In order to investigate the effects of spermidine (Spd) and spermine (Spm) on cadmium stress, the content of chlorophyll, hydrogen peroxide (H2O2), malondialdehyde (MDA), soluble protein and proline, the rate of O2·− generation, and activities of antioxidant enzymes (superoxide dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX), and glutathione reductase (GR)) in Potamogeton malaianus Miq. were measured. Exogenous application of Spd or Spm significantly enhanced the level of proline, retarded the loss of chlorophyll, enhanced photosynthesis, decreased the rate of O2·− generation and H2O2 content, and prevented Cd-induced lipid peroxidation. Spd and Spm also effectively maintained the balance of antioxidant enzyme activities under Cd stress; however, GR activity was found to increase only slightly in response to polyamines (PAs). The antioxidant systems, which were modified by PAs, were able to moderate the radical-scavenging system and to lessen in this way the oxidative stress. These results suggest that both Spd and Spm can enhance Cd tolerance of P. malaianus.  相似文献   

11.
Three AtHSP90 isoforms, cytosol-localized AtHSP90.2, chloroplast-localized AtHSP90.5, and endoplasmic reticulum (ER)-localized AtHSP90.7 genes, were constitutively overexpressed in Arabidopsis thaliana to study their functional mechanisms under oxidative stress. Overexpression of AtHSP90 genes reduced germination of transgenic seeds under oxidative stress. When exposed to 10 mM H2O2, AtHSP90 transgenic seedlings displayed lower activities of superoxide dismutase, catalase, and peroxidase; higher content of malondialdehyde; and higher levels of protein damage than detected in the wild type. This indicated that overexpression of AtHSP90.2, AtHSP90.5, and AtHSP90.7 in Arabidopsis impaired plant tolerance to oxidative stress. Moreover, overexpression of chloroplast- and ER-localized AtHSP90 resulted in lower resistance to oxidative stress than that of cytosolic AtHSP90. This suggested that HSP90.2, HSP90.5, and HSP90.7 localized in different cellular compartments were involved in different functional mechanisms during oxidative stress.  相似文献   

12.
The expression of the rpoS gene during PHA depolymerization was monitored in Pseudomonas oleovorans GPo1 and its mutant defective in PHA degradation by analyzing the tolerance to oxidative and thermal stresses and the RpoS intracellular content. An increase in the tolerance to H2O2 and heat shock was observed coincidentally with PHA degradation. Western blotting experiments performed in carbon-starved cultures showed that the RpoS levels were higher in the wild type than in the mutant strain. Complementation of the phaZ mutation restores the wild-type RpoS levels. These results suggest a probable association between PHA depolymerization and the stress tolerance phenotype controlled by RpoS.  相似文献   

13.
14.
Two cultivars of peanut (Arachis hypogaea L.) which were designated as resistant (Florispan) and sensitive (Gazipasa) according to their growth retardation under drought stress conditions were compared for their oxidative damage and antioxidant responses. Sixteen days-old peanut seedlings were subjected to PEG-6000 solutions of two different osmotic potentials; −0.4 and −0.8 MPa, and various growth parameters, photosystem II activity, changes in malondialdehyde (MDA), hydrogen peroxide (H2O2) and proline levels, activities of ascorbate peroxidase (APX), catalase (CAT), peroxidase (POX) and gluthatione reductase (GR) enzymes were determined. Both cultivars exhibited water deficit at −0.8 MPa osmotic potential of PEG-6000 and H2O2 levels significantly increased during exposure to −0.4 MPa osmotic potential. However, H2O2 levels were under control in both cultivars at exposure to −0.8 MPa osmotic potential. Significant proline accumulation was observed in the tissues of cv. Florispan at −0.8 MPa osmotic potential, whereas proline accumulation did not appear to be an essential part of the protection mechanism against drought in cv. Gazipasa. No significant variation in chlorophyll fluorescence values were detected in neither of the cultivars. Enzyme activity measurements revealed that Gazipasa copes well with lesser magnitudes of drought stress by increasing the activity of mainly APX, and during harsh stress conditions, only APX maintains its activity in the tissues. In cultivar Florispan, GR activity appears to take role in lesser magnitudes of drought stress, whereas CAT and APX activities appear to be very crucial antioxidative defenses during intense stress conditions. The results indicate that, the level of proline and activities of the enzymes CAT and APX are important mechanisms for the maintenance of drought tolerance in peanut plants.  相似文献   

15.
In the present investigation, we studied the possible potentiating effect of salicylic acid (SA) under Cd toxicity in Oryza sativa L. leaves. Cd treatments for 24 h reduced the shoot length, dry biomass and total chlorophyll content followed by high Cd accumulation in shoots. About 16 h presoaking with SA resulted in partial protection against Cd, as observed by minor changes in length, biomass and total chlorophyll. SA priming resulted in low Cd accumulation. Enhanced thiobarbituric acid reactive substances (TBARS), hydrogen peroxide (H2O2) and superoxide anion (O2 ) content were seen when Cd was applied alone, while under SA priming the extent of TBARS, H2O2 and O2 were significantly low, suggesting SA-regulated protection against oxidative stress. The antioxidant enzymes like Catalase (CAT), guaiacol peroxidase (GPx), glutathione reductase (GR) and superoxide dismutase (SOD) showed varied activities under Cd alone. CAT activity increased after Cd treatment, followed by a decline in GPX and GR activity. SOD also declined at the highest concentrations with an initial increase. Under SA-priming conditions, the efficiency of the antioxidant enzymes was significantly elevated. GPx and SOD activity showed significant increase in activity. The ascorbate activity increased after Cd treatment, followed by a decline in glutathione under SA-free condition. SA priming showed gradual increase in these non-enzymic antioxidants. Our results indicate that Cd-induced oxidative stress can be regulated by SA.  相似文献   

16.
The present study examined the possibility of increasing the contents of some bioactive compounds of Spirulina platensis cultivated in medium containing various hydrogen peroxide concentrations (2, 4, 6 and 8 mM) as a model for environmental stress. A positive correlation was observed between the increase of H2O2 and increasing amounts of cellular lipophilic antioxidants (total carotenoids and α-tocopherol) and hydrophilic antioxidants [glutathione (GSH) and ascorbic acid (AsA)]. HPLC profile of carotenoids revealed that algae responded to the change of H2O2 exposure by the accumulation of higher amounts of β-carotene, astaxanthine, luteine, zeaxanthin and cryptoxanthin. S. platensis showed significant linear increase in activities of antioxidant enzymes, i.e., catalase (CAT), peroxidase (PX), ascorbate peroxidase (APX) and superoxide dismutase (SOD), with increasing H2O2 concentrations. A pronounced increase of oxidative lesions’ indexes [thiobarbituric acid reactive substances (TBARS) and paramagnetic radical-EPR signal] was found in algal grown at 8 mM H2O2. These data revealed that S. platensis behaved with different strategies against H2O2 exposure which is dose dependent and their response strongly correlated with the scavenging enzymes (SOD, CAT, PX and APX) and antioxidant compounds (GSH, AsA, β-carotene, astaxanthine and α-tocopherol) in the antioxidant defense systems. Therefore, S. platensis could be considered as good candidates for successful cultivation in artificial open ponds under different environmental conditions, as high value health foods, functional foods and as source of wide spectrum of nutrients.  相似文献   

17.
18.
The mechanisms of plant tolerance to cadmium stress were studied by short-term exposure of Potamogeton crispus L. to various concentrations of Cd ranging from 0 to 0.09 mM. The accumulation of Cd and its influence on nutrient elements, chlorophyll pigments, ultrastructure, proline and MDA contents, and free radical production, as well as the activities of superoxide dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX), and glutathione reductase (GR) were investigated. The higher Cd concentration in the medium resulted in a significant enhancement of Cd accumulation. Photosynthetic pigment content decreased and ultrastructural damage to the leaf cells was aggravated with the increase in the Cd concentrations. Disruption of chloroplasts and mitochondria was observed even at the lowest concentration of Cd. Meantime, the rate of O2*− generation and the contents of H2O2 and MDA significantly increased under Cd stress, suggesting that Cd caused oxidative stress. In addition, the antioxidant system was clearly activated following Cd exposure. SOD and POD activities increased initially and then decreased, while APX and GR activities markedly increased. Simultaneously, mineral nutrition was disturbed. While K, P, Ca, and Cu contents decreased, Na, Fe, and Mn contents increased. Induction of antioxidant enzyme activities in leaves exposed to elevated Cd concentrations may be involved in Cd tolerance of P. crispus.  相似文献   

19.
Interest in, and use of, bifidobacteria as a probiotic delivered in functional foods has increased dramatically in recent years. As a result of their anaerobic nature, oxidative stress can pose a major challenge to maintaining viability of bifidobacteria during functional food storage. To better understand the oxidative stress response in two industrially important bifidobacteria species, we examined the response of three strains of B. longum and three strains of B. animalis subsp. lactis to hydrogen peroxide (H2O2). Each strain was exposed to a range of H2O2 concentrations (0–10 mM) to evaluate and compare intrinsic resistance to H2O2. Next, strains were tested for the presence of an inducible oxidative stress response by exposure to a sublethal H2O2 concentration for 20 or 60 min followed by challenge at a lethal H2O2 concentration. Results showed B. longum subsp. infantis ATCC 15697 had the highest level of intrinsic H2O2 resistance of all strains tested and B. animalis subsp. lactis BL-04 had the highest resistance among B. lactis strains. Inducible H2O2 resistance was detected in four strains, B. longum NCC2705, B. longum D2957, B. lactis RH-1, and B. lactis BL-04. Other strains showed either no difference or increased sensitivity to H2O2 after induction treatments. These data indicate that intrinsic and inducible resistance to hydrogen peroxide is strain specific in B. longum and B. lactis and suggest that for some strains, sublethal H2O2 treatments might help increase cell resistance to oxidative damage during production and storage of probiotic-containing foods.  相似文献   

20.
The effect of mutations in the genes encoding glutathione, glutaredoxin, thioredoxin, and thioredoxin reductase on the response of growing Escherichia coli to oxidative stress was studied. The gshA mutants defective in glutathione synthesis had the lowest resistance to high doses of H2O2, whereas the trxB mutants defective in thioredoxin reductase synthesis had the highest resistance to this oxidant, exceeding that of the parent strain. Among the studied mutants, the trxB cells demonstrated the highest basic levels of catalase activity and intracellular glutathione; they were able to rapidly reach the normal GSH level after oxidative stress. At the same time, these bacteria showed high frequency of induced mutations. The expression of the katG and sulA genes suggests that, having different sensitivity to high oxidant concentrations, the studied mutants differ primarily in their ability to induce the antioxidant genes of the OxyR and SOS regulons.  相似文献   

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