Glutathione, glutathione-related enzymes, and oxidative stress in individuals with subacute occupational exposure to lead.

PubMed

Dobrakowski, Michał; Pawlas, Natalia; Hudziec, Edyta; Kozłowska, Agnieszka; Mikołajczyk, Agnieszka; Birkner, Ewa; Kasperczyk, Sławomir

2016-07-01

The aim of the study was to investigate the influence of subacute exposure to lead on the glutathione-related antioxidant defense and oxidative stress parameters in 36 males occupationally exposed to lead for 40±3.2days. Blood lead level in the examined population increased significantly by 359% due to lead exposure. Simultaneously, erythrocyte glutathione level decreased by 16%, whereas the activity of glutathione-6-phosphate dehydrogenase in erythrocytes and leukocytes decreased by 28% and 10%, respectively. Similarly, the activity of glutathione-S-transferase in erythrocytes decreased by 45%. However, the activity of glutathione reductase in erythrocytes and leukocytes increased by 26% and 6%, respectively, whereas the total oxidant status value in leukocytes increased by 37%. Subacute exposure to lead results in glutathione pool depletion and accumulation of lipid peroxidation products; however, it does not cause DNA damage. Besides, subacute exposure to lead modifies the activity of glutathione-related enzymes. Copyright © 2016 Elsevier B.V. All rights reserved.

Glutathione conjugation and contaminant transformation

USGS Publications Warehouse

Field, Jennifer A.; Thurman, E.M.

1996-01-01

The recent identification of a novel sulfonated metabolite of alachlor in groundwater and metolachlor in soil is likely the result of glutathione conjugation. Glutathione conjugation is an important biochemical reaction that leads, in the case of alachlor, to the formation of a rather difficult to detect, water-soluble, and therefore highly mobile, sulfonated metabolite. Research from weed science, toxicology, and biochemistry is discussed to support the hypothesis that glutathione conjugation is a potentially important detoxification pathway carried out by aquatic and terrestrial plants and soil microorganisms. A brief review of the biochemical basis for glutathione conjugation is presented. We recommend that multidisciplinary research focus on the occurrence and expression of glutathione and its attendant enzymes in plants and microorganisms, relationships between electrophilic substrate structure and enzyme activity, and the potential exploitation of plants and microorganisms that are competent in glutathione conjugation for phytoremediation and bioremediation.

Correction of glutathione deficiency in the lower respiratory tract of HIV seropositive individuals by glutathione aerosol treatment.

PubMed

Holroyd, K J; Buhl, R; Borok, Z; Roum, J H; Bokser, A D; Grimes, G J; Czerski, D; Cantin, A M; Crystal, R G

1993-10-01

Concentrations of glutathione, a ubiquitous tripeptide with immune enhancing and antioxidant properties, are decreased in the blood and lung epithelial lining fluid of human immunodeficiency virus (HIV) seropositive individuals. Since the lung is the most common site of infection in those who progress to AIDS it is rational to consider whether it is possible to safely augment glutathione levels in the epithelial lining fluid of HIV seropositive individuals, thus potentially improving local host defence. Purified reduced glutathione was delivered by aerosol to HIV seropositive individuals (n = 14) and the glutathione levels in lung epithelial lining fluid were compared before and at one, two, and three hours after aerosol administration. Before treatment total glutathione concentrations in the epithelial lining fluid were approximately 60% of controls. After three days of twice daily doses each of 600 mg reduced glutathione, total glutathione levels in the epithelial lining fluid increased and remained in the normal range for at least three hours after treatment. Strikingly, even though > 95% of the glutathione in the aerosol was in its reduced form, the percentage of oxidised glutathione in epithelial lining fluid increased from 5% before treatment to about 40% three hours after treatment, probably reflecting the use of glutathione as an antioxidant in vivo. No adverse effects were observed. It is feasible and safe to use aerosolised reduced glutathione to augment the deficient glutathione levels of the lower respiratory tract of HIV seropositive individuals. It is rational to evaluate further the efficacy of this tripeptide in improving host defence in HIV seropositive individuals.

Correction of glutathione deficiency in the lower respiratory tract of HIV seropositive individuals by glutathione aerosol treatment.

PubMed Central

Holroyd, K. J.; Buhl, R.; Borok, Z.; Roum, J. H.; Bokser, A. D.; Grimes, G. J.; Czerski, D.; Cantin, A. M.; Crystal, R. G.

1993-01-01

BACKGROUND--Concentrations of glutathione, a ubiquitous tripeptide with immune enhancing and antioxidant properties, are decreased in the blood and lung epithelial lining fluid of human immunodeficiency virus (HIV) seropositive individuals. Since the lung is the most common site of infection in those who progress to AIDS it is rational to consider whether it is possible to safely augment glutathione levels in the epithelial lining fluid of HIV seropositive individuals, thus potentially improving local host defence. METHODS--Purified reduced glutathione was delivered by aerosol to HIV seropositive individuals (n = 14) and the glutathione levels in lung epithelial lining fluid were compared before and at one, two, and three hours after aerosol administration. RESULTS--Before treatment total glutathione concentrations in the epithelial lining fluid were approximately 60% of controls. After three days of twice daily doses each of 600 mg reduced glutathione, total glutathione levels in the epithelial lining fluid increased and remained in the normal range for at least three hours after treatment. Strikingly, even though > 95% of the glutathione in the aerosol was in its reduced form, the percentage of oxidised glutathione in epithelial lining fluid increased from 5% before treatment to about 40% three hours after treatment, probably reflecting the use of glutathione as an antioxidant in vivo. No adverse effects were observed. CONCLUSIONS--It is feasible and safe to use aerosolised reduced glutathione to augment the deficient glutathione levels of the lower respiratory tract of HIV seropositive individuals. It is rational to evaluate further the efficacy of this tripeptide in improving host defence in HIV seropositive individuals. PMID:8256245

Identification of Protein-Protein Interactions with Glutathione-S-Transferase (GST) Fusion Proteins.

PubMed

Einarson, Margret B; Pugacheva, Elena N; Orlinick, Jason R

2007-08-01

INTRODUCTIONGlutathione-S-transferase (GST) fusion proteins have had a wide range of applications since their introduction as tools for synthesis of recombinant proteins in bacteria. GST was originally selected as a fusion moiety because of several desirable properties. First and foremost, when expressed in bacteria alone, or as a fusion, GST is not sequestered in inclusion bodies (in contrast to previous fusion protein systems). Second, GST can be affinity-purified without denaturation because it binds to immobilized glutathione, which provides the basis for simple purification. Consequently, GST fusion proteins are routinely used for antibody generation and purification, protein-protein interaction studies, and biochemical analysis. This article describes the use of GST fusion proteins as probes for the identification of protein-protein interactions.

Glutathione and multidrug resistance protein transporter mediate a self-propelled disposal of bismuth in human cells

PubMed Central

Hong, Yifan; Lai, Yau-Tsz; Chan, Godfrey Chi-Fung; Sun, Hongzhe

2015-01-01

Glutathione and multidrug resistance protein (MRP) play an important role on the metabolism of a variety of drugs. Bismuth drugs have been used to treat gastrointestinal disorder and Helicobacter pylori infection for decades without exerting acute toxicity. They were found to interact with a wide variety of biomolecules, but the major metabolic pathway remains unknown. For the first time (to our knowledge), we systematically and quantitatively studied the metabolism of bismuth in human cells. Our data demonstrated that over 90% of bismuth was passively absorbed, conjugated to glutathione, and transported into vesicles by MRP transporter. Mathematical modeling of the system reveals an interesting phenomenon. Passively absorbed bismuth consumes intracellular glutathione, which therefore activates de novo biosynthesis of glutathione. Reciprocally, sequestration by glutathione facilitates the passive uptake of bismuth and thus completes a self-sustaining positive feedback circle. This mechanism robustly removes bismuth from both intra- and extracellular space, protecting critical systems of human body from acute toxicity. It elucidates the selectivity of bismuth drugs between human and pathogens that lack of glutathione, such as Helicobacter pylori, opening new horizons for further drug development. PMID:25737551

Anethole dithiolethione, a putative neuroprotectant, increases intracellular and extracellular glutathione levels during starvation of cultured astroglial cells.

PubMed

Dringen, R; Hamprecht, B; Drukarch, B

1998-12-01

Astroglial cells protect neurons against oxidative damage. The antioxidant glutathione plays a pivotal role in the neuroprotective action of astroglial cells which is impaired following loss of glutathione. Anethole dithiolethione (ADT), a sulfur-containing compound which is used in humans as a secretagogue, increases glutathione levels in cultured astroglial cells under "physiological" conditions and is thought thereby to protect against oxidative damage. Presently, we report the effect of ADT (3-100 microM) on glutathione content of and efflux from rat primary astroglia-rich cultures under "pathological" conditions, i.e., extended deprivation of glucose and amino acids. Although cellular viability was not affected significantly, starvation of these cultures for 24 h in a bicarbonate buffer lacking glucose and amino acids led to a decrease in glutathione and protein content of approximately 43% and 40%, respectively. Although no effect on the protein loss occurred, the presence of ADT during starvation counteracted the starvation-induced loss of intracellular glutathione in a concentration-dependent way. At a concentration of 100 microM ADT even a significant increase in astroglial glutathione content was noted after 24 h of starvation. Alike intracellular glutathione levels, the amount of glutathione found in the buffer was elevated substantially if ADT was present during starvation. This ADT-mediated, apparent increase in glutathione efflux was additive to the stimulatory effect on extracellular glutathione levels of acivicin (100 microM), an inhibitor of extracellular enzymatic glutathione breakdown. However, the ADT-induced elevation of both intra- and extracellular glutathione content during starvation was prevented completely by coincubation with buthionine sulfoximine (10 microM), an inhibitor of glutathione synthesis. These results demonstrate that, most likely through stimulation of glutathione synthesis, ADT enables astroglial cells to maintain higher

Methyl-esterified 3-hydroxybutyrate oligomers protect bacteria from hydroxyl radicals

USDA-ARS?s Scientific Manuscript database

Bacteria rely mainly on enzymes, glutathione and other low-molecular weight thiols to overcome oxidative stress. However, hydroxyl radicals are the most cytotoxic reactive oxygen species, and no known enzymatic system exists for their detoxification. We now show that methyl-esterified dimers and tri...

The evolution of glutathione metabolism in phototrophic microorganisms

NASA Technical Reports Server (NTRS)

Fahey, Robert C.; Buschbacher, Ralph M.; Newton, Gerald L.

1988-01-01

The low molecular weight thiol composition of a variety of phototropic microorganisms is examined in order to ascertain how evolution of glutathione (GSH) production is related to the evolution of oxygenic photosynthesis. Cells were extracted in the presence of monobromobimane (mBBr) to convert thiols (RSH) to fluorescent derivatives (RSmB) which were analyzed by high performance liquid chromatography (HPLC). Significant levels of GSH were not found in green sulfur bacteria. Substantial levels were present in purple bacteria, cyanobacteria, and eukaryotic algae. Other thiols measured included cysteine, gamma-glutamylcysteine, thiosulfate, coenzyme A, and sulfide. Many of the organisms also exhibited a marked ability to reduce mBBr to syn-(methyl,methyl)bimane, an ability which was quenched by treatment with 2-pyridyl disulfide or 5,5 prime-bisdithio - (2-nitrobenzoic acid) prior to reaction with mBBr. These observations indicate the presence of a reducing system capable of electron transfer to mBBr and reduction of reactive disulfides. The distribution of GSH in phototropic eubacteria indicates that GSH synthesis evolved at or around the time that oxygenic photosynthesis evolved.

Determining Glutathione Levels in Plants.

PubMed

Sahoo, Smita; Awasthi, Jay Prakash; Sunkar, Ramanjulu; Panda, Sanjib Kumar

2017-01-01

Upon exposure to abiotic stresses, plants tend to accumulate excessive amounts of reactive oxygen species (ROS) that inturn react with cellular lipids, proteins, and DNA. Therefore, decreasing ROS accumulation is indispensible to survive under stress, which is accomplished by inducing enzymatic and nonenzymatic antioxidant defense pathways. Glutathione, particularly reduced glutathione (GSH), represents a principal anitioxidant that could decrease ROS through scavenging them directly or indirectly through ascorbate-glutathione cycle or GSH peroxidases. Glutathione content can be determined using HPLC or spectrophotometric assays. In this chapter, we provided detailed assays to determine total, reduced, and oxidized gluathione using spectrophotometric method.

Dysregulation of Glutathione Homeostasis in Neurodegenerative Diseases

PubMed Central

Johnson, William M.; Wilson-Delfosse, Amy L.; Mieyal, John. J.

2012-01-01

Dysregulation of glutathione homeostasis and alterations in glutathione-dependent enzyme activities are increasingly implicated in the induction and progression of neurodegenerative diseases, including Alzheimer’s, Parkinson’s and Huntington’s diseases, amyotrophic lateral sclerosis, and Friedreich’s ataxia. In this review background is provided on the steady-state synthesis, regulation, and transport of glutathione, with primary focus on the brain. A brief overview is presented on the distinct but vital roles of glutathione in cellular maintenance and survival, and on the functions of key glutathione-dependent enzymes. Major contributors to initiation and progression of neurodegenerative diseases are considered, including oxidative stress, protein misfolding, and protein aggregation. In each case examples of key regulatory mechanisms are identified that are sensitive to changes in glutathione redox status and/or in the activities of glutathione-dependent enzymes. Mechanisms of dysregulation of glutathione and/or glutathione-dependent enzymes are discussed that are implicated in pathogenesis of each neurodegenerative disease. Limitations in information or interpretation are identified, and possible avenues for further research are described with an aim to elucidating novel targets for therapeutic interventions. The pros and cons of administration of N-acetylcysteine or glutathione as therapeutic agents for neurodegenerative diseases, as well as the potential utility of serum glutathione as a biomarker, are critically evaluated. PMID:23201762

Glutathione oxidation in response to intracellular H2O2: Key but overlapping roles for dehydroascorbate reductases.

PubMed

Rahantaniaina, Marie-Sylviane; Li, Shengchun; Chatel-Innocenti, Gilles; Tuzet, Andrée; Mhamdi, Amna; Vanacker, Hélène; Noctor, Graham

2017-08-03

Glutathione is a pivotal molecule in oxidative stress, during which it is potentially oxidized by several pathways linked to H 2 O 2 detoxification. We have investigated the response and functional importance of 3 potential routes for glutathione oxidation pathways mediated by glutathione S-transferases (GST), glutaredoxin-dependent peroxiredoxins (PRXII), and dehydroascorbate reductases (DHAR) in Arabidopsis during oxidative stress. Loss-of-function gstU8, gstU24, gstF8, prxIIE and prxIIF mutants as well as double gstU8 gstU24, gstU8 gstF8, gstU24 gstF8, prxIIE prxIIF mutants were obtained. No mutant lines showed marked changes in their phenotype and glutathione profiles in comparison to the wild-type plants in either optimal conditions or oxidative stress triggered by catalase inhibition. By contrast, multiple loss of DHAR functions markedly decreased glutathione oxidation triggered by catalase deficiency. To assess whether this effect was mediated directly by loss of DHAR enzyme activity, or more indirectly by upregulation of other enzymes involved in glutathione and ascorbate recycling, we measured expression of glutathione reductase (GR) and expression and activity of monodehydroascorbate reductases (MDHAR). No evidence was obtained that either GRs or MDHARs were upregulated in plants lacking DHAR function. Hence, interplay between different DHARs appears to be necessary to couple ascorbate and glutathione pools and to allow glutathione-related signaling during enhanced H 2 O 2 metabolism.

Passive regeneration of glutathione: glutathione reductase regulation in the freeze-tolerant North American wood frog, Rana sylvatica.

PubMed

Dawson, Neal J; Storey, Kenneth B

2017-09-01

Wood frogs inhabit a broad range across North America, extending from the southern tip of the Appalachian Mountains to the northern boreal forest. Remarkably, they can survive the winter in a frozen state, where as much as 70% of their body water is converted into ice. Whilst in the frozen state, their hearts cease to pump blood, causing their cells to experience ischemia, which can dramatically increase the production of reactive oxygen species within the cell. To overcome this, wood frogs have elevated levels of glutathione, a primary antioxidant. We examined the regulation of glutathione reductase, the enzyme involved in recycling glutathione, in both the frozen and unfrozen (control) state. Glutathione reductase activity from both the control and frozen state showed a dramatic reduction in substrate specificity ( K m ) for oxidized glutathione (50%) when measured in the presence of glucose (300 mmol l -1 ) and a increase (157%) when measured in the presence of levels of urea (75 mmol l -1 ) encountered in the frozen state. However, when we tested the synergistic effect of urea and glucose simultaneously, we observed a substantial reduction in the K m for oxidized glutathione (43%) to a value similar to that with glucose alone. In fact, we found no observable differences in the kinetic and structural properties of glutathione reductase between the two states. Therefore, a significant increase in the affinity for oxidized glutathione in the presence of endogenous levels of glucose suggests that increased glutathione recycling may occur as a result of passive regulation of glutathione reductase by rising levels of glucose during freezing. © 2017. Published by The Company of Biologists Ltd.

Antibacterial activity of exogenous glutathione and its synergism on antibiotics sensitize carbapenem-associated multidrug resistant clinical isolates of Acinetobacter baumannii.

PubMed

Alharbe, Roaa; Almansour, Ayidh; Kwon, Dong H

2017-10-01

A major clinical impact of A. baumannii is hospital-acquired infections including ventilator-associated pneumonia. The treatment of this pathogen is often difficult due to its innate and acquired resistance to almost all commercially available antibiotics. Infections with carbapenem-associated multidrug resistant A. baumannii is the most problematic. Glutathione is a tripeptide thiol-antioxidant and antibacterial activity of exogenous glutathione was reported in some bacteria. However, clinical relevance and molecular details of the antibacterial activity of glutathione are currently unclear. Seventy clinical isolates of A. baumannii including 63 carbapenem-associated multidrug resistant isolates and a type strain A. baumannii ATCC 19606 were used to determine minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). Fractional inhibitory concentration (FIC) and time-killing activity with meropenem and/or glutathione were also determined in the carbapenem-associated multidrug resistant isolates. In addition, the roles of exogenous glutathione in multidrug efflux pumps and β-lactamase production were examined. Levels of MIC and MBC were ranged from 10 to 15mM of exogenous glutathione. All tested carbapenem-associated multidrug resistant isolates were sensitized by all tested antibiotics in combination with subinhibitory concentrations of glutathione. FIC levels of glutathione with carbapenem (meropenem) were all<0.5 and the carbapenem-associated multidrug resistant isolates were killed by subinhibitory concentrations of both glutathione and meropenem at>2log10 within 12h, suggesting glutathione synergistically interacts with meropenem. The roles of multidrug efflux pumps and β-lactamase production were excluded for the glutathione-mediated antibiotic susceptibility. Overall results demonstrate that the antibacterial activity of glutathione is clinically relevant and its synergism on antibiotics sensitizes clinical isolates of A

Low levels of glutathione are sufficient for survival of keratinocytes after UV irradiation and for healing of mouse skin wounds.

PubMed

Telorack, Michèle; Abplanalp, Jeannette; Werner, Sabine

2016-08-01

Reduced levels of the cellular antioxidant glutathione are associated with premature skin aging, cancer and impaired wound healing, but the in vivo functions of glutathione in the skin remain largely unknown. Therefore, we analyzed mice lacking the modifier subunit of the glutamate cysteine ligase (Gclm), the enzyme that catalyzes the rate-limiting step of glutathione biosynthesis. Glutathione levels in the skin of these mice were reduced by 70 %. However, neither skin development and homeostasis, nor UVA- or UVB-induced apoptosis in the epidermis were affected. Histomorphometric analysis of excisional wounds did not reveal wound healing abnormalities in young Gclm-deficient mice, while the area of hyperproliferative epithelium as well as keratinocyte proliferation were affected in aged mice. These findings suggest that low levels of glutathione are sufficient for wound repair in young mice, but become rate-limiting upon aging.

Distribution and abundance of organic thiols

NASA Technical Reports Server (NTRS)

Fahey, R.

1985-01-01

The role of glutathione (GSH) in protecting against the toxicity of oxygen and oxygen by products is well established for all eukaryotes studied except Entamoeba histolytica which lacks mitochrondria, chloroplasts, and microtubules. The GSH is not universal among prokaryotes. Entamoeba histolytica does not produce GSH or key enzymes of GSH metabolism. A general method of thiol analysis based upon fluorescent labeling with monobromobimane and HPLC separation of the resulting thiol derivatives was developed to determine the occurrence of GSH and other low molecular weight thiols in bacteria. Glutathione is the major thiol in cyanobacteria and in most bacteria closely related to the purple photosynthetic bacteria, but GSH was not found in archaebacteria, green bacteria, or GRAM positive bacteria. It suggested that glutathione metabolism was incorporated into eukaryotes at the time that mitochondria and chloroplasts were acquired by endosymbiosis. In Gram positive aerobes, coenzyme A occurs at millimolar levels and CoA disulfide reductases are identified. The CoA, rather than glutathione, may function in the oxygen detoxification processes of these organisms.

[Alternative nutrition and glutathione levels].

PubMed

Krajcovicová-Kudlácková, M; Simoncic, R; Béderová, A; Brtková, A; Magálová, T; Barteková, S

1999-08-30

Low protein quality and quantity is reported to be a possible risk of alternative nutrition. Pulses contain 18-41% of methionine in relation to reference protein, moreover, its content in cereals is by one half lower. Therefore vegetarians and vegans may have an insufficient intake of sulphur-containing amino acids that may subsequently affect glutathione values (precursors of its synthesis). In groups of adults on an alternative diet--lactoovovegetarians (n = 47) and vegans (n = 44) aged 19-62 years with average duration on a vegetarian or vegan diet of 7.6 and 4.9 years, respectively, glutathione levels (GSH) were measured in erythrocytes (spectrophotometrically), as well as the activity of GSH-dependent enzymes. As nutritional control (n = 42) served an average sample of omnivores selected from a group of 489 examined, apparently healthy subjects of the same age range living in the same region. One to low protein intake (56% of RDA) exclusively of plant origin significantly lower levels of total proteins were observed in vegans with a 16% frequency of hypoproteinaemia (vs 0% in omnivores). In comparison to omnivores a significantly lower glutathione level was found (4.28 +/- 0.12 vs 4.84 +/- 0.14 mumol/g Hb, P < 0.01). Lactoovovegetarians because of their protein intake in adequate amounts with a 27% proportion of animal proteins (dairy products, eggs) consume a balanced mixture of amino acids, which is reflected in total protein levels similar to omnivores and significantly higher values of glutathione -5.26 +/- 0.12 mumol/g Hb, P < 0.05 (intake of glutathione in diet, higher consumption of fruit and vegetable in comparison to omnivores). A sufficient supply of glutathione as the substrate for enzymatic reactions of hydrogen peroxide or lipid hydroperoxide catabolism, as well as for detoxication of xenobiotics, was reflected in lactoovovegetarians in a significantly higher activity of glutathione-peroxidase and glutathione-S-transferase in erythrocytes. Low

Changes of reduced glutathion, glutathion reductase, and glutathione peroxidase after radiation in guinea pigs

DOE Office of Scientific and Technical Information (OSTI.GOV)

Erden, M.; Bor, N.M.

1984-04-01

In this series of experiments the protective action of reduced glutathion due to ionizing radiation has been studied. In the experimental group 18 guinea pigs were exposed to successive radiations of 150 rad 3 or 4 days apart. Total dose given amounted to 750 rad which is the LD50 for guinea pigs. Blood samples were taken 30 min after each exposure. The control series were sham radiated but otherwise treated identically. The cells of the removed blood samples were separated by centrifugation and were subjected to the reduced glutathion stability test. GSSGR, GPer, and LDH enzyme activities were also measuredmore » of which the latter served as a marked enzyme. It was found that LDH did not show any alteration after radiation. The reduced glutathion stability test showed a consistent but minor reduction (P greater than 0.05), in the experimental group. GSSGR enzyme activity on the other hand was reduced significantly (from 176.48 +/- 11.32 to 41.34 +/- 1.17 IU/ml of packed erythrocytes, P less than 0.001) in the same group. GPer activity showed a consistent but minor elevation during the early phase of the experimental group. It was later increased significantly beginning after 600 rad total radiation on the fourth session (P less than 0.050).« less

Optimization of glutathione production in batch and fed-batch cultures by the wild-type and recombinant strains of the methylotrophic yeast Hansenula polymorpha DL-1

PubMed Central

2011-01-01

Background Tripeptide glutathione (gamma-glutamyl-L-cysteinyl-glycine) is the most abundant non-protein thiol that protects cells from metabolic and oxidative stresses and is widely used as medicine, food additives and in cosmetic industry. The methylotrophic yeast Hansenula polymorpha is regarded as a rich source of glutathione due to the role of this thiol in detoxifications of key intermediates of methanol metabolism. Cellular and extracellular glutathione production of H. polymorpha DL-1 in the wild type and recombinant strains which overexpress genes of glutathione biosynthesis (GSH2) and its precursor cysteine (MET4) was studied. Results Glutathione producing capacity of H. polymorpha DL-1 depending on parameters of cultivation (dissolved oxygen tension, pH, stirrer speed), carbon substrate (glucose, methanol) and type of overexpressed genes of glutathione and its precursor biosynthesis during batch and fed-batch fermentations were studied. Under optimized conditions of glucose fed-batch cultivation, the glutathione productivity of the engineered strains was increased from ~900 up to ~ 2300 mg of Total Intracellular Glutathione (TIG) or GSH+GSSGin, per liter of culture medium. Meantime, methanol fed-batch cultivation of one of the recombinant strains allowed achieving the extracellular glutathione productivity up to 250 mg of Total Extracellular Glutathione (TEG) or GSH+GSSGex, per liter of the culture medium. Conclusions H. polymorpha is an competitive glutathione producer as compared to other known yeast and bacteria strains (Saccharomyces cerevisiae, Candida utilis, Escherichia coli, Lactococcus lactis etc.) with good perspectives for further improvement especially for production of extracellular form of glutathione. PMID:21255454

Subcellular Distribution of Glutathione Precursors in Arabidopsis thaliana

PubMed Central

Koffler, Barbara Eva; Maier, Romana; Zechmann, Bernd

2011-01-01

Abstract Glutathione is an important antioxidant and has many important functions in plant development, growth and defense. Glutathione synthesis and degradation is highly compartment-specific and relies on the subcellular availability of its precursors, cysteine, glutamate, glycine and γ-glutamylcysteine especially in plastids and the cytosol which are considered as the main centers for glutathione synthesis. The availability of glutathione precursors within these cell compartments is therefore of great importance for successful plant development and defense. The aim of this study was to investigate the compartment-specific importance of glutathione precursors in Arabidopsis thaliana. The subcellular distribution was compared between wild type plants (Col-0), plants with impaired glutathione synthesis (glutathione deficient pad2-1 mutant, wild type plants treated with buthionine sulfoximine), and one complemented line (OE3) with restored glutathione synthesis. Immunocytohistochemistry revealed that the inhibition of glutathione synthesis induced the accumulation of the glutathione precursors cysteine, glutamate and glycine in most cell compartments including plastids and the cytosol. A strong decrease could be observed in γ-glutamylcysteine (γ-EC) contents in these cell compartments. These experiments demonstrated that the inhibition of γ-glutamylcysteine synthetase (GSH1) – the first enzyme of glutathione synthesis – causes a reduction of γ-EC levels and an accumulation of all other glutathione precursors within the cells. PMID:22050910

Glutathione-related enzymes and the eye.

PubMed

Ganea, Elena; Harding, John J

2006-01-01

Glutathione and the related enzymes belong to the defence system protecting the eye against chemical and oxidative stress. This review focuses on GSH and two key enzymes, glutathione reductase and glucose-6-phosphate dehydrogenase in lens, cornea, and retina. Lens contains a high concentration of reduced glutathione, which maintains the thiol groups in the reduced form. These contribute to lens complete transparency as well as to the transparent and refractive properties of the mammalian cornea, which are essential for proper image formation on the retina. In cornea, gluthatione also plays an important role in maintaining normal hydration level, and in protecting cellular membrane integrity. In retina, glutathione is distributed in the different types of retinal cells. Intracellular enzyme, glutathione reductase, involved in reducing the oxidized glutathione has been found at highest activity in human and primate lenses, as compared to other species. Besides the enzymes directly involved in maintaining the normal redox status of the cell, glucose-6-phosphate dehydrogenase which catalyzes the first reaction of the pentose phosphate pathway, plays a key role in protection of the eye against reactive oxygen species. Cornea has a high activity of the pentose phosphate pathway and glucose-6-phosphate dehydrogenase activity. Glycation, the non-enzymic reaction between a free amino group in proteins and a reducing sugar, slowly inactivates gluthathione-related and other enzymes. In addition, glutathione can be also glycated. The presence of glutathione, and of the related enzymes has been also reported in other parts of the eye, such as ciliary body and trabecular meshwork, suggesting that the same enzyme systems are present in all tissues of the eye to generate NADPH and to maintain gluthatione in the reduced form. Changes of glutathione and related enzymes activity in lens, cornea, retina and other eye tissues, occur with ageing, cataract, diabetes, irradiation and

Glutathione synthetase deficiency: a family report.

PubMed Central

Pejaver, R K; Watson, A H

1994-01-01

Glutathione synthetase deficiency is a rare inborn error of metabolism. Low levels of and at times unstable molecules of glutathione synthetase leads to glutathione deficiency affecting various systems of the body. The inheritance is thought to be of autosomal recessive variety. We diagnosed the condition in a neonate and proceeded to investigate the family. The results are discussed below. PMID:8158601

Biochemical heterogeneity in glutathione synthetase deficiency.

PubMed Central

Spielberg, S P; Garrick, M D; Corash, L M; Butler, J D; Tietze, F; Rogers, L; Schulman, J D

1978-01-01

Two different clinical syndromes are associated with glutathione synthetase deficiency, one presenting with hemolytic anemia and 5-oxoprolinuria, the other with isolated hemolysis. We have differentiated these disorders on an enzymatic basis. In 5-oxoprolinuria, all cell types examined have grossly deficient enzyme activity and glutathione content. In contrast, in the nonoxoprolinuric variant, erythrocytes have decreased enzyme activity and glutathione content, whereas nucleated cells maintain substantial levels of both. The enzyme in this disorder is unstable in vitro and has shortened survival in intact erythrocytes. Nucleated cells appear able to maintain sufficient enzyme activity and concentrations of glutathione to suppress overproduction of 5-oxoproline. PMID:659603

Methylmercury alters glutathione homeostasis by inhibiting glutaredoxin 1 and enhancing glutathione biosynthesis in cultured human astrocytoma cells.

PubMed

Robitaille, Stephan; Mailloux, Ryan J; Chan, Hing Man

2016-08-10

Methylmercury (MeHg) is a neurotoxin that binds strongly to thiol residues on protein and low molecular weight molecules like reduced glutathione (GSH). The mechanism of its effects on GSH homeostasis particularly at environmentally relevant low doses is not fully known. We hypothesized that exposure to MeHg would lead to a depletion of reduced glutathione (GSH) and an accumulation of glutathione disulfide (GSSG) leading to alterations in S-glutathionylation of proteins. Our results showed exposure to low concentrations of MeHg (1μM) did not significantly alter GSH levels but increased GSSG levels by ∼12-fold. This effect was associated with a significant increase in total cellular glutathione content and a decrease in GSH/GSSG. Immunoblot analyses revealed that proteins involved in glutathione synthesis were upregulated accounting for the increase in cellular glutathione. This was associated an increase in cellular Nrf2 protein levels which is required to induce the expression of antioxidant genes in response to cellular stress. Intriguingly, we noted that a key enzyme involved in reversing protein S-glutathionylation and maintaining glutathione homeostasis, glutaredoxin-1 (Grx1), was inhibited by ∼50%. MeHg treatment also increased the S-glutathionylation of a high molecular weight protein. This observation is consistent with the inhibition of Grx1 and elevated H2O2 production however; contrary to our original hypothesis we found few S-glutathionylated proteins in the astrocytoma cells. Collectively, MeHg affects multiple arms of glutathione homeostasis ranging from pool management to protein S-glutathionylation and Grx1 activity. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Glutathione synthesis is essential for pollen germination in vitro

PubMed Central

2011-01-01

Background The antioxidant glutathione fulfills many important roles during plant development, growth and defense in the sporophyte, however the role of this important molecule in the gametophyte generation is largely unclear. Bioinformatic data indicate that critical control enzymes are negligibly transcribed in pollen and sperm cells. Therefore, we decided to investigate the role of glutathione synthesis for pollen germination in vitro in Arabidopsis thaliana accession Col-0 and in the glutathione deficient mutant pad2-1 and link it with glutathione status on the subcellular level. Results The depletion of glutathione by buthionine sulfoximine (BSO), an inhibitor of glutathione synthesis, reduced pollen germination rates to 2-5% compared to 71% germination in wildtype controls. The application of reduced glutathione (GSH), together with BSO, restored pollen germination and glutathione contents to control values, demonstrating that inhibition of glutathione synthesis is responsible for the decrease of pollen germination in vitro. The addition of indole-3-acetic acid (IAA) to media containing BSO restored pollen germination to control values, which demonstrated that glutathione depletion in pollen grains triggered disturbances in auxin metabolism which led to inhibition of pollen germination. Conclusions This study demonstrates that glutathione synthesis is essential for pollen germination in vitro and that glutathione depletion and auxin metabolism are linked in pollen germination and early elongation of the pollen tube, as IAA addition rescues glutathione deficient pollen. PMID:21439079

21 CFR 862.1365 - Glutathione test system.

Code of Federal Regulations, 2011 CFR

2011-04-01

....1365 Glutathione test system. (a) Identification. A glutathione test system is a device intended to measure glutathione (the tripeptide of glycine, cysteine, and glutamic acid) in erythrocytes (red blood... I (general controls). The device is exempt from the premarket notification procedures in subpart E...

21 CFR 862.1365 - Glutathione test system.

Code of Federal Regulations, 2010 CFR

2010-04-01

....1365 Glutathione test system. (a) Identification. A glutathione test system is a device intended to measure glutathione (the tripeptide of glycine, cysteine, and glutamic acid) in erythrocytes (red blood... I (general controls). The device is exempt from the premarket notification procedures in subpart E...

21 CFR 862.1365 - Glutathione test system.

Code of Federal Regulations, 2013 CFR

2013-04-01

....1365 Glutathione test system. (a) Identification. A glutathione test system is a device intended to measure glutathione (the tripeptide of glycine, cysteine, and glutamic acid) in erythrocytes (red blood... I (general controls). The device is exempt from the premarket notification procedures in subpart E...

Farnesol-Induced Apoptosis in Candida albicans Is Mediated by Cdr1-p Extrusion and Depletion of Intracellular Glutathione

PubMed Central

Zhu, Jingsong; Krom, Bastiaan P.; Sanglard, Dominique; Intapa, Chaidan; Dawson, Clinton C.; Peters, Brian M.; Shirtliff, Mark E.; Jabra-Rizk, Mary Ann

2011-01-01

Farnesol is a key derivative in the sterol biosynthesis pathway in eukaryotic cells previously identified as a quorum sensing molecule in the human fungal pathogen Candida albicans. Recently, we demonstrated that above threshold concentrations, farnesol is capable of triggering apoptosis in C. albicans. However, the exact mechanism of farnesol cytotoxicity is not fully elucidated. Lipophilic compounds such as farnesol are known to conjugate with glutathione, an antioxidant crucial for cellular detoxification against damaging compounds. Glutathione conjugates act as substrates for ATP-dependent ABC transporters and are extruded from the cell. To that end, this current study was undertaken to validate the hypothesis that farnesol conjugation with intracellular glutathione coupled with Cdr1p-mediated extrusion of glutathione conjugates, results in total glutathione depletion, oxidative stress and ultimately fungal cell death. The combined findings demonstrated a significant decrease in intracellular glutathione levels concomitant with up-regulation of CDR1 and decreased cell viability. However, addition of exogenous reduced glutathione maintained intracellular glutathione levels and enhanced viability. In contrast, farnesol toxicity was decreased in a mutant lacking CDR1, whereas it was increased in a CDR1-overexpressing strain. Further, gene expression studies demonstrated significant up-regulation of the SOD genes, primary enzymes responsible for defense against oxidative stress, with no changes in expression in CDR1. This is the first study describing the involvement of Cdr1p-mediated glutathione efflux as a mechanism preceding the farnesol-induced apoptotic process in C. albicans. Understanding of the mechanisms underlying farnesol-cytotoxicity in C. albicans may lead to the development of this redox-cycling agent as an alternative antifungal agent. PMID:22205973

Determination of glutathione and glutathione disulfide in biological samples: an in-depth review.

PubMed

Monostori, Péter; Wittmann, Gyula; Karg, Eszter; Túri, Sándor

2009-10-15

Glutathione (GSH) is a thiol-containing tripeptide, which plays central roles in the defence against oxidative damage and in signaling pathways. Upon oxidation, GSH is transformed to glutathione disulfide (GSSG). The concentrations of GSH and GSSG and their molar ratio are indicators of cell functionality and oxidative stress. Assessment of redox homeostasis in various clinical states and medical applications for restoration of the glutathione status are of growing importance. This review is intended to provide a state-of-the-art overview of issues relating to sample pretreatment and choices for the separation and detection of GSH and GSSG. High-performance liquid chromatography, capillary electrophoresis and gas chromatography (as techniques with a separation step) with photometric, fluorimetric, electrochemical and mass spectrometric detection are discussed, stress being laid on novel approaches.

p-aminophenol nephrotoxicity: biosynthesis of toxic glutathione conjugates.

PubMed

Klos, C; Koob, M; Kramer, C; Dekant, W

1992-07-01

p-Aminophenol causes necrosis of the pars recta of the proximal tubules in rats, and its nephrotoxicity may be due to glutathione-dependent bioactivation reactions. We have investigated the hepatic metabolism of p-aminophenol in Wistar rats and the cytotoxicity of formed glutathione S-conjugates in rat renal epithelial cells. After ip application of p-aminophenol (100 mg/kg), the following metabolites were identified in rat bile: 4-amino-2-(glutathion-S-yl)phenol, 4-amino-3-(glutathion-S-yl)-phenol, 4-amino-2,5-bis(glutathion-S-yl)phenol, 4-amino-2,3,5(or 6)-tris(glutathion-S-yl)phenol, an aminophenol conjugate (likely a sulfate or glucuronide), acetaminophen glucuronide, and 3-(glutathion-S-yl)acetaminophen. 4-Amino-3-(glutathion-S-yl)phenol, 4-amino-2,5-bis(glutathion-S-yl)phenol, and 4-amino-2,3,5(or 6)-tris(glutathion-S-yl)phenol induced a dose- and time-dependent loss of cell viability in rat kidney cortical cells. Cell killing was significantly reduced by inhibition of gamma-glutamyl transpeptidase with Acivicin. p-Aminophenol was also toxic to renal epithelial cells. Coincubation of p-aminophenol with tetraethylammonium bromide, a competitive inhibitor of the organic cation transporter, and with SKF-525A, an inhibitor of cytochrome P450, protected cells from p-aminophenol-induced toxicity. p-Aminophenol would thus be accumulated in the kidney mainly by organic cation transport systems, which are concentrated in the S-1 segment of the proximal tubule. However, p-aminophenol toxicity in vivo is directed toward the S-2 and S-3 segments, which are rich in gamma-glutamyl transpeptidase. These results and the observation that biliary cannulation and glutathione depletion reduce p-aminophenol nephrotoxicity suggest that the biosynthesis of toxic glutathione conjugates is responsible for p-aminophenol nephrotoxicity in vivo. The aminophenol glutathione S-conjugates formed induce p-aminophenol nephrotoxicity by a pathway dependent on gamma-glutamyl transpeptidase.

Plastid-Localized Glutathione Reductase2–Regulated Glutathione Redox Status Is Essential for Arabidopsis Root Apical Meristem Maintenance[C][W

PubMed Central

Yu, Xin; Pasternak, Taras; Eiblmeier, Monika; Ditengou, Franck; Kochersperger, Philip; Sun, Jiaqiang; Wang, Hui; Rennenberg, Heinz; Teale, William; Paponov, Ivan; Zhou, Wenkun; Li, Chuanyou; Li, Xugang; Palme, Klaus

2013-01-01

Glutathione is involved in thiol redox signaling and acts as a major redox buffer against reactive oxygen species, helping to maintain a reducing environment in vivo. Glutathione reductase (GR) catalyzes the reduction of glutathione disulfide (GSSG) into reduced glutathione (GSH). The Arabidopsis thaliana genome encodes two GRs: GR1 and GR2. Whereas the cytosolic/peroxisomal GR1 is not crucial for plant development, we show here that the plastid-localized GR2 is essential for root growth and root apical meristem (RAM) maintenance. We identify a GR2 mutant, miao, that displays strong inhibition of root growth and severe defects in the RAM, with GR activity being reduced to ∼50%. miao accumulates high levels of GSSG and exhibits increased glutathione oxidation. The exogenous application of GSH or the thiol-reducing agent DTT can rescue the root phenotype of miao, demonstrating that the RAM defects in miao are triggered by glutathione oxidation. Our in silico analysis of public microarray data shows that auxin and glutathione redox signaling generally act independently at the transcriptional level. We propose that glutathione redox status is essential for RAM maintenance through both auxin/PLETHORA (PLT)-dependent and auxin/PLT-independent redox signaling pathways. PMID:24249834

Evidence of Dual Mechanisms of Glutathione Uptake in the Rodent Lens: A Novel Role for Vitreous Humor in Lens Glutathione Homeostasis

PubMed Central

Whitson, Jeremy A.; Sell, David R.; Goodman, Michael C.; Monnier, Vincent M.; Fan, Xingjun

2016-01-01

Purpose Lens glutathione synthesis knockout (LEGSKO) mouse lenses lack de novo glutathione (GSH) synthesis but still maintain >1 mM GSH. We sought to determine the source of this residual GSH and the mechanism by which it accumulates in the lens. Methods Levels of GSH, glutathione disulfide (GSSG), and GSH-related compounds were measured in vitro and in vivo using isotope standards and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Results Wild-type (WT) lenses could accumulate GSH from γ-glutamylcysteine and glycine or from intact GSH, but LEGSKO lenses could only accumulate GSH from intact GSH, indicating that LEGSKO lens GSH content is not due to synthesis by a salvage pathway. Uptake of GSH in cultured lenses occurred at the same rate for LEGSKO and WT lenses, could not be inhibited, and occurred primarily through cortical fiber cells. In contrast, uptake of GSH from aqueous humor could be competitively inhibited and showed an enhanced Km in LEGSKO lenses. Mouse vitreous had >1 mM GSH, whereas aqueous had <20 μM GSH. Testing physiologically relevant GSH concentrations for uptake in vivo, we found that both LEGSKO and WT lenses could obtain GSH from the vitreous but not from the aqueous. Vitreous rapidly accumulated GSH from the circulation, and depletion of circulating GSH reduced vitreous but not aqueous GSH. Conclusions The above data provide, for the first time, evidence for the existence of dual mechanisms of GSH uptake into the lens, one mechanism being a passive, high-flux transport through the vitreous exposed side of the lens versus an active, carrier-mediated uptake mechanism at the anterior of the lens. PMID:27472077

Effect of glutathione on phytochelatin synthesis in tomato cells.

PubMed

Mendum, M L; Gupta, S C; Goldsbrough, P B

1990-06-01

Growth of cell suspension cultures of tomato, Lycopersicon esculentum Mill. cv VFNT-Cherry, in the presence of cadmium is inhibited by buthionine sulfoximine, an inhibitor of glutathione synthesis. Cell growth and phytochelatin synthesis are restored to cells treated with buthionine sulfoximine by the addition of glutathione to the medium. Glutathione stimulates the accumulation of phytochelatins in cadmium treated cells, indicating that availability of glutathione can limit synthesis of these peptides. Exogenous glutathione causes a disproportionate increase in the level of smaller phytochelatins, notably [gamma-Glu-Cys](2)-Gly. In the presence of buthionine sulfoximine and glutathione, phytochelatins that are produced upon exposure to cadmium incorporate little [(35)S]cysteine, indicating that these peptides are probably not synthesized by sequential addition of cysteine and glutamate to glutathione.

Melatonin and nitric oxide modulate glutathione content and glutathione reductase activity in sunflower seedling cotyledons accompanying salt stress.

PubMed

Kaur, Harmeet; Bhatla, Satish C

2016-09-30

The present findings demonstrate significant modulation of total glutathione content, reduced glutathione (GSH) content, oxidized glutathione (GSSG) content, GSH/GSSG ratio and glutathione reductase (GR; EC 1.6.4.2) activity in dark-grown seedling cotyledons in response to salt-stress (120 mM NaCl) in sunflower (Helianthus annuus L.) seedlings. A differential spatial distribution of GR activity (monitored by confocal laser scanning microscopic (CLSM) imaging) is also evident. Melatonin and nitric oxide (NO) differentially ameliorate salt stress effect by modulating GR activity and GSH content in seedling cotyledons. Total glutathione content (GSH + GSSG) exhibit a seedling age-dependent increase in the cotyledons, more so in salt-stressed conditions and when subjected to melatonin treatment. Seedlings raised in presence of 15 μM of melatonin exhibit significant increase in GR activity in cotyledon homogenates (10,000 g supernatant) coinciding with significant increase in GSH content. GSSG content and GSH/GSSG ratio also increased due to melatonin treatment. A correlation is thus evident in NaCl-sensitized modulation of GSH content and GR activity by melatonin. GSH content is down regulated by NO provided as 250 μM of sodium nitroprusside (SNP) although total glutathione content remained in similar range. A reversal of response (enhanced total glutathione accumulation) by NO scavenger (cPTIO) highlights the critical role of NO in modulating glutathione homeostasis. SNP lowers the activity of hydroxyindole-O-methyltransferase (HIOMT) - a regulatory enzyme in melatonin biosynthesis in control seedlings whereas its activity is upregulated in salt-stressed seedling cotyledons. Melatonin content of seedling cotyledons is also modulated by NO. NO and melatonin thus seem to modulate GR activity and GSH content during seedling growth under salt stress. Copyright © 2016 Elsevier Inc. All rights reserved.

Quantitative real-time imaging of glutathione

USDA-ARS?s Scientific Manuscript database

Glutathione plays many important roles in biological processes; however, the dynamic changes of glutathione concentrations in living cells remain largely unknown. Here, we report a reversible reaction-based fluorescent probe—designated as RealThiol (RT)—that can quantitatively monitor the real-time ...

Protein disulfide isomerase mediates glutathione depletion-induced cytotoxicity.

PubMed

Okada, Kazushi; Fukui, Masayuki; Zhu, Bao-Ting

2016-08-26

Glutathione depletion is a distinct cause underlying many forms of pathogenesis associated with oxidative stress and cytotoxicity. Earlier studies showed that glutamate-induced glutathione depletion in immortalized murine HT22 hippocampal neuronal cells leads to accumulation of reactive oxygen species (ROS) and ultimately cell death, but the precise mechanism underlying these processes is not clear. Here we show that during the induction of glutathione depletion, nitric oxide (NO) accumulation precedes ROS accumulation. While neuronal NO synthase (nNOS) in untreated HT22 cells exists mostly as a monomer, glutathione depletion results in increased formation of the dimer nNOS, accompanied by increases in the catalytic activity. We identified that nNOS dimerization is catalyzed by protein disulfide isomerase (PDI). Inhibition of PDI's isomerase activity effectively abrogates glutathione depletion-induced conversion of monomer nNOS into dimer nNOS, accumulation of NO and ROS, and cytotoxicity. Furthermore, we found that PDI is present in untreated cells in an inactive S-nitrosylated form, which becomes activated following glutathione depletion via S-denitrosylation. These results reveal a novel role for PDI in mediating glutathione depletion-induced oxidative cytotoxicity, as well as its role as a valuable therapeutic target for protection against oxidative cytotoxicity. Copyright © 2016. Published by Elsevier Inc.

Quantitation of protein S-glutathionylation by liquid chromatograph-tandem mass spectrometry: Correction for contaminating glutathione and glutathione disulfide

USDA-ARS?s Scientific Manuscript database

Protein S-glutathionylation is a posttranslational modification that links oxidative stimuli to reversible changes in cellular function. Protein-glutathione mixed disulfides (PSSG) are commonly quantified by the reduction of the disulfide and detection of the resultant glutathione species. This met...

Defects in a New Class of Sulfate/Anion Transporter Link Sulfur Acclimation Responses to Intracellular Glutathione Levels and Cell Cycle Control1[W][OPEN

PubMed Central

Fang, Su-Chiung; Chung, Chin-Lin; Chen, Chun-Han; Lopez-Paz, Cristina; Umen, James G.

2014-01-01

We previously identified a mutation, suppressor of mating type locus3 15-1 (smt15-1), that partially suppresses the cell cycle defects caused by loss of the retinoblastoma tumor suppressor-related protein encoded by the MAT3 gene in Chlamydomonas reinhardtii. smt15-1 single mutants were also found to have a cell cycle defect leading to a small-cell phenotype. SMT15 belongs to a previously uncharacterized subfamily of putative membrane-localized sulfate/anion transporters that contain a sulfate transporter domain and are found in a widely distributed subset of eukaryotes and bacteria. Although we observed that smt15-1 has a defect in acclimation to sulfur-limited growth conditions, sulfur acclimation (sac) mutants, which are more severely defective for acclimation to sulfur limitation, do not have cell cycle defects and cannot suppress mat3. Moreover, we found that smt15-1, but not sac mutants, overaccumulates glutathione. In wild-type cells, glutathione fluctuated during the cell cycle, with highest levels in mid G1 phase and lower levels during S and M phases, while in smt15-1, glutathione levels remained elevated during S and M. In addition to increased total glutathione levels, smt15-1 cells had an increased reduced-to-oxidized glutathione redox ratio throughout the cell cycle. These data suggest a role for SMT15 in maintaining glutathione homeostasis that impacts the cell cycle and sulfur acclimation responses. PMID:25361960

The mitochondrial dicarboxylate and 2-oxoglutarate carriers do not transport glutathione

PubMed Central

Booty, Lee M.; King, Martin S.; Thangaratnarajah, Chancievan; Majd, Homa; James, Andrew M.; Kunji, Edmund R.S.; Murphy, Michael P.

2015-01-01

Glutathione carries out vital protective roles within mitochondria, but is synthesised in the cytosol. Previous studies have suggested that the mitochondrial dicarboxylate and 2-oxoglutarate carriers were responsible for glutathione uptake. We set out to characterise the putative glutathione transport by using fused membrane vesicles of Lactococcus lactis overexpressing the dicarboxylate and 2-oxoglutarate carriers. Although transport of the canonical substrates could be measured readily, an excess of glutathione did not compete for substrate uptake nor could transport of glutathione be measured directly. Thus these mitochondrial carriers do not transport glutathione and the identity of the mitochondrial glutathione transporter remains unknown. PMID:25637873

[Glutathione participation in regulating the secretory function of the stomach].

PubMed

Shlygin, G K; Vasilevskaia, L S; Martinchik, A N; Ignatenko, L G

1987-01-01

In experiments on dogs with Pavlov's pouches it was shown that glutathione infusion into the blood produced a highly pronounced stimulating effect on the gastric secretion induced by pentagastrin. Endogenous glutathione produced similar effect. It was found that intake as a drink of mono-sodium glutamate led to a significant increase of glutathione concentration in the dogs' blood, that was, probably, the result of its intensified production in the intestinal wall and passing into the blood. The growth of glutathione concentration in the blood coincided with its stimulating effect on the gastric secretion. Glutathione administered separately into the blood, or intake of glutathione without pentagastrin did not produce stimulating effect on gastric secretion. The data presented have evidenced that glutathione, besides its known functions, plays a role of the factor engaged in the regulation of gastric secretion.

The evolution of glutathione metabolism in phototrophic microorganisms

NASA Technical Reports Server (NTRS)

Fahey, R. C.; Buschbacher, R. M.; Newton, G. L.

1987-01-01

Of the many roles ascribed to glutathione (GSH) the one most clearly established is its role in the protection of higher eucaryotes against oxygen toxicity through destruction of thiol-reactive oxygen byproducts. If this is the primary function of GSH then GSH metabolism should have evolved during or after the evolution of oxygenic photosynthesis. That many bacteria do not produce GSH is consistent with this view. In the present study we have examined the low-molecular-weight thiol composition of a variety of phototrophic microorganisms to ascertain how evolution of GSH production is related to evolution of oxygenic photosynthesis. Cells were extracted in the presence of monobromobimane (mBBr) to convert thiols to fluorescent derivatives, which were analyzed by high-pressure liquid chromatography. Significant levels of GSH were not found in the green bacteria (Chlorobium thiosulfatophilum and Chloroflexus aurantiacus). Substantial levels of GSH were present in the purple bacteria (Chromatium vinosum, Rhodospirillum rubrum, Rhodobacter sphaeroides, and Rhodocyclus gelatinosa), the cyanobacteria [Anacystis nidulans, Microcoleus chthonoplastes S.G., Nostoc muscorum, Oscillatoria amphigranulata, Oscillatoria limnetica, Oscillatoria sp. (Stinky Spring, Utah), Oscillatoria terebriformis, Plectonema boryanum, and Synechococcus lividus], and eucaryotic algae (Chlorella pyrenoidsa, Chlorella vulgaris, Euglena gracilis, Scenedesmus obliquus, and Chlamydomonas reinhardtii). Other thiols measured included cysteine, gamma-glutamylcysteine, thiosulfate, coenzyme A, and sulfide; several unidentified thiols were also detected. Many of the organisms examined also exhibited a marked ability to reduce mBBr to syn-(methyl,methyl)bimane, an ability that was quenched by treatment with 2-pyridyl disulfide or 5,5'-bisdithio-(2-nitrobenzoic acid) prior to reaction with mBBr. These observations indicate the presence of a reducing system capable of electron transfer to mBBr and reduction of

The Incomplete Glutathione Puzzle: Just Guessing at Numbers and Figures?

PubMed

Deponte, Marcel

2017-11-20

Glutathione metabolism is comparable to a jigsaw puzzle with too many pieces. It is supposed to comprise (i) the reduction of disulfides, hydroperoxides, sulfenic acids, and nitrosothiols, (ii) the detoxification of aldehydes, xenobiotics, and heavy metals, and (iii) the synthesis of eicosanoids, steroids, and iron-sulfur clusters. In addition, glutathione affects oxidative protein folding and redox signaling. Here, I try to provide an overview on the relevance of glutathione-dependent pathways with an emphasis on quantitative data. Recent Advances: Intracellular redox measurements reveal that the cytosol, the nucleus, and mitochondria contain very little glutathione disulfide and that oxidative challenges are rapidly counterbalanced. Genetic approaches suggest that iron metabolism is the centerpiece of the glutathione puzzle in yeast. Furthermore, recent biochemical studies provide novel insights on glutathione transport processes and uncoupling mechanisms. Which parts of the glutathione puzzle are most relevant? Does this explain the high intracellular concentrations of reduced glutathione? How can iron-sulfur cluster biogenesis, oxidative protein folding, or redox signaling occur at high glutathione concentrations? Answers to these questions not only seem to depend on the organism, cell type, and subcellular compartment but also on different ideologies among researchers. A rational approach to compare the relevance of glutathione-dependent pathways is to combine genetic and quantitative kinetic data. However, there are still many missing pieces and too little is known about the compartment-specific repertoire and concentration of numerous metabolites, substrates, enzymes, and transporters as well as rate constants and enzyme kinetic patterns. Gathering this information might require the development of novel tools but is crucial to address potential kinetic competitions and to decipher uncoupling mechanisms to solve the glutathione puzzle. Antioxid. Redox Signal

Transgenic plants and associated bacteria for phytoremediation of chlorinated compounds.

PubMed

Van Aken, Benoit; Doty, Sharon Lafferty

2010-01-01

Phytoremediation is the use of plants for the treatment of environmental pollution, including chlorinated organics. Although conceptually very attractive, removal and biodegradation of chlorinated pollutants by plants is a rather slow and inefficient process resulting in incomplete treatment and potential release of toxic metabolites into the environment. In order to overcome inherent limitations of plant metabolic capabilities, plants have been genetically modified, following a strategy similar to the development of transgenic crops: genes from bacteria, fungi, and mammals involved in the metabolism of organic contaminants, such as cytochrome P-450 and glutathione S-transferase, have been introduced into higher plants, resulting in significant improvement of tolerance, removal, and degradation of pollutants. Recently, plant-associated bacteria have been recognized playing a significant role in phytoremediation, leading to the development of genetically modified rhizospheric and endophytic bacteria with improved biodegradation capabilities. Transgenic plants and associated bacteria constitute a new generation of genetically modified organisms for efficient and environmental-friendly treatment of polluted soil and water. This review focuses on recent advances in the development of transgenic plants and bacteria for the treatment of chlorinated pollutants, including chlorinated solvents, polychlorinated phenols, and chlorinated herbicides.

The mitochondrial dicarboxylate and 2-oxoglutarate carriers do not transport glutathione.

PubMed

Booty, Lee M; King, Martin S; Thangaratnarajah, Chancievan; Majd, Homa; James, Andrew M; Kunji, Edmund R S; Murphy, Michael P

2015-02-27

Glutathione carries out vital protective roles within mitochondria, but is synthesised in the cytosol. Previous studies have suggested that the mitochondrial dicarboxylate and 2-oxoglutarate carriers were responsible for glutathione uptake. We set out to characterise the putative glutathione transport by using fused membrane vesicles of Lactococcus lactis overexpressing the dicarboxylate and 2-oxoglutarate carriers. Although transport of the canonical substrates could be measured readily, an excess of glutathione did not compete for substrate uptake nor could transport of glutathione be measured directly. Thus these mitochondrial carriers do not transport glutathione and the identity of the mitochondrial glutathione transporter remains unknown. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Effects of polysaccharide peptides from COV-1 strain of Coriolus versicolor on glutathione and glutathione-related enzymes in the mouse.

PubMed

Yeung, John H K; Or, Penelope M Y

2007-06-01

The effects of polysaccharide peptide (PSP), an immunomodulator isolated from Coriolus versicolor COV-1, on glutathione (GSH) and GSH-related enzymes was investigated in C57 mouse. Administration of PSP (1-4 micromole/kg, i.p.) produced a transient, dose-dependent depletion (10-37%) of hepatic GSH, with no effect on serum glutamic-pyruvic transaminase (SGPT) activity. Blood GSH was depleted (6-25%) at 3 h, followed by a rebound increase above the control GSH level (20%) at 18 h. The GSSG/GSH ratio, a measure of oxidative stress, was increased 3 h after PSP treatment but returned to normal levels at 24 h. Sub-chronic treatment of PSP (1-4 micromole/kg/day, i.p.) for seven days did not produce any significant changes in hepatic GSH levels and the GSSG/GSH ratio when measured 24 h after the final dose of PSP. PSP had little effect on glutathione transferase (GST), glutathione reductase (GSSG reductase) and glutathione peroxidase (GPX) activities in the liver. However, a dose-dependent increase in blood GPX activity (30-48%) was observed at 3h, which coincided with the increase in the GSSG/GSH ratio. The increase in blood GPX activity may be a responsive measure to deal with the transient oxidative stress induced by PSP treatment. The results showed that PSP only caused a transient perturbation on hepatic glutathione without affecting the GSH-related enzymes such as GST, GSSG reductase and GPX. The observed changes in blood GSH simply reflected the intra-organ translocation of glutathione, as the glutathione-related enzymes were not significantly affected by PSP treatment.

The antioxidant master glutathione and periodontal health

PubMed Central

Bains, Vivek Kumar; Bains, Rhythm

2015-01-01

Glutathione, considered to be the master antioxidant (AO), is the most-important redox regulator that controls inflammatory processes, and thus damage to the periodontium. Periodontitis patients have reduced total AO capacity in whole saliva, and lower concentrations of reduced glutathione (GSH) in serum and gingival crevicular fluid, and periodontal therapy restores the redox balance. Therapeutic considerations for the adjunctive use of glutathione in management of periodontitis, in limiting the tissue damage associated with oxidative stress, and enhancing wound healing cannot be underestimated, but need to be evaluated further through multi-centered randomized controlled trials. PMID:26604952

The Incomplete Glutathione Puzzle: Just Guessing at Numbers and Figures?

PubMed Central

2017-01-01

Abstract Significance: Glutathione metabolism is comparable to a jigsaw puzzle with too many pieces. It is supposed to comprise (i) the reduction of disulfides, hydroperoxides, sulfenic acids, and nitrosothiols, (ii) the detoxification of aldehydes, xenobiotics, and heavy metals, and (iii) the synthesis of eicosanoids, steroids, and iron–sulfur clusters. In addition, glutathione affects oxidative protein folding and redox signaling. Here, I try to provide an overview on the relevance of glutathione-dependent pathways with an emphasis on quantitative data. Recent Advances: Intracellular redox measurements reveal that the cytosol, the nucleus, and mitochondria contain very little glutathione disulfide and that oxidative challenges are rapidly counterbalanced. Genetic approaches suggest that iron metabolism is the centerpiece of the glutathione puzzle in yeast. Furthermore, recent biochemical studies provide novel insights on glutathione transport processes and uncoupling mechanisms. Critical Issues: Which parts of the glutathione puzzle are most relevant? Does this explain the high intracellular concentrations of reduced glutathione? How can iron–sulfur cluster biogenesis, oxidative protein folding, or redox signaling occur at high glutathione concentrations? Answers to these questions not only seem to depend on the organism, cell type, and subcellular compartment but also on different ideologies among researchers. Future Directions: A rational approach to compare the relevance of glutathione-dependent pathways is to combine genetic and quantitative kinetic data. However, there are still many missing pieces and too little is known about the compartment-specific repertoire and concentration of numerous metabolites, substrates, enzymes, and transporters as well as rate constants and enzyme kinetic patterns. Gathering this information might require the development of novel tools but is crucial to address potential kinetic competitions and to decipher uncoupling

Conjugations with glutathione. The enzymic conjugation of some chlorocyclohexenes

PubMed Central

Sims, P.; Grover, P. L.

1965-01-01

1. α-3,4,5,6-Tetrachlorocyclohex-1-ene and γ-2,3,4,5,6-pentachlorocyclohex-1-ene are conjugated with glutathione in vitro by a rat-liver enzyme that is probably glutathione S-aryltransferase. 2. Chlorocyclohexane and the α-, β-, γ- and δ-isomers of hexachlorocyclohexane were not substrates for rat-liver glutathione S-aryltransferase. 3. Glutathione-S-aryltransferase activity was present in tissue preparations of houseflies of insecticide-resistant and -susceptible strains. More activity was found in a dieldrin-resistant strain of houseflies fed on dieldrin than in either a dieldrin-resistant strain not fed on dieldrin or a control strain of dieldrin-susceptible houseflies. 4. Housefly soluble supernatant preparations converted S-(2-chloro-4-nitrophenyl)glutathione into the corresponding cysteine and mercapturic acid derivatives. PMID:14333551

Widespread Fosfomycin Resistance in Gram-Negative Bacteria Attributable to the Chromosomal fosA Gene

PubMed Central

Ito, Ryota; Tomich, Adam D.; Callaghan, Jake D.; McElheny, Christi L.; Mettus, Roberta T.; Sluis-Cremer, Nicolas

2017-01-01

ABSTRACT Fosfomycin is a decades-old antibiotic which is being revisited because of its perceived activity against many extensively drug-resistant Gram-negative pathogens. FosA proteins are Mn2+ and K+-dependent glutathione S-transferases which confer fosfomycin resistance in Gram-negative bacteria by conjugation of glutathione to the antibiotic. Plasmid-borne fosA variants have been reported in fosfomycin-resistant Escherichia coli strains. However, the prevalence and distribution of fosA in other Gram-negative bacteria are not known. We systematically surveyed the presence of fosA in Gram-negative bacteria in over 18,000 published genomes from 18 Gram-negative species and investigated their contribution to fosfomycin resistance. We show that FosA homologues are present in the majority of genomes in some species (e.g., Klebsiella spp., Enterobacter spp., Serratia marcescens, and Pseudomonas aeruginosa), whereas they are largely absent in others (e.g., E. coli, Acinetobacter baumannii, and Burkholderia cepacia). FosA proteins in different bacterial pathogens are highly divergent, but key amino acid residues in the active site are conserved. Chromosomal fosA genes conferred high-level fosfomycin resistance when expressed in E. coli, and deletion of chromosomal fosA in S. marcescens eliminated fosfomycin resistance. Our results indicate that FosA is encoded by clinically relevant Gram-negative species and contributes to intrinsic fosfomycin resistance. PMID:28851843

21 CFR 864.7375 - Glutathione reductase assay.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Glutathione reductase assay. 864.7375 Section 864.7375 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7375 Glutathione...

21 CFR 864.7375 - Glutathione reductase assay.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Glutathione reductase assay. 864.7375 Section 864.7375 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7375 Glutathione...

21 CFR 864.7375 - Glutathione reductase assay.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Glutathione reductase assay. 864.7375 Section 864.7375 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7375 Glutathione...

21 CFR 864.7375 - Glutathione reductase assay.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Glutathione reductase assay. 864.7375 Section 864.7375 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7375 Glutathione...

21 CFR 864.7375 - Glutathione reductase assay.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Glutathione reductase assay. 864.7375 Section 864.7375 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7375 Glutathione...

The aerobic activity of metronidazole against anaerobic bacteria.

PubMed

Dione, Niokhor; Khelaifia, Saber; Lagier, Jean-Christophe; Raoult, Didier

2015-05-01

Recently, the aerobic growth of strictly anaerobic bacteria was demonstrated using antioxidants. Metronidazole is frequently used to treat infections caused by anaerobic bacteria; however, to date its antibacterial activity was only tested in anaerobic conditions. Here we aerobically tested using antioxidants the in vitro activities of metronidazole, gentamicin, doxycycline and imipenem against 10 common anaerobic and aerobic bacteria. In vitro susceptibility testing was performed by the disk diffusion method, and minimum inhibitory concentrations (MICs) were determined by Etest. Aerobic culture of the bacteria was performed at 37°C using Schaedler agar medium supplemented with 1mg/mL ascorbic acid and 0.1mg/mL glutathione; the pH was adjusted to 7.2 by 10M KOH. Growth of anaerobic bacteria cultured aerobically using antioxidants was inhibited by metronidazole after 72h of incubation at 37°C, with a mean inhibition diameter of 37.76mm and an MIC of 1μg/mL; however, strains remained non-sensitive to gentamicin. No growth inhibition of aerobic bacteria was observed after 24h of incubation at 37°C with metronidazole; however, inhibition was observed with doxycycline and imipenem used as controls. These results indicate that bacterial sensitivity to metronidazole is not related to the oxygen tension but is a result of the sensitivity of the micro-organism. In future, both culture and antibiotic susceptibility testing of strictly anaerobic bacteria will be performed in an aerobic atmosphere using antioxidants in clinical microbiology laboratories. Copyright © 2015 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

Effect of glutathione on phytochelatin synthesis in tomato cells. [Lycopersicon esculentum

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mendum, M.L.; Gupta, S.C.; Goldsbrough, P.B.

Growth of cell suspension cultures of tomato, Lycopersicon esculentum Mill. cv VFNT-Cherry, in the presence of cadmium is inhibited by buthionine sulfoximine, an inhibitor of glutathione synthesis. Cell growth and phytochelatin synthesis are restored to cells treated with buthionine sulfoximine by the addition of glutathione to the medium. Glutathione stimulates the accumulation of phytochelatins in cadmium treated cells, indicating that availability of glutathione can limit synthesis of these peptides. Exogenous glutathione causes a disproportionate increase in the level of smaller phytochelatins, notably ({gamma}-Glu-Cys){sub 2}-Gly. In the presence of buthionine sulfoximine and glutathione, phytochelatins that are produced upon exposure to cadmiummore » incorporate little ({sup 35}S)cysteine, indicating that these peptides are probably not synthesized by sequential addition of cysteine and glutamate to glutathione.« less

Glutathione transferases, regulators of cellular metabolism and physiology.

PubMed

Board, Philip G; Menon, Deepthi

2013-05-01

The cytosolic glutathione transferases (GSTs) comprise a super family of proteins that can be categorized into multiple classes with a mixture of highly specific and overlapping functions. The review covers the genetics, structure and function of the human cytosolic GSTs with particular attention to their emerging roles in cellular metabolism. All the catalytically active GSTs contribute to the glutathione conjugation or glutathione dependant-biotransformation of xenobiotics and many catalyze glutathione peroxidase or thiol transferase reactions. GSTs also catalyze glutathione dependent isomerization reactions required for the synthesis of several prostaglandins and steroid hormones and the catabolism of tyrosine. An increasing body of work has implicated several GSTs in the regulation of cell signaling pathways mediated by stress-activated kinases like Jun N-terminal kinase. In addition, some members of the cytosolic GST family have been shown to form ion channels in intracellular membranes and to modulate ryanodine receptor Ca(2+) channels in skeletal and cardiac muscle. In addition to their well established roles in the conjugation and biotransformation of xenobiotics, GSTs have emerged as significant regulators of pathways determining cell proliferation and survival and as regulators of ryanodine receptors that are essential for muscle function. This article is part of a Special Issue entitled Cellular functions of glutathione. Copyright © 2012 Elsevier B.V. All rights reserved.

Mechanism-based biomarker gene sets for glutathione depletion-related hepatotoxicity in rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao Weihua; Mizukawa, Yumiko; Nakatsu, Noriyuki

Chemical-induced glutathione depletion is thought to be caused by two types of toxicological mechanisms: PHO-type glutathione depletion [glutathione conjugated with chemicals such as phorone (PHO) or diethyl maleate (DEM)], and BSO-type glutathione depletion [i.e., glutathione synthesis inhibited by chemicals such as L-buthionine-sulfoximine (BSO)]. In order to identify mechanism-based biomarker gene sets for glutathione depletion in rat liver, male SD rats were treated with various chemicals including PHO (40, 120 and 400 mg/kg), DEM (80, 240 and 800 mg/kg), BSO (150, 450 and 1500 mg/kg), and bromobenzene (BBZ, 10, 100 and 300 mg/kg). Liver samples were taken 3, 6, 9 andmore » 24 h after administration and examined for hepatic glutathione content, physiological and pathological changes, and gene expression changes using Affymetrix GeneChip Arrays. To identify differentially expressed probe sets in response to glutathione depletion, we focused on the following two courses of events for the two types of mechanisms of glutathione depletion: a) gene expression changes occurring simultaneously in response to glutathione depletion, and b) gene expression changes after glutathione was depleted. The gene expression profiles of the identified probe sets for the two types of glutathione depletion differed markedly at times during and after glutathione depletion, whereas Srxn1 was markedly increased for both types as glutathione was depleted, suggesting that Srxn1 is a key molecule in oxidative stress related to glutathione. The extracted probe sets were refined and verified using various compounds including 13 additional positive or negative compounds, and they established two useful marker sets. One contained three probe sets (Akr7a3, Trib3 and Gstp1) that could detect conjugation-type glutathione depletors any time within 24 h after dosing, and the other contained 14 probe sets that could detect glutathione depletors by any mechanism. These two sets, with appropriate

Do glutathione levels decline in aging human brain?

PubMed

Tong, Junchao; Fitzmaurice, Paul S; Moszczynska, Anna; Mattina, Katie; Ang, Lee-Cyn; Boileau, Isabelle; Furukawa, Yoshiaki; Sailasuta, Napapon; Kish, Stephen J

2016-04-01

For the past 60 years a major theory of "aging" is that age-related damage is largely caused by excessive uncompensated oxidative stress. The ubiquitous tripeptide glutathione is a major antioxidant defense mechanism against reactive free radicals and has also served as a marker of changes in oxidative stress. Some (albeit conflicting) animal data suggest a loss of glutathione in brain senescence, which might compromise the ability of the aging brain to meet the demands of oxidative stress. Our objective was to establish whether advancing age is associated with glutathione deficiency in human brain. We measured reduced glutathione (GSH) levels in multiple regions of autopsied brain of normal subjects (n=74) aged one day to 99 years. Brain GSH levels during the infancy/teenage years were generally similar to those in the oldest examined adult group (76-99 years). During adulthood (23-99 years) GSH levels remained either stable (occipital cortex) or increased (caudate nucleus, frontal and cerebellar cortices). To the extent that GSH levels represent glutathione antioxidant capacity, our postmortem data suggest that human brain aging is not associated with declining glutathione status. We suggest that aged healthy human brains can maintain antioxidant capacity related to glutathione and that an age-related increase in GSH levels in some brain regions might possibly be a compensatory response to increased oxidative stress. Since our findings, although suggestive, suffer from the generic limitations of all postmortem brain studies, we also suggest the need for "replication" investigations employing the new (1)H MRS imaging procedures in living human brain. Copyright © 2016 Elsevier Inc. All rights reserved.

Involvement of glutathione/glutathione S-transferase antioxidant system in butyrate-inhibited vascular smooth muscle cell proliferation.

PubMed

Ranganna, Kasturi; Mathew, Omana P; Yatsu, Frank M; Yousefipour, Zivar; Hayes, Barbara E; Milton, Shirlette G

2007-11-01

Vascular smooth muscle cell (VSMC) proliferation is an important etiological factor in vascular proliferative diseases such as primary atherosclerosis, hypertension, arterial and in-stent restenosis, and transplant vasculopathy. Our studies established that butyrate, a bacterial fermentation product of dietary fiber and a chromatin modulator, is a potent inhibitor of VSMC proliferation. The cardiovascular health benefits of a high-fiber diet, the principle source of butyrate in the body, have been known for a long time, however, very little is known about the antiatherogenic potential of butyrate. Because oxidative stress plays an important role in the pathogenesis of atherosclerosis, we examined involvement of the glutathione/glutathione S-transferase (GST) antioxidant system in butyrate's inhibition of VSMC proliferation. Treatment of proliferating VSMCs with butyrate leads to the induction of several GSTs. Interestingly, our study also demonstrated the nuclear localization of GST-P1 (GST-7-7), which is considered to be a cytosolic protein; this was demonstrated using immunostaining and was corroborated by western blotting. Also, the butyrate-induced antiproliferative action, and the induction of GST-P1 and its nuclear localization are downregulated when butyrate is withdrawn. Furthermore, assessment of intracellular glutathione levels reveals their augmentation by butyrate. Conversely, butyrate treatment reduces the levels of reactive oxygen species in VSMCs. Collectively, the butyrate-treatment-related increase in glutathione content, the reduction in reactive oxygen species, the upregulation of GST and the nuclear localization of GST-P1 in growth-arrested VSMCs imply that butyrate's antiproliferative action involves modulation of the cellular redox state. Thus, induction of the glutathione/GST antioxidant system appears to have other regulatory role(s) besides detoxification and regulation of the cellular redox state, for example, cell-cycle control and cell

Lack of genomic evidence of AI-2 receptors suggests a non-quorum sensing role for luxS in most bacteria

PubMed Central

Rezzonico, Fabio; Duffy, Brion

2008-01-01

Background Great excitement accompanied discoveries over the last decade in several Gram-negative and Gram-positive bacteria of the LuxS protein, which catalyzes production of the AI-2 autoinducer molecule for a second quorum sensing system (QS-2). Since the luxS gene was found to be widespread among the most diverse bacterial taxa, it was hypothesized that AI-2 may constitute the basis of a universal microbial language, a kind of bacterial Esperanto. Many of the studies published in this field have drawn a direct correlation between the occurrence of the luxS gene in a given organism and the presence and functionality of a QS-2 therein. However, rarely hathe existence of potential AI-2 receptors been examined. This is important, since it is now well recognized that LuxS also holds a central role as a metabolic enzyme in the activated methyl cycle which is responsible for the generation of S-adenosyl-L-methionine, the major methyl donor in the cell. Results In order to assess whether the role of LuxS in these bacteria is indeed related to AI-2 mediated quorum sensing we analyzed genomic databases searching for established AI-2 receptors (i.e., LuxPQ-receptor of Vibrio harveyi and Lsr ABC-transporter of Salmonella typhimurium) and other presumed QS-related proteins and compared the outcome with published results about the role of QS-2 in these organisms. An unequivocal AI-2 related behavior was restricted primarily to organisms bearing known AI-2 receptor genes, while phenotypes of luxS mutant bacteria lacking these genes could often be explained simply by assuming deficiencies in sulfur metabolism. Conclusion Genomic analysis shows that while LuxPQ is restricted to Vibrionales, the Lsr-receptor complex is mainly present in pathogenic bacteria associated with endotherms. This suggests that QS-2 may play an important role in interactions with animal hosts. In most other species, however, the role of LuxS appears to be limited to metabolism, although in a few cases

Lack of genomic evidence of AI-2 receptors suggests a non-quorum sensing role for luxS in most bacteria.

PubMed

Rezzonico, Fabio; Duffy, Brion

2008-09-20

Great excitement accompanied discoveries over the last decade in several Gram-negative and Gram-positive bacteria of the LuxS protein, which catalyzes production of the AI-2 autoinducer molecule for a second quorum sensing system (QS-2). Since the luxS gene was found to be widespread among the most diverse bacterial taxa, it was hypothesized that AI-2 may constitute the basis of a universal microbial language, a kind of bacterial Esperanto. Many of the studies published in this field have drawn a direct correlation between the occurrence of the luxS gene in a given organism and the presence and functionality of a QS-2 therein. However, rarely hathe existence of potential AI-2 receptors been examined. This is important, since it is now well recognized that LuxS also holds a central role as a metabolic enzyme in the activated methyl cycle which is responsible for the generation of S-adenosyl-L-methionine, the major methyl donor in the cell. In order to assess whether the role of LuxS in these bacteria is indeed related to AI-2 mediated quorum sensing we analyzed genomic databases searching for established AI-2 receptors (i.e., LuxPQ-receptor of Vibrio harveyi and Lsr ABC-transporter of Salmonella typhimurium) and other presumed QS-related proteins and compared the outcome with published results about the role of QS-2 in these organisms. An unequivocal AI-2 related behavior was restricted primarily to organisms bearing known AI-2 receptor genes, while phenotypes of luxS mutant bacteria lacking these genes could often be explained simply by assuming deficiencies in sulfur metabolism. Genomic analysis shows that while LuxPQ is restricted to Vibrionales, the Lsr-receptor complex is mainly present in pathogenic bacteria associated with endotherms. This suggests that QS-2 may play an important role in interactions with animal hosts. In most other species, however, the role of LuxS appears to be limited to metabolism, although in a few cases the presence of yet unknown

High resolution imaging of subcellular glutathione concentrations by quantitative immunoelectron microscopy in different leaf areas of Arabidopsis

PubMed Central

Koffler, Barbara E.; Bloem, Elke; Zellnig, Günther; Zechmann, Bernd

2013-01-01

Glutathione is an important antioxidant and redox buffer in plants. It fulfills many important roles during plant development, defense and is essential for plant metabolism. Even though the compartment specific roles of glutathione during abiotic and biotic stress situations have been studied in detail there is still great lack of knowledge about subcellular glutathione concentrations within the different leaf areas at different stages of development. In this study a method is described that allows the calculation of compartment specific glutathione concentrations in all cell compartments simultaneously in one experiment by using quantitative immunogold electron microscopy combined with biochemical methods in different leaf areas of Arabidopsis thaliana Col-0 (center of the leaf, leaf apex, leaf base and leaf edge). The volume of subcellular compartments in the mesophyll of Arabidopsis was found to be similar to other plants. Vacuoles covered the largest volume within a mesophyll cell and increased with leaf age (up to 80% in the leaf apex of older leaves). Behind vacuoles, chloroplasts covered the second largest volume (up to 20% in the leaf edge of the younger leaves) followed by nuclei (up to 2.3% in the leaf edge of the younger leaves), mitochondria (up to 1.6% in the leaf apex of the younger leaves), and peroxisomes (up to 0.3% in the leaf apex of the younger leaves). These values together with volumes of the mesophyll determined by stereological methods from light and electron micrographs and global glutathione contents measured with biochemical methods enabled the determination of subcellular glutathione contents in mM. Even though biochemical investigations did not reveal differences in global glutathione contents, compartment specific differences could be observed in some cell compartments within the different leaf areas. Highest concentrations of glutathione were always found in mitochondria, where values in a range between 8.7 mM (in the apex of younger

Induction of glutathione synthesis and glutathione reductase activity by abiotic stresses in maize and wheat.

PubMed

Kocsy, Gábor; Szalai, Gabriella; Galiba, Gábor

2002-06-21

The effect of different abiotic stresses (extreme temperatures and osmotic stress) on the synthesis of glutathione and hydroxymethylglutathione, on the ratio of the reduced to oxidised forms of these thiols (GSH/GSSG, hmGSH/hmGSSG), and on the glutathione reductase (GR) activity was studied in maize and wheat genotypes having different sensitivity to low temperature stress. Cold treatment induced a greater increase in total glutathione (TG) content and in GR activity in tolerant genotypes of both species than in sensitive ones. The GSH/GSSG and hmGSH/hmGSSG ratios were increased by this treatment only in the frost-tolerant wheat variety. High-temperature stress increased the TG content and the GSH/GSSG ratio only in the chilling-sensitive maize genotype, but GR activity was greater after this treatment in both maize genotypes. Osmotic stress resulted in a great increase in the TG content in wheat and the GR activity in maize. The amount of total hydroxymethylglutathione increased following all stress treatments. These results indicate the involvement of these antioxidants in the stress responses of wheat and maize.

Interaction of glutathione with bovine serum albumin: Spectroscopy and molecular docking.

PubMed

Jahanban-Esfahlan, Ali; Panahi-Azar, Vahid

2016-07-01

This study aims to investigate the interaction between glutathione and bovine serum albumin (BSA) using ultraviolet-visible (UV-vis) absorption, fluorescence spectroscopies under simulated physiological conditions (pH 7.4) and molecular docking methods. The results of fluorescence spectroscopy indicated that the fluorescence intensity of BSA was decreased considerably upon the addition of glutathione through a static quenching mechanism. The fluorescence quenching obtained was related to the formation of BSA-glutathione complex. The values of KSV, Ka and Kb for the glutathione and BSA interaction were in the order of 10(5). The thermodynamic parameters including enthalpy change (ΔH), entropy change (ΔS) and also Gibb's free energy (ΔG) were determined using Van't Hoff equation. These values showed that hydrogen bonding and van der Waals forces were the main interactions in the binding of glutathione to BSA and the stabilization of the complex. Also, the interaction of glutathione and BSA was spontaneous. The effects of glutathione on the BSA conformation were determined using UV-vis spectroscopy. Moreover, glutathione was docked in BSA using ArgusLab as a molecular docking program. It was recognized that glutathione binds within the sub-domain IIA pocket in domain II of BSA. Copyright © 2016 Elsevier Ltd. All rights reserved.

Impact of glutathione metabolism on zinc homeostasis in Saccharomyces cerevisiae.

PubMed

Steiger, Matthias G; Patzschke, Anett; Holz, Caterina; Lang, Christine; Causon, Tim; Hann, Stephan; Mattanovich, Diethard; Sauer, Michael

2017-06-01

Zinc is a crucial mineral for all organisms as it is an essential cofactor for the proper function of a plethora of proteins and depletion of zinc causes oxidative stress. Glutathione is the major redox buffering agent in the cell and therefore important for mitigation of the adverse effects of oxidative stress. In mammalian cells, zinc deficiency is accompanied by a glutathione depletion. In the yeast Saccharomyces cerevisiae, the opposite effect is observed: under low zinc conditions, an elevated glutathione concentration is found. The main regulator to overcome zinc deficiency is Zap1p. However, we show that Zap1p is not involved in this glutathione accumulation phenotype. Furthermore, we found that in glutathione-accumulating strains also the metal ion-binding phytochelatin-2, which is an oligomer of glutathione, is accumulated. This increased phytochelatin concentration correlates with a lower free zinc level in the vacuole. These results suggest that phytochelatin is important for zinc buffering in S. cerevisiae and thus explains how zinc homeostasis is connected with glutathione metabolism. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Targeting Aberrant Glutathione Metabolism to Eradicate Human Acute Myelogenous Leukemia Cells*

PubMed Central

Pei, Shanshan; Minhajuddin, Mohammad; Callahan, Kevin P.; Balys, Marlene; Ashton, John M.; Neering, Sarah J.; Lagadinou, Eleni D.; Corbett, Cheryl; Ye, Haobin; Liesveld, Jane L.; O'Dwyer, Kristen M.; Li, Zheng; Shi, Lei; Greninger, Patricia; Settleman, Jeffrey; Benes, Cyril; Hagen, Fred K.; Munger, Joshua; Crooks, Peter A.; Becker, Michael W.; Jordan, Craig T.

2013-01-01

The development of strategies to eradicate primary human acute myelogenous leukemia (AML) cells is a major challenge to the leukemia research field. In particular, primitive leukemia cells, often termed leukemia stem cells, are typically refractory to many forms of therapy. To investigate improved strategies for targeting of human AML cells we compared the molecular mechanisms regulating oxidative state in primitive (CD34+) leukemic versus normal specimens. Our data indicate that CD34+ AML cells have elevated expression of multiple glutathione pathway regulatory proteins, presumably as a mechanism to compensate for increased oxidative stress in leukemic cells. Consistent with this observation, CD34+ AML cells have lower levels of reduced glutathione and increased levels of oxidized glutathione compared with normal CD34+ cells. These findings led us to hypothesize that AML cells will be hypersensitive to inhibition of glutathione metabolism. To test this premise, we identified compounds such as parthenolide (PTL) or piperlongumine that induce almost complete glutathione depletion and severe cell death in CD34+ AML cells. Importantly, these compounds only induce limited and transient glutathione depletion as well as significantly less toxicity in normal CD34+ cells. We further determined that PTL perturbs glutathione homeostasis by a multifactorial mechanism, which includes inhibiting key glutathione metabolic enzymes (GCLC and GPX1), as well as direct depletion of glutathione. These findings demonstrate that primitive leukemia cells are uniquely sensitive to agents that target aberrant glutathione metabolism, an intrinsic property of primary human AML cells. PMID:24089526

Mushrooms: A rich source of the antioxidants ergothioneine and glutathione.

PubMed

Kalaras, Michael D; Richie, John P; Calcagnotto, Ana; Beelman, Robert B

2017-10-15

While mushrooms are the highest dietary source for the unique sulfur-containing antioxidant ergothioneine, little is known regarding levels of the major biological antioxidant glutathione. Thus, our objectives were to determine and compare levels of glutathione, as well as ergothioneine, in different species of mushrooms. Glutathione levels varied >20-fold (0.11-2.41mg/gdw) with some varieties having higher levels than reported for other foods. Ergothioneine levels also varied widely (0.15-7.27mg/gdw) and were highly correlated with those of glutathione (r=0.62, P<0.001). Both antioxidants were more concentrated in pileus than stipe tissues in selected mushrooms species. Agaricus bisporus harvested during the third cropping flush contained higher levels of ergothioneine and glutathione compared to the first flush, possibly as a response to increased oxidative stress. This study demonstrated that certain mushroom species are high in glutathione and ergothioneine and should be considered an excellent dietary source of these important antioxidants. Copyright © 2017 Elsevier Ltd. All rights reserved.

Glutathione, Glutaredoxins, and Iron.

PubMed

Berndt, Carsten; Lillig, Christopher Horst

2017-11-20

Glutathione (GSH) is the most abundant cellular low-molecular-weight thiol in the majority of organisms in all kingdoms of life. Therefore, functions of GSH and disturbed regulation of its concentration are associated with numerous physiological and pathological situations. Recent Advances: The function of GSH as redox buffer or antioxidant is increasingly being questioned. New functions, especially functions connected to the cellular iron homeostasis, were elucidated. Via the formation of iron complexes, GSH is an important player in all aspects of iron metabolism: sensing and regulation of iron levels, iron trafficking, and biosynthesis of iron cofactors. The variety of GSH coordinated iron complexes and their functions with a special focus on FeS-glutaredoxins are summarized in this review. Interestingly, GSH analogues that function as major low-molecular-weight thiols in organisms lacking GSH resemble the functions in iron homeostasis. Since these iron-related functions are most likely also connected to thiol redox chemistry, it is difficult to distinguish between mechanisms related to either redox or iron metabolisms. The ability of GSH to coordinate iron in different complexes with or without proteins needs further investigation. The discovery of new Fe-GSH complexes and their physiological functions will significantly advance our understanding of cellular iron homeostasis. Antioxid. Redox Signal. 27, 1235-1251.

Stabilization of anthocyanins in blackberry juice by glutathione fortification.

PubMed

Stebbins, Nathan B; Howard, Luke R; Prior, Ronald L; Brownmiller, Cindi; Mauromoustakos, Andy

2017-10-18

Blackberry anthocyanins provide attractive color and antioxidant activity. However, anthocyanins degrade during juice processing and storage, so maintaining high anthocyanin concentrations in berry juices may lead to greater antioxidant and health benefits for the consumer. This study evaluated potential additives to stabilize anthocyanins during blackberry juice storage. The anthocyanin stabilizing agents used were: glutathione, galacturonic acid, diethylenetriaminepentaacetic acid and tannic acid, which were added at a level of 500 mg L -1 . Juice anthocyanin, flavonol, and ellagitannin content and percent polymeric color were measured over five weeks of accelerated storage at 30 °C. Glutathione had the greatest protective effect on total anthocyanins and polymeric color. Therefore a second study was performed with glutathione in combination with lipoic and ascorbic acids in an effort to use antioxidant recycling to achieve a synergistic effect. However, the antioxidant recycling system had no protective effect relative to glutathione alone. Glutathione appears to be a promising blackberry juice additive to protect against anthocyanin degradation during storage.

Enzymatic activity necessary to restore the lethality due to Escherichia coli RNase E deficiency is distributed among bacteria lacking RNase E homologues

PubMed Central

Kageyama, Daisuke; Honda, Naoko; Fujimoto, Hirofumi; Kato, Atsushi

2017-01-01

Escherichia coli RNase E (Eco-RNase E), encoded by rne (Eco-rne), is considered the global RNA decay initiator. Although Eco-RNase E is an essential gene product in E. coli, some bacterial species, such as Bacillus subtilis, do not possess Eco-RNase E sequence homologues. B. subtilis instead possesses RNase J1/J2 (Bsu-RNase J1/J2) and RNase Y (Bsu-RNase Y) to execute RNA decay. Here we found that E. coli lacking the Eco-rne gene (Δrne E. coli) was viable conditional on M9 minimal media by introducing Bsu-RNase J1/J2 or Bsu-RNase Y. We also cloned an extremely short Eco-RNase E homologue (Wpi-RNase E) and a canonical sized Bsu-RNase J1/J2 homologue (Wpi-RNase J) from Wolbachia pipientis, an α-proteobacterial endosymbiont of arthropods. We found that Wpi-RNase J restored the colony-forming ability (CFA) of Δrne E. coli, whereas Wpi-RNase E did not. Unexpectedly, Wpi-RNase E restored defective CFA due to lack of Eco-RNase G, a paralogue of Eco-RNase E. Our results indicate that bacterial species that lack Eco-RNase E homologues or bacterial species that possess Eco-RNase E homologues which lack Eco-RNase E-like activities have a modest Eco-RNase E-like function using RNase J and/or RNase Y. These results suggest that Eco-RNase E-like activities might distribute among a wide array of bacteria and that functions of RNases may have changed dynamically during evolutionary divergence of bacterial lineages. PMID:28542621

Glutathione maintenance is crucial for survival of melanocytes after exposure to rhododendrol.

PubMed

Kondo, Masatoshi; Kawabata, Keigo; Sato, Kohji; Yamaguchi, Sayuri; Hachiya, Akira; Takahashi, Yoshito; Inoue, Shintaro

2016-09-01

Rhododendrol is a phenolic compound that shows a tyrosinase-dependent toxicity for melanocytes and occasionally induces a vitiligo-like skin depigmentation. The post-tyrosinase mechanisms determining melanocyte death or survival, however, are far from clear. Here, we find that rhododendrol treatment leads to a reduction in the levels of cellular glutathione but also induces a cellular antioxidant response that eventually increases glutathione levels. We further find that rhododendrol toxicity is enhanced when glutathione levels are experimentally reduced and alleviated when glutathione levels are increased. Hence, it appears that the size of the preexisting glutathione pool along with the capacity to supply glutathione via the antioxidant response determines whether melanocytes survive or die after rhododendrol exposure. It is conceivable, therefore, that rhododendrol-induced leukoderma depends on the capacity to maintain appropriate glutathione levels and that enhancement of glutathione levels may preserve a patient's melanocytes and potentially help in repigmentation. © 2016 The Authors. Pigment Cell & Melanoma Research published by John Wiley & Sons Ltd.

Glutathione Efflux and Cell Death

PubMed Central

2012-01-01

Abstract Significance: Glutathione (GSH) depletion is a central signaling event that regulates the activation of cell death pathways. GSH depletion is often taken as a marker of oxidative stress and thus, as a consequence of its antioxidant properties scavenging reactive species of both oxygen and nitrogen (ROS/RNS). Recent Advances: There is increasing evidence demonstrating that GSH loss is an active phenomenon regulating the redox signaling events modulating cell death activation and progression. Critical Issues: In this work, we review the role of GSH depletion by its efflux, as an important event regulating alterations in the cellular redox balance during cell death independent from oxidative stress and ROS/RNS formation. We discuss the mechanisms involved in GSH efflux during cell death progression and the redox signaling events by which GSH depletion regulates the activation of the cell death machinery. Future Directions: The evidence summarized here clearly places GSH transport as a central mechanism mediating redox signaling during cell death progression. Future studies should be directed toward identifying the molecular identity of GSH transporters mediating GSH extrusion during cell death, and addressing the lack of sensitive approaches to quantify GSH efflux. Antioxid. Redox Signal. 17, 1694–1713. PMID:22656858

Directed evolution of glutathione transferases towards a selective glutathione-binding site and improved oxidative stability.

PubMed

Axarli, Irine; Muleta, Abdi W; Chronopoulou, Evangelia G; Papageorgiou, Anastassios C; Labrou, Nikolaos E

2017-01-01

Glutathione transferases (GSTs) are a family of detoxification enzymes that catalyze the conjugation of glutathione (GSH) to electrophilic compounds. A library of alpha class GSTs was constructed by DNA shuffling using the DNA encoding the human glutathione transferase A1-1 (hGSTA1-1) and the rat glutathione transferase A1-1 (rGSTA1-1). Activity screening of the library allowed the selection of a chimeric enzyme variant (GSTD4) that displayed high affinity towards GSH and GSH-Sepharose affinity adsorbent, higher k cat /K m and improved thermal stability, compared to the parent enzymes. The crystal structures of the GSTD4 enzyme in free form and in complex with GSH were determined to 1.6Šand 2.3Šresolution, respectively. Analysis of the GSTD4 structure showed subtle conformational changes in the GSH-binding site and in electron-sharing network that may contribute to the increased GSH affinity. The shuffled variant GSTD4 was further optimized for improved oxidative stability employing site-saturation mutagenesis. The Cys112Ser mutation confers optimal oxidative stability and kinetic properties in the GSTD4 enzyme. DNA shuffling allowed the creation of a chimeric enzyme variant with improved properties, compared to the parent enzymes. X-ray crystallography shed light on how recombination of a specific segment from homologous GSTA1-1 together with point mutations gives rise to a new functionally competent enzyme with improved binding, catalytic properties and stability. Such an engineered GST would be useful in biotechnology as affinity tool in affinity chromatography as well as a biocatalytic matrix for the construction of biochips or enzyme biosensors. Copyright © 2016 Elsevier B.V. All rights reserved.

Effect of fish oil on glutathione redox system in multiple sclerosis

PubMed Central

Sorto-Gomez, Tania E; Ortiz, Genaro G; Pacheco-Moises, Fermín P; Torres-Sanchez, Erandis D; Ramirez-Ramirez, Viridiana; Macias-Islas, Miguel A; de la Rosa, Alfredo Celis; Velázquez-Brizuela, Irma E

2016-01-01

Multiple sclerosis (MS) is a chronic, inflammatory and autoimmune disease of the central nervous system. Dysregulation of glutathione homeostasis and alterations in glutathione-dependent enzyme activities are implicated in the induction and progression of MS. Evidence suggests that Omega-3 polyunsaturated fatty acids (PUFAs) have anti-inflammatory, antioxidant and neuroprotective effects. The aim of the present work was to evaluate the effect of fish oil on the activity of glutathione reductase (GR), content of reduced and oxidized glutathione, and GSH/GSSG ratio in MS. 50 patients with relapsing-remitting MS were enrolled. The experimental group received orally 4 g/day of fish oil for 12 months. Fish oil supplementation resulted in a significant increase in n-3 fatty acids and a decrease n-6 fatty acids. No differences in glutathione reductase activity, content of reduced and oxidized glutathione, and GSH/GSSG ratio were found. Conclusion: Glutathione reductase activity was not significantly different between the groups; however, fish oil supplementation resulted in smaller increase in GR compared with control group, suggesting a possible effect on antioxidant defence mechanisms. PMID:27335704

Glutathione and glutamate in schizophrenia: a 7T MRS study.

PubMed

Kumar, Jyothika; Liddle, Elizabeth B; Fernandes, Carolina C; Palaniyappan, Lena; Hall, Emma L; Robson, Siân E; Simmonite, Molly; Fiesal, Jan; Katshu, Mohammad Z; Qureshi, Ayaz; Skelton, Michael; Christodoulou, Nikolaos G; Brookes, Matthew J; Morris, Peter G; Liddle, Peter F

2018-06-22

In schizophrenia, abnormal neural metabolite concentrations may arise from cortical damage following neuroinflammatory processes implicated in acute episodes. Inflammation is associated with increased glutamate, whereas the antioxidant glutathione may protect against inflammation-induced oxidative stress. We hypothesized that patients with stable schizophrenia would exhibit a reduction in glutathione, glutamate, and/or glutamine in the cerebral cortex, consistent with a post-inflammatory response, and that this reduction would be most marked in patients with "residual schizophrenia", in whom an early stage with positive psychotic symptoms has progressed to a late stage characterized by long-term negative symptoms and impairments. We recruited 28 patients with stable schizophrenia and 45 healthy participants matched for age, gender, and parental socio-economic status. We measured glutathione, glutamate and glutamine concentrations in the anterior cingulate cortex (ACC), left insula, and visual cortex using 7T proton magnetic resonance spectroscopy (MRS). Glutathione and glutamate were significantly correlated in all three voxels. Glutamine concentrations across the three voxels were significantly correlated with each other. Principal components analysis (PCA) produced three clear components: an ACC glutathione-glutamate component; an insula-visual glutathione-glutamate component; and a glutamine component. Patients with stable schizophrenia had significantly lower scores on the ACC glutathione-glutamate component, an effect almost entirely leveraged by the sub-group of patients with residual schizophrenia. All three metabolite concentration values in the ACC were significantly reduced in this group. These findings are consistent with the hypothesis that excitotoxicity during the acute phase of illness leads to reduced glutathione and glutamate in the residual phase of the illness.

Glutathione protects Candida albicans against horseradish volatile oil.

PubMed

Bertóti, Regina; Vasas, Gábor; Gonda, Sándor; Nguyen, Nhat Minh; Szőke, Éva; Jakab, Ágnes; Pócsi, István; Emri, Tamás

2016-10-01

Horseradish essential oil (HREO; a natural mixture of different isothiocyanates) had strong fungicide effect against Candida albicans both in volatile and liquid phase. In liquid phase this antifungal effect was more significant than those of its main components allyl, and 2-phenylethyl isothiocyanate. HREO, at sublethal concentration, induced oxidative stress which was characterized with elevated superoxide content and up-regulated specific glutathione reductase, glutathione peroxidase, catalase and superoxide dismutase activities. Induction of specific glutathione S-transferase activities as marker of glutathione (GSH) dependent detoxification was also observed. At higher concentration, HREO depleted the GSH pool, increased heavily the superoxide production and killed the cells rapidly. HREO and the GSH pool depleting agent, 1-chlore-2,4-dinitrobenzene showed strong synergism when they were applied together to kill C. albicans cells. Based on all these, we assume that GSH metabolism protects fungi against isothiocyanates. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effect of glutathione aerosol on oxidant-antioxidant imbalance in idiopathic pulmonary fibrosis.

PubMed

Borok, Z; Buhl, R; Grimes, G J; Bokser, A D; Hubbard, R C; Holroyd, K J; Roum, J H; Czerski, D B; Cantin, A M; Crystal, R G

1991-07-27

Idiopathic pulmonary fibrosis (IPF) is characterised by alveolar inflammation, exaggerated release of oxidants, and subnormal concentrations of the antioxidant glutathione in respiratory epithelial lining fluid (ELF). Glutathione (600 mg twice daily for 3 days) was given by aerosol to 10 patients with IPF. Total ELF glutathione rose transiently, ELF oxidised glutathione concentrations increased, and there was a decrease in spontaneous superoxide anion release by alveolar macrophages. Thus, glutathione by aerosol could be a means of reversing the oxidant-antioxidant imbalance in IPF.

Glutathione S-transferase-encoding gene as a potential probe for environmental bacterial isolates capable of degrading polycyclic aromatic hydrocarbons.

PubMed Central

Lloyd-Jones, G; Lau, P C

1997-01-01

Homologs of the glutathione S-transferase (GST)-encoding gene were identified in a collection of aromatic hydrocarbon-degrading Sphingomonas spp. isolated from New Zealand, Antarctica, and the United States by using PCR primers designed from the GST-encoding gene of Sphingomonas paucimobilis EPA505. Sequence analysis of PCR fragments generated from these isolates and of the GST gene amplified from DNA extracted from polycyclic aromatic hydrocarbon (PAH)-contaminated soil revealed a high degree of conservation, which may make the GST-encoding gene a potentially useful marker for PAH-degrading bacteria. PMID:9251217

Metabolic cooperation of ascorbic acid and glutathione in normal and vitamin C-deficient ODS rats.

PubMed

Wang, Y; Kashiba, M; Kasahara, E; Tsuchiya, M; Sato, E F; Utsumi, K; Inoue, M

2001-01-01

Although the coordination of various antioxidants is important for the protection of organisms from oxidative stress, dynamic aspects of the interaction of endogenous antioxidants in vivo remain to be elucidated. We studied the metabolic coordination of two naturally occurring water-soluble antioxidants, ascorbic acid (AA) and reduced glutathione (GSH), in liver, kidney and plasma of control and scurvy-prone osteogenic disorder Shionogi (ODS) rats that hereditarily lack the ability to synthesize AA. When supplemented with AA, its levels in liver and kidney of ODS rats increased to similar levels of those in control rats. Hepato-renal levels of glutathione were similar with the two animal groups except for the slight increase in its hepatic levels in AA-supplemented ODS rats. Administration of L-buthionine sulfoximine (BSO), a specific inhibitor of GSH synthesis, rapidly decreased the hepato-renal levels of glutathione in a biphasic manner, a rapid phase followed by a slower phase. Kinetic analysis revealed that glutathione turnover was enhanced significantly in liver mitochondria and renal cytosol of ODS rats. Administration of BSO significantly increased AA levels in the liver and kidney of control rats but decreased them in AA-supplemented ODS rats. Kinetic analysis revealed that AA is synthesized by control rat liver by some BSO-enhanced mechanism and the de novo synthesized AA is transferred to the kidney. Such a coordination of the metabolism of GSH and AA in liver and kidney is suppressed in AA-deficient ODS rats. These and other results suggest that the metabolism of AA and GSH forms a compensatory network by which oxidative stress can be decreased.

Lack of direct effects of agrochemicals on zoonotic pathogens and fecal indicator bacteria.

PubMed

Staley, Zachery R; Senkbeil, Jacob K; Rohr, Jason R; Harwood, Valerie J

2012-11-01

Agrochemicals, fecal indicator bacteria (FIB), and pathogens frequently contaminate water simultaneously. No significant direct effects of fertilizer, atrazine, malathion, and chlorothalonil on the survival of Escherichia coli, Enterococcus faecalis, Salmonella enterica, human polyomaviruses, and adenovirus were detected, supporting the assertion that previously observed effects of agrochemicals on FIB were indirect.

ChaC2, an Enzyme for Slow Turnover of Cytosolic Glutathione*

PubMed Central

Kaur, Amandeep; Gautam, Ruchi; Srivastava, Ritika; Chandel, Avinash; Kumar, Akhilesh; Karthikeyan, Subramanian; Bachhawat, Anand Kumar

2017-01-01

Glutathione degradation plays an important role in glutathione and redox homeostasis, and thus it is imperative to understand the enzymes and the mechanisms involved in glutathione degradation in detail. We describe here ChaC2, a member of the ChaC family of γ-glutamylcyclotransferases, as an enzyme that degrades glutathione in the cytosol of mammalian cells. ChaC2 is distinct from the previously described ChaC1, to which ChaC2 shows ∼50% sequence identity. Human and mouse ChaC2 proteins purified in vitro show 10–20-fold lower catalytic efficiency than ChaC1, although they showed comparable Km values (Km of 3.7 ± 0.4 mm and kcat of 15.9 ± 1.0 min−1 toward glutathione for human ChaC2; Km of 2.2 ± 0.4 mm and kcat of 225.2 ± 15 min−1 toward glutathione for human ChaC1). The ChaC1 and ChaC2 proteins also shared the same specificity for reduced glutathione, with no activity against either γ-glutamyl amino acids or oxidized glutathione. The ChaC2 proteins were found to be expressed constitutively in cells, unlike the tightly regulated ChaC1. Moreover, lower eukaryotes have a single member of the ChaC family that appears to be orthologous to ChaC2. In addition, we determined the crystal structure of yeast ChaC2 homologue, GCG1, at 1.34 Å resolution, which represents the first structure of the ChaC family of proteins. The catalytic site is defined by a fortuitous benzoic acid molecule bound to the crystal structure. The mechanism for binding and catalytic activity of this new enzyme of glutathione degradation, which is involved in continuous but basal turnover of cytosolic glutathione, is proposed. PMID:27913623

Glutathione Metabolism and Parkinson’s Disease

PubMed Central

Smeyne, Michelle

2013-01-01

It has been established that oxidative stress, defined as the condition when the sum of free radicals in a cell exceeds the antioxidant capacity of the cell, contributes to the pathogenesis of Parkinson’s disease. Glutathione is a ubiquitous thiol tripeptide that acts alone, or in concert with enzymes within cells to reduce superoxide radicals, hydroxyl radicals and peroxynitrites. In this review, we examine the synthesis, metabolism and functional interactions of glutathione, and discuss how this relates to protection of dopaminergic neurons from oxidative damage and its therapeutic potential in Parkinson’s disease. PMID:23665395

The effects of glutathione depletion on thermotolerance and heat stress protein synthesis.

PubMed Central

Russo, A.; Mitchell, J. B.; McPherson, S.

1984-01-01

The effects of cellular glutathione depletion by buthionine sulfoximine on the development of thermotolerance and synthesis of heat stress protein was studied. Cellular glutathione levels were found to increase rapidly following an acute heat treatment of either 12 min at 45.5 degrees C or 1 h at 43 degrees C and remain elevated for prolonged periods. Glutathione depletion and prevention of glutathione synthesis by buthionine sulfoximine resulted in inhibition of the development of thermotolerance and a decrease in total protein as well as specific heat stress proteins. While the degree of inhibition of thermotolerance was similar for both glutathione depletion protocols, inhibition in heat stress protein synthesis was greater when glutathione was depleted to low levels prior to heating. The possible role of glutathione and the cellular redox state to thermotolerance and synthesis of heat stress protein is discussed. Images Figure 2 PMID:6733022

Lack of Direct Effects of Agrochemicals on Zoonotic Pathogens and Fecal Indicator Bacteria

PubMed Central

Staley, Zachery R.; Senkbeil, Jacob K.; Rohr, Jason R.

2012-01-01

Agrochemicals, fecal indicator bacteria (FIB), and pathogens frequently contaminate water simultaneously. No significant direct effects of fertilizer, atrazine, malathion, and chlorothalonil on the survival of Escherichia coli, Enterococcus faecalis, Salmonella enterica, human polyomaviruses, and adenovirus were detected, supporting the assertion that previously observed effects of agrochemicals on FIB were indirect. PMID:22961900

Glutathione metabolic status in the aged rabbit aorta.

PubMed

Lapenna, Domenico; Ciofani, Giuliano; Giamberardino, Maria Adele

2017-05-01

It is not known whether aging alters glutathione metabolic status of the mammalian arterial tissue favoring vascular oxidative stress and dysfunction. Thus we assessed total, reduced and oxidized glutathione (TG, GSH and GSSG, respectively), the glutathione redox ratio (GRR, namely [GSSG]/[GSH+2GSSG]×100), and the activities of the glutathione status-regulating enzymes glutathione reductase (GSSG-Red), γ-glutamylcysteine synthetase (γ-GCS) and γ-glutamyl transpeptidase (γ-GT) in the aortic tissue of 9 young adult control rabbits (YACR, about 4months old) and 9 aged rabbits (AR, about 4.5years old); aortic lipid and protein oxidation and H 2 O 2 were also determined as oxidative stress indicators. Vascular function was assessed on aortic ring preparations. TG and GSH concentrations, together with γ-GCS and γ-GT activities, were significantly lower, while GSSG content and the GRR higher, in the AR than in the YACR aortas; GSSG-Red activity did not differ significantly between the two groups. Heightened levels of lipid and protein oxidation and H 2 O 2 occurred in the AR aortas, indicating age-dependent vascular oxidative stress. Moreover, in the whole population of 18 rabbits, the aortic values of GSH and related enzyme activities were inversely and significantly correlated with those of lipid and protein oxidation and H 2 O 2 , highlighting the antioxidant role of GSH and related enzymes in the vascular tissue. Aortic endothelium-dependent vasodilation was lower in the AR than in the YACR. In conclusion, glutathione metabolic status is altered in the aged rabbit aorta reflecting depressed γ-GCS- and γ-GT-related GSH biosynthesis and GSSG burden eventually favoring vascular oxidative stress and dysfunction. Copyright © 2017 Elsevier Inc. All rights reserved.

Amodiaquine failure associated with erythrocytic glutathione in Plasmodium falciparum malaria

PubMed Central

Zuluaga, Lina; Pabón, Adriana; López, Carlos; Ochoa, Aleida; Blair, Silvia

2007-01-01

Objective To establish the relationship between production of glutathione and the therapeutic response to amodiaquine (AQ) monotherapy in Plasmodium falciparum non-complicated malaria patients. Methodology Therapeutic response to AQ was evaluated in 32 patients with falciparum malaria in two townships of Antioquia, Colombia, and followed-up for 28 days. For every patient, total glutathione and enzymatic activity (glutathione reductase, GR, and γ-glutamylcysteine synthetase, γ-GCS) were determined in parasitized erythrocytes, non-infected erythrocytes and free parasites, on the starting day (day zero, before ingestion of AQ) and on the day of failure (in case of occurrence). Results There was found an AQ failure of 31.25%. Independent of the therapeutic response, on the starting day and on the day of failure, lower total glutathione concentration and higher GR activities in parasitized erythrocytes were found, compared with non-infected erythrocytes (p < 0.003). In addition, only on the day of failure, γ-GCS activity of parasitized erythrocytes was higher, compared with that of healthy erythrocytes (p = 0.01). Parasitized and non-parasitized erythrocytes in therapeutic failure patients (TF) had higher total glutathione on the starting day compared with those of adequate clinical response (ACR) (p < 0.02). Parasitized erythrocytes of TF patients showed lower total glutathione on the failure day, compared with starting day (p = 0.017). No differences was seen in the GR and γ-GCS activities by compartment, neither between the two therapeutic response groups nor between the two treatment days. Conclusion This study is a first approach to explaining P. falciparum therapeutic failure in humans through differences in glutathione metabolism in TF and ACR patients. These results suggest a role for glutathione in the therapeutic failure to antimalarials. PMID:17451604

ChaC2, an Enzyme for Slow Turnover of Cytosolic Glutathione.

PubMed

Kaur, Amandeep; Gautam, Ruchi; Srivastava, Ritika; Chandel, Avinash; Kumar, Akhilesh; Karthikeyan, Subramanian; Bachhawat, Anand Kumar

2017-01-13

Glutathione degradation plays an important role in glutathione and redox homeostasis, and thus it is imperative to understand the enzymes and the mechanisms involved in glutathione degradation in detail. We describe here ChaC2, a member of the ChaC family of γ-glutamylcyclotransferases, as an enzyme that degrades glutathione in the cytosol of mammalian cells. ChaC2 is distinct from the previously described ChaC1, to which ChaC2 shows ∼50% sequence identity. Human and mouse ChaC2 proteins purified in vitro show 10-20-fold lower catalytic efficiency than ChaC1, although they showed comparable K m values (K m of 3.7 ± 0.4 mm and k cat of 15.9 ± 1.0 min -1 toward glutathione for human ChaC2; K m of 2.2 ± 0.4 mm and k cat of 225.2 ± 15 min -1 toward glutathione for human ChaC1). The ChaC1 and ChaC2 proteins also shared the same specificity for reduced glutathione, with no activity against either γ-glutamyl amino acids or oxidized glutathione. The ChaC2 proteins were found to be expressed constitutively in cells, unlike the tightly regulated ChaC1. Moreover, lower eukaryotes have a single member of the ChaC family that appears to be orthologous to ChaC2. In addition, we determined the crystal structure of yeast ChaC2 homologue, GCG1, at 1.34 Å resolution, which represents the first structure of the ChaC family of proteins. The catalytic site is defined by a fortuitous benzoic acid molecule bound to the crystal structure. The mechanism for binding and catalytic activity of this new enzyme of glutathione degradation, which is involved in continuous but basal turnover of cytosolic glutathione, is proposed. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Glutathione suppresses the enzymatic and non-enzymatic browning in grape juice.

PubMed

Wu, Shengjun

2014-10-01

Browning tends to occur in grape juice during processing and storage and decreases the commercial value of it. Thus, browning inhibition is an important objective for manufacturers. This study aims to investigate the efficacy of glutathione as a browning inhibitor for use on grape juice. Grape juice browning treated with glutathione was monitored during processing and accelerated browning. 0.04% of glutathione inhibited 99.4% of the polyphenoloxidase activity in the grape juice. Consequently, during processing at room temperature and accelerated browning at 80 °C, the browning in the grape juice treated with glutathione was significantly lower than that in the control (p<0.05). The results indicate that glutathione is a promising browning inhibitor used in grape juice. Copyright © 2014 Elsevier Ltd. All rights reserved.

Multiscale modelling approach combining a kinetic model of glutathione metabolism with PBPK models of paracetamol and the potential glutathione-depletion biomarkers ophthalmic acid and 5-oxoproline in humans and rats.

PubMed

Geenen, Suzanne; Yates, James W T; Kenna, J Gerry; Bois, Frederic Y; Wilson, Ian D; Westerhoff, Hans V

2013-06-01

A key role of the antioxidant glutathione is detoxification of chemically reactive electrophilic drug metabolites within the liver. Therefore glutathione depletion can have severe toxic consequences. Ophthalmic acid and 5-oxoproline are metabolites involved in glutathione metabolism, which can be measured readily in the blood and urine and have been proposed as candidate biomarkers of hepatic glutathione content. However, currently it is unclear whether their concentrations in plasma exhibit a robust correlation with hepatic glutathione content. To explore this important question, we have developed a novel approach which combines a physiologically based pharmacokinetic (PBPK) model of metabolism and disposition of paracetamol (acetaminophen) with a previously developed mathematical systems model of hepatic glutathione homeostasis. Paracetamol is metabolised to reactive intermediates which deplete glutathione and cause toxicity when given at high doses. Our model correctly predicted that hepatic glutathione depletion following paracetamol administration resulted in elevated concentrations of 5-oxoproline and ophthalmic acid in blood and of 5-oxoproline in urine. However, we also found from the model that concentrations of both of the compounds were likely to be influenced by prolonged administration of paracetamol and by the concentrations of intracellular metabolites such as methionine. We conclude that care must be taken when extrapolating from concentrations of these biomarkers to hepatic glutathione status.

Feedback inhibition by thiols outranks glutathione depletion: a luciferase-based screen reveals glutathione-deficient γ -ECS and glutathione synthetase mutants impaired in cadmium-induced sulfate assimilation

PubMed Central

Jobe, Timothy O.; Sung, Dong-Yul; Akmakjian, Garo; Pham, Allis; Komives, Elizabeth A.; Mendoza-Cózatl, David G.; Schroeder, Julian I.

2015-01-01

Summary Plants exposed to heavy metals rapidly induce changes in gene expression that activate and enhance detoxification mechanisms, including toxic-metal chelation and the scavenging of reactive oxygen species. However, the mechanisms mediating toxic heavy metal-induced gene expression remain largely unknown. To genetically elucidate cadmium-specific transcriptional responses in Arabidopsis, we designed a genetic screen based on the activation of a cadmium-inducible reporter gene. Microarray studies identified a high-affinity sulfate transporter (SULTR1;2) among the most robust and rapid cadmium-inducible transcripts. The SULTR1;2 promoter (2.2 kb) was fused with the firefly luciferase reporter gene to quantitatively report the transcriptional response of plants exposed to cadmium. Stably transformed luciferase reporter lines were ethyl methanesulfonate (EMS) mutagenized, and stable M2 seedlings were screened for an abnormal luciferase response during exposure to cadmium. The screen identified non-allelic mutant lines that fell into one of three categories: (i) super response to cadmium (SRC) mutants; (ii) constitutive response to cadmium (CRC) mutants; or (iii) non-response and reduced response to cadmium (NRC) mutants. Two nrc mutants, nrc1 and nrc2, were mapped, cloned and further characterized. The nrc1 mutation was mapped to the γ-glutamylcysteine synthetase gene and the nrc2 mutation was identified as the first viable recessive mutant allele in the glutathione synthetase gene. Moreover, genetic, HPLC mass spectrometry, and gene expression analysis of the nrc1 and nrc2 mutants, revealed that intracellular glutathione depletion alone would be insufficient to induce gene expression of sulfate uptake and assimilation mechanisms. Our results modify the glutathione-depletion driven model for sulfate assimilation gene induction during cadmium stress, and suggest that an enhanced oxidative state and depletion of upstream thiols, in addition to glutathione

Extraction of glutathione from EFB fermentation waste using methanol with sonication process

NASA Astrophysics Data System (ADS)

Muryanto, Muryanto; Alvin, Nurdin, Muhammad; Hanifah, Ummu; Sudiyani, Yanni

2017-11-01

Glutathione is important compound on the human body. Glutathione have a widely use at pharmacy and cosmetics as detoxification, skin whitening agent, antioxidant and many other. This study aims to obtain glutathione from Saccharomyces cerevisiae in fermentation waste of second generation bioethanol. The remaining yeast in the empty fruit bunch (EFB) fermentation was separated from the fermentation solution use centrifugation process and then extracted using a methanol-water solution. The extraction process was done by maceration which was assisted by sonication process. Solvent concentration and time of sonication were varied to see its effect on glutathione concentration. The concentration of glutathione from the extraction process was analyzed using alloxan method with UV-Vis spectrophotometer. The results show that the highest glutathione concentration was approximately 1.32 g/L obtained with methanol solvent at 90 minutes of maceration following with 15 minutes sonication.

Endophytic bacterium Sphingomonas SaMR12 promotes cadmium accumulation by increasing glutathione biosynthesis in Sedum alfredii Hance.

PubMed

Pan, Fengshan; Meng, Qian; Wang, Qiong; Luo, Sha; Chen, Bao; Khan, Kiran Yasmin; Yang, Xiaoe; Feng, Ying

2016-07-01

A hydroponic experiment was conducted to verify the effects of inoculation with endophytic bacteria Sphingomonas SaMR12 on root growth, cadmium (Cd) uptake, reactive oxygen species (ROS), antioxidases, glutathione (GSH) and the related gene expression of Sedum alfredii Hance under different levels of Cd such as 0, 10, 25, 100 and 400 μM. The results showed that inoculation of SaMR12 improved Cd accumulation and upregulated glutathione synthase (GS) expression, but slightly reduced malondialdehyde (MDA) concentration and alleviated Cd-induced damage in roots. However it didn't alter the activities of antioxidant enzymes. When Cd concentration exceeded 25 μM, SaMR12 increased the concentration of GSH and the expression level of GSH1. At high Cd treatment levels (100 and 400 μM), SaMR12 significantly reduced H2O2 concentration and enhanced expression level of 1-Cys peroxiredoxin PER1 and ATPS genes. These results indicate that although SaMR12 has no significant effects on antioxidases activities, it reduces H2O2 concentration by enhancing GSH concentration and relevant genes expression, and subsequently improves Cd tolerance and accumulation. Copyright © 2016 Elsevier Ltd. All rights reserved.

Glutathione metabolism as a determinant of therapeutic efficacy: a review.

PubMed

Arrick, B A; Nathan, C F

1984-10-01

Glutathione, as the chief nonprotein intracellular sulfhydryl, affects the efficacy and interactions of a variety of antineoplastic interventions, mainly through nucleophilic thioether formation or oxidation-reduction reactions. Thus, glutathione plays a role in the detoxification and repair of cellular injury by such diverse agents as mechlorethamine, melphalan, cyclophosphamide, nitrosoureas, 6-thiopurine, 4'-(9-acridinylamino)methanesulfon-m-anisidide, the quinone antibiotics (including Adriamycin, daunorubicin, and mitomycin C), the sesquiterpene lactones (such as vernolepin), and other sulfhydryl-reactive diterpenes (like jatrophone). Glutathione may play a similar role in host and tumor cell responses to radiation, hyperthermia, and the reactive reduction products of oxygen secreted by inflammatory cells. Further, glutathione participates in the formation of toxic metabolites of such chemotherapeutics as azathioprine and bleomycin and may affect the cellular uptake of other agents, such as methotrexate. It seems likely that alterations in glutathione metabolism of tumor or host as a result of one therapeutic intervention may affect the outcome of concurrent treatments. Knowledge of these interactions may be useful in designing combination therapy for neoplastic disease.

Expression of glutathione peroxidase I gene in selenium-deficient rats.

PubMed Central

Reddy, A P; Hsu, B L; Reddy, P S; Li, N Q; Thyagaraju, K; Reddy, C C; Tam, M F; Tu, C P

1988-01-01

We have characterized a cDNA pGPX1211 encoding rat glutathione peroxidase I. The selenocysteine in the protein corresponded to a TGA codon in the coding region of the cDNA, similar to earlier findings in mouse and human genes, and a gene encoding the formate dehydrogenase from E. coli, another selenoenzyme. The rat GSH peroxidase I has a calculated subunit molecular weight of 22,155 daltons and shares 95% and 86% sequence homology with the mouse and human subunits, respectively. The 3'-noncoding sequence (greater than 930 bp) in pGPX1211 is much longer than that of the human sequences. We found that glutathione peroxidase I mRNA, but not the polypeptide, was expressed under nutritional stress of selenium deficiency where no glutathione peroxidase I activity can be detected. The failure of detecting any apoprotein for the glutathione peroxidase I under selenium deficiency and results published from other laboratories supports the proposal that selenium may be incorporated into the glutathione peroxidase I co-translationally. Images PMID:2838821

Effect of glutathione during bottle storage of sparkling wine.

PubMed

Webber, Vanessa; Dutra, Sandra Valduga; Spinelli, Fernanda Rodrigues; Carnieli, Gilberto João; Cardozo, Alejandro; Vanderlinde, Regina

2017-02-01

Reduced glutathione (GSH) is an efficient antioxidant on limiting browning, losing varietal aromas and off-flavor formation. Therefore, this study aims to evaluate the effect of GSH addition (10, 20 and 30mgL(-1)) after the disgorging of the sparkling wine during storage. The sparkling wines were analyzed at 1, 6, 12 and 18months of storage according to the color index, concentration of the free SO2, phenolic compounds, catechin, epicatechin, caffeic acid, coumaric acid, acetaldehyde, total and reduced glutathione. The results show that GSH concentration decreased to the level of the control sparkling wine during the first 6months, and the total glutathione gradually declined up to 12months. The GSH reduces browning and acetaldehyde formation for up to 12months. However, the presence of glutathione had low or no influence on the concentration of free SO2, total phenolics, catechin, epicatechin, caffeic and coumaric acids. Copyright © 2016 Elsevier Ltd. All rights reserved.

Inactivation of Bacteria in Oil Field Injected Water by a Pulsed Plasma Discharge Process

NASA Astrophysics Data System (ADS)

Xin, Qing; Li, Zhongjian; Lei, Lecheng; Yang, Bin

2016-09-01

Pulsed plasma discharge was employed to inactivate bacteria in the injection water for an oil field. The effects of water conductivity and initial concentration of bacteria on elimination efficiency were investigated in the batch and continuous flow modes. It was demonstrated that Fe2+ contained in injection water could enhance the elimination efficiency greatly. The addition of reducing agent glutathione (GSH) indicated that active radicals generated by pulsed plasma discharges played an important role in the inactivation of bacteria. Moreover, it was found that the microbial inactivation process for both batch and continuous flow mode well fitted the model based on the Weibull's survival function. supported by Zhejiang Province Welfare Technology Applied Research Project of China (No. 2014C31137), National Natural Science Foundation of China (Nos. 21436007 and U1462201), and the Fundamental Research Funds for the Central Universities of China (No. 2015QNA4032)

Polymorphisms of glutathione S-transferase Mu 1, glutathione S-transferase theta 1 and glutathione S-transferase Pi 1 genes in Hodgkin's lymphoma susceptibility and progression.

PubMed

Lourenço, Gustavo J; Néri, Iramaia A; Sforni, Vitor C S; Kameo, Rodolfo; Lorand-Metze, Irene; Lima, Carmen S P

2009-06-01

We tested in this study whether the polymorphisms of the glutathione S-transferase Mu1 (GSTM1), glutathione S-transferase Theta 1 (GSTT1) and glutathione S-transferase Pi 1 (GSTP1), involved in metabolism of chemical agents, cell proliferation and cell survival, alter the risk for Hodgkin lymphoma (HL). Genomic DNA from 110 consecutive patients with HL and 226 controls was analysed by polymerase chain reaction and restriction digestion for the polymorphism analyses. Similar frequencies of the GSTM1 and GSTT1 genotypes were seen in patients and controls. In contrast, the frequency of the GSTP1 wild genotype (59.1%versus 36.3%, P = 0.004) was higher in patients than in controls. Individuals with the wild genotype had a 2.68 (95%CI: 1.38-5.21)-fold increased risk for the disease than others. An excess of the GSTP1 wild genotype was also observed in patients with tumors of stages III + IV when compared with those with tumors of stages I + II (39.1%versus 20.0%, P = 0.03). These results suggest that the wild allele of the GSTP1 gene is linked to an increased risk and high aggressiveness of the HL in our cases but they should be confirmed by further studies with larger cohorts of patients and controls.

Photoaffinity labelling of the active site of the rat glutathione transferases 3-3 and 1-1 and human glutathione transferase A1-1.

PubMed

Cooke, R J; Björnestedt, R; Douglas, K T; McKie, J H; King, M D; Coles, B; Ketterer, B; Mannervik, B

1994-09-01

The glutathione transferases (GSTs) form a group of enzymes responsible for a wide range of molecular detoxications. The photoaffinity label S-(2-nitro-4-azidophenyl)glutathione was used to study the hydrophobic region of the active site of the rat liver GST 1-1 and 2-2 isoenzymes (class Alpha) as well as the rat class-Mu GST 3-3. Photoaffinity labelling was carried out using a version of S-(2-nitro-4-azidophenyl)glutathione tritiated in the arylazido ring. The labelling occurred with higher levels of radioisotope incorporation for the Mu than the Alpha families. Taking rat GST 3-3, 1.18 (+/- 0.05) mol of radiolabel from S-(2-nitro-4-azidophenyl)glutathione was incorporated per mol of dimeric enzyme, which could be blocked by the presence of the strong competitive inhibitor, S-tritylglutathione (Ki = 1.4 x 10(-7) M). Radiolabelling of the protein paralleled the loss of enzyme activity. Photoaffinity labelling by tritiated S-(2-nitro-4-azidophenyl)glutathione on a preparative scale (in the presence and absence of S-tritylglutathione) followed by tryptic digestion and purification of the labelled peptides indicated that GST 3-3 was specifically photolabelled; the labelled peptides were sequenced. Similarly, preparative photoaffinity labelling by S-(2-nitro-4-azidophenyl)glutathione of the rat liver 1-1 isoenzyme, the human GST A1-1 and the human-rat chimaeric GST, H1R1/1, was carried out with subsequent sequencing of radiolabelled h.p.l.c.-purified tryptic peptides. The results were interpreted by means of molecular-graphics analysis to locate photoaffinity-labelled peptides using the X-ray-crystallographic co-ordinates of rat GST 3-3 and human GST A1-1. The molecular-graphical analysis indicated that the labelled peptides are located within the immediate vicinity of the region occupied by S-substituted glutathione derivatives bound in the active-site cavity of the GSTs investigated.

Reactions of benzene oxide with thiols including glutathione.

PubMed

Henderson, Alistair P; Barnes, Martine L; Bleasdale, Christine; Cameron, Richard; Clegg, William; Heath, Sarah L; Lindstrom, Andrew B; Rappaport, Stephen M; Waidyanatha, Suramya; Watson, William P; Golding, Bernard T

2005-02-01

S-Phenylmercapturic acid is a minor metabolite of benzene used as a biomarker for human benzene exposures. The reaction of intracellular glutathione with benzene oxide-oxepin, the initial metabolite of benzene, is presumed to give 1-(S-glutathionyl)-cyclohexa-3,5-dien-2-ol, which undergoes dehydration to S-phenylglutathione, the precursor of S-phenylmercapturic acid. To validate the proposed route to S-phenylglutathione, reactions of benzene oxide-oxepin with glutathione and other sulfur nucleophiles have been studied. The reaction of benzene oxide with an excess of aqueous sodium sulfide, followed by acetylation, gave bis-(6-trans-5-acetoxycyclohexa-1,3-dienyl)sulfide, the structure of which was proved by X-ray crystallography. Reactions of benzene oxide-oxepin in a 95:5 (v/v) mixture of phosphate buffer in D2O with (CD3)2SO were monitored by 1H NMR spectroscopy. In the absence of glutathione, the half-life of benzene oxide-oxepin was ca. 34 min at 25 degrees C and pD 7.0. The half-life was not affected in the range of 2-15 mM glutathione in the presence and absence of a commercial sample of human glutathione S-transferase (at pH 7.0, 8.0, 8.5, or 10.0). The adduct 1-(S-glutathionyl)-cyclohexa-3,5-diene-2-ol was identified in these reaction mixtures, especially at higher pH, by mass spectrometry and by its acid-catalyzed decomposition to S-phenylglutathione. Incubation of benzene oxide with N-acetyl-L-cysteine at 37 degrees C and pH 10.0 and subsequent mass spectrometric analysis of the mixture showed formation of pre-S-phenylmercapturic acid and the dehydration product, S-phenylmercapturic acid. The data validate the premise that benzene oxide-oxepin can be captured by glutathione to give (1R,2R)- and/or (1S,2S)-1-(S-glutathionyl)-cyclohexa-3,5-dien-2-ol, which dehydrate to S-phenylglutathione. The capture is a relatively inefficient process at pH 7 that is accelerated at higher pH. These studies account for the observation that the metabolism of benzene is

Glutathione redox dynamics and expression of glutathione-related genes in the developing embryo

PubMed Central

Timme-Laragy, Alicia R.; Goldstone, Jared V.; Imhoff, Barry R.; Stegeman, John J.; Hahn, Mark E.; Hansen, Jason M.

2013-01-01

Embryonic development involves dramatic changes in cell proliferation and differentiation that must be highly coordinated and tightly regulated. Cellular redox balance is critical for cell fate decisions, but it is susceptible to disruption by endogenous and exogenous sources of oxidative stress. The most abundant endogenous non-protein antioxidant defense molecule is the tri-peptide glutathione (γ-glutamyl-cysteinylglycine, GSH), but the ontogeny of GSH concentration and redox state during early life stages is poorly understood. Here, we describe the GSH redox dynamics during embryonic and early larval development (0–5 days post-fertilization) in the zebrafish (Danio rerio), a model vertebrate embryo. We measured reduced and oxidized glutathione (GSH, GSSG) using HPLC, and calculated the whole embryo total glutathione (GSHT) concentrations and redox potentials (Eh) over 0–120 hours of zebrafish development (including mature oocytes, fertilization, mid-blastula transition, gastrulation, somitogenesis, pharyngula, pre-hatch embryos, and hatched eleutheroembryos). GSHT concentration doubled between 12 hours post fertilization (hpf) and hatching. The GSH Eh increased, becoming more oxidizing during the first 12 h, and then oscillated around −190 mV through organogenesis, followed by a rapid change, associated with hatching, to a more negative (more reducing) Eh (−220 mV). After hatching, Eh stabilized and remained steady through 120 hpf. The dynamic changes in GSH redox status and concentration defined discrete windows of development: primary organogenesis, organ differentiation, and larval growth. We identified the set of zebrafish genes involved in the synthesis, utilization, and recycling of GSH, including several novel paralogs, and measured how expression of these genes changes during development. Ontogenic changes in the expression of GSH-related genes support the hypothesis that GSH redox state is tightly regulated early in development. This study

Feedback inhibition by thiols outranks glutathione depletion: a luciferase-based screen reveals glutathione-deficient γ-ECS and glutathione synthetase mutants impaired in cadmium-induced sulfate assimilation.

PubMed

Jobe, Timothy O; Sung, Dong-Yul; Akmakjian, Garo; Pham, Allis; Komives, Elizabeth A; Mendoza-Cózatl, David G; Schroeder, Julian I

2012-06-01

Plants exposed to heavy metals rapidly induce changes in gene expression that activate and enhance detoxification mechanisms, including toxic-metal chelation and the scavenging of reactive oxygen species. However, the mechanisms mediating toxic heavy metal-induced gene expression remain largely unknown. To genetically elucidate cadmium-specific transcriptional responses in Arabidopsis, we designed a genetic screen based on the activation of a cadmium-inducible reporter gene. Microarray studies identified a high-affinity sulfate transporter (SULTR1;2) among the most robust and rapid cadmium-inducible transcripts. The SULTR1;2 promoter (2.2 kb) was fused with the firefly luciferase reporter gene to quantitatively report the transcriptional response of plants exposed to cadmium. Stably transformed luciferase reporter lines were ethyl methanesulfonate (EMS) mutagenized, and stable M(2) seedlings were screened for an abnormal luciferase response during exposure to cadmium. The screen identified non-allelic mutant lines that fell into one of three categories: (i) super response to cadmium (SRC) mutants; (ii) constitutive response to cadmium (CRC) mutants; or (iii) non-response and reduced response to cadmium (NRC) mutants. Two nrc mutants, nrc1 and nrc2, were mapped, cloned and further characterized. The nrc1 mutation was mapped to the γ-glutamylcysteine synthetase gene and the nrc2 mutation was identified as the first viable recessive mutant allele in the glutathione synthetase gene. Moreover, genetic, HPLC mass spectrometry, and gene expression analysis of the nrc1 and nrc2 mutants, revealed that intracellular glutathione depletion alone would be insufficient to induce gene expression of sulfate uptake and assimilation mechanisms. Our results modify the glutathione-depletion driven model for sulfate assimilation gene induction during cadmium stress, and suggest that an enhanced oxidative state and depletion of upstream thiols, in addition to glutathione depletion

A Western diet induced NAFLD in LDLR(-/)(-) mice is associated with reduced hepatic glutathione synthesis.

PubMed

Li, Ling; Zhang, Guo-Fang; Lee, Kwangwon; Lopez, Rocio; Previs, Stephen F; Willard, Belinda; McCullough, Arthur; Kasumov, Takhar

2016-07-01

Oxidative stress plays a key role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). Glutathione is the major anti-oxidant involved in cellular oxidative defense, however there are currently no simple non-invasive methods for assessing hepatic glutathione metabolism in patients with NAFLD. As a primary source of plasma glutathione, liver plays an important role in interorgan glutathione homeostasis. In this study, we have tested the hypothesis that measurements of plasma glutathione turnover could be used to assess the hepatic glutathione metabolism in LDLR(-/)(-) mice, a mouse model of diet-induced NAFLD. Mice were fed a standard low fat diet (LFD) or a high fat diet containing cholesterol (a Western type diet (WD)). The kinetics of hepatic and plasma glutathione were quantified using the (2)H2O metabolic labeling approach. Our results show that a WD leads to reduced fractional synthesis rates (FSR) of hepatic (25%/h in LFD vs. 18%/h in WD, P<0.05) and plasma glutathione (43%/h in LFD vs. 21%/h in WD, P<0.05), without any significant effect on their absolute production rates (PRs). WD-induced concordant changes in both hepatic and plasma glutathione turnover suggest that the plasma glutathione turnover measurements could be used to assess hepatic glutathione metabolism. The safety, simplicity, and low cost of the (2)H2O-based glutathione turnover approach suggest that this method has the potential for non-invasive probing of hepatic glutathione metabolism in patients with NAFLD and other diseases. Copyright © 2016 Elsevier Inc. All rights reserved.

Neutrophil-generated HOCl leads to non-specific thiol oxidation in phagocytized bacteria

PubMed Central

Degrossoli, Adriana; Müller, Alexandra; Xie, Kaibo; Schneider, Jannis F; Bader, Verian; Winklhofer, Konstanze F; Meyer, Andreas J

2018-01-01

Phagocytic immune cells kill pathogens in the phagolysosomal compartment with a cocktail of antimicrobial agents. Chief among them are reactive species produced in the so-called oxidative burst. Here, we show that bacteria exposed to a neutrophil-like cell line experience a rapid and massive oxidation of cytosolic thiols. Using roGFP2-based fusion probes, we could show that this massive breakdown of the thiol redox homeostasis was dependent on phagocytosis, presence of NADPH oxidase and ultimately myeloperoxidase. Interestingly, the redox-mediated fluorescence change in bacteria expressing a glutathione-specific Grx1-roGFP2 fusion protein or an unfused roGFP2 showed highly similar reaction kinetics to the ones observed with roGFP2-Orp1, under all conditions tested. We recently observed such an indiscriminate oxidation of roGFP2-based fusion probes by HOCl with fast kinetics in vitro. In line with these observations, abating HOCl production in immune cells with a myeloperoxidase inhibitor significantly attenuated the oxidation of all three probes in bacteria. PMID:29506649

The dyad palindromic glutathione transferase P enhancer binds multiple factors including AP1.

PubMed Central

Diccianni, M B; Imagawa, M; Muramatsu, M

1992-01-01

Glutathione Transferase P (GST-P) gene expression is dominantly regulated by an upstream enhancer (GPEI) consisting of a dyad of palindromically oriented imperfect TPA (12-O-tetradecanoyl-phorbol-13-acetate)-responsive elements (TRE). GPEI is active in AP1-lacking F9 cells as well in AP1-containing HeLa cells. Despite GPEI's similarity to a TRE, c-jun co-transfection has only a minimal effect on transactivation. Antisense c-jun and c-fos co-transfection experiments further demonstrate the lack of a role for AP1 in GPEI mediated trans-activation in F9 cells, although endogenously present AP1 can influence GPEI in HeLa cells. Co-transfection of delta fosB with c-jun, which forms an inactive c-Jun/delta FosB heterodimer that binds TRE sequences, inhibits GPEI-mediated transcription in AP1-lacking F9 cells as well as AP1-containing HeLa cells. These data suggest novel factor(s) other than AP1 are influencing GPEI. Binding studies reveal multiple nucleoproteins bind to GPEI. These factors are likely responsible for the high level of GPEI-mediated transcription observed in the absence of AP1 and during hepatocarcinogenesis. Images PMID:1408831

The dyad palindromic glutathione transferase P enhancer binds multiple factors including AP1.

PubMed

Diccianni, M B; Imagawa, M; Muramatsu, M

1992-10-11

Glutathione Transferase P (GST-P) gene expression is dominantly regulated by an upstream enhancer (GPEI) consisting of a dyad of palindromically oriented imperfect TPA (12-O-tetradecanoyl-phorbol-13-acetate)-responsive elements (TRE). GPEI is active in AP1-lacking F9 cells as well in AP1-containing HeLa cells. Despite GPEI's similarity to a TRE, c-jun co-transfection has only a minimal effect on transactivation. Antisense c-jun and c-fos co-transfection experiments further demonstrate the lack of a role for AP1 in GPEI mediated trans-activation in F9 cells, although endogenously present AP1 can influence GPEI in HeLa cells. Co-transfection of delta fosB with c-jun, which forms an inactive c-Jun/delta FosB heterodimer that binds TRE sequences, inhibits GPEI-mediated transcription in AP1-lacking F9 cells as well as AP1-containing HeLa cells. These data suggest novel factor(s) other than AP1 are influencing GPEI. Binding studies reveal multiple nucleoproteins bind to GPEI. These factors are likely responsible for the high level of GPEI-mediated transcription observed in the absence of AP1 and during hepatocarcinogenesis.

Effect of natural ageing and antioxidant inhibition on liver antioxidant enzymes, glutathione system, peroxidation, and oxygen consumption in Rana perezi.

PubMed

López-Torres, M; Pérez-Campo, R; Barja de Quiroga, G

1991-01-01

A study of the physiological role of oxygen free radicals in relation to the ageing process was performed using the liver of Rana perezi, an animal with a moderate rate of oxygen consumption and a life span substantially longer than that of laboratory rodents. Among the five different antioxidant enzymes only superoxide dismutase (SOD) showed an age-dependent decrease. Cytochrome oxidase (COX), glutathione status, in vivo and in vitro liver peroxidation, and metabolic rate did not vary as a function of age. Long-term (2.5 months) treatment with aminotriazole and diethyldithiocarbamate depleted catalase (CAT) activity and did not change both glutathione peroxidases (GPx), COX, reduced (GSH) and oxidized (GSSG) glutathione, or metabolic rate. This treatment resulted in great compensatory increases in SOD (to 250-460% of controls) and glutathione reductase (GR) (to 200%) which are possibly responsible for the lack of increase of in vivo and in vitro liver peroxidation and for the absence of changes in survival rate. The comparison of these results with previous data from other species suggests the possibility that decreases in antioxidant capacity in old age are restricted to animal species with high metabolic rates. Nevertheless, ageing can still be due to the continuous presence of small concentrations of O2 radicals in the tissues throughout life in animals with either high or low metabolic rates, because radical scavenging can not be 100% effective. Compensatory homeostasis among antioxidants seems to be a general phenomenon in different species.

Characterization of a Novel Dithiocarbamate Glutathione Reductase Inhibitor and Its Use as a Tool to Modulate Intracellular Glutathione*

PubMed Central

Seefeldt, Teresa; Zhao, Yong; Chen, Wei; Raza, Ashraf S.; Carlson, Laura; Herman, Jocqueline; Stoebner, Adam; Hanson, Sarah; Foll, Ryan; Guan, Xiangming

2009-01-01

Thiol redox state (TRS) is an important parameter to reflect intracellular oxidative stress and is associated with various normal and abnormal biochemical processes. Agents that can be used to increase intracellular TRS will be valuable tools in TRS-related research. Glutathione reductase (GR) is a critical enzyme in the homeostasis of TRS. The enzyme catalyzes the reduction of GSSG to GSH to maintain a high GSH:GSSG ratio. Inhibition of the enzyme can be used to increase TRS. Despite the reports of various GR inhibitors, N,N-bis(2-chloroethyl)-N-nitrosourea, an anticancer drug with IC50 = 647 μm against yeast GR, remains the most commonly used GR inhibitor in the literature. However, the toxicity caused by nonspecific interactions, as well as inhibition of DNA synthesis, complicates the use of N,N-bis(2-chloroethyl)-N-nitrosourea as a GR inhibitor. We report 2-acetylamino-3-[4-(2-acetylamino-2-carboxyethylsulfanylthiocarbonylamino)phenylthiocarbamoylsulfanyl]propionic acid (2-AAPA) as a novel irreversible GR inhibitor. 2-AAPA was prepared by one-step synthesis from commercially available reagents. The Ki and kinact of 2-AAPA against yeast GR were determined to be 56 μm and 0.1 min–1, respectively. At the concentration that produced >80% yeast GR inhibition, 2-AAPA showed no inhibition against glutamylcysteine synthetase, glutathione synthetase, catalase, and superoxide dismutase, but minimal inhibition against glutathione S-transferase and glutathione peroxidase. In CV-1 cells, 2-AAPA (0.1 mm) produced 97% GR inhibition, 25% GSH reduction, and a 5-fold increase in GSSG in 20 min. The compound can be a useful tool in TRS-related research. PMID:19049979

Different roles of glutathione in copper and zinc chelation in Brassica napus roots.

PubMed

Zlobin, Ilya E; Kartashov, Alexander V; Shpakovski, George V

2017-09-01

We investigated the specific features of copper and zinc excess action on the roots of canola (Brassica napus L.) plants. Copper rapidly accumulated in canola root cells and reached saturation during several hours of treatment, whereas the root zinc content increased relatively slowly. Excessive copper and zinc entry inside the cell resulted in significant cell damage, as evidenced by alterations in plasmalemma permeability and decreases in cellular enzymatic activity. Zinc excess specifically damaged root hair cells, which correlated with a pronounced elevation of their labile zinc level. In vitro, we showed that reduced glutathione (GSH) readily reacted with copper ions to form complexes with blocked sulfhydryl groups. In contrast, zinc ions were ineffective as glutathione blockers, and glutathione molecules did not lose their specific chemical activity in the presence of Zn 2+ ions. The effect of copper and zinc excess on the glutathione pool in canola root cells was analysed by a combination of biochemical determination of total and oxidized glutathione contents and fluorescent staining of free reduced glutathione with monochlorobimane dye. Excess copper led to dose-dependent diminution of free reduced glutathione contents in the root cells, which could not be explained by the loss of total cellular glutathione or its oxidation. In contrast, we observed little effect of much higher intracellular zinc concentrations on the free reduced glutathione content. We concluded that GSH plays an important role in copper excess, but not zinc excess chelation, in canola root cells. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Endoplasmic Reticulum Transport of Glutathione by Sec61 Is Regulated by Ero1 and Bip.

PubMed

Ponsero, Alise J; Igbaria, Aeid; Darch, Maxwell A; Miled, Samia; Outten, Caryn E; Winther, Jakob R; Palais, Gael; D'Autréaux, Benoit; Delaunay-Moisan, Agnès; Toledano, Michel B

2017-09-21

In the endoplasmic reticulum (ER), Ero1 catalyzes disulfide bond formation and promotes glutathione (GSH) oxidation to GSSG. Since GSSG cannot be reduced in the ER, maintenance of the ER glutathione redox state and levels likely depends on ER glutathione import and GSSG export. We used quantitative GSH and GSSG biosensors to monitor glutathione import into the ER of yeast cells. We found that glutathione enters the ER by facilitated diffusion through the Sec61 protein-conducting channel, while oxidized Bip (Kar2) inhibits transport. Increased ER glutathione import triggers H 2 O 2 -dependent Bip oxidation through Ero1 reductive activation, which inhibits glutathione import in a negative regulatory loop. During ER stress, transport is activated by UPR-dependent Ero1 induction, and cytosolic glutathione levels increase. Thus, the ER redox poise is tuned by reciprocal control of glutathione import and Ero1 activation. The ER protein-conducting channel is permeable to small molecules, provided the driving force of a concentration gradient. Copyright © 2017 Elsevier Inc. All rights reserved.

Glutathione in the human brain: Review of its roles and measurement by magnetic resonance spectroscopy.

PubMed

Rae, Caroline D; Williams, Stephen R

2017-07-15

We review the transport, synthesis and catabolism of glutathione in the brain as well as its compartmentation and biochemistry in different brain cells. The major reactions involving glutathione are reviewed and the factors limiting its availability in brain cells are discussed. We also describe and critique current methods for measuring glutathione in the human brain using magnetic resonance spectroscopy, and review the literature on glutathione measurements in healthy brains and in neurological, psychiatric, neurodegenerative and neurodevelopmental conditions In summary: Healthy human brain glutathione concentration is ∼1-2 mM, but it varies by brain region, with evidence of gender differences and age effects; in neurological disease glutathione appears reduced in multiple sclerosis, motor neurone disease and epilepsy, while being increased in meningiomas; in psychiatric disease the picture is complex and confounded by methodological differences, regional effects, length of disease and drug-treatment. Both increases and decreases in glutathione have been reported in depression and schizophrenia. In Alzheimer's disease and mild cognitive impairment there is evidence for a decrease in glutathione compared to age-matched healthy controls. Improved methods to measure glutathione in vivo will provide better precision in glutathione determination and help resolve the complex biochemistry of this molecule in health and disease. Copyright © 2017 Elsevier Inc. All rights reserved.

Effect of Glutathione on the Taste and Texture of Type I Sourdough Bread.

PubMed

Tang, Kai Xing; Zhao, Cindy J; Gänzle, Michael G

2017-05-31

Type I sourdough fermentations with Lactobacillus sanfranciscensis as predominant organism accumulate reduced glutathione through glutathione reductase (GshR) activity of L. sanfranciscensis. Reduced glutathione acts as chain terminator for gluten polymerization but is also kokumi-active and may thus enhance bread taste. This study implemented a type I model sourdough fermentations to quantitate glutathione accumulation sourdough, bread dough, and bread and to assess the effect of L. sanfranciscensis GshR on bread volume by comparison of L. sanfranciscensis and an isogenic strain devoid of GshR. L. sanfranciscensis sourdough accumulated the highest amount of reduced glutathione during proofing. Bread produced with the wild type strain had a lower volume when compared to the gshR deficient mutant. The accumulation of γ-glutamyl-cysteine was also higher in L. sanfranciscensis sourdoughs when compared to doughs fermented with the gshR mutant strain. The accumulation of reduced glutathione in L. sanfranciscensis bread did not enhance the saltiness of bread.

Functional analysis and localisation of a delta-class glutathione S-transferase from Sarcoptes scabiei.

PubMed

Pettersson, Eva U; Ljunggren, Erland L; Morrison, David A; Mattsson, Jens G

2005-01-01

The mite Sarcoptes scabiei causes sarcoptic mange, or scabies, a disease that affects both animals and humans worldwide. Our interest in S. scabiei led us to further characterise a glutathione S-transferase. This multifunctional enzyme is a target for vaccine and drug development in several parasitic diseases. The S. scabiei glutathione S-transferase open reading frame reported here is 684 nucleotides long and yields a protein with a predicted molecular mass of 26 kDa. Through phylogenetic analysis the enzyme was classified as a delta-class glutathione S-transferase, and our paper is the first to report that delta-class glutathione S-transferases occur in organisms other than insects. The recombinant S. scabiei glutathione S-transferase was expressed in Escherichia coli via three different constructs and purified for biochemical analysis. The S. scabiei glutathione S-transferase was active towards the substrate 1-chloro-2,4-dinitrobenzene, though the positioning of fusion partners influenced the kinetic activity of the enzyme. Polyclonal antibodies raised against S. scabiei glutathione S-transferase specifically localised the enzyme to the integument of the epidermis and cavities surrounding internal organs in adult parasites. However, some minor staining of parasite intestines was observed. No staining was seen in host tissues, nor could we detect any antibody response against S. scabiei glutathione S-transferase in sera from naturally S. scabiei infected dogs or pigs. Additionally, the polyclonal sera raised against recombinant S. scabiei glutathione S-transferase readily detected a protein from mites, corresponding to the predicted size of native glutathione S-transferase.

1-3-A Resolution Structure of Human Glutathione S-Transferase With S-Hexyl Glutathione Bound Reveals Possible Extended Ligandin Binding Site

DOE Office of Scientific and Technical Information (OSTI.GOV)

Trong, I.Le; Stenkamp, R.E.; Ibarra, C.

2005-08-22

Cytosolic glutathione S-transferases (GSTs) play a critical role in xenobiotic binding and metabolism, as well as in modulation of oxidative stress. Here, the high-resolution X-ray crystal structures of homodimeric human GSTA1-1 in the apo form and in complex with S-hexyl glutathione (two data sets) are reported at 1.8, 1.5, and 1.3A respectively. At this level of resolution, distinct conformations of the alkyl chain of S-hexyl glutathione are observed, reflecting the nonspecific nature of the hydrophobic substrate binding site (H-site). Also, an extensive network of ordered water, including 75 discrete solvent molecules, traverses the open subunit-subunit interface and connects the glutathionemore » binding sites in each subunit. In the highest-resolution structure, three glycerol moieties lie within this network and directly connect the amino termini of the glutathione molecules. A search for ligand binding sites with the docking program Molecular Operating Environment identified the ordered water network binding site, lined mainly with hydrophobic residues, suggesting an extended ligand binding surface for nonsubstrate ligands, the so-called ligandin site. Finally, detailed comparison of the structures reported here with previously published X-ray structures reveal a possible reaction coordinate for ligand-dependent conformational changes in the active site and the C-terminus.« less

Glutathione Preservation during Storage of Rat Lenses in Optisol-GS and Castor Oil

PubMed Central

Holm, Thomas; Brøgger-Jensen, Martin Rocho; Johnson, Leif; Kessel, Line

2013-01-01

Background Glutathione concentration in the lens decreases in aging and cataractous lenses, providing a marker for tissue condition. Experimental procedures requiring unfrozen lenses from donor banks rely on transportation in storage medium, affecting lens homeostasis and alterations in glutathione levels. The aim of the study was to examine the effects of Optisol-GS and castor oil on lens condition, determined from their ability to maintain glutathione concentrations. Methodology/Principal Findings Rat lenses were stored in the two types of storage media at varying time intervals up to 3 days. Glutathione concentration was afterwards determined in an enzymatic detection assay, specific for both reduced and oxidized forms. Lenses removed immediately after death exhibited a glutathione concentration of 4.70±0.29 mM. In vitro stored lenses in Optisol-GS lost glutathione quickly, ending with a concentration of 0.60±0.34 mM after 3 days while castor oil stored lenses exhibited a slower decline and ended at 3 times the concentration. A group of lenses were additionally stored under post mortem conditions within the host for 6 hours before its removal. Total glutathione after 6 hours was similar to that of lenses removed immediately after death, but with altered GSH and GSSG concentrations. Subsequent storage of these lenses in media showed changes similar to those in the first series of experiments, albeit to a lesser degree. Conclusions/Significance It was determined that storage in Optisol-GS resulted in a higher loss of glutathione than lenses stored in castor oil. Storage for more than 12 hours reduced glutathione to half its original concentration, and was considered unusable after 24 hours. PMID:24260265

One-pot synthesis of glutathione by a two-enzyme cascade using a thermophilic ATP regeneration system.

PubMed

Zhang, Xing; Wu, Hui; Huang, Bing; Li, Zhimin; Ye, Qin

2017-01-10

In vitro cascade catalysis using enzyme-based system is becoming a promising biomanufacturing platform for biofuels and biochemicals production. Glutathione is a pivotal non-protein thiol compound and widely applied in food and pharmaceutical industries. In this study, glutathione was synthesized by a bifunctional glutathione synthetase together with a thermophilic ATP regeneration system through a two-enzyme cascade in vitro. Four bifunctional glutathione synthetases from Streptococcus sanguinis, S. gordonii, S. uberis and Bacillus cereus were applied for glutathione synthesis. The bifunctional glutathione synthetase from S. sanguinis was selected and coupled with the polyphosphate kinase from Thermosynechococcus elongatus BP-1 for regenerating ATP to produce glutathione in one pot. In the optimized system, 28.5mM glutathione was produced within 5h due to efficient ATP regeneration from low-cost polyphosphate. The yield based on added l-cysteine reached 81.4% and the productivity of glutathione achieved 5.7mM/h. The one-pot system indicated a potential biotransformation platform for industrial production of glutathione. Copyright © 2016 Elsevier B.V. All rights reserved.

Effect of T-2 Toxin, Fasting, and 2-Methyl-thiazolidine-4-carboxylate, a Glutathione Prodrug, on Hepatic Glutathione Levels1,2

DTIC Science & Technology

1986-11-14

Iftnvi fnr fard mirn: O Fo, M73 3 r0tiON OF I MOV 6s IS OBSOLETE SECURITY CLASSIFICATION OF THIS PACE (When Does Entoeed) , I I I I I 4.5± 0.39...Glende, 1973). An important cellular defense against peroxida- tive damage is the presence of glutathione and its use as an enzyme substrate or...cofactor. Even though intracellular glutathione concentration is in the millimolar range (Kosower and Kosower, 1978), there are conditions which lead to

Phytochelatin synthesis and glutathione levels in response to heavy metals in tomato cells.

PubMed

Scheller, H V; Huang, B; Hatch, E; Goldsbrough, P B

1987-12-01

Cell suspension cultures of tomato, Lycopersicon esculentum Mill. cv VFNT-Cherry, produce phytochelatins (poly[gamma-glutamylcysteinyl]glycines) when exposed to cadmium. The synthesis of these peptides is accompanied by a decline in cellular levels of glutathione. Buthionine sulfoximine, an inhibitor of glutathione synthesis, inhibits the sustained production of phytochelatins. However, phytochelatin synthesis can occur in the presence of buthionine sulfoximine provided that sufficient glutathione is available. These results indicate that glutathione is a substrate for phytochelatin synthesis. The protein synthesis inhibitor cycloheximide does not affect the initial production of phytochelatin.

The glutathione defense system in the pathogenesis of rheumatoid arthritis.

PubMed

Hassan, M Q; Hadi, R A; Al-Rawi, Z S; Padron, V A; Stohs, S J

2001-01-01

In order to assess a possible role of the natural glutathione defense system in the pathogenesis of rheumatoid arthritis (RA), serum reduced glutathione levels (GSH), glutathione reductase (GSR), glutathione S-transferase (GST), glutathione peroxidase (GSH-Px) and alkaline phosphatase (ALP) activities, lipid peroxidation (MDA content) and indexes of inflammation were evaluated in 58 rheumatic patients. Rheumatoid athritis was associated with significant depletion (ca. 50%) in GSH levels compared with normal control subjects. Serum levels of the detoxifying enzymes GSR and GSH-Px decreased by ca. 50% and 45%, respectively, whereas a threefold increase in the activity of GST was observed. A 1.2-fold increase in ALP was observed in patients with RA. These effects were accompanied by a 3.1-fold increase in serum MDA content. The MDA content was higher in RA patients who were seropositive for rheumatoid factor as well as positive for C-reactive proteins. The erythrocyte sedimentation rate for all patients with RA was approximately 13.8-fold higher than for the control group, and was higher among RA patients who were positive for C-reactive proteins and exhibited seropositivity for rheumatoid factor. Patients with RA receiving gold therapy exhibited significantly lower MDA levels whereas all other factors that were measured were not effected. The results support a hypothesis that defense mechanisms against reactive oxygen species are impaired in RA. Copyright 2001 John Wiley & Sons, Ltd.

Reactivity of 9-aminoacridine drug quinacrine with glutathione limits its antiprion activity.

PubMed

Šafařík, Martin; Moško, Tibor; Zawada, Zbigniew; Šafaříková, Eva; Dračínský, Martin; Holada, Karel; Šebestík, Jaroslav

2017-06-01

Quinacrine-the drug based on 9-aminoacridine-failed in clinical trials for prion diseases, whereas it was active in in vitro studies. We hypothesize that aromatic nucleophilic substitution at C9 could be contributing factor responsible for this failure because of the transfer of acridine moiety from quinacrine to abundant glutathione. Here, we described the semi-large-scale synthesis of the acridinylated glutathione and the consequences of its formation on biological and biophysical activities. The acridinylated glutathione is one order of magnitude weaker prion protein binder than the parent quinacrine. Moreover, according to log D pH 7.4 , the glutathione conjugate is two orders of magnitude more hydrophilic than quinacrine. Its higher hydrophilicity and higher dsDNA binding potency will significantly decrease its bioavailability in membrane-like environment. The glutathione deactivates quinacrine not only directly but also decreases its bioavailability. Furthermore, the conjugate can spontaneously decompose to practically insoluble acridone, which is precipitated out from the living systems. © 2016 John Wiley & Sons A/S.

A Western diet induced NAFLD in LDLR−/− mice is associated with reduced hepatic glutathione synthesis

PubMed Central

Li, Ling; Zhang, Guo-Fang; Lee, Kwangwon; Lopez, Rocio; Previs, Stephen F.; Willard, Belinda; McCullough, Arthur; Kasumov, Takhar

2017-01-01

Oxidative stress plays a key role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). Glutathione is the major anti-oxidant involved in cellular oxidative defense, however there are currently no simple non-invasive methods for assessing hepatic glutathione metabolism in patients with NAFLD. As a primary source of plasma glutathione, liver plays an important role in interorgan glutathione homeostasis. In this study, we have tested the hypothesis that measurements of plasma glutathione turnover could be used to assess the hepatic glutathione metabolism in LDLR−/− mice, a mouse model of diet-induced NAFLD. Mice were fed a standard low fat diet (LFD) or a high fat diet containing cholesterol (a Western type diet (WD)). The kinetics of hepatic and plasma glutathione were quantified using the 2H2O metabolic labeling approach. Our results show that a WD leads to reduced fractional synthesis rates (FSR) of hepatic (25%/h in LFD vs. 18%/h in WD, P < 0.05) and plasma glutathione (43%/h in LFD vs. 21%/h in WD, P <0.05), without any significant effect on their absolute production rates (PRs). WD-induced concordant changes in both hepatic and plasma glutathione turnover suggest that the plasma glutathione turnover measurements could be used to assess hepatic glutathione metabolism. The safety, simplicity, and low cost of the 2H2O-based glutathione turnover approach suggest that this method has the potential for non-invasive probing of hepatic glutathione metabolism in patients with NAFLD and other diseases. PMID:27036364

Inducible NAD(H)-linked methylglyoxal oxidoreductase regulates cellular methylglyoxal and pyruvate through enhanced activities of alcohol dehydrogenase and methylglyoxal-oxidizing enzymes in glutathione-depleted Candida albicans.

PubMed

Kwak, Min-Kyu; Ku, MyungHee; Kang, Sa-Ouk

2018-01-01

High methylglyoxal content disrupts cell physiology, but mammals have scavengers to prevent glycolytic and mitochondrial dysfunctions. In yeast, methylglyoxal accumulation triggers methylglyoxal-oxidizing alcohol dehydrogenase (Adh1) activity. While methylglyoxal reductases and glyoxalases have been well studied in prokaryotes and eukaryotes, experimental evidence for methylglyoxal dehydrogenase (Mgd) and other catalytic activities of this enzyme affecting glycolysis and the tricarboxylic acid cycle is lacking. A glycine-rich cytoplasmic Mgd protein, designated as Mgd1/Grp2, was isolated from glutathione-depleted Candida albicans. The effects of Mgd1/Grp2 activities on metabolic pathophysiology were investigated using knockout and overexpression mutants. We measured glutathione-(in)dependent metabolite contents and metabolic effects, including viability, oxygen consumption, ADH1 transcripts, and glutathione reductase and α-ketoglutarate dehydrogenase activities in the mutants. Based on the findings, methylglyoxal-oxidizing proteins were monitored to determine effects of MGD1/GRP2 disruption on methylglyoxal-scavenging traits during glutathione deprivation. Methylglyoxal-oxidizing NAD(H)-linked Mgd1/Grp2 was found solely in glutathione auxotrophs, and it catalyzed the reduction of both methylglyoxal and pyruvate. MGD1/GRP2 disruptants showed growth defects, cell-cycle arrest, and methylglyoxal and pyruvate accumulation with mitochondrial impairment, regardless of ADH1 compensation. Other methylglyoxal-oxidizing enzymes were identified as key glycolytic enzymes with enhanced activity and transcription in MGD1/GRP2 disruptants, irrespective of glutathione content. Failure of methylglyoxal and pyruvate dissimilation by Mgd1/Grp2 deficiency leads to poor glutathione-dependent redox regulation despite compensation by Adh1. This is the first report that multifunctional Mgd activities contribute to scavenging methylglyoxal and pyruvate to maintain metabolic homeostasis

A mathematical modelling approach to assessing the reliability of biomarkers of glutathione metabolism.

PubMed

Geenen, Suzanne; du Preez, Franco B; Reed, Michael; Nijhout, H Frederik; Kenna, J Gerry; Wilson, Ian D; Westerhoff, Hans V; Snoep, Jacky L

2012-07-16

One of the main pathways for the detoxification of reactive metabolites in the liver involves glutathione conjugation. Metabolic profiling studies have shown paradoxical responses in glutathione-related biochemical pathways. One of these is the increase in 5-oxoproline and ophthalmic acid concentrations with increased dosage of paracetamol. Experimental studies have thus far failed to resolve these paradoxes and the robustness of how these proposed biomarkers correlate with liver glutathione levels has been questioned. To better understand how these biomarkers behave in the glutathione system a kinetic model of this pathway was made. By using metabolic control analysis and by simulating biomarker levels under a variety of conditions, we found that 5-oxoproline and ophthalmic acid concentrations may not only depend on the glutathione but also on the methionine status of the cell. We show that neither of the two potential biomarkers are reliable on their own since they need additional information about the methionine status of the system to relate them uniquely to intracellular glutathione concentration. However, when both biomarkers are measured simultaneously a direct inference of the glutathione concentration can be made, irrespective of the methionine concentration in the system. Copyright © 2011 Elsevier B.V. All rights reserved.

Effects of glutathione on sperm quality during liquid storage in boars.

PubMed

Zhang, Xiao-Gang; Liu, Qi; Wang, Li-Qiang; Yang, Gong-She; Hu, Jian-Hong

2016-10-01

The aim of this study was to investigate the effects of different concentrations of glutathione in Modena on boar sperm quality during liquid storage at 17°C. Boar semen samples were collected and diluted with Modena containing different concentrations (0, 1, 5, 10, 15 mmol/L) of glutathione. Sperm motility, effective survival period, plasma membrane integrity, acrosome integrity, total antioxidant capacity (T-AOC) activity, malondialdehyde (MDA) content and hydrogen peroxide (H 2 O 2 ) content were measured and analyzed. The results showed that Modena supplemented with 1, 5 and 10 mmol/L glutathione improved sperm motility, effective survival period, plasma membrane integrity and T-AOC, and decreased MDA content and H 2 O 2 content. Meanwhile, the semen sample diluted with Modena containing 1 mmol/L glutathione achieved optimum effect, and effective survival period was 6.1 days. After 5 days preservation, sperm motility, plasma membrane integrity and T-AOC of the group treated with 1 mmol/L glutathione were all higher than that of other groups. Meanwhile, MDA content and H 2 O 2 content were lower than that of other groups. In conclusion, Modena supplemented with glutathione decreased the oxidative stress and improved the quality of boar semen during liquid storage at 17°C, and 1 mmol/L concentration was the optimum concentration. © 2016 Japanese Society of Animal Science. © 2016 Japanese Society of Animal Science.

Formation of 5-Oxoproline from Glutathione in Erythrocytes by the γ-Glutamyltranspeptidase-Cyclotransferase Pathway

PubMed Central

Palekar, Anil G.; Tate, Suresh S.; Meister, Alton

1974-01-01

γ-Glutamyltranspeptidase activity was demonstrated in the membrane fraction of rabbit erythrocytes. The activity observed (with glutathione and various amino-acid acceptors) was similar in magnitude to that of the γ-glutamylcyclotransferase and γ-glutamylcysteine synthetase activities found in the soluble fraction of the cell. No transpeptidase activity was observed with either γ-glutamyl p-nitroanilide or oxidized glutathione in contrast to the rabbit-kidney enzyme for which these compounds and glutathione serve as substrates. Erythrocyte suspensions and hemolysates formed 5-oxoproline (pyroglutamate; pyrrolidone carboxylate); the rate of 5-oxoproline formation from glutathione by hemolysates was increased by addition of methionine. The findings indicate that 5-oxoproline is an end-product of glutathione metabolism in erythrocytes, and that 5-oxoproline passes out of the erythrocyte and is metabolized in other tissues. The observed rate of 5-oxoproline formation is consistent with the conclusion that the γ-glutamyltranspeptidase-cyclotransferase pathway, together with the synthesis of glutathione from glycine, cysteine, and glutamate, account for a large fraction of the observed amino-acid turnover of erythrocyte glutathione. PMID:4150022

Glutathione reductase-mediated synthesis of tellurium-containing nanostructures exhibiting antibacterial properties.

PubMed

Pugin, Benoit; Cornejo, Fabián A; Muñoz-Díaz, Pablo; Muñoz-Villagrán, Claudia M; Vargas-Pérez, Joaquín I; Arenas, Felipe A; Vásquez, Claudio C

2014-11-01

Tellurium, a metalloid belonging to group 16 of the periodic table, displays very interesting physical and chemical properties and lately has attracted significant attention for its use in nanotechnology. In this context, the use of microorganisms for synthesizing nanostructures emerges as an eco-friendly and exciting approach compared to their chemical synthesis. To generate Te-containing nanostructures, bacteria enzymatically reduce tellurite to elemental tellurium. In this work, using a classic biochemical approach, we looked for a novel tellurite reductase from the Antarctic bacterium Pseudomonas sp. strain BNF22 and used it to generate tellurium-containing nanostructures. A new tellurite reductase was identified as glutathione reductase, which was subsequently overproduced in Escherichia coli. The characterization of this enzyme showed that it is an NADPH-dependent tellurite reductase, with optimum reducing activity at 30°C and pH 9.0. Finally, the enzyme was able to generate Te-containing nanostructures, about 68 nm in size, which exhibit interesting antibacterial properties against E. coli, with no apparent cytotoxicity against eukaryotic cells. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Glutathione Reductase-Mediated Synthesis of Tellurium-Containing Nanostructures Exhibiting Antibacterial Properties

PubMed Central

Pugin, Benoit; Cornejo, Fabián A.; Muñoz-Díaz, Pablo; Muñoz-Villagrán, Claudia M.; Vargas-Pérez, Joaquín I.; Arenas, Felipe A.

2014-01-01

Tellurium, a metalloid belonging to group 16 of the periodic table, displays very interesting physical and chemical properties and lately has attracted significant attention for its use in nanotechnology. In this context, the use of microorganisms for synthesizing nanostructures emerges as an eco-friendly and exciting approach compared to their chemical synthesis. To generate Te-containing nanostructures, bacteria enzymatically reduce tellurite to elemental tellurium. In this work, using a classic biochemical approach, we looked for a novel tellurite reductase from the Antarctic bacterium Pseudomonas sp. strain BNF22 and used it to generate tellurium-containing nanostructures. A new tellurite reductase was identified as glutathione reductase, which was subsequently overproduced in Escherichia coli. The characterization of this enzyme showed that it is an NADPH-dependent tellurite reductase, with optimum reducing activity at 30°C and pH 9.0. Finally, the enzyme was able to generate Te-containing nanostructures, about 68 nm in size, which exhibit interesting antibacterial properties against E. coli, with no apparent cytotoxicity against eukaryotic cells. PMID:25193000

Changes in glutathione redox cycle during diapause determination and termination in the bivoltine silkworm, Bombyx mori.

PubMed

Zhao, Lin-Chuan; Hou, Yi-Sheng; Sima, Yang-Hu

2014-02-01

To explore whether glutathione regulates diapause determination and termination in the bivoltine silkworm Bombyx mori, we monitored the changes in glutathione redox cycle in the ovary of both diapause- and nondiapause-egg producers, as well as those in diapause eggs incubated at different temperatures. The activity of thioredoxin reductase (TrxR) was detected in ovaries but not in eggs, while neither ovaries nor eggs showed activity of glutathione peroxidase. A lower reduced glutathione/oxidized glutathione (GSH/GSSG) ratio was observed in the ovary of diapause-egg producers, due to weaker reduction of oxidized glutathione (GSSG) to the reduced glutathione (GSH) catalyzed by glutathione reductase (GR) and TrxR. This indicates an oxidative shift in the glutathione redox cycle during diapause determination. Compared with the 25°C-treated diapause eggs, the 5°C-treated diapause eggs showed lower GSH/GSSG ratio, a result of stronger oxidation of GSH catalyzed by thioredoxin peroxidase and weaker reduction of GSSG catalyzed by GR. Our study demonstrated the important regulatory role of glutathione in diapause determination and termination of the bivoltine silkworm. © 2013 Institute of Zoology, Chinese Academy of Sciences.

Subcellular distribution of glutathione and its dynamic changes under oxidative stress in the yeast Saccharomyces cerevisiae

PubMed Central

Zechmann, Bernd; Liou, Liang-Chun; Koffler, Barbara E; Horvat, Lucija; Tomašić, Ana; Fulgosi, Hrvoje; Zhang, Zhaojie

2011-01-01

Glutathione is an important antioxidant in most prokaryotes and eukaryotes. It detoxifies reactive oxygen species and is also involved in the modulation of gene expression, in redox signaling, and in the regulation of enzymatic activities. In this study, the subcellular distribution of glutathione was studied in Saccharomyces cerevisiae by quantitative immunoelectron microscopy. Highest glutathione contents were detected in mitochondria and subsequently in the cytosol, nuclei, cell walls, and vacuoles. The induction of oxidative stress by hydrogen peroxide (H2O2) led to changes in glutathione-specific labeling. Three cell types were identified. Cell types I and II contained more glutathione than control cells. Cell type II differed from cell type I in showing a decrease in glutathione-specific labeling solely in mitochondria. Cell type III contained much less glutathione contents than the control and showed the strongest decrease in mitochondria, suggesting that high and stable levels of glutathione in mitochondria are important for the protection and survival of the cells during oxidative stress. Additionally, large amounts of glutathione were relocated and stored in vacuoles in cell type III, suggesting the importance of the sequestration of glutathione in vacuoles under oxidative stress. PMID:22093747

Glutathione cycle activity and pyridine nucleotide levels in oxidant-induced injury of cells.

PubMed Central

Schraufstätter, I U; Hinshaw, D B; Hyslop, P A; Spragg, R G; Cochrane, C G

1985-01-01

Exposure of target cells to a bolus of H2O2 induced cell lysis after a latent period of several hours, which was prevented only when the H2O2 was removed within the first 30 min of injury by addition of catalase. This indicated that early metabolic events take place that are important in the fate of the cell exposed to oxidants. In this study, we described two early and independent events of H2O2-induced injury in P388D1 macrophagelike tumor cells: activation of the glutathione cycle and depletion of cellular NAD. Glutathione cycle and hexose monophosphate shunt (HMPS) were activated within seconds after the addition of H2O2. High HMPS activity maintained glutathione that was largely reduced. However, when HMPS activity was inhibited--by glucose depletion or by incubation at 4 degrees C--glutathione remained in the oxidized state. Total pyridine nucleotide levels were diminished when cells were exposed to H2O2, and the breakdown product, nicotinamide, was recovered in the extracellular medium. Intracellular NAD levels fell by 80% within 20 min of exposure of cells to H2O2. The loss of NADP(H) and stimulation of the HMPS could be prevented when the glutathione cycle was inhibited by either blocking glutathione synthesis with buthionine sulfoximine (BSO) or by inhibiting glutathione reductase with (1,3-bis) 2 chlorethyl-1-nitrosourea. The loss of NAD developed independently of glutathione cycle and HMPS activity, as it also occurred in BSO-treated cells. PMID:3840176

Elevation of Glutathione as a Therapeutic Strategy in Alzheimer Disease

PubMed Central

Pocernich, Chava B.; Butterfield, D. Allan

2011-01-01

Oxidative stress has been associated with the onset and progression of mild cognitive impairment (MCI) and Alzheimer disease (AD). AD and MCI brain and plasma display extensive oxidative stress as indexed by protein oxidation, lipid peroxidation, free radical formation, DNA oxidation, and decreased antioxidants. The most abundant endogenous antioxidant, glutathione, plays a significant role in combating oxidative stress. The ratio of oxidized to reduced glutathione is utilized as a measure of intensity of oxidative stress. Antioxidants have long been considered as an approach to slow down AD progression. In this review, we focus on the elevation on glutathione through N-acytl-cysteine (NAC) and γ-glutamylcysteine ethyl ester (GCEE) as a potential therapeutic approach for Alzheimer disease. PMID:22015471

Rat lung glutathione S-transferases. Evidence for two distinct types of 22000-Mr subunits.

PubMed Central

Singh, S V; Partridge, C A; Awasthi, Y C

1984-01-01

Two immunologically distinct types of 22000-Mr subunits are present in rat lung glutathione S-transferases. One of these subunits is probably similar to Ya subunits of rat liver glutathione S-transferases, whereas the other subunit Ya' is immunologically distinct. Glutathione S-transferase II (pI7.2) of rat lung is a heterodimer (YaYa') of these subunits, and glutathione S-transferase VI (pI4.8) of rat lung is a homodimer of Ya' subunits. On hybridization in vitro of the subunits of glutathione S-transferase II of rat lung three active dimers having pI values 9.4, 7.2 and 4.8 are obtained. Immunological properties and substrate specificities indicate that the hybridized enzymes having pI7.2 and 4.8 correspond to glutathione S-transferases II and VI of rat lung respectively. Images Fig. 1. Fig. 5. PMID:6433888

Regression of Aflatoxin B1-Induced Hepatocellular Carcinomas by Reduced Glutathione

NASA Astrophysics Data System (ADS)

Novi, Anna M.

1981-05-01

Reduced glutathione administered to rats bearing aflatoxin B1-induced liver tumors caused regression of tumor growth and resulted in survival of the animals. Since glutathione is a harmless natural product, it merits further investigation as a potential antitumor drug for humans.

[Effect of bemethyl on the glutathione system in the rat liver in acute hypoxia].

PubMed

Zarubina, I V; Mironova, O P

2002-01-01

The effect of bemithyl on the state of liver glutathione system was studied in rats under acute hypoxic hypoxia conditions modeled by "elevating" animals in a pressure chamber up to an altitude of 8000-11,000 m for 30 min. Bemithyl (25 mg/kg, i.p.) administered 30 min before the hypoxia onset, prevents a decrease in the content of reduced glutathione and SH groups and impedes a drop in the activity of glutathione reductase and glutathione peroxidase. By means of the inhibition analysis using actinomycin D (a protein synthesis inhibitor), it was established that the protective action of bemithyl is related to the ability of enhancing the synthesis of antioxidant enzymes in the liver glutathione system.

Crystallization and preliminary X-ray crystallographic analysis of Escherichia coli glutaredoxin 2 in complex with glutathione and of a cysteine-less variant without glutathione

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sheng, Ju; Ye, Jun; Rosen, Barry P., E-mail: brosen@med.wayne.edu

2007-04-01

Glutaredoxin 2 from E. coli was cocrystallized with glutathione and data were collected to 1.60 Å. A mutant with the active-site residues Cys9 and Cys12 changed to serine was crystallized in the absence of glutathione and data were collected to 2.4 Å. Glutaredoxin 2 (Grx2) from Escherichia coli is larger in size than classical glutaredoxins. It is extremely efficient in the catalysis of reduced glutathione-dependent disulfide reduction. A complex of Grx2 with reduced glutathione (GSH) has been crystallized. Data were collected to 1.60 Å. The crystals belong to space group P3{sub 2}21, with one Grx2–GSH complex in the asymmetric unit.more » The unit-cell parameters are a = b = 50.10, c = 152.47 Å. A Grx2 mutant, C9S/C12S, which cannot form a disulfide bond with GSH was also crystallized. The crystals diffracted to 2.40 Å and belong to space group P2{sub 1}2{sub 1}2{sub 1}, with one molecule in the asymmetric unit. The unit-cell parameters are a = 28.16, b = 78.65, c = 89.16 Å.« less

Genetics Home Reference: glutathione synthetase deficiency

MedlinePlus

... Ristoff E, Larsson A. Inborn errors in the metabolism of glutathione. Orphanet J Rare Dis. 2007 Mar 30;2:16. Review. Citation on PubMed or Free article on PubMed Central Wu G, Fang YZ, Yang S, ...

Long-term dietary supplementation with cystathionine improves tissue glutathione in ageing rats.

PubMed

Pouget, Mélanie; Perrot, Marie; Denis, Philippe; Vuichoud, Jacques; Dardevet, Dominique; Vidal, Karine; Breuillé, Denis; Papet, Isabelle

2016-08-01

Ageing is associated with decrease in tissue glutathione that can be reduced by food fortification with the amino acid cysteine. However, cysteine is not stable in solution and generates bad taste. Cystathionine, the direct precursor of cysteine, could be a valuable alternative. This study aimed to determine whether long-term dietary supplementation with cystathionine induces an increase in glutathione pools. Aged rats (20.5-month-old) were fed ad libitum during 29 weeks with either a cystathionine-supplemented diet (7.3 g/kg, n = 90 rats) or a control iso-nitrogenous alanine-supplemented diet (2.9 g/kg, n = 90 rats). Cystathionine was detected in the plasma of the cystathionine-supplemented rats but not in the control alanine-supplemented rats. Cystathionine increased glutathione concentrations in liver, small intestine and gastrocnemius muscle (P < 0.03). No adverse effect was observed. Cystathionine supplementation being able to increase moderately glutathione in healthy old rats could be considered as a candidate for nutritional supports aiming to revert the stronger glutathione depletions occurring in unhealthy elderly.

Enhancement of Combined Umami and Salty Taste by Glutathione in the Human Tongue and Brain.

PubMed

Goto, Tazuko K; Yeung, Andy Wai Kan; Tanabe, Hiroki C; Ito, Yuki; Jung, Han-Sung; Ninomiya, Yuzo

2016-09-01

Glutathione, a natural substance, acts on calcium receptors on the tongue and is known to enhance basic taste sensations. However, the effects of glutathione on brain activity associated with taste sensation on the tongue have not been determined under standardized taste delivery conditions. In this study, we investigated the sensory effect of glutathione on taste with no effect of the smell when glutathione added to a combined umami and salty taste stimulus. Twenty-six volunteers (12 women and 14 men; age 19-27 years) performed a sensory evaluation of taste of a solution of monosodium L-glutamate and sodium chloride, with and without glutathione. The addition of glutathione changed taste qualities and significantly increased taste intensity ratings under standardized taste delivery conditions (P < 0.001). Functional magnetic resonance imaging showed that glutathione itself elicited significant activation in the left ventral insula. These results are the first to demonstrate the enhancing effect of glutathione as reflected by brain data while tasting an umami and salty mixture. © The Author 2016. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Glutathione system participation in thoracic aneurysms from patients with Marfan syndrome.

PubMed

Zúñiga-Muñoz, Alejandra María; Pérez-Torres, Israel; Guarner-Lans, Verónica; Núñez-Garrido, Elías; Velázquez Espejel, Rodrigo; Huesca-Gómez, Claudia; Gamboa-Ávila, Ricardo; Soto, María Elena

2017-05-01

Aortic dilatation in Marfan syndrome (MFS) is progressive. It is associated with oxidative stress and endothelial dysfunction that contribute to the early acute dissection of the vessel and can result in rupture of the aorta and sudden death. We evaluated the participation of the glutathione (GSH) system, which could be involved in the mechanisms that promote the formation and progression of the aortic aneurysms in MFS patients. Aortic aneurysm tissue was obtained during chest surgery from eight control subjects and 14 MFS patients. Spectrophotometrical determination of activity of glutathione peroxidase (GPx), glutathione-S-transferase (GST), glutathione reductase (GR), lipid peroxidation (LPO) index, carbonylation, total antioxidant capacity (TAC), and concentration of reduced and oxidized glutathione (GSH and GSSG respectively), was performed in the homogenate from aortic aneurysm tissue. LPO index, carbonylation, TGF-β1, and GR activity were increased in MFS patients (p < 0.04), while TAC, GSH/GSSG ratio, GPx, and GST activity were significantly decreased (p < 0.04). The depletion of GSH, in spite of the elevated activity of GR, not only diminished the activity of GSH-depend GST and GPx, but increased LPO, carbonylation and decreased TAC. These changes could promote the structural and functional alterations in the thoracic aorta of MFS patients.

Glutathione level after long-term occupational elemental mercury exposure

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kobal, Alfred Bogomir; Prezelj, Marija; Horvat, Milena

2008-05-15

Many in vitro and in vivo studies have elucidated the interaction of inorganic mercury (Hg) and glutathione. However, human studies are limited. In this study, we investigated the potential effects of remote long-term intermittent occupational elemental Hg vapour (Hg{sup o}) exposure on erythrocyte glutathione levels and some antioxidative enzyme activities in ex-mercury miners in the period after exposure. The study included 49 ex-mercury miners divided into subgroups of 28 still active, Hg{sup o}-not-exposed miners and 21 elderly retired miners, and 41 controls, age-matched to the miners subgroup. The control workers were taken from 'mercury-free works'. Reduced glutathione (GSH) and oxidizedmore » disulphide glutathione (GSSG) concentrations in haemolysed erythrocytes were determined by capillary electrophoresis, while total glutathione (total GSH) and the GSH/GSSG ratio were calculated from the determined values. Catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR) activities in erythrocytes were measured using commercially available reagent kits, while urine Hg (U-Hg) concentrations were determined by cold vapour atomic absorption (CVAAS). No correlation of present U-Hg levels, GSH, GSSG, and antioxidative enzymes with remote occupational biological exposure indices were found. The mean CAT activity in miners and retired miners was significantly higher (p<0.05) than in the controls. No differences in mean GPx activity among the three groups were found, whereas the mean GR activity was significantly higher (p<0.05) in miners than in retired miners. The mean concentrations of GSH (mmol/g Hb) in miners (13.03{+-}3.71) were significantly higher (p<0.05) than in the control group (11.68{+-}2.66). No differences in mean total GSH, GSSG levels, and GSH/GSSG ratio between miners and controls were found. A positive correlation between GSSG and present U-Hg excretion (r=0.41, p=0.001) in the whole group of ex-mercury miners was observed. The

Glutathione Transferase U13 Functions in Pathogen-Triggered Glucosinolate Metabolism.

PubMed

Piślewska-Bednarek, Mariola; Nakano, Ryohei Thomas; Hiruma, Kei; Pastorczyk, Marta; Sanchez-Vallet, Andrea; Singkaravanit-Ogawa, Suthitar; Ciesiołka, Danuta; Takano, Yoshitaka; Molina, Antonio; Schulze-Lefert, Paul; Bednarek, Paweł

2018-01-01

Glutathione (GSH) and indole glucosinolates (IGs) exert key functions in the immune system of the model plant Arabidopsis ( Arabidopsis thaliana ). Appropriate GSH levels are important for execution of both pre- and postinvasive disease resistance mechanisms to invasive pathogens, whereas an intact PENETRATION2 (PEN2)-pathway for IG metabolism is essential for preinvasive resistance in this species. Earlier indirect evidence suggested that the latter pathway involves conjugation of GSH with unstable products of IG metabolism and further processing of the resulting adducts to biologically active molecules. Here we describe the identification of Glutathione- S -Transferase class-tau member 13 (GSTU13) as an indispensable component of the PEN2 immune pathway for IG metabolism. gstu13 mutant plants are defective in the pathogen-triggered biosynthesis of end products of the PEN2 pathway, including 4-O-β-d-glucosyl-indol-3-yl formamide, indole-3-ylmethyl amine, and raphanusamic acid. In line with this metabolic defect, lack of functional GSTU13 results in enhanced disease susceptibility toward several fungal pathogens including Erysiphe pisi , Colletotrichum gloeosporioides , and Plectosphaerella cucumerina Seedlings of gstu13 plants fail also to deposit the (1,3)-β-glucan cell wall polymer, callose, after recognition of the bacterial flg22 epitope. We show that GSTU13 mediates specifically the role of GSH in IG metabolism without noticeable impact on other immune functions of this tripeptide. We postulate that GSTU13 connects GSH with the pathogen-triggered PEN2 pathway for IG metabolism to deliver metabolites that may have numerous functions in the innate immune system of Arabidopsis. © 2018 American Society of Plant Biologists. All Rights Reserved.

Identification of Small-Molecule Frequent Hitters of Glutathione S-Transferase-Glutathione Interaction.

PubMed

Brenke, Jara K; Salmina, Elena S; Ringelstetter, Larissa; Dornauer, Scarlett; Kuzikov, Maria; Rothenaigner, Ina; Schorpp, Kenji; Giehler, Fabian; Gopalakrishnan, Jay; Kieser, Arnd; Gul, Sheraz; Tetko, Igor V; Hadian, Kamyar

2016-07-01

In high-throughput screening (HTS) campaigns, the binding of glutathione S-transferase (GST) to glutathione (GSH) is used for detection of GST-tagged proteins in protein-protein interactions or enzyme assays. However, many false-positives, so-called frequent hitters (FH), arise that either prevent GST/GSH interaction or interfere with assay signal generation or detection. To identify GST-FH compounds, we analyzed the data of five independent AlphaScreen-based screening campaigns to classify compounds that inhibit the GST/GSH interaction. We identified 53 compounds affecting GST/GSH binding but not influencing His-tag/Ni(2+)-NTA interaction and general AlphaScreen signals. The structures of these 53 experimentally identified GST-FHs were analyzed in chemoinformatic studies to categorize substructural features that promote interference with GST/GSH binding. Here, we confirmed several existing chemoinformatic filters and more importantly extended them as well as added novel filters that specify compounds with anti-GST/GSH activity. Selected compounds were also tested using different antibody-based GST detection technologies and exhibited no interference clearly demonstrating specificity toward their GST/GSH interaction. Thus, these newly described GST-FH will further contribute to the identification of FH compounds containing promiscuous substructures. The developed filters were uploaded to the OCHEM website (http://ochem.eu) and are publicly accessible for analysis of future HTS results. © 2016 Society for Laboratory Automation and Screening.

Glutathione reduces cytotoxicity of polyethyleneimine coated magnetic nanoparticles in CHO cells.

PubMed

Strojan, Klemen; Lojk, Jasna; Bregar, Vladimir B; Veranič, Peter; Pavlin, Mojca

2017-06-01

Polyethyleneimine (PEI) is a polycationic compound frequently used as a transfection agent. However, cytotoxicity of PEI and PEI-coated nanoparticles (PEI NPs) is still a major obstacle in its use. In this study we report a method for reducing cytotoxicity of PEI NPs by addition of glutathione in NPs synthesis. Glutathione reduced cytotoxic effects for at least 30% and decreased observed oxidative stress response compared to standard formulation. Results showed that the effect was partially due to reduced zeta potential and partially due to protective antioxidant properties of glutathione. Addition of glutathione to cell culture media with concurrent exposure to PEI NPs proved to be insufficient for cytotoxicity reduction. Additionally, we compared internalization pathways of both PEI NPs and GSH NPs. NPs were only found in endosomes and no NPs were found free in the cytosol, as would be expected according to so called proton sponge hypothesis. Copyright © 2017 Elsevier B.V. All rights reserved.

S-glutathionyl-(chloro)hydroquinone reductases: a new class of glutathione transferases functioning as oxidoreductases

PubMed Central

Belchik, Sara M.; Xun, Luying

2011-01-01

Glutathione transferases (GSTs) are best known for transferring glutathione (GSH) to hydrophobic organic compounds, making the conjugates more soluble. However, the omega-class GSTs of animals and the lambda-class GSTs and dehydroascorbate reductases (DHARs) of plants have little or no activity for GSH transfer. Instead, they catalyze GSH-dependent oxidoreductions. The lambda-class GSTs reduce disulfide bonds, the DHARs reduce the disulfide bonds and dehydroascorbate, and the omega-class GSTs can reduce more substrates, including disulfide bonds, dehydroascorbate, and dimethylarsinate. Glutathionyl-(chloro)hydroquinone reductases (GS-HQRs) are the newest class of GSTs that mainly catalyze oxidoreductions. Besides the activities of the other three classes, GS-HQRs also reduce GS-hydroquinones, including GS-trichloro-p-hydroquinone, GS-dichloro-p-hydroquinone, GS-2-hydroxy-p-hydroquinone, and GS-p-hydroquinone. They are conserved and widely distributed in bacteria, fungi, protozoa, and plants, but not in animals. The four classes are phylogenetically more related to each other than to other GSTs, and they share a Cys-Pro motif at the GSH-binding site. Hydroquinones are metabolic intermediates of certain aromatic compounds. They can be auto-oxidized by O2 to benzoquinones, which spontaneously react with GSH to form GS-hydroquinones via Michael’s addition. GS-HQRs are expected to channel GS-hydroquinones, formed spontaneously or enzymatically, back to hydroquinones. When the released hydroquinones are intermediates of metabolic pathways, GS-HQRs play a maintenance role for the pathways. Further, the common presence of GS-HQRs in plants, green algae, cyanobacteria, and halobacteria suggest a beneficial role in the light-using organisms. PMID:21425927

[Effects of dissolved oxygen and pH on Candida utilis batch fermentation of glutathione].

PubMed

Wei, Gong-Yuan; Li, Yin; Du, Guo-Cheng; Chen, Jian

2003-11-01

The effects of dissolved oxygen (DO) and pH on glutathione batch fermentation by Candida utilis WSH-02-08 in a 7 liters stirred fermentor were investigated. It was shown that DO concentration is an important factor in glutathione production. With the initial glucose concentration of 30 g/L and a 5 L/min air flow rate, and the agitation rate less than 250 r/min, the DO concentration was not sufficient to satisfy the oxygen requirement during the fermentation. With an agitation rate of more than 300 r/min, the cell growth and glutathione production were enhanced significantly, with the dry cell mass and glutathione production were 20% and 25% higher than that at 200 r/min. When C. utilis WSH 02-08 was cultivated in a batch process without pH control, cell growth and glutathione production were inhibited, likely due to a dramatic decrease in the pH. Intracellular glutathione leakages were observed when the pH was 1.5 or less. To assess the effect of pH on glutathione production, six batch processes controlled at pH 4.0, 4.5, 5.0, 5.5, 6.0 and 6.5 were conducted. The yield was highest at pH 5.5, when the dry cell mass and yield were 27% and 95% respectively higher than fermentation without pH control. The maximal intracellular glutathione content (2.15 %) was also achieved at the pH. To improve our understandings on the effect of pH on the batch glutathione production, a modified Logistic equation and Luedeking-Piret equation were used to simulate cell growth and glutathione production, respectively, under different pH. Based on the parameters obtained by the nonlinear estimation, kinetic analysis was performed to elucidate the effect of pH on the batch glutathione production. The process controlled at pH 5.5 was proven to be the best due to the higher value of K(I) (substrate inhibitory constant in the Logistic equation), lower value of a and higher value of beta (slope and intercept in the Luedeking-Piret equation, respectively).

Improvements in the bread-making quality of gluten-free rice batter by glutathione.

PubMed

Yano, Hiroyuki

2010-07-14

The wide prevalence of celiac disease and wheat allergy has led to a growing demand for gluten-free foods. Rice proteins do not possess the viscoelastic properties typically found in gluten, thus making rice flour unsuitable for the production of yeast-leavened products. In the present study, we found that the addition of glutathione to rice batter improves its gas-retaining properties. Glutathione was found to prevent the formation of the disulfide-linked macromolecular protein barrier, which is reported to confer resistance to the deformation of rice batter in the baking process. Also, glutathione appeared to gelatinize rice starch at lower temperatures. Microstructure analyses of glutathione-added rice bread revealed it to have a perforated structure like wheat bread but with a smoother-looking surface. These data collectively suggest that glutathione facilitates the deformation of rice batter, thus increasing its elasticity in the early stages of bread baking and the volume of the resulting bread.

Purification and kinetic analysis of cytosolic and mitochondrial thioredoxin glutathione reductase extracted from Taenia solium cysticerci.

PubMed

Plancarte, Agustin; Nava, Gabriela

2015-02-01

Thioredoxin glutathione reductases (TGRs) (EC 1.8.1.9) were purified to homogeneity from the cytosolic (cTsTGR) and mitochondrial (mTsTGR) fractions of Taenia solium, the agent responsible for neurocysticercosis, one of the major central nervous system parasitic diseases in humans. TsTGRs had a relative molecular weight of 132,000, while the corresponding value per subunit obtained under denaturing conditions, was of 62,000. Specific activities for thioredoxin reductase and glutathione reductase substrates for both TGRs explored were in the range or lower than values obtained for other platyhelminths and mammalian TGRs. cTsTGR and mTsTGR also showed hydroperoxide reductase activity using hydroperoxide as substrate. Km(DTNB) and Kcat(DTNB) values for cTsTGR and mTsTGR (88 µM and 1.9 s(-1); 45 µM and 12.6 s(-1), respectively) and Km(GSSG) and Kcat(GSSG) values for cTsTGR and mTsTGR (6.3 µM and 0.96 s(-1); 4 µM and 1.62 s(-1), respectively) were similar to or lower than those reported for mammalian TGRs. Mass spectrometry analysis showed that 12 peptides from cTsTGR and seven from mTsTGR were a match for gi|29825896 thioredoxin glutathione reductase [Echinococcus granulosus], confirming that both enzymes are TGRs. Both T. solium TGRs were inhibited by the gold compound auranofin, a selective inhibitor of thiol-dependent flavoreductases (I₅₀ = 3.25, 2.29 nM for DTNB and GSSG substrates, respectively for cTsTGR; I₅₀ = 5.6, 25.4 nM for mTsTGR toward the same substrates in the described order). Glutathione reductase activity of cTsTGR and mTsTGR exhibited hysteretic behavior with moderate to high concentrations of GSSG; this result was not observed either with thioredoxin, DTNB or NADPH. However, the observed hysteretic kinetics was suppressed with increasing amounts of both parasitic TGRs. These data suggest the existence of an effective substitute which may account for the lack of the detoxification enzymes glutathione reductase

Intact protein folding in the glutathione-depleted endoplasmic reticulum implicates alternative protein thiol reductants

PubMed Central

Tsunoda, Satoshi; Avezov, Edward; Zyryanova, Alisa; Konno, Tasuku; Mendes-Silva, Leonardo; Pinho Melo, Eduardo; Harding, Heather P; Ron, David

2014-01-01

Protein folding homeostasis in the endoplasmic reticulum (ER) requires efficient protein thiol oxidation, but also relies on a parallel reductive process to edit disulfides during the maturation or degradation of secreted proteins. To critically examine the widely held assumption that reduced ER glutathione fuels disulfide reduction, we expressed a modified form of a cytosolic glutathione-degrading enzyme, ChaC1, in the ER lumen. ChaC1CtoS purged the ER of glutathione eliciting the expected kinetic defect in oxidation of an ER-localized glutathione-coupled Grx1-roGFP2 optical probe, but had no effect on the disulfide editing-dependent maturation of the LDL receptor or the reduction-dependent degradation of misfolded alpha-1 antitrypsin. Furthermore, glutathione depletion had no measurable effect on induction of the unfolded protein response (UPR); a sensitive measure of ER protein folding homeostasis. These findings challenge the importance of reduced ER glutathione and suggest the existence of alternative electron donor(s) that maintain the reductive capacity of the ER. DOI: http://dx.doi.org/10.7554/eLife.03421.001 PMID:25073928

Hemolytic anemia and metabolic acidosis: think about glutathione synthetase deficiency.

PubMed

Ben Ameur, Salma; Aloulou, Hajer; Nasrallah, Fehmi; Kamoun, Thouraya; Kaabachi, Naziha; Hachicha, Mongia

2015-02-01

Glutathione synthetase deficiency (GSSD) is a rare disorder of glutathione metabolism with varying clinical severity. Patients may present with hemolytic anemia alone or together with acidosis and central nervous system impairment. Diagnosis is made by clinical presentation and detection of elevated concentrations of 5-oxoproline in urine and low glutathione synthetase activity in erythrocytes or cultured skin fibroblasts. The prognosis seems to depend on early diagnosis and treatment. We report a 4 months old Tunisian male infant who presented with severe metabolic acidosis with high anion gap and hemolytic anemia. High level of 5-oxoproline was detected in her urine and diagnosis of GSSD was made. Treatment consists of the correction of acidosis, blood transfusion, and supplementation with antioxidants. He died of severe metabolic acidosis and sepsis at the age of 15 months.

Mercury(II) Complex Formation With Glutathione in Alkaline Aqueous Solution

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mah, V.; Jalilehvand, F.

2009-05-19

The structure and speciation of the complexes formed between mercury(II) ions and glutathione (GSH = L-glutamyl-L-cysteinyl-glycine) have been studied for a series of alkaline aqueous solutions (C{sub Hg{sup 2+}} {approx} 18 mmol dm{sup -3} and C{sub GSH} = 40-200 mmol dm{sup -3} at pH {approx} 10.5) by means of extended X-ray absorption fine structure (EXAFS) and {sup 199}Hg NMR spectroscopy at ambient temperature. The dominant complexes are [Hg(GS){sub 2}]{sup 4-} and [Hg(GS){sub 3}]{sup 7-}, with mean Hg-S bond distances of 2.32(1) and 2.42(2) {angstrom} observed in digonal and trigonal Hg-S coordination, respectively. The proportions of the Hg{sup 2+}-glutathione complexes weremore » evaluated by fitting linear combinations of model EXAFS oscillations representing each species to the experimental EXAFS spectra. The [Hg(GS){sub 4}]{sup 10-} complex, with four sulfur atoms coordinated at a mean Hg-S bond distance of 2.52(2) {angstrom}, is present in minor amounts (<30%) in solutions containing a large excess of glutathione (C{sub GSH} {ge} 160 mmol dm{sup -3}). Comparable alkaline mercury(II) cysteine (H{sub 2}Cys) solutions were also investigated and a reduced tendency to form higher complexes was observed, because the deprotonated amino group of Cys{sup 2-} allows the stable [Hg(S,N-Cys){sub 2}]{sup 2-} chelate to form. The effect of temperature on the distribution of the Hg{sup 2+}-glutathione complexes was studied by comparing the EXAFS spectra at ambient temperature and at 25 K of a series of glycerol/water (33/67, v/v) frozen glasses with and C{sub Hg{sup 2+}} {approx} 7 mmol dm{sup -3} and C{sub GSH} = 16-81 mmol dm{sup -3}. Complexes with high Hg-S coordination numbers, [Hg(GS){sub 3}]{sup 7-} and [Hg(GS){sub 4}]{sup 10-}, became strongly favored when just a moderate excess of glutathione (C{sub GSH} {ge} 28 mmol dm{sup -3}) was used in the glassy samples, as expected for a stepwise exothermic bond formation. Addition of glycerol had no effect on the Hg(II)-glutathione

[Effects of glutamic acid and glutathione on the secretory function of the stomach].

PubMed

Shlygin, G K; Vasilevskaia, L S; Ignatenko, L G

1988-10-01

Experiments on dogs with Pavlov isolated pouches and gastric fistulas have shown that the ingested solution of MSG produces a potentiating effect on maximal gastric secretion caused by pentagastrin. This effect is apparently connected with the formation of glutathione in intestine. The glutathione concentration in blood after the intake of MSG is significantly elevated. It has been established that reduced glutathione administered in blood produced the similar potentiating effect on gastric secretion caused by pentagastrin.

Glutathione affects the transport activity of Rhizobium leguminosarum 3841 and is essential for efficient nodulation.

PubMed

Cheng, Guojun; Karunakaran, Ramakrishnan; East, Alison K; Munoz-Azcarate, Olaya; Poole, Philip S

2017-04-01

As glutathione (GSH) plays an essential role in growth and symbiotic capacity of rhizobia, a glutathione synthetase (gshB) mutant of Rhizobium leguminosarum biovar viciae 3841 (Rlv3841) was characterised. It fails to efficiently utilise various compounds as a sole carbon source, including glucose, succinate, glutamine and histidine, and shows 60%-69% reduction in uptake rates of glucose, succinate and the non-metabolisable substrate α-amino isobutyric acid. The defect in glucose uptake can be overcome by addition of exogenous GSH, indicating GSH, but not its bacterial synthesis, is required for efficient transport. GSH is not involved in the regulation of the activity of Rlv3841's transporters via the global regulator of transport, PtsNTR. Although lack of GSH reduces transcription of the branched amino acid transporter, this was not the case for all uptake transport systems, for example, the amino acid permease. This suggests GSH alters activity and/or assembly of transport systems by an unknown mechanism. In interaction with plants, the gshB mutant is not only severely impaired in rhizosphere colonisation, but also shows a 50% reduction in dry weight of plants and nitrogen-fixation ability. This reveals that changes in GSH metabolism affect the bacterial-plant interactions required for symbiosis. © FEMS 2017.

Glutathione in combination with trehalose has supplementary beneficial effects on cryopreserved red deer (cervus elaphus) sperm.

PubMed

Wang, Yan; Dong, Shude

2017-01-01

In this study, we evaluated the effects of glutathione in combination with trehalose addition to semen extenders on the quality parameters of frozen-thawed red deer (cervus elaphus) spermatozoa. The semen samples collected from six mature red deer once a week were diluted with Tris-egg yolk-based extenders. The diluted semen samples were supplemented with glutathione (8 mmol L -1 ) and or trehalose (5%, w/v), cryopreserved, thawed and then subjected to sperm quality parameter evaluation. Both glutathione and trehalose addition to the extender significantly improved progressive motility, acrosome integrity, membrane integrity, superoxide dismutase and glutathione peroxidase activity and decreased percentage abnormality and sperm malondialdehyde level compared with the control group (P<.05). Moreover, glutathione in combination with trehalose addition to semen extenders had higher efficiency compared with the glutathione or trehalose addition alone (P<.05). Therefore, glutathione in combination with trehalose could be a promising cryoprotectant for red deer sperm. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Glutathione and glutathione S-transferase levels in patients with liver metastases of colorectal cancer and other hepatic disorders.

PubMed

Baltruskeviciene, Edita; Kazbariene, Birutė; Badaras, Robertas; Bagdonaitė, Loreta; Krikštaponienė, Aurelija; Zdanavičius, Linas; Aleknavicius, Eduardas; Didziapetrienė, Janina

2016-07-01

Glutathione and glutathione S-transferases (GST) are involved in cell defence against reactive oxygen species, which induces oxidative stress and are associated with different chronic diseases. The aim of the present study was to determine the differences in reduced glutathione (GSH) and GST levels in patients with different liver diseases. Overall, 114 patients were enrolled in this study: 58 patients with colorectal cancer (18 without and 40 with liver metastases), 27 with liver steatosis, 29 with alcoholic cirrhosis and a group of 40 healthy volunteers. The levels of GSH and GST in blood serum were evaluated by enzyme-linked immunosorbent assay (ELISA) according to the manufacturer's guidelines. Significant differences in GSH and GST levels were observed in most of the groups compared to the healthy volunteers (GSH: 52.72 µg/mL, GST: 0.53 ng/mL): with hepatic steatosis (GSH: 17.04 µg/mL, p < 0.001; GST: 5.89 ng/mL, p < 0.001), alcoholic cirrhosis (GSH: 62.04 µg/mL, p < 0.003; GST: 0.94 ng/mL, p < 0.001) and liver metastases (GSH: 37.84 µg/mL, p < 0.001, GST: 1.25 ng/mL, p=0.747). The different GSH and GST levels in patients with colorectal cancer liver metastases, liver steatosis and alcoholic cirrhosis indicate the differences in antioxidative system damage and its compensatory possibilities and could serve as potential biomarkers for its correction.

Phi Class of Glutathione S-transferase Gene Superfamily Widely Exists in Nonplant Taxonomic Groups.

PubMed

Munyampundu, Jean-Pierre; Xu, You-Ping; Cai, Xin-Zhong

2016-01-01

Glutathione S-transferases (GSTs) constitute a superfamily of enzymes involved in detoxification of noxious compounds and protection against oxidative damage. GST class Phi (GSTF), one of the important classes of plant GSTs, has long been considered as plant specific but was recently found in basidiomycete fungi. However, the range of nonplant taxonomic groups containing GSTFs remains unknown. In this study, the distribution and phylogenetic relationships of nonplant GSTFs were investigated. We identified GSTFs in ascomycete fungi, myxobacteria, and protists Naegleria gruberi and Aureococcus anophagefferens. GSTF occurrence in these bacteria and protists correlated with their genome sizes and habitats. While this link was missing across ascomycetes, the distribution and abundance of GSTFs among ascomycete genomes could be associated with their lifestyles to some extent. Sequence comparison, gene structure, and phylogenetic analyses indicated divergence among nonplant GSTFs, suggesting polyphyletic origins during evolution. Furthermore, in silico prediction of functional partners suggested functional diversification among nonplant GSTFs.

Phi Class of Glutathione S-transferase Gene Superfamily Widely Exists in Nonplant Taxonomic Groups

PubMed Central

Munyampundu, Jean-Pierre; Xu, You-Ping; Cai, Xin-Zhong

2016-01-01

Glutathione S-transferases (GSTs) constitute a superfamily of enzymes involved in detoxification of noxious compounds and protection against oxidative damage. GST class Phi (GSTF), one of the important classes of plant GSTs, has long been considered as plant specific but was recently found in basidiomycete fungi. However, the range of nonplant taxonomic groups containing GSTFs remains unknown. In this study, the distribution and phylogenetic relationships of nonplant GSTFs were investigated. We identified GSTFs in ascomycete fungi, myxobacteria, and protists Naegleria gruberi and Aureococcus anophagefferens. GSTF occurrence in these bacteria and protists correlated with their genome sizes and habitats. While this link was missing across ascomycetes, the distribution and abundance of GSTFs among ascomycete genomes could be associated with their lifestyles to some extent. Sequence comparison, gene structure, and phylogenetic analyses indicated divergence among nonplant GSTFs, suggesting polyphyletic origins during evolution. Furthermore, in silico prediction of functional partners suggested functional diversification among nonplant GSTFs. PMID:26884677

Rapid accumulation of glutathione during light stress in Arabidopsis.

PubMed

Choudhury, Feroza K; Devireddy, Amith R; Azad, Rajeev K; Shulaev, Vladimir; Mittler, Ron

2018-05-25

Environmental stress conditions can drastically affect plant growth and productivity. In contrast to soil moisture or salinity that can gradually change over a period of days or weeks, changes in light intensity or temperature can occur very rapidly, sometimes over the course of minutes or seconds. We previously reported that in response to rapid changes in light intensity (0-60 sec), Arabidopsis thaliana plants mount a large-scale transcriptomic response that includes several different transcripts essential for light stress acclimation. Here, we expand our analysis of the rapid response of Arabidopsis to light stress using a metabolomics approach and identify 111 metabolites that significantly alter in their level during the first 90 sec of light stress exposure. We further show that the levels of free and total glutathione accumulate rapidly during light stress in Arabidopsis and that the accumulation of total glutathione during light stress is associated with an increase in nitric oxide (NO) levels. We further suggest that the increase in precursors for glutathione biosynthesis could be linked to alterations in photorespiration, and that phosphoenolpyruvate could represent a major energy and carbon source for rapid metabolic responses. Taken together, our analysis could be used as an initial road map for the identification of different pathways that could be used to augment the rapid response of plants to abiotic stress. In addition, it highlights the important role of glutathione in these responses.

A mitochondria-targeted turn-on fluorescent probe for the detection of glutathione in living cells.

PubMed

Zhang, Jian; Bao, Xiaolong; Zhou, Junliang; Peng, Fangfang; Ren, Hang; Dong, Xiaochun; Zhao, Weili

2016-11-15

A novel turn-on red fluorescent BODIPY-based probe (Probe 1) for the detection of glutathione was developed. Such a probe carries a para-dinitrophenoxy benzyl pyridinium moiety at the meso position of a BODIPY dye as self-immolative linker. Probe 1 responds selectively to glutathione with the detection limit of 109nM over other amino acids, common metal ions, reactive oxygen species, reactive nitrogen species, and reactive sulfur species. A novel electrostatic interaction to modulate the SNAr attack of glutathione was believed to play significant role for the observed selective response to glutathione. The cleavage of dinitrophenyl ether by glutathione leads to the production of para-hydroxybenzyl moiety which is able to self-immolate through an intramolecular 1,4-elimination reaction to release the fluorescent BODIPY dye. The low toxic probe has been successfully used to detect mitochondrial glutathione in living cells. Copyright © 2016 Elsevier B.V. All rights reserved.

Glutathione and zebrafish: Old assays to address a current issue.

PubMed

Massarsky, Andrey; Kozal, Jordan S; Di Giulio, Richard T

2017-02-01

Several xenobiotic agents (e.g. metals, polycyclic aromatic hydrocarbons, nanoparticles, etc.) commonly involve the generation of reactive oxygen species (ROS) and oxidative stress as part of their toxic mode of action. Among piscine models, the zebrafish is a popular vertebrate model to study toxicity of various xenobiotic agents. Similarly to other vertebrates, zebrafish possess an extensive antioxidant system, including the reduced form of glutathione (GSH), which is an important antioxidant that acts alone or in conjunction with enzymes, such as glutathione peroxidase (GPx). Upon interaction with ROS, GSH is oxidized, resulting in the formation of glutathione disulfide (GSSG). GSSG is recycled by an auxiliary antioxidant enzyme glutathione reductase (GR). This article outlines detailed methods to measure the concentrations of GSH and GSSG, as well as the activities of GPx and GR in zebrafish larvae as robust and economical means to assess oxidative stress. The studies that have assessed these endpoints in zebrafish and alternative methods are also discussed. We conclude that the availability of these robust and economical methods support the use of zebrafish as a model organism in studies evaluating redox biology, as well as the induction of oxidative stress following exposure to toxic agents. Copyright © 2016 Elsevier Ltd. All rights reserved.

Glutathione and redox signaling in substance abuse.

PubMed

Uys, Joachim D; Mulholland, Patrick J; Townsend, Danyelle M

2014-07-01

Throughout the last couple decades, the cause and consequences of substance abuse has expanded to identify the underlying neurobiological signaling mechanisms associated with addictive behavior. Chronic use of drugs, such as cocaine, methamphetamine and alcohol leads to the formation of oxidative or nitrosative stress (ROS/RNS) and changes in glutathione and redox homeostasis. Of importance, redox-sensitive post-translational modifications on cysteine residues, such as S-glutathionylation and S-nitrosylation could impact on the structure and function of addiction related signaling proteins. In this commentary, we evaluate the role of glutathione and redox signaling in cocaine-, methamphetamine- and alcohol addiction and conclude by discussing the possibility of targeting redox pathways for the therapeutic intervention of these substance abuse disorders. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Investigation of electroacupuncture and manual acupuncture on carnitine and glutathione in muscle.

PubMed

Toda, Shizuo

2011-01-01

Electroacupuncture (EA) and manual acupuncture (MA) have therapeutic effects on muscle fatigue in muscle disease. The deficiencies of carnitine and glutathione induce muscle fatigue. This report investigated the effects of EA and MA on carnitine and glutathione in muscle. After the mice of EA group were fixed in the animal cage, right Zusanli (ST36) and Jiexi (ST41) were acupunctured and stimulated with uniform reinforcing and reducing method by twirling the acupuncture needle for 15 min. And then, the needle handles were connected to an electric stimulator for stimulating the acupoint with dense-sparse waves. After the mice of MA group were fixed in an animal cage, right ST36 and ST41 were acupunctured and allowed for 15 min. The mice of normal control group were not acupunctured and stimulated for 15 min. The mice of all groups were killed for collecting muscle tissue 1 h after the final treatment. Carnitine and glutathione in homogenate of muscle tissue were determined with carnitine (Kainos Laboratories Co., Tokyo, Japan) and glutathione assay kit (Dojin Chemicals Co., Kumamoto, Japan). Carnitine level in muscle tissue of MA group was significantly higher than those of EA group and normal control group. Carnitine level in muscle tissue of EA group was not significantly different from that of normal control group. Glutathione levels in muscle tissue of EA group and MA group were significantly higher than that of normal control group. This report presented that carnitine in muscle is increased by MA, and not increased by EA, and that glutathione in muscle is increased by EA and MA.

Chloride-associated adaptive response in aerobic methylotrophic dichloromethane-utilising bacteria.

PubMed

Torgonskaya, Maria L; Doronina, Nina V; Hourcade, Edith; Trotsenko, Yuri A; Vuilleumier, Stéphane

2011-06-01

Aerobic methylotrophic bacteria able to grow with dichloromethane (DCM) as the sole carbon and energy source possess a specific glutathione S-transferase, DCM dehalogenase, which transforms DCM to formaldehyde, used for biomass and energy production, and hydrochloric acid, which is excreted. Evidence is presented for chloride-specific responses for three DCM-degrading bacteria, Methylobacterium extorquens DM4, Methylopila helvetica DM6 and Albibacter methylovorans DM10. Chloride release into the medium was inhibited by sodium azide and m -chlorophenylhydrazone, suggesting an energy-dependent process. In contrast, only nigericin affected chloride excretion in Mb. extorquens DM4 and Mp. helvetica DM6, while valinomycin had the same effect in A. methylovorans DM10 only. Chloride ions stimulated DCM-dependent induction of DCM dehalogenase expression for Mp. helvetica DM6 and A. methylovorans DM10, and shortened the time for onset of chloride release into the medium. Striking chloride-containing structures were observed by electron microscopy and X-ray microanalysis on the cell surface of Mp. helvetica DM6 and A. methylovorans DM10 during growth with DCM, and with methanol in medium supplemented with sodium chloride. Taken together, these data suggest the existence of both general and specific chloride-associated adaptations in aerobic DCM-degrading bacteria. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Enzyme activity of α-chymotrypsin: Deactivation by gold nano-cluster and reactivation by glutathione.

PubMed

Ghosh, Catherine; Mondal, Tridib; Bhattacharyya, Kankan

2017-05-15

Effect of gold nanoclusters (Au-NCs) on the circular dichroism (CD) spectra and enzymatic activity of α-chymotrypsin (ChT) (towards hydrolysis of a substrate, N-succinyl-l-phenylalanine p-nitroanilide) are studied. The CD spectra indicate that on binding to Au-NC, ChT is completely unfolded, resulting in nearly zero ellipticity. α-chymotrypsin (ChT) coated gold nano-clusters exhibit almost no enzymatic activity. Addition of glutathione (GSH) or oxidized glutathione (GSSG) restore the enzyme activity of α-chymotrypsin by 30-45%. ChT coated Au-NC exhibits two emission maxima-one at 480nm (corresponding to Au 10 ) and one at 640nm (Au 25 ). On addition of glutathione (GSH) or oxidized glutathione (GSSG) the emission peak at 640nm vanishes and only one peak at 480nm (Au 10 ) remains. MALDI mass spectrometry studies suggest addition of glutathione (GSH) to α-chymotrypsin capped Au-NCs results in the formation of glutathione-capped Au-NCs and α-chymotrypsin is released from Au-NCs. CD spectroscopy indicates that the conformation of the released α-chymotrypsin is different from that of the native α-chymotrypsin. Copyright © 2017 Elsevier Inc. All rights reserved.

Sulfur partitioning between glutathione and protein synthesis determines plant growth.

PubMed

Speiser, Anna; Silbermann, Marleen; Dong, Yihan; Haberland, Stefan; Uslu, Veli Vural; Wang, Shanshan; Bangash, Sajid A K; Reichelt, Michael; Meyer, Andreas J; Wirtz, Markus; Hell, Ruediger

2018-05-11

Photoautotrophic organisms must efficiently allocate their resources between stress-response pathways and growth-promoting pathways to be successful in a constantly changing environment. In this study, we addressed the coordination of sulfur flux between biosynthesis of the reactive oxygen species scavenger glutathione (GSH) and protein translation as one example of a central resource allocation switch. We crossed the Arabidopsis (Arabidopsis thaliana) GSH synthesis depleted cadmium-sensitive cad2-1 mutant, which lacks glutamate Cys ligase, into the sulfite reductase (SiR) sir1-1 mutant, which suffers from a significantly decreased flux of sulfur into cysteine and is consequently retarded in growth. Surprisingly, depletion of GSH synthesis promoted the growth of the sir1-1 cad2-1 double mutant (s1c2) when compared to sir1-1. Determination of GSH levels and in vivo live-cell imaging of the reduction-oxidation sensitive green fluorescent protein (roGFP2)-sensor demonstrated significant oxidation of the plastidic GSH redox-potential in cad2-1 and s1c2. This oxidized GSH redox-potential aligned with significant activation of plastid-localized sulfate reduction and a significantly higher flux of sulfur into proteins. The specific activation of the serine/threonine sensor kinase Target Of Rapamycin (TOR) in cad2-1 and s1c2 was the trigger for reallocation of cysteine from GSH biosynthesis into protein translation. Activation of TOR in s1c2 enhanced ribosome abundance and partially rescued the decreased meristematic activity observed in sir1-1 mutants. Therefore, we found that the coordination of sulfur flux between glutathione biosynthesis and protein translation determines growth via regulation of TOR. {copyright, serif} 2018 American Society of Plant Biologists. All rights reserved.

Variable Levels of Glutathione S-Transferases Are Responsible for the Differential Tolerance to Metolachlor between Maize (Zea mays) Shoots and Roots.

PubMed

Li, Dongzhi; Xu, Li; Pang, Sen; Liu, Zhiqian; Wang, Kai; Wang, Chengju

2017-01-11

Glutathione S-transferases (GSTs) play important roles in herbicide tolerance. However, studies on GST function in herbicide tolerance among plant tissues are still lacking. To explore the mechanism of metolachlor tolerance difference between maize shoots and roots, the effects of metolachlor on growth, GST activity, and the expression of the entire GST gene family were investigated. It was found that this differential tolerance to metolachlor was correlated with contrasting GST activity between the two tissues and can be eliminated by a GST inhibitor. An in vitro metolachlor-glutathione conjugation assay confirmed that the transformation of metolachlor is 2-fold faster in roots than in shoots. The expression analysis of the GST gene family revealed that most GST genes are expressed much higher in roots than shoots, both in control and in metolachlor-treated plants. Taken together, higher level expression of most GST genes, leading to higher GST activity and faster herbicide transformation, appears to be responsible for the higher tolerance to metolachlor of maize roots than shoots.

CHARACTERIZATION OF DANSYLATED CYSTEINE, CYSTINE, GLUTATHIONE, AND GLUTATHIONE DISULFIDE BY NARROW BORE LIQUID CHROMATOGRAPHY - ELECTROSPRAY IONIZATION MASS SPECTROMETRY

EPA Science Inventory

A method using reversed phase high performance liquid chromtography/electrospray ionization-mass spectrometry (RP-LC/ESI-MS) has been developed to confirm the dientity of dansylated derivatives of cysteine (C) and glutathione (GSH), and their respective dimers, cystine (CSSC) and...

Fast and Selective Two-Stage Ratiometric Fluorescent Probes for Imaging of Glutathione in Living Cells.

PubMed

Gong, Deyan; Han, Shi-Chong; Iqbal, Anam; Qian, Jing; Cao, Ting; Liu, Wei; Liu, Weisheng; Qin, Wenwu; Guo, Huichen

2017-12-19

Two fluorescent, m-nitrophenol-substituted difluoroboron dipyrromethene dyes have been designed by nucleophilic substitution reaction of 3,5-dichloro-4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY). Nonsymmetric and symmetric probes, that is. BODIPY 1 (with one nitrophenol group at the position 3) and BODIPY 2 (with two nitrophenol groups at the positions 3 and 5) were applied to ratiometric fluorescent glutathione detection. The detection is based on the two-step nucleophilic aromatic substitution of the nitrophenol groups of the probes by glutathione in buffer solution containing CTAB. In the first stage, probe 1 showed ratiometric fluorescent color change from green (λ em = 530 nm) to yellow (λ em = 561 nm) because of monosubstitution with glutathione (I 561nm /I 530nm ). Addition of excess glutathione caused the second stage of ratiometric fluorescent color change from yellow to reddish orange (λ em = 596 nm, I 596nm /I 561nm ) due to disubstitution with glutathione. Therefore, different concentration ranges of glutathione (from less to excess) could be rapidly detected by the two-stage ratiometric fluorescent probe 1 in 5 min. While, probe 2 shows single-stage ratiometric fluorescent detection to GSH (from green to reddish orange, I 596nm /I 535nm ). Probes 1 and 2 exhibit excellent properties with sensitive, specific colorimetric response and ratiometric fluorescent response to glutathione over other sulfur nucleophiles. Application to cellular ratiometric fluorescence imaging indicated that the probes were highly responsive to intracellular glutathione.

Chemotherapy triggers HIF-1–dependent glutathione synthesis and copper chelation that induces the breast cancer stem cell phenotype

PubMed Central

Lu, Haiquan; Samanta, Debangshu; Xiang, Lisha; Zhang, Huimin; Hu, Hongxia; Chen, Ivan; Bullen, John W.; Semenza, Gregg L.

2015-01-01

Triple negative breast cancer (TNBC) accounts for 10–15% of all breast cancer but is responsible for a disproportionate share of morbidity and mortality because of its aggressive characteristics and lack of targeted therapies. Chemotherapy induces enrichment of breast cancer stem cells (BCSCs), which are responsible for tumor recurrence and metastasis. Here, we demonstrate that chemotherapy induces the expression of the cystine transporter xCT and the regulatory subunit of glutamate-cysteine ligase (GCLM) in a hypoxia-inducible factor (HIF)-1–dependent manner, leading to increased intracellular glutathione levels, which inhibit mitogen-activated protein kinase kinase (MEK) activity through copper chelation. Loss of MEK-ERK signaling causes FoxO3 nuclear translocation and transcriptional activation of the gene encoding the pluripotency factor Nanog, which is required for enrichment of BCSCs. Inhibition of xCT, GCLM, FoxO3, or Nanog blocks chemotherapy-induced enrichment of BCSCs and impairs tumor initiation. These results suggest that, in combination with chemotherapy, targeting BCSCs by inhibiting HIF-1–regulated glutathione synthesis may improve outcome in TNBC. PMID:26229077

Fluorescence detection of glutathione and oxidized glutathione in blood with a NIR-excitable cyanine probe

NASA Astrophysics Data System (ADS)

Liu, Chang-hui; Qi, Feng-pei; Wen, Fu-bin; Long, Li-ping; Liu, Ai-juan; Yang, Rong-hua

2018-04-01

Cyanine has been widely utilized as a near infrared (NIR) fluorophore for detection of glutathione (GSH). However, the excitation of most of the reported cyanine-based probes was less than 800 nm, which inevitably induce biological background absorption and lower the sensitivity, limiting their use for detection of GSH in blood samples. To address this issue, here, a heptamethine cyanine probe (DNIR), with a NIR excitation wavelength at 804 nm and a NIR emission wavelength at 832 nm, is employed for the detection of GSH and its oxidized form (GSSG) in blood. The probe displays excellent selectivity for GSH over GSSG and other amino acids, and rapid response to GSH, in particular a good property for indirect detection of GSSG in the presence of enzyme glutathione reductase and the reducing agent nicotinamideadenine dinucleotide phosphate, without further separation prior to fluorescent measurement. To the best of our knowledge, this is the first attempt to explore NIR fluorescent approach for the simultaneous assay of GSH and GSSG in blood. As such, we expect that our fluorescence sensors with both NIR excitation and NIR emission make this strategy suitable for the application in complex physiological systems.

High performance liquid chromatographic assay for the quantitation of total glutathione in plasma

NASA Technical Reports Server (NTRS)

Abukhalaf, Imad K.; Silvestrov, Natalia A.; Menter, Julian M.; von Deutsch, Daniel A.; Bayorh, Mohamed A.; Socci, Robin R.; Ganafa, Agaba A.

2002-01-01

A simple and widely used homocysteine HPLC procedure was applied for the HPLC identification and quantitation of glutathione in plasma. The method, which utilizes SBDF as a derivatizing agent utilizes only 50 microl of sample volume. Linear quantitative response curve was generated for glutathione over a concentration range of 0.3125-62.50 micromol/l. Linear regression analysis of the standard curve exhibited correlation coefficient of 0.999. Limit of detection (LOD) and limit of quantitation (LOQ) values were 5.0 and 15 pmol, respectively. Glutathione recovery using this method was nearly complete (above 96%). Intra-assay and inter-assay precision studies reflected a high level of reliability and reproducibility of the method. The applicability of the method for the quantitation of glutathione was demonstrated successfully using human and rat plasma samples.

Detection of Glutathione by Glutathione-S-Transferase-Nanoconjugate Ensemble Electrochemical Device.

PubMed

Barman, Ujjwol; Mukhopadhyay, Gargi; Goswami, Namami; Ghosh, Siddhartha Sankar; Paily, Roy P

2017-06-01

This paper reports a novel electrochemical method for detection of Glutathione (GSH) using Glutathione-S-Transferase (GST) - ZnO composite nanoparticles to investigate the prospects of the method for detection of cancer at an early stage. The purified GST enzyme was bound with ZnO nanoparticles by electrostatic interactions and the nanocomposite was dropcast on a silicon dioxide wafer. The GST functionalized deposited layer was then used as a chemiresistive channel to detect conjugation reaction between GSH and 1-Chloro-2, 4-Dinitrobenzene (CDNB). The zeta potential values of the ZnO nanoparticles and the GST were found to be 13.4 mV and-6.21 mV, respectively. Around 73.8% binding was observed between the enzyme and ZnO nanoparticles. I - V analysis of the chemiresistive channel showed an increase in conductivity of the channel due to conjugation reaction between GSH and CDNB as compared with that of GSH or CDNB alone. I - V characterization of the GST functionalized layer was performed at various concentrations of GSH and a sensitivity and limit of detection of 5.68 nA/ [Formula: see text] and 41.9 nM were obtained, respectively. Thus from I - V analysis of the chemiresistivechannel, the detectionand quantification of GSH could be obtained. The kinetic parameters of both GST and nanoconjugate of ZnO nanoparticles andGSTwere determinedwith respect to its substrates, GSH and CDNB, using Michaelis-Mentenmodel. This novel approach of detection of GSH bymeans of ZnO nanoparticle and GST enzyme composite can be further analyzed for in vitro experiments, which will lead us to a new and efficient way of detecting certain types of cancers at an early stage.

Elevated oxidized glutathione in cystinotic proximal tubular epithelial cells.

PubMed

Wilmer, Martijn J G; de Graaf-Hess, Adriana; Blom, Henk J; Dijkman, Henry B P M; Monnens, Leo A; van den Heuvel, Lambertus P; Levtchenko, Elena N

2005-11-18

Cystinosis, the most frequent cause of inborn Fanconi syndrome, is characterized by the lysosomal cystine accumulation, caused by mutations in the CTNS gene. To elucidate the pathogenesis of cystinosis, we cultured proximal tubular cells from urine of cystinotic patients (n = 9) and healthy controls (n = 9), followed by immortalization with human papilloma virus (HPV E6/E7). Obtained cell lines displayed basolateral polarization, alkaline phosphatase activity, and presence of aminopeptidase N (CD-13) and megalin, confirming their proximal tubular origin. Cystinotic cell lines exhibited elevated cystine levels (0.86 +/- 0.95 nmol/mg versus 0.09 +/- 0.01 nmol/mg protein in controls, p = 0.03). Oxidized glutathione was elevated in cystinotic cells (1.16 +/- 0.83 nmol/mg versus 0.29 +/- 0.18 nmol/mg protein, p = 0.04), while total glutathione, free cysteine, and ATP contents were normal in these cells. In conclusion, elevated oxidized glutathione in cystinotic proximal tubular epithelial cell lines suggests increased oxidative stress, which may contribute to tubular dysfunction in cystinosis.

Identification of residues critical for proton-coupled glutathione translocation in the yeast glutathione transporter, Hgt1p.

PubMed

Zulkifli, Mohammad; Bachhawat, Anand Kumar

2017-05-16

The proton gradient acts as the driving force for the transport of many metabolites across fungal and plant plasma membranes. Identifying the mechanism of proton relay is critical for understanding the mechanism of transport mediated by these transporters. We investigated two strategies for identifying residues critical for proton-dependent substrate transport in the yeast glutathione transporter, Hgt1p, a member of the poorly understood oligopeptide transporter family of transporters. In the first strategy, we tried to identify the pH-independent mutants that could grow at higher pH when dependant on glutathione transport. Screening a library of 269 alanine mutants of the transmembrane domains (TMDs) along with a random mutagenesis strategy yielded two residues (E135K on the cusp of TMD2 and N710S on TMD12) that permitted growth on glutathione at pH 8.0. Further analysis revealed that these residues were not involved in proton symport even though they conferred better transport at a higher pH. The second strategy involved a knowledge-driven approach, targeting 31 potential residues based on charge, conservation and location. Mutation of these residues followed by functional and biochemical characterization revealed E177A, Y193A, D335A, Y374A, H445A and R554A as being defective in proton transport. Further analysis enabled possible roles of these residues to be assigned in proton relay. The implications of these findings in relation to Hgt1p and the suitability of these strategic approaches for identifying such residues are discussed. © 2017 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

"Use of acidophilic bacteria of the genus Acidithiobacillus to biosynthesize CdS fluorescent nanoparticles (quantum dots) with high tolerance to acidic pH".

PubMed

Ulloa, G; Collao, B; Araneda, M; Escobar, B; Álvarez, S; Bravo, D; Pérez-Donoso, J M

2016-12-01

The use of bacterial cells to produce fluorescent semiconductor nanoparticles (quantum dots, QDs) represents a green alternative with promising economic potential. In the present work, we report for the first time the biosynthesis of CdS QDs by acidophilic bacteria of the Acidithiobacillus genus. CdS QDs were obtained by exposing A. ferrooxidans, A. thiooxidans and A. caldus cells to sublethal Cd 2+ concentrations in the presence of cysteine and glutathione. The fluorescence of cadmium-exposed cells moves from green to red with incubation time, a characteristic property of QDs associated with nanocrystals growth. Biosynthesized nanoparticles (NPs) display an absorption peak at 360nm and a broad emission spectra between 450 and 650nm when excited at 370nm, both characteristic of CdS QDs. Average sizes of 6 and 10nm were determined for green and red NPs, respectively. The importance of cysteine and glutathione on QDs biosynthesis in Acidithiobacillus was related with the generation of H 2 S. Interestingly, QDs produced by acidophilic bacteria display high tolerance to acidic pH. Absorbance and fluorescence properties of QDs was not affected at pH 2.0, a condition that totally inhibits the fluorescence of QDs produced chemically or biosynthesized by mesophilic bacteria (stable until pH 4.5-5.0). Results presented here constitute the first report of the generation of QDs with improved properties by using extremophile microorganisms. Copyright © 2016 Elsevier Inc. All rights reserved.

Concomitant ingestion of lactic acid bacteria and black tea synergistically enhances flavonoid bioavailability and attenuates d-galactose-induced oxidative stress in mice via modulating glutathione antioxidant system.

PubMed

Zhao, Danyue; Shah, Nagendra P

2016-12-01

Black tea (BT) has been positively linked to improved redox status, while its efficacy is limited due to the low bioavailability of BT flavonoids. In addition to the direct antioxidant activity, flavonoids regulate redox balance via inducing endogenous antioxidants, particularly glutathione (GSH) and GSH-dependent antioxidant enzymes. This work first examined the effect of lactic acid bacteria (LAB) and BT alone or in combination on flavonoid bioavailability and metabolism; next, the effect of LAB-fermented BT diet in attenuating oxidative stress in mice and the underlying mechanisms were studied. Phenolic profiles of plasma, urine and feces from healthy mice consuming plain yogurt, BT milk (BTM) or BT yogurt (BTY) were acquired using LC-MS/MS. Plasma antioxidant capacity, lipid peroxidation level, content of nonprotein thiols and expression of GSH-related antioxidant enzymes and Nrf2 were examined in d-galactose-treated mice. Total flavonoid content in plasma following a single dose of BTY attained 0.657 μmol/l, increased by 50% compared with the BTM group. Increased excretion of phenolic metabolite and hippuric acid in urine and feces indicated enhanced metabolism of flavonoids in BTY-fed mice. In the second study, 8-week concomitant LAB-BT treatment of oxidatively stressed mice effectively restored plasma antioxidant capacity and GSH levels, and mitigated lipid peroxidation, which were associated with significant induction of GSH-dependent antioxidant enzymes and nuclear accumulation of Nrf2. Our results demonstrated the effect of LAB fermentation in enhancing BT flavonoid bioavailability in vivo. The synergistic antioxidant efficacy of LAB-BT diet implied its therapeutic potential in enhancing antioxidant defenses and protecting organisms from oxidative damage. Copyright © 2016. Published by Elsevier Inc.

Glutathione redox regulates airway hyperresponsiveness and airway inflammation in mice.

PubMed

Koike, Yoko; Hisada, Takeshi; Utsugi, Mitsuyoshi; Ishizuka, Tamotsu; Shimizu, Yasuo; Ono, Akihiro; Murata, Yukie; Hamuro, Junji; Mori, Masatomo; Dobashi, Kunio

2007-09-01

Glutathione is the major intracellular redox buffer. We have shown that glutathione redox status, which is the balance between intracellular reduced (GSH) and oxidized (GSSG) glutathione, in antigen-presenting cells (APC) regulates the helper T cell type 1 (Th1)/Th2 balance due to the production of IL-12. Bronchial asthma is a typical Th2 disease. Th2 cells and Th2 cytokines are characteristic of asthma and trigger off an inflammation. Accordingly, we studied the effects of the intracellular glutathione redox status on airway hyperresponsiveness (AHR) and allergen-induced airway inflammation in a mouse model of asthma. We used gamma-Glutamylcysteinylethyl ester (gamma-GCE), which is a membrane-permeating GSH precursor, to elevate the intracellular GSH level and GSH/GSSG ratio of mice. In vitro, gamma-GCE pretreatment of human monocytic THP-1 cells elevated the GSH/GSSG ratio and enhanced IL-12(p70) production induced by LPS. In the mouse asthma model, intraperitoneal injection of gamma-GCE elevated the GSH/GSSG ratio of lung tissue and reduced AHR. gamma-GCE reduced levels of IL-4, IL-5, IL-10, and the chemokines eotaxin and RANTES (regulated on activation, normal T cell expressed and secreted) in bronchoalveolar lavage fluid, whereas it enhanced the production of IL-12 and IFN-gamma. Histologically, gamma-GCE suppressed eosinophils infiltration. Interestingly, we also found that gamma-GCE directly inhibited chemokine-induced eosinophil chemotaxis without affecting eotaxin receptor chemokine receptor 3 (CCR3) expressions. Taken together, these findings suggest that changing glutathione redox balance, increase in GSH level, and the GSH/GSSG ratio by gamma-GCE, ameliorate bronchial asthma by altering the Th1/Th2 imbalance through IL-12 production from APC and suppressing chemokine production and eosinophil migration itself.

Prolonged fasting increases glutathione biosynthesis in postweaned northern elephant seals.

PubMed

Vázquez-Medina, José Pablo; Zenteno-Savín, Tania; Forman, Henry Jay; Crocker, Daniel E; Ortiz, Rudy M

2011-04-15

Northern elephant seals experience prolonged periods of absolute food and water deprivation (fasting) while breeding, molting or weaning. The postweaning fast in elephant seals is characterized by increases in the renin-angiotensin system, expression of the oxidant-producing protein Nox4, and NADPH oxidase activity; however, these increases are not correlated with increased oxidative damage or inflammation. Glutathione (GSH) is a potent reductant and a cofactor for glutathione peroxidases (GPx), glutathione-S transferases (GST) and 1-cys peroxiredoxin (PrxVI) and thus contributes to the removal of hydroperoxides, preventing oxidative damage. The effects of prolonged food deprivation on the GSH system are not well described in mammals. To test our hypothesis that GSH biosynthesis increases with fasting in postweaned elephant seals, we measured circulating and muscle GSH content at the early and late phases of the postweaning fast in elephant seals along with the activity/protein content of glutamate-cysteine ligase [GCL; catalytic (GCLc) and modulatory (GCLm) subunits], γ-glutamyl transpeptidase (GGT), glutathione disulphide reductase (GR), glucose-6-phosphate dehydrogenase (G6PDH), GST and PrxVI, as well as plasma changes in γ-glutamyl amino acids, glutamate and glutamine. GSH increased two- to four-fold with fasting along with a 40-50% increase in the content of GCLm and GCLc, a 75% increase in GGT activity, a two- to 2.5-fold increase in GR, G6PDH and GST activities and a 30% increase in PrxVI content. Plasma γ-glutamyl glutamine, γ-glutamyl isoleucine and γ-glutamyl methionine also increased with fasting whereas glutamate and glutamine decreased. Results indicate that GSH biosynthesis increases with fasting and that GSH contributes to counteracting hydroperoxide production, preventing oxidative damage in fasting seals.

Prolonged fasting increases glutathione biosynthesis in postweaned northern elephant seals

PubMed Central

Vázquez-Medina, José Pablo; Zenteno-Savín, Tania; Forman, Henry Jay; Crocker, Daniel E.; Ortiz, Rudy M.

2011-01-01

SUMMARY Northern elephant seals experience prolonged periods of absolute food and water deprivation (fasting) while breeding, molting or weaning. The postweaning fast in elephant seals is characterized by increases in the renin–angiotensin system, expression of the oxidant-producing protein Nox4, and NADPH oxidase activity; however, these increases are not correlated with increased oxidative damage or inflammation. Glutathione (GSH) is a potent reductant and a cofactor for glutathione peroxidases (GPx), glutathione-S transferases (GST) and 1-cys peroxiredoxin (PrxVI) and thus contributes to the removal of hydroperoxides, preventing oxidative damage. The effects of prolonged food deprivation on the GSH system are not well described in mammals. To test our hypothesis that GSH biosynthesis increases with fasting in postweaned elephant seals, we measured circulating and muscle GSH content at the early and late phases of the postweaning fast in elephant seals along with the activity/protein content of glutamate-cysteine ligase [GCL; catalytic (GCLc) and modulatory (GCLm) subunits], γ-glutamyl transpeptidase (GGT), glutathione disulphide reductase (GR), glucose-6-phosphate dehydrogenase (G6PDH), GST and PrxVI, as well as plasma changes in γ-glutamyl amino acids, glutamate and glutamine. GSH increased two- to four-fold with fasting along with a 40–50% increase in the content of GCLm and GCLc, a 75% increase in GGT activity, a two- to 2.5-fold increase in GR, G6PDH and GST activities and a 30% increase in PrxVI content. Plasma γ-glutamyl glutamine, γ-glutamyl isoleucine and γ-glutamyl methionine also increased with fasting whereas glutamate and glutamine decreased. Results indicate that GSH biosynthesis increases with fasting and that GSH contributes to counteracting hydroperoxide production, preventing oxidative damage in fasting seals. PMID:21430206

Occurrence of low molecular weight thiols in biological systems

NASA Technical Reports Server (NTRS)

Fahey, Robert C.; Newton, Gerald L.

1983-01-01

Bromobimane labeling and high performance chromatography analysis were applied to various species of bacteria, plant tissues, and animal tissues. The reaction between thiols and monobromobimane is studied. Chromatograms revealing peaks produced by nonthiols and thiols are analyzed and compared. It is observed that all the bacteria species contain hydrogen sulfide, and glutathione is contained in facultative and aerobic gram-negative bacteria. For the plant tissues, the data reveal that mung bean sprouts contain homoglutathione and no glutathione; alfalfa sprouts contain homoglutathione and glutathione; the pea seed, nonlegumes, and fungi contain glutathione and no homoglutathione. It is detected that the main thiol in the animal tissues is glutathione. Based on the data, it is suggested that glutathione has an essential function in higher organisms.

Investigation of Electroacupuncture and Manual Acupuncture on Carnitine and Glutathione in Muscle

PubMed Central

Toda, Shizuo

2011-01-01

Electroacupuncture (EA) and manual acupuncture (MA) have therapeutic effects on muscle fatigue in muscle disease. The deficiencies of carnitine and glutathione induce muscle fatigue. This report investigated the effects of EA and MA on carnitine and glutathione in muscle. After the mice of EA group were fixed in the animal cage, right Zusanli (ST36) and Jiexi (ST41) were acupunctured and stimulated with uniform reinforcing and reducing method by twirling the acupuncture needle for 15 min. And then, the needle handles were connected to an electric stimulator for stimulating the acupoint with dense-sparse waves. After the mice of MA group were fixed in an animal cage, right ST36 and ST41 were acupunctured and allowed for 15 min. The mice of normal control group were not acupunctured and stimulated for 15 min. The mice of all groups were killed for collecting muscle tissue 1 h after the final treatment. Carnitine and glutathione in homogenate of muscle tissue were determined with carnitine (Kainos Laboratories Co., Tokyo, Japan) and glutathione assay kit (Dojin Chemicals Co., Kumamoto, Japan). Carnitine level in muscle tissue of MA group was significantly higher than those of EA group and normal control group. Carnitine level in muscle tissue of EA group was not significantly different from that of normal control group. Glutathione levels in muscle tissue of EA group and MA group were significantly higher than that of normal control group. This report presented that carnitine in muscle is increased by MA, and not increased by EA, and that glutathione in muscle is increased by EA and MA. PMID:19592478

Scanning electrochemical microscopy of menadione-glutathione conjugate export from yeast cells

PubMed Central

Mauzeroll, Janine; Bard, Allen J.

2004-01-01

The uptake of menadione (2-methyl-1,4-naphthoquinone), which is toxic to yeast cells, and its expulsion as a glutathione complex were studied by scanning electrochemical microscopy. The progression of the in vitro reaction between menadione and glutathione was monitored electrochemically by cyclic voltammetry and correlated with the spectroscopic (UV–visible) behavior. By observing the scanning electrochemical microscope tip current of yeast cells suspended in a menadione-containing solution, the export of the conjugate from the cells with time could be measured. Similar experiments were performed on immobilized yeast cell aggregates stressed by a menadione solution. From the export of the menadione-glutathione conjugate detected at a 1-μm-diameter electrode situated 10 μm from the cells, a flux of about 30,000 thiodione molecules per second per cell was extracted. Numerical simulations based on an explicit finite difference method further revealed that the observation of a constant efflux of thiodione from the cells suggested the rate was limited by the uptake of menadione and that the efflux through the glutathione-conjugate pump was at least an order of magnitude faster. PMID:15148374

[35S]-LABELING OF THE SALMONELLA TYPHIMURIUM GLUTATHIONE POOL TO ASSESS GLUTATHIONE-MEDIATED DNA BINDING BY 1,2-DIBROMOETHANE

EPA Science Inventory

Biotransformation of drugs and environmental chemicals to reactive intermediates is often studied with the use of radiolabeled compounds that are synthesized by expensive and technically difficult procedures. In general, glutathione (GSH) conjugation serves as a detoxification m...

The diverse roles of glutathione-associated cell resistance against hypericin photodynamic therapy.

PubMed

Theodossiou, Theodossis A; Olsen, Cathrine E; Jonsson, Marte; Kubin, Andreas; Hothersall, John S; Berg, Kristian

2017-08-01

The diverse responses of different cancers to treatments such as photodynamic therapy of cancer (PDT) have fueled a growing need for reliable predictive markers for treatment outcome. In the present work we have studied the differential response of two phenotypically and genotypically different breast adenocarcinoma cell lines, MCF7 and MDA-MB-231, to hypericin PDT (HYP-PDT). MDA-MB-231 cells were 70% more sensitive to HYP PDT than MCF7 cells at LD 50 . MCF7 were found to express a substantially higher level of glutathione peroxidase (GPX4) than MDA-MB-231, while MDA-MB-231 differentially expressed glutathione-S-transferase (GSTP1), mainly used for xenobiotic detoxification. Eighty % reduction of intracellular glutathione (GSH) by buthionine sulfoximine (BSO), largely enhanced the sensitivity of the GSTP1 expressing MDA-MB-231 cells to HYP-PDT, but not in MCF7 cells. Further inhibition of the GSH reduction however by carmustine (BCNU) resulted in an enhanced sensitivity of MCF7 to HYP-PDT. HYP loading studies suggested that HYP can be a substrate of GSTP for GSH conjugation as BSO enhanced the cellular HYP accumulation by 20% in MDA-MB-231 cells, but not in MCF7 cells. Studies in solutions showed that L-cysteine can bind the GSTP substrate CDNB in the absence of GSTP. This means that the GSTP-lacking MCF7 may use L-cysteine for xenobiotic detoxification, especially during GSH synthesis inhibition, which leads to L-cysteine build-up. This was confirmed by the lowered accumulation of HYP in both cell lines in the presence of BSO and the L-cysteine source NAC. NAC reduced the sensitivity of MCF7, but not MDA-MB-231, cells to HYP PDT which is in accordance with the antioxidant effects of L-cysteine and its potential as a GSTP substrate. As a conclusion we have herein shown that the different GSH based cell defense mechanisms can be utilized as predictive markers for the outcome of PDT and as a guide for selecting optimal combination strategies. Copyright © 2017 The

Reaction of rat liver glutathione S-transferases and bacterial dichloromethane dehalogenase with dihalomethanes.

PubMed

Blocki, F A; Logan, M S; Baoli, C; Wackett, L P

1994-03-25

Dichloromethane dehalogenase from Methylophilus sp. DM11 is a glutathione S-transferase homolog that is specifically active with dihalomethane substrates. This bacterial enzyme and rat liver glutathione S-transferases were purified to investigate their relative reactivity with CH2Cl2 and related substrates. Rat liver alpha class glutathione transferases were inactive and mu class enzymes showed low activity (7-23 nmol/min/mg of protein) with CH2Cl2. theta class glutathione transferase 5-5 from rat liver and Methylophilus sp. dichloromethane dehalogenase showed specific activities of > or = 1 mumol/min/mg of protein. Apparent Kcat/Km were determined to be 3.3 x 10(4) and 6.0 x 10(4) L M-1 S-1 for the two enzymes, respectively. Dideutero-dichloromethane was processed to dideutereo-formaldehyde, consistent with a nucleophilic halide displacement mechanism. The possibility of a GSCH2X reaction intermediate (GS, glutathione; X, halide) was probed using CH2ClF to generate a more stable halomethylglutathione species (GSCH2F). The reaction of CH2ClF with dichloromethane dehalogenase produced a kinetically identifiable intermediate that decomposed to formaldehyde at a similar rate to synthetic HOCH2CH2SCH2F. 19F-NMR revealed the transient formation of an intermediate identified as GSCH2F by its chemical shift, its triplet resonance, and H-F coupling constant consistent with a fluoromethylthioether. Its decomposition was matched by a stoichiometric formation of fluoride. These studies indicated that the bacterial dichloromethane dehalogenase directs a nucleophilic attack of glutathione on CH2Cl2 to produce a halomethylthioether intermediate. This focuses attention on the mechanism used by theta class glutathione transferases to generate a halomethylthioeter from relatively unreactive dihalomethanes.

Auranofin-induced oxidative stress causes redistribution of the glutathione pool in Taenia crassiceps cysticerci.

PubMed

Martínez-González, J J; Guevara-Flores, A; Rendón, J L; Arenal, I P Del

2015-05-01

Previously, we have studied the effect of the gold-compound auranofin (AF) on both thioredoxin-glutathione reductasa (TGR) activity and viability of Taenia crassiceps cysticerci. It was demonstrated that micromolar concentrations of AF were high enough to fully inhibit TGR and kill the parasites. In this work, the dynamics of changes in the glutathione pool of T. crassiceps cysticerci following the addition of AF, was analyzed. A dose-dependent decrease in the internal glutathione concentration, concomitant with an increase in ROS production was observed. These changes were simultaneous with the formation of glutathione-protein complexes and the export of glutathione disulfide (GSSG) to the culture medium. Incubation of cysticerci in the presence of both AF and N-acetyl cysteine (NAC) prevents all the above changes, maintaining cysticerci viability. By contrast, the presence of both AF and buthionine sulfoximine (BSO) resulted in a potentiation of the effects of the gold compound, jeopardizing cysticerci viability. These results suggest the lethal effect of AF on T. crassiceps cysticerci, observed at micromolar concentrations, can be explained as a consequence of major changes in the glutathione status, which results in a significant increase in the oxidative stress of the parasites. Copyright © 2015 Elsevier B.V. All rights reserved.

Vacuolar transport of the glutathione conjugate of trans-cinnamic acid.

PubMed

Walczak, H A; Dean, J V

2000-02-01

Red beet (Beta vulgaris L.) tonoplast membrane vesicles and [14C]trans-cinnamic acid-glutatione were used to study the vacuolar transport of phynylpropanoid-glutathione conjugates which are formed in peroxidase-mediated reactions. It was determined that the uptake of [14C]trans-cinnamic acid-glutathione into the tonoplast membrane vesicles was MgATP dependent and was 10-fold faster than the uptake of non-conjugated [14C]trans-cinnamic acid. Uptake of the conjugate in the presence of MgATP was not dependent on a trans-tonoblast H+-electrochemical gradient, because uptake was not affected by the addition of NH4Cl (1 mM; 0% inhibition) and was only slightly affected by gramicidin-D (5 microM; 14% inhibition). Uptake of the conjugate was inhibited 92% by the addition of vanadate (1 mM) and 71% by the addition of the model substrate S-(2,4-dinitrophenyl) glutathione (500 microM). Uptake did not occur when a nonhydrolyzable analog of ATP was used in place of MgATP. The calculated Km and Vmax values for uptake were 142 microM amd 5.95 nmol mg(-1) min(-1), respectively. Based on these results, phenylpropanoid-glutation conjugates formed in peroxidase-mediated reactions appear to be transported into the vacuole by the glutathione S-conjugate pump(s) located in the tonoplast membrane.

Balneotherapy and platelet glutathione metabolism in type II diabetic patients

NASA Astrophysics Data System (ADS)

Ohtsuka, Yoshinori; Yabunaka, Noriyuki; Watanabe, Ichiro; Noro, Hiroshi; Agishi, Yuko

1996-09-01

Effects of balneotherapy on platelet glutathione metabolism were investigated in 12 type II (non-insulin-dependent) diabetic patients. Levels of the reduced form of glutathione (GSH) on admission were well correlated with those of fasting plasma glucose (FPG; r=0.692, P<0.02). After 4 weeks of balneotherapy, the mean level of GSH showed no changes; however, in well-controlled patients (FPG <150 mg/dl), the level increased ( P<0.01) and in poorly controlled patients (FPG >150 mg/dl), the value decreased ( P<0.05). There was a negative correlation between glutathione peroxidase (GPX) activities and the levels of FPG ( r=-0.430, P<0.05). After balneotherapy, the activity increased in 5 patients, decreased in 3 patients and showed no changes (alteration within ±3%) in all the other patients. From these findings in diabetic patients we concluded: (1) platelet GSH synthesis appeared to be induced in response to oxidative stress; (2) lowered GPX activities indicated that the antioxidative defense system was impaired; and (3) platelet glutathione metabolism was partially improved by 4 weeks balneotherapy, an effect thought to be dependent on the control status of plasma glucose levels. It is suggested that balneotherapy is beneficial for patients whose platelet antioxidative defense system is damaged, such as those with diabetes mellitus and coronary heart disease.

Development and characterization of glutathione-conjugated albumin nanoparticles for improved brain delivery of hydrophilic fluorescent marker.

PubMed

Patel, Prerak J; Acharya, Niyati S; Acharya, Sanjeev R

2013-01-01

The glutathione-conjugated bovine serum albumin (BSA) nanoparticles were constructed in the present exploration as a novel biodegradable carrier for brain-specific drug delivery with evaluation of its in vitro and in vivo delivery properties. BSA nanocarriers were activated and conjugated to the distal amine functions of the glutathione via carbodiimide chemistry using EDAC as a mediator. These nanoparticles were characterized for particle shape, average size, SPAN value, drug entrapment and in vitro drug release. Further, presence of glutathione on the surface of BSA nanoparticles was confirmed by Ellman's assay, which has suggested that approximately 750 units of glutathione were conjugated per BSA nanoparticle. To evaluate the brain delivery properties of the glutathione-conjugated BSA nanoparticles fluorescein sodium was used as a model hydrophilic compound. Permeability and neuronal uptake properties of developed formulations were evaluated against the MDCK-MDR1 endothelial and neuro-glial cells, respectively. The permeability of glutathione-conjugated BSA nanoparticles across the monolayer of MDCK-MDR1 endothelial tight junction was shown significantly higher than that of unconjugated nanoparticles and fluorescein sodium solution. Similarly, glutathione-conjugated nanoparticles exhibited considerably higher uptake by neuro-glial cells which was inferred by high fluorescence intensity under microscope in comparison to unconjugated nanoparticles and fluorescein sodium solution. Following an intravenous administration, nearly three folds higher fluorescein sodium was carried to the rat brain by glutathione-conjugated nanoparticles as compared to unconjugated nanoparticles. The significant in vitro and in vivo results suggest that glutathione-conjugated BSA nanoparticles is a promising brain drug delivery system with low toxicity.

Plasma L-5-oxoproline kinetics and whole blood glutathione synthesis rates in severely burned adult humans.

PubMed

Yu, Yong-Ming; Ryan, Colleen M; Fei, Zhe-Wei; Lu, Xiao-Ming; Castillo, Leticia; Schultz, John T; Tompkins, Ronald G; Young, Vernon R

2002-02-01

Compromised glutathione homeostasis is associated with increased morbidity in various disease states. We evaluated the kinetics of L-5-oxoproline, an intermediate in the gamma-glutamyl cycle of glutathione production, in fourteen severely burned adults by use of a primed, constant intravenous infusion of L-5-[1-(13)C]oxoproline. In nine of these patients, whole blood glutathione synthesis and plasma kinetics of glycine and leucine were also measured with [(15)N]glycine and L-[(2)H(3)]leucine tracers. Patients were studied under a "basal" condition that provided a low dose of glucose and total parenteral nutrition. For comparison with control subjects, whole blood glutathione synthesis was estimated in six healthy adults. Burn patients in a basal condition showed significantly higher rates of plasma oxoproline clearance and urinary D- and L-oxoproline excretion compared with fasting healthy control subjects. Whole blood glutathione concentration and absolute synthesis rate in the basal state were lower than for control subjects. Total parenteral feeding without cysteine but with generous methionine did not affect oxoproline kinetics or whole blood glutathione synthesis. The estimated rate of glycine de novo synthesis was also lower in burn patients, suggesting a possible change in glycine availability for glutathione synthesis. The roles of precursor amino acid availability, as well as alterations in metabolic capacity, in modulating whole blood glutathione production in burns now require investigation.

GLUTATHIONE S-TRANSFERASE-MEDIATED METABOLISM OF BROMODICHLOROMETHANE

EPA Science Inventory

GLUTATHIONE s-TRANSFERASE-MEDIATED METABOLISM OF BROMODICHLOROMETHANE. M K Ross1 and R A Pegram2. 1Curriculum in Toxicology, University of North Carolina at Chapel Hill; 2Experimental Toxicology Division, NHEERL/ORD, United States Environmental Protection Agency, Research Triangl...

Radiosensitization of Hypoxic Tumor Cells by Depletion of Intracellular Glutathione

NASA Astrophysics Data System (ADS)

Bump, Edward A.; Yu, Ning Y.; Brown, J. Martin

1982-08-01

Depletion of glutathione in Chinese hamster ovary cells in vitro by diethyl maleate resulted in enhancement of the effect of x-rays on cell survival under hypoxic conditions but not under oxygenated conditions. Hypoxic EMT6 tumor cells were similarly sensitized in vivo. The action of diethyl maleate is synergistic with the effect of the electron-affinic radiosensitizer misonidazole, suggesting that the effectiveness of misonidazole in cancer radiotherapy may be improved by combining it with drugs that deplete intracellular glutathione.

Radiosensitization of hypoxic tumor cells by depletion of intracellular glutathione

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bump, E.A.; Yu, N.Y.; Brown, J.M.

1982-08-06

Depletion of glutathione in Chinese hamster ovary cells in vitro by diethyl maleate resulted in enhancement of the effect of x-rays on cell survival under hypoxic conditions but not under oxygenated conditions. Hypoxic EMT6 tumor cells were similarly sensitized in vivo. The action of diethyl maleate is synergistic with the effect of the electron-affinic radiosensitizer misonidazole, suggesting that the effectiveness of misonidazole in cancer radiotherapy may be improved by combining it with drugs that deplete intracellular glutathione.

Transcriptional analysis of disk abalone (Haliotis discus discus) antioxidant enzymes against marine bacteria and virus challenge.

PubMed

De Zoysa, Mahanama; Whang, Ilson; Nikapitiya, Chamilani; Oh, Chulhong; Choi, Cheol Young; Lee, Jehee

2011-07-01

Diverse antioxidant enzymes are essential for marine organisms to overcome oxidative stress as well as for the fine-tuning of immune reactions through activating different signal transduction pathways. This study describes the transcriptional analysis of antioxidant enzymes of disk abalone by challenging with bacteria (Vibrio alginolyticus, Vibrio parahemolyticus, and Listeria monocytogenes) and viral hemorrhagic septicemia virus (VHSV). Upon bacteria and VHSV challenge, Manganese superoxide dismutase (MnSOD), Copper, Zinc superoxide dismutase (CuZnSOD), catalase, thioredoxin peroxidase (TPx), Selenium-dependent glutathione peroxidase (SeGPx), and thioredoxin-2 (TRx-2) expression levels were altered in gills, and hemocytes at different magnitudes. In gills, only MnSOD, catalase, and SeGPx genes were completely upregulated by post-challenge of bacterial and VHSV. Among them, SeGPx demonstrated strong upregulation by 16-fold (bacteria) and 2-fold (VHSV) in gills, and 5-fold (bacteria) and 3.0-fold (VHSV) in hemocytes. None of the genes examined were downregulated (in gills and hemocytes) by bacteria challenge even though CuZnSOD and TPx showed downregulation (completely) in hemocytes by VHSV. In general, abalone hemocytes had lower potential to induce antioxidant enzyme transcripts upon bacteria and VHSV challenge than gills. Based upon these results, we suggest that abalones induce oxidative stress in tissues during the bacteria and VHSV challenge, and the identified response of antioxidant enzymes could be supported for maintaining a low-level of reactive oxygen species (ROS) that may serve as a signal for activating immune reactions against pathogenic conditions. Copyright © 2011 Elsevier Ltd. All rights reserved.

[The activity of glutathione antioxidant system at melaksen and valdoxan action under experimental hyperthyroidism in rats].

PubMed

Gorbenko, M V; Popova, T N; Shul'gin, K K; Popov, S S

2013-01-01

Investigation of glutathione antioxidant system activity and diene conjugates content in rats liver and blood serum at the influence of melaksen and valdoxan under experimental hyperthyroidism (EG) has been revealed. It has been established that the activities of glutathione reductase (GR), glutathione peroxidase (GP) and glutathione transferase (GT), growing at pathological conditions, change to the side of control value at these substunces introduction. Reduced glutathione content (GSH) at melaxen and valdoxan action increased compared with values under the pathology, that, obviously, could be associated with a reduction of its spending on the detoxication of free radical oxidation (FRO) toxic products. Diene conjugates level in rats liver and blood serum, increasing at experimental hyperthyroidism conditions, under introduction of melatonin level correcting drugs, also approached to the control meaning. Results of the study indicate on positive effect of melaxen and valdoxan on free radical homeostasis, that appears to be accompanied by decrease of load on the glutathione antioxidant system in comparison with the pathology.

Regional modulation of the response to glutathione in Hydra vulgaris.

PubMed

Pierobon, Paola

2015-07-01

In the presence of prey, or upon exposure to reduced glutathione (GSH), Hydra polyps open a mouth to ingest the captured prey and close it after feeding; at rest the mouth is not evident. In previous papers we have shown that GABA, glycine and NMDA modulate the mechanisms of mouth closure through ligand-gated-ion-channel receptors that are similar to their mammalian analogues in terms of biochemical and pharmacological properties. In order to study the regional distribution of these receptors, we have applied the GSH assay to polyps amputated at different levels of the body column. The response to 1-10 µmol l(-1) GSH of polyps lacking either peduncle and foot or the entire body columns (heads) was not different from control, whole animals. In the presence of GABA or muscimol, duration of the response was significantly decreased in heads; the decrease was suppressed by the GABA antagonists gabazine and bicuculline. By contrast, in animals lacking peduncle and foot, duration of the response did not vary upon GABA administration. Conversely, in the presence of glycine, duration of the response in heads preparations was similar to control, whereas in footless polyps, it was significantly reduced. The decrease was mimicked by the glycine agonists taurine and β-alanine, and counteracted by strychnine. These results suggest a regional distribution of receptors to GABA and glycine in the neuromuscular circuitry modulating the feeding behaviour. © 2015. Published by The Company of Biologists Ltd.

Glutathione S-transferase M1 and glutathione S-transferase T1 genotype in chronic pancreatitis: a meta-analysis.

PubMed

Zhong, Yanjun; Zou, Runmei; Cao, Jie; Peng, Mou

2015-02-01

A meta-analysis to determine the association between chronic pancreatitis and glutathione-S transferase (GST) mu 1 (GSTM1) and theta 1 (GSTT1) deletions. Case-control studies concerning the relationship between chronic pancreatitis and GSTM1 or GSTT1 deletions were identified (up to October 2013). Meta-analyses of the association between GSTM1 and GSTT1 genotype and chronic pancreatitis or alcoholic chronic pancreatitis (ACP) were performed. Seven studies were included in the meta-analysis (650 patients/1382 controls for GSTM1 and 536 patients/1304 controls for GSTT1). There were no significant relationships between GSTM1/GSTT1 and chronic pancreatitis or GSTT1 and ACP. There was a significant association between GSTM1 null genotype and ACP (odds ratio 1.16, 95% confidence intervals 1.03, 1.30). The GSTM1 null genotype was significantly associated with ACP risk. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

The crucial protective role of glutathione against tienilic acid hepatotoxicity in rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nishiya, Takayoshi; Mori, Kazuhiko; Hattori, Chiharu

2008-10-15

To investigate the hepatotoxic potential of tienilic acid in vivo, we administered a single oral dose of tienilic acid to Sprague-Dawley rats and performed general clinicopathological examinations and hepatic gene expression analysis using Affymetrix microarrays. No change in the serum transaminases was noted at up to 1000 mg/kg, although slight elevation of the serum bile acid and bilirubin, and very mild hepatotoxic changes in morphology were observed. In contrast to the marginal clinicopathological changes, marked upregulation of the genes involved in glutathione biosynthesis [glutathione synthetase and glutamate-cysteine ligase (Gcl)], oxidative stress response [heme oxygenase-1 and NAD(P)H dehydrogenase quinone 1] andmore » phase II drug metabolism (glutathione S-transferase and UDP glycosyltransferase 1A6) were noted after 3 or 6 h post-dosing. The hepatic reduced glutathione level decreased at 3-6 h, and then increased at 24 or 48 h, indicating that the upregulation of NF-E2-related factor 2 (Nrf2)-regulated gene and the late increase in hepatic glutathione are protective responses against the oxidative and/or electrophilic stresses caused by tienilic acid. In a subsequent experiment, tienilic acid in combination with L-buthionine-(S,R)-sulfoximine (BSO), an inhibitor of Gcl caused marked elevation of serum alanine aminotransferase (ALT) with extensive centrilobular hepatocyte necrosis, whereas BSO alone showed no hepatotoxicity. The elevation of ALT by this combination was observed at the same dose levels of tienilic acid as the upregulation of the Nrf2-regulated genes by tienilic acid alone. In conclusion, these results suggest that the impairment of glutathione biosynthesis may play a critical role in the development of tienilic acid hepatotoxicity through extensive oxidative and/or electrophilic stresses.« less

Pleiotropic Functions of Glutathione S-Transferase P

PubMed Central

Zhang, Jie; Grek, Christina; Ye, Zhi-Wei; Manevich, Yefim; Tew, Kenneth D.; Townsend, Danyelle M.

2016-01-01

Glutathione S-transferase P (GSTP) is one member of the GST superfamily that is prevalently expressed in mammals. Known to possess catalytic activity through deprotonating glutathione allowing formation of thioether bonds with electrophilic substrates, more recent discoveries have broadened our understanding of the biological roles of this protein. In addition to catalytic detoxification, other properties so far ascribed to GSTP include chaperone functions, regulation of nitric oxide pathways, regulation of a variety of kinase signaling pathways, and participation in the forward reaction of protein S-glutathionylation. The expression of GSTP has been linked with cancer and other human pathologies and more recently even with drug addiction. With respect to human health, polymorphic variants of GSTP may determine individual susceptibility to oxidative stress and/or be critical in the design and development of drugs that have used redox pathways as a discovery platform. PMID:24974181

Glutathione determination by the Tietze enzymatic recycling assay and its relationship to cellular radiation response.

PubMed Central

Eady, J. J.; Orta, T.; Dennis, M. F.; Stratford, M. R.; Peacock, J. H.

1995-01-01

Large fluctuations in glutathione content were observed on a daily basis using the Tietze enzyme recycling assay in a panel of six human cell lines of varying radiosensitivity. Glutathione content tended to increase to a maximum during exponential cell proliferation, and then decreased at different rates as the cells approached plateau phase. By reference to high-performance liquid chromatography and flow cytometry of the fluorescent bimane derivative we were able to verify that these changes were real. However, the Tietze assay was occasionally unable to detect glutathione in two of our cell lines (MGH-U1 and AT5BIVA), although the other methods indicated its presence. The existence of an inhibitory activity responsible for these anomalies was confirmed through spiking our samples with known amounts of glutathione. We were unable to detect a direct relationship between cellular glutathione concentration and aerobic radiosensitivity in our panel of cell lines. PMID:7577452

Targeting brain cells with glutathione-modulated nanoliposomes: in vitro and in vivo study

PubMed Central

Salem, Heba F; Ahmed, Sayed M; Hassaballah, Ashraf E; Omar, Mahmoud M

2015-01-01

Background The blood–brain barrier prevents many drug moieties from reaching the central nervous system. Therefore, glutathione-modulated nanoliposomes have been engineered to enhance the targeting of flucytosine to the brain. Methods Glutathione-modulated nanoliposomes were prepared by thin-film hydration technique and evaluated in the primary brain cells of rats. Lecithin, cholesterol, and span 65 were mixed at 1:1:1 molar ratio. The molar percentage of PEGylated glutathione varied from 0 mol% to 0.75 mol%. The cellular binding and the uptake of the targeted liposomes were both monitored by epifluorescent microscope and flow cytometry techniques. A biodistribution and a pharmacokinetic study of flucytosine and flucytosine-loaded glutathione–modulated liposomes was carried out to evaluate the in vivo brain-targeting efficiency. Results The size of glutathione-modulated nanoliposomes was <100 nm and the zeta potential was more than −65 mV. The cumulative release reached 70% for certain formulations. The cellular uptake increased as molar percent of glutathione increased to reach the maximum at 0.75 mol%. The uptake of the targeted liposomes by brain cells of the rats was three times greater than that of the nontargeted liposomes. An in vivo study showed that the relative efficiency was 2.632±0.089 and the concentration efficiency was 1.590±0.049, and also, the drug-targeting index was 3.670±0.824. Conclusion Overall, these results revealed that glutathione-PEGylated nanoliposomes enhance the effective delivery of flucytosine to brain and could become a promising new therapeutic option for the treatment of the brain infections. PMID:26229435

The proportional lack of archaeal pathogens: Do viruses/phages hold the key?

PubMed Central

Gill, Erin E; Brinkman, Fiona S L

2011-01-01

Although Archaea inhabit the human body and possess some characteristics of pathogens, there is a notable lack of pathogenic archaeal species identified to date. We hypothesize that the scarcity of disease-causing Archaea is due, in part, to mutually-exclusive phage and virus populations infecting Bacteria and Archaea, coupled with an association of bacterial virulence factors with phages or mobile elements. The ability of bacterial phages to infect Bacteria and then use them as a vehicle to infect eukaryotes may be difficult for archaeal viruses to evolve independently. Differences in extracellular structures between Bacteria and Archaea would make adsorption of bacterial phage particles onto Archaea (i.e. horizontal transfer of virulence) exceedingly hard. If phage and virus populations are indeed exclusive to their respective host Domains, this has important implications for both the evolution of pathogens and approaches to infectious disease control. PMID:21328413

Activity-Based Probes for Isoenzyme- and Site-Specific Functional Characterization of Glutathione S -Transferases

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stoddard, Ethan G.; Killinger, Bryan J.; Nair, Reji N.

Glutathione S-transferases (GSTs) comprise a highly diverse family of phase II drug metabolizing enzymes whose shared function is the conjugation of reduced glutathione to various endo- and xenobiotics. Although the conglomerate activity of these enzymes can be measured by colorimetric assays, measurement of the individual contribution from specific isoforms and their contribution to the detoxification of xenobiotics in complex biological samples has not been possible. For this reason, we have developed two activity-based probes that characterize active glutathione transferases in mammalian tissues. The GST active site is comprised of a glutathione binding “G site” and a distinct substrate binding “Hmore » site”. Therefore, we developed (1) a glutathione-based photoaffinity probe (GSH-ABP) to target the “G site”, and (2) a probe designed to mimic a substrate molecule and show “H site” activity (GST-ABP). The GSH-ABP features a photoreactive moiety for UV-induced covalent binding to GSTs and glutathione-binding enzymes. The GST-ABP is a derivative of a known mechanism-based GST inhibitor that binds within the active site and inhibits GST activity. Validation of probe targets and “G” and “H” site specificity was carried out using a series of competitors in liver homogenates. Herein, we present robust tools for the novel characterization of enzyme- and active site-specific GST activity in mammalian model systems.« less

Evidence for the Presence of the Ascorbate-Glutathione Cycle in Mitochondria and Peroxisomes of Pea Leaves.

PubMed Central

Jimenez, A.; Hernandez, J. A.; Del Rio, L. A.; Sevilla, F.

1997-01-01

The presence of the enzymes of the ascorbate-glutathione cycle was investigated in mitochondria and peroxisomes purified from pea (Pisum sativum L.) leaves. All four enzymes, ascorbate peroxidase (APX; EC 1.11.1.11), monodehydroascorbate reductase (EC 1.6.5.4), dehydroascorbate reductase (EC 1.8.5.1), and glutathione reductase (EC 1.6.4.2), were present in mitochondria and peroxisomes, as well as in the antioxidants ascorbate and glutathione. The activity of the ascorbate-glutathione cycle enzymes was higher in mitochondria than in peroxisomes, except for APX, which was more active in peroxisomes than in mitochondria. Intact mitochondria and peroxisomes had no latent APX activity, and this remained in the membrane fraction after solubilization assays with 0.2 M KCl. Monodehydroascorbate reductase was highly latent in intact mitochondria and peroxisomes and was membrane-bound, suggesting that the electron acceptor and donor sites of this redox protein are not on the external side of the mitochondrial and peroxisomal membranes. Dehydroascorbate reductase was found mainly in the soluble peroxisomal and mitochondrial fractions. Glutathione reductase had a high latency in mitochondria and peroxisomes and was present in the soluble fractions of both organelles. In intact peroxisomes and mitochondria, the presence of reduced ascorbate and glutathione and the oxidized forms of ascorbate and glutathione were demonstrated by high-performance liquid chromatography analysis. The ascorbate-glutathione cycle of mitochondria and peroxisomes could represent an important antioxidant protection system against H2O2 generated in both plant organelles. PMID:12223704

Withanolide A Prevents Neurodegeneration by Modulating Hippocampal Glutathione Biosynthesis during Hypoxia

PubMed Central

Baitharu, Iswar; Jain, Vishal; Deep, Satya Narayan; Shroff, Sabita; Sahu, Jayanta Kumar; Naik, Pradeep Kumar; Ilavazhagan, Govindasamy

2014-01-01

Withania somnifera root extract has been used traditionally in ayurvedic system of medicine as a memory enhancer. Present study explores the ameliorative effect of withanolide A, a major component of withania root extract and its molecular mechanism against hypoxia induced memory impairment. Withanolide A was administered to male Sprague Dawley rats before a period of 21 days pre-exposure and during 07 days of exposure to a simulated altitude of 25,000 ft. Glutathione level and glutathione dependent free radicals scavenging enzyme system, ATP, NADPH level, γ-glutamylcysteinyl ligase (GCLC) activity and oxidative stress markers were assessed in the hippocampus. Expression of apoptotic marker caspase 3 in hippocampus was investigated by immunohistochemistry. Transcriptional alteration and expression of GCLC and Nuclear factor (erythroid-derived 2)–related factor 2 (Nrf2) were investigated by real time PCR and immunoblotting respectively. Exposure to hypobaric hypoxia decreased reduced glutathione (GSH) level and impaired reduced gluatathione dependent free radical scavenging system in hippocampus resulting in elevated oxidative stress. Supplementation of withanolide A during hypoxic exposure increased GSH level, augmented GSH dependent free radicals scavenging system and decreased the number of caspase and hoescht positive cells in hippocampus. While withanolide A reversed hypoxia mediated neurodegeneration, administration of buthionine sulfoximine along with withanolide A blunted its neuroprotective effects. Exogenous administration of corticosterone suppressed Nrf2 and GCLC expression whereas inhibition of corticosterone synthesis upregulated Nrf2 as well as GCLC. Thus present study infers that withanolide A reduces neurodegeneration by restoring hypoxia induced glutathione depletion in hippocampus. Further, Withanolide A increases glutathione biosynthesis in neuronal cells by upregulating GCLC level through Nrf2 pathway in a corticosterone dependenet manner

Withanolide A prevents neurodegeneration by modulating hippocampal glutathione biosynthesis during hypoxia.

PubMed

Baitharu, Iswar; Jain, Vishal; Deep, Satya Narayan; Shroff, Sabita; Sahu, Jayanta Kumar; Naik, Pradeep Kumar; Ilavazhagan, Govindasamy

2014-01-01

Withania somnifera root extract has been used traditionally in ayurvedic system of medicine as a memory enhancer. Present study explores the ameliorative effect of withanolide A, a major component of withania root extract and its molecular mechanism against hypoxia induced memory impairment. Withanolide A was administered to male Sprague Dawley rats before a period of 21 days pre-exposure and during 07 days of exposure to a simulated altitude of 25,000 ft. Glutathione level and glutathione dependent free radicals scavenging enzyme system, ATP, NADPH level, γ-glutamylcysteinyl ligase (GCLC) activity and oxidative stress markers were assessed in the hippocampus. Expression of apoptotic marker caspase 3 in hippocampus was investigated by immunohistochemistry. Transcriptional alteration and expression of GCLC and Nuclear factor (erythroid-derived 2)-related factor 2 (Nrf2) were investigated by real time PCR and immunoblotting respectively. Exposure to hypobaric hypoxia decreased reduced glutathione (GSH) level and impaired reduced gluatathione dependent free radical scavenging system in hippocampus resulting in elevated oxidative stress. Supplementation of withanolide A during hypoxic exposure increased GSH level, augmented GSH dependent free radicals scavenging system and decreased the number of caspase and hoescht positive cells in hippocampus. While withanolide A reversed hypoxia mediated neurodegeneration, administration of buthionine sulfoximine along with withanolide A blunted its neuroprotective effects. Exogenous administration of corticosterone suppressed Nrf2 and GCLC expression whereas inhibition of corticosterone synthesis upregulated Nrf2 as well as GCLC. Thus present study infers that withanolide A reduces neurodegeneration by restoring hypoxia induced glutathione depletion in hippocampus. Further, Withanolide A increases glutathione biosynthesis in neuronal cells by upregulating GCLC level through Nrf2 pathway in a corticosterone dependenet manner.

Selenium concentrations and enzyme activities of glutathione metabolism in wild long-tailed ducks and common eiders

USGS Publications Warehouse

Franson, J. Christian; Hoffman, David J.; Flint, Paul L.

2011-01-01

The relationships of selenium (Se) concentrations in whole blood with plasma activities of total glutathione peroxidase, Se-dependent glutathione peroxidase, and glutathione reductase were studied in long-tailed ducks (Clangula hyemalis) and common eiders (Somateria mollissima) sampled along the Beaufort Sea coast of Alaska, USA. Blood Se concentrations were >8 μg/g wet weight in both species. Linear regression revealed that the activities of total and Se-dependent glutathione peroxidase were significantly related to Se concentrations only in long-tailed ducks, raising the possibility that these birds were experiencing early oxidative stress.

Chlortetracycline detoxification in maize via induction of glutathione S-transferases after antibiotic exposure.

PubMed

Farkas, Michael H; Berry, James O; Aga, Diana S

2007-02-15

Soil contamination with nonmetabolized antibiotics is an emerging environmental concern, especially on agricultural croplands that receive animal manure as fertilizer. In this study, phytotoxicity of chlortetracycline (CTC) antibiotics on pinto beans (Phaseolus vulgaris) and maize (Zea mays) was investigated under controlled conditions. When grown in CTC-treated soil, a significant increase in the activities of the plant stress proteins glutathione S-transferases (GST) and peroxidases (POX) were observed in maize plants, but not in pinto beans. In vitro conjugation reactions demonstrated that the induced GST in maize catalyzed the conjugation of glutathione (GSH) with CTC, producing stable conjugates that were structurally characterized using liquid chromatography/mass spectrometry. The antibiotic-induced GST produced CTC-glutathione conjugate at relative concentrations 2-fold higher than that produced by constitutively expressed GST extracted from untreated maize. On the other hand, GST extracted from pinto beans (both treated and untreated) did not efficiently catalyze glutathione conjugation with CTC. These results suggest that maize is able to detoxify chlortetracycline via the glutathione pathway, whereas pinto beans cannot. This may explain the observed stunted growth of pinto beans after antibiotic treatment. This study demonstrates the importance of plant uptake in determining the fate of antibiotics in soil and their potential phytotoxicity to susceptible plants.

Glycation of human erythrocyte glutathione peroxidase: effect on the physical and kinetic properties.

PubMed

Suravajjala, Sreekanth; Cohenford, Menashi; Frost, Leslie R; Pampati, Praveen K; Dain, Joel A

2013-06-05

Glutathione peroxidase (GPx) is a significant antioxidant enzyme that plays a key role in protecting the body from reactive oxygen species (ROS) and their toxicity. As a biocatalyst, the enzyme has been shown to reduce hydrogen peroxide to water and lipid hydroperoxides to their respective alcohols. The increased levels of ROS in patients with diabetes have been speculated to arise, in part, from alterations in the activity of glutathione antioxidant enzymes, perhaps, by mechanisms such as the glycation of the protein, in vivo. Under physiological conditions of temperature and pH, we investigated the susceptibility of human glutathione peroxidase to glycation, determined the effects of glycation on the physical and kinetic properties of the enzyme, and identified the protein's vulnerable amino acid sites of glycation. Circular dichroism, UV and mass spectrometry studies revealed that methylglyoxal and DL-glyceraldehyde are potent glycators of glutathione peroxidase; destabilizing its structure, altering its pH activity and stability profiles and increasing its Km value. In comparison to DL-glyceraldehyde, methylglyxol was a more potent glycator of the enzyme and was found to nonenzymatically condense with Arg-177, located near the glutathione binding site of GPx. Copyright © 2013 Elsevier B.V. All rights reserved.

Diagnosis of glutathione synthetase deficiency in newborn screening.

PubMed

Simon, E; Vogel, M; Fingerhut, R; Ristoff, E; Mayatepek, E; Spiekerkötter, U

2009-12-01

Glutathione synthetase (GSS) deficiency is a rare disorder of glutathione metabolism with varying clinical severity. Patients may present with haemolytic anaemia alone or together with acidosis and central nervous system impairment. Diagnosis is made by clinical presentation and detection of elevated concentrations of 5-oxoproline in urine and low GSS activity in erythrocytes or cultured skin fibroblasts. Diagnosis can be confirmed by mutational analysis. Treatment consists of the correction of acidosis, blood transfusion, and supplementation with antioxidants. The most important determinants for outcome and survival in patients with GSS deficiency are early diagnosis and early initiation of treatment. The case of a newborn with GSS deficiency diagnosed by tandem mass spectrometry (MS/MS)-based newborn screening is described. After onset of clinical symptoms on the 2nd day of life, expanded newborn screening revealed normal results for all disorders included in the German screening programme; however, selective MS/MS screening revealed a >10-fold elevation of 5-oxoproline in dried blood, leading to the presumptive diagnosis of GSS deficiency by the 5th day of life. Diagnosis was later confirmed by detection of markedly reduced glutathione concentration in erythrocytes and mutational analysis of the GSS gene. Presently, GSS deficiency is not included in newborn screening programmes in Europe. As outcome depends significantly on early start of treatment, routine inclusion of this disorder in newborn screening panels should be considered.

Comparison of glutathione levels measured using optimized monochlorobimane assay with those from ortho-phthalaldehyde assay in intact cells.

PubMed

Čapek, Jan; Hauschke, Martina; Brůčková, Lenka; Roušar, Tomáš

2017-11-01

Fluorometric glutathione assays have been generally preferred for their high specificity and sensitivity. An additional advantage offered by fluorescent bimane dyes is their ability to penetrate inside the cell. Their ability to react with glutathione within intact cells is frequently useful in flow cytometry and microscopy. Hence, the aims of our study were to use monochlorobimane for optimizing a spectrofluorometric glutathione assay in cells and then to compare that assay with the frequently used ortho-phthalaldehyde assay. We used glutathione-depleting agents (e.g., cisplatin and diethylmalonate) to induce cell impairment. For glutathione assessment, monochlorobimane (40μM) was added to cells and fluorescence was detected at 394/490nm. In addition to the regularly used calculation of glutathione levels from fluorescence change after 60min, we used an optimized calculation from the linear part of the fluorescence curve after 10min of measurement. We found that 10min treatment of cells with monochlorobimane is sufficient for evaluating cellular glutathione concentration and provides results entirely comparable with those from the standard ortho-phthalaldehyde assay. In contrast, the results obtained by the standardly used evaluation after 60min of monochlorobimane treatment provided higher glutathione values. We conclude that measuring glutathione using monochlorobimane with the here-described optimized evaluation of fluorescence signal could be a simple and useful method for routine and rapid assessment of glutathione within intact cells in large numbers of samples. Copyright © 2017 Elsevier Inc. All rights reserved.

Differential Responses of the Antioxidant System of Ametryn and Clomazone Tolerant Bacteria

PubMed Central

Peters, Leila Priscila; Carvalho, Giselle; Martins, Paula Fabiane; Dourado, Manuella Nóbrega; Vilhena, Milca Bartz; Pileggi, Marcos; Azevedo, Ricardo Antunes

2014-01-01

The herbicides ametryn and clomazone are widely used in sugarcane cultivation, and following microbial degradation are considered as soil and water contaminants. The exposure of microorganisms to pesticides can result in oxidative damage due to an increase in the production of reactive oxygen species (ROS). This study investigated the response of the antioxidant systems of two bacterial strains tolerant to the herbicides ametryn and clomazone. Bacteria were isolated from soil with a long history of ametryn and clomazone application. Comparative analyses based on 16S rRNA gene sequences revealed that strain CC07 is phylogenetically related to Pseudomonas aeruginosa and strain 4C07 to P. fulva. The two bacterial strains were grown for 14 h in the presence of separate and combined herbicides. Lipid peroxidation, reduced glutathione content (GSH) and antioxidant enzymes activities were evaluated. The overall results indicated that strain 4C07 formed an efficient mechanism to maintain the cellular redox balance by producing reactive oxygen species (ROS) and subsequently scavenging ROS in the presence of the herbicides. The growth of bacterium strain 4C07 was inhibited in the presence of clomazone alone, or in combination with ametryn, but increased glutathione reductase (GR) and glutathione S-transferase (GST) activities, and a higher GSH concentration were detected. Meanwhile, reduced superoxide dismutase (SOD), catalase (CAT) and GST activities and a lower concentration of GSH were detected in the bacterium strain CC07, which was able to achieve better growth in the presence of the herbicides. The results suggest that the two bacterial strains tolerate the ametryn and clomazone herbicides with distinctly different responses of the antioxidant systems. PMID:25380132

Functional characterisation of ganglioside-induced differentiation-associated protein 1 as a glutathione transferase.

PubMed

Shield, Alison J; Murray, Tracy P; Board, Philip G

2006-09-08

Mutations in the ganglioside-induced differentiation-associated protein 1 (GDAP1) gene have been linked with Charcot-Marie-Tooth (CMT) disease. This protein, and its paralogue GDAP1L1, appear to be structurally related to the cytosolic glutathione S-transferases (GST) including an N-terminal thioredoxin fold domain with conserved active site residues. The specific function, of GDAP1 remains unknown. To further characterise their structure and function we purified recombinant human GDAP1 and GDAP1L1 proteins using bacterial expression and immobilised metal affinity chromatography. Like other cytosolic GSTs, GDAP1 protein has a dimeric structure. Although the full-length proteins were largely insoluble, the deletion of a proposed C-terminal transmembrane domain allowed the preparation of soluble protein. The purified proteins were assayed for glutathione-dependent activity against a library of 'prototypic' GST substrates. No evidence of glutathione-dependent activity or an ability to bind glutathione immobilised on agarose was found.

Glutathione and growth inhibition of Mycobacterium tuberculosis in healthy and HIV infected subjects

PubMed Central

Venketaraman, Vishwanath; Rodgers, Tatanisha; Linares, Rafael; Reilly, Nancy; Swaminathan, Shobha; Hom, David; Millman, Ariel C; Wallis, Robert; Connell, Nancy D

2006-01-01

Intracellular levels of glutathione are depleted in patients with acquired immunodeficiency syndrome in whom the risk of tuberculosis, particularly disseminated disease is many times that of healthy individuals. In this study, we examined the role of glutathione in immunity against tuberculosis infection in samples derived from healthy and human immunodeficiency virus infected subjects. Our studies confirm that glutathione levels are reduced in peripheral blood mononuclear cells and in red blood cells isolated from human immunodeficiency virus-infected subjects (CD4>400/cumm). Furthermore, treatment of blood cultures from human immunodeficiency virus infected subjects with N-acetyl cysteine, a glutathione precursor, caused improved control of intracellular M. tuberculosis infection. N-acetyl cysteine treatment decreased the levels of IL-1, TNF-α, and IL-6, and increased the levels of IFN-γ in blood cultures derived from human immunodeficiency virus-infected subjects, promoting the host immune responses to contain M. tuberculosis infection successfully. PMID:16504020

Intercellular Distribution of Glutathione Synthesis in Maize Leaves and Its Response to Short-Term Chilling1

PubMed Central

Gómez, Leonardo D.; Vanacker, Hélène; Buchner, Peter; Noctor, Graham; Foyer, Christine H.

2004-01-01

To investigate the intercellular control of glutathione synthesis and its influence on leaf redox state in response to short-term chilling, genes encoding γ-glutamylcysteine synthetase (γ-ECS) and glutathione synthetase (GSH-S) were cloned from maize (Zea mays) and specific antibodies produced. These tools were used to provide the first information on the intercellular distribution of γ-ECS and GSH-S transcript and protein in maize leaves, in both optimal conditions and chilling stress. A 2-d exposure to low growth temperatures (chill) had no effect on leaf phenotype, whereas return to optimal temperatures (recovery) caused extensive leaf bleaching. The chill did not affect total leaf GSH-S transcripts but strongly induced γ-ECS mRNA, an effect reversed during recovery. The chilling-induced increase in γ-ECS transcripts was not accompanied by enhanced total leaf γ-ECS protein or extractable activity. In situ hybridization and immunolocalization of leaf sections showed that γ-ECS and GSH-S transcripts and proteins were found in both the bundle sheath (BS) and the mesophyll cells under optimal conditions. Chilling increased γ-ECS transcript and protein in the BS but not in the mesophyll cells. Increased BS γ-ECS was correlated with a 2-fold increase in both leaf Cys and γ-glutamylcysteine, but leaf total glutathione significantly increased only in the recovery period, when the reduced glutathione to glutathione disulfide ratio decreased 3-fold. Thus, while there was a specific increase in the potential contribution of the BS cells to glutathione synthesis during chilling, it did not result in enhanced leaf glutathione accumulation at low temperatures. Return to optimal temperatures allowed glutathione to increase, particularly glutathione disulfide, and this was associated with leaf chlorosis. PMID:15047902

Effect of glutathione on the cadmium chelation of EDTA in a patient with cadmium intoxication.

PubMed

Gil, Hyo-wook; Kang, Eun-jung; Lee, Kwon-hyun; Yang, Jong-oh; Lee, Eun-young; Hong, Sae-yong

2011-01-01

In order to evaluate the efficiency and renal protective effects of glutathione during Ca(++)-EDTA chelation therapy for chronic cadmium intoxication, we measured the renal excretion of cadmium, β(2)-microglobulin, proteinuria, and hematuria during intravenous administration of glutathione with Ca(++)-EDTA in a 54-year-old patient with chronic cadmium intoxication. We administered 500 mg of Ca(++)-EDTA and 50 mg/kg of glutathione alone or in 1 L of normal saline over the next 24 hours and repeated this over 12 consecutive days. During the first 3 days, the basal levels (only saline administration) were determined; during the second 3 days, Ca(++)-EDTA only was administered, for the third sequence of 3 days, Ca(++)-EDTA with glutathione was provided, and for the last 3 days, glutathione alone was given. One month later, the same protocol was repeated. There were six blood and urine samples to analyze in each group. The blood cadmium level was higher when the EDTA was infused together with glutathione (7.44 ± 0.73 µg/L, p < 0.01) compared to the basal level of 4.6 ± 0.44 µg/L. Also, the renal cadmium excretion was significantly higher in the EDTA with glutathione group than in the basal group (23.4 ± 15.81 µg/g creatinine vs 89.23 ± 58.52 µg/g creatinine, p < 0.01). There was no difference in the protein/creatinine and β(2)-microglobulin/creatinine ratio in the urine (p > 0.05) among the groups. Furthermore, microhematuria and proteinuria did not develop over the observation period of 6 months. These results suggest that glutathione administration with EDTA might be an effective treatment modality for patients with cadmium intoxication.

Unbalanced Activation of Glutathione Metabolic Pathways Suggests Potential Involvement in Plant Defense against the Gall Midge Mayetiola destructor in Wheat

PubMed Central

Liu, Xuming; Zhang, Shize; Whitworth, R. Jeff; Stuart, Jeffrey J.; Chen, Ming-Shun

2015-01-01

Glutathione, γ-glutamylcysteinylglycine, exists abundantly in nearly all organisms. Glutathione participates in various physiological processes involved in redox reactions by serving as an electron donor/acceptor. We found that the abundance of total glutathione increased up to 60% in resistant wheat plants within 72 hours following attack by the gall midge Mayetiola destructor, the Hessian fly. The increase in total glutathione abundance, however, is coupled with an unbalanced activation of glutathione metabolic pathways. The activity and transcript abundance of glutathione peroxidases, which convert reduced glutathione (GSH) to oxidized glutathione (GSSG), increased in infested resistant plants. However, the enzymatic activity and transcript abundance of glutathione reductases, which convert GSSG back to GSH, did not change. This unbalanced regulation of the glutathione oxidation/reduction cycle indicates the existence of an alternative pathway to regenerate GSH from GSSG to maintain a stable GSSG/GSH ratio. Our data suggest the possibility that GSSG is transported from cytosol to apoplast to serve as an oxidant for class III peroxidases to generate reactive oxygen species for plant defense against Hessian fly larvae. Our results provide a foundation for elucidating the molecular processes involved in glutathione-mediated plant resistance to Hessian fly and potentially other pests as well. PMID:25627558

An extracytoplasmic function sigma factor-dependent periplasmic glutathione peroxidase is involved in oxidative stress response of Shewanella oneidensis

DOE PAGES

Dai, Jingcheng; Wei, Hehong; Tian, Chunyuan; ...

2015-01-01

Background: Bacteria use alternative sigma factors (σs) to regulate condition-specific gene expression for survival and Shewanella harbors multiple ECF (extracytoplasmic function) σ genes and cognate anti-sigma factor genes. Here we comparatively analyzed two of the rpoE-like operons in the strain MR-1: rpoE-rseA-rseB-rseC and rpoE2-chrR. Results: RpoE was important for bacterial growth at low and high temperatures, in the minimal medium, and high salinity. The degP/htrA orthologue, required for growth of Escherichia coli and Pseudomonas aeruginosa at high temperature, is absent in Shewanella, while the degQ gene is RpoE-regulated and is required for bacterial growth at high temperature. RpoE2 was essentialmore » for the optimal growth in oxidative stress conditions because the rpoE2 mutant was sensitive to hydrogen peroxide and paraquat. The operon encoding a ferrochelatase paralogue (HemH2) and a periplasmic glutathione peroxidase (PgpD) was identified as RpoE2-dependent. PgpD exhibited higher activities and played a more important role in the oxidative stress responses than the cytoplasmic glutathione peroxidase CgpD under tested conditions. The rpoE2-chrR operon and the identified regulon genes, including pgpD and hemH2, are coincidently absent in several psychrophilic and/or deep-sea Shewanella strains. Conclusion: In S. oneidensis MR-1, the RpoE-dependent degQ gene is required for optimal growth under high temperature. The rpoE2 and RpoE2-dependent pgpD gene encoding a periplasmic glutathione peroxidase are involved in oxidative stress responses. But rpoE2 is not required for bacterial growth at low temperature and it even affected bacterial growth under salt stress, indicating that there is a tradeoff between the salt resistance and RpoE2-mediated oxidative stress responses.« less

An extracytoplasmic function sigma factor-dependent periplasmic glutathione peroxidase is involved in oxidative stress response of Shewanella oneidensis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dai, Jingcheng; Wei, Hehong; Tian, Chunyuan

Background: Bacteria use alternative sigma factors (σs) to regulate condition-specific gene expression for survival and Shewanella harbors multiple ECF (extracytoplasmic function) σ genes and cognate anti-sigma factor genes. Here we comparatively analyzed two of the rpoE-like operons in the strain MR-1: rpoE-rseA-rseB-rseC and rpoE2-chrR. Results: RpoE was important for bacterial growth at low and high temperatures, in the minimal medium, and high salinity. The degP/htrA orthologue, required for growth of Escherichia coli and Pseudomonas aeruginosa at high temperature, is absent in Shewanella, while the degQ gene is RpoE-regulated and is required for bacterial growth at high temperature. RpoE2 was essentialmore » for the optimal growth in oxidative stress conditions because the rpoE2 mutant was sensitive to hydrogen peroxide and paraquat. The operon encoding a ferrochelatase paralogue (HemH2) and a periplasmic glutathione peroxidase (PgpD) was identified as RpoE2-dependent. PgpD exhibited higher activities and played a more important role in the oxidative stress responses than the cytoplasmic glutathione peroxidase CgpD under tested conditions. The rpoE2-chrR operon and the identified regulon genes, including pgpD and hemH2, are coincidently absent in several psychrophilic and/or deep-sea Shewanella strains. Conclusion: In S. oneidensis MR-1, the RpoE-dependent degQ gene is required for optimal growth under high temperature. The rpoE2 and RpoE2-dependent pgpD gene encoding a periplasmic glutathione peroxidase are involved in oxidative stress responses. But rpoE2 is not required for bacterial growth at low temperature and it even affected bacterial growth under salt stress, indicating that there is a tradeoff between the salt resistance and RpoE2-mediated oxidative stress responses.« less

Effect of cyanamide on toxicity and glutathione depletion in rat hepatocyte cultures: differences between two dichloropropanol isomers.

PubMed

Hammond, A H; Fry, J R

1999-09-30

The effect of aldehyde dehydrogenase inhibition by cyanamide pre-treatment in vitro on dichloropropanol-dependent toxicity and glutathione depletion was investigated in 24 h rat hepatocyte cultures. Cyanamide pre-treatment had no effect on nitrophenol hydroxylase, 7-methoxy-, 7-ethoxy- or 7-benzyloxyresorufin O-dealkylase activities in 24 h cultures from untreated rats, and had no effect on intracellular glutathione content in cultures from untreated rats, or in cultures from isoniazid-treated rats in which cytochrome P4502E1 (CYP2E1) is increased. In cultures from untreated animals the primary alcohol, 2,3-dichloropropanol, was not toxic and did not significantly deplete glutathione. Cyanamide pre-treatment however, potentiated both toxicity and glutathione depletion. Induction of CYP2E1 also potentiated the toxicity of 2,3-dichloropropanol, and in these cultures cyanamide pre-treatment significantly increased both toxicity and glutathione depletion. Cyanamide did not alter the toxicity or glutathione depletion due to the secondary alcohol, 1,3-dichloropropanol, irrespective of CYP2E1 induction. These results indicate that the primary alcohol isomer is metabolised to an aldehyde intermediate which depletes glutathione. Under basal conditions this metabolite appears to be effectively detoxified, but increased CYP2E1 activity and/or decreased aldehyde dehydrogenase activity promotes accumulation of metabolite, and therefore increases glutathione depletion and toxicity.

Blood selenium concentrations and enzyme activities related to glutathione metabolism in wild emperor geese

USGS Publications Warehouse

Franson, J. Christian; Hoffman, David J.; Schmutz, Joel A.

2002-01-01

In 1998, we collected blood samples from 63 emperor geese (Chen canagica) on their breeding grounds on the Yukon-Kuskokwim Delta (YKD) in western Alaska, USA. We studied the relationship between selenium concentrations in whole blood and the activities of glutathione peroxidase and glutathione reductase in plasma. Experimental studies have shown that plasma activities of these enzymes are useful biomarkers of selenium-induced oxidative stress, but little information is available on their relationship to selenium in the blood of wild birds. Adult female emperor geese incubating their eggs in mid-June had a higher mean concentration of selenium in their blood and a greater activity of glutathione peroxidase in their plasma than adult geese or goslings that were sampled during the adult flight feathermolting period in late July and early August. Glutathione peroxidase activity was positively correlated with the concentration of selenium in the blood of emperor geese, and the rate of increase relative to selenium was greater in goslings than in adults. The activity of glutathione reductase was greatest in the plasma of goslings and was greater in molting adults than incubating females but was not significantly correlated with selenium in the blood of adults or goslings. Incubating female emperor geese had high selenium concentrations in their blood, accompanied by increased glutathione peroxidase activity consistent with early oxidative stress. These findings indicate that further study of the effects of selenium exposure, particularly on reproductive success, is warranted in this species.

An energy-saving glutathione production method from low-temperature cooked rice using amylase-expressing Saccharomyces cerevisiae.

PubMed

Hara, Kiyotaka Y; Kim, Songhee; Kiriyama, Kentaro; Yoshida, Hideyo; Arai, Shogo; Ishii, Jun; Ogino, Chiaki; Fukuda, Hideki; Kondo, Akihiko

2012-05-01

Glutathione is a valuable tripeptide that is widely used in the pharmaceutical, food, and cosmetic industries. Glutathione is industrially produced by fermentation using Saccharomyces cerevisiae. Before the glutathione fermentation process with S. cerevisiae, a glucose extraction process from starchy materials is required. This glucose extraction is usually carried out by converting starchy materials to starch using high-temperature cooking and subsequent hydrolysis by amylases to convert starch to glucose. In this study, to develop an energy-saving glutathione production process by reducing energy consumption during the cooking step, we efficiently produced glutathione from low-temperature cooked rice using amylase-expressing S. cerevisiae. The combination of the amylase-expressing yeast with low-temperature cooking is potentially applicable to a variety of energy-saving bio-production methods of chemicals from starchy bio-resources. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Glutathione S-conjugates as prodrugs to target drug-resistant tumors

PubMed Central

Ramsay, Emma E.; Dilda, Pierre J.

2014-01-01

Living organisms are continuously exposed to xenobiotics. The major phase of enzymatic detoxification in many species is the conjugation of activated xenobiotics to reduced glutathione (GSH) catalyzed by the glutathione-S-transferase (GST). It has been reported that some compounds, once transformed into glutathione S-conjugates, enter the mercapturic acid pathway whose end products are highly reactive and toxic for the cell responsible for their production. The cytotoxicity of these GSH conjugates depends essentially on GST and gamma-glutamyl transferases (γGT), the enzymes which initiate the mercapturic acid synthesis pathway. Numerous studies support the view that the expression of GST and γGT in cancer cells represents an important factor in the appearance of a more aggressive and resistant phenotype. High levels of tumor GST and γGT expression were employed to selectively target tumor with GST- or γGT-activated drugs. This strategy, explored over the last two decades, has recently been successful using GST-activated nitrogen mustard (TLK286) and γGT-activated arsenic-based (GSAO and Darinaparsin) prodrugs confirming the potential of GSH-conjugates as anticancer drugs. PMID:25157234

Dorsal Anterior Cingulate Lactate and Glutathione Levels in Euthymic Bipolar I Disorder: 1H-MRS Study.

PubMed

Soeiro-de-Souza, Márcio Gerhardt; Pastorello, Bruno F; Leite, Cláudia da Costa; Henning, Anke; Moreno, Ricardo A; Garcia Otaduy, Maria Concepción

2016-08-01

Oxidative stress and mitochondrial dysfunction are 2 closely integrated processes implicated in the physiopathology of bipolar disorder. Advanced proton magnetic resonance spectroscopy techniques enable the measurement of levels of lactate, the main marker of mitochondrial dysfunction, and glutathione, the predominant brain antioxidant. The objective of this study was to measure brain lactate and glutathione levels in bipolar disorder and healthy controls. Eighty-eight individuals (50 bipolar disorder and 38 healthy controls) underwent 3T proton magnetic resonance spectroscopy in the dorsal anterior cingulate cortex (2x2x4.5cm(3)) using a 2-D JPRESS sequence. Lactate and glutathione were quantified using the ProFit software program. Bipolar disorder patients had higher dorsal anterior cingulate cortex lactate levels compared with controls. Glutathione levels did not differ between euthymic bipolar disorder and controls. There was a positive correlation between lactate and glutathione levels specific to bipolar disorder. No influence of medications on metabolites was observed. This is the most extensive magnetic resonance spectroscopy study of lactate and glutathione in bipolar disorder to date, and results indicated that euthymic bipolar disorder patients had higher levels of lactate, which might be an indication of altered mitochondrial function. Moreover, lactate levels correlated with glutathione levels, indicating a compensatory mechanism regardless of bipolar disorder diagnosis. © The Author 2016. Published by Oxford University Press on behalf of CINP.

Dorsal Anterior Cingulate Lactate and Glutathione Levels in Euthymic Bipolar I Disorder: 1H-MRS Study

PubMed Central

Pastorello, Bruno F.; Leite, Cláudia da Costa; Henning, Anke; Moreno, Ricardo A.; Garcia Otaduy, Maria Concepción

2016-01-01

Objective: Oxidative stress and mitochondrial dysfunction are 2 closely integrated processes implicated in the physiopathology of bipolar disorder. Advanced proton magnetic resonance spectroscopy techniques enable the measurement of levels of lactate, the main marker of mitochondrial dysfunction, and glutathione, the predominant brain antioxidant. The objective of this study was to measure brain lactate and glutathione levels in bipolar disorder and healthy controls. Methods: Eighty-eight individuals (50 bipolar disorder and 38 healthy controls) underwent 3T proton magnetic resonance spectroscopy in the dorsal anterior cingulate cortex (2x2x4.5cm3) using a 2-D JPRESS sequence. Lactate and glutathione were quantified using the ProFit software program. Results: Bipolar disorder patients had higher dorsal anterior cingulate cortex lactate levels compared with controls. Glutathione levels did not differ between euthymic bipolar disorder and controls. There was a positive correlation between lactate and glutathione levels specific to bipolar disorder. No influence of medications on metabolites was observed. Conclusion: This is the most extensive magnetic resonance spectroscopy study of lactate and glutathione in bipolar disorder to date, and results indicated that euthymic bipolar disorder patients had higher levels of lactate, which might be an indication of altered mitochondrial function. Moreover, lactate levels correlated with glutathione levels, indicating a compensatory mechanism regardless of bipolar disorder diagnosis. PMID:27207914

21 CFR 862.1365 - Glutathione test system.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Glutathione test system. 862.1365 Section 862.1365 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862...

21 CFR 862.1365 - Glutathione test system.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Glutathione test system. 862.1365 Section 862.1365 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862...

The evolution of new enzyme function: lessons from xenobiotic metabolizing bacteria versus insecticide-resistant insects

PubMed Central

Russell, Robyn J; Scott, Colin; Jackson, Colin J; Pandey, Rinku; Pandey, Gunjan; Taylor, Matthew C; Coppin, Christopher W; Liu, Jian-Wei; Oakeshott, John G

2011-01-01

Here, we compare the evolutionary routes by which bacteria and insects have evolved enzymatic processes for the degradation of four classes of synthetic chemical insecticide. For insects, the selective advantage of such degradative activities is survival on exposure to the insecticide, whereas for the bacteria the advantage is simply a matter of access to additional sources of nutrients. Nevertheless, bacteria have evolved highly efficient enzymes from a wide variety of enzyme families, whereas insects have relied upon generalist esterase-, cytochrome P450- and glutathione-S-transferase-dependent detoxification systems. Moreover, the mutant insect enzymes are less efficient kinetically and less diverged in sequence from their putative ancestors than their bacterial counterparts. This presumably reflects several advantages that bacteria have over insects in the acquisition of new enzymatic functions, such as a broad biochemical repertoire from which new functions can be evolved, large population sizes, high effective mutation rates, very short generation times and access to genetic diversity through horizontal gene transfer. Both the insect and bacterial systems support recent theory proposing that new biochemical functions often evolve from ‘promiscuous’ activities in existing enzymes, with subsequent mutations then enhancing those activities. Study of the insect enzymes will help in resistance management, while the bacterial enzymes are potential bioremediants of insecticide residues in a range of contaminated environments. PMID:25567970

METABOLISM OF 1,1- AND 1,3- DICHLOROPROPENE: A MECHANISM OF BIOACTIVATION BY GLUTATHIONE

EPA Science Inventory

Glutathione transferases (GST) catalyze the reaction of glutathione (GSH) with haloalkenes via a nucleophilic vinylic substitution mechanism (SNV reaction). The source water contaminants 1,1-dichloropropene and 1,3-dichloropropene, which are under scrutiny by the U.S.EPA, were...

Assessment of glutathione levels in model solution and grape ferments supplemented with glutathione-enriched inactive dry yeast preparations using a novel UPLC-MS/MS method.

PubMed

Kritzinger, E C; Stander, M A; Du Toit, W J

2013-01-01

A novel, robust and fast ultra-high performance liquid chromatography-MS method has been developed for the simultaneous quantification of reduced glutathione (GSH) and oxidised glutathione (GSSG) in grape juice, wine and model wine solution. Sample preparation is minimal and does not require derivatisation. The method has very good performance in terms of sensitivity and selectivity. The limit of detection was 0.002 and 0.001 mg L(-1) for GSH and GSSG, respectively. The amount of GSH and GSSG released by commercial glutathione-enriched inactivated dry yeast preparations (GSH-IDYs) into a model solution was assessed. Significant differences in the amount of GSH and/or GSSG released into a model wine by different GSH-IDYs were observed, with ethanol influencing this release under certain conditions. The GSH and GSSG levels in grape juice fermentations supplemented with GSH-IDY were also assessed in relation to different addition times during fermentation. GSH-IDY addition can lead to elevated wine GSH levels, provided the supplementation is done early during alcoholic fermentation.

Impact of glutathione supplementation of parenteral nutrition on hepatic methionine adenosyltransferase activity.

PubMed

Elremaly, Wesam; Mohamed, Ibrahim; Rouleau, Thérèse; Lavoie, Jean-Claude

2016-08-01

The oxidation of the methionine adenosyltransferase (MAT) by the combined impact of peroxides contaminating parenteral nutrition (PN) and oxidized redox potential of glutathione is suspected to explain its inhibition observed in animals. A modification of MAT activity is suspected to be at origin of the PN-associated liver disease as observed in newborns. We hypothesized that the correction of redox potential of glutathione by adding glutathione in PN protects the MAT activity. To investigate whether the addition of glutathione to PN can reverse the inhibition of MAT observed in animal on PN. Three days old guinea pigs received through a jugular vein catheter 2 series of solutions. First with methionine supplement, (1) Sham (no infusion); (2) PN: amino acids, dextrose, lipids and vitamins; (3) PN-GSSG: PN+10μM GSSG. Second without methionine, (4) D: dextrose; (5) D+180μM ascorbylperoxide; (6) D+350μM H2O2. Four days later, liver was sampled for determination of redox potential of glutathione and MAT activity in the presence or absence of 1mM DTT. Data were compared by ANOVA, p<0.05. MAT activity was 45±4% lower in animal infused with PN and 23±7% with peroxides generated in PN. The inhibition by peroxides was associated with oxidized redox potential and was reversible by DTT. Correction of redox potential (PN+GSSG) or DTT was without effect on the inhibition of MAT by PN. The slope of the linear relation between MAT activity and redox potential was two fold lower in animal infused with PN than in others groups. The present study suggests that prevention of peroxide generation in PN and/or correction of the redox potential by adding glutathione in PN are not sufficient, at least in newborn guinea pigs, to restore normal MAT activity. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Gastric spiral bacteria in small felids.

PubMed

Kinsel, M J; Kovarik, P; Murnane, R D

1998-06-01

Nine small cats, including one bobcat (Felis rufus), one Pallas cat (F. manul), one Canada lynx (F. lynx canadensis), two fishing cats (F. viverrina), two margays (F. wiedii), and two sand cats (F. margarita), necropsied between June 1995 and March 1997 had large numbers of gastric spiral bacteria, whereas five large cats, including one African lion (Panthera leo), two snow leopards (P. uncia), one Siberian tiger (P. tigris altaica), and one jaguar (P. onca), necropsied during the same period had none. All of the spiral organisms from the nine small cats were histologically and ultrastructurally similar. Histologically, the spiral bacteria were 5-14 microm long with five to nine coils per organism and were located both extracellularly within gastric glands and surface mucus, and intracellularly in parietal cells. Spiral bacteria in gastric mucosal scrapings from the Canada lynx, one fishing cat, and the two sand cats were gram negative and had corkscrewlike to tumbling motility when viewed with phase contrast microscopy. The bacteria were 0.5-0.7 microm wide, with a periodicity of 0.65-1.1 microm in all cats. Bipolar sheathed flagella were occasionally observed, and no periplasmic fibrils were seen. The bacteria were extracellular in parietal cell canaliculi and intracellular within parietal cells. Culture of mucosal scrapings from the Canada lynx and sand cats was unsuccessful. Based on morphology, motility, and cellular tropism, the bacteria were probably Helicobacter-like organisms. Although the two margays had moderate lymphoplasmacytic gastritis, the other cats lacked or had only mild gastric lymphoid infiltrates, suggesting that these organisms are either commensals or opportunistic pathogens.

Reduced glutathione and glutathione disulfide in the blood of glucose-6-phosphate dehydrogenase-deficient newborns.

PubMed

Gong, Zhen-Hua; Tian, Guo-Li; Huang, Qi-Wei; Wang, Yan-Min; Xu, Hong-Ping

2017-07-20

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is commonly detected during mass screening for neonatal disease. We developed a method to measure reduced glutathione (GSH) and glutathione disulfide (GSSG) using tandem mass spectrometry (MS/MS) for detecting G6PD deficiency. The concentration of GSH and the GSH/GSSG ratio in newborn dry-blood-spot (DBS) screening and in blood plus sodium citrate for test confirmation were examined by MS/MS using labeled glycine as an internal standard. G6PD-deficient newborns had a lower GSH content (242.9 ± 15.9 μmol/L)and GSH/GSSG ratio (14.9 ± 7.2) than neonatal controls (370.0 ± 53.2 μmol/L and 46.7 ± 19.6, respectively). Although the results showed a significance of P < 0.001 for DBS samples plus sodium citrate that were examined the first day after preparation, there were no significant differences in the mean GSH concentration and GSH/GSSG ratio between the G6PD deficiency-positive and negative groups when examined three days after sample preparation. The concentration of GSH and the ratio of GSH/GSSG in blood measured using MS/MS on the first day of sample preparation are consistent with G6PD activity and are helpful for diagnosing G6PD deficiency.

The Triple Roles of Glutathione for a DNA-Cleaving DNAzyme and Development of a Fluorescent Glutathione/Cu2+-Dependent DNAzyme Sensor for Detection of Cu2+ in Drinking Water.

PubMed

Wang, Shijin; Liu, Chengcheng; Li, Guiying; Sheng, Yongjie; Sun, Yanhong; Rui, Hongyue; Zhang, Jin; Xu, Jiacui; Jiang, Dazhi

2017-03-24

Pistol-like DNAzyme (PLDz) is an oxidative DNA-cleaving catalytic DNA with ascorbic acid as cofactor. Herein, glutathione was induced into the reaction system to maintain reduced ascorbic acid levels for higher efficient cleavage. However, data indicated that glutathione played triple roles in PLDz-catalyzed reactions. Glutathione alone had no effect on PLDz, and showed inhibitory effect on ascorbic acid-induced PLDz catalysis, but exhibited stimulating effect on Cu 2+ -promoted self-cleavage of PLDz. Further analysis of the effect of glutathione/Cu 2+ on PLDz indicated that H 2 O 2 played a key role in PLDz catalysis. Finally, we developed a fluorescent Cu 2+ sensor (PL-Cu 1.0) based on the relationship between glutathione/Cu 2+ and catalytic activity of PLDz. The fluorescent intensity showed a linear response toward the logarithm concentration of Cu 2+ over the range from 80 nM to 30 μM, with a detection limit of 21.1 nM. PL-Cu 1.0 provided only detection of Cu 2+ over other divalent metal ions. Ca 2+ and Mg 2+ could not interfere with Cu 2+ detection even at a 1000-fold concentration. We further applied PL-Cu 1.0 for Cu 2+ detection in tap and bottled water. Water stored in copper taps overnight had relatively high Cu 2+ concentrations, with a maximum 22.3 μM. Trace Cu 2+ (52.2 nM) in deep spring was detected among the tested bottled water. Therefore, PL-Cu 1.0 is feasible to detect Cu 2+ in drinking water, with a practical application.

Study of Linkage between Glutathione Pathway and the Antibiotic Resistance of Escherichia coli from Patients’ Swabs

PubMed Central

Kominkova, Marketa; Michalek, Petr; Cihalova, Kristyna; Guran, Roman; Cernei, Natalia; Nejdl, Lukas; Smerkova, Kristyna; Dostalova, Simona; Chudobova, Dagmar; Heger, Zbynek; Vesely, Radek; Gumulec, Jaromir; Kynicky, Jindrich; Xhaxhiu, Kledi; Zitka, Ondrej; Adam, Vojtech; Kizek, Rene

2015-01-01

In this work, we focused on the differences between bacterial cultures of E. coli obtained from swabs of infectious wounds of patients compared to laboratory E. coli. In addition, blocking of the protein responsible for the synthesis of glutathione (γ-glutamylcysteine synthase—GCL) using 10 mM buthionine sulfoximine was investigated. Each E. coli showed significant differences in resistance to antibiotics. According to the determined resistance, E. coli were divided into experimental groups based on a statistical evaluation of their properties as more resistant and more sensitive. These groups were also used for finding the differences in a dependence of the glutathione pathway on resistance to antibiotics. More sensitive E. coli showed the same kinetics of glutathione synthesis while blocking GCL (Km 0.1 µM), as compared to non-blocking. In addition, the most frequent mutations in genes of glutathione synthetase, glutathione peroxidase and glutathione reductase were observed in this group compared to laboratory E.coli. The group of “more resistant” E. coli exhibited differences in Km between 0.3 and 0.8 µM. The number of mutations compared to the laboratory E. coli was substantially lower compared to the other group. PMID:25837469

Downregulation of Glutathione Biosynthesis Contributes to Oxidative Stress and Liver Dysfunction in Acute Kidney Injury

PubMed Central

Siow, Yaw L.; Isaak, Cara K.

2016-01-01

Ischemia-reperfusion is a common cause for acute kidney injury and can lead to distant organ dysfunction. Glutathione is a major endogenous antioxidant and its depletion directly correlates to ischemia-reperfusion injury. The liver has high capacity for producing glutathione and is a key organ in modulating local and systemic redox balance. In the present study, we investigated the mechanism by which kidney ischemia-reperfusion led to glutathione depletion and oxidative stress. The left kidney of Sprague-Dawley rats was subjected to 45 min ischemia followed by 6 h reperfusion. Ischemia-reperfusion impaired kidney and liver function. This was accompanied by a decrease in glutathione levels in the liver and plasma and increased hepatic lipid peroxidation and plasma homocysteine levels. Ischemia-reperfusion caused a significant decrease in mRNA and protein levels of hepatic glutamate-cysteine ligase mediated through the inhibition of transcription factor Nrf2. Ischemia-reperfusion inhibited hepatic expression of cystathionine γ-lyase, an enzyme responsible for producing cysteine (an essential precursor for glutathione synthesis) through the transsulfuration pathway. These results suggest that inhibition of glutamate-cysteine ligase expression and downregulation of the transsulfuration pathway lead to reduced hepatic glutathione biosynthesis and elevation of plasma homocysteine levels, which, in turn, may contribute to oxidative stress and distant organ injury during renal ischemia-reperfusion. PMID:27872680

The photochemical thiol–ene reaction as a versatile method for the synthesis of glutathione S-conjugates targeting the bacterial potassium efflux system Kef† †Electronic supplementary information (ESI) available: Further experimental details and NMR spectra. See DOI: 10.1039/c5qo00436e Click here for additional data file.

PubMed Central

Rasmussen, Tim; Miller, Samantha; Booth, Ian R.

2016-01-01

The thiol–ene coupling reaction is emerging as an important conjugation reaction that is suitable for use in a biological setting. Here, we explore the utility of this reaction for the synthesis of glutathione-S-conjugates (GSX) and present a general, operationally simple, protocol with a wide substrate scope. The GSX afforded are an important class of compounds and provide invaluable molecular tools to study glutathione-binding proteins. In this study we apply the diverse library of GSX synthesised to further our understanding of the structural requirements for binding to the glutathione-binding protein, Kef, a bacterial K+ efflux system, found in many bacterial pathogens. This system is vital to the survival of bacteria upon exposure to electrophiles, and plays an essential role in the maintenance of intracellular pH and K+ homeostasis. Consequently, Kef is an appealing target for the development of novel antibacterial drugs. PMID:27110363

Antioxidant Protection of NADPH-Depleted Oligodendrocyte Precursor Cells Is Dependent on Supply of Reduced Glutathione.

PubMed

Kilanczyk, Ewa; Saraswat Ohri, Sujata; Whittemore, Scott R; Hetman, Michal

2016-08-01

The pentose phosphate pathway is the main source of NADPH, which by reducing oxidized glutathione, contributes to antioxidant defenses. Although oxidative stress plays a major role in white matter injury, significance of NADPH for oligodendrocyte survival has not been yet investigated. It is reported here that the NADPH antimetabolite 6-amino-NADP (6AN) was cytotoxic to cultured adult rat spinal cord oligodendrocyte precursor cells (OPCs) as well as OPC-derived oligodendrocytes. The 6AN-induced necrosis was preceded by increased production of superoxide, NADPH depletion, and lower supply of reduced glutathione. Moreover, survival of NADPH-depleted OPCs was improved by the antioxidant drug trolox. Such cells were also protected by physiological concentrations of the neurosteroid dehydroepiandrosterone (10(-8) M). The protection by dehydroepiandrosterone was associated with restoration of reduced glutathione, but not NADPH, and was sensitive to inhibition of glutathione synthesis. A similar protective mechanism was engaged by the cAMP activator forskolin or the G protein-coupled estrogen receptor (GPER/GPR30) ligand G1. Finally, treatment with the glutathione precursor N-acetyl cysteine reduced cytotoxicity of 6AN. Taken together, NADPH is critical for survival of OPCs by supporting their antioxidant defenses. Consequently, injury-associated inhibition of the pentose phosphate pathway may be detrimental for the myelination or remyelination potential of the white matter. Conversely, steroid hormones and cAMP activators may promote survival of NADPH-deprived OPCs by increasing a NADPH-independent supply of reduced glutathione. Therefore, maintenance of glutathione homeostasis appears as a critical effector mechanism for OPC protection against NADPH depletion and preservation of the regenerative potential of the injured white matter. © The Author(s) 2016.

Is antioxidant potential of the mitochondrial targeted ubiquinone derivative MitoQ conserved in cells lacking mtDNA?

PubMed

Lu, Chao; Zhang, Dawei; Whiteman, Matthew; Armstrong, Jeffrey S

2008-03-01

MitoQ has been developed as a mitochondrial targeted antioxidant for diseases associated with oxidative stress. Here we show that MitoQ blocks the generation of reactive oxygen species (ROS) and mitochondrial protein thiol oxidation, and preserves mitochondrial function and ultrastructure after glutathione (GSH) depletion. Furthermore, the antioxidant effect of MitoQ is conserved in cells lacking mitochondrial DNA, indicating that its antioxidant properties do not depend on a functional electron transport chain (ETC). Our results elucidate the antioxidant mechanism of MitoQ and suggest that it may be a useful therapeutic for disorders associated with a dysfunctional ETC and increased ROS production.

α-Lipoic acid protects against the cytotoxicity and oxidative stress induced by cadmium in HepG2 cells through regeneration of glutathione by glutathione reductase via Nrf2/ARE signaling pathway.

PubMed

Shi, Chunli; Zhou, Xue; Zhang, Jiayu; Wang, Jiachun; Xie, Hong; Wu, Zhigang

2016-07-01

α-Lipoic acid (α-LA) is a potent natural antioxidant, which is capable of regenerating glutathione (GSH). However, the mechanisms by which α-LA regenerates reduced glutathione (rGSH) via the reduction of oxidized glutathione (GSSG) by glutathione reductase (GR) are still not well understood. In the present study, we investigated if α-LA replenished rGSH by GR via Nrf2/ARE signaling pathway in cadmium-treated HepG2 cells. We found that α-LA antagonized the oxidative damage and alleviated the cytotoxicity in cadmium-induced HepG2 cells by regeneration of rGSH. α-LA regenerated rGSH by activating Nrf2 signaling pathway via promoting the nuclear translocation of Nrf2, which upregulates the transcription of GR, and thus increased the activity of GR. Our results indicated that α-LA was an effective agent to antagonize the oxidative stress and alleviate the cytotoxicity in cadmium-treated HepG2 cells by regenerating rGSH through activating Nrf2 signaling pathway. Copyright © 2016. Published by Elsevier B.V.

Luminous bacteria cultured from fish guts in the Gulf of Oman.

PubMed

Makemson, J C; Hermosa, G V

1999-01-01

The incidence of culturable luminous bacteria in Omani market fish guts was correlated to habitat type amongst 109 species of fish. Isolated representative luminous bacteria were compared to known species using the Biolog system (95 traits/isolate) and cluster analysis, which showed that the main taxa present in fish guts were clades related to Vibrio harveyi and Photobacterium species with sporadic incidence of P. phosphoreum. The luminous isolates from gut of the slip-mouth (barred pony fish), Leiognathus fasciatus, were mainly a type related to Photobacterium but phenotypically different from known species. These luminous gut bacteria were identical with the bacteria in the light organ, indicating that the light organ supplies a significant quantity of luminous bacteria to the gut. In many of the fish that lack light organs, luminous bacteria were also the dominant bacterial type in the gut, while in some others luminous bacteria were encountered sporadically and at low densities, reflecting the incidence of culturable luminous bacteria in seawater. Pelagic fish contained the highest incidence of culturable luminous bacteria and reef-associated fish the lowest. No correlation was found between the incidence of culturable luminous bacteria and the degree to which fish produce a melanin-covered gut. Copyright 1999 John Wiley & Sons, Ltd.

Microbial β-etherases and glutathione lyases for lignin valorisation in biorefineries: current state and future perspectives.

PubMed

Husarcíková, Jana; Voß, Hauke; Domínguez de María, Pablo; Schallmey, Anett

2018-05-04

Lignin is the major aromatic biopolymer in nature, and it is considered a valuable feedstock for the future supply of aromatics. Hence, its valorisation in biorefineries is of high importance, and various chemical and enzymatic approaches for lignin depolymerisation have been reported. Among the enzymes known to act on lignin, β-etherases offer the possibility for a selective cleavage of the β-O-4 aryl ether bonds present in lignin. These enzymes, together with glutathione lyases, catalyse a reductive, glutathione-dependent ether bond cleavage displaying high stereospecificity. β-Etherases and glutathione lyases both belong to the superfamily of glutathione transferases, and several structures have been solved recently. Additionally, different approaches for their application in lignin valorisation have been reported in the last years. This review gives an overview on the current knowledge on β-etherases and glutathione lyases, their biochemical and structural features, and critically discusses their potential for application in biorefineries.

Oxidative stress protection and glutathione metabolism in response to hydrogen peroxide and menadione in riboflavinogenic fungus Ashbya gossypii.

PubMed

Kavitha, S; Chandra, T S

2014-11-01

Ashbya gossypii is a plant pathogen and a natural overproducer of riboflavin and is used for industrial riboflavin production. A few literature reports depict a link between riboflavin overproduction and stress in this fungus. However, the stress protection mechanisms and glutathione metabolism are not much explored in A. gossypii. In the present study, an increase in the activity of catalase and superoxide dismutase was observed in response to hydrogen peroxide and menadione. The lipid peroxide and membrane lipid peroxide levels were increased by H2O2 and menadione, indicating oxidative damage. The glutathione metabolism was altered with a significant increase in oxidized glutathione (GSSG), glutathione peroxidase (GPX), glutathione S transferase (GST), and glutathione reductase (GR) and a decrease in reduced glutathione (GSH) levels in the presence of H2O2 and menadione. Expression of the genes involved in stress mechanism was analyzed in response to the stressors by semiquantitative RT-PCR. The messenger RNA (mRNA) levels of CTT1, SOD1, GSH1, YAP1, and RIB3 were increased by H2O2 and menadione, indicating the effect of stress at the transcriptional level. A preliminary bioinformatics study for the presence of stress response elements (STRE)/Yap response elements (YRE) depicted that the glutathione metabolic genes, stress genes, and the RIB genes hosted either STRE/YRE, which may enable induction of these genes during stress.

A Dispersion-Dominated Chromogenic Strategy for Colorimetric Sensing of Glutathione at the Nanomolar Level Using Gold Nanoparticles.

PubMed

Xianyu, Yunlei; Xie, Yangzhouyun; Wang, Nuoxin; Wang, Zhuo; Jiang, Xingyu

2015-11-04

A dispersion-dominated chromogenic strategy for glutathione sensing is developed. Glutathione prevents the aggregation of arginine-modified gold nanoparticles via mercury-thiol interaction, which allows for glutathione sensing at the nanomolar level (10.9 × 10(-9) m) with facile operation and naked-eye readout. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Bcl-2 is a novel interacting partner for the 2-oxoglutarate carrier and a key regulator of mitochondrial glutathione

PubMed Central

Wilkins, Heather M.; Marquardt, Kristin; Lash, Lawrence H.; Linseman, Daniel A.

2011-01-01

Despite making up only a minor fraction of the total cellular glutathione, recent studies indicate that the mitochondrial glutathione pool is essential for cell survival. Selective depletion of mitochondrial glutathione is sufficient to sensitize cells to mitochondrial oxidative stress (MOS)1 and intrinsic apoptosis. Glutathione is synthesized exclusively in the cytoplasm and must be actively transported into mitochondria. Therefore, regulation of mitochondrial glutathione transport is a key factor in maintaining the antioxidant status of mitochondria. Bcl-2 is resident in the outer mitochondrial membrane where it acts as a central regulator of the intrinsic apoptotic cascade. In addition, Bcl-2 displays an antioxidant-like function that has been linked experimentally to the regulation of cellular glutathione content. We have previously demonstrated a novel interaction between recombinant Bcl-2 and reduced glutathione (GSH) which was antagonized by either Bcl-2 homology-3 domain (BH3) mimetics or a BH3-only protein, recombinant Bim. These previous findings prompted us to investigate if this novel Bcl-2/GSH interaction might play a role in regulating mitochondrial glutathione transport. Incubation of primary cultures of cerebellar granule neurons (CGNs) with the BH3 mimetic, HA14-1, induced MOS and caused specific depletion of the mitochondrial glutathione pool. Bcl-2 was co-immunoprecipitated with GSH following chemical cross-linking in CGNs and this Bcl-2/GSH interaction was antagonized by pre-incubation with HA14-1. Moreover, both HA14-1 and recombinant Bim inhibited GSH transport into isolated rat brain mitochondria. To further investigate a possible link between Bcl-2 function and mitochondrial glutathione transport, we next examined if Bcl-2 associated with the 2-oxoglutarate carrier (OGC), an inner mitochondrial membrane protein known to transport glutathione in liver and kidney. Following co-transfection of CHO cells, Bcl-2 was co-immunoprecipitated with OGC

High Glutathione and Glutathione Peroxidase-2 Levels Mediate Cell-Type-Specific DNA Damage Protection in Human Induced Pluripotent Stem Cells

PubMed Central

Dannenmann, Benjamin; Lehle, Simon; Hildebrand, Dominic G.; Kübler, Ayline; Grondona, Paula; Schmid, Vera; Holzer, Katharina; Fröschl, Mirjam; Essmann, Frank; Rothfuss, Oliver; Schulze-Osthoff, Klaus

2015-01-01

Summary Pluripotent stem cells must strictly maintain genomic integrity to prevent transmission of mutations. In human induced pluripotent stem cells (iPSCs), we found that genome surveillance is achieved via two ways, namely, a hypersensitivity to apoptosis and a very low accumulation of DNA lesions. The low apoptosis threshold was mediated by constitutive p53 expression and a marked upregulation of proapoptotic p53 target genes of the BCL-2 family, ensuring the efficient iPSC removal upon genotoxic insults. Intriguingly, despite the elevated apoptosis sensitivity, both mitochondrial and nuclear DNA lesions induced by genotoxins were less frequent in iPSCs compared to fibroblasts. Gene profiling identified that mRNA expression of several antioxidant proteins was considerably upregulated in iPSCs. Knockdown of glutathione peroxidase-2 and depletion of glutathione impaired protection against DNA lesions. Thus, iPSCs ensure genomic integrity through enhanced apoptosis induction and increased antioxidant defense, contributing to protection against DNA damage. PMID:25937369

Organophosphorus insecticides chlorpyrifos and diazinon and oxidative stress in neuronal cells in a genetic model of glutathione deficiency

DOE Office of Scientific and Technical Information (OSTI.GOV)

Giordano, Gennaro; Afsharinejad, Zhara; Guizzetti, Marina

2007-03-15

Over the past several years evidence has been accumulating from in vivo animal studies, observations in humans, and in vitro studies, that organophosphorus (OP) insecticides may induce oxidative stress. Such effects may contribute to some of the toxic manifestations of OPs, particularly upon chronic or developmental exposures. The aim of this study was to investigate the role of oxidative stress in the neurotoxicity of two commonly used OPs, chlorpyrifos (CPF) and diazinon (DZ), their oxygen analogs (CPO and DZO), and their 'inactive' metabolites (TCP and IMP), in neuronal cells from a genetic model of glutathione deficiency. Cerebellar granule neurons frommore » wild type mice (Gclm +/+) and mice lacking the modifier subunit of glutamate cysteine ligase (Gclm -/-), the first and limiting step in the synthesis of glutathione (GSH), were utilized. The latter display very low levels of GSH and are more susceptible to the toxicity of agents that increase oxidative stress. CPO and DZO were the most cytotoxic compounds, followed by CPF and DZ, while TCP and IMP displayed lower toxicity. Toxicity was significantly higher (10- to 25-fold) in neurons from Gclm (-/-) mice, and was antagonized by various antioxidants. Depletion of GSH from Gclm (+/+) neurons significantly increased their sensitivity to OP toxicity. OPs increased intracellular levels of reactive oxygen species and lipid peroxidation and in both cases the effects were greater in neurons from Gclm (-/-) mice. OPs did not alter intracellular levels of GSH, but significantly increased those of oxidized glutathione (GSSG). Cytotoxicity was not antagonized by cholinergic antagonists, but was decreased by the calcium chelator BAPTA-AM. These studies indicate that cytotoxicity of OPs involves generation of reactive oxygen species and is modulated by intracellular GSH, and suggest that it may involve disturbances in intracellular homeostasis of calcium.« less

Structure-activity relationships for the impact of selected isothiazol-3-one biocides on glutathione metabolism and glutathione reductase of the human liver cell line Hep G2.

PubMed

Arning, Jürgen; Dringen, Ralf; Schmidt, Maike; Thiessen, Anette; Stolte, Stefan; Matzke, Marianne; Bottin-Weber, Ulrike; Caesar-Geertz, Birgit; Jastorff, Bernd; Ranke, Johannes

2008-04-18

To investigate the toxic mode of action of isothiazol-3-one biocides the four compounds N-methylisothiazol-3-one (MIT), 5-chloro-N-methylisothiazol-3-one (CIT), N-octylisothiazol-3-one (OIT) and 4,5-dichloro-N-octylisothiazol-3-one (DCOIT) were purified and tested as single chemical entities for their effects on the human hepatoblastoma cell line Hep G2 and on isolated and cellular glutathione reductase GR). The two chlorinated substances CIT and DCOIT significantly decreased the amount of total cellular glutathione (GSx) in a dose and time dependent manner. Concomitantly, an increase in the level of oxidised glutathione (GSSG) was observed. The resulting shift in the GSH/GSSG ratio entailing the breakdown of the cellular thiol reduction potential was accompanied by necrotic morphological changes like swelling of the plasma membrane and subsequent lysis of the cells. Additionally, CIT and DCOIT were found to inhibit cellular GR in the cells in a concentration dependent manner. The T-SAR-based (thinking in terms of structure-activity relationships) comparison of the chlorine-substituted structures CIT and DCOIT with their non-chlorinated and less active analogues MIT and OIT identified the chlorine substituents and the resulting reaction mechanisms to be the key structural mediators of the observed toxic effects. Furthermore, differences in the activity of both chlorinated substances could be explained using the T-SAR approach to link the lipophilicity and the intrinsic glutathione-reactivity of the compounds to the expected target site concentrations inside the cells.

Glutamine drives glutathione synthesis and contributes to radiation sensitivity of A549 and H460 lung cancer cell lines

PubMed Central

Sappington, Daniel R.; Siegel, Eric R.; Hiatt, Gloria; Desai, Abhishek; Penney, Rosalind B.; Jamshidi-Parsian, Azemat; Griffin, Robert J.; Boysen, Gunnar

2016-01-01

Background Increased glutamine uptake is known to drive cancer cell proliferation, making tumor cells glutamine-dependent. Glutamine provides additional carbon and nitrogen sources for cell growth. The first step in glutamine utilization is its conversion to glutamate by glutaminase (GLS). Glutamate is a precursor for glutathione synthesis, and we investigated the hypothesis that glutamine drives glutathione synthesis and thereby contributes to cellular defense systems. Methods The importance of glutamine for glutathione synthesis was studied in H460 and A549 lung cancer cell lines using glutamine-free medium and Bis-2-(5-phenyl-acetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide (BPTES) a GLS inhibitor. Metabolic activities were determined by targeted mass spectrometry. Results A significant correlation between glutamine consumption and glutathione excretion was demonstrated in H460 and A549 tumor cells. Culturing in the presence of [13C5]glutamine demonstrated that by 12 hrs >50% of excreted glutathione is derived from glutamine. Culturing in glutamine-free medium or treatment with BPTES, a glutaminase (GLS)-specific inhibitor, reduced cell proliferation and viability, and abolished glutathione excretion. Treatment with glutathione-ester prevented BPTES induced cytotoxicity. Inhibition of GLS markedly radiosensitized the lung tumor cell lines, suggesting an important role of glutamine-derived glutathione in determining radiation sensitivity. Conclusions We demonstrate here for the first time that a significant amount of extracellular glutathione is directly derived from glutamine. This finding adds yet another important function to the already known glutamine dependence of tumor cells and probably tumors as well. General significance Glutamine is essential for synthesis and excretion of glutathione to promote cell growth and viability. PMID:26825773

Rice protein improves oxidative stress by regulating glutathione metabolism and attenuating oxidative damage to lipids and proteins in rats.

PubMed

Yang, Lin; Chen, Jia-Hou; Xu, Tong; Zhou, Ai-Shen; Yang, Hong-Kun

2012-10-05

To evaluate the effects of rice protein (RP) on glutathione metabolism and oxidative damage. Seven-week-old male Wistar rats were fed diets containing casein and RP without cholesterol for 3weeks. Plasma and liver lipid levels, hepatic accumulation of total glutathione (T-GSH), oxidized glutathione (GSSG), reduced glutathione (GSH), malondialdehyde (MDA) and protein carbonyl (PCO) were measured. In the liver, the total antioxidative capacity (T-AOC), mRNA levels of glutamate cysteine ligase catalytic subunit (GCLC) and glutamate cysteine ligase modulatory subunit (GCLM), and the activities of hepatic catalase (CAT), total superoxide dismutase (T-SOD), γ-glutamylcysteine synthetase (γ-GCS), glutathione S-transferase (GST), glutathione reductase (GR) and glutathione peroxidase (GSHPx) were also measured. T-AOC, GCLC and GCLM mRNA levels, antioxidative enzyme activities (T-SOD and CAT) and glutathione metabolism related enzyme activities (γ-GCS, GST, GR and GSHPx) were effectively stimulated by RP feeding compared to casein, and RP significantly reduced the hepatic accumulation of MDA and PCO in rats. These results indicate that lipid-lowering activity was induced by RP feeding. The present study demonstrates that RP improves oxidative stress primarily through enzymatic and non-enzymatic antioxidative defense mechanisms, reflected by enhancing the antioxidative status and attenuating the oxidative damage to lipids and proteins. These results suggest that RP can prevent hyperlipidemia in part through modifying glutathione metabolism, and sulfur amino acids may be the main modulator of this antioxidative mechanism. Copyright © 2012 Elsevier Inc. All rights reserved.

REACTION OF BENZENE OXIDE WITH THIOLS INCLUDING GLUTATHIONE

EPA Science Inventory

This study accounts for the observations that the metabolism of benzene is dominated by the formation of phenol. As demonstrated here, the pathway leading to S-phenylmercapturic acid is necessarily minor on account of the low efficiency of benzene oxide capture by glutathione at ...

The effect of prolonged oral contraceptive steroid use on erythrocyte glutathione peroxidase activity.

PubMed

Capel, I D; Jenner, M; Williams, D C; Donaldson, D; Nath, A

1981-08-01

A clinical study was undertaken to determine whether oral contraceptives (OCs) affect the activity of the enzyme glutathione peroxidase. OC users recruited for the study were volunteers attending the Redhill Family Planning Clinic in England. Their demographic characteristics were noted. Pre- and postmenopausal comparative subjects were also used. The laboratory procedures involved in the study are described. Findings are tabulated. The average erythrocyte glutathione peroxidase levels of women using OCs for more than 7 months were significantly higher than those of the pre- and postmenopausal subjects. These levels increased progressively with duration of OC use. These levels did not fluctuate with the menstrual cycle in either OC or non-OC users. Levels of erythrocyte selenium and plasma pyridoxal were not significantly altered by OC use. Riboflavin status, however, as estimated by glutathione reductase activity was substantially lower in OC users and was lowest in women who had used OCs for the longest amount of time. Riboflavin status was found to be directly correlated with erythrocyte glutathione peroxidase levels. These findings may be important because selenium is currently believed to offer protective benefits against carcinogenesis, especially breast cancer. All the OCs studied produced the same effects.

Semen Quality of Post-Thawed Local Ram’s in Tris-Egg Yolk Extender with Different Glutathione Level

NASA Astrophysics Data System (ADS)

Solihati, N.; Rasad, S. D.; Setiawan, R.; Foziah, E. N.; Wigiyanti, E. T.

2018-02-01

The aims of this research were to find out the effect of glutathione level on semen quality of local ram and to find out the glutathione level that provide the best quality of local ram semen. This research use Completely Randomized Design (CRD) with five treatments of glutathione level (0 mM, 4 mM, 5 mM, 6 mM and 7mM) and was repeated for 10 times. Data were analyzed using analysis of varians (ANOVA) and differences between treatment was analyzed using Duncan test. The parameter were semen quality consist of motility, intact plasma membrane (IPM), abnormality, and recovery rate. Result of this research showed that glutathione level significantly (p<0.05) affect on motility, IPM, abnormality and recovery rate. Duncan test result showed that recovery rate from treatment of 5 mM glutathione level have significantly higher than other treatment, whereas for motility, IPM and abnormality were not significant different with 4 mM, and 6 mM. It is concluded that semen quality of local ram was affected by glutathione level, and the level of 5 mM in egg yolk tris extender provide the best quality of local ram semen.

Monobromobimane occupies a distinct xenobiotic substrate site in glutathione S-transferase π

PubMed Central

Ralat, Luis A.; Colman, Roberta F.

2003-01-01

Monobromobimane (mBBr), functions as a substrate of porcine glutathione S-transferase π (GST π): The enzyme catalyzes the reaction of mBBr with glutathione. S-(Hydroxyethyl)bimane, a nonreactive analog of monobromobimane, acts as a competitive inhibitor with respect to mBBr as substrate but does not affect the reaction of GST π with another substrate, 1-chloro-2,4-dinitrobenzene (CDNB). In the absence of glutathione, monobromobimane inactivates GST π at pH 7.0 and 25°C as assayed using mBBr as substrate, with a lesser effect on the enzyme’s use of CDNB as substrate. These results indicate that the sites occupied by CDNB and mBBr are not identical. Inactivation is proportional to the incorporation of 2 moles of bimane/mole of subunit. Modification of GST π with mBBr does not interfere with its binding of 8-anilino-1-naphthalene sulfonate, indicating that this hydrophobic site is not the target of monobromobimane. S-Methylglutathione and S-(hydroxyethyl)bimane each yield partial protection against inactivation and decrease reagent incorporation, while glutathionyl-bimane protects completely against inactivation. Peptide analysis after trypsin digestion indicates that mBBr modifies Cys45 and Cys99 equally. Modification of Cys45 is reduced in the presence of S-methylglutathione, indicating that this residue is at or near the glutathione binding region. In contrast, modification of Cys99 is reduced in the presence of S-(hydroxyethyl)bimane, suggesting that this residue is at or near the mBBr xenobiotic substrate binding site. Modification of Cys99 can best be understood by reaction with monobromobimane while it is bound to its xenobiotic substrate site in an alternate orientation. These results support the concept that glutathione S-transferase accomplishes its ability to react with a diversity of substrates in part by harboring distinct xenobiotic substrate sites. PMID:14573868

Monobromobimane occupies a distinct xenobiotic substrate site in glutathione S-transferase pi.

PubMed

Ralat, Luis A; Colman, Roberta F

2003-11-01

Monobromobimane (mBBr), functions as a substrate of porcine glutathione S-transferase pi (GST pi): The enzyme catalyzes the reaction of mBBr with glutathione. S-(Hydroxyethyl)bimane, a nonreactive analog of monobromobimane, acts as a competitive inhibitor with respect to mBBr as substrate but does not affect the reaction of GST pi with another substrate, 1-chloro-2,4-dinitrobenzene (CDNB). In the absence of glutathione, monobromobimane inactivates GST pi at pH 7.0 and 25 degrees C as assayed using mBBr as substrate, with a lesser effect on the enzyme's use of CDNB as substrate. These results indicate that the sites occupied by CDNB and mBBr are not identical. Inactivation is proportional to the incorporation of 2 moles of bimane/mole of subunit. Modification of GST pi with mBBr does not interfere with its binding of 8-anilino-1-naphthalene sulfonate, indicating that this hydrophobic site is not the target of monobromobimane. S-Methylglutathione and S-(hydroxyethyl)bimane each yield partial protection against inactivation and decrease reagent incorporation, while glutathionyl-bimane protects completely against inactivation. Peptide analysis after trypsin digestion indicates that mBBr modifies Cys45 and Cys99 equally. Modification of Cys45 is reduced in the presence of S-methylglutathione, indicating that this residue is at or near the glutathione binding region. In contrast, modification of Cys99 is reduced in the presence of S-(hydroxyethyl)bimane, suggesting that this residue is at or near the mBBr xenobiotic substrate binding site. Modification of Cys99 can best be understood by reaction with monobromobimane while it is bound to its xenobiotic substrate site in an alternate orientation. These results support the concept that glutathione S-transferase accomplishes its ability to react with a diversity of substrates in part by harboring distinct xenobiotic substrate sites.

Purification and Thermal Dependence of Glutathione Reductase from Two Forage Legume Species 1

PubMed Central

Kidambi, Saranga P.; Mahan, James R.; Matches, Arthur G.

1990-01-01

Alfalfa (Medicago sativa L.) and sainfoin (Onobrychis viciifolia Scop.) are forage legumes that differ in their responses to high and low temperature stresses. Thermal limitations on the function of glutathione reductase (EC 1.6.4.2) could adversely affect the ability of the plant to cope with adverse temperatures. Our objectives were to (a) purify glutathione reductase from `Cimarron' alfalfa and `PI 212241' sainfoin and (b) investigate the intraspecies variation in the thermal dependency of glutathione reductase from each of three cultivars of alfalfa and two cultivars and an introduction of sainfoin. Glutathione reductase was purified 1222-and 1948-fold to a specific activity of 281 and 273 units per milligram of protein, from one species each of alfalfa and sainfoin, respectively. The relative molecular mass of the protein was approximately 140 kilodaltons with subunits of 57 and 37 kilodaltons under denaturing conditions. The activation energies were approximately 50 kilojoules per mole for both species. Over a 5 to 45°C temperature gradient, large variation among species and genotypes within species was found for: (a) the minimum apparent Michaelis constant (0.6-2.1 micromoles of NADPH), (b) the temperature at which the minimum apparent Michaelis constant was observed (10-25°C), and (c) the thermal kinetic windows (6-19°C width). Future studies will focus on relating the thermal dependence of the Michaelis constant of the glutathione reductases and plant growth rates and forage quality of these species throughout the growing season. PMID:16667283

Purification and Biochemical Characterization of Glutathione S-Transferase from Down Syndrome and Normal Children Erythrocytes: A Comparative Study

ERIC Educational Resources Information Center

Hamed, Ragaa R.; Maharem, Tahany M.; Abdel-Meguid, Nagwa; Sabry, Gilane M.; Abdalla, Abdel-Monem; Guneidy, Rasha A.

2011-01-01

Down syndrome (DS) is the phenotypic manifestation of trisomy 21. Our study was concerned with the characterization and purification of glutathione S-transferase enzyme (GST) from normal and Down syndrome (DS) erythrocytes to illustrate the difference in the role of this enzyme in the cell. Glutathione S-transferase and glutathione (GSH) was…

Molecular mimicry between cockroach and helminth glutathione S-transferases promotes cross-reactivity and cross-sensitization

USDA-ARS?s Scientific Manuscript database

The extensive similarities between helminth proteins and allergens are thought to contribute to helminth-driven allergic sensitization. We investigated the molecular and structural similarities between Bla g 5, a major glutathione-S transferase (GST) allergen of cockroaches, and the GST of Wucherer...

Structure-activity relationships of 4-hydroxyalkenals in the conjugation catalysed by mammalian glutathione transferases.

PubMed Central

Danielson, U H; Esterbauer, H; Mannervik, B

1987-01-01

The substrate specificities of 15 cytosolic glutathione transferases from rat, mouse and man have been explored by use of a homologous series of 4-hydroxyalkenals, extending from 4-hydroxypentenal to 4-hydroxypentadecenal. Rat glutathione transferase 8-8 is exceptionally active with the whole range of 4-hydroxyalkenals, from C5 to C15. Rat transferase 1-1, although more than 10-fold less efficient than transferase 8-8, is the second most active transferase with the longest chain length substrates. Other enzyme forms showing high activities with these substrates are rat transferase 4-4 and human transferase mu. The specificity constants, kcat./Km, for the various enzymes have been determined with the 4-hydroxyalkenals. From these constants the incremental Gibbs free energy of binding to the enzyme has been calculated for the homologous substrates. The enzymes responded differently to changes in the length of the hydrocarbon side chain and could be divided into three groups. All glutathione transferases displayed increased binding energy in response to increased hydrophobicity of the substrate. For some of the enzymes, steric limitations of the active site appear to counteract the increase in binding strength afforded by increased chain length of the substrate. Comparison of the activities with 4-hydroxyalkenals and other activated alkenes provides information about the active-site properties of certain glutathione transferases. The results show that the ensemble of glutathione transferases in a given species may serve an important physiological role in the conjugation of the whole range of 4-hydroxyalkenals. In view of its high catalytic efficiency with all the homologues, rat glutathione transferase 8-8 appears to have evolved specifically to serve in the detoxication of these reactive compounds of oxidative metabolism. PMID:3426557

Decreased glutathione levels and impaired antioxidant enzyme activities in drug-naive first-episode schizophrenic patients

PubMed Central

2011-01-01

Background The aim of this study was to determine glutathione levels and antioxidant enzyme activities in the drug-naive first-episode patients with schizophrenia in comparison with healthy control subjects. Methods It was a case-controlled study carried on twenty-three patients (20 men and 3 women, mean age = 29.3 ± 7.5 years) recruited in their first-episode of schizophrenia and 40 healthy control subjects (36 men and 9 women, mean age = 29.6 ± 6.2 years). In patients, the blood samples were obtained prior to the initiation of neuroleptic treatments. Glutathione levels: total glutathione (GSHt), reduced glutathione (GSHr) and oxidized glutathione (GSSG) and antioxidant enzyme activities: superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) were determined by spectrophotometry. Results GSHt and reduced GSHr were significantly lower in patients than in controls, whereas GSSG was significantly higher in patients. GPx activity was significantly higher in patients compared to control subjects. CAT activity was significantly lower in patients, whereas the SOD activity was comparable to that of controls. Conclusion This is a report of decreased plasma levels of GSHt and GSHr, and impaired antioxidant enzyme activities in drug-naive first-episode patients with schizophrenia. The GSH deficit seems to be implicated in psychosis, and may be an important indirect biomarker of oxidative stress in schizophrenia early in the course of illness. Finally, our results provide support for further studies of the possible role of antioxidants as neuroprotective therapeutic strategies for schizophrenia from early stages. PMID:21810251

Effects of mitochondrial poisons on glutathione redox potential and carotid body chemoreceptor activity.

PubMed

Gomez-Niño, A; Agapito, M T; Obeso, A; Gonzalez, C

2009-01-01

Low oxygen sensing in chemoreceptor cells involves the inhibition of specific plasma membrane K(+) channels, suggesting that mitochondria-derived reactive oxygen species (ROS) link hypoxia to K(+) channel inhibition, subsequent cell depolarization and activation of neurotransmitter release. We have used several mitochondrial poisons, alone and in combination with the antioxidant N-acetylcysteine (NAC), and quantify their capacity to alter GSH/GSSG levels and glutathione redox potential (E(GSH)) in rat diaphragm. Selected concentrations of mitochondrial poisons with or without NAC were tested for their capacity to activate neurotransmitter release in chemoreceptor cells and to alter ATP levels in intact rat carotid body (CB). We found that rotenone (1 microM), antimycin A (0.2 microg/ml) and sodium azide (5mM) decreased E(GSH); NAC restored E(GSH) to control values. At those concentrations mitochondrial poisons activated neurotransmitter release from CB chemoreceptor cells and decreased CB ATP levels, NAC being ineffective to modify these responses. Additional experiments with 3-nitroprionate (5mM), lower concentrations of rotenone and dinitrophenol revealed variable relationships between E(GSH) and chemoreceptor cell neurotransmitter release responses and ATP levels. These findings indicate a lack of correlation between mitochondrial-generated modifications of E(GSH) and chemoreceptor cells activity. This lack of correlation renders unlikely that alteration of mitochondrial production of ROS is the physiological pathway chemoreceptor cells use to signal hypoxia.

Glutathione reductase gsr-1 is an essential gene required for Caenorhabditis elegans early embryonic development.

PubMed

Mora-Lorca, José Antonio; Sáenz-Narciso, Beatriz; Gaffney, Christopher J; Naranjo-Galindo, Francisco José; Pedrajas, José Rafael; Guerrero-Gómez, David; Dobrzynska, Agnieszka; Askjaer, Peter; Szewczyk, Nathaniel J; Cabello, Juan; Miranda-Vizuete, Antonio

2016-07-01

Glutathione is the most abundant thiol in the vast majority of organisms and is maintained in its reduced form by the flavoenzyme glutathione reductase. In this work, we describe the genetic and functional analysis of the Caenorhabditis elegans gsr-1 gene that encodes the only glutathione reductase protein in this model organism. By using green fluorescent protein reporters we demonstrate that gsr-1 produces two GSR-1 isoforms, one located in the cytoplasm and one in the mitochondria. gsr-1 loss of function mutants display a fully penetrant embryonic lethal phenotype characterized by a progressive and robust cell division delay accompanied by an aberrant distribution of interphasic chromatin in the periphery of the cell nucleus. Maternally expressed GSR-1 is sufficient to support embryonic development but these animals are short-lived, sensitized to chemical stress, have increased mitochondrial fragmentation and lower mitochondrial DNA content. Furthermore, the embryonic lethality of gsr-1 worms is prevented by restoring GSR-1 activity in the cytoplasm but not in mitochondria. Given the fact that the thioredoxin redox systems are dispensable in C. elegans, our data support a prominent role of the glutathione reductase/glutathione pathway in maintaining redox homeostasis in the nematode. Copyright © 2016 Elsevier Inc. All rights reserved.

Role of glutathione S-transferase Pi in cisplatin-induced nephrotoxicity.

PubMed

Townsend, Danyelle M; Tew, Kenneth D; He, Lin; King, Jarrod B; Hanigan, Marie H

2009-02-01

One of the dose-limiting toxicities of cisplatin is nephrotoxicity. Renal toxicity is localized to quiescent proximal tubule cells, where the formation of DNA-adducts cannot account for the dose-limiting toxicity. Our earlier results have shown that a glutathione conjugate of cisplatin is metabolized to a nephrotoxicant via gamma-glutamyl transpeptidase (GGT) and a cysteine S-conjugate beta-lyase. The present study was designed to evaluate the potential role of glutathione S-transferase Pi (GSTP) in the initial steps of the bioactivation of cisplatin. Wild-type mice and mice deficient in both murine GSTP genes (GstP1/P2) were treated with cisplatin. Toxicity in both male and female mice was evaluated 5 days after treatment and renal damage was most severe in wild-type male mice. Wild-type males have approximately 10-fold higher levels of GSTP expression in the liver than females, suggesting that hepatic GSTP in the wild-type males contributed to the formation of the nephrotoxic platinum-glutathione conjugate. In GstP1/P2 null mice the gender difference in toxicity was eliminated. Our data show that GSTP expression is a determinant in cisplatin-induced nephrotoxicity and its levels contribute to sex-dependent differences.

Role of Glutathione S-Transferase Pi in Cisplatin Induced Nephrotoxicity

PubMed Central

Townsend, Danyelle M.; Tew, Kenneth D.; He, Lin; King, Jarrod B.; Hanigan, Marie H.

2009-01-01

SUMMARY One of the dose-limiting toxicities of cisplatin is nephrotoxicity. Renal toxicity is localized to quiescent proximal tubule cells, where the formation of DNA-adducts cannot account for the dose-limiting toxicity. Our earlier results have shown that a glutathione-conjugate of cisplatin is metabolized to a nephrotoxicant via gamma-glutamyltranspeptidase (GGT) and a cysteine S-conjugate beta-lyase. The present study was designed to evaluate the potential role of glutathione-S-transferase Pi (GSTP) in the initial steps of the bioactivation of cisplatin. Wild-type mice and mice deficient in both murine GSTP genes (GstP1/P2) were treated with cisplatin. Toxicity in both male and female mice was evaluated 5 days after treatment and renal damage was most severe in wild-type male mice. Wild-type males have ~10-fold higher levels of GSTP expression in the liver than females, suggesting that hepatic GSTP in the wild-type males contributed to the formation of the nephrotoxic platinum-glutathione conjugate. In GstP1/P2 null mice the gender difference in toxicity was eliminated. Our data show that GSTP expression is a determinant in cisplatin-induced nephrotoxicity and its levels contribute to sex-dependent differences. PMID:18819770

FAD-induced in vitro activation of glutathione reductase in the lens of B2 deficient rats.

PubMed

Ono, S; Hirano, H

1984-04-01

We studied the FAD-induced in vitro stimulation of lenticular glutathione reductase in riboflavin-deficient rats. The stimulatory effect of FAD on lenticular glutathione reductase in rats fed a B2-deficient diet for 4 weeks was remarkably higher than in paired control rats fed a B2-supplemented basal diet and control rats had ad libitum access to a B2-supplemented basal diet. The in vitro FAD stimulation effect on rat lenticular glutathione reductase represents a sensitive indicator of the B2 deficient status.

1-Methyl-4-phenylpyridinium-induced alterations of glutathione status in immortalized rat dopaminergic neurons

DOE Office of Scientific and Technical Information (OSTI.GOV)

Drechsel, Derek A.; Liang, L.-P.; Patel, Manisha

2007-05-01

Decreased glutathione levels associated with increased oxidative stress are a hallmark of numerous neurodegenerative diseases, including Parkinson's disease. GSH is an important molecule that serves as an anti-oxidant and is also a major determinant of cellular redox environment. Previous studies have demonstrated that neurotoxins can cause changes in reduced and oxidized GSH levels; however, information regarding steady state levels remains unexplored. The goal of this study was to characterize changes in cellular GSH levels and its regulatory enzymes in a dopaminergic cell line (N27) following treatment with the Parkinsonian toxin, 1-methyl-4-phenylpyridinium (MPP{sup +}). Cellular GSH levels were initially significantly decreasedmore » 12 h after treatment, but subsequently recovered to values greater than controls by 24 h. However, oxidized glutathione (GSSG) levels were increased 24 h following treatment, concomitant with a decrease in GSH/GSSG ratio prior to cell death. In accordance with these changes, ROS levels were also increased, confirming the presence of oxidative stress. Decreased enzymatic activities of glutathione reductase and glutamate-cysteine ligase by 20-25% were observed at early time points and partly account for changes in GSH levels after MPP{sup +} exposure. Additionally, glutathione peroxidase activity was increased 24 h following treatment. MPP{sup +} treatment was not associated with increased efflux of glutathione to the medium. These data further elucidate the mechanisms underlying GSH depletion in response to the Parkinsonian toxin, MPP{sup +}.« less

Cellular and mitochondrial glutathione redox imbalance in lymphoblastoid cells derived from children with autism

PubMed Central

James, S. Jill; Rose, Shannon; Melnyk, Stepan; Jernigan, Stefanie; Blossom, Sarah; Pavliv, Oleksandra; Gaylor, David W.

2009-01-01

Research into the metabolic phenotype of autism has been relatively unexplored despite the fact that metabolic abnormalities have been implicated in the pathophysiology of several other neurobehavioral disorders. Plasma biomarkers of oxidative stress have been reported in autistic children; however, intracellular redox status has not yet been evaluated. Lymphoblastoid cells (LCLs) derived from autistic children and unaffected controls were used to assess relative concentrations of reduced glutathione (GSH) and oxidized disulfide glutathione (GSSG) in cell extracts and isolated mitochondria as a measure of intracellular redox capacity. The results indicated that the GSH/GSSG redox ratio was decreased and percentage oxidized glutathione increased in both cytosol and mitochondria in the autism LCLs. Exposure to oxidative stress via the sulfhydryl reagent thimerosal resulted in a greater decrease in the GSH/GSSG ratio and increase in free radical generation in autism compared to control cells. Acute exposure to physiological levels of nitric oxide decreased mitochondrial membrane potential to a greater extent in the autism LCLs, although GSH/GSSG and ATP concentrations were similarly decreased in both cell lines. These results suggest that the autism LCLs exhibit a reduced glutathione reserve capacity in both cytosol and mitochondria that may compromise antioxidant defense and detoxification capacity under prooxidant conditions.—James, S. J., Rose, S., Melnyk, S., Jernigan, S., Blossom, S., Pavliv, O., Gaylor, D. W. Cellular and mitochondrial glutathione redox imbalance in lymphoblastoid cells derived from children with autism. PMID:19307255

Competitive reactions among glutathione, cisplatin and copper-phenanthroline complexes.

PubMed

Cadoni, Enzo; Valletta, Elisa; Caddeo, Graziano; Isaia, Francesco; Cabiddu, Maria Grazia; Vascellari, Sarah; Pivetta, Tiziana

2017-08-01

A large number of cancers are treated with cisplatin (CDDP). However, its use is limited by drug resistance, which is often related to intracellular levels of thiol-containing molecules such as glutathione (GSH). The role of GSH in cisplatin-resistant cancer cells is still unclear. GSH may form adducts with CDDP which results in the deactivation of the drug, and, actually, a high intracellular level of GSH was observed in some cisplatin-resistant cancers. To overcome drug resistance, CDDP is often administered in combination with one or more drugs to exploit a possible synergistic effect. In previous studies, we observed that the sensitivity to CDDP of leukemic and ovarian cisplatin-resistant cancer cells was restored in the presence of [Cu(phen) 2 (H 2 O)](ClO 4 ) 2 (C0) (phen is 1,10-phenathroline). In order to clarify the possible interactions between GSH and CDDP, the reactivity and competitive reactions among CDDP, C0 and GSH in binary and ternary mixtures were studied. The investigation was extended also to [Cu(phen)(H 2 O) 2 (ClO 4 ) 2 ] (C10) and GSSG, the oxidized form of GSH. It was observed that CDDP was able to react with the studied copper complexes and with GSH or GSSG. However, in mixtures containing CDDP, GSH or GSSG and C0 or C10, only copper-glutathione complexes were detected, while no platinum-glutathione adducts were found. Copyright © 2017 Elsevier Inc. All rights reserved.

Proteomics analysis of human placenta reveals glutathione metabolism dysfunction as the underlying pathogenesis for preeclampsia.

PubMed

Jin, Xiaohan; Xu, Zhongwei; Cao, Jin; Shao, Ping; Zhou, Maobin; Qin, Zhe; Liu, Yan; Yu, Fang; Zhou, Xin; Ji, Wenjie; Cai, Wei; Ma, Yongqiang; Wang, Chengyan; Shan, Nana; Yang, Ning; Chen, Xu; Li, Yuming

2017-09-01

Hypertensive disorder in pregnancy (HDP) refers to a series of diseases that cause the hypertension during pregnancy, including HDP, preeclampsia (PE) and eclampsia. This study screens differentially expressed proteins of placenta tissues in PE cases using 2D LC-MS/MS quantitative proteomics strategy. A total of 2281 proteins are quantified, of these, 145 altering expression proteins are successfully screened between PE and control cases (p<0.05). Bioinformatics analysis suggests that these proteins are mainly involved in many biological processes, such as oxidation reduction, mitochondrion organization, and acute inflammatory response. Especially, the glutamine metabolic process related molecules, GPX1, GPX3, SMS, GGCT, GSTK1, NFκB, GSTT2, SOD1 and GCLM, are involved in the switching process from oxidized glutathione (GSSG) conversion to the reduced glutathione (GSH) by glutathione, mercapturic acid and arginine metabolism process. Results of this study revealed that glutathione metabolism disorder of placenta tissues may contribute to the occurrence of PE disease. Copyright © 2017. Published by Elsevier B.V.

Sulforaphane Restores Cellular Glutathione Levels and Reduces Chronic Periodontitis Neutrophil Hyperactivity In Vitro

PubMed Central

Dias, Irundika H. K.; Chapple, Ian L. C.; Milward, Mike; Grant, Melissa M.; Hill, Eric; Brown, James; Griffiths, Helen R.

2013-01-01

The production of high levels of reactive oxygen species by neutrophils is associated with the local and systemic destructive phenotype found in the chronic inflammatory disease periodontitis. In the present study, we investigated the ability of sulforaphane (SFN) to restore cellular glutathione levels and reduce the hyperactivity of circulating neutrophils associated with chronic periodontitis. Using differentiated HL60 cells as a neutrophil model, here we show that generation of extracellular O2 . - by the nicotinamide adenine dinucleotide (NADPH) oxidase complex is increased by intracellular glutathione depletion. This may be attributed to the upregulation of thiol regulated acid sphingomyelinase driven lipid raft formation. Intracellular glutathione was also lower in primary neutrophils from periodontitis patients and, consistent with our previous findings, patients neutrophils were hyper-reactive to stimuli. The activity of nuclear factor erythroid-2-related factor 2 (Nrf2), a master regulator of the antioxidant response, is impaired in circulating neutrophils from chronic periodontitis patients. Although patients’ neutrophils exhibit a low reduced glutathione (GSH)/oxidised glutathione (GSSG) ratio and a higher total Nrf2 level, the DNA-binding activity of nuclear Nrf2 remained unchanged relative to healthy controls and had reduced expression of glutamate cysteine ligase catalytic (GCLC), and modifier (GCLM) subunit mRNAs, compared to periodontally healthy subjects neutrophils. Pre-treatment with SFN increased expression of GCLC and GCM, improved intracellular GSH/GSSG ratios and reduced agonist-activated extracellular O2 . - production in both dHL60 and primary neutrophils from patients with periodontitis and controls. These findings suggest that a deficiency in Nrf2-dependent pathways may underpin susceptibility to hyper-reactivity in circulating primary neutrophils during chronic periodontitis. PMID:23826097

Sulforaphane restores cellular glutathione levels and reduces chronic periodontitis neutrophil hyperactivity in vitro.

PubMed

Dias, Irundika H K; Chapple, Ian L C; Milward, Mike; Grant, Melissa M; Hill, Eric; Brown, James; Griffiths, Helen R

2013-01-01

The production of high levels of reactive oxygen species by neutrophils is associated with the local and systemic destructive phenotype found in the chronic inflammatory disease periodontitis. In the present study, we investigated the ability of sulforaphane (SFN) to restore cellular glutathione levels and reduce the hyperactivity of circulating neutrophils associated with chronic periodontitis. Using differentiated HL60 cells as a neutrophil model, here we show that generation of extracellular O2 (. -) by the nicotinamide adenine dinucleotide (NADPH) oxidase complex is increased by intracellular glutathione depletion. This may be attributed to the upregulation of thiol regulated acid sphingomyelinase driven lipid raft formation. Intracellular glutathione was also lower in primary neutrophils from periodontitis patients and, consistent with our previous findings, patients neutrophils were hyper-reactive to stimuli. The activity of nuclear factor erythroid-2-related factor 2 (Nrf2), a master regulator of the antioxidant response, is impaired in circulating neutrophils from chronic periodontitis patients. Although patients' neutrophils exhibit a low reduced glutathione (GSH)/oxidised glutathione (GSSG) ratio and a higher total Nrf2 level, the DNA-binding activity of nuclear Nrf2 remained unchanged relative to healthy controls and had reduced expression of glutamate cysteine ligase catalytic (GCLC), and modifier (GCLM) subunit mRNAs, compared to periodontally healthy subjects neutrophils. Pre-treatment with SFN increased expression of GCLC and GCM, improved intracellular GSH/GSSG ratios and reduced agonist-activated extracellular O2 (. -) production in both dHL60 and primary neutrophils from patients with periodontitis and controls. These findings suggest that a deficiency in Nrf2-dependent pathways may underpin susceptibility to hyper-reactivity in circulating primary neutrophils during chronic periodontitis.

Effect of catalase inactivation on levels of inorganic peroxides, superoxide dismutase, glutathione, oxygen consumption and life span in adult houseflies (Musca domestica).

PubMed Central

Allen, R G; Farmer, K J; Sohal, R S

1983-01-01

The effects of total inhibition of catalase, induced by 3-amino-1,2,4-triazole, on the adult housefly (Musca domestica) were examined. The lack of catalase activity had no effect on the longevity of the houseflies. Inorganic-peroxide concentration was elevated at younger ages, but declined in older flies. The rate of oxygen consumption by the flies was greatly decreased and the levels of oxidized as well as reduced glutathione were augmented. Superoxide dismutase activity showed a slight increase. This study suggests that loss of catalase activity does not affect survival of houseflies due to adaptive responses. PMID:6661212

Reduced glutathione enhances fertility of frozen/thawed C57BL/6 mouse sperm after exposure to methyl-beta-cyclodextrin.

PubMed

Takeo, Toru; Nakagata, Naomi

2011-11-01

Sperm cryopreservation is useful for the effective storage of genomic resources derived from genetically engineered mice. However, freezing the sperm of C57BL/6 mice, the most commonly used genetic background for genetically engineered mice, considerably reduces its fertility. We previously reported that methyl-beta-cyclodextrin dramatically improved the fertility of frozen/thawed C57BL/6 mouse sperm. Recently, it was reported that exposing sperm to reduced glutathione may alleviate oxidative stress in frozen/thawed mouse sperm, thereby enhancing in vitro fertilization (IVF); however, the mechanism underlying this effect is poorly understood. In the present study, we examined the combined effects of methyl-beta-cyclodextrin and reduced glutathione on the fertilization rate of IVF with frozen/thawed C57BL/6 mouse sperm and the characteristic changes in the zona pellucida induced by reduced glutathione. Adding reduced glutathione to the fertilization medium increased the fertilization rate. Methyl-beta-cyclodextrin and reduced glutathione independently increased fertilization rates, and their combination produced the strongest effect. We found that reduced glutathione increased the amount of free thiols in the zona pellucida and promoted zona pellucida enlargement. Finally, 2-cell embryos produced by IVF with the addition of reduced glutathione developed normally and produced live offspring. In summary, we have established a novel IVF method using methyl-beta-cyclodextrin during sperm preincubation and reduced glutathione during the IVF procedure to enhance fertility of frozen/thawed C57BL/6 mouse sperm.

Crystal structure and functional characterization of selenocysteine-containing glutathione peroxidase 4 suggests an alternative mechanism of peroxide reduction.

PubMed

Borchert, Astrid; Kalms, Jacqueline; Roth, Sophia R; Rademacher, Marlena; Schmidt, Andrea; Holzhutter, Hermann-Georg; Kuhn, Hartmut; Scheerer, Patrick

2018-06-05

Glutathione peroxidases (GPX) are anti-oxidative enzymes that reduce organic and inorganic hydroperoxides to the corresponding alcohols at the expense of reduced glutathione. The human genome involves eight GPX genes and five of them encode for selenocysteine-containing enzymes. Among the human GPX-isoforms, GPX4 is unique since it is capable of reducing complex hydroperoxy ester lipids such as hydroperoxy phospholipids and hydroperoxy cholesterolesters. Using a number of genetically modified mouse strains the biological role of GPX4 has comprehensively characterized but the molecular enzymology is less well explored. This lack of knowledge is partly related to the fact that mammalian selenoproteins are not high-level expressed in conventional overexpression systems. To explore the structural and functional properties of human GPX4 we expressed this selenoprotein in a cysteine-auxotrophic E. coli strain using a semi-chemical expression strategy. The recombinant enzyme was purified in mg amounts from the bacterial lysate to electrophoretic homogeneity and characterized with respect to its protein-chemical and enzymatic properties. Its crystal structure was solved at 1.3 Å resolution and the X-ray data indicated a monomeric protein, which contains the catalytic selenium at the redox level of the seleninic acid. These data suggest an alternative reaction mechanism involving three different redox states (selenol, selenenic acid, seleninic acid) of the catalytically active selenocysteine. Copyright © 2018 Elsevier B.V. All rights reserved.

Unbalanced activation of glutathione metabolic pathways suggests potential involvement in plant defense against the gall midge mayetiola destructor in wheat

USDA-ARS?s Scientific Manuscript database

Glutathione, a thiol tripeptide of '-glutamylcysteinylglycine, exists abundantly in nearly all organisms. Glutathione participates in various physiological processes involved in redox reactions by serving as an electron donor/acceptor. In this study, we found that the abundance of total glutathion...

Mammalian proapoptotic factor ChaC1 and its homologues function as γ-glutamyl cyclotransferases acting specifically on glutathione

PubMed Central

Kumar, Akhilesh; Tikoo, Shweta; Maity, Shuvadeep; Sengupta, Shantanu; Sengupta, Sagar; Kaur, Amandeep; Kumar Bachhawat, Anand

2012-01-01

ChaC1 is a mammalian proapoptic protein of unknown function induced during endoplasmic reticulum stress. We show using in vivo studies and novel in vitro assays that the ChaC family of proteins function as γ-glutamyl cyclotransferases acting specifically to degrade glutathione but not other γ-glutamyl peptides. The overexpression of these proteins (but not the catalytically dead E>Q mutants) led to glutathione depletion and enhanced apoptosis in yeast. The ChaC family is conversed across all phyla and represents a new pathway for glutathione degradation in living cells, and the first cytosolic pathway for glutathione degradation in mammalian cells. PMID:23070364

Diversification in substrate usage by glutathione synthetases from soya bean (Glycine max), wheat (Triticum aestivum) and maize (Zea mays)

PubMed Central

2005-01-01

Unlike animals which accumulate glutathione (γ-glutamyl-L-cysteinyl-glycine) alone as their major thiol antioxidant, several crops synthesize alternative forms of glutathione by varying the carboxy residue. The molecular basis of this variation is not well understood, but the substrate specificity of the respective GSs (glutathione synthetases) has been implicated. To investigate their substrate tolerance, five GS-like cDNAs have been cloned from plants that can accumulate alternative forms of glutathione, notably soya bean [hGSH (homoglutathione or γ-glutamyl-L-cysteinyl-β-alanine)], wheat (hydroxymethylglutathione or γ-glutamyl-L-cysteinyl-serine) and maize (γ-Glu-Cys-Glu). The respective recombinant GSs were then assayed for the incorporation of differing C-termini into γ-Glu-Cys. The soya bean enzyme primarily incorporated β-alanine to form hGSH, whereas the GS enzymes from cereals preferentially catalysed the formation of glutathione. However, when assayed with other substrates, several GSs and one wheat enzyme in particular were able to synthesize a diverse range of glutathione variants by incorporating unusual C-terminal moieties including D-serine, non-natural amino acids and α-amino alcohols. Our results suggest that plant GSs are capable of producing a diverse range of glutathione homologues depending on the availability of the acyl acceptor. PMID:16008521

Yeast mitochondrial glutathione is an essential antioxidant with mitochondrial thioredoxin providing a back-up system

PubMed Central

Gostimskaya, Irina; Grant, Chris M.

2016-01-01

Glutathione is an abundant, low-molecular-weight tripeptide whose biological importance is dependent upon its redox-active free sulphydryl moiety. Its role as the main determinant of thiol-redox control has been challenged such that it has been proposed to play a crucial role in iron–sulphur clusters maturation, and only a minor role in thiol redox regulation, predominantly as a back-up system for the cytoplasmic thioredoxin system. Here, we have tested the importance of mitochondrial glutathione in thiol-redox regulation. Glutathione reductase (Glr1) is an oxidoreductase which converts oxidized glutathione to its reduced form. Yeast Glr1 localizes to both the cytosol and mitochondria and we have used a Glr1M1L mutant that is constitutively localized to the cytosol to test the requirement for mitochondrial Glr1. We show that the loss of mitochondrial Glr1 specifically accounts for oxidant sensitivity of a glr1 mutant. Loss of mitochondrial Glr1 does not influence iron–sulphur cluster maturation and we have used targeted roGFP2 fluorescent probes to show that oxidant sensitivity is linked to an altered redox environment. Our data indicate mitochondrial glutathione is crucial for mitochondrial thiol-redox regulation, and the mitochondrial thioredoxin system provides a back-up system, but cannot bear the redox load of the mitochondria on its own. PMID:26898146

Glutathione upregulates cAMP signalling via G protein alpha 2 during the development of Dictyostelium discoideum.

PubMed

Lee, Hyang-Mi; Kim, Ji-Sun; Kang, Sa-Ouk

2016-12-01

Despite the importance of glutathione in Dictyostelium, the role of glutathione synthetase (gshB/GSS) has not been clearly investigated. In this study, we observed that increasing glutathione content by constitutive expression of gshB leads to mound-arrest and defects in 3',5'-cyclic adenosine monophosphate (cAMP)-mediated aggregation and developmental gene expression. The overexpression of gpaB encoding G protein alpha 2 (Gα2), an essential component of the cAMP signalling pathway, results in a phenotype similar to that caused by gshB overexpression, whereas gpaB knockdown in gshB-overexpressing cells partially rescues the above-mentioned phenotypic defects. Furthermore, Gα2 is highly enriched at the plasma membrane of gshB-overexpressing cells compared to wild-type cells. Therefore, our findings suggest that glutathione upregulates cAMP signalling via Gα2 modulation during Dictyostelium development. © 2016 Federation of European Biochemical Societies.

The Roles of Glutathione Peroxidases during Embryo Development

PubMed Central

Ufer, Christoph; Wang, Chi Chiu

2011-01-01

Embryo development relies on the complex interplay of the basic cellular processes including proliferation, differentiation, and apoptotic cell death. Precise regulation of these events is the basis for the establishment of embryonic structures and the organ development. Beginning with fertilization of the oocyte until delivery the developing embryo encounters changing environmental conditions such as varying levels of oxygen, which can give rise to reactive oxygen species (ROS). These challenges are met by the embryo with metabolic adaptations and by an array of anti-oxidative mechanisms. ROS can be deleterious by modifying biological molecules including lipids, proteins, and nucleic acids and may induce abnormal development or even embryonic lethality. On the other hand ROS are vital players of various signaling cascades that affect the balance between cell growth, differentiation, and death. An imbalance or dysregulation of these biological processes may generate cells with abnormal growth and is therefore potentially teratogenic and tumorigenic. Thus, a precise balance between processes generating ROS and those decomposing ROS is critical for normal embryo development. One tier of the cellular protective system against ROS constitutes the family of selenium-dependent glutathione peroxidases (GPx). These enzymes reduce hydroperoxides to the corresponding alcohols at the expense of reduced glutathione. Of special interest within this protein family is the moonlighting enzyme glutathione peroxidase 4 (Gpx4). This enzyme is a scavenger of lipophilic hydroperoxides on one hand, but on the other hand can be transformed into an enzymatically inactive cellular structural component. GPx4 deficiency – in contrast to all other GPx family members – leads to abnormal embryo development and finally produces a lethal phenotype in mice. This review is aimed at summarizing the current knowledge on GPx isoforms during embryo development and tumor development with an emphasis

The Roles of Glutathione Peroxidases during Embryo Development.

PubMed

Ufer, Christoph; Wang, Chi Chiu

2011-01-01

Embryo development relies on the complex interplay of the basic cellular processes including proliferation, differentiation, and apoptotic cell death. Precise regulation of these events is the basis for the establishment of embryonic structures and the organ development. Beginning with fertilization of the oocyte until delivery the developing embryo encounters changing environmental conditions such as varying levels of oxygen, which can give rise to reactive oxygen species (ROS). These challenges are met by the embryo with metabolic adaptations and by an array of anti-oxidative mechanisms. ROS can be deleterious by modifying biological molecules including lipids, proteins, and nucleic acids and may induce abnormal development or even embryonic lethality. On the other hand ROS are vital players of various signaling cascades that affect the balance between cell growth, differentiation, and death. An imbalance or dysregulation of these biological processes may generate cells with abnormal growth and is therefore potentially teratogenic and tumorigenic. Thus, a precise balance between processes generating ROS and those decomposing ROS is critical for normal embryo development. One tier of the cellular protective system against ROS constitutes the family of selenium-dependent glutathione peroxidases (GPx). These enzymes reduce hydroperoxides to the corresponding alcohols at the expense of reduced glutathione. Of special interest within this protein family is the moonlighting enzyme glutathione peroxidase 4 (Gpx4). This enzyme is a scavenger of lipophilic hydroperoxides on one hand, but on the other hand can be transformed into an enzymatically inactive cellular structural component. GPx4 deficiency - in contrast to all other GPx family members - leads to abnormal embryo development and finally produces a lethal phenotype in mice. This review is aimed at summarizing the current knowledge on GPx isoforms during embryo development and tumor development with an emphasis on

Glutathione S - transferases class Pi and Mi and their significance in oncology.

PubMed

Marchewka, Zofia; Piwowar, Agnieszka; Ruzik, Sylwia; Długosz, Anna

2017-06-19

In this article the current data, which shows that glutathione S-transferases (GST) class Pi and Mi are interesting and promising biomarkers in acute and chronic inflammatory processes as well as in the oncology, were presented based on the review of the latest experimental and clinical studies. The article shows their characteristics, functions and participation (direct - GST Pi, indirect - GST Mi) in the regulation of signaling pathways of JNK kinases, which are involved in cell differentiation. Overexpression of glutathione S-transferases class Pi and Mi in many cancer cells plays a key role in cancer treatment, making them resistant to chemotherapy. GST isoenzymes are involved in the metabolism of various types of xenobiotics and endogenous substrates, so their altered expression in cancer tissues as well as in serum and urine could be an important potential marker of the cancer and an indicator of oxidative stress. The study shows the role of glutathione S-transferases in redox homeostasis of tumor cells and in the mechanism of resistance to anticancer drugs.

Anaerobic Ammonium-Oxidizing Bacteria: Unique Microorganisms with Exceptional Properties

PubMed Central

Jetten, Mike S. M.

2012-01-01

Summary: Anaerobic ammonium-oxidizing (anammox) bacteria defy many microbiological concepts and share numerous properties with both eukaryotes and archaea. Among their most intriguing characteristics are their compartmentalized cell plan and archaeon-like cell wall. Here we review our current knowledge about anammox cell biology. The anammox cell is divided into three separate compartments by bilayer membranes. The anammox cell consists of (from outside to inside) the cell wall, paryphoplasm, riboplasm, and anammoxosome. Not much is known about the composition or function of both the anammox cell wall and the paryphoplasm compartment. The cell wall is proposed to be proteinaceous and to lack both peptidoglycan and an outer membrane typical of Gram-negative bacteria. The function of the paryphoplasm is unknown, but it contains the cell division ring. The riboplasm resembles the standard cytoplasmic compartment of other bacteria; it contains ribosomes and the nucleoid. The anammoxosome occupies most of the cell volume and is a so-called “prokaryotic organelle” analogous to the eukaryotic mitochondrion. This is the site where the anammox reaction takes place, coupled over the curved anammoxosome membrane, possibly giving rise to a proton motive force and subsequent ATP synthesis. With these unique properties, anammox bacteria are food for thought concerning the early evolution of the domains Bacteria, Archaea, and Eukarya. PMID:22933561

Transmutation of human glutathione transferase A2-2 with peroxidase activity into an efficient steroid isomerase.

PubMed

Pettersson, Par L; Johansson, Ann-Sofie; Mannervik, Bengt

2002-08-16

A major goal in protein engineering is the tailor-making of enzymes for specified chemical reactions. Successful attempts have frequently been based on directed molecular evolution involving libraries of random mutants in which variants with desired properties were identified. For the engineering of enzymes with novel functions, it would be of great value if the necessary changes of the active site could be predicted and implemented. Such attempts based on the comparison of similar structures with different substrate selectivities have previously met with limited success. However, the present work shows that the knowledge-based redesign restricted to substrate-binding residues in human glutathione transferase A2-2 can introduce high steroid double-bond isomerase activity into the enzyme originally characterized by glutathione peroxidase activity. Both the catalytic center activity (k(cat)) and catalytic efficiency (k(cat)/K(m)) match the values of the naturally evolved glutathione transferase A3-3, the most active steroid isomerase known in human tissues. The substrate selectivity of the mutated glutathione transferase was changed 7000-fold by five point mutations. This example demonstrates the functional plasticity of the glutathione transferase scaffold as well as the potential of rational active-site directed mutagenesis as a complement to DNA shuffling and other stochastic methods for the redesign of proteins with novel functions.

Water stress amelioration and plant growth promotion in wheat plants by osmotic stress tolerant bacteria.

PubMed

Chakraborty, U; Chakraborty, B N; Chakraborty, A P; Dey, P L

2013-05-01

Soil microorganisms with potential for alleviation of abiotic stresses in combination with plant growth promotion would be extremely useful tools in sustainable agriculture. To this end, the present study was initiated where forty-five salt tolerant bacterial isolates with ability to grow in high salt medium were obtained from the rhizosphere of Triticum aestivum and Imperata cylindrica. These bacteria were tested for plant-growth-promoting rhizobacteria traits in vitro such as phosphate solubilization, siderophore, ACC deaminase and IAA production. Of the forty-five isolates, W10 from wheat rhizosphere and IP8 from blady grass rhizosphere, which tested positive in all the tests were identified by morpholological, biochemical and 16SrDNA sequencing as Bacillus safensis and Ochrobactrum pseudogregnonense respectively and selected for in vivo studies. Both the bacteria could promote growth in six varieties of wheat tested in terms of increase in root and shoot biomass, height of plants, yield, as well as increase in chlorophyll content. Besides, the wheat plants could withstand water stress more efficiently in presence of the bacteria as indicated by delay in appearance of wilting symptoms increases in relative water content of treated water stressed plants in comparison to untreated stressed ones, and elevated antioxidant responses. Enhanced antioxidant responses were evident as elevated activities of enzymes such as catalase, peroxidase, ascorbate peroxidase, superoxide dismutase and glutathione reductase as well as increased accumulation of antioxidants such as carotenoids and ascorbate. Results clearly indicate that the ability of wheat plants to withstand water stress is enhanced by application of these bacteria which also function as plant growth promoting rhizobacteria.

Effect of antioxidant-enriched diets on glutathione redox status in tissue homogenates and mitochondria of the senescence-accelerated mouse

PubMed Central

Rebrin, Igor; Zicker, Steven; Wedekind, Karen J.; Paetau-Robinson, Inke; Packer, Lester; Sohal, Rajindar S.

2010-01-01

The main purpose of this study was to investigate whether consumption of diets enriched in antioxidants attenuates the level of oxidative stress in the senescence-accelerated mouse (SAM). In separate and independent studies, two different dietary mixtures, one enriched with vitamin E, vitamin C, L-carnitine, and lipoic acid (Diet I) and another diet including vitamins E and C and 13 additional ingredients containing micronutrients with bioflavonoids, polyphenols, and carotenoids (Diet II), were fed for 8 and 10 months, respectively. The amounts of glutathione (GSH) and glutathione disulfides (GSSG) and GSH:GSSG ratios were determined in plasma, tissue homogenates, and mitochondria isolated from five different tissues of SAM (P8) mice. Both diets had a reductive effect in plasma; however Diet I had relatively little effect on the glutathione redox status in tissue homogenates or mitochondria. Remarkably, Diet II caused a large increase in the amount of glutathione and a marked reductive shift in glutathione redox state in mitochondria. Overall, the effects of Diet II were tissue and gender specific. Results indicated that the glutathione redox state in mitochondria and tissues can be altered by supplemental intake of a relatively complex mixture of dietary antioxidants that contains substances known to induce phase 2 enzymes, glutathione, and antioxidant defenses. Whether corresponding attenuations occur in age-associated deleterious changes in physiological functions or life span remains unknown. PMID:16043026

Effect of antioxidant-enriched diets on glutathione redox status in tissue homogenates and mitochondria of the senescence-accelerated mouse.

PubMed

Rebrin, Igor; Zicker, Steven; Wedekind, Karen J; Paetau-Robinson, Inke; Packer, Lester; Sohal, Rajindar S

2005-08-15

The main purpose of this study was to investigate whether consumption of diets enriched in antioxidants attenuates the level of oxidative stress in the senescence-accelerated mouse (SAM). In separate and independent studies, two different dietary mixtures, one enriched with vitamin E, vitamin C, L-carnitine, and lipoic acid (Diet I) and another diet including vitamins E and C and 13 additional ingredients containing micronutrients with bioflavonoids, polyphenols, and carotenoids (Diet II), were fed for 8 and 10 months, respectively. The amounts of glutathione (GSH) and glutathione disulfides (GSSG) and GSH:GSSG ratios were determined in plasma, tissue homogenates, and mitochondria isolated from five different tissues of SAM (P8) mice. Both diets had a reductive effect in plasma; however Diet I had relatively little effect on the glutathione redox status in tissue homogenates or mitochondria. Remarkably, Diet II caused a large increase in the amount of glutathione and a marked reductive shift in glutathione redox state in mitochondria. Overall, the effects of Diet II were tissue and gender specific. Results indicated that the glutathione redox state in mitochondria and tissues can be altered by supplemental intake of a relatively complex mixture of dietary antioxidants that contains substances known to induce phase 2 enzymes, glutathione, and antioxidant defenses. Whether corresponding attenuations occur in age-associated deleterious changes in physiological functions or life span remains unknown.

Unearthing the secrets of mitochondrial ROS and glutathione in bioenergetics.

PubMed

Mailloux, Ryan J; McBride, Skye L; Harper, Mary-Ellen

2013-12-01

During the cellular oxidation of fuels, electrons are used to power the proton pumps of the mitochondrial electron transport chain (ETC) and ultimately drive ATP synthesis and the reduction of molecular oxygen to water. During these oxidative processes, some electrons can 'spin off' during fuel oxidation and electron transport to univalently reduce O2, forming reactive oxygen species (ROS). In excess, ROS can be detrimental; however, at low concentrations oxyradicals are essential signaling molecules. Mitochondria thus use a battery of systems to finely control types and levels of ROS, including antioxidants. Several antioxidant systems depend on glutathione. Here, we review mitochondrial ROS homeostatic systems, including emerging knowledge about roles of glutathione in redox balance and the control of protein function by post-translational modification. Copyright © 2013 Elsevier Ltd. All rights reserved.

Glutathione redox balance in hibernating Chinese soft-shelled turtle Pelodiscus sinensis hatchlings.

PubMed

Zhang, Wenyi; Niu, Cuijuan; Liu, Yukun; Chen, Bojian

2017-05-01

Glutathione (GSH) system is a critical component of antioxidant defense, which is important for hibernating survive of turtle hatchlings. The present work measured changes at the mRNA level of genes involved in GSH synthesis, GSH reduction and GSH utilization, as well as enzyme activity, in Pelodiscus sinensis hatchlings during hibernation. Samples were taken in the field at pre-hibernation (17°C, Mud temperature (MT)), hibernation (5.8°C, MT) and arousal (20.1°C, MT). Cerebral total GSH content decreased during hibernation, recovered after arousal along with a stable ratio of GSH/GSSG. Hepatic total GSH increased after arousal and pushed the ratio of GSH/GSSG to a more reduced status. Cerebral glutathione reductase (GR) mRNA and activity were depressed during hibernation then recovered after arousal. However, hepatic GR mRNA elevated during hibernation but its activity did not change. Tissue-specific changes of GR activity and mRNA may promote these tissue-specific changes of GSH redox. Hibernation caused little effect on mRNA level of glutathione synthetase (GS) while arousal induced them in the brain and liver. Most Glutathione-S-transferase (GST) isoform mRNAs did not change in both brain and liver during hibernation, then induced after arousal. Cerebral and hepatic GST activities kept stable throughout the entire experiment. Our results showed that GSH system may play a more important role in antioxidant defense in the liver while mainly maintaining stable redox balance in the brain of hibernating P. sinensis hatchings. Copyright © 2017 Elsevier Inc. All rights reserved.

Enalapril and captopril enhance glutathione-dependent antioxidant defenses in mouse tissues.

PubMed

de Cavanagh, E M; Inserra, F; Ferder, L; Fraga, C G

2000-03-01

The effect of enalapril and captopril on total glutathione content (GSSG + GSH) and selenium-dependent glutathione peroxidase (Se-GPx) and glutathione reductase (GSSG-Rd) activities was investigated in mouse tissues. CF-1 mice (4-mo-old females) received water containing enalapril (20 mg/l) or captopril (50 mg/l) for 11 wk. Enalapril increased GSSG + GSH content (P < 0.05) in erythrocytes (147%), brain (112%), and lung (67%), and captopril increased GSSG + GSH content in erythrocytes (190%) and brain (132%). Enalapril enhanced Se-GPx activity in kidney cortex (42%) and kidney medulla (23%) and captopril in kidney cortex (30%). GSSG-Rd activity was enhanced by enalapril in erythrocytes (21%), brain (21%), liver (18%), and kidney cortex (53%) and by captopril in erythrocytes (25%), brain (19%), and liver (34%). In vitro erythrocyte oxidant stress was evaluated by thiobarbituric acid-reactive substances (TBARS) production (control 365 +/- 11, enalapril 221 +/- 26, captopril 206 +/- 17 nmol TBARS x g Hb(-1) x h(-1); both P < 0.05 vs. control) and phenylhydrazine-induced methemoglobin (MetHb) formation (control 66.5 +/- 3.5, enalapril 52.9 +/- 0.4, captopril: 56.4 +/- 2.9 micromol MetHb/g Hb; both P < 0.05 vs. control). Both angiotensin-converting enzyme inhibitor treatments were associated with increased nitric oxide production, as assessed by plasma NO-(3) + NO-(2) level determination (control 9.22 +/- 0.64, enalapril 13.7 +/- 1.9, captopril 17.3 +/- 3.0 micromol NO-(3) + NO-(2)/l plasma; both P < 0.05 vs. control). These findings support our previous reports on the enalapril- and captopril-induced enhancement of endogenous antioxidant defenses and include new data on glutathione-dependent defenses, thus furthering current knowledge on the association of ACE inhibition and antioxidants.

FMISO accumulation in tumor is dependent on glutathione conjugation capacity in addition to hypoxic state.

PubMed

Masaki, Yukiko; Shimizu, Yoichi; Yoshioka, Takeshi; Nishijima, Ken-Ichi; Zhao, Songji; Higashino, Kenichi; Numata, Yoshito; Tamaki, Nagara; Kuge, Yuji

2017-10-01

18 F-fluoromisonidazole (FMISO), a well-known PET imaging probe for diagnosis of hypoxia, is believed to accumulate in hypoxic cells via covalent binding with macromolecules after reduction of the nitro group. Previously, we showed the majority of 18 F-FMISO was incorporated into low-molecular-weight metabolites in hypoxic tumors, and the glutathione conjugate of reduced FMISO (amino-FMISO-GS) distributed in the tumor hypoxic regions as revealed by imaging mass spectrometry (IMS). The present study was conducted to clarify whether FMISO is metabolized to amino-FMISO-GS within tumor cells and how amino-FMISO-GS contributes to FMISO accumulation in hypoxic cells. We also evaluated the relationship between FMISO accumulation and the glutathione conjugation-related factors in the cells. Tumor cells (FaDu, LOVO, and T24) were treated with 18 F-FMISO and incubated under normoxic or hypoxic conditions for 4 h. The FMISO metabolites were analyzed with LC-ESI-MS. Several glutathione conjugation-related factors of tumor cells were evaluated in vitro. FaDu tumor-bearing mice were intravenously injected with 18 F-FMISO and the tumors were excised at 4 h post-injection. Autoradiography, IMS and histologic studies were performed. Amino-FMISO-GS was the main contributor to FMISO incorporated in hypoxic FaDu cells in vitro and in vivo. Total FMISO uptake levels and amino-FMISO-GS levels were highest in FaDu, followed by LOVO, and then T24 (total uptake: 0.851 ± 0.009 (FaDu), 0.617 ± 0.021 (LOVO) and 0.167 ± 0.006 (T24) % dose/mg protein; amino-FMISO-GS: 0.502 ± 0.035 (FaDu), 0.158 ± 0.013 (LOVO), and 0.007 ± 0.001 (T24) % dose/mg protein). The glutathione level of FaDu was significantly higher than those of LOVO and T24. The enzyme activity of glutathione-S-transferase catalyzing the glutathione conjugation reaction in FaDu was similar levels to that in LOVO, and was higher than that in T24. Quantitative RT-PCR analysis revealed that the expression levels of

Detecting cell division of Pseudomonas aeruginosa bacteria from bright-field microscopy images with hidden conditional random fields.

PubMed

Ong, Lee-Ling S; Xinghua Zhang; Kundukad, Binu; Dauwels, Justin; Doyle, Patrick; Asada, H Harry

2016-08-01

An approach to automatically detect bacteria division with temporal models is presented. To understand how bacteria migrate and proliferate to form complex multicellular behaviours such as biofilms, it is desirable to track individual bacteria and detect cell division events. Unlike eukaryotic cells, prokaryotic cells such as bacteria lack distinctive features, causing bacteria division difficult to detect in a single image frame. Furthermore, bacteria may detach, migrate close to other bacteria and may orientate themselves at an angle to the horizontal plane. Our system trains a hidden conditional random field (HCRF) model from tracked and aligned bacteria division sequences. The HCRF model classifies a set of image frames as division or otherwise. The performance of our HCRF model is compared with a Hidden Markov Model (HMM). The results show that a HCRF classifier outperforms a HMM classifier. From 2D bright field microscopy data, it is a challenge to separate individual bacteria and associate observations to tracks. Automatic detection of sequences with bacteria division will improve tracking accuracy.

Characterization of bifunctional L-glutathione synthetases from Actinobacillus pleuropneumoniae and Actinobacillus succinogenes for efficient glutathione biosynthesis.

PubMed

Yang, Jianhua; Li, Wei; Wang, Dezheng; Wu, Hui; Li, Zhimin; Ye, Qin

2016-07-01

Glutathione (GSH), an important bioactive substance, is widely applied in pharmaceutical and food industries. In this work, two bifunctional L-glutathione synthetases (GshF) from Actinobacillus pleuropneumoniae (GshFAp) and Actinobacillus succinogenes (GshFAs) were successfully expressed in Escherichia coli BL-21(DE3). Similar to the GshF from Streptococcus thermophilus (GshFSt), GshFAp and GshFAs can be applied for high titer GSH production because they are less sensitive to end-product inhibition (Ki values 33 and 43 mM, respectively). The active catalytic forms of GshFAs and GshFAp are dimers, consistent with those of GshFPm (GshF from Pasteurella multocida) and GshFSa (GshF from Streptococcus agalactiae), but are different from GshFSt (GshF from S. thermophilus) which is an active monomer. The analysis of the protein sequences and three dimensional structures of GshFs suggested that the binding sites of GshFs for substrates, L-cysteine, L-glutamate, γ-glutamylcysteine, adenosine-triphosphate, and glycine are highly conserved with only very few differences. With sufficient supply of the precursors, the recombinant strains BL-21(DE3)/pET28a-gshFas and BL-21(DE3)/pET28a-gshFap were able to produce 36.6 and 34.1 mM GSH, with the molar yield of 0.92 and 0.85 mol/mol, respectively, based on the added L-cysteine. The results showed that GshFAp and GshFAs are potentially good candidates for industrial GSH production.

Desired and side effects of the supplementation with l-glutamine and l-glutathione in enteric glia of diabetic rats.

PubMed

Panizzon, Cynthia Priscilla do Nascimento Bonato; Zanoni, Jacqueline Nelisis; Hermes-Uliana, Catchia; Trevizan, Aline Rosa; Sehaber, Camila Caviquioli; Pereira, Renata Virginia Fernandes; Linden, David Robert; Neto, Marcílio Hubner de Miranda

2016-07-01

Enteric neuropathy associated with Diabetes Mellitus causes dysfunction in the digestive system, such as: nausea, diarrhea, constipation, vomiting, among others. The aim of this study was to compare the effects of supplementation with 2% l-glutamine and 1% l-glutathione on neurons and enteric glial cells of ileum of diabetic rats. Thirty male Wistar rats have been used according to these group distributions: Normoglycemic (N), Normoglycemic supplemented with l-glutamine (NG), Normoglycemic supplemented with l-glutathione (NGO), Diabetic (D), Diabetic supplemented with l-glutamine (DG) and Diabetic supplemented with l-glutathione (DGO). After 120days, the ileum was processed for immunohistochemistry of HuC/D and S100β. Quantitative and morphometric analysis have been performed. Diabetic rats presented a decrease in the number of neurons when compared to normoglycemic animals. However, diabetes was not associated with a change in glial density. l-Glutathione prevented the neuronal death in diabetic rats. l-Glutathione increased a glial proliferation in diabetic rats. The neuronal area in diabetic rats increased in relation to the normoglycemics. The diabetic rats supplemented with l-glutamine and l-glutathione showed a smaller neuronal area in comparison to diabetic group. The glial cell area was a decreased in the diabetics. The diabetic rats supplemented with l-glutamine and l-glutathione did not have significant difference in the glial cell body area when compared to diabetic rats. It is concluded that the usage of l-glutamine and l-glutathione as supplements presents both desired and side effects that are different for the same substance in considering normoglycemic or diabetic animals. Copyright © 2016 Elsevier GmbH. All rights reserved.

Effects of glutamine supplementation on gut barrier, glutathione content and acute phase response in malnourished rats during inflammatory shock.

PubMed

Belmonte, Liliana; Coëffier, Moïse; Le Pessot, Florence; Miralles-Barrachina, Olga; Hiron, Martine; Leplingard, Antony; Lemeland, Jean-François; Hecketsweiler, Bernadette; Daveau, Maryvonne; Ducrotté, Philippe; Déchelotte, Pierre

2007-05-28

To evaluate the effect of glutamine on intestinal mucosa integrity, glutathione stores and acute phase response in protein-depleted rats during an inflammatory shock. Plasma acute phase proteins (APP), jejunal APP mRNA levels, liver and jejunal glutathione concentrations were measured before and one, three and seven days after turpentine injection in 4 groups of control, protein-restricted, protein-restricted rats supplemented with glutamine or protein powder. Bacterial translocation in mesenteric lymph nodes and intestinal morphology were also assessed. Protein deprivation and turpentine injection significantly reduced jejunal villus height, and crypt depths. Mucosal glutathione concentration significantly decreased in protein-restricted rats. Before turpentine oil, glutamine supplementation restored villus heights and glutathione concentration (3.24 +/- 1.05 vs 1.72 +/- 0.46 mumol/g tissue, P<0.05) in the jejunum, whereas in the liver glutathione remained low. Glutamine markedly increased jejunal alpha1-acid glycoprotein mRNA level after turpentine oil but did not affect its plasma concentration. Bacterial translocation in protein-restricted rats was not prevented by glutamine or protein powder supplementation. Glutamine restored gut glutathione stores and villus heights in malnourished rats but had no preventive effect on bacterial translocation in our model.

Effects of mercury and selenium on glutathione metabolism and oxidative stress in mallard ducks

USGS Publications Warehouse

Hoffman, D.J.; Heinz, G.H.

1998-01-01

Earlier studies reported on the toxicity and related oxidative stress of different forms of Se, including seleno-D,L-methionine, in mallards (Anas platyrhynchos). This study compares the effects of Se (seleno-D,L-methionine) and Hg (methylmercury chloride) separately and in combination. Mallard drakes received one of the following diets: untreated feed (controls), or feed containing 10 ppm Se, 10 ppm Hg, or 10 ppm Se in combination with 10 ppm Hg. After 10 weeks, blood, liver, and brain samples were collected for biochemical assays. The following clinical and biochemical alterations occurred in response to mercury exposure: hematocrit and hemoglobin concentrations decreased; activities of the enzymes glutathione (GSH) peroxidase (plasma and liver), glutathione-S-transferase (liver), and glucose-6-phosphate dehydrogenase (G-6-PDH) (liver and brain) decreased; hepatic oxidized glutathione (GSSG) concentration increased relative to reduced glutathione (GSH); and lipid peroxidation in the brain was evident as detected by increased thiobarbituric reactive substances (TBARS). Effects of Se alone included increased hepatic GSSG reductase activity and brain TBARS concentration. Se in combination with Hg partially or totally alleviated effects of Hg on GSH peroxidase, G-6-PDH, and GSSG. These findings are compared in relation to field observations for diving ducks and other aquatic birds. It is concluded that since both Hg and excess Se can affect thiol status, measurement of associated enzymes in conjunction with thiol status may be a useful bioindicator to discriminate between Hg and Se effects. The ability of Se to restore the activities of G-6-PDH, GSH peroxidase, and glutathione status involved in antioxidative defense mechanisms may be crucial to biological protection from the toxic effects of methyl mercury.

The peptide transporter PepT2 mediates the uptake of the glutathione precursor CysGly in astroglia-rich primary cultures.

PubMed

Dringen, R; Hamprecht, B; Bröer, S

1998-07-01

The intracellular content of glutathione in astroglia-rich primary cultures derived from the brains of newborn rats was used as an indicator for the ability of these cultures to utilize cysteinylglycine (CysGly) for glutathione synthesis. After a 24-h starvation period in the absence of glucose and amino acids, CysGly was able to substitute for cysteine plus glycine in the restoration of glutathione. Glutathione restoration from CysGly plus glutamate was only slightly affected by the dipeptides carnosine or serylglycine in a 200-fold excess. Captopril, a substrate of the peptide transporter PepT1, had almost no effect on glutathione restoration. In contrast, with increasing concentrations of alanylalanine or cefadroxil, known substrates of the peptide transporter PepT2, the amount of glutathione restored in the presence of CysGly and glutamate was strongly reduced. Cefadroxil in a 200-fold excess totally prevented the utilization of CysGly for glutathione restoration. The presence of mRNA for PepT2 in astroglia-rich primary cultures was demonstrated by application of RT-PCR. These results demonstrate that PepT2 is expressed in astroglia-rich primary cultures and that this transporter is highly likely to be responsible for the uptake of CysGly in these cultures.

In vitro interactions of thallium with components of the glutathione-dependent antioxidant defence system.

PubMed

Villaverde, Marcela S; Hanzel, Cecilia E; Verstraeten, Sandra V

2004-09-01

We investigated the hypothesis that thallium (Tl) interactions with the glutathione-dependent antioxidant defence system could contribute to the oxidative stress associated with Tl toxicity. Working in vitro with reduced glutathione (GSH), glutathione reductase (GR) or glutathione peroxidase (GPx) in solution, we studied the effects of Tl+ and Tl3+ (1-25 microM) on: (a) the amount of free GSH, investigating whether the metal binds to GSH and/or oxidizes it; (b) the activity of the enzyme GR, that catalyzes GSH regeneration; and (c) the enzyme GPx, that reduces hydroperoxide at expense of GSH oxidation. We found that, while Tl+ had no effect on GSH concentration, Tl3+ oxidized it. Both cations inhibited the reduction of GSSG by GR and the diaphorase activity of this enzyme. In addition, Tl3+ per se oxidized NADPH, the cofactor of GR. The effects of Tl on GPx activity depended on the metal charge: Tl+ inhibited GPx when cumene hydroperoxide (CuOOH) was the substrate, while Tl(3+)-mediated GPx inhibition occurred with both substrates. The present results show that Tl interacts with all the components of GSH/GSSG antioxidant defence system. Alterations of this protective pathway could be partially responsible for the oxidative stress associated with Tl toxicity.

Effects of Combined Low Glutathione with Mild Oxidative and Low Phosphorus Stress on the Metabolism of Arabidopsis thaliana

PubMed Central

Fukushima, Atsushi; Iwasa, Mami; Nakabayashi, Ryo; Kobayashi, Makoto; Nishizawa, Tomoko; Okazaki, Yozo; Saito, Kazuki; Kusano, Miyako

2017-01-01

Plants possess highly sensitive mechanisms that monitor environmental stress levels for a dose-dependent fine-tuning of their growth and development. Differences in plant responses to severe and mild abiotic stresses have been recognized. Although many studies have revealed that glutathione can contribute to plant tolerance to various environmental stresses, little is known about the relationship between glutathione and mild abiotic stress, especially the effect of stress-induced altered glutathione levels on the metabolism. Here, we applied a systems biology approach to identify key pathways involved in the gene-to-metabolite networks perturbed by low glutathione content under mild abiotic stress in Arabidopsis thaliana. We used glutathione synthesis mutants (cad2-1 and pad2-1) and plants overexpressing the gene encoding γ-glutamylcysteine synthetase, the first enzyme of the glutathione biosynthetic pathway. The plants were exposed to two mild stress conditions—oxidative stress elicited by methyl viologen and stress induced by the limited availability of phosphate. We observed that the mutants and transgenic plants showed similar shoot growth as that of the wild-type plants under mild abiotic stress. We then selected the synthesis mutants and performed multi-platform metabolomics and microarray experiments to evaluate the possible effects on the overall metabolome and the transcriptome. As a common oxidative stress response, several flavonoids that we assessed showed overaccumulation, whereas the mild phosphate stress resulted in increased levels of specific kaempferol- and quercetin-glycosides. Remarkably, in addition to a significant increased level of sugar, osmolytes, and lipids as mild oxidative stress-responsive metabolites, short-chain aliphatic glucosinolates over-accumulated in the mutants, whereas the level of long-chain aliphatic glucosinolates and specific lipids decreased. Coordinated gene expressions related to glucosinolate and flavonoid

Symbiotic Bacteria Direct Expression of an Intestinal Bactericidal Lectin

PubMed Central

Cash, Heather L.; Whitham, Cecilia V.; Behrendt, Cassie L.; Hooper, Lora V.

2009-01-01

The mammalian intestine harbors complex societies of beneficial bacteria that are maintained in the lumen with minimal penetration of mucosal surfaces. Microbial colonization of germ-free mice triggers epithelial expression of RegIIIγ, a secreted C-type lectin. RegIIIγ binds intestinal bacteria but lacks the complement recruitment domains present in other microbe-binding mammalian C-type lectins. We show that RegIIIγ and its human counterpart, HIP/PAP, are directly antimicrobial proteins that bind their bacterial targets via interactions with peptidoglycan carbohydrate. We propose that these proteins represent an evolutionarily primitive form of lectin-mediated innate immunity, and that they reveal intestinal strategies for maintaining symbiotic host-microbial relationships. PMID:16931762

Glutathione redox potential in the mitochondrial intermembrane space is linked to the cytosol and impacts the Mia40 redox state

PubMed Central

Kojer, Kerstin; Bien, Melanie; Gangel, Heike; Morgan, Bruce; Dick, Tobias P; Riemer, Jan

2012-01-01

Glutathione is an important mediator and regulator of cellular redox processes. Detailed knowledge of local glutathione redox potential (EGSH) dynamics is critical to understand the network of redox processes and their influence on cellular function. Using dynamic oxidant recovery assays together with EGSH-specific fluorescent reporters, we investigate the glutathione pools of the cytosol, mitochondrial matrix and intermembrane space (IMS). We demonstrate that the glutathione pools of IMS and cytosol are dynamically interconnected via porins. In contrast, no appreciable communication was observed between the glutathione pools of the IMS and matrix. By modulating redox pathways in the cytosol and IMS, we find that the cytosolic glutathione reductase system is the major determinant of EGSH in the IMS, thus explaining a steady-state EGSH in the IMS which is similar to the cytosol. Moreover, we show that the local EGSH contributes to the partially reduced redox state of the IMS oxidoreductase Mia40 in vivo. Taken together, we provide a comprehensive mechanistic picture of the IMS redox milieu and define the redox influences on Mia40 in living cells. PMID:22705944

Antibacterial clay against gram-negative antibiotic resistant bacteria.

PubMed

Zarate-Reyes, Luis; Lopez-Pacheco, Cynthia; Nieto-Camacho, Antonio; Palacios, Eduardo; Gómez-Vidales, Virginia; Kaufhold, Stephan; Ufer, Kristian; García Zepeda, Eduardo; Cervini-Silva, Javiera

2018-01-15

Antibiotic resistant bacteria persist throughout the world because they have evolved the ability to express various defense mechanisms to cope with antibiotics and the immune system; thus, low-cost strategies for the treatment of these bacteria are needed, such as the usage of environmental minerals. This paper reports the antimicrobial properties of a clay collected from Brunnenberg, Germany, that is composed of ferroan saponite with admixtures of quartz, feldspar and calcite as well as exposed or hidden (layered at inner regions) nano Fe(0). Based on the growth curves (log phase) of six antibiotic resistant bacteria (4 gram-negative and 2 gram-positive), we concluded that the clay acted as a bacteriostat; however, the clay was only active against the gram-negative bacteria (except for resilient Klebsiella pneumonia). The bacteriostatic mode of action was evidenced by the initial lack of Colony Forming Units on agar plates with growth registered afterward, certainly after 24h, and can be explained because interactions between membrane lipopolysaccharides and the siloxane surfaces of the clay. Labile or bioavailable Fe in the clay (extracted by EDTA or DFO-B) induced the quantitative production of HO as well as oxidative stress, which, nevertheless, did not account for by its bacteriostatic activity. Copyright © 2017 Elsevier B.V. All rights reserved.

Anaerobic bacteria

MedlinePlus

Anaerobic bacteria are bacteria that do not live or grow when oxygen is present. In humans, these bacteria ... Brook I. Diseases caused by non-spore-forming anaerobic bacteria. In: Goldman L, Schafer AI, eds. Goldman-Cecil ...

Impaired synthesis and antioxidant defense of glutathione in the cerebellum of autistic subjects: alterations in the activities and protein expression of glutathione-related enzymes.

PubMed

Gu, Feng; Chauhan, Ved; Chauhan, Abha

2013-12-01

Autism is a neurodevelopmental disorder associated with social deficits and behavioral abnormalities. Recent evidence in autism suggests a deficit in glutathione (GSH), a major endogenous antioxidant. It is not known whether the synthesis, consumption, and/or regeneration of GSH is affected in autism. In the cerebellum tissues from autism (n=10) and age-matched control subjects (n=10), the activities of GSH-related enzymes glutathione peroxidase (GPx), glutathione-S-transferase (GST), glutathione reductase (GR), and glutamate cysteine ligase (GCL) involved in antioxidant defense, detoxification, GSH regeneration, and synthesis, respectively, were analyzed. GCL is a rate-limiting enzyme for GSH synthesis, and the relationship between its activity and the protein expression of its catalytic subunit GCLC and its modulatory subunit GCLM was also compared between the autistic and the control groups. Results showed that the activities of GPx and GST were significantly decreased in autism compared to that of the control group (P<0.05). Although there was no significant difference in GR activity between autism and control groups, 40% of autistic subjects showed lower GR activity than 95% confidence interval (CI) of the control group. GCL activity was also significantly reduced by 38.7% in the autistic group compared to the control group (P=0.023), and 8 of 10 autistic subjects had values below 95% CI of the control group. The ratio of protein levels of GCLC to GCLM in the autism group was significantly higher than that of the control group (P=0.022), and GCLM protein levels were reduced by 37.3% in the autistic group compared to the control group. A positive strong correlation was observed between GCL activity and protein levels of GCLM (r=0.887) and GCLC (r=0.799) subunits in control subjects but not in autistic subjects, suggesting that regulation of GCL activity is affected in autism. These results suggest that enzymes involved in GSH homeostasis have impaired

Glutathione Is Involved in Environmental Stress Responses in Rhizobium tropici, Including Acid Tolerance

PubMed Central

Riccillo, Pablo M.; Muglia, Cecilia I.; de Bruijn, Frans J.; Roe, Andrew J.; Booth, Ian R.; Aguilar, O. Mario

2000-01-01

The isolation of rhizobial strains which exhibit an intrinsic tolerance to acidic conditions has been reported and has facilitated studies on the basic mechanisms underlying acid tolerance. Rhizobium tropici strain CIAT899 displays a high intrinsic tolerance to acidity and therefore was used in this work to study the molecular basis of bacterial responses to acid conditions and other environmental stresses. We generated a collection of R. tropici CIAT899 mutants affected in acid tolerance using Tn5-luxAB mutagenesis, and one mutant strain (CIAT899-13T2), which fails to grow under acid conditions, was characterized in detail. Strain CIAT899-13T2 was found to contain a single Tn5-luxAB insertion in a gene showing a high degree of similarity with the Escherichia coli gshB gene, encoding the enzyme glutathione synthetase. Intracellular potassium pools and intracellular pH levels were found to be lower in the mutant than in the parent. The glutathione-deficient mutant was shown to be sensitive to weak organic acids, osmotic and oxidative stresses, and the presence of methylglyoxal. Glutathione restores responses to these stresses almost to wild-type levels. Our data show that in R. tropici the production of glutathione is essential for growth in extreme environmental conditions. The mutant strain CIAT899-13T2 induced effective nodules; however, it was found to be outcompeted by the wild-type strain in coinoculation experiments. PMID:10692382

Influence of galactose cataract on erythrocytic and lenticular glutathione metabolism in albino rats.

PubMed

Jyothi, M; Sanil, R; Shashidhar, S

2011-01-01

Glutathione depletion has been postulated to be the prime reason for galactose cataract. The current research seeks the prospect of targeting erythrocytes to pursue the lens metabolism by studying the glutathione system. To study the activity of the glutathione-linked scavenger enzyme system in the erythrocyte and lens of rats with cataract. Experiments were conducted in 36 male albino rats weighing 80 ± 20 g of 28 days of age. The rats were divided into two major groups, viz. experimental and control. Six rats in each group were sacrificed every 10 days, for 30 days. Cataract was induced in the experimental group by feeding the rats 30% galactose (w/w). The involvement of reduced glutathione (GSH) and the linked enzymes was studied in the erythrocytes and lens of cataractous as well as control rats. Parametric tests like one-way ANOVA and Student's 't' test were used for comparison. Correlation linear plot was used to compare the erythrocyte and lens metabolism. The concentration of GSH and the activity of linked enzymes were found decreased with the progression of cataract, and also in comparison to the control. The same linear fashion was also observed in the erythrocytes. Depletion of GSH was the prime factor for initiating galactose cataract in the rat model. This depletion may in turn result in enzyme inactivation leading to cross-linking of protein and glycation. The correlation analysis specifies that the biochemical mechanism in the erythrocytes and lens is similar in the rat model.

Lacking "Lack": A Reply to Joldersma

ERIC Educational Resources Information Center

Marshall, James D.

2007-01-01

First I would like to thank Clarence Joldersma for his review of our "Poststructuralism, Philosophy, Pedagogy" (Marshall, 2004-PPP). In particular, I would thank him for his opening sentence: "[t]his book is a response to a lack." It is the notion of a lack, noted again later in his review, which I wish to take up mainly in this response. Rather…

Antiprotozoal glutathione derivatives with flagellar membrane binding activity against T. brucei rhodesiense.

PubMed

Daunes, Sylvie; Yardley, Vanessa; Croft, Simon L; D'Silva, Claudius

2017-02-15

A new series of N-substituted S-(2,4-dinitrophenyl)glutathione dibutyl diesters were synthesized to improve in vitro anti-protozoal activity against the pathogenic parasites Trypanosoma brucei rhodesiense, Trypanosoma cruzi and Leishmania donovani. The results obtained indicate that N-substituents enhance the inhibitory properties of glutathione diesters whilst showing reduced toxicity against KB cells as in the cases of compounds 5, 9, 10, 16, 18 and 19. We suggest that the interaction of N-substituted S-(2,4-dinitrophenyl) glutathione dibutyl diesters with T. b. brucei occurs mainly by weak hydrophobic interactions such as London and van der Waals forces. A QSAR study indicated that the inhibitory activity of the peptide is associated negatively with the average number of C atoms, N C and positively to S ZX, the ZX shadow a geometric descriptor related to molecular size and orientation of the compound. HPLC-UV studies in conjunction with optical microscopy indicate that the observed selectivity of inhibition of these compounds against bloodstream form T. b. brucei parasites in comparison to L. donovani under the same conditions is due to intracellular uptake via endocytosis in the flagellar pocket. Copyright © 2016. Published by Elsevier Ltd.

High level of reduced glutathione contributes to detoxification of lipid peroxide-derived reactive carbonyl species in transgenic Arabidopsis overexpressing glutathione reductase under aluminum stress.

PubMed

Yin, Lina; Mano, Jun'ichi; Tanaka, Kiyoshi; Wang, Shiwen; Zhang, Meijuan; Deng, Xiping; Zhang, Suiqi

2017-10-01

Lipid peroxide-derived reactive carbonyl species (RCS), generated downstream of reactive oxygen species (ROS), are critical damage-inducing species in plant aluminum (Al) toxicity. In mammals, RCS are scavenged primarily by glutathione (reduced form of glutathione, GSH), but in plant Al stress, contribution of GSH to RCS detoxification has not been evaluated. In this study, Arabidopsis plants overexpressing the gene AtGR1 (accession code At3g24170), encoding glutathione reductase (GR), were generated, and their performance under Al stress was examined. These transgenic plants (GR-OE plants) showed higher GSH levels and GSH/GSSG (oxidized form of GSH) ratio, and an improved Al tolerance as they suffered less inhibition of root growth than wild-type under Al stress. Exogenous application of 4-hydroxy-2-nonenal, an RCS responsible for Al toxicity in roots, markedly inhibited root growth in wild-type plants. GR-OE plants suffered significantly smaller inhibition, indicating that the enhanced GSH level increased the capacity of RCS detoxification. The generation of H 2 O 2 due to Al stress in GR-OE plants was lower by 26% than in wild-type. Levels of various RCS, such as malondialdehyde, butyraldehyde, phenylacetaldehyde, (E)-2-heptenal and n-octanal, were suppressed by more than 50%. These results indicate that high levels of GSH and GSH/GSSG ratio by GR overexpression contributed to the suppression of not only ROS, but also RCS. Thus, the maintenance of GSH level by overexpressing GR reinforces dual detoxification functions in plants and is an efficient approach to enhance Al tolerance. © 2017 Scandinavian Plant Physiology Society.

Metagenomic analysis of nitrate-reducing bacteria in the oral cavity: implications for nitric oxide homeostasis.

PubMed

Hyde, Embriette R; Andrade, Fernando; Vaksman, Zalman; Parthasarathy, Kavitha; Jiang, Hong; Parthasarathy, Deepa K; Torregrossa, Ashley C; Tribble, Gena; Kaplan, Heidi B; Petrosino, Joseph F; Bryan, Nathan S

2014-01-01

The microbiota of the human lower intestinal tract helps maintain healthy host physiology, for example through nutrient acquisition and bile acid recycling, but specific positive contributions of the oral microbiota to host health are not well established. Nitric oxide (NO) homeostasis is crucial to mammalian physiology. The recently described entero-salivary nitrate-nitrite-nitric oxide pathway has been shown to provide bioactive NO from dietary nitrate sources. Interestingly, this pathway is dependent upon oral nitrate-reducing bacteria, since humans lack this enzyme activity. This pathway appears to represent a newly recognized symbiosis between oral nitrate-reducing bacteria and their human hosts in which the bacteria provide nitrite and nitric oxide from nitrate reduction. Here we measure the nitrate-reducing capacity of tongue-scraping samples from six healthy human volunteers, and analyze metagenomes of the bacterial communities to identify bacteria contributing to nitrate reduction. We identified 14 candidate species, seven of which were not previously believed to contribute to nitrate reduction. We cultivated isolates of four candidate species in single- and mixed-species biofilms, revealing that they have substantial nitrate- and nitrite-reduction capabilities. Colonization by specific oral bacteria may thus contribute to host NO homeostasis by providing nitrite and nitric oxide. Conversely, the lack of specific nitrate-reducing communities may disrupt the nitrate-nitrite-nitric oxide pathway and lead to a state of NO insufficiency. These findings may also provide mechanistic evidence for the oral systemic link. Our results provide a possible new therapeutic target and paradigm for NO restoration in humans by specific oral bacteria.

Metagenomic Analysis of Nitrate-Reducing Bacteria in the Oral Cavity: Implications for Nitric Oxide Homeostasis

PubMed Central

Hyde, Embriette R.; Andrade, Fernando; Vaksman, Zalman; Parthasarathy, Kavitha; Jiang, Hong; Parthasarathy, Deepa K.; Torregrossa, Ashley C.; Tribble, Gena; Kaplan, Heidi B.; Petrosino, Joseph F.; Bryan, Nathan S.

2014-01-01

The microbiota of the human lower intestinal tract helps maintain healthy host physiology, for example through nutrient acquisition and bile acid recycling, but specific positive contributions of the oral microbiota to host health are not well established. Nitric oxide (NO) homeostasis is crucial to mammalian physiology. The recently described entero-salivary nitrate-nitrite-nitric oxide pathway has been shown to provide bioactive NO from dietary nitrate sources. Interestingly, this pathway is dependent upon oral nitrate-reducing bacteria, since humans lack this enzyme activity. This pathway appears to represent a newly recognized symbiosis between oral nitrate-reducing bacteria and their human hosts in which the bacteria provide nitrite and nitric oxide from nitrate reduction. Here we measure the nitrate-reducing capacity of tongue-scraping samples from six healthy human volunteers, and analyze metagenomes of the bacterial communities to identify bacteria contributing to nitrate reduction. We identified 14 candidate species, seven of which were not previously believed to contribute to nitrate reduction. We cultivated isolates of four candidate species in single- and mixed-species biofilms, revealing that they have substantial nitrate- and nitrite-reduction capabilities. Colonization by specific oral bacteria may thus contribute to host NO homeostasis by providing nitrite and nitric oxide. Conversely, the lack of specific nitrate-reducing communities may disrupt the nitrate-nitrite-nitric oxide pathway and lead to a state of NO insufficiency. These findings may also provide mechanistic evidence for the oral systemic link. Our results provide a possible new therapeutic target and paradigm for NO restoration in humans by specific oral bacteria. PMID:24670812

Dual pharmacological inhibition of glutathione and thioredoxin systems synergizes to kill colorectal carcinoma stem cells.

PubMed

Tanaka, Genki; Inoue, Ken-Ichi; Shimizu, Takayuki; Akimoto, Kazumi; Kubota, Keiichi

2016-09-01

NRF2 stabilizes redox potential through genes for glutathione and thioredoxin antioxidant systems. Whether blockade of glutathione and thioredoxin is useful in eliminating cancer stem cells remain unknown. We used xenografts derived from colorectal carcinoma patients to investigate the pharmacological inhibition of glutathione and thioredoxin systems. Higher expression of five glutathione S-transferase isoforms (GSTA1, A2, M4, O2, and P1) was observed in xenograft-derived spheroids than in fibroblasts. Piperlongumine (2.5-10 μmol/L) and auranofin (0.25-4 μmol/L) were used to inhibit glutathione S-transferase π and thioredoxin reductase, respectively. Piperlongumine or auranofin alone up-regulated the expression of NRF2 target genes, but not TP53 targets. While piperlongumine showed modest cancer-specific cell killing (IC50 difference between cancer spheroids and fibroblasts: P = 0.052), auranofin appeared more toxic to fibroblasts (IC50 difference between cancer spheroids and fibroblasts: P = 0.002). The synergism of dual inhibition was evaluated by determining the Combination Index, based on the number of surviving cells with combination treatments. Molar ratios indicated synergism in cancer spheroids, but not in fibroblasts: (auranofin:piperlongumine) = 2:5, 1:5, 1:10, and 1:20. Cancer-specific cell killing was achieved at the following drug concentrations (auranofin:piperlongumine): 0.25:2.5 μmol/L, 0.5:2.5 μmol/L, or 0.25:5 μmol/L. The dual inhibition successfully decreased CD44v9 surface presentation and delayed tumor emergence in nude mouse. However, a small subpopulation persistently survived and accumulated phosphorylated histone H2A. Such "persisters" still retained lesser but significant tumorigenicity. Thus, dual inhibition of glutathione S-transferase π and thioredoxin reductase could be a feasible option for decreasing the tumor mass and CD44v9-positive fraction by disrupting redox regulation. © 2016 The Authors. Cancer Medicine

The cell-based L-glutathione protection assays to study endocytosis and recycling of plasma membrane proteins.

PubMed

Cihil, Kristine M; Swiatecka-Urban, Agnieszka

2013-12-13

Membrane trafficking involves transport of proteins from the plasma membrane to the cell interior (i.e. endocytosis) followed by trafficking to lysosomes for degradation or to the plasma membrane for recycling. The cell based L-glutathione protection assays can be used to study endocytosis and recycling of protein receptors, channels, transporters, and adhesion molecules localized at the cell surface. The endocytic assay requires labeling of cell surface proteins with a cell membrane impermeable biotin containing a disulfide bond and the N-hydroxysuccinimide (NHS) ester at 4 ºC - a temperature at which membrane trafficking does not occur. Endocytosis of biotinylated plasma membrane proteins is induced by incubation at 37 ºC. Next, the temperature is decreased again to 4 ºC to stop endocytic trafficking and the disulfide bond in biotin covalently attached to proteins that have remained at the plasma membrane is reduced with L-glutathione. At this point, only proteins that were endocytosed remain protected from L-glutathione and thus remain biotinylated. After cell lysis, biotinylated proteins are isolated with streptavidin agarose, eluted from agarose, and the biotinylated protein of interest is detected by western blotting. During the recycling assay, after biotinylation cells are incubated at 37 °C to load endocytic vesicles with biotinylated proteins and the disulfide bond in biotin covalently attached to proteins remaining at the plasma membrane is reduced with L-glutathione at 4 ºC as in the endocytic assay. Next, cells are incubated again at 37 °C to allow biotinylated proteins from endocytic vesicles to recycle to the plasma membrane. Cells are then incubated at 4 ºC, and the disulfide bond in biotin attached to proteins that recycled to the plasma membranes is reduced with L-glutathione. The biotinylated proteins protected from L-glutathione are those that did not recycle to the plasma membrane.

Effects of glutamine supplementation on gut barrier, glutathione content and acute phase response in malnourished rats during inflammatory shock

PubMed Central

Belmonte, Liliana; Coëffier, Moïse; Pessot, Florence Le; Miralles-Barrachina, Olga; Hiron, Martine; Leplingard, Antony; Lemeland, Jean-François; Hecketsweiler, Bernadette; Daveau, Maryvonne; Ducrotté, Philippe; Déchelotte, Pierre

2007-01-01

AIM: To evaluate the effect of glutamine on intestinal mucosa integrity, glutathione stores and acute phase response in protein-depleted rats during an inflammatory shock. METHODS: Plasma acute phase proteins (APP), jejunal APP mRNA levels, liver and jejunal glutathione concentrations were measured before and one, three and seven days after turpentine injection in 4 groups of control, protein-restricted, protein-restricted rats supplemented with glutamine or protein powder. Bacterial translocation in mesenteric lymph nodes and intestinal morphology were also assessed. RESULTS: Protein deprivation and turpentine injection significantly reduced jejunal villus height, and crypt depths. Mucosal glutathione concentration significantly decreased in protein-restricted rats. Before turpentine oil, glutamine supplementation restored villus heights and glutathione concentration (3.24 ± 1.05 vs 1.72 ± 0.46 μmol/g tissue, P < 0.05) in the jejunum, whereas in the liver glutathione remained low. Glutamine markedly increased jejunal α1-acid glycoprotein mRNA level after turpentine oil but did not affect its plasma concentration. Bacterial translocation in protein-restricted rats was not prevented by glutamine or protein powder supplementation. CONCLUSION: Glutamine restored gut glutathione stores and villus heights in malnourished rats but had no preventive effect on bacterial translocation in our model. PMID:17569119

The effect of alcohol and hydrogen peroxide on liver hepcidin gene expression in mice lacking antioxidant enzymes, glutathione peroxidase-1 or catalase.

PubMed

Harrison-Findik, Duygu Dee; Lu, Sizhao

2015-05-06

This study investigates the regulation of hepcidin, the key iron-regulatory molecule, by alcohol and hydrogen peroxide (H2O2) in glutathione peroxidase-1 (gpx-1(-/-)) and catalase (catalase(-/-)) knockout mice. For alcohol studies, 10% ethanol was administered in the drinking water for 7 days. Gpx-1(-/-) displayed significantly higher hepatic H2O2 levels than catalase(-/-) compared to wild-type mice, as measured by 2'-7'-dichlorodihydrofluorescein diacetate (DCFH-DA). The basal level of liver hepcidin expression was attenuated in gpx-1(-/-) mice. Alcohol increased H2O2 production in catalase(-/-) and wild-type, but not gpx-1(-/-), mice. Hepcidin expression was inhibited in alcohol-fed catalase(-/-) and wild-type mice. In contrast, alcohol elevated hepcidin expression in gpx-1(-/-) mice. Gpx-1(-/-) mice also displayed higher level of basal liver CHOP protein expression than catalase(-/-) mice. Alcohol induced CHOP and to a lesser extent GRP78/BiP expression, but not XBP1 splicing or binding of CREBH to hepcidin gene promoter, in gpx-1(-/-) mice. The up-regulation of hepatic ATF4 mRNA levels, which was observed in gpx-1(-/-) mice, was attenuated by alcohol. In conclusion, our findings strongly suggest that H2O2 inhibits hepcidin expression in vivo. Synergistic induction of CHOP by alcohol and H2O2, in the absence of gpx-1, stimulates liver hepcidin gene expression by ER stress independent of CREBH.

Lack of surface-associated microorganisms in a mixed species community of freshwater Unionidae

USGS Publications Warehouse

Nichols, S. Jerrine; Allen, J.; Walker, G.; Yokoyama, M.; Garling, D.

2001-01-01

To determine whether unionids contain surface-attached endosymbiotic bacteria, ciliates, or fungi, we used scanning electron microscopy to examine the epithelial surface of various organs within the digestive systems and mantle cavity of temperate river and lake unionids on a seasonal basis. We also cultured material removed from the lumen of these same organs and from the mantle cavity to detect cellobiose-, cellulose-, and chitin- degrading microbes. No true endosymbiotic fauna were observed attached to the surface of the digestive or mantle tissues of any species of unionid. Microbial growth on cellulose or chitin bacteriological media varied by season and habitat, but not by unionid species or source of the isolate. Lake unionids did not contain microbes capable of digesting cellulose or chitin, whereas unionids from the river site did in March and August, but not in December. Since these cultured cellulose- and chitin-degrading bacteria were never found attached to any unionid tissues, they appeared to be transient forms, not true endosymbionts. Microbes capable of digesting cellobiose were found in every unionid collected in all seasons and habitats, but again, no microbes were directly observed attached to unionid tissues. If unionids, like most other vertebrates, lack digestive enzymes required to initiate primary bond breakage, then the lack of cellulolytic and chitinolytic endosymbionts would affect the ability to utilize cellulose or chitin foods. Thus, in captivity dry feeds based on corn, soybeans, or nauplii should be pre-digested to ensure maximum absorption of nutrients by unionids. The lack of celluloytic or chitinolytic endosymbionts should not affect relocation success, though the seasonal role of transient microbes in unionid nutrition requires further investigation.

Cell Proliferation, Reactive Oxygen and Cellular Glutathione

PubMed Central

Day, Regina M.; Suzuki, Yuichiro J.

2005-01-01

A variety of cellular activities, including metabolism, growth, and death, are regulated and modulated by the redox status of the environment. A biphasic effect has been demonstrated on cellular proliferation with reactive oxygen species (ROS)—especially hydrogen peroxide and superoxide—in which low levels (usually submicromolar concentrations) induce growth but higher concentrations (usually >10–30 micromolar) induce apoptosis or necrosis. This phenomenon has been demonstrated for primary, immortalized and transformed cell types. However, the mechanism of the proliferative response to low levels of ROS is not well understood. Much of the work examining the signal transduction by ROS, including H2O2, has been performed using doses in the lethal range. Although use of higher ROS doses have allowed the identification of important signal transduction pathways, these pathways may be activated by cells only in association with ROS-induced apoptosis and necrosis, and may not utilize the same pathways activated by lower doses of ROS associated with increased cell growth. Recent data has shown that low levels of exogenous H2O2 up-regulate intracellular glutathione and activate the DNA binding activity toward antioxidant response element. The modulation of the cellular redox environment, through the regulation of cellular glutathione levels, may be a part of the hormetic effect shown by ROS on cell growth. PMID:18648617

Safety of long-term feeding of dl-alpha-lipoic acid and its effect on reduced glutathione:oxidized glutathione ratios in beagles.

PubMed

Zicker, Steven C; Hagen, Tory M; Joisher, Neha; Golder, Christina; Joshi, Dinesh K; Miller, E Phillip

2002-01-01

Alpha-lipoic acid is touted as a powerful antioxidant and possibly a conditionally essential nutrient in older mammals. The safety and efficacy of dl-alpha-lipoic acid was evaluated in 30 adult beagles that were evenly randomized into five groups, each of which was fed one of five different foods with varying inclusion rates of dl-alpha-lipoic acid (0, 150, 1500, 3000, and 4500 ppm). All dogs were fed their respective portion of food daily as their sole source of nutrition for 6 months. Evaluations included general health, body weight, food intake, hematologic and serum biochemical parameters, and glutathione:oxidized glutathione (GSH:GSSG) ratios in lymphocytes. No signs of toxicity were observed at any except the highest level of dl-alpha-lipoic acid inclusion, and no consistent abnormalities were noted in hematologic or biochemical measures at any level. There was a significant overall effect (P< .05) of food on the difference of GSH:GSSG ratio between Day 84 and Day 0. All inclusions of dl-alpha-lipoic acid increased the ratio of GSH:GSSG with the largest numeric improvement occurring at the lowest inclusion rate (150 ppm).

Deficient synthesis of glutathione underlies oxidative stress in aging and can be corrected by dietary cysteine and glycine supplementation1234

PubMed Central

Patel, Sanjeet G; Guthikonda, Anuradha P; Reid, Marvin; Balasubramanyam, Ashok; Taffet, George E; Jahoor, Farook

2011-01-01

Background: Aging is associated with oxidative stress, but underlying mechanisms remain poorly understood. Objective: We tested whether glutathione deficiency occurs because of diminished synthesis and contributes to oxidative stress in aging and whether stimulating glutathione synthesis with its precursors cysteine and glycine could alleviate oxidative stress. Design: Eight elderly and 8 younger subjects received stable-isotope infusions of [2H2]glycine, after which red blood cell (RBC) glutathione synthesis and concentrations, plasma oxidative stress, and markers of oxidant damage (eg, F2-isoprostanes) were measured. Elderly subjects were restudied after 2 wk of glutathione precursor supplementation. Results: Compared with younger control subjects, elderly subjects had markedly lower RBC concentrations of glycine (486.7 ± 28.3 compared with 218.0 ± 23.7 μmol/L; P < 0.01), cysteine (26.2 ± 1.4 compared with 19.8 ± 1.3 μmol/L; P < 0.05), and glutathione (2.08 ± 0.12 compared with 1.12 ± 0.18 mmol/L RBCs; P < 0.05); lower glutathione fractional (83.14 ± 6.43% compared with 45.80 ± 5.69%/d; P < 0.01) and absolute (1.73 ± 0.16 compared with 0.55 ± 0.12 mmol/L RBCs per day; P < 0.01) synthesis rates; and higher plasma oxidative stress (304 ± 16 compared with 346 ± 20 Carratelli units; P < 0.05) and plasma F2-isoprostanes (97.7 ± 8.3 compared with 136.3 ± 11.3 pg/mL; P < 0.05). Precursor supplementation in elderly subjects led to a 94.6% higher glutathione concentration, a 78.8% higher fractional synthesis rate, a 230.9% higher absolute synthesis rate, and significantly lower plasma oxidative stress and F2-isoprostanes. No differences in these measures were observed between younger subjects and supplemented elderly subjects. Conclusions: Glutathione deficiency in elderly humans occurs because of a marked reduction in synthesis. Dietary supplementation with the glutathione precursors cysteine and glycine fully restores glutathione synthesis and

3-Sulfanyl-4-methylpentan-1-ol in Dry-Hopped Beers: First Evidence of Glutathione S-Conjugates in Hop (Humulus lupulus L.).

PubMed

Kankolongo Cibaka, Marie-Lucie; Decourrière, Laura; Lorenzo-Alonso, Celso-José; Bodart, Etienne; Robiette, Raphaël; Collin, Sonia

2016-11-16

Monovarietal dry-hopped beers were produced with the dual-purpose hop cultivars Amarillo, Hallertau Blanc, and Mosaic. The grapefruit-like 3-sulfanyl-4-methylpentan-1-ol was found in all three beers at concentrations much higher than expected on the basis of the free thiol content in hop. Even cysteinylated precursors proved unable to explain our results. As observed in wine, the occurrence of S-glutathione precursors was therefore suspected in hop. The analytical standards of S-3-(4-methyl-1-hydroxypentyl)glutathione, never described before, and of S-3-(1-hydroxyhexyl)glutathione, previously evidenced in grapes, were chemically synthesized. An optimized extraction of glutathionylated precursors was then applied to Amarillo, Hallertau Blanc, and Mosaic hop samples. HPLC-ESI(+)MS/MS revealed, for the first time, the occurrence of S-3-(1-hydroxyhexyl)glutathione and S-3-(4-methyl-1-hydroxypentyl)glutathione in hop, at levels well above those reported for their cysteinylated counterparts. S-3-(1-Hydroxyhexyl)glutathione emerged in all cases as the major adduct in hop. Yet, although 3-sulfanylhexan-1-ol seems relatively ubiquitous in free, cysteinylated, and glutathionylated forms, the glutathione adduct of 3-sulfanyl-4-methylpentan-1-ol, never evidenced in other plants up to now, was found only in the Hallertau Blanc variety.

Effects of Bacterial Host and Dichloromethane Dehalogenase on the Competitiveness of Methylotrophic Bacteria Growing with Dichloromethane

PubMed Central

Gisi, Daniel; Willi, Laurent; Traber, Hubert; Leisinger, Thomas; Vuilleumier, Stéphane

1998-01-01

Methylobacterium sp. strain DM4 and Methylophilus sp. strain DM11 can grow with dichloromethane (DCM) as the sole source of carbon and energy by virtue of homologous glutathione-dependent DCM dehalogenases with markedly different kinetic properties (the kcat values of the enzymes of these strains are 0.6 and 3.3 s−1, respectively, and the Km values are 9 and 59 μM, respectively). These strains, as well as transconjugant bacteria expressing the DCM dehalogenase gene (dcmA) from DM11 or DM4 on a broad-host-range plasmid in the background of dcmA mutant DM4-2cr, were investigated by growing them under growth-limiting conditions and in the presence of an excess of DCM. The maximal growth rates and maximal levels of dehalogenase for chemostat-adapted bacteria were higher than the maximal growth rates and maximal levels of dehalogenase for batch-grown bacteria. The substrate saturation constant of strain DM4 was much lower than the Km of its associated dehalogenase, suggesting that this strain is adapted to scavenge low concentrations of DCM. Strains and transconjugants expressing the DCM dehalogenase from strain DM11, on the other hand, had higher growth rates than bacteria expressing the homologous dehalogenase from strain DM4. Competition experiments performed with pairs of DCM-degrading strains revealed that a strain expressing the dehalogenase from DM4 had a selective advantage in continuous culture under substrate-limiting conditions, while strains expressing the DM11 dehalogenase were superior in batch culture when there was an excess of substrate. Only DCM-degrading bacteria with a dcmA gene similar to that from strain DM4, however, were obtained in batch enrichment cultures prepared with activated sludge from sewage treatment plants. PMID:9546153

In vivo toxicity, biodistribution, and clearance of glutathione-coated gold nanoparticles.

PubMed

Simpson, Carrie A; Salleng, Kenneth J; Cliffel, David E; Feldheim, Daniel L

2013-02-01

Gold nanoparticles are emerging as promising materials from which to construct nanoscale therapeutics and therapeutic delivery systems. However, animal studies have shown that gold nanoparticles modified with certain thiol monolayers such as tiopronin can cause renal complications and morbidity. Although these effects may be eliminated by coadsorbing small amounts of polyethylene glycol (PEG) onto the nanoparticle surface, PEG can also lower cellular internalization efficiency and binding interactions with protein disease targets, significantly reducing the potential for using gold nanoparticles as therapeutics. Using ICP-MS analysis of blood, urine, and several organs, we show in this article that glutathione-coated gold nanoparticles (1.2 nm ± 0.9 nm) cause no morbidity at any concentration up to and including 60 μM and target primary organs although providing gradual dissipation and clearance over time. This study suggests that glutathione may be an attractive alternative to PEG in the design of gold nanoparticle therapeutics. This study describes the utility and toxicity of glutathione coated gold nanoparticles in comparison to PEGylated counterparts that are commonly used to increase "Stealth" properties and lower cytotoxicity. Too much PEG on the NPs can lead to lower cellular internalization efficiency and less efficient binding interactions with protein disease targets, significantly reducing the potential for using gold nanoparticles as therapeutics. Copyright © 2013 Elsevier Inc. All rights reserved.

Energetics of Glutathione Binding to Human Eukaryotic Elongation Factor 1 Gamma: Isothermal Titration Calorimetry and Molecular Dynamics Studies.

PubMed

Tshabalala, Thabiso N; Tomescu, Mihai-Silviu; Prior, Allan; Balakrishnan, Vijayakumar; Sayed, Yasien; Dirr, Heini W; Achilonu, Ikechukwu

2016-12-01

The energetics of ligand binding to human eukaryotic elongation factor 1 gamma (heEF1γ) was investigated using reduced glutathione (GSH), oxidised glutathione (GSSG), glutathione sulfonate and S-hexylglutathione as ligands. The experiments were conducted using isothermal titration calorimetry, and the findings were supported using computational studies. The data show that the binding of these ligands to heEF1γ is enthalpically favourable and entropically driven (except for the binding of GSSG). The full length heEF1γ binds GSSG with lower affinity (K d  = 115 μM), with more hydrogen-bond contacts (ΔH = -73.8 kJ/mol) and unfavourable entropy (-TΔS = 51.7 kJ/mol) compared to the glutathione transferase-like N-terminus domain of heEF1γ, which did not show preference to any specific ligand. Computational free binding energy calculations from the 10 ligand poses show that GSSG and GSH consistently bind heEF1γ, and that both ligands bind at the same site with a folded bioactive conformation. This study reveals the possibility that heEF1γ is a glutathione-binding protein.

The relation of blood pressure and carotid intima-media thickness with the glutathione cycle in a young bi-ethnic population: the African-PREDICT study.

PubMed

Myburgh, Caitlynd; Huisman, Hugo W; Mels, Catharina M C

2018-04-01

Oxidative stress has been implicated in the development of hypertension, arterial stiffness and atherosclerosis. Optimal functioning of the enzymatic antioxidant system is central to prevent increased oxidative stress and its consequences. We aimed to investigate the relationships of ambulatory blood pressure and carotid intima-media thickness with enzyme activities of the glutathione cycle in 396 young, black and white South Africans of the African-PREDICT study. Ambulatory blood pressure and carotid intima-media thickness were measured and glutathione peroxidase and glutathione reductase activities were analyzed. Black participants had higher reactive oxygen species (men: p = 0.019; women: borderline p = 0.064) and total glutathione (both p < 0.001), but lower glutathione peroxidase activity and total antioxidant status (all p < 0.001). In black men, ambulatory pulse pressure was negatively associated with glutathione peroxidase activity (R 2  = 0.19; β = -0.25; p = 0.06). Black and white women displayed positive associations of ambulatory systolic blood pressure (black: R 2  = 0.25; β = 0.21; p = 0.048; white: R 2  = 0.44; β = 0.18; p = 0.016) with glutathione reductase activity, whereas white men displayed a positive association of ambulatory pulse pressure with glutathione reductase activity (R 2  = 0.25; β = 0.29; p = 0.01). The lower glutathione peroxidase activity and total antioxidant status, the higher reactive oxygen species, as well as the negative association between ambulatory pulse pressure and glutathione peroxidase activity in the black men suggest that oxidative stress may be associated with early vascular changes in this group. In the other three groups, the positive associations of blood pressure with glutathione reductase activity suggest a possible role for adequate glutathione reductase activity in preventing or delaying the development of hypertension.

Enzymatic Production of Glutathione by Bifunctional γ-Glutamylcysteine Synthetase/Glutathione Synthetase Coupled with In Vitro Acetate Kinase-Based ATP Generation.

PubMed

Jiang, Yu; Tao, Rongsheng; Shen, Zhengquan; Sun, Liangdong; Zhu, Fuyun; Yang, Sheng

2016-12-01

Glutathione (γ-glutamyl-L-cysteinylglycine, GSH) is a pharmaceutical compound often used in food additives and the cosmetics industry. GSH can be produced biologically from L-glutamic acid, L-cysteine, and glycine through an enzymatic process traditionally involving two sequential adenosine triphosphate (ATP)-dependent reactions catalyzed by γ-glutamylcysteine synthetase (γ-GCS or GSHI, EC 6.3.2.2) and GSH synthetase (GS or GSHII, EC 6.3.2.3). Here, we report the enzymatic production of GSH by recombinant cell-free bifunctional γ-glutamylcysteine synthetase/glutathione synthetase (γ-GCS-GS or GshF) coupled with in vitro acetate kinase-based ATP generation. GSH production by an acetate kinase-integrated Escherichia coli Rosetta(DE3) mutant expressing Streptococcus thermophilus GshF reached 18.3 ± 0.1 g l -1 (59.5 ± 0.3 mM) within 3 h, with a molar yield of 0.75 ± 0.00 mol mol -1 added cysteine and a productivity of 6.1 ± 0.0 g l -1  h -1 . This is the highest GSH titer reported to date. This newly developed biocatalytic process offers a promising approach for meeting the industrial requirements for GSH production.

Zinc and copper in animal feed – development of resistance and co-resistance to antimicrobial agents in bacteria of animal origin

PubMed Central

Yazdankhah, Siamak; Rudi, Knut; Bernhoft, Aksel

2014-01-01

Farmed animals such as pig and poultry receive additional Zn and Cu in their diets due to supplementing elements in compound feed as well as medical remedies. Enteral bacteria in farmed animals are shown to develop resistance to trace elements such as Zn and Cu. Resistance to Zn is often linked with resistance to methicillin in staphylococci, and Zn supplementation to animal feed may increase the proportion of multiresistant E. coli in the gut. Resistance to Cu in bacteria, in particular enterococci, is often associated with resistance to antimicrobial drugs like macrolides and glycopeptides (e.g. vancomycin). Such resistant bacteria may be transferred from the food-producing animals to humans (farmers, veterinarians, and consumers). Data on dose-response relation for Zn/Cu exposure and resistance are lacking; however, it seems more likely that a resistance-driven effect occurs at high trace element exposure than at more basal exposure levels. There is also lack of data which could demonstrate whether Zn/Cu-resistant bacteria may acquire antibiotic resistance genes/become antibiotics resistant, or if antibiotics-resistant bacteria are more capable to become Zn/Cu resistant than antibiotics-susceptible bacteria. Further research is needed to elucidate the link between Zn/Cu and antibiotic resistance in bacteria. PMID:25317117

Reversal of hypermethylation and reactivation of glutathione S-transferase pi 1 gene by curcumin in breast cancer cell line.

PubMed

Kumar, Umesh; Sharma, Ujjawal; Rathi, Garima

2017-02-01

One of the mechanisms for epigenetic silencing of tumor suppressor genes is hypermethylation of cytosine residue at CpG islands at their promoter region that contributes to malignant progression of tumor. Therefore, activation of tumor suppressor genes that have been silenced by promoter methylation is considered to be very attractive molecular target for cancer therapy. Epigenetic silencing of glutathione S-transferase pi 1, a tumor suppressor gene, is involved in various types of cancers including breast cancer. Epigenetic silencing of tumor suppressor genes can be reversed by several molecules including natural compounds such as polyphenols that can act as a hypomethylating agent. Curcumin has been found to specifically target various tumor suppressor genes and alter their expression. To check the effect of curcumin on the methylation pattern of glutathione S-transferase pi 1 gene in MCF-7 breast cancer cell line in dose-dependent manner. To check the reversal of methylation pattern of hypermethylated glutathione S-transferase pi 1, MCF-7 breast cancer cell line was treated with different concentrations of curcumin for different time periods. DNA and proteins of treated and untreated cell lines were isolated, and methylation status of the promoter region of glutathione S-transferase pi 1 was analyzed using methylation-specific polymerase chain reaction assay, and expression of this gene was analyzed by immunoblotting using specific antibodies against glutathione S-transferase pi 1. A very low and a nontoxic concentration (10 µM) of curcumin treatment was able to reverse the hypermethylation and led to reactivation of glutathione S-transferase pi 1 protein expression in MCF-7 cells after 72 h of treatment, although the IC 50 value of curcumin was found to be at 20 µM. However, curcumin less than 3 µM of curcumin could not alter the promoter methylation pattern of glutathione S-transferase pi 1. Treatment of breast cancer MCF-7 cells with curcumin

Efficacy of Jian'ganle () versus Hugan Pian (), glucuronolactone and reduced glutathione in prevention of antituberculosis drug-induced liver injury.

PubMed

Zhang, Quan; Zhong, Fang-ying; Wu, Meng; Zhang, Xin-ping

2014-06-01

Evidence-based medicine is advocated by WHO and adopted by developed countries for many years. In China, however, the selection of essential medicine and various medical insurance reimbursement schemes medicine is usually based on experts' experience of prescription practice which is under heavy critics resulting from the lack of related comparative efficacy and evidence-based research. The efficacy of Jian'ganle in prevention of drug-induced liver injury (DILI) caused by antituberculotics was evaluated in this study by comparison with Hugan Pian, glucuronolactone and reduced glutathione. Evidence was provided for relevant sectors such as Ministry for Human Resources and Social Security of the People's Republic of China and National Health and Family Planning Commission of the People's Republic of China to select and renew the Essential Medicine List (EML), the new rural cooperative medical scheme in China (NRCMS) list or the reimbursement list of industrial injury insurance. A total of 189 patients with initial pulmonary tuberculosis were divided into four groups who took antituberculotics combined with Jian'ganle, Hugan Pian, glucuronolactone and reduced glutathione respectively. Their liver function profile including alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), direct bilirubin (DBIL), total protein (TP), albumin (A) and globulin (G) were detected at admission as baseline and after treatment. The Jian'ganle group was compared with the three others by chi-square tests. In an aspect of maintaining bilirubin indexes normal, Jian'ganle was more efficacious than glucuronolactone. And Jian'ganle had a little more efficacy than reduced glutathione to maintain protein indexes normal as well. And the therapeutic regimen of antituberculotics combined with Jian'ganle was the best in treating tuberculosis and preventing DILI at the same time. The study showed that among the four hepatinicas which demonstrated similar prevention

Molecularly imprinted polymer for glutathione by modified precipitation polymerization and its application to determination of glutathione in supplements.

PubMed

Nakamura, Yukari; Masumoto, Shizuka; Matsunaga, Hisami; Haginaka, Jun

2017-09-10

Molecularly imprinted polymers (MIP) particles for glutathione (GSH) with a narrow particle size distribution were prepared by modified precipitation polymerization using methacrylic acid as a functional monomer, divinylbenzene as a crosslinker and water as a co-solvent. The particle diameters of the MIP and non-imprinted polymer (NIP) prepared under the optimum conditions were 3.81±0.95 (average±standard deviation) and 3.39±1.22μm, respectively. The retention and molecular-recognition properties of the prepared MIP were evaluated using a mixture of acetonitrile and water as a mobile phase in hydrophilic interaction chromatography. With an increase of acetonitrile content, the retention factor of GSH was increased on the MIP. In addition to shape recognition, hydrophilic interactions seem to work for the recognition of GSH on the MIP. The MIP had a specific molecular-recognition ability for GSH, while glutathione disulfide, l-Glu, l-Cys, Gly-Gly and l-Cys-Gly could not be retained or recognized on the MIP. The effect of column temperature revealed that the separation of GSH on the MIP was entropically driven. Binding experiments and Scatchard analyses revealed that one binding sites were formed on both the MIP and NIP, while the MIP gave higher affinity and capacity for GSH than the NIP. Furthermore, the MIP was successfully applied for determination of GSH in the supplements. Copyright © 2016 Elsevier B.V. All rights reserved.

Metabolic plasticity for isoprenoid biosynthesis in bacteria.

PubMed

Pérez-Gil, Jordi; Rodríguez-Concepción, Manuel

2013-05-15

Isoprenoids are a large family of compounds synthesized by all free-living organisms. In most bacteria, the common precursors of all isoprenoids are produced by the MEP (methylerythritol 4-phosphate) pathway. The MEP pathway is absent from archaea, fungi and animals (including humans), which synthesize their isoprenoid precursors using the completely unrelated MVA (mevalonate) pathway. Because the MEP pathway is essential in most bacterial pathogens (as well as in the malaria parasites), it has been proposed as a promising new target for the development of novel anti-infective agents. However, bacteria show a remarkable plasticity for isoprenoid biosynthesis that should be taken into account when targeting this metabolic pathway for the development of new antibiotics. For example, a few bacteria use the MVA pathway instead of the MEP pathway, whereas others possess the two full pathways, and some parasitic strains lack both the MVA and the MEP pathways (probably because they obtain their isoprenoids from host cells). Moreover, alternative enzymes and metabolic intermediates to those of the canonical MVA or MEP pathways exist in some organisms. Recent work has also shown that resistance to a block of the first steps of the MEP pathway can easily be developed because several enzymes unrelated to isoprenoid biosynthesis can produce pathway intermediates upon spontaneous mutations. In the present review, we discuss the major advances in our knowledge of the biochemical toolbox exploited by bacteria to synthesize the universal precursors for their essential isoprenoids.

Effects of exercise training on the glutathione antioxidant system.

PubMed

Elokda, Ahmed S; Nielsen, David H

2007-10-01

The glutathione (GSH) antioxidant system has been shown to play an important role in the maintenance of good health and disease prevention. Various approaches have been used to enhance GSH availability including diet, nutritional supplementation, and drug administration, with minor to moderate success. Exercise training has evolved as a new approach. The purpose of this study was to investigate the effects of aerobic exercise training (AET), circuit weight training (CWT), and combined training (AET+CWT) on general adaptations, and resistance to acutely induced oxidative stress, as assessed by changes in the GSH antioxidant system. Eighty healthy sedentary volunteers participated in the study who were randomly assigned to four groups: control (no exercise); AET, CWT, and AET+CWT. Exercise training programs were designed to simulate outpatient cardiac rehabilitation (40 min x 3 days x 6 weeks). Venous blood sampling was taken at rest and post maximal graded exercise test (GXT). A new improved spectrophotometric venous assay analysis technique was used. A mixed model repeated measures analysis of variance design was used with t-tests for preplanned comparisons evaluated at Bonferroni-adjusted alpha levels. Effectiveness of the exercise training programs was demonstrated by significant between-group (exercise group versus control) comparisons. AET, CWT, and AET+CWT showed significant pretraining-posttraining increases in resting GSH and glutathione-glutathione disulfide ratio (GSH:GSSG), and significant decreases in GSSG levels (P<0.005). AET+CWT showed the most pronounced effect compared with AET or CWT alone (P<0.025). This study represents the first longitudinal investigation involving the effects of multiple modes of exercise training on the GSH antioxidant system with evidence, suggesting the GHS:GSSG ratio as the most sensitive change marker. The significant findings of this study have potential clinical implications to individuals involved in cardiac and

Glutathione binding to dirhodium tetraacetate: a spectroscopic, mass spectral and computational study of an anti-tumour compound.

PubMed

Wong, Daisy L; Zhang, Angel; Faponle, Abayomi S; de Visser, Sam P; Stillman, Martin J

2017-05-24

Glutathione (γ-l-glutamyl-l-cysteinyl-glycine) is a ubiquitous tripeptide found in all plants and animals. Glutathione has key roles as a metallochaperone and as a cellular thiol involved in metabolism. Little is known about how glutathione interacts with organometallic compounds in vivo. Here, we report the reactions of glutathione in vitro with dirhodium(ii) tetraacetate (tetrakis(μ-acetato)dirhodium(ii), Rh 2 (OAc) 4 ), a compound with anti-tumour properties. Electrospray ionization mass spectrometry, UV-Visible absorption and circular dichroism spectroscopic methods were used to determine the stoichiometries and optical properties of the final conjugate. Computational analyses were used to predict the binding modes of glutathione to the Rh 2 (OAc) 4 , and report on the orbital assignments for the resulting products. We explored the competition by GSH for methionine-bound axial sites on Rh 2 (OAc) 4 to investigate the use of weak thioether to protect its cellular-based anti-cancer activity. Our study highlights the important role that axial ligation would play in deactivating or significantly decreasing the efficacy of this bimetallic anti-tumor drug. The computational data explain the stability of the mono-adduct and the appearance of new absorption bands in the UV region including retention of the Rh-Rh single bond. Additionally, these data show that glutathione can effectively disable the potency of these metallo-drugs through orbital overlap of the entire Rh-Rh core as a result of the strong binding. Electronic absorption spectroscopy, mass spectrometry and computational analysis are a powerful combination in understanding possible chemical reactions in vivo and this information can be used to synthetically tune dirhodium complexes for use in the fight against cancer.

[Dinitrosyl iron complexes with glutathione recover rats with experimental endometriosis].

PubMed

Adamian, L V; Burgova, E N; Tkachev, N A; Mikoian, V D; Stepanian, A A; Sonova, M M; Vanin, A F

2013-01-01

The effect of binuclear dinitrosyl iron complexes (DNIC) with glutathione on endometrioid tumors in rats with experimental endometriosis has been studied. The latter was induced by an autotransplantation model, where two fragments of endometrium with myometrium (2 x 2 mm) from the left uterine horn was grafted to the inner surface of the anterior abdominal wall. The test animals received intraperitoneal injections of 0.5 ml DNIC-glutathione at the dose of 12.5 micromole per kg daily for 12 days 28 days after operation. The injections resulted in more than a 2-fold decrease in the total volume of both large tumors formed from grafts and small additive tumors formed nearby grafts. The disappearance of the additive tumors was also observed in test animals. The EPR signal with g(av) = 2.03 characteristic of protein bound DNIC with thiol-containing ligands was recorded in livers, graft and additive tumors of test and control animals pointing out intensive generation of nitric oxide in rats with experimental endometriosis. Ribonucleotide reductase activation discovered by doublet the EPR signal at g = 2.0 with 2.3 mT hyperfine structure splitting was found in small tumors. The cytotoxic effect of DNIC-glutathione on endometrioid tumors was suggested to be due to DNIC degradation nearby the tumors induced by iron chelating compounds released from the tumors. The degradation resulted in release of a high amount of nitric oxide molecules and nitrosonium ions from DNICs affecting the tumors by way of the cytotoxic effect.

Effect of asoka on the intracellular glutathione levels and skin tumour promotion in mice.

PubMed

Varghese, C D; Nair, S C; Panikkar, B; Panikkar, K R

1993-04-15

The bark of Saraka asoca (asoka) is commonly used to treat various diseases by the Indian system of medicine and in Sri Lanka. Further purification and chemical analysis of the active compound from the bark extract of asoka showed that (-)-epicatechin was responsible for the observed antitumour/anticarcinogenic activity. Papilloma formation in mice initiated with 7,12-dimethylbenz[a]anthracene (DMBA) and promoted using croton oil was inhibited by the topical application of 100 mg/kg body weight (b.w.) of (-)-epicatechin isolated from asoka bark extract. Oral administration of the same dose restricted the growth of s.c. injected 20 methylcholanthrene (MCA) induced soil tissue fibrosarcomas significantly in mice. Elevations of almost 2-4-fold in the intracellular reduced glutathione and related enzymes viz., glutathione reductase and glutathione S-transferase of sarcoma-180 tumour cells were noted in the presence of 1 microgram/ml of (-)-epicatechin, further highlighting its antiproliferative effect.

A cadmium-sensitive, glutathione-deficient mutant of Arabidopsis thaliana.

PubMed Central

Howden, R; Andersen, C R; Goldsbrough, P B; Cobbett, C S

1995-01-01

The roots of the cadmium-sensitive mutant of Arabidopsis thaliana, cad1-1, become brown in the presence of cadmium. A new cadmium-sensitive mutant affected at a second locus, cad2, has been identified using this phenotype. Genetic analysis has grown that the sensitive phenotype is recessive to the wild type and segregates as a single Mendelian locus. Assays of cadmium accumulation by intact plants indicated that the mutant is deficient in its ability to sequester cadmium. Undifferentiated callus tissue was also cadmium sensitive, suggesting that the mutant phenotype is expressed at the cellular level. The level of cadmium-binding complexes formed in vivo was decreased compared with the wild type and accumulation of phytochelatins was about 10% of that in the wild type. The level of glutathione, the substrate for phytochelatin biosynthesis, in tissues of the mutant was decreased to about 15 to 30% of that in the wild type. Thus, the deficiency in phytochelatin biosynthesis can be explained by a deficiency in glutathione. PMID:7770518

Glutathione-supported arsenate reduction coupled to arsenolysis catalyzed by ornithine carbamoyl transferase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nemeti, Balazs; Gregus, Zoltan

2009-09-01

Three cytosolic phosphorolytic/arsenolytic enzymes, (purine nucleoside phosphorylase [PNP], glycogen phosphorylase, glyceraldehyde-3-phosphate dehydrogenase) have been shown to mediate reduction of arsenate (AsV) to the more toxic arsenite (AsIII) in a thiol-dependent manner. With unknown mechanism, hepatic mitochondria also reduce AsV. Mitochondria possess ornithine carbamoyl transferase (OCT), which catalyzes phosphorolytic or arsenolytic citrulline cleavage; therefore, we examined if mitochondrial OCT facilitated AsV reduction in presence of glutathione. Isolated rat liver mitochondria were incubated with AsV, and AsIII formed was quantified. Glutathione-supplemented permeabilized or solubilized mitochondria reduced AsV. Citrulline (substrate for OCT-catalyzed arsenolysis) increased AsV reduction. The citrulline-stimulated AsV reduction was abolished bymore » ornithine (OCT substrate inhibiting citrulline cleavage), phosphate (OCT substrate competing with AsV), and the OCT inhibitor norvaline or PALO, indicating that AsV reduction is coupled to OCT-catalyzed arsenolysis of citrulline. Corroborating this conclusion, purified bacterial OCT mediated AsV reduction in presence of citrulline and glutathione with similar responsiveness to these agents. In contrast, AsIII formation by intact mitochondria was unaffected by PALO and slightly stimulated by citrulline, ornithine, and norvaline, suggesting minimal role for OCT in AsV reduction in intact mitochondria. In addition to OCT, mitochondrial PNP can also mediate AsIII formation; however, its role in AsV reduction appears severely limited by purine nucleoside supply. Collectively, mitochondrial and bacterial OCT promote glutathione-dependent AsV reduction with coupled arsenolysis of citrulline, supporting the hypothesis that AsV reduction is mediated by phosphorolytic/arsenolytic enzymes. Nevertheless, because citrulline cleavage is disfavored physiologically, OCT may have little role in AsV reduction in vivo.« less

Ascorbic acid glucoside reduces neurotoxicity and glutathione depletion in mouse brain induced by nitrotriazole radiosensitazer.

PubMed

Cherdyntseva, Nadezda V; Ivanova, Anna A; Ivanov, Vladimir V; Cherdyntsev, Evgeny; Nair, Cherupally Krishnan Krishnan; Kagiya, Tsutomu V

2013-01-01

To investigate the potential of the anti-oxidant ascorbic acid glucoside (AA-2G) to modulate neurotoxicity induced by high doses of nitrotriazole radiosensitizer. Male and female C56Bl/6xCBA hybrid mice aged 8-14 weeks (weight 18-24 g) were used. Nitrotriazole drug radiosensitizer sanazole at a high dose of 2, 1 g/kg was per os administered to induce neurotoxicity at mice. Ascorbic acid glucoside was given 30 min before the sanazole administration. Serum ascorbic acid, brain glutathione level, as well as behavioral performance using open field apparatus were measured. Administration of high (non-therapeutic) doses of the nitrotriazole drug sanazole results in neurotoxicity in mice as evidenced from behavioral performance, emotional activity and depletion of the cellular antioxidant, glutathione, in the brain. The serum levels of ascorbic acid was also found reduced in high dose sanazole treated animals. Per os administration of ascorbic acid glucoside significantly reduced the neurotoxicity. This effect was associated with the prevention of glutathione depletion in mouse brain and restoring the ascorbic acid level in serum. Administration of ascorbic acid glucoside, but not ascorbic acid, before sanazole administration protected from sanazole-induced neurotoxicity by preventing the decrease in the brain reduced glutathione level and providing high level of ascorbic acid in plasma.

Ferrocene labelings as inhibitors and dual electrochemical sensors of human glutathione S-transferase P1-1.

PubMed

Martos-Maldonado, Manuel C; Quesada-Soriano, Indalecio; García-Maroto, Federico; Vargas-Berenguel, Antonio; García-Fuentes, Luís

2012-12-01

The inhibitory and sensor properties of two ferrocene conjugates, in which the ferrocene and glutathione are linked through a spacer arm of different length and chemical structure, on human Pi glutathione S-transferase, were examined by activity assays, ITC, fluorescence spectroscopy and voltammetry. Such ferrocene conjugates are strong competitive inhibitors of this enzyme with an enhanced binding affinity, the one bearing the longest spacer arm being the most potent inhibitor. Voltammetric measurements showed a strong decrease of the peak current intensity and an increase of the oxidation potential upon binding of ferrocene-glutathione conjugates to GST P1-1 showing that both conjugates can be used as dual electrochemical sensors for GST P1-1. Copyright © 2012 Elsevier Ltd. All rights reserved.

Nitric oxide alleviates aluminum-induced oxidative damage through regulating the ascorbate-glutathione cycle in roots of wheat.

PubMed

Sun, Chengliang; Liu, Lijuan; Yu, Yan; Liu, Wenjing; Lu, Lingli; Jin, Chongwei; Lin, Xianyong

2015-06-01

The possible association with nitric oxide (NO) and ascorbate-glutathione (AsA-GSH) cycle in regulating aluminum (Al) tolerance of wheat (Triticum aestivum L.) was investigated using two genotypes with different Al resistance. Exposure to Al inhibited root elongation, and triggered lipid peroxidation and oxidation of AsA to dehydroascorbate and GSH to glutathione disulfide in wheat roots. Exogenous NO significantly increased endogenous NO levels, and subsequently alleviated Al-induced inhibition of root elongation and oxidation of AsA and GSH to maintain the redox molecules in the reduced form in both wheat genotypes. Under Al stress, significantly increased activities and gene transcriptional levels of ascorbate peroxidase, glutathione reductase, and dehydroascorbate reductase, were observed in the root tips of the Al-tolerant genotype Jian-864. Nitric oxide application enhanced the activity and gene transcriptional level of these enzymes in both wheat genotypes. γ-Glutamylcysteine synthetase was not significantly affected by Al or NO, but NO treatments increased the activity of glutathione peroxidase and glutathione S-transferase to a greater extent than the Al-treated wheat seedlings. Proline was significantly decreased by Al, while it was not affected by NO. These results clearly suggest that NO protects wheat root against Al-induced oxidative stress, possibly through its regulation of the AsA-GSH cycle. © 2014 Institute of Botany, Chinese Academy of Sciences.

Modeling of Cd adsorption to goethite-bacteria composites

DOE PAGES

Qu, Chenchen; Ma, Mingkai; Chen, Wenli; ...

2017-11-21

The accurate modeling of heavy metal adsorption in complex systems is fundamental for risk assessments in soils and associated environments. Bacteria-iron (hydr)oxide associations in soils and sediments play a critical role in heavy metal immobilization. The reduced adsorption of heavy metals on these composites have been widely reported using the component additivity (CA) method. However, there is a lack of a mechanism model to account for these deviations. In this study, we established models for Cd adsorption on goethite-Pseudomonas putida composites at 1:1 and 5:1 mass ratios. Cadmium adsorption on the 5:1 composite was consistent with the additivity method. But,more » the CA method over predicted Cd adsorption by approximately 8% on the 1:1 composite at high Cd concentration. The deviation was corrected by adding the site blockage reactions between P. putida and goethite. Both CA and “CA-site masking” models for Cd adsorption onto the composites were in line with the ITC data. These results indicate that CA method in simulating Cd adsorption on bacteria-iron oxides composites is limited to low bacterial and Cd concentrations. Thus the interfacial complexation reactions that occur between iron (hydr)oxides and bacteria should be taken into account when high concentrations of bacteria and heavy metals are present.« less

Modeling of Cd adsorption to goethite-bacteria composites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Qu, Chenchen; Ma, Mingkai; Chen, Wenli

The accurate modeling of heavy metal adsorption in complex systems is fundamental for risk assessments in soils and associated environments. Bacteria-iron (hydr)oxide associations in soils and sediments play a critical role in heavy metal immobilization. The reduced adsorption of heavy metals on these composites have been widely reported using the component additivity (CA) method. However, there is a lack of a mechanism model to account for these deviations. In this study, we established models for Cd adsorption on goethite-Pseudomonas putida composites at 1:1 and 5:1 mass ratios. Cadmium adsorption on the 5:1 composite was consistent with the additivity method. But,more » the CA method over predicted Cd adsorption by approximately 8% on the 1:1 composite at high Cd concentration. The deviation was corrected by adding the site blockage reactions between P. putida and goethite. Both CA and “CA-site masking” models for Cd adsorption onto the composites were in line with the ITC data. These results indicate that CA method in simulating Cd adsorption on bacteria-iron oxides composites is limited to low bacterial and Cd concentrations. Thus the interfacial complexation reactions that occur between iron (hydr)oxides and bacteria should be taken into account when high concentrations of bacteria and heavy metals are present.« less

Preferential glutathione conjugation of a reverse diol epoxide compared to a bay region diol epoxide of phenanthrene in human hepatocytes: relevance to molecular epidemiology studies of glutathione-s-transferase polymorphisms and cancer.

PubMed

Hecht, Stephen S; Berg, Jeannette Zinggeler; Hochalter, J Bradley

2009-03-16

Bay region diol epoxides are recognized ultimate carcinogens of polycyclic aromatic hydrocarbons (PAH), and in vitro studies have demonstrated that they can be detoxified by conjugation with glutathione, leading to the widely investigated hypothesis that individuals with low activity forms of glutathione-S-transferases are at higher risk of PAH induced cancer, a hypothesis that has found at most weak support in molecular epidemiology studies. A weakness in this hypothesis was that the mercapturic acids resulting from the conjugation of PAH bay region diol epoxides had never been identified in human urine. We recently analyzed smokers' urine for mercapturic acids derived from phenanthrene, the simplest PAH with a bay region. The only phenanthrene diol epoxide-derived mercapturic acid in smokers' urine was produced from the reverse diol epoxide, anti-phenanthrene-3,4-diol-1,2-epoxide (11), not the bay region diol epoxide, anti-phenanthrene-1,2-diol-3,4-epoxide (10), which does not support the hypothesis noted above. In this study, we extended these results by examining the conjugation of phenanthrene metabolites with glutathione in human hepatocytes. We identified the mercapturic acid N-acetyl-S-(r-4,t-2,3-trihydroxy-1,2,3,4-tetrahydro-c-1-phenanthryl)-L-cysteine (14a), (0.33-35.9 pmol/mL at 10 microM 8, 24 h incubation, N = 10) in all incubations with phenanthrene-3,4-diol (8) and the corresponding diol epoxide 11, but no mercapturic acids were detected in incubations with phenanthrene-1,2-diol (7), and only trace amounts were observed in incubations with the corresponding bay region diol epoxide 10. Taken together with our previous results, these studies clearly demonstrate that glutathione conjugation of a reverse diol epoxide of phenanthrene is favored over conjugation of a bay region diol epoxide. Since reverse diol epoxides of PAH are generally weakly or nonmutagenic/carcinogenic, these results, if generalizable to other PAH, do not support the widely held

Lactic acid bacteria found in fermented fish in Thailand.

PubMed

Tanasupawat, Somboon; Okada, Sanae; Komagata, Kazuo

1998-06-01

Forty-seven strains of homofermentative rod-shaped and 5 heterofermentative sphere-shaped lactic acid bacteria were isolated from 4 kinds of fermented fish (pla-ra, pla-chom, kung-chom, and hoi-dong) in Thailand. These bacteria were separated into four groups by phenotypic and chemotaxonomic characteristics, including fluorometric DNA-DNA hybridization. Five strains (Group I) contained meso-diaminopimelic acid in the cell wall. Four strains were identified as Lactobacillus pentosus, and one strain was L. plantarum. Tested strains of this group produced DL-lactic acid. The rest of the rod-shaped bacteria, 23 strains (Group II) and 19 strains (Group III), lacked meso-diaminopimelic acid in the cell wall and were identified as L. farciminis and Lactobacillus species, respectively. The tested strains of these groups produced L-lactic acid. The amount of cellular fatty acids of C16:0 and C18:1, and the DNA base compositions were significant for differentiating the strains in Groups II and III. Five strains of cocci in chains (Group IV) produced gas from glucose. The tested strains of this group produced d-lactic acid. They were identified as a Leuconostoc species. The distribution of these bacteria in fermented fish in Thailand is discussed.

Brain glutathione reductase induction increases early survival and decreases lipofuscin accumulation in aging frogs.

PubMed

López-Torres, M; Pérez-Campo, R; Fernandez, A; Barba, C; Barja de Quiroga, G

1993-02-01

Brain catalase was continuously depleted throughout the life span starting with a large population of initially young and old frogs. Free radical-related parameters were measured in the brain tissue once per year after 2.5, 14.5, and 26.5 months of experimentation. Brain lipofuscin accumulation was observed after 14.5 and 26.5 months, and survival was continuously followed during 33 months. The age of the animal did not decrease endogenous antioxidants nor increase tissue peroxidation either in cross-sectional or longitudinal comparisons. Continuous catalase depletion similarly affected young and old animals, inducing glutathione reductase, tending to decrease oxidized glutathione/reduced glutathione (GSSG/GSH) ratio, decreasing lipofuscin accumulation in the brain, and increasing survival from 46% to 91% after 14.5 months. At 26.5 months of experimentation the loss of the glutathione reductase induction in catalase-depleted animals was accompanied by the presence of higher lipofuscin deposits than in controls and was followed by a great increase in mortality rate. Even though the maximal life span (7 years) was the same in the control and treated animals which were already old (4.2 years) at the beginning of the experiment, the treated animals showed a strong reduction in the rates of early death. It is proposed that the maintenance of a high antioxidant/prooxidant balance in the vertebrate brain greatly increases the probability of the individual to reach the final segments of its species-specific life span.

Glutathione-mediated biodegradable polyurethanes derived from L-arabinitol.

PubMed

de Paz, M Violante; Zamora, Francisca; Begines, Belén; Ferris, Cristina; Galbis, Juan A

2010-01-11

The synthesis, characterization, and some properties of new glutathione-mediated biodegradable sugar-based copolyurethanes are described. These copolyurethanes were obtained by polyaddition reaction of mixtures of 2,2'-dithiodiethanol (DiT) and 2,3,4-tri-O-benzyl-L-arabinitol (ArBn) or 2,3,4-tri-O-methyl-L-arabinitol (ArMe) to 1,6-hexamethylene diisocyanate (HMDI). The copolymer compositions were studied by elemental microanalyses and (1)H NMR, revealing that the content of the copolymer units is in all cases very similar to that of their corresponding feed. The PU(DiT-HMDI) homopolymer exhibited a high crystallinity, but the introduction of the arabinitol-based diols led to a reduction in the crystallinity of the copolymers. In their TG curves, the copolymers exhibited a mixed trend of the related homopolymers, and all of them were thermally stable, with degradation temperatures above 220 degrees C. The degradation properties of the macromolecules under physiological conditions in the presence of glutathione were tested. All the copolyurethanes proved to be biodegradable under the experimental conditions (pH = 7.02 and 37 degrees C). The degradation pattern of the copolymers depended not only on the dithiodiethanol (DiT) reactive units ratio in the polymer backbone, but also on the crystallinity of the macromolecule.

Measuring protein-bound glutathioine (PSSG): Critical correction for cytosolic glutathione species

USDA-ARS?s Scientific Manuscript database

Introduction: Protein glutathionylation is gaining recognition as an important posttranslational protein modification. The common first step in measuring protein glutathionylation is the denaturation and precipitation of protein away from soluble, millimolar quantities of glutathione (GSH) and glut...

The Effect of Alcohol and Hydrogen Peroxide on Liver Hepcidin Gene Expression in Mice Lacking Antioxidant Enzymes, Glutathione Peroxidase-1 or Catalase

PubMed Central

Harrison-Findik, Duygu Dee; Lu, Sizhao

2015-01-01

This study investigates the regulation of hepcidin, the key iron-regulatory molecule, by alcohol and hydrogen peroxide (H2O2) in glutathione peroxidase-1 (gpx-1−/−) and catalase (catalase−/−) knockout mice. For alcohol studies, 10% ethanol was administered in the drinking water for 7 days. Gpx-1−/− displayed significantly higher hepatic H2O2 levels than catalase−/− compared to wild-type mice, as measured by 2'-7'-dichlorodihydrofluorescein diacetate (DCFH-DA). The basal level of liver hepcidin expression was attenuated in gpx-1−/− mice. Alcohol increased H2O2 production in catalase−/− and wild-type, but not gpx-1−/−, mice. Hepcidin expression was inhibited in alcohol-fed catalase−/− and wild-type mice. In contrast, alcohol elevated hepcidin expression in gpx-1−/− mice. Gpx-1−/− mice also displayed higher level of basal liver CHOP protein expression than catalase−/− mice. Alcohol induced CHOP and to a lesser extent GRP78/BiP expression, but not XBP1 splicing or binding of CREBH to hepcidin gene promoter, in gpx-1−/− mice. The up-regulation of hepatic ATF4 mRNA levels, which was observed in gpx-1−/− mice, was attenuated by alcohol. In conclusion, our findings strongly suggest that H2O2 inhibits hepcidin expression in vivo. Synergistic induction of CHOP by alcohol and H2O2, in the absence of gpx-1, stimulates liver hepcidin gene expression by ER stress independent of CREBH. PMID:25955433

Lung glutathione reductase induction in aging catalase-depleted frogs correlates with early survival throughout the life span.

PubMed

Perez-Campo, R; Lopez-Torres, M; Rojas, C; Cadenas, S; Barja de Quiroga, G

1993-02-01

A comprehensive experimental study on free radical-related parameters was performed in the lung throughout the life span of 220 initially young or old frogs. No age related differences were found transversely or longitudinally for lung superoxide dismutase, catalase, Se-dependent and -independent glutathione peroxidases, glutathione reductase, GSH, GSSG, or GSSG/GSH ratio. Continuous catalase depletion with aminotriazole led to glutathione reductase induction in the lung after 14.5 months of experimentation. This was accompanied by a great increase in survival rate of treated animals in relation to controls (especially in the old group). After 26.5 months of experimentation, glutathione reductase induction was lost and GSSG/GSH values tended to increase. This was followed by a 3-month long period of acute decrease in survival rate of treated animals. It is suggested that a high antioxidant/prooxidant balance is of protective value against causes of early death and can possibly be used in the future (when appropriately controlled) to increase the number of healthy years of the normal life span.

Insights into ligand binding to a Glutathione S-transferase from mango: structure, thermodynamics and kinetics

PubMed Central

Valenzuela-Chavira, Ignacio; Contreras-Vergara, Carmen A.; Arvizu-Flores, Aldo A.; Serrano-Posada, Hugo; Lopez-Zavala, Alonso A.; García-Orozco, Karina D.; Hernandez-Paredes, Javier; Rudiño-Piñera, Enrique; Stojanoff, Vivian; Sotelo-Mundo, Rogerio R.; Islas-Osuna, Maria A.

2017-01-01

We studied a mango glutathione S-transferase (GST) (Mangifera indica) bound to glutathione (GSH) and S-hexyl glutathione (GSX). This GST Tau class (MiGSTU) had a molecular mass of 25.5 kDa. MiGSTU Michaelis-Menten kinetic constants were determined for their substrates obtaining a Km, Vmax and kcat for CDNB of 0.792 mM, 80.58 mM·min−1 and 68.49 s−1 respectively and 0.693 mM, 105.32 mM·min−1 and 89.57 s−1, for reduced GSH respectively. MiGSTU had a micromolar affinity towards GSH (5.2 μM) or GSX (7.8 μM). The crystal structure of the MiGSTU in apo or bound to GSH or GSX generated a model that explains the thermodynamic signatures of binding and showed the importance of enthalpic-entropic compensation in ligand binding to Tau-class GST enzymes. PMID:28104507

S-Adenosylmethionine Attenuates Oxidative Stress and Neuroinflammation Induced by Amyloid-β Through Modulation of Glutathione Metabolism.

PubMed

Li, Qian; Cui, Jing; Fang, Chen; Liu, Min; Min, Guowen; Li, Liang

2017-01-01

Oxidative stress and neuroinflammation are mainly involved in the pathogenic mechanisms of Alzheimer's disease (AD). Amyloid-β (Aβ), the main component of senile plaques, is a kind of strong inducer of oxidative stress. Glutathione is an endogenous antioxidant protecting cells from oxidative injury. S-adenosylmethionine (SAM) produced in the methionine cycle is the primary methyl donor and the precursor of glutathione. In this study, the Aβ intrahippocampal injection rat model and cultured SH-SY5Y cells were used to explore the neuroprotective effect of SAM. We found that SAM could protect cells against Aβ-induced cellular injury by inhibition of oxidative stress and neuroinflammation. SAM administration could increase the endogenous antioxidant glutathione and potentiate the antioxidant enzymes activities. SAM might act as an antioxidant and be a potential candidate therapy for AD patients.

Immunoregulatory effects of glutathione during mesenchymal stem cell differentiation to hepatocyte-like cells.

PubMed

Ahmadi-Ashtiani, Hamid-Reza; Allameh, Abdolamir; Rastegar, Hossein; Mortaz, Esmaeil; Saraf, Zahir

2012-09-01

The role of mesenchymal stem cell in cellular therapy is the subject of interest for many researchers. The differentiation potential of MSCs and abilities in modulations of the recipient's immune system makes them important cells in tissue regenerative studies. MSCs by releasing the proinflammatory cytokines play important role in immunomodulatory systems; however the signaling pathways for releasing of these mediators are not well understood. Glutathione has been shown to play a role in modulation of cytokines in hepatogenic differentiation. In the current study we aimed to investigate the effects of buthionine sulfoximine (BSO, inhibitor for glutathione synthesis) and N-acetylecystin (NAC, an inhibitor for ROS generation) on proinflammatory cytokines production in a hepatogenic differentiation model. BSO and NAC significantly decreased IL-6 and TNF-α levels at 14 days of differentiation, whereas, NAC decreased the levels of IL-8 at days 2 and 14 of differentiation. Moreover, intracellular glutathione level during the differentiation was depleted. Our current study suggests a novel role of GSH as an immunopharmacological regulatory molecule during hepatogenic differentiation. Finally, this information may shed some light on the understanding of MSCs responses in transplantation and cell therapy in diseases such as chronic hepatic diseases.

Glutathione levels in and total antioxidant capacity of Candida sp. cells exposed to oxidative stress caused by hydrogen peroxide.

PubMed

Abegg, Maxwel Adriano; Alabarse, Paulo Vinícius Gil; Schüller, Artur Krumberg; Benfato, Mara Silveira

2012-10-01

The capacity to overcome the oxidative stress imposed by phagocytes seems to be critical for Candida species to cause invasive candidiasis. To better characterize the oxidative stress response (OSR) of 8 clinically relevant Candida sp., glutathione, a vital component of the intracellular redox balance, was measured using the 5,5'-dithiobis-(2-nitrobenzoic acid (DTNB)-glutathione disulfide (GSSG) reductase reconversion method; the total antioxidant capacity (TAC) was measured using a modified method based on the decolorization of the 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic) acid radical cation (ABTS*+). Both methods were used with cellular Candida sp. extracts treated or not with hydrogen peroxide (0.5 mM). Oxidative stress induced by hydrogen peroxide clearly reduced intracellular glutathione levels. This depletion was stronger in Candida albicans and the levels of glutathione in untreated cells were also higher in this species. The TAC demonstrated intra-specific variation. Glutathione levels did not correlate with the measured TAC values, despite this being the most important non-enzymatic intracellular antioxidant molecule. The results indicate that the isolated measurement of TAC does not give a clear picture of the ability of a given Candida sp. to respond to oxidative stress.

Detection of intracellular glutathione using ThiolTracker violet stain and fluorescence microscopy.

PubMed

Mandavilli, Bhaskar S; Janes, Michael S

2010-07-01

Glutathione plays an important role in protecting mammalian cells from oxidative stress and cell death. Because reduced glutathione (GSH) represents the large majority of intracellular free thiols, cell-permeant, thiol-reactive fluorescent probes represent potentially useful indicators of intracellular GSH. The ThiolTracker Violet stain (a registered trademark of Invitrogen) is a bright fluorescent probe that is highly reactive to thiols and can be used as a convenient and effective indicator of intracellular GSH and general redox status by a variety of detection modalities. While this probe has been validated in flow cytometry and microplate fluorimetry assays, the following method will describe details on the use of the ThiolTracker Violet dye in traditional fluorescence microscopy, as well as high-content imaging and analysis.

Cellular glutathione level does not predict ovarian cancer cells' resistance after initial or repeated exposure to cisplatin.

PubMed

Nikounezhad, Nastaran; Nakhjavani, Maryam; Shirazi, Farshad H

2017-05-01

Cisplatin resistance development is a major obstacle in ovarian cancer treatment. One of the most important mechanisms underlying cisplatin resistance is drug detoxification by glutathione. In the present study, the importance of initial or repeated exposure to cisplatin in glutathione dependent resistance was investigated. To this purpose, some cisplatin sensitive and resistant variants of human ovarian cancer cell lines providing an appropriate range of cisplatin sensitivity were selected. Clonogenic survival assay was performed to evaluate cisplatin resistance and intracellular contents of reduced (GSH) and oxidized (GSSG) glutathione were analyzed using an HPLC method. Our results indicated that the intracellular GSH and GSSG concentrations were nearly equal in A2780 and A2780CP cells, while the A2780CP cells showed 14 times more resistance than the A2780 cells after initial exposure to cisplatin. A2780-R1 and A2780-R3 cells which have been repeatedly exposed to cisplatin also showed no significant difference in glutathione content, even though A2780-R3 was about two times more resistant than A2780-R1. Moreover, intracellular GSH/GSSG ratio decreased in the resistant cells, reflecting a shift towards a more oxidizing intracellular environment indicative of oxidative stress. As a conclusion, it seems that although the intracellular glutathione concentration increases after repeated exposure to cisplatin, there is no clear correlation between the intracellular GSH content in ovarian cancer cells and their resistance to cisplatin neither after initial nor after repeated exposure to this drug.

Role of the Ascorbate-Glutathione Cycle of Mitochondria and Peroxisomes in the Senescence of Pea Leaves1

PubMed Central

Jiménez, Ana; Hernández, José A.; Pastori, Gabriela; del Río, Luis A.; Sevilla, Francisca

1998-01-01

We investigated the relationship between H2O2 metabolism and the senescence process using soluble fractions, mitochondria, and peroxisomes from senescent pea (Pisum sativum L.) leaves. After 11 d of senescence the activities of Mn-superoxide dismutase, dehydroascorbate reductase (DHAR), and glutathione reductase (GR) present in the matrix, and ascorbate peroxidase (APX) and monodehydroascorbate reductase (MDHAR) activities localized in the mitochondrial membrane, were all substantially decreased in mitochondria. The mitochondrial ascorbate and dehydroascorbate pools were reduced, whereas the oxidized glutathione levels were maintained. In senescent leaves the H2O2 content in isolated mitochondria and the NADH- and succinate-dependent production of superoxide (O2·−) radicals by submitochondrial particles increased significantly. However, in peroxisomes from senescent leaves both membrane-bound APX and MDHAR activities were reduced. In the matrix the DHAR activity was enhanced and the GR activity remained unchanged. As a result of senescence, the reduced and the oxidized glutathione pools were considerably increased in peroxisomes. A large increase in the glutathione pool and DHAR activity were also found in soluble fractions of senescent pea leaves, together with a decrease in GR, APX, and MDHAR activities. The differential response to senescence of the mitochondrial and peroxisomal ascorbate-glutathione cycle suggests that mitochondria could be affected by oxidative damage earlier than peroxisomes, which may participate in the cellular oxidative mechanism of leaf senescence longer than mitochondria. PMID:9847106

[Effects of melaxen and valdoxan on the activity of glutathione antioxidant system and NADPH-producing enzymes in rat heart under experimental hyperthyroidism conditions].

PubMed

Gorbenko, M V; Popova, T N; Shul'gin, K K; Popov, S S

2013-01-01

The effects of melaxen and valdoxan on the activity of glutathione antioxidant system and some NADPH-producing enzymes have been studied under conditions of experimental hyperthyroidism in rat heart. Under the action of these drugs, reduced glutathione (GSH) content increased as compared to values observed under the conditions of pathology. It has been established that the activities of glutathione reductase (GR), glutathione peroxidase (GP), glucose-6-phosphate dehydrogenase, and NADP isocitrate dehydrogenase (increased under pathological conditions) change toward the intact control values upon the introduction of both drugs. The influence of melaxen and valdoxan, capable of producing antioxidant effect, leads apparently to the inhibition of free-radical oxidation processes and, as a consequence, the reduction of mobilization degree of the glutathione antioxidant system.

Bacteria isolated from amoebae/bacteria consortium

DOEpatents

Tyndall, R.L.

1995-05-30

New protozoan derived microbial consortia and method for their isolation are provided. Consortia and bacteria isolated therefrom are useful for treating wastes such as trichloroethylene and trinitrotoluene. Consortia, bacteria isolated therefrom, and dispersants isolated therefrom are useful for dispersing hydrocarbons such as oil, creosote, wax, and grease.

Bacteria isolated from amoebae/bacteria consortium

DOEpatents

Tyndall, Richard L.

1995-01-01

New protozoan derived microbial consortia and method for their isolation are provided. Consortia and bacteria isolated therefrom are useful for treating wastes such as trichloroethylene and trinitrotoluene. Consortia, bacteria isolated therefrom, and dispersants isolated therefrom are useful for dispersing hydrocarbons such as oil, creosote, wax, and grease.

Depletion of pulmonary glutathione using diethylmaleic acid accelerates the development of oxygen-induced lung injury in term and preterm guinea-pig neonates.

PubMed

Langley, S C; Kelly, F J

1994-02-01

Dietary or chemical depletion of pulmonary glutathione in adult rats and mice, has been demonstrated to exacerbate the toxic effects of high oxygen concentrations. The present paper has examined this phenomenon in a guinea-pig model of prematurity, using the electrophilic agent diethylmaleic acid (DEM) to provide a transient (up to 12 h) pulmonary glutathione depletion. Full-term and 3-days preterm guinea-pig pups were studied to assess the possible role for glutathione deficiency as a mechanism mediating the increased susceptibility of the immature lung to oxygen free-radical damage. The administration of DEM to guinea-pig neonates depleted lung glutathione by 90% (term) or 68% (preterm) over 2 h. On exposure of pups to 95% oxygen for 48 h, DEM increased the incidence of oxygen-related death to 31% in term pups and 100% in preterm pups. Term pups exposed to hyperoxia and treated with DEM showed evidence of pulmonary injury, indicated by an influx of neutrophils into the lung airspaces, and elevated microvascular permeability. Control pups exposed to 95% oxygen were found to have uninjured lungs after 48 h. We conclude that glutathione is an essential component of the pulmonary antioxidant array in neonates. Glutathione may be of particular importance in the early phase of oxygen exposure. The deficiency of lung glutathione observed in preterm animals may account for their increased susceptibility to oxygen-induced pulmonary injury.

Ethacrynic acid and a derivative enhance apoptosis in arsenic trioxide-treated myeloid leukemia and lymphoma cells: the role of glutathione S-transferase P1-1

PubMed Central

Wang, Rui; Liu, Changda; Xia, Lijuan; Zhao, Guisen; Gabrilove, Janice; Waxman, Samuel; Jing, Yongkui

2012-01-01

Purpose Arsenic trioxide (ATO) as a single agent is used for treatment of acute promyelocytic leukemia (APL) with minimal toxicity but therapeutic effect of ATO in other types of malignancies has not been achieved. We tested whether a combination with ethacrynic acid (EA), a glutathione S-transferase P1-1 (GSTP1-1) inhibitor and a reactive oxygen species (ROS) inducer will extend the therapeutic effect of ATO beyond APL. Experimental Design The combined apoptotic effects of ATO plus EA were tested in non-APL leukemia and lymphoma cell lines. The role of ROS, GSTP1-1, glutathione, and Mcl-1 in apoptosis was determined. The selective response to this combination of cells with and without GSTP1-1 expression was compared. Results ATO/EA combination synergistically induced apoptosis in myeloid leukemia and lymphoma cells. This treatment produced high ROS levels, activated c-jun-NH2-terminal kinase and reduced Mcl-1 protein. This led to the decrease of mitochondrial transmembrane potential, release of cytochrome c and, subsequently, to activation of caspase 3 and 9. Induction of apoptosis in leukemia and lymphoma cells expressing GSTP1-1 required that high EA concentrations be combined with ATO. Silencing of GSTP1 in leukemia cells sensitized them to ATO/EA-induced apoptosis. In a sub-group of B-cell lymphoma which do not express GSTP1-1, lower concentrations of EA and its more potent derivative, ethacrynic acid butyl-ester, decreased intracellular glutathione levels and synergistically induced apoptosis when combined with ATO. Conclusion B-cell lymphoma cells lacking GSTP1-1 are more sensitive than myeloid leukemia cells to ATO/EA-induced apoptosis. PMID:23082001

Insights into ligand binding to a glutathione S-transferase from mango: Structure, thermodynamics and kinetics

DOE PAGES

Valenzuela-Chavira, Ignacio; Contreras-Vergara, Carmen A.; Arvizu-Flores, Aldo A.; ...

2017-01-17

We studied a mango glutathione S-transferase (GST) ( Mangifera indica) bound to glutathione (GSH) and S-hexyl glutathione (GSX). This GST Tau class (MiGSTU) had a molecular mass of 25.5 kDa. MiGSTU Michaelis-Menten kinetic constants were determined for their substrates obtaining a K m, V max and k cat for CDNB of 0.792 mM, 80.58 mM min -1 and 68.49 s -1 respectively and 0.693 mM, 105.32 mM min -1 and 89.57 s -1, for reduced GSH respectively. MiGSTU had a micromolar affinity towards GSH (5.2 mM) or GSX (7.8 mM). As a result, the crystal structure of the MiGSTU inmore » apo or bound to GSH or GSX generated a model that explains the thermodynamic signatures of binding and showed the importance of enthalpic-entropic compensation in ligand binding to Tau-class GST enzymes.« less

Insights into ligand binding to a glutathione S-transferase from mango: Structure, thermodynamics and kinetics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Valenzuela-Chavira, Ignacio; Contreras-Vergara, Carmen A.; Arvizu-Flores, Aldo A.

We studied a mango glutathione S-transferase (GST) ( Mangifera indica) bound to glutathione (GSH) and S-hexyl glutathione (GSX). This GST Tau class (MiGSTU) had a molecular mass of 25.5 kDa. MiGSTU Michaelis-Menten kinetic constants were determined for their substrates obtaining a K m, V max and k cat for CDNB of 0.792 mM, 80.58 mM min -1 and 68.49 s -1 respectively and 0.693 mM, 105.32 mM min -1 and 89.57 s -1, for reduced GSH respectively. MiGSTU had a micromolar affinity towards GSH (5.2 mM) or GSX (7.8 mM). As a result, the crystal structure of the MiGSTU inmore » apo or bound to GSH or GSX generated a model that explains the thermodynamic signatures of binding and showed the importance of enthalpic-entropic compensation in ligand binding to Tau-class GST enzymes.« less

Glutathione adducts on sarcoplasmic/endoplasmic reticulum Ca2+ ATPase Cys-674 regulate endothelial cell calcium stores and angiogenic function as well as promote ischemic blood flow recovery.

PubMed

Thompson, Melissa D; Mei, Yu; Weisbrod, Robert M; Silver, Marcy; Shukla, Praphulla C; Bolotina, Victoria M; Cohen, Richard A; Tong, Xiaoyong

2014-07-18

The sarco/endoplasmic reticulum Ca(2+) ATPase (SERCA) is key to Ca(2+) homeostasis and is redox-regulated by reversible glutathione (GSH) adducts on the cysteine (C) 674 thiol that stimulate Ca(2+) uptake activity and endothelial cell angiogenic responses in vitro. We found that mouse hind limb muscle ischemia induced S-glutathione adducts on SERCA in both whole muscle tissue and endothelial cells. To determine the role of S-glutathiolation, we used a SERCA 2 C674S heterozygote knock-in (SKI) mouse lacking half the key thiol. Following hind limb ischemia, SKI animals had decreased SERCA S-glutathione adducts and impaired blood flow recovery. We studied SKI microvascular endothelial cells in which total SERCA 2 expression was unchanged. Cultured SKI microvascular endothelial cells showed impaired migration and network formation compared with wild type (WT). Ca(2+) studies showed decreased nitric oxide (·NO)-induced (45)Ca(2+) uptake into the endoplasmic reticulum (ER) of SKI cells, while Fura-2 studies revealed lower Ca(2+) stores and decreased vascular endothelial growth factor (VEGF)- and ·NO-induced Ca(2+) influx. Adenoviral overexpression of calreticulin, an ER Ca(2+) binding protein, increased ionomycin-releasable stores, VEGF-induced Ca(2+) influx and endothelial cell migration. Taken together, these data indicate that the redox-sensitive Cys-674 thiol on SERCA 2 is required for normal endothelial cell Ca(2+) homeostasis and ischemia-induced angiogenic responses, revealing a novel redox control of angiogenesis via Ca(2+) stores. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Bee Venom (Apis Mellifera) an Effective Potential Alternative to Gentamicin for Specific Bacteria Strains: Bee Venom an Effective Potential for Bacteria.

PubMed

Zolfagharian, Hossein; Mohajeri, Mohammad; Babaie, Mahdi

2016-09-01

Mellitine, a major component of bee venom (BV, Apis mellifera ), is more active against gram positive than gram negative bacteria. Moreover, BV has been reported to have multiple effects, including antibacterial, antivirus, and anti-inflammation effects, in various types of cells. In addition, wasp venom has been reported to have antibacterial properties. The aim of this study was to evaluate the antibacterial activity of BV against selected gram positive and gram negative bacterial strains of medical importance. This investigation was set up to evaluate the antibacterial activity of BV against six grams positive and gram negative bacteria, including Staphylococcus aureus ( S. aureus ), Salmonella typhimurium , Escherichia coli ( E. coli ) O157:H7, Pseudomonas aeruginosa , Burkholderia mallei and Burkholderia pseudomallei. Three concentrations of crude BV and standard antibiotic (gentamicin) disks as positive controls were tested by using the disc diffusion method. BV was found to have a significant antibacterial effect against E. coli , S. aureus , and Salmonella typhyimurium in all three concentrations tested. However, BV had no noticeable effect on other tested bacteria for any of the three doses tested. The results of the current study indicate that BV inhibits the growth and survival of bacterial strains and that BV can be used as a complementary antimicrobial agent against pathogenic bacteria. BV lacked the effective proteins necessary for it to exhibit antibacterial activity for some specific strains while being very effective against other specific strains. Thus, one may conclude, that Apis mellifera venom may have a specific mechanism that allows it to have an antibacterial effect on certain susceptible bacteria, but that mechanism is not well understood.

Lack of antimicrobial effect on periodontopathic bacteria by ultrasonic and sonic scalers in vitro.

PubMed

Schenk, G; Flemmig, T F; Lob, S; Ruckdeschel, G; Hickel, R

2000-02-01

The purpose of this study was to assess the antimicrobial effects of a sonic and ultrasonic scaler generally used for subgingival scaling on gram-negative and gram-positive periodontopathic bacteria. Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Campylobacter rectus, or Peptostreptococcus micros were suspended in Schaedler's broth medium and treated by a sonic or a magnetostrictive ultrasonic scaler for 30 s and 150 s in vitro. Bacterial suspensions treated by an ultrasonic cell disruptor served as a positive control and untreated bacterial suspensions served as a negative control. Following sonication, samples were serially diluted, streaked on blood agar plates and incubated for 2-5 days at 37 degrees C. Treatment by the sonic or ultrasonic scaler for up to 150 s did not reduce the viability of any of the tested periodontal pathogens. Compared to untreated controls, the viability of A. actinomycetemcomitans and P. gingivalis was significantly (p<0.05) reduced only following ultrasonication with the cell disruptor after 30 s (0.72 and 0.54 log CFU/ml, respectively) and of A. actinomycetemcomitans, P. gingivalis, C. rectus, and P. micros after 150 s (1.98, 1.34, 1.95 and 1.98 log CFU/ml, respectively). The data of the study may indicate that the assessed sonic and ultrasonic scaler used for subgingival debridement do not result in killing of the tested periodontal pathogens.

Glutathione depletion activates the yeast vacuolar transient receptor potential channel, Yvc1p, by reversible glutathionylation of specific cysteines

PubMed Central

Chandel, Avinash; Das, Krishna K.; Bachhawat, Anand K.

2016-01-01

Glutathione depletion and calcium influx into the cytoplasm are two hallmarks of apoptosis. We have been investigating how glutathione depletion leads to apoptosis in yeast. We show here that glutathione depletion in yeast leads to the activation of two cytoplasmically inward-facing channels: the plasma membrane, Cch1p, and the vacuolar calcium channel, Yvc1p. Deletion of these channels partially rescues cells from glutathione depletion–induced cell death. Subsequent investigations on the Yvc1p channel, a homologue of the mammalian TRP channels, revealed that the channel is activated by glutathionylation. Yvc1p has nine cysteine residues, of which eight are located in the cytoplasmic regions and one on the transmembrane domain. We show that three of these cysteines, Cys-17, Cys-79, and Cys-191, are specifically glutathionylated. Mutation of these cysteines to alanine leads to a loss in glutathionylation and a concomitant loss in calcium channel activity. We further investigated the mechanism of glutathionylation and demonstrate a role for the yeast glutathione S-transferase Gtt1p in glutathionylation. Yvc1p is also deglutathionylated, and this was found to be mediated by the yeast thioredoxin, Trx2p. A model for redox activation and deactivation of the yeast Yvc1p channel is presented. PMID:27708136

A role for CFTR in the elevation of glutathione levels in the lung by oral glutathione administration

PubMed Central

Kariya, Chirag; Leitner, Heather; Min, Elysia; van Heeckeren, Christiaan; van Heeckeren, Anna; Day, Brian J.

2014-01-01

The cystic fibrosis transmembrane conductance regulator (CFTR) protein is the only known apical glutathione (GSH) transporter in the lung. The purpose of these studies was to determine whether oral GSH or glutathione disulfide (GSSG) treatment could increase lung epithelial lining fluid (ELF) GSH levels and whether CFTR plays a role in this process. The pharmacokinetic profile of an oral bolus dose of GSH (300 mg/kg) was determined in mice. Plasma, ELF, bronchoalveolar lavage (BAL) cells, and lung tissue were analyzed for GSH content. There was a rapid elevation in the GSH levels that peaked at 30 min in the plasma and 60 min in the lung, ELF, and BAL cells after oral GSH dosing. Oral GSH treatment produced a selective increase in the reduced and active form of GSH in all lung compartments examined. Oral GSSG treatment (300 mg/kg) resulted in a smaller increase of GSH levels. To evaluate the role of CFTR in this process, Cftr knockout (KO) mice and gut-corrected Cftr KO-transgenic (Tg) mice were given an oral bolus dose of GSH (300 mg/kg) and compared with wild-type mice for changes in GSH levels in plasma, lung, ELF, and BAL cells. There was a twofold increase in plasma, a twofold increase in lung, a fivefold increase in ELF, and a threefold increase in BAL cell GSH levels at 60 min in wild-type mice; however, GSH levels only increased by 40% in the plasma, 60% in the lung, 50% in the ELF, and twofold in the BAL cells within the gut-corrected Cftr KO-Tg mice. No change in GSH levels was observed in the uncorrected Cftr KO mice. These studies suggest that CFTR plays an important role in GSH uptake from the diet and transport processes in the lung. PMID:17369290

Choroid plexus glutathione peroxidases are instrumental in protecting the brain fluid environment from hydroperoxides during postnatal development.

PubMed

Saudrais, Elodie; Strazielle, Nathalie; Ghersi-Egea, Jean-Francois

2018-06-27

Hydrogen peroxide, released at low physiological concentration, is involved in different cell signaling pathways during brain development. When released at supraphysiological concentrations in brain fluids following an inflammatory, hypoxic or toxic stress, it can initiate lipid peroxidation, protein and nucleic acid damage and contribute to long-term neurological impairment associated with perinatal diseases. We found high glutathione peroxidase and glutathione reductase enzymatic activities in both lateral and fourth ventricle choroid plexus tissue isolated from developing rats, in comparison to the cerebral cortex and liver. Consistent with these, a high protein expression of glutathione peroxidases 1 and 4 was observed in choroid plexus epithelial cells, which form the blood-cerebrospinal fluid barrier. Live choroid plexuses isolated from newborn rats were highly efficient in detoxifying H2O2 from mock cerebrospinal fluid, illustrating the capacity of the choroid plexuses to control H2O2 concentration in the ventricular system of the brain. We used a differentiated cellular model of the blood-cerebrospinal fluid barrier coupled to kinetic and inhibition analyses to show that glutathione peroxidases are more potent than catalase to detoxify extracellular H2O2 at concentrations up to 250 µM. The choroidal cells also formed an enzymatic barrier preventing blood-borne hydroperoxides to reach the cerebrospinal fluid. These data point out the choroid plexuses as key structures in the control of hydroperoxide levels in the cerebral fluid environment during development, at a time when the protective glial cell network is still immature. Glutathione peroxidases are the main effectors of this choroidal hydroperoxide inactivation.

Beta-carotene reduces oxidative stress, improves glutathione metabolism and modifies antioxidant defense systems in lead-exposed workers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kasperczyk, Sławomir, E-mail: kaslav@mp.pl; Dobrakowski, Michał; Kasperczyk, Janusz

2014-10-01

The aim of this study was to determine whether beta-carotene administration reduces oxidative stress and influences antioxidant, mainly glutathione-related, defense systems in workers chronically exposed to lead. The population consisted of two randomly divided groups of healthy male volunteers exposed to lead. Workers in the first group (reference group) were not administered any antioxidants, while workers in the second group (CAR group) were treated orally with 10 mg of beta-carotene once a day for 12 weeks. Biochemical analysis included measuring markers of lead-exposure and oxidative stress in addition to the levels and activities of selected antioxidants. After treatment, levels ofmore » malondialdehyde, lipid hydroperoxides and lipofuscin significantly decreased compared with the reference group. However, the level of glutathione significantly increased compared with the baseline. Treatment with beta-carotene also resulted in significantly decreased glutathione peroxidase activity compared with the reference group, while the activities of other glutathione-related enzymes and of superoxide dismutase were not significantly changed. However, the activities of glucose-6-phosphate dehydrogenase and catalase, as well as the level of alpha-tocopherol, were significantly higher after treatment compared with the baseline. Despite controversy over the antioxidant properties of beta-carotene in vivo, our findings showed reduced oxidative stress after beta-carotene supplementation in chronic lead poisoning. - Highlights: • Beta-carotene reduces oxidative stress in lead-exposed workers. • Beta-carotene elevates glutathione level in lead-exposed workers. • Beta-carotene administration could be beneficial in lead poisoning.« less

The Ascorbate-glutathione-α-tocopherol Triad in Abiotic Stress Response

PubMed Central

Szarka, András; Tomasskovics, Bálint; Bánhegyi, Gábor

2012-01-01

The life of any living organism can be defined as a hurdle due to different kind of stresses. As with all living organisms, plants are exposed to various abiotic stresses, such as drought, salinity, extreme temperatures and chemical toxicity. These primary stresses are often interconnected, and lead to the overproduction of reactive oxygen species (ROS) in plants, which are highly reactive and toxic and cause damage to proteins, lipids, carbohydrates and DNA, which ultimately results in oxidative stress. Stress-induced ROS accumulation is counteracted by enzymatic antioxidant systems and non-enzymatic low molecular weight metabolites, such as ascorbate, glutathione and α-tocopherol. The above mentioned low molecular weight antioxidants are also capable of chelating metal ions, reducing thus their catalytic activity to form ROS and also scavenge them. Hence, in plant cells, this triad of low molecular weight antioxidants (ascorbate, glutathione and α-tocopherol) form an important part of abiotic stress response. In this work we are presenting a review of abiotic stress responses connected to these antioxidants. PMID:22605990

[Association between glutathione peroxidase levels and clinical manifestations of dengue].

PubMed

Rojas, Elsa Marina; Díaz-Quijano, Fredi Alexander; Coronel-Ruiz, Carolina; Martínez-Vega, Ruth Aralf; Rueda, Ernesto; Villar-Centeno, Luis Angel

2007-06-01

Glutathione peroxidase (GP) can be used as a marker of oxidative stress in infectious diseases. To evaluate the association between the levels of glutathione peroxidase (GP) and the manifestations and complications of dengue. Between April 2003 and December 2004, 161 patients with dengue were prospectively evaluated. In the first evaluation, within 48 and 96 hours of disease onset, a plasma sample was obtained to measure the GP levels. The association between GP levels, clinical manifestations and complications was evaluated during the follow up. Mean GP values were 1198 U/L (95% confidence interval 1089-1306). Values greater than 1200 U/L were associated with headache, arthralgias and increased heart rate. There was a negative association between GP levels and serum triglycerides. During follow up, patients with GP >1200 U/L had a higher frequency of spontaneous hemorrhages. In a logistic regression analysis arthralgias, fever and increased heart rate, were independently associated with levels >1200 U/L. GP levels was associated to some of the manifestations of dengue. This finding suggests that the intensity of oxidative stress can influence the clinical presentation of dengue.

Dynamic compartment specific changes in glutathione and ascorbate levels in Arabidopsis plants exposed to different light intensities

PubMed Central

2013-01-01

Background Excess light conditions induce the generation of reactive oxygen species (ROS) directly in the chloroplasts but also cause an accumulation and production of ROS in peroxisomes, cytosol and vacuoles. Antioxidants such as ascorbate and glutathione occur in all cell compartments where they detoxify ROS. In this study compartment specific changes in antioxidant levels and related enzymes were monitored among Arabidopsis wildtype plants and ascorbate and glutathione deficient mutants (vtc2-1 and pad2-1, respectively) exposed to different light intensities (50, 150 which was considered as control condition, 300, 700 and 1,500 μmol m-2 s-1) for 4 h and 14 d. Results The results revealed that wildtype plants reacted to short term exposure to excess light conditions with the accumulation of ascorbate and glutathione in chloroplasts, peroxisomes and the cytosol and an increased activity of catalase in the leaves. Long term exposure led to an accumulation of ascorbate and glutathione mainly in chloroplasts. In wildtype plants an accumulation of ascorbate and hydrogen peroxide (H2O2) could be observed in vacuoles when exposed to high light conditions. The pad2-1 mutant reacted to long term excess light exposure with an accumulation of ascorbate in peroxisomes whereas the vtc2-1 mutant reacted with an accumulation of glutathione in the chloroplasts (relative to the wildtype) and nuclei during long term high light conditions indicating an important role of these antioxidants in these cell compartments for the protection of the mutants against high light stress. Conclusion The results obtained in this study demonstrate that the accumulation of ascorbate and glutathione in chloroplasts, peroxisomes and the cytosol is an important reaction of plants to short term high light stress. The accumulation of ascorbate and H2O2 along the tonoplast and in vacuoles during these conditions indicates an important route for H2O2 detoxification under these conditions. PMID

Lymphocyte DNA damage and plasma antioxidant status in Korean subclinical hypertensive patients by glutathione S-transferase polymorphism

PubMed Central

Han, Jeong-Hwa; Lee, Hye-Jin; Choi, Hee Jeong; Yun, Kyung Eun

2017-01-01

BACKGROUND/OBJECTIVES Glutathione S-transferase (GST) forms a multigene family of phase II detoxification enzymes which are involved in the detoxification of xenobiotics by conjugating substances with glutathione. The aim of this study is to assess the antioxidative status and the degree of DNA damage in the subclinical hypertensive patients in Korea using glutathione S-transferase polymorphisms. SUBJECTS/METHODS We examined whether DNA damage and antioxidative status show a difference between GSTM1 or GSTT1 genotype in 227 newly diagnosed, untreated (systolic blood pressure (BP) ≥ 130 mmHg or diastolic BP ≥ 85 mmHg) subclinical hypertensive patients and 130 normotensive subjects (systolic BP < 120 mmHg and diastolic BP < 80 mmHg). From the blood of the subjects, the degree of the DNA damage in lymphocyte, the activities of erythrocyte superoxide dismutase, the catalase, and the glutathione peroxidase, the level of glutathione, plasma total radical-trapping antioxidant potential (TRAP), anti-oxidative vitamins, as well as plasma lipid profiles and conjugated diene (CD) were analyzed. RESULTS Of the 227 subjects studied, 68.3% were GSTM1 null genotype and 66.5% were GSTT1 null genotype. GSTM1 null genotype had an increased risk of hypertension (OR: 2.104, CI: 1.38-3.35), but no significant association in GSTT1 null genotype (OR 0.982, CI: 0.62-1.55). No difference in erythrocyte activities of superoxide dismutase, catalase, or glutathione peroxidase, and plasma TRAP, CD, lipid profiles, and GSH levels were observed between GSTM1 or GSTT1 genotype. Plasma levels of α-tocopherol increased significantly in GSTT1 wild genotype (P < 0.05); however, plasma level of β-carotene increased significantly in GSTT1 null genotype (P < 0.01). DNA damage assessed by the Comet assay was significantly higher in GSTM1 null genotype than wild genotype (P < 0.05). CONCLUSIONS These results confirm the association between GSTM1 null genotype and risk of hypertension as they suggest

Recycling antibiotics into GUMBOS: A new combination strategy to combat multi-drug resistant bacteria

USDA-ARS?s Scientific Manuscript database

The emergence of multi-drug resistant bacteria, coupled with the lack of new antibiotics in development, is fast evolving into a global crisis. New strategies utilizing existing antibacterial agents are urgently needed. We propose one such strategy in which four outmoded ß-lactam antibiotics (amp...

[GLUTATHIONE SYSTEM ACTIVITY IN RAT TISSUES UNDER PHENYLETHYL BIGUANIDE ACTION ON THE BACKGROUND OF EXPERIMENTAL BRAIN ISCHEMIA/REPERFUSION DEVELOPMENT].

PubMed

Safonova, O A; Popova, T N; Kryl'skii, D V

2016-01-01

It was studied the total antioxidant activity, content of primary lipid peroxidation (LPO) products and reduced glutathione, and the activity of glutathione peroxidase, glutathione reductase, glucose-6-phosphate dehydrogenase, and NADP-isocitrate dehydrogenase in rat tissues under phenylethyl biguanide (phenfor- min) action on the background of experimental brain ischemia/reperfusion development. It is stablished the analyzed parameters, increasing under ischemia/reperfusion conditions in the brain and blood serum of animals, exhibit a decrease upon the introduction of this biguanide derivative. The obtained data can be explained by a decrease in degree of mobilization of the antioxidant system--in particular, of its glutathione chain--in the pathologic state. Hence, there is a need in NADPH supply for the system functioning compared with the pathology. Thus, phenylethyl biguanide demonstrates its antioxidant and protective properties under oxidative stress development that is accompanied by accumulation of the products of free radical oxidation of biomolecules during the ischemic brain injury.

Glutathione peroxidase contributes with heme oxygenase-1 to redox balance in mouse brain during the course of cerebral malaria.

PubMed

Linares, María; Marín-García, Patricia; Martínez-Chacón, Gabriela; Pérez-Benavente, Susana; Puyet, Antonio; Diez, Amalia; Bautista, José M

2013-12-01

Oxidative stress has been attributed both a key pathogenic and rescuing role in cerebral malaria (CM). In a Plasmodium berghei ANKA murine model of CM, host redox signaling and functioning were examined during the course of neurological damage. Host antioxidant defenses were early altered at the transcriptional level indicated by the gradually diminished expression of superoxide dismutase-1 (sod-1), sod-2, sod-3 and catalase genes. During severe disease, this led to the dysfunctional activity of superoxide dismutase and catalase enzymes in damaged brain regions. Vitagene associated markers (heat shock protein 70 and thioredoxin-1) also showed a decaying expression pattern that paralleled reduced expression of the transcription factors Parkinson disease 7, Forkhead box O 3 and X-box binding protein 1 with a role in preserving brain redox status. However, the oxidative stress markers reactive oxygen/nitrogen species were not accumulated in the brains of CM mice and redox proteomics and immunohistochemistry failed to detect quantitative or qualitative differences in protein carbonylation. Thus, the loss of antioxidant capacity was compensated for in all cerebral regions by progressive upregulation of heme oxygenase-1, and in specific regions by early glutathione peroxidase-1 induction. This study shows for the first time a scenario of cooperative glutathione peroxidase and heme oxygenase-1 upregulation to suppress superoxide dismutase, catalase, heat shock protein-70 and thioredoxin-1 downregulation effects in experimental CM, counteracting oxidative damage and maintaining redox equilibrium. Our findings reconcile the apparent inconsistency between the lack of oxidative metabolite build up and reported protective effect of antioxidant therapy against CM. © 2013.

The effect of hemodialysis and dialyzer biocompatibility on erythrocyte glutathione-defense system in chronic hemodialysis patients.

PubMed

Alhamdani, M S; Al-Najjar, A F; Al-Kassir, A H

2005-06-01

Uremic patients, especially those receiving regular hemodialysis (HD) treatment, are at high risk of oxidative damage by noxious free radicals and reactive oxygen species (ROS). The erythrocyte glutathione-defense system (GSH-DS) is one of the major enzymatic means of scavenging and detoxifying ROS. This study aimed to elucidate the effect of HD and dialyzer biocompatibility on erythrocyte GSH-DS in uremic patients on maintenance HD treatment. Twenty-five healthy volunteers and 42 HD patients were enrolled in this study. Blood samples were drawn immediately before and after HD session, and erythrocyte glutathione (GSH) level as well as the activities of the enzymes glucose-6-phosphate dehydrogenase (G6PD), glutathione peroxidase (GSH-Px), glutathione reductase (GSSG-Rd), and glutathione S-transferase (GST) were measured. To evaluate the effect of dialyzer type on the studied parameters the patients were were subdivided into two groups: those who had dialysis with cuprophane (CU) membranes (n=23) and those who received dialysis with the aid of polysulfone (PS) membranes (n=19). The activities of G6PD and GSH-Px as well as GSH level were significantly decreased in HD patients as compared with controls. On the other hand, the activities of GSSG-Rd and GST were significantly elevated among HD patients in comparison with control values. A single HD session, regardless of the type of dialyzer, did not induce any significant effect on any of the measured parameters, although G6PD activity increased significantly after dialysis. CU membrane did not result in any change in GSH or its metabolizing enzymes, while PS dialyzers exerted a minor but significant restoration in GSH-DS. The antioxidant pool, as represented by GSH-DS, is significantly affected by dialyzer type in HD patients being significantly corrected with polysulfone dialyzer.

Effect of associated bacteria on the growth and toxicity of Alexandrium catenella.

PubMed

Uribe, Paulina; Espejo, Romilio T

2003-01-01

Saprophytic bacteria in cultures of the marine dinoflagellate Alexandrium catenella were removed to assess their effect on growth and paralytic shellfish poisoning toxin production of this dinoflagellate. The actual axenic status was demonstrated by the lack of observable bacteria both immediately after treatment and following extended incubation in the absence of antibiotics. Bacteria were measured by counting CFU and also by epifluorescence microscopy and PCR amplification of bacterial 16S-23S spacer ribosomal DNA to detect noncultivable bacteria. Removal of bacteria did not have any effect on the growth of the dinoflagellate except for the inhibition of A. catenella disintegration after reaching the stationary phase. Toxicity was determined in dinoflagellate cell extracts by different methods: high-performance liquid chromatography (HPLC); an electrophysiological test called the Electrotest, which measures the inhibition of saxitoxin-sensitive Na(+) channels expressed in a cell line; and a mouse bioassay, which measures the toxic effect on the whole mammal neuromuscular system. A lower toxicity of the dinoflagellates in axenic culture was observed by these three methods, though the difference was significant only by the mouse bioassay and HPLC methods. Altogether the results indicate that axenic cultures of A. catenella are able to produce toxin, though the total toxicity is probably diminished to about one-fifth of that in nonaxenic cultures.

Glutathione and thioredoxin systems contribute to recombinant monoclonal antibody interchain disulfide bond reduction during bioprocessing.

PubMed

Handlogten, Michael W; Zhu, Min; Ahuja, Sanjeev

2017-07-01

Antibody interchain disulfide bond reduction during biopharmaceutical manufacturing has received increased attention since it was first reported in 2010. Antibody reduction leads to loss of product and reduced product stability. It is therefore critical to understand the underlying mechanisms of reduction. To date, the thioredoxin system has been reported as the sole contributor to antibody reduction during bioprocessing. In this work, we show that the glutathione system, in addition to the thioredoxin system, is involved in reducing antibody molecules and the contributions of the two systems can vary depending upon the cell culture process. The roles of the glutathione and thioredoxin systems were evaluated for three molecules with different IgG subclass where reduction was observed during manufacturing: mAb A, mAb B, and mAb C representing an IgG 1 , IgG 2 , and IgG 4, respectively. The expression of enzymes for both the thioredoxin and glutathione systems were confirmed in all three cell lines. Inhibitors were evaluated using purified mammalian reductases to evaluate their specificity. The optimized experimental conditions enabled both the determination of reductase activity contributed from as well as the amount of antibody reduced by each enzymatic system. Our results demonstrate that the underlying enzymatic mechanisms are different depending upon the cell culture process; one of the two systems may be the dominant mechanism, or both enzymatic systems may be involved. Specifically, the glutathione system was found to be the major contributor to mAb A reduction while the thioredoxin system was the major contributor to mAb C reduction. Intriguingly, mAb B experienced significant reduction from both enzymatic systems. In summary, we have demonstrated that in addition to the thioredoxin pathway, the glutathione system is a second major pathway contributing to antibody reduction and this knowledge can be leveraged to develop more specific antibody reduction

Functional complementation of anthocyanin sequestration in the vacuole by widely divergent glutathione S-transferases.

PubMed Central

Alfenito, M R; Souer, E; Goodman, C D; Buell, R; Mol, J; Koes, R; Walbot, V

1998-01-01

Glutathione S-transferases (GSTs) traditionally have been studied in plants and other organisms for their ability to detoxify chemically diverse herbicides and other toxic organic compounds. Anthocyanins are among the few endogenous substrates of plant GSTs that have been identified. The Bronze2 (Bz2) gene encodes a type III GST and performs the last genetically defined step of the maize anthocyanin pigment pathway. This step is the conjugation of glutathione to cyanidin 3-glucoside (C3G). Glutathionated C3G is transported to the vacuole via a tonoplast Mg-ATP-requiring glutathione pump (GS-X pump). Genetically, the comparable step in the petunia anthocyanin pathway is controlled by the Anthocyanin9 (An9) gene. An9 was cloned by transposon tagging and found to encode a type I plant GST. Bz2 and An9 have evolved independently from distinct types of GSTs, but each is regulated by the conserved transcriptional activators of the anthocyanin pathway. Here, a phylogenetic analysis is presented, with special consideration given to the origin of these genes and their relaxed substrate requirements. In particle bombardment tests, An9 and Bz2 functionally complement both mutants. Among several other GSTs tested, only soybean GmGST26A (previously called GmHsp26A and GH2/4) and maize GSTIII were found to confer vacuolar sequestration of anthocyanin. Previously, these genes had not been associated with the anthocyanin pathway. Requirements for An9 and Bz2 gene function were investigated by sequencing functional and nonfunctional germinal revertants of an9-T3529, bz2::Ds, and bz2::Mu1. PMID:9668133

Clonorchis sinensis omega-class glutathione transferases play major roles in the protection of the reproductive system during maturation and the response to oxidative stress.

PubMed

Kim, Jeong-Geun; Ahn, Chun-Seob; Kim, Seon-Hee; Bae, Young-An; Kwon, Na-Young; Kang, Insug; Yang, Hyun-Jong; Sohn, Woon-Mok; Kong, Yoon

2016-06-13

Clonorchis sinensis causes a major food-borne helminthic infection. This species locates in mammalian hepatobiliary ducts, where oxidative stressors and hydrophobic substances are profuse. To adapt to the hostile micromilieu and to ensure its long-term survival, the parasite continuously produces a diverse repertoire of antioxidant enzymes including several species of glutathione transferases (GSTs). Helminth GSTs play pertinent roles during sequestration of harmful xenobiotics since most helminths lack the cytochrome P-450 detoxifying enzyme. We isolated and analyzed the biochemical properties of two omega-class GSTs of C. sinensis (CsGSTo1 and CsGSTo2). We observed spatiotemporal expression patterns in accordance with the maturation of the worm's reproductive system. Possible biological protective roles of CsGSTos in these organs under oxidative stress were investigated. The full-length cDNAs of CsGSTo1 and 2 constituted 965 bp and 1,061 bp with open reading frames of 737 bp (246 amino acids) and 669 bp (223 amino acids). They harbored characteristic N-terminal thioredoxin-like and C-terminal α-helical domains. A cysteine residue, which constituted omega-class specific active site, and the glutathione-binding amino acids, were recognized in appropriate positions. They shared 44 % sequence identity with each other and 14.8-44.8 % with orthologues/homologues from other organisms. Bacterially expressed recombinant proteins (rCsGSTo1 and 2) exhibited dehydroascorbate reductase (DHAR) and thioltransferase activities. DHAR activity was higher than thioltransferase activity. They showed weak canonical GST activity toward 1-chloro-2,4-dinitrobenzene. S-hexylglutathione potently and competitively inhibited the active-site at nanomolar concentrations (0.63 and 0.58 nM for rCsGSTo1 and 2). Interestingly, rCsGSTos exhibited high enzyme activity toward mu- and theta-class GST specific substrate, 4-nitrobenzyl chloride. Expression of CsGSTo transcripts and proteins

Identification and characterization of the zebrafish glutathione S-transferase Pi-1.

PubMed

Abunnaja, Maryam S; Kurogi, Katsuhisa; Mohammed, Yasir I; Sakakibara, Yoichi; Suiko, Masahito; Hassoun, Ezdihar A; Liu, Ming-Cheh

2017-10-01

Zebrafish has in recent years emerged as a popular vertebrate model for use in pharmacological and toxicological studies. While there have been sporadic studies on the zebrafish glutathione S-transferases (GSTs), the zebrafish GST gene superfamily still awaits to be fully elucidated. We report here the identification of 15 zebrafish cytosolic GST genes in NCBI GenBank database and the expression, purification, and enzymatic characterization of the zebrafish cytosolic GST Pi-1 (GSTP1). The cDNA encoding the zebrafish GSTP1 was cloned from a 3-month-old female zebrafish, expressed in Eschelichia coli host cells, and purified. Purified GSTP1 displayed glutathione-conjugating activity toward 1-chloro-2,4-dinitrobenzene as a representative substrate. The enzymatic characteristics of the zebrafish GSTP1, including pH-dependency, effects of metal cations, and kinetic parameters, were studied. Moreover, the expression of zebrafish GSTP1 at different developmental stages during embryogenesis, throughout larval development, onto maturity was examined. © 2017 Wiley Periodicals, Inc.

Formation of the accumulative human metabolite and human-specific glutathione conjugate of diclofenac in TK-NOG chimeric mice with humanized livers.

PubMed

Kamimura, Hidetaka; Ito, Satoshi; Nozawa, Kohei; Nakamura, Shota; Chijiwa, Hiroyuki; Nagatsuka, Shin-ichiro; Kuronuma, Miyuki; Ohnishi, Yasuyuki; Suemizu, Hiroshi; Ninomiya, Shin-ichi

2015-03-01

3'-Hydroxy-4'-methoxydiclofenac (VI) is a human-specific metabolite known to accumulate in the plasma of patients after repeated administration of diclofenac sodium. Diclofenac also produces glutathione-conjugated metabolites, some of which are human-specific. In the present study, we investigated whether these metabolites could be generated in humanized chimeric mice produced from TK-NOG mice. After a single oral administration of diclofenac to humanized mice, the unchanged drug in plasma peaked at 0.25 hour and then declined with a half-life (t1/2) of 2.4 hours. 4'-Hydroxydiclofenac (II) and 3'-hydroxydiclofenac also peaked at 0.25 hour and were undetectable within 24 hours. However, VI peaked at 8 hours and declined with a t1/2 of 13 hours. When diclofenac was given once per day, peak and trough levels of VI reached plateau within 3 days. Studies with administration of II suggested VI was generated via II as an intermediate. Among six reported glutathione-conjugated metabolites of diclofenac, M1 (5-hydroxy-4-(glutathion-S-yl)diclofenac) to M6 (2'-(glutathion-S-yl)monoclofenac), we found three dichlorinated conjugates [M1, M2 (4'-hydroxy-3'-(glutathion-S-yl)diclofenac), and M3 (5-hydroxy-6-(glutathion-S-yl)diclofenac)], and a single monochlorinated conjugate [M4 (2'-hydroxy-3'-(glutathion-S-yl)monoclofenac) or M5 (4'-hydroxy-2'-(glutathion-S-yl)monoclofenac)], in the bile of humanized chimeric mice. M4 and M5 are positional isomers and have been previously reported as human-specific in vitro metabolites likely generated via arene oxide and quinone imine-type intermediates, respectively. The biliary monochlorinated metabolite exhibited the same mass spectrum as those of M4 and M5, and we discuss whether this conjugate corresponded to M4 or M5. Overall, humanized TK-NOG chimeric mice were considered to be a functional tool for the study of drug metabolism of diclofenac in humans. Copyright © 2015 by The American Society for Pharmacology and Experimental

Characterization of inter-tissue and inter-strain variability of TCE glutathione conjugation metabolites DCVG, DCVC, and NAcDCVC in the mouse.

PubMed

Luo, Yu-Syuan; Furuya, Shinji; Chiu, Weihsueh; Rusyn, Ivan

2018-01-01

Trichloroethylene (TCE) is a ubiquitous environmental toxicant that is a liver and kidney carcinogen. Conjugation of TCE with glutathione (GSH) leads to formation of nepthrotoxic and mutagenic metabolites postulated to be critical for kidney cancerdevelopment; however, relatively little is known regarding their tissue levels as previous analytical methods for their detection lacked sensitivity. Here, an LC-MS/MS-based method for simultaneous detection of S-(1,2-dichlorovinyl)-glutathione (DCVG), S-(1,2-dichlorovinyl)-L-cysteine (DCVC), and N-acetyl-S-(1,2-dichlorovinyl)-L-cysteine (NAcDCVC) in multiple mouse tissues was developed. This analytical method is rapid, sensitive (limits of detection (LOD) 3-30 fmol across metabolites and tissues), and robust to quantify all three metabolites in liver, kidneys, and serum. The method was used to characterize inter-tissue and inter-strain variability in formation of conjugative metabolites of TCE. Single oral dose of TCE (24, 240 or 800 mg/kg) was administered to male mice from 20 inbred strains of Collaborative Cross. Inter-strain variability in the levels of DCVG, DCVC, and NAcDCVC (GSD = 1.6-2.9) was observed. Whereas NAcDCVC was distributed equally among analyzed tissues, highest levels of DCVG were detected in liver and DCVC in kidneys. Evidence indicated that inter-strain variability in conjugative metabolite formation of TCE might affect susceptibility to adverse health effects and that this method might aid in filling data gaps in human health assessment of TCE.

Glutathione peroxidase deficiency exacerbates ischemia-reperfusion injury in male but not female myocardium: insights into antioxidant compensatory mechanisms.

PubMed

Lim, Chee Chew; Bryan, Nathan S; Jain, Mohit; Garcia-Saura, Maria F; Fernandez, Bernadette O; Sawyer, Douglas B; Handy, Diane E; Loscalzo, Joseph; Feelisch, Martin; Liao, Ronglih

2009-12-01

The female sex has been associated with increased resistance to acute myocardial ischemia-reperfusion (I/R) injury. While enhanced antioxidant capacity has been implicated in female cardioprotection, there is little evidence to support this assumption. Previous studies have shown an important role of cellular glutathione peroxidase (GPx1) in protection of the myocardium from I/R injury. Whether GPx1 is mechanistic in the protection of female myocardium, post-I/R, has not been examined. We utilized a murine model with homozygous deletion of GPx1 and examined its impact on postischemic myocardial recovery in male and female mice. Following I/R, male GPx1(-/-) hearts were more susceptible to contractile and diastolic dysfunction, and this was associated with increased protein carbonyls, a marker of oxidative stress. In contrast, GPx1 deficiency in female hearts did not exacerbate dysfunction or oxidative stress post-I/R. Both wild-type and GPx1(-/-) female hearts exhibited reduced creatine kinase leakage and a more favorable ascorbate redox status compared with males. Following I/R, female GPx1(-/-) hearts showed a comparable decrease in glutathione redox status as their male counterparts; however, they exhibited a greater decrease in nitrate-to-nitrite ratio, suggesting a higher consumption of nitrate in female GPx1(-/-) hearts. Our findings demonstrate that GPx1 is critical for cardioprotection during I/R in male, but not female, mice. The maintenance of cardioprotection in female mice lacking GPx1 post-I/R may be due to an improved ascorbate redox homeostasis and enhanced nitrate-to-nitrite conversion, which would predictably be accompanied by enhanced production of cardioprotective nitric oxide.

Light controlled 3D micromotors powered by bacteria

NASA Astrophysics Data System (ADS)

Vizsnyiczai, Gaszton; Frangipane, Giacomo; Maggi, Claudio; Saglimbeni, Filippo; Bianchi, Silvio; di Leonardo, Roberto

2017-06-01

Self-propelled bacteria can be integrated into synthetic micromachines and act as biological propellers. So far, proposed designs suffer from low reproducibility, large noise levels or lack of tunability. Here we demonstrate that fast, reliable and tunable bio-hybrid micromotors can be obtained by the self-assembly of synthetic structures with genetically engineered biological propellers. The synthetic components consist of 3D interconnected structures having a rotating unit that can capture individual bacteria into an array of microchambers so that cells contribute maximally to the applied torque. Bacterial cells are smooth swimmers expressing a light-driven proton pump that allows to optically control their swimming speed. Using a spatial light modulator, we can address individual motors with tunable light intensities allowing the dynamic control of their rotational speeds. Applying a real-time feedback control loop, we can also command a set of micromotors to rotate in unison with a prescribed angular speed.

Light controlled 3D micromotors powered by bacteria

PubMed Central

Vizsnyiczai, Gaszton; Frangipane, Giacomo; Maggi, Claudio; Saglimbeni, Filippo; Bianchi, Silvio; Di Leonardo, Roberto

2017-01-01

Self-propelled bacteria can be integrated into synthetic micromachines and act as biological propellers. So far, proposed designs suffer from low reproducibility, large noise levels or lack of tunability. Here we demonstrate that fast, reliable and tunable bio-hybrid micromotors can be obtained by the self-assembly of synthetic structures with genetically engineered biological propellers. The synthetic components consist of 3D interconnected structures having a rotating unit that can capture individual bacteria into an array of microchambers so that cells contribute maximally to the applied torque. Bacterial cells are smooth swimmers expressing a light-driven proton pump that allows to optically control their swimming speed. Using a spatial light modulator, we can address individual motors with tunable light intensities allowing the dynamic control of their rotational speeds. Applying a real-time feedback control loop, we can also command a set of micromotors to rotate in unison with a prescribed angular speed. PMID:28656975

Relationships among Cortical Glutathione Levels, Brain Amyloidosis, and Memory in Healthy Older Adults Investigated In Vivo with 1H-MRS and Pittsburgh Compound-B PET.

PubMed

Chiang, G C; Mao, X; Kang, G; Chang, E; Pandya, S; Vallabhajosula, S; Isaacson, R; Ravdin, L D; Shungu, D C

2017-06-01

Oxidative stress has been implicated as an important pathologic mechanism in the development of Alzheimer disease. The purpose of this study was to assess whether glutathione levels, detected noninvasively with proton MR spectroscopy, are associated with brain amyloidosis and memory in a community-dwelling cohort of healthy older adults. Fifteen cognitively healthy subjects were prospectively enrolled in this study. All subjects underwent 1 H-MR spectroscopy of glutathione, a positron-emission tomography scan with an amyloid tracer, and neuropsychological testing by using the Repeatable Battery for the Assessment of Neuropsychological Status. Associations among glutathione levels, brain amyloidosis, and memory were assessed by using multivariate regression models. Lower glutathione levels were associated with greater brain amyloidosis in the temporal ( P = .03) and parietal ( P = .05) regions, adjusted for apolipoprotein E ε4 carrier status. There were no significant associations between glutathione levels and cognitive scores. This study found an association between cortical glutathione levels and brain amyloidosis in healthy older adults, suggesting a potential role for 1 H-MR spectroscopy measures of glutathione as a noninvasive biomarker of early Alzheimer disease pathogenesis. © 2017 by American Journal of Neuroradiology.

Quercetin affects glutathione levels and redox ratio in human aortic endothelial cells not through oxidation but formation and cellular export of quercetin-glutathione conjugates and upregulation of glutamate-cysteine ligase.

PubMed

Li, Chuan; Zhang, Wei-Jian; Choi, Jaewoo; Frei, Balz

2016-10-01

Endothelial dysfunction due to vascular inflammation and oxidative stress critically contributes to the etiology of atherosclerosis. The intracellular redox environment plays a key role in regulating endothelial cell function and is intimately linked to cellular thiol status, including and foremost glutathione (GSH). In the present study we investigated whether and how the dietary flavonoid, quercetin, affects GSH status of human aortic endothelial cells (HAEC) and their response to oxidative stress. We found that treating cells with buthionine sulfoximine to deplete cellular GSH levels significantly reduced the capacity of quercetin to inhibit lipopolysaccharide (LPS)-induced oxidant production. Furthermore, incubation of HAEC with quercetin caused a transient decrease and then full recovery of cellular GSH concentrations. The initial decline in GSH was not accompanied by a corresponding increase in glutathione disulfide (GSSG). To the contrary, GSSG levels, which were less than 0.5% of GSH levels at baseline (0.26±0.01 vs. 64.7±1.9nmol/mg protein, respectively), decreased by about 25% during incubation with quercetin. As a result, the GSH: GSSG ratio increased by about 70%, from 253±7 to 372±23. These quercetin-induced changes in GSH and GSSG levels were not affected by treating HAEC with 500µM ascorbic acid phosphate for 24h to increase intracellular ascorbate levels. Incubation of HAEC with quercetin also led to the appearance of extracellular quercetin-glutathione conjugates, which was paralleled by upregulation of the multidrug resistance protein 1 (MRP1). Furthermore, quercetin slightly but significantly increased mRNA and protein levels of glutamate-cysteine ligase (GCL) catalytic and modifier subunits. Taken together, our results suggest that quercetin causes loss of GSH in HAEC, not because of oxidation but due to formation and cellular export of quercetin-glutathione conjugates. Induction by quercetin of GCL subsequently restores GSH levels, thereby

Barley yellow dwarf virus infection and elevated CO2 alter the antioxidants ascorbate and glutathione in wheat.

PubMed

Vandegeer, Rebecca K; Powell, Kevin S; Tausz, Michael

2016-07-20

Plant antioxidants ascorbate and glutathione play an important role in regulating potentially harmful reactive oxygen species produced in response to virus infection. Barley yellow dwarf virus is a widespread viral pathogen that systemically infects cereal crops including wheat, barley and oats. In addition, rising atmospheric CO 2 will alter plant growth and metabolism, including many potential but not well understood effects on plant-virus interactions. In order to better understand the wheat-BYDV interaction and any potential changes under elevated CO 2 , the total concentration and oxidised fraction of ascorbate and glutathione was measured in leaves of a susceptible wheat cultivar (Triticum aestivum L. 'Yitpi') infected with Barley yellow dwarf virus-PAV (Padi Avenae virus) and grown under elevated CO 2 in controlled environment chambers. Virus infection decreased total leaf ascorbate and glutathione concentrations and increased the fraction of oxidised ascorbate (dehydroascorbate). Elevated CO 2 decreased the fraction of oxidised ascorbate. In this work, we demonstrate that systemic infection by a phloem-restricted virus weakens the antioxidant pools of ascorbate and glutathione. In addition, elevated CO 2 may decrease oxidative stress, for example, from virus infection, but there was no direct evidence for an interactive effect between treatments. Copyright © 2016 Elsevier GmbH. All rights reserved.

Lack of association between the glutathione-s-transferase genes (GSTT1 and GSTM1) and nasal polyposis.

PubMed

Arbag, Hamdi; Cora, Tulin; Acar, Hasan; Ozturk, Kayhan; Sari, Fatih; Ulusoy, Bulent

2006-03-01

To evaluate the glutation-S-transferase (GST) polymorphisms (GSTM1 and GSTT1) in nasal polyposis (NP). The study population consisted of 102 unrelated healthy individuals and 98 patients with NP (67 without asthma, 31 with asthma). Genotyping of the polymorphism in the GSTM1 and GSTT1 genes was performed using the multiplex polymerase chain reaction (PCR)-based method. GSTM1 and GSTT1 null-genotypes were found in 46.1% and 23.5% of the controls, and in 43.9% and 33.7% of the NP patients, respectively. These differences were not significant (for GSTM1 null odds ratio (OR) = 0.92; 95% confidence interval (CI) = 0.52-1.6 and for GSTT1, OR = 1.65; 95% CI = 0.89-3.07). Although no significant difference for combined GSTM1 and GSTT1 null genotypes between control (8.8%) and NP patients (17.3%) was found, there was a 2.16-fold increased proportion in the NP with the combined GSTM1-null and GSTT1-null genotype (OR = 2.16; 95% CI = 0.91-5.13). These results suggest that there is lack of association between GSTM1 and GSTT1 polymorphisms and NP. The GSTM1 or GSTT1 polymorphisms had also no relevant developing effect on NP patients without or with asthma.

N-acetylcysteine stimulates protein synthesis in enterocytes independently of glutathione synthesis.

PubMed

Yi, Dan; Hou, Yongqing; Wang, Lei; Long, Minhui; Hu, Shengdi; Mei, Huimin; Yan, Liqiong; Hu, Chien-An Andy; Wu, Guoyao

2016-02-01

Dietary supplementation with N-acetylcysteine (NAC) has been reported to improve intestinal health and treat gastrointestinal diseases. However, the underlying mechanisms are not fully understood. According to previous reports, NAC was thought to exert its effect through glutathione synthesis. This study tested the hypothesis that NAC enhances enterocyte growth and protein synthesis independently of cellular glutathione synthesis. Intestinal porcine epithelial cells were cultured for 3 days in Dulbecco's modified Eagle medium containing 0 or 100 μM NAC. To determine a possible role for GSH (the reduced form of glutathione) in mediating the effect of NAC on cell growth and protein synthesis, additional experiments were conducted using culture medium containing 100 μM GSH, 100 μM GSH ethyl ester (GSHee), diethylmaleate (a GSH-depletion agent; 10 μM), or a GSH-synthesis inhibitor (buthionine sulfoximine, BSO; 20 μM). NAC increased cell proliferation, GSH concentration, and protein synthesis, while inhibiting proteolysis. GSHee enhanced cell proliferation and GSH concentration without affecting protein synthesis but inhibited proteolysis. Conversely, BSO or diethylmaleate reduced cell proliferation and GSH concentration without affecting protein synthesis, while promoting protein degradation. At the signaling level, NAC augmented the protein abundance of total mTOR, phosphorylated mTOR, and phosphorylated 70S6 kinase as well as mRNA levels for mTOR and p70S6 kinase in IPEC-1 cells. Collectively, these results indicate that NAC upregulates expression of mTOR signaling proteins to stimulate protein synthesis in enterocytes independently of GSH generation. Our findings provide a hitherto unrecognized biochemical mechanism for beneficial effects of NAC in intestinal cells.

Antibiotic Resistance in Plant-Pathogenic Bacteria.

PubMed

Sundin, George W; Wang, Nian

2018-06-01

Antibiotics have been used for the management of relatively few bacterial plant diseases and are largely restricted to high-value fruit crops because of the expense involved. Antibiotic resistance in plant-pathogenic bacteria has become a problem in pathosystems where these antibiotics have been used for many years. Where the genetic basis for resistance has been examined, antibiotic resistance in plant pathogens has most often evolved through the acquisition of a resistance determinant via horizontal gene transfer. For example, the strAB streptomycin-resistance genes occur in Erwinia amylovora, Pseudomonas syringae, and Xanthomonas campestris, and these genes have presumably been acquired from nonpathogenic epiphytic bacteria colocated on plant hosts under antibiotic selection. We currently lack knowledge of the effect of the microbiome of commensal organisms on the potential of plant pathogens to evolve antibiotic resistance. Such knowledge is critical to the development of robust resistance management strategies to ensure the safe and effective continued use of antibiotics in the management of critically important diseases. Expected final online publication date for the Annual Review of Phytopathology Volume 56 is August 25, 2018. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Hepatic glutathione metabolism and lipid peroxidation in response to excess dietary selenomethionine and selenite in mallard ducklings

USGS Publications Warehouse

Hoffman, D.J.; Heinz, G.H.; Krynitsky, A.J.

1989-01-01

Selenium from selenomethionine accumulated in a dose-dependent manner in the liver, resulting in a decrease in hepatic-reduced glutathione with a corresponding decrease in total hepatic thiols. There was a dose-dependent increase in the oxidized to reduced glutathione ratio, and an increase in lipid peroxidation. These findings indicate that Se in the diet at 10 ppm and higher causes significant sublethal alterations in mallard ducklings, and 20-40 ppm causes significant hepatotoxicity.

DehydroalanylGly, a new post translational modification resulting from the breakdown of glutathione.

PubMed

Friedrich, Michael G; Wang, Zhen; Schey, Kevin L; Truscott, Roger J W

2018-04-01

The human body contains numerous long-lived proteins which deteriorate with age, typically by racemisation, deamidation, crosslinking and truncation. Previously we elucidated one reaction responsible for age-related crosslinking, the spontaneous formation of dehydroalanine (DHA) intermediates from phosphoserine and cysteine. This resulted in non-disulphide covalent crosslinks. The current paper outlines a novel posttranslational modification (PTM) in human proteins, which involves the addition of dehydroalanylglycine (DHAGly) to Lys residues. Human lens digests were examined by mass spectrometry for the presence of (DHA)Gly (+144.0535 Da) adducts to Lys residues. Peptide model studies were undertaken to elucidate the mechanism of formation. In the lens, this PTM was detected at 18 lysine sites in 7 proteins. Using model peptides, a pathway for its formation was found to involve initial formation of the glutathione degradation product, γ-Glu(DHA)Gly from oxidised glutathione (GSSG). Once the Lys adduct formed, the Glu residue was lost in a hydrolytic mechanism apparently catalysed by the ε-amino group of the Lys. This discovery suggests that within cells, the functional groups of amino acids in proteins may be susceptible to modification by reactive metabolites derived from GSSG. Our finding demonstrates a novel +144.0535 Da PTM arising from the breakdown of oxidised glutathione. Copyright © 2018. Published by Elsevier B.V.

Glutathione exposes sequential IgE-epitopes in ovomucoid relevant in persistent egg allergy.

PubMed

Roth-Walter, Franziska; Starkl, Philipp; Zuberbier, Torsten; Hummel, Karin; Nöbauer, Karin; Razzazi-Fazeli, Ebrahim; Brunner, Richard; Pali-Schöll, Isabella; Kinkel, Janis; Felix, Ferdinand; Jensen-Jarolim, Erika; Kinaciyan, Tamar

2013-03-01

Patients with persistent egg allergy have more immunoglobulin E (IgE) against sequential than conformational epitopes of ovomucoid (OVO). Here, we aimed to identify compounds capable to render sequential epitopes in egg. Glutathione was used for in vitro reduction of OVO and circular dichroism analyses were performed. Glutathione reduced OVO in a concentration-dependent manner. Egg white was analyzed for reduced proteins with a thiol probe and by MALDI-TOF/TOF. In unprocessed total egg white, several reduced proteins were detected by the thiol probe, among them reduced ovalbumin could be confirmed with MS analyses. Egg-allergics or sensitized controls were tested serologically (n = 19) for IgE against native and reduced OVO and in skin prick tests (n = 9). More patients had IgE against reduced than native OVO in Western blots. In skin prick test, five out of seven persistent egg-allergics and none of the controls reacted with reduced OVO. Reduced egg proteins are present in natural egg white. Glutathione, which is present in egg and furthermore is used as texture-improving additive in processed food, is capable of reducing OVO. Patients with persistent egg allergy reacted rather to reduce the native OVO. Hence, our data indicate that reduction is a novel natural and processing-associated principle, which contributes to the allergenicity of food. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Cellular copper homeostasis: current concepts on its interplay with glutathione homeostasis and its implication in physiology and human diseases.

PubMed

Bhattacharjee, Ashima; Chakraborty, Kaustav; Shukla, Aditya

2017-10-18

Copper is a trace element essential for almost all living organisms. But the level of intracellular copper needs to be tightly regulated. Dysregulation of cellular copper homeostasis leading to various diseases demonstrates the importance of this tight regulation. Copper homeostasis is regulated not only within the cell but also within individual intracellular compartments. Inactivation of export machinery results in excess copper being redistributed into various intracellular organelles. Recent evidence suggests the involvement of glutathione in playing an important role in regulating copper entry and intracellular copper homeostasis. Therefore interplay of both homeostases might play an important role within the cell. Similar to copper, glutathione balance is tightly regulated within individual cellular compartments. This review explores the existing literature on the role of glutathione in regulating cellular copper homeostasis. On the one hand, interplay of glutathione and copper homeostasis performs an important role in normal physiological processes, for example neuronal differentiation. On the other hand, perturbation of the interplay might play a key role in the pathogenesis of copper homeostasis disorders.

Cigarette smoke inhibits macrophage sensing of Gram-negative bacteria and lipopolysaccharide: relative roles of nicotine and oxidant stress.

PubMed

McMaster, S K; Paul-Clark, M J; Walters, M; Fleet, M; Anandarajah, J; Sriskandan, S; Mitchell, J A

2008-02-01

Smoking cigarettes is a major risk factor for the development of cardiovascular and respiratory disease. Moreover, smokers are more prone to infections. This has been associated with a suppression of the immune system by smoke. However, it is not clear how cigarette smoke affects the ability of immune cells to sense pathogens. Cigarette smoke contains a large number of molecules which may mediate responses on immune cells and of these, nicotine and oxidants have both been identified as inhibitory for the sensing of bacterial lipopolysaccharide (LPS). Nitric oxide synthase (NOS) and tumour necrosis factor (TNF)-alpha are both induced in macrophages on stimulation with Gram negative bacteria or LPS. We used murine macrophages stimulated with whole heat-killed bacteria or LPS. We measured output of NO (as nitrite) and TNFalpha, NOS protein by Western blotting and cellular oxidant stress. Cigarette smoke extract suppressed the ability of murine macrophages to release NO, but not TNFalpha in response to whole bacteria. Cigarette smoke extract also inhibited nitric oxide synthase II protein expression in response to LPS. The effects of cigarette smoke extract on nitrite formation stimulated by LPS were unaffected by inhibition of nicotinic receptors with alpha-bungarotoxin (100 units ml(-1)). However, the effects of cigarette smoke extract on LPS-induced nitrite formation were mimicked by hydrogen peroxide and reversed by the anti-oxidants N-acetyl cysteine and glutathione. We suggest that cigarette smoke exerts its immunosuppressive effects through an oxidant-dependent and not a nicotine-dependent mechanism.

Commensal bacteria and essential amino acids control food choice behavior and reproduction

PubMed Central

Fioreze, Gabriela Tondolo; Anjos, Margarida; Baltazar, Célia; Elias, Ana Paula; Itskov, Pavel M.; Piper, Matthew D. W.

2017-01-01

Choosing the right nutrients to consume is essential to health and wellbeing across species. However, the factors that influence these decisions are poorly understood. This is particularly true for dietary proteins, which are important determinants of lifespan and reproduction. We show that in Drosophila melanogaster, essential amino acids (eAAs) and the concerted action of the commensal bacteria Acetobacter pomorum and Lactobacilli are critical modulators of food choice. Using a chemically defined diet, we show that the absence of any single eAA from the diet is sufficient to elicit specific appetites for amino acid (AA)-rich food. Furthermore, commensal bacteria buffer the animal from the lack of dietary eAAs: both increased yeast appetite and decreased reproduction induced by eAA deprivation are rescued by the presence of commensals. Surprisingly, these effects do not seem to be due to changes in AA titers, suggesting that gut bacteria act through a different mechanism to change behavior and reproduction. Thus, eAAs and commensal bacteria are potent modulators of feeding decisions and reproductive output. This demonstrates how the interaction of specific nutrients with the microbiome can shape behavioral decisions and life history traits. PMID:28441450

Commensal bacteria and essential amino acids control food choice behavior and reproduction.

PubMed

Leitão-Gonçalves, Ricardo; Carvalho-Santos, Zita; Francisco, Ana Patrícia; Fioreze, Gabriela Tondolo; Anjos, Margarida; Baltazar, Célia; Elias, Ana Paula; Itskov, Pavel M; Piper, Matthew D W; Ribeiro, Carlos

2017-04-01

Choosing the right nutrients to consume is essential to health and wellbeing across species. However, the factors that influence these decisions are poorly understood. This is particularly true for dietary proteins, which are important determinants of lifespan and reproduction. We show that in Drosophila melanogaster, essential amino acids (eAAs) and the concerted action of the commensal bacteria Acetobacter pomorum and Lactobacilli are critical modulators of food choice. Using a chemically defined diet, we show that the absence of any single eAA from the diet is sufficient to elicit specific appetites for amino acid (AA)-rich food. Furthermore, commensal bacteria buffer the animal from the lack of dietary eAAs: both increased yeast appetite and decreased reproduction induced by eAA deprivation are rescued by the presence of commensals. Surprisingly, these effects do not seem to be due to changes in AA titers, suggesting that gut bacteria act through a different mechanism to change behavior and reproduction. Thus, eAAs and commensal bacteria are potent modulators of feeding decisions and reproductive output. This demonstrates how the interaction of specific nutrients with the microbiome can shape behavioral decisions and life history traits.

Identification and clarification of the role of key active site residues in bacterial glutathione S-transferase zeta/maleylpyruvate isomerase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fang, Ti; Li, De-Feng; Zhou, Ning-Yi, E-mail: n.zhou@pentium.whiov.ac.cn

2011-07-08

Highlights: {yields} Application of site-directed mutagenesis to probe the active site residues of glutathione-dependent maleylpyruvate isomerase. {yields} Two conserved residues, Arg8 and Arg176, in zeta class glutathione S-transferases are critical for maleylpyruvate orientation and enolization. {yields} Arg109, found exclusively in NagL, participates in k{sub cat} regulation. {yields} The T11A mutant exhibited a significantly decreased K{sub m} value for glutathione with little impact on maleylpyruvate kinetics. {yields} The Thr11 residue appears to have significance in the evolution of glutathione S-transferase classes. -- Abstract: The maleylpyruvate isomerase NagL from Ralstonia sp. strain U2, which has been structurally characterized previously, catalyzes the isomerizationmore » of maleylpyruvate to fumarylpyruvate. It belongs to the class zeta glutathione S-transferases (GSTZs), part of the cytosolic GST family (cGSTs). In this study, site-directed mutagenesis was conducted to probe the functions of 13 putative active site residues. Steady-state kinetic information for mutants in the reduced glutathione (GSH) binding site, suggested that (a) Gln64 and Asp102 interact directly with the glutamyl moiety of glutathione, (b) Gln49 and Gln64 are involved in a potential electron-sharing network that influences the ionization of the GSH thiol. The information also suggests that (c) His38, Asn108 and Arg109 interact with the GSH glycine moiety, (d) His104 has a role in the ionization of the GSH sulfur and the stabilization of the maleyl terminal carboxyl group in the reaction intermediate and (e) Arg110 influences the electron distribution in the active site and therefore the ionization of the GSH thiolate. Kinetic data for mutants altered in the substrate-binding site imply that (a) Arg8 and Arg176 are critical for maleylpyruvate orientation and enolization, and (b) Arg109 (exclusive to NagL) participates in k{sub cat} regulation. Surprisingly, the T11A mutant

The role of nuclear factor E2-Related factor 2 and uncoupling protein 2 in glutathione metabolism: Evidence from an in vivo gene knockout study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Yanyan; The Hamner Institutes for Health Sciences, Research Triangle Park, NC; Xu, Yuanyuan, E-mail: yyxu@cmu.edu.cn

Nuclear factor E2-related factor 2 (NRF2) and uncoupling protein 2 (UCP2) are indicated to protect from oxidative stress. They also play roles in the homeostasis of glutathione. However, the detailed mechanisms are not well understood. In the present study, we found Nrf2-knockout (Nrf2-KO) mice exhibited altered glutathione homeostasis and reduced expression of various genes involved in GSH biosynthesis, regeneration, utilization and transport in the liver. Ucp2-knockout (Ucp2-KO) mice exhibited altered glutathione homeostasis in the liver, spleen and blood, as well as increased transcript of cystic fibrosis transmembrane conductance regulator in the liver, a protein capable of mediating glutathione efflux. Nrf2-Ucp2-doublemore » knockout (DKO) mice showed characteristics of both Nrf2-KO and Ucp2-KO mice. But no significant difference was observed in DKO mice when compared with Nrf2-KO or Ucp2-KO mice, except in blood glutathione levels. These data suggest that ablation of Nrf2 and Ucp2 leads to disrupted GSH balance, which could result from altered expression of genes involved in GSH metabolism. DKO may not evoke more severe oxidative stress than the single gene knockout. - Highlights: • Nrf2/Ucp2 deficiency leads to alteration of glutathione homeostasis. • Nrf2 regulates expression of genes in glutathione generation and utilization. • Ucp2 affects glutathione metabolism by regulating hepatic efflux of glutathione. • Nrf2 deficiency may not aggravate oxidative stress in Ucp2-deficient mice.« less

The effect of aqueous speciation and cellular ligand binding on the biotransformation and bioavailability of methylmercury in mercury-resistant bacteria.

PubMed

Ndu, Udonna; Barkay, Tamar; Schartup, Amina Traore; Mason, Robert P; Reinfelder, John R

2016-02-01

Mercury resistant bacteria play a critical role in mercury biogeochemical cycling in that they convert methylmercury (MeHg) and inorganic mercury to elemental mercury, Hg(0). To date there are very few studies on the effects of speciation and bioavailability of MeHg in these organisms, and even fewer studies on the role that binding to cellular ligands plays on MeHg uptake. The objective of this study was to investigate the effects of thiol complexation on the uptake of MeHg by measuring the intracellular demethylation-reduction (transformation) of MeHg to Hg(0) in Hg-resistant bacteria. Short-term intracellular transformation of MeHg was quantified by monitoring the loss of volatile Hg(0) generated during incubations of bacteria containing the complete mer operon (including genes from putative mercury transporters) exposed to MeHg in minimal media compared to negative controls with non-mer or heat-killed cells. The results indicate that the complexes MeHgOH, MeHg-cysteine, and MeHg-glutathione are all bioavailable in these bacteria, and without the mer operon there is very little biological degradation of MeHg. In both Pseudomonas stutzeri and Escherichia coli, there was a pool of MeHg that was not transformed to elemental Hg(0), which was likely rendered unavailable to Mer enzymes by non-specific binding to cellular ligands. Since the rates of MeHg accumulation and transformation varied more between the two species of bacteria examined than among MeHg complexes, microbial bioavailability, and therefore microbial demethylation, of MeHg in aquatic systems likely depends more on the species of microorganism than on the types and relative concentrations of thiols or other MeHg ligands present.

Treatment of antimony mine drainage: challenges and opportunities with special emphasis on mineral adsorption and sulfate reducing bacteria.

PubMed

Li, Yongchao; Hu, Xiaoxian; Ren, Bozhi

2016-01-01

The present article summarizes antimony mine distribution, antimony mine drainage generation and environmental impacts, and critically analyses the remediation approach with special emphasis on iron oxidizing bacteria and sulfate reducing bacteria. Most recent research focuses on readily available low-cost adsorbents, such as minerals, wastes, and biosorbents. It is found that iron oxides prepared by chemical methods present superior adsorption ability for Sb(III) and Sb(V). However, this process is more costly and iron oxide activity can be inhibited by plenty of sulfate in antimony mine drainage. In the presence of sulfate reducing bacteria, sulfate can be reduced to sulfide and form Sb(2)S(3) precipitates. However, dissolved oxygen and lack of nutrient source in antimony mine drainage inhibit sulfate reducing bacteria activity. Biogenetic iron oxide minerals from iron corrosion by iron-oxidizing bacteria may prove promising for antimony adsorption, while the micro-environment generated from iron corrosion by iron oxidizing bacteria may provide better growth conditions for symbiotic sulfate reducing bacteria. Finally, based on biogenetic iron oxide adsorption and sulfate reducing bacteria followed by precipitation, the paper suggests an alternative treatment for antimony mine drainage that deserves exploration.

Role of protein-glutathione contacts in defining glutaredoxin-3 [2Fe-2S] cluster chirality, ligand exchange and transfer chemistry.

PubMed

Sen, Sambuddha; Cowan, J A

2017-10-01

Monothiol glutaredoxins (Grx) serve as intermediate cluster carriers in iron-sulfur cluster trafficking. The [2Fe-2S]-bound holo forms of Grx proteins display cysteinyl coordination from exogenous glutathione (GSH), in addition to contact from protein-derived Cys. Herein, we report mechanistic studies that investigate the role of exogenous glutathione in defining cluster chirality, ligand exchange, and the cluster transfer chemistry of Saccharomyces cerevisiae Grx3. Systematic perturbations were introduced to the glutathione-binding site by substitution of conserved charged amino acids that form crucial electrostatic contacts with the glutathione molecule. Native Grx3 could also be reconstituted in the absence of glutathione, with either DTT, BME or free L-cysteine as the source of the exogenous Fe-S ligand contact, while retaining full functional reactivity. The delivery of the [2Fe-2S] cluster to Grx3 from cluster donor proteins such as Isa, Nfu, and a [2Fe-2S](GS) 4 complex, revealed that electrostatic contacts are of key importance for positioning the exogenous glutathione that in turn influences the chiral environment of the cluster. All Grx3 derivatives were reconstituted by standard chemical reconstitution protocols and found to transfer cluster to apo ferredoxin 1 (Fdx1) at rates comparable to native protein, even when using DTT, BME or free L-cysteine as a thiol source in place of GSH during reconstitution. Kinetic analysis of cluster transfer from holo derivatives to apo Fdx1 has led to a mechanistic model for cluster transfer chemistry of native holo Grx3, and identification of the likely rate-limiting step for the reaction.

Negative effect of combined cysteine and glutathione in soy lecithin-based extender on post-thawed ram spermatozoa.

PubMed

Zhandi, Mahdi; Sharafi, Mohsen

2015-09-01

This study was conducted to evaluate the effect of combined cysteine and glutathione in soy lecithin-based semen extender on post-thawed ram sperm quality. A total of 28 ejaculates were collected twice a week (from four rams) during breeding season. In each replicate, semen samples (n = 4, one ejaculate for each ram) were pooled and divided into three equal parts, and each part was diluted with one of following extender: (1) soy lecithin-based extender containing no cysteine and no glutathione (C0-G0), (2) soy lecithin-based extender containing cysteine (5 mM) and glutathione (5 mM) (C5-G5), and (3) soy lecithin-based extender containing cysteine (10 mM) and glutathione (10 mM) (C10-G10). After freeze-thawing process, motility and velocity parameters, plasma membrane integrity and functionality, mitochondrial activity, and apoptosis features of spermatozoa were evaluated. The obtained results showed that total and progressive motility, plasma membrane integrity and functionality, and live post-thawed spermatozoa was lower in C10-G10 extender compared to C0-G0 and C5-G5 extenders (P < 0.05). Also, the percentage of dead spermatozoa was higher in C10-G10 extender compared to C0-G0 extender (P < 0.05). Apoptotic spermatozoa was lower in C10-G10 extender compared to C0-G0 and C5-G5 extenders (P < 0.05). All velocity parameters, exception of BCF, did not different between extenders (P > 0.05). In conclusion, it seems that high concentration of combined cysteine and glutathione in soy lecithin-based semen extender has a detrimental effect of post-thawed ram sperm quality.

Modulation of nitric oxide-mediated metal release from metallothionein by the redox state of glutathione in vitro.

PubMed

Khatai, Leila; Goessler, Walter; Lorencova, Helena; Zangger, Klaus

2004-06-01

Metallothioneins (MTs) release bound metals when exposed to nitric oxide. At inflammatory sites, both metallothionein and inducible nitric oxide synthase (iNOS) are induced by the same factors and the zinc released from metallothionein by NO suppresses both the induction and activity of iNOS. In a search for a possible modulatory mechanism of this coexpression of counteracting proteins, we investigated the role of the glutathione redox state in vitro because the oxidation state of thiols is involved in the metal binding in Cd-S or Zn-S clusters found in metallothioneins, and NO also binds to reduced glutathione via S-nitrosation. Using a variety of techniques, we found that NO and also ONOO(-)-mediated metal release from purified MTs is suppressed by reduced glutathione (GSH), but not by oxidized glutathione. Considering the millimolar concentrations of GSH present in mammalian cells, the metal release from MTs by NO should play no role in living systems. Therefore, the fact that it has been observed in vivo points to a hitherto unknown mechanism or additional compound(s) being involved in this physiologically relevant reaction and as long as this additional factor is not found experimental results on the MT-NO interaction should be treated with caution. Contrary to the peroxynitrite-induced activation of guanylyl cyclase, where GSH is needed, we found that the metal release from metallothionein by peroxynitrite is not enhanced, but also suppressed by reduced glutathione. In addition, we show that zinc, the major natural metal ligand in mammalian MTs and suppressor of iNOS, is released more readily under the influence of NO than cadmium, but in contrast to the MT isoform 1, the amount of metal released from the beta-domain of MT-2 is comparable to that from the alpha-domain.

Pharmacodynamics of S-dimethylarsino-glutathione, a putative metabolic intermediate of inorganic arsenic, in mice.

PubMed

Kato, Ayaka; Kobayashi, Yayoi; Udagawa, Osamu; Hirano, Seishiro

2017-02-15

Inorganic arsenicals are well-known carcinogens, whereas arsenite (iAs III ) compounds are now recognized as potent therapeutic agents for several leukemias, and arsenic trioxide has been used for the treatment of recurrent acute promyelocytic leukemia (APL). However, recent clinical trials revealed that arsenite is not always effective for non-APL malignancies. Another arsenical, S-dimethylarsino-glutathione ([DMA III (GS)]), which is a putative metabolic intermediate in the hepatic metabolism of iAs III , shows promise for treating several types of lymphoma. However, the metabolism of [DMA III (GS)] has not been well investigated, probably because [DMA III (GS)] is not stable in biological fluids where the concentration of glutathione is low. In the present study, we injected [DMA III (GS)] intravenously into mice and compared the tissue distribution and metabolic dynamics of [DMA III (GS)] with those of sodium arsenite (NaAsO 2 ). We found a unique organ preference for the distribution of [DMA III (GS)] to the lung and brain in comparison to NaAsO 2 . Furthermore, [DMA III (GS)] appeared to bind to serum albumin by exchanging its glutathione moiety quickly after administration, providing novel insights into the longer retention of [DMA III (GS)] in plasma. Copyright © 2016 Elsevier Inc. All rights reserved.

Expression of a Drosophila glutathione transferase in Arabidopsis confers the ability to detoxify the environmental pollutant, and explosive, 2,4,6-trinitrotoluene.

PubMed

Tzafestas, Kyriakos; Razalan, Maria M; Gyulev, Ivan; Mazari, Aslam M A; Mannervik, Bengt; Rylott, Elizabeth L; Bruce, Neil C

2017-04-01

The explosive 2,4,6-trinitrotoluene (TNT) is a significant, global environmental pollutant that is both toxic and recalcitrant to degradation. Given the sheer scale and inaccessible nature of contaminated areas, phytoremediation may be a viable clean-up approach. Here, we have characterized a Drosophila melanogaster glutathione transferase (DmGSTE6) which has activity towards TNT. Recombinantly expressed, purified DmGSTE6 produces predominantly 2-glutathionyl-4,6-dinitrotoluene, and has a 2.5-fold higher Maximal Velocity (V max ), and five-fold lower Michaelis Constant (K m ) than previously characterized TNT-active Arabidopsis thaliana (Arabidopsis) GSTs. Expression of DmGSTE6 in Arabidopsis conferred enhanced resistance to TNT, and increased the ability to remove TNT from contaminated soil relative to wild-type plants. Arabidopsis lines overexpressing TNT-active GSTs AtGST-U24 and AtGST-U25 were compromised in biomass production when grown in the absence of TNT. This yield drag was not observed in the DmGSTE6-expressing Arabidopsis lines. We hypothesize that increased levels of endogenous TNT-active GSTs catalyse excessive glutathionylation of endogenous substrates, depleting glutathione pools, an activity that DmGST may lack. In conclusion, DmGSTE6 has activity towards TNT, producing a compound with potential for further biodegradation. Selecting or manipulating plants to confer DmGSTE6-like activity could contribute towards development of phytoremediation strategies to clean up TNT from polluted military sites. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

Selection of Surrogate Bacteria for Use in Food Safety Challenge Studies: A Review.

PubMed

Hu, Mengyi; Gurtler, Joshua B

2017-09-01

Nonpathogenic surrogate bacteria are prevalently used in a variety of food challenge studies in place of foodborne pathogens such as Listeria monocytogenes, Salmonella, Escherichia coli O157:H7, and Clostridium botulinum because of safety and sanitary concerns. Surrogate bacteria should have growth characteristics and/or inactivation kinetics similar to those of target pathogens under given conditions in challenge studies. It is of great importance to carefully select and validate potential surrogate bacteria when verifying microbial inactivation processes. A validated surrogate responds similar to the targeted pathogen when tested for inactivation kinetics, growth parameters, or survivability under given conditions in agreement with appropriate statistical analyses. However, a considerable number of food studies involving putative surrogate bacteria lack convincing validation sources or adequate validation processes. Most of the validation information for surrogates in these studies is anecdotal and has been collected from previous publications but may not be sufficient for given conditions in the study at hand. This review is limited to an overview of select studies and discussion of the general criteria and approaches for selecting potential surrogate bacteria under given conditions. The review also includes a list of documented bacterial pathogen surrogates and their corresponding food products and treatments to provide guidance for future studies.

[Selective-differential nutrient medium "Shewanella IRHLS agar" for isolation of Shewanella genus bacteria].

PubMed

Sivolodsky, E P

2015-01-01

Development of a selective-differential nutrient medium for isolation of Shewanella genus bacteria. 73 strains of Shewanella bacteria (S. algae--3, S. baltica--26, S. putrefaciens--44) and 80 strains of 22 other bacteria genera were used. Shewanella species were identified by methods and criteria proposed by Nozue H. et al., 1992; Khashe S. et al., 1998. Nutrient media "Shewanella IRHLS Agar" for shewanella isolation was developed. Medium selective factors: irgazan DP-300 (I). 0.14-0.2 g/l and rifampicin (R) 0.0005-0.001 g/l. Shevanella colonies were detected by the production of hydrogen sulfide (H), lipase presence (L), lack of sorbitol fermentation (S). The medium suppressed the growth of hydrogen sulfide producers (Salmonella, Proteus) and blocked hydrogen sulfide production by Citrobacter. Growth of Escherichia, Enterobacter, Klebsiella, Shigella, Staphylococcus, Bacillus was also suppressed, Analytical sensitivity of the medium was 1-2 CFU/ml for Shewanella and Stenotrophomonas, Aerombnas, Serratia genera bacteria. 72 strains of Shewanella were isolated from water of Neva river in this medium, 91.7 ± 3.2% of those produced H2S. 1 strain of S. algae was isolated from clinical material. The developed media allows to use it in a complex for Stenotrophomo- nas sp., Aeromonas sp., Serratia sp., Citrobactersp. and Shewanella bacteria isolation.

Glutathione in Cancer Cell Death

PubMed Central

Ortega, Angel L.; Mena, Salvador; Estrela, Jose M.

2011-01-01

Glutathione (L-γ-glutamyl-L-cysteinyl-glycine; GSH) in cancer cells is particularly relevant in the regulation of carcinogenic mechanisms; sensitivity against cytotoxic drugs, ionizing radiations, and some cytokines; DNA synthesis; and cell proliferation and death. The intracellular thiol redox state (controlled by GSH) is one of the endogenous effectors involved in regulating the mitochondrial permeability transition pore complex and, in consequence, thiol oxidation can be a causal factor in the mitochondrion-based mechanism that leads to cell death. Nevertheless GSH depletion is a common feature not only of apoptosis but also of other types of cell death. Indeed rates of GSH synthesis and fluxes regulate its levels in cellular compartments, and potentially influence switches among different mechanisms of death. How changes in gene expression, post-translational modifications of proteins, and signaling cascades are implicated will be discussed. Furthermore, this review will finally analyze whether GSH depletion may facilitate cancer cell death under in vivo conditions, and how this can be applied to cancer therapy. PMID:24212662

Expression Patterns of Genes Involved in Ascorbate-Glutathione Cycle in Aphid-Infested Maize (Zea mays L.) Seedlings

PubMed Central

Sytykiewicz, Hubert

2016-01-01

Reduced forms of ascorbate (AsA) and glutathione (GSH) are among the most important non-enzymatic foliar antioxidants in maize (Zea mays L.). The survey was aimed to evaluate impact of bird cherry-oat aphid (Rhopalosiphum padi L.) or grain aphid (Sitobion avenae F.) herbivory on expression of genes related to ascorbate-glutathione (AsA-GSH) cycle in seedlings of six maize varieties (Ambrozja, Nana, Tasty Sweet, Touran, Waza, Złota Karłowa), differing in resistance to the cereal aphids. Relative expression of sixteen maize genes encoding isoenzymes of ascorbate peroxidase (APX1, APX2, APX3, APX4, APX5, APX6, APX7), monodehydroascorbate reductase (MDHAR1, MDHAR2, MDHAR3, MDHAR4), dehydroascorbate reductase (DHAR1, DHAR2, DHAR3) and glutathione reductase (GR1, GR2) was quantified. Furthermore, effect of hemipterans’ attack on activity of APX, MDHAR, DHAR and GR enzymes, and the content of reduced and oxidized ascorbate and glutathione in maize plants were assessed. Seedling leaves of more resistant Z. mays varieties responded higher elevations in abundance of target transcripts. In addition, earlier and stronger aphid-triggered changes in activity of APX, MDHAR, DHAR and GR enzymes, and greater modulations in amount of the analyzed antioxidative metabolites were detected in foliar tissues of highly resistant Ambrozja genotype in relation to susceptible Tasty Sweet plants. PMID:26907270

Expression Patterns of Genes Involved in Ascorbate-Glutathione Cycle in Aphid-Infested Maize (Zea mays L.) Seedlings.

PubMed

Sytykiewicz, Hubert

2016-02-23

Reduced forms of ascorbate (AsA) and glutathione (GSH) are among the most important non-enzymatic foliar antioxidants in maize (Zea mays L.). The survey was aimed to evaluate impact of bird cherry-oat aphid (Rhopalosiphum padi L.) or grain aphid (Sitobion avenae F.) herbivory on expression of genes related to ascorbate-glutathione (AsA-GSH) cycle in seedlings of six maize varieties (Ambrozja, Nana, Tasty Sweet, Touran, Waza, Złota Karłowa), differing in resistance to the cereal aphids. Relative expression of sixteen maize genes encoding isoenzymes of ascorbate peroxidase (APX1, APX2, APX3, APX4, APX5, APX6, APX7), monodehydroascorbate reductase (MDHAR1, MDHAR2, MDHAR3, MDHAR4), dehydroascorbate reductase (DHAR1, DHAR2, DHAR3) and glutathione reductase (GR1, GR2) was quantified. Furthermore, effect of hemipterans' attack on activity of APX, MDHAR, DHAR and GR enzymes, and the content of reduced and oxidized ascorbate and glutathione in maize plants were assessed. Seedling leaves of more resistant Z. mays varieties responded higher elevations in abundance of target transcripts. In addition, earlier and stronger aphid-triggered changes in activity of APX, MDHAR, DHAR and GR enzymes, and greater modulations in amount of the analyzed antioxidative metabolites were detected in foliar tissues of highly resistant Ambrozja genotype in relation to susceptible Tasty Sweet plants.

Release of mitochondrial glutathione and calcium by a cyclosporin A-sensitive mechanism occurs without large amplitude swelling.

PubMed

Savage, M K; Reed, D J

1994-11-15

Treatment of isolated mitochondria with calcium and inorganic phosphate induces inner membrane permeability that is thought to be mediated through a non-selective, calcium-dependent pore. The inner membrane permeability results in the rapid efflux of small matrix solutes such as glutathione and calcium, loss of coupled functions, and large amplitude swelling. We have identified conditions of permeability transition without large amplitude swelling, a parameter often used to assess inner membrane permeability. The addition of either oligomycin, antimycin, or sulfide to incubation buffer containing calcium and inorganic phosphate abolished large-amplitude swelling of mitochondria but did not prevent inner membrane permeability as demonstrated by the release of mitochondrial glutathione and calcium. The release of both glutathione and calcium was inhibited by the addition of cyclosporin A, a potent inhibitor of permeability transition. Transmission electron microscopy analysis, combined with the glutathione and calcium release data, indicate that permeability transition can be observed in the absence of large-amplitude swelling. Permeability transition occurring both with and without large-amplitude swelling was accompanied by a collapse of the membrane potential. We conclude that cyclosporin A-sensitive permeability transition can occur without obvious morphological changes such as large-amplitude swelling. Monitoring the cyclosporin A-sensitive release of concentrated endogenous matrix solutes, such as GSH, may be a sensitive and useful indicator of permeability transition.

Determination of Glutathione, Selenium, and Malondialdehyde in Different Edible Mushroom Species.

PubMed

Dogan, Hacer; Coteli, Ebru; Karatas, Fikret

2016-12-01

In this study, the amount of reduced glutathione (GSH), oxidized glutathione (GSSG), and malondialdehyde (MDA) were determined by high performance liquid chromatography (HPLC), and selenium was determined by using the fluorescence spectrophotometer in eight different species of edible mushrooms. Brittlegill mushroom (Russula delica), meadow mushroom (Agaricus campestris), dryad's saddle mushroom (Polyporus squamosus), white button mushroom (Agaricus bisporus), Pleurotus spp., ink mushroom (Coprinus atramentarius), ebekari mushroom (slimy) (Elazığ local) and çaşır mushroom (Pleurotus eryngii) (Tunceli local) were used for analysis. The amounts of GSH, GSSG, Se, and MDA with GSH/GSSG ratio in the eight different species of edible mushrooms were observed in between 269.10 ± 16.94-1554.83 ± 58.12 μg/g; 23.55 ± 1.89-841.90 ± 20.03 μg/g; 15.06 ± 1.56-82.10 ± 3.84 μg/g; 5.46 ± 0.50-27.45 ± 2.58 μg/g wet weight and 0.32-41.35, respectively. There is a weak correlation (R 2  = 0.389) between MDA and Se, on the other hand, the correlation (R 2  = 0.831) between GSH/GSSG ratio and selenium in mushrooms are reasonable well. In a similar manner, there is a weak correlation (R 2  = 0551) between GSH/GSSG and MDA ratios in mushrooms. It was found that these edible mushroom species are good source of glutathione (GSH, GSSG), and selenium (Se) in terms of quantities obtained; therefore, it can be said that mushrooms are a rich source of antioxidants.

Characterization of glutathione transferases involved in the pathogenicity of Alternaria brassicicola.

PubMed

Calmes, Benoit; Morel-Rouhier, Mélanie; Bataillé-Simoneau, Nelly; Gelhaye, Eric; Guillemette, Thomas; Simoneau, Philippe

2015-06-18

Glutathione transferases (GSTs) represent an extended family of multifunctional proteins involved in detoxification processes and tolerance to oxidative stress. We thus anticipated that some GSTs could play an essential role in the protection of fungal necrotrophs against plant-derived toxic metabolites and reactive oxygen species that accumulate at the host-pathogen interface during infection. Mining the genome of the necrotrophic Brassica pathogen Alternaria brassicicola for glutathione transferase revealed 23 sequences, 17 of which could be clustered into the main classes previously defined for fungal GSTs and six were 'orphans'. Five isothiocyanate-inducible GSTs from five different classes were more thoroughly investigated. Analysis of their catalytic properties revealed that two GSTs, belonging to the GSTFuA and GTT1 classes, exhibited GSH transferase activity with isothiocyanates (ITC) and peroxidase activity with cumene hydroperoxide, respectively. Mutant deficient for these two GSTs were however neither more susceptible to ITC nor less aggressive than the wild-type parental strain. By contrast mutants deficient for two other GSTs, belonging to the Ure2pB and GSTO classes, were distinguished by their hyper-susceptibility to ITC and low aggressiveness against Brassica oleracea. In particular AbGSTO1 could participate in cell tolerance to ITC due to its glutathione-dependent thioltransferase activity. The fifth ITC-inducible GST belonged to the MAPEG class and although it was not possible to produce the soluble active form of this protein in a bacterial expression system, the corresponding deficient mutant failed to develop normal symptoms on host plant tissues. Among the five ITC-inducible GSTs analyzed in this study, three were found essential for full aggressiveness of A. brassicicola on host plant. This, to our knowledge is the first evidence that GSTs might be essential virulence factors for fungal necrotrophs.

Avocado oil induces long-term alleviation of oxidative damage in kidney mitochondria from type 2 diabetic rats by improving glutathione status.

PubMed

Ortiz-Avila, Omar; Figueroa-García, María Del Consuelo; García-Berumen, Claudia Isabel; Calderón-Cortés, Elizabeth; Mejía-Barajas, Jorge A; Rodriguez-Orozco, Alain R; Mejía-Zepeda, Ricardo; Saavedra-Molina, Alfredo; Cortés-Rojo, Christian

2017-04-01

Hyperglycemia and mitochondrial ROS overproduction have been identified as key factors involved in the development of diabetic nephropathy. This has encouraged the search for strategies decreasing glucose levels and long-term improvement of redox status of glutathione, the main antioxidant counteracting mitochondrial damage. Previously, we have shown that avocado oil improves redox status of glutathione in liver and brain mitochondria from streptozotocin-induced diabetic rats; however, the long-term effects of avocado oil and its hypoglycemic effect cannot be evaluated because this model displays low survival and insulin depletion. Therefore, we tested during 1 year the effects of avocado oil on glycemia, ROS levels, lipid peroxidation and glutathione status in kidney mitochondria from type 2 diabetic Goto-Kakizaki rats. Diabetic rats exhibited glycemia of 120-186 mg/dL the first 9 months with a further increase to 250-300 mg/dL. Avocado oil decreased hyperglycemia at intermediate levels between diabetic and control rats. Diabetic rats displayed augmented lipid peroxidation and depletion of reduced glutathione throughout the study, while increased ROS generation was observed at the 3rd and 12th months along with diminished content of total glutathione at the 6th and 12th months. Avocado oil ameliorated all these defects and augmented the mitochondrial content of oleic acid. The beneficial effects of avocado oil are discussed in terms of the hypoglycemic effect of oleic acid and the probable dependence of glutathione transport on lipid peroxidation and thiol oxidation of mitochondrial carriers.

S-Glutathionyl-(chloro)hydroquinone reductases: a novel class of glutathione transferases

PubMed Central

XUN, Luying; BELCHIK, Sara M.; XUN, Randy; HUANG, Yan; ZHOU, Huina; SANCHEZ, Emiliano; KANG, ChulHee; BOARD, Philip G.

2010-01-01

Sphingobium chlorophenolicum completely mineralizes PCP (pentachlorophenol). Two GSTs (glutathione transferases), PcpC and PcpF, are involved in the degradation. PcpC uses GSH to reduce TeCH (tetrachloro-p-hydroquinone) to TriCH (trichloro-p-hydroquinone) and then to DiCH (dichloro-p-hydroquinone) during PCP degradation. However, oxidatively damaged PcpC produces GS-TriCH (S-glutathionyl-TriCH) and GS-DiCH (S-glutathionyl-TriCH) conjugates. PcpF converts the conjugates into TriCH and DiCH, re-entering the degradation pathway. PcpF was further characterized in the present study. It catalysed GSH-dependent reduction of GS-TriCH via a Ping Pong mechanism. First, PcpF reacted with GS-TriCH to release TriCH and formed disulfide bond between its Cys53 residue and the GS moiety. Then, a GSH came in to regenerate PcpF and release GS–SG. A TBLASTN search revealed that PcpF homologues were widely distributed in bacteria, halobacteria (archaea), fungi and plants, and they belonged to ECM4 (extracellular mutant 4) group COG0435 in the conserved domain database. Phylogenetic analysis grouped PcpF and homologues into a distinct group, separated from Omega class GSTs. The two groups shared conserved amino acid residues, for GSH binding, but had different residues for the binding of the second substrate. Several recombinant PcpF homologues and two human Omega class GSTs were produced in Escherichia coli and purified. They had zero or low activities for transferring GSH to standard substrates, but all had reasonable activities for GSH-dependent reduction of disulfide bond (thiol transfer), dehydroascorbate and dimethylarsinate. All the tested PcpF homologues reduced GS-TriCH, but the two Omega class GSTs did not. Thus PcpF homologues were tentatively named S-glutathionyl-(chloro)hydroquinone reductases for catalysing the GSH-dependent reduction of GS-TriCH. PMID:20388120

Exogenous salicylic acid improves photosynthesis and growth through increase in ascorbate-glutathione metabolism and S assimilation in mustard under salt stress

PubMed Central

Nazar, Rahat; Umar, Shahid; Khan, Nafees A.

2015-01-01

Ascorbate (AsA)–glutathione (GSH) cycle metabolism has been regarded as the most important defense mechanism for the resistance of plants under stress. In this study the influence of salicylic acid (SA) was studied on ascorbate-glutathione pathway, S-assimilation, photosynthesis and growth of mustard (Brassica juncea L.) plants subjected to 100 mM NaCl. Treatment of SA (0.5 mM) alleviated the negative effects of salt stress and improved photosynthesis and growth through increase in enzymes of ascorbate-glutathione pathway which suggest that SA may participate in the redox balance under salt stress. The increase in leaf sulfur content through higher activity of ATP sulfurylase (ATPS) and serine acetyl transferase (SAT) by SA application was associated with the increased accumulation of glutathione (GSH) and lower levels of oxidative stress. These effects of SA were substantiated by the findings that application of SA-analog, 2,6, dichloro-isonicotinic acid (INA) and 1 mM GSH treatment produced similar results on rubisco, photosynthesis and growth of plants establishing that SA application alleviates the salt-induced decrease in photosynthesis mainly through inducing the enzyme activity of ascorbate-glutathione pathway and increased GSH production. Thus, SA/GSH could be a promising tool for alleviation of salt stress in mustard plants. PMID:25730495

Modeling Fate and Transport of Fecal Coliform Bacteria Using SWAT 2005 (Case Study: Jajrood River Watershed, Iran)

NASA Astrophysics Data System (ADS)

Maghrebi, M.; Tajrishy, M.

2010-12-01

Jajrood River watershed is one of the main drinking water resources of the capital city of Tehran, Iran. In addition it has been available as many recreational usages especially in the warm months. As a result of being located near one of the crowded cities of the world, a variety of microbial pollutions is commonly perceived in the Jajrood River. Among them, there are strong concerns about fecal coliform bacteria concentration. This article aimed to model fate and transport of fecal coliform bacteria in Jajrood River watershed using Soil and Water Assessment Tool (SWAT) model version 2005. Potential pollutant sources in the study area were detected and quantified for modeling purposes. In spite of being lack of knowledge about bacteria die-off rate in small river bodies, as well as in other watershed-based forms, fecal coliform bacteria die-off rates were estimated using both laboratory and field data investigations with some simplifications. The SWAT model was calibrated over an extended time period (1997-2002) for this watershed. The river flow calibrated using SUFI-2 software and resulted in a very good outputs (R2=0.82, E=0.81). Furthermore SWAT model was validated over January 2003 to September 2005 in the study area and has resulted in good outputs (R2=0.61, E=0.57). This research illustrates SWAT 2005 capability to model fecal coliform bacteria in a populated watershed, and deals with most of watershed microbial pollution sources that are usually observed in developing countries. Fecal coliform concentration simulation results were mostly in the same order in comparison with real data. However, Differences were judged to be related to lack of input data. In this article different aspects of SWAT capabilities for modeling of fecal coliform bacteria concentration will be reviewed and it will present new insights in bacteria modeling procedures especially for mountainous, high populated and small sized watersheds.

Glutathione biosynthesis plays an important role against 4-tert-octylphenol-induced oxidative stress in Ceratophyllum demersum.

PubMed

Cahyanurani, Annisa' Bias; Chiu, Kuo-Hsun; Wu, Tsung-Meng

2017-09-01

4-tert-octylphenol (OP) is a persistent environmental pollutant with an endocrine-disrupting property. In the present study, we examined the effect of various concentrations of OP (0, 0.5, 1, 1.5, 2 and 3 mg L -1 ) applied to an aquatic plant, the submersed macrophyte Ceratophyllum demersum. The toxic effect caused by OP inhibited the plant's growth rate, reduced total chlorophyll content and increased levels of the reactive oxygen species (ROS) O 2 •- and H 2 O 2 . OP treatment significantly increased the activities of antioxidant enzymes including superoxide dismutase, guaiacol peroxidase, glutathione reductase and ascorbate peroxidase. The contents of the non-enzymatic antioxidant glutathione (GSH) and ratio of GSH to glutathione disulfide were markedly increased with OP treatment. Pretreatment with buthionine sulfoximine, a specific and potent inhibitor of GSH biosynthesis, significantly reduced total GSH content and conferred a more severe toxic phenotype on OP exposure. Thus, with OP-induced oxidative stress, C. demersum might actively regulate the antioxidant machinery, especially the biosynthesis and redox state of GSH. Copyright © 2017 Elsevier Ltd. All rights reserved.

Global deletion of glutathione S-Transferase A4 exacerbates developmental nonalcoholic steatohepatitis

USDA-ARS?s Scientific Manuscript database

We established a mouse model of developmental nonalcoholic steatohepatitis (NASH) by feeding a high polyunsaturated fat liquid diet to female glutathione-S-transferase 4-4 (Gsta4-/-)/peroxisome proliferator activated receptor a (Ppara-/-) double knockout 129/SvJ mice for 12 weeks from weaning. We us...

The chemistry of the S-nitrosoglutathione/glutathione system

PubMed Central

Singh, S. P.; Wishnok, J. S.; Keshive, M.; Deen, W. M.; Tannenbaum, S. R.

1996-01-01

S-Nitrosothiols have generated considerable interest due to their ability to act as nitric oxide (NO) donors and due to their possible involvement in bioregulatory systems—e.g., NO transfer reactions. Elucidation of the reaction pathways involved in the modification of the thiol group by S-nitrosothiols is important for understanding the role of S-nitroso compounds in vivo. The modification of glutathione (GSH) in the presence of S-nitrosoglutathione (GSNO) was examined as a model reaction. Incubation of GSNO (1 mM) with GSH at various concentrations (1–10 mM) in phosphate buffer (pH 7.4) yielded oxidized glutathione, nitrite, nitrous oxide, and ammonia as end products. The product yields were dependent on the concentrations of GSH and oxygen. Transient signals corresponding to GSH conjugates, which increased by one mass unit when the reaction was carried out with 15N-labeled GSNO, were identified by electrospray ionization mass spectrometry. When morpholine was present in the reaction system, N-nitrosomorpholine was formed. Increasing concentrations of either phosphate or GSH led to lower yields of N-nitrosomorpholine. The inhibitory effect of phosphate may be due to reaction with the nitrosating agent, nitrous anhydride (N2O3), formed by oxidation of NO. This supports the release of NO during the reaction of GSNO with GSH. The products noted above account quantitatively for virtually all of the GSNO nitrogen consumed during the reaction, and it is now possible to construct a complete set of pathways for the complex transformations arising from GSNO + GSH. PMID:8962068

Apathy in aging: are lack of interest and lack of initiative dissociable?

PubMed

Esposito, Fabienne; Rochat, Lucien; Juillerat Van der Linden, Anne-Claude; Lekeu, Françoise; Charnallet, Annik; Van der Linden, Martial

2014-01-01

Apathy is common in aging and generally defined on the basis of three dimensions: lack of initiative, lack of interest and emotional blunting. Curiously, no study until now has examined the associations and dissociations between these dimensions in elderly people (with or without dementia). These questions were addressed in two studies. In the first study, we explored the distribution of scores and the relationships between the three dimensions of apathy in 56 patients with dementia, focusing mainly on lack of initiative and lack of interest. Apathy was hetero-evaluated with the Apathy Inventory (AI), a scale widely used to assess the apathy dimensions in aging. In the second study, given the AI's limitations, we investigated in more detail the relationship between lack of initiative and interest in 115 elderly people using a new questionnaire specifically designed to assess these two dimensions. Results showed that lack of initiative was closely related to lack of interest (Study 1). Although we used a more specific questionnaire, these facets of apathy did not constitute two separable dimensions, but reflected a common main factor of apathy in aging (Study 2). Thus, the distinction between lack of initiative and lack of interest seems questionable. Only a multifactorial approach that includes the various psychological factors involved in apathy would enable one to gain a better understanding of the different manifestations of apathy and to highlight possible dissociations between them. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Secreted CLIC3 drives cancer progression through its glutathione-dependent oxidoreductase activity

PubMed Central

Hernandez-Fernaud, Juan R.; Ruengeler, Elena; Casazza, Andrea; Neilson, Lisa J.; Pulleine, Ellie; Santi, Alice; Ismail, Shehab; Lilla, Sergio; Dhayade, Sandeep; MacPherson, Iain R.; McNeish, Iain; Ennis, Darren; Ali, Hala; Kugeratski, Fernanda G.; Al Khamici, Heba; van den Biggelaar, Maartje; van den Berghe, Peter V.E.; Cloix, Catherine; McDonald, Laura; Millan, David; Hoyle, Aoisha; Kuchnio, Anna; Carmeliet, Peter; Valenzuela, Stella M.; Blyth, Karen; Yin, Huabing; Mazzone, Massimiliano; Norman, Jim C.; Zanivan, Sara

2017-01-01

The secretome of cancer and stromal cells generates a microenvironment that contributes to tumour cell invasion and angiogenesis. Here we compare the secretome of human mammary normal and cancer-associated fibroblasts (CAFs). We discover that the chloride intracellular channel protein 3 (CLIC3) is an abundant component of the CAF secretome. Secreted CLIC3 promotes invasive behaviour of endothelial cells to drive angiogenesis and increases invasiveness of cancer cells both in vivo and in 3D cell culture models, and this requires active transglutaminase-2 (TGM2). CLIC3 acts as a glutathione-dependent oxidoreductase that reduces TGM2 and regulates TGM2 binding to its cofactors. Finally, CLIC3 is also secreted by cancer cells, is abundant in the stromal and tumour compartments of aggressive ovarian cancers and its levels correlate with poor clinical outcome. This work reveals a previously undescribed invasive mechanism whereby the secretion of a glutathione-dependent oxidoreductase drives angiogenesis and cancer progression by promoting TGM2-dependent invasion. PMID:28198360

Connexin hemichannels mediate glutathione transport and protect lens fiber cells from oxidative stress.

PubMed

Shi, Wen; Riquelme, Manuel A; Gu, Sumin; Jiang, Jean X

2018-03-21

Elevated oxidized stress contributes to lens cataracts, and gap junctions play important roles in maintaining lens transparency. As well as forming gap junctions, connexin (Cx) proteins also form hemichannels. Here, we report a new mechanism whereby hemichannels mediate transport of reductant glutathione into lens fiber cells and protect cells against oxidative stress. We found that Cx50 (also known as GJA8) hemichannels opened in response to H 2 O 2 in lens fiber cells but that transport through the channels was inhibited by two dominant-negative mutants in Cx50, Cx50P88S, which inhibits transport through both gap junctions and hemichannels, and Cx50H156N, which only inhibits transport through hemichannels and not gap junctions. Treatment with H 2 O 2 increased the number of fiber cells undergoing apoptosis, and this increase was augmented with dominant-negative mutants that disrupted both hemichannels formed from Cx46 (also known as GJA3) and Cx50, while Cx50E48K, which only impairs gap junctions, did not have such an effect. Moreover, hemichannels mediate uptake of glutathione, and this uptake protected lens fiber cells against oxidative stress, while hemichannels with impaired transport had less protective benefit from glutathione. Taken together, these results show that oxidative stress activates connexin hemichannels in the lens fiber cells and that hemichannels likely protect lens cell against oxidative damage through transporting extracellular reductants. © 2018. Published by The Company of Biologists Ltd.

Plants as sources of airborne bacteria, including ice nucleation-active bacteria.

PubMed

Lindemann, J; Constantinidou, H A; Barchet, W R; Upper, C D

1982-11-01

Vertical wind shear and concentration gradients of viable, airborne bacteria were used to calculate the upward flux of viable cells above bare soil and canopies of several crops. Concentrations at soil or canopy height varied from 46 colony-forming units per m over young corn and wet soil to 663 colony-forming units per m over dry soil and 6,500 colony-forming units per m over a closed wheat canopy. In simultaneous samples, concentrations of viable bacteria in the air 10 m inside an alfalfa field were fourfold higher than those over a field with dry, bare soil immediately upwind. The upward flux of viable bacteria over alfalfa was three- to fourfold greater than over dry soil. Concentrations of ice nucleation-active bacteria were higher over plants than over soil. Thus, plant canopies may constitute a major source of bacteria, including ice nucleation-active bacteria, in the air.

Glutathione-Capped Gold Nanoparticles-Based Photoacoustic Sensor for Label-Free Detection of Lead Ions

NASA Astrophysics Data System (ADS)

Shi, R.; Liu, X.-J.; Ying, Y.

2017-07-01

The photoacoustic signal generated by laser-induced nanobubbles (PA-LINB) proved to be a sensitive tool to monitor the aggregation of gold nanoparticles. Here, a simple and label-free photoacoustic method for the rapid detection of Pb2+ in the aqueous phase was developed. Due to the high affinity of Pb2+ ions to glutathione, the presence of Pb2+ led to the aggregation of glutathione-conjugated gold nanoparticles (GSH-GNPs). Hence, by measuring the variation of the PA-LINB signal after the aggregation of GSH-GNPs, Pb2+ can be quantified. A low detection limit for Pb2+ (42 nM) and a wide linear working range ( 42-1000 nM) were achieved. Furthermore, the proposed method showed good selectivity against other metal ions.

Arabidopsis dehydroascorbate reductase 1 and 2 modulate redox states of ascorbate-glutathione cycle in the cytosol in response to photooxidative stress.

PubMed

Noshi, Masahiro; Yamada, Hiroki; Hatanaka, Risa; Tanabe, Noriaki; Tamoi, Masahiro; Shigeoka, Shigeru

2017-03-01

Ascorbate and glutathione are indispensable cellular redox buffers and allow plants to acclimate stressful conditions. Arabidopsis contains three functional dehydroascorbate reductases (DHAR1-3), which catalyzes the conversion of dehydroascorbate into its reduced form using glutathione as a reductant. We herein attempted to elucidate the physiological role in DHAR1 and DHAR2 in stress responses. The total DHAR activities in DHAR knockout Arabidopsis plants, dhar1 and dhar2, were 22 and 92%, respectively, that in wild-type leaves. Under high light (HL), the levels of total ascorbate and dehydroascorbate were only reduced and increased, respectively, in dhar1. The oxidation of glutathione under HL was significantly inhibited in both dhar1 and dhar2, while glutathione contents were only enhanced in dhar1. The dhar1 showed stronger visible symptoms than the dhar2 under photooxidative stress conditions. Our results demonstrated a pivotal role of DHAR1 in the modulation of cellular redox states under photooxidative stress.

Immunoreactivities for glutathione S-transferases and glutathione peroxidase in the lateral wall of pigmented and albino guinea pig cochlea.

PubMed

Fujimura, Takeyuki; Suzuki, Hideaki; Udaka, Tsuyoshi; Shiomori, Teruo; Mori, Takanori; Inaba, Tsuyoshi; Hiraki, Nobuaki; Kayashima, Kotaro; Doi, Yoshiaki

2008-09-01

Dark-skinned people are known to be more tolerant of ototraumatic noise than are light-skinned people, and pigmented animals are more tolerant of ototraumatic noise and aminoglycoside ototoxicity than are albino animals. Such tolerance may be dependent on the local ability of detoxification and antioxidant enzymes, including glutathione S-transferase (GST) and glutathione peroxidase (GSPx). In the present study, we examined the difference in GST/GSPx expression in the lateral wall of the cochlea between pigmented and albino guinea pigs. Eight-week-old male pigmented and albino guinea pigs were killed by transcardiac perfusion with 2% paraformaldehyde. The cochlear ducts were isolated, further fixed with 4% paraformaldehyde, decalcified, and then embedded in paraffin. Sections prepared at 5-microm thickness were incubated with anti-GST-alpha,-mu,-pi, or anti-GSPx antibody, reacted with Alexa Fluorconjugated secondary antibody, and examined under a Carl Zeiss Axioskop 2 plus fluorescence microscope. The cochlea ducts were also subjected to immunoelectron microscopy for GST-pi by the postembedment method. The stria vascularis of pigmented guinea pigs was strongly immunoreactive for GST-alpha,-mu,-pi, and GSPx, whereas no or only weak immunoreactivities were seen in the stria vascularis of albino guinea pigs. The spiral ligament showed positive but different immunoreactivities for these enzymes between the strains. Double-stained immunofluorescence micrographs for GST-pi and GSPx showed a close resemblance of localization between the two enzymes in both pigmented and albino guinea pigs. At the ultrastructural level, immunoreactivity for GST-pi was localized preferentially in the melanin cells of pigmented guinea pigs. These results suggest that correlation between pigmentation and inner ear susceptibility is, at least partially, attributed to the different distribution of GST/GSPx in the stria vascularis.

Pharmacogenetics of azathioprine in inflammatory bowel disease: A role for glutathione-S-transferase?

PubMed Central

Stocco, Gabriele; Pelin, Marco; Franca, Raffaella; De Iudicibus, Sara; Cuzzoni, Eva; Favretto, Diego; Martelossi, Stefano; Ventura, Alessandro; Decorti, Giuliana

2014-01-01

Azathioprine is a purine antimetabolite drug commonly used to treat inflammatory bowel disease (IBD). In vivo it is active after reaction with reduced glutathione (GSH) and conversion to mercaptopurine. Although this reaction may occur spontaneously, the presence of isoforms M and A of the enzyme glutathione-S-transferase (GST) may increase its speed. Indeed, in pediatric patients with IBD, deletion of GST-M1, which determines reduced enzymatic activity, was recently associated with reduced sensitivity to azathioprine and reduced production of azathioprine active metabolites. In addition to increase the activation of azathioprine to mercaptopurine, GSTs may contribute to azathioprine effects even by modulating GSH consumption, oxidative stress and apoptosis. Therefore, genetic polymorphisms in genes for GSTs may be useful to predict response to azathioprine even if more in vitro and clinical validation studies are needed. PMID:24707136

Magnetically separable nanoferrite-anchored glutathione: Aqueous homocoupling of arylboronic acids under microwave irradiation

EPA Science Inventory

A highly active, stable and magnetically separable glutathione based organocatalyst provided good to excellent yields to symmetric biaryls in the homocoupling of arylboronic acids under microwave irradiation. Symmetrical biaryl motifs are present in a wide range of natural p...

Determination of intracellular glutathione and cysteine using HPLC with a monolithic column after derivatization with monobromobimane.

PubMed

Conlan, Xavier A; Stupka, Nicole; McDermott, Geoffrey P; Francis, Paul S; Barnett, Neil W

2010-05-01

An optimized high-performance liquid chromatography (HPLC) method is used to show that, as myoblasts differentiate into multinucleated muscle fibers, there is a shift to a more oxidized cell redox state. The HPLC method incorporated derivatization with monobromobimane for the determination of the reduced (GSH) and oxidized (GSSG) forms of glutathione and the reduced (Cys) and oxidized (CysSS) forms of cysteine. The derivatization was optimized to improve the sensitivity of the approach; the limits of detection for glutathione and cysteine were 3 x 10(-8) and 5 x 10(-8) M, respectively. 2009 John Wiley & Sons, Ltd.

Tyrosine 8 contributes to catalysis but is not required for activity of rat liver glutathione S-transferase, 1-1.

PubMed Central

Wang, J.; Barycki, J. J.; Colman, R. F.

1996-01-01

Reaction of rat liver glutathione S-transferase, isozyme 1-1, with 4-(fluorosulfonyl)benzoic acid (4-FSB), a xenobiotic substrate analogue, results in a time-dependent inactivation of the enzyme to a final value of 35% of its original activity when assayed at pH 6.5 with 1-chloro-2,4-dinitrobenzene (CDNB) as substrate. The rate of inactivation exhibits a nonlinear dependence on the concentration of 4-FSB from 0.25 mM to 9 mM, characterized by a KI of 0.78 mM and kmax of 0.011 min-1. S-Hexylglutathione or the xenobiotic substrate analogue, 2,4-dinitrophenol, protects against inactivation of the enzyme by 4-FSB, whereas S-methylglutathione has little effect on the reaction. These experiments indicate that reaction occurs within the active site of the enzyme, probably in the binding site of the xenobiotic substrate, close to the glutathione binding site. Incorporation of [3,5-3H]-4-FSB into the enzyme in the absence and presence of S-hexylglutathione suggests that modification of one residue is responsible for the partial loss of enzyme activity. Tyr 8 and Cys 17 are shown to be the reaction targets of 4-FSB, but only Tyr 8 is protected against 4-FSB by S-hexylglutathione. DTT regenerates cysteine from the reaction product of cysteine and 4-FSB, but does not reactivate the enzyme. These results show that modification of Tyr 8 by 4-FSB causes the partial inactivation of the enzyme. The Michaelis constants for various substrates are not changed by the modification of the enzyme. The pH dependence of the enzyme-catalyzed reaction of glutathione with CDNB for the modified enzyme, as compared with the native enzyme, reveals an increase of about 0.9 in the apparent pKa, which has been interpreted as representing the ionization of enzyme-bound glutathione; however, this pKa of about 7.4 for modified enzyme remains far below the pK of 9.1 for the -SH of free glutathione. Previously, it was considered that Tyr 8 was essential for GST catalysis. In contrast, we conclude that

Association of mercury and selenium with altered glutathione metabolism and oxidative stress in diving ducks from the San Francisco Bay region

USGS Publications Warehouse

Hoffman, D.J.; Ohlendorf, H.M.; Marn, C.M.; Pendleton, G.W.

1998-01-01

Adult male greater scaup (Aythya marila) (GS), surf scoters (Melanitta perspicillata)(SS), and ruddy ducks (Oxyura jamaicensis) (RD) were collected from Suisun Bay and coastal Tomales Bay in the greater San Francisco Bay area to assess exposure to inorganic contaminants. Hepatic selenium (Se) concentrations were highest in GS (geometric mean = 67 ppm, dw) and SS (119 ppm) in Suisun Bay, whereas hepatic mercury (Hg) was highest (19 ppm) in GS and SS from Tomales Bay. Hepatic Se and Hg were lower in RD and did not differ between locations. Hepatic supernatants were assayed for enzymes related to glutathione metabolism and antioxidant activity including: glucose-6-phosphate dehydrogenase (G-6-PDH), glutathione peroxidase (GSH-peroxidase), glutathione reductase (GSSG-reductase), and glutathione-S-transferase (GSH-transferase). GSH-peroxidase activity was higher in SS and RD, and G-6-PDH higher in GS and SS from Suisun Bay than Tomales Bay. GSSG-reductase was higher in SS from Suisun Bay. The ratio of oxidized glutathione (GSSG) to reduced glutathione (GSH) was greater in all species from Tomales Bay. The following significant relationships were found in one or more species with increasing hepatic Hg concentration: lower body, liver and heart weights; decreased hepatic GSH concentration, G-6-PDH and GSH-peroxidase activities; increased ratio of GSSG to GSH, and increased GSSG-reductase activity. With increasing hepatic Se concentration, GSH-peroxidase increased but GSH decreased. It is concluded that measurement of associated enzymes in conjunction with thiol status may be a useful bioindicator to discriminate between Hg and Se effects. Concentrations of mercury and selenium and variable affected have been associated with adverse effects on reproduction and neurological function in experimental studies with mallards.

Neural Tube Defects and Maternal Biomarkers of Folate, Homocysteine, and Glutathione Metabolism

PubMed Central

Zhao, Weizhi; Mosley, Bridget S.; Cleves, Mario A.; Melnyk, Stepan; James, S. Jill; Hobbs, Charlotte A.

2010-01-01

Background Alterations in maternal folate and homocysteine metabolism are associated with neural tube defects (NTDs). The role that specific micronutrients and metabolites play in the causal pathway leading to NTDs is not fully understood. Methods We conducted a case-control study to investigate the association between NTDs and maternal alterations in plasma micronutrients and metabolites in two metabolic pathways, the methionine remethylation and glutathione transsulfuration. Biomarkers were measured in a population-based sample of women who had NTD-affected pregnancies (n = 43) and a control group of women who had a pregnancy unaffected by a birth defect (n = 160). Plasma concentrations of folate, Vitamin B12, Vitamin B6, methionine, S-adenosylmethionine (SAM), s- adenosylhomocysteine (SAH), adenosine, homocysteine, cysteine, and reduced and oxidized glutathione were compared between cases and controls after adjusting for lifestyle and sociodemographic factors. Results Women with NTD-affected pregnancies had significantly higher plasma concentrations of SAH (29.12 vs. 23.13 nmol/L, P = 0.0011), adenosine (0.323 vs. 0.255 μmol/L, P = 0.0269), homocysteine (9.40 vs. 7.56 μmol/L, P < 0.001), and oxidized glutathione (0.379 vs. 0.262μmol/L, P = 0.0001), but lower plasma SAM concentration (78.99 vs. 83.16 nmol/L, P = 0.0172) than controls. This metabolic profile is consistent with reduced methylation capacity and increased oxidative stress in women with affected pregnancies. Conclusions Increased maternal oxidative stress and decreased methylation capacity may contribute to the occurrence of NTDs. Further analysis of relevant genetic and environmental factors is required to define the basis for these observed alterations. PMID:16575882

Inherited glutathione-S-transferase deficiency is a risk factor for pulmonary asbestosis.

PubMed

Smith, C M; Kelsey, K T; Wiencke, J K; Leyden, K; Levin, S; Christiani, D C

1994-09-01

Pulmonary diseases attributable to asbestos exposure constitute a significant public health burden, yet few studies have investigated potential genetic determinants of susceptibility to asbestos-related diseases. The glutathione-S-transferases are a family of conjugating enzymes that both catalyze the detoxification of a variety of potentially cytotoxic electrophilic agents and act in the generation of sulfadipeptide leukotriene inflammatory mediators. The gene encoding glutathione-S-transferase class mu (GSTM-1) is polymorphic; approximately 50% of Caucasian individuals have a homozygous deletion of this gene and do not produce functional enzyme. Glutathione-S-transferase mu (GST-mu) deficiency has been previously reported to be associated with smoking-induced lung cancer. We conducted a cross-sectional study to examine the prevalence of the homozygous deletion for the GSTM-1 gene in members of the carpentry trade occupationally exposed to asbestos. Members of the United Brotherhood of Carpenters and Joiners of America attending their 1991 National Union conference were invited to participate. Each participant was offered a chest X-ray and was asked to complete a comprehensive questionnaire and have their blood drawn. All radiographs were assessed for the presence of pneumoconiosis in a blinded fashion by a National Institute for Occupational Safety and Health-certified International Labor Office "B" reader. Individual GSTM-1 status was determined using polymerase chain reaction methods. Six hundred fifty-eight workers were studied. Of these, 80 (12.2%) had X-ray abnormalities associated with asbestos exposure. Individuals genetically deficient in GST-mu were significantly more likely to have radiographic evidence of nonmalignant asbestos-related disease than those who were not deficient (chi 2 = 5.0; P < 0.03).(ABSTRACT TRUNCATED AT 250 WORDS)

Enzymatic improvement of mitochondrial thiol oxidase Erv1 for oxidized glutathione fermentation by Saccharomyces cerevisiae.

PubMed

Kobayashi, Jyumpei; Sasaki, Daisuke; Hara, Kiyotaka Y; Hasunuma, Tomohisa; Kondo, Akihiko

2017-03-15

Oxidized glutathione (GSSG) is the preferred form for industrial mass production of glutathione due to its high stability compared with reduced glutathione (GSH). In our previous study, over-expression of the mitochondrial thiol oxidase ERV1 gene was the most effective for high GSSG production in Saccharomyces cerevisiae cells among three types of different thiol oxidase genes. We improved Erv1 enzyme activity for oxidation of GSH and revealed that S32 and N34 residues are critical for the oxidation. Five engineered Erv1 variant proteins containing S32 and/or N34 replacements exhibited 1.7- to 2.4-fold higher in vitro GSH oxidation activity than that of parental Erv1, whereas the oxidation activities of these variants for γ-glutamylcysteine were comparable. According to three-dimensional structures of Erv1 and protein stability assays, S32 and N34 residues interact with nearby residues through hydrogen bonding and greatly contribute to protein stability. These results suggest that increased flexibility by amino acid replacements around the active center decrease inhibitory effects on GSH oxidation. Over-expressions of mutant genes coding these Erv1 variants also increased GSSG and consequently total glutathione production in S. cerevisiae cells. Over-expression of the ERV1 S32A gene was the most effective for GSSG production in S. cerevisiae cells among the parent and other mutant genes, and it increased GSSG production about 1.5-fold compared to that of the parental ERV1 gene. This is the first study demonstrating the pivotal effects of S32 and N34 residues to high GSH oxidation activity of Erv1. Furthermore, in vivo validity of Erv1 variants containing these S32 and N34 replacements were also demonstrated. This study indicates potentials of Erv1 for high GSSG production.

Glutathione regulation of redox-sensitive signals in tumor necrosis factor-{alpha}-induced vascular endothelial dysfunction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tsou, T.-C.; Yeh, S.C.; Tsai, F.-Y.

2007-06-01

We investigated the regulatory role of glutathione in tumor necrosis factor-alpha (TNF-{alpha})-induced vascular endothelial dysfunction as evaluated by using vascular endothelial adhesion molecule expression and monocyte-endothelial monolayer binding. Since TNF-{alpha} induces various biological effects on vascular cells, TNF-{alpha} dosage could be a determinant factor directing vascular cells into different biological fates. Based on the adhesion molecule expression patterns responding to different TNF-{alpha} concentrations, we adopted the lower TNF-{alpha} (0.2 ng/ml) to rule out the possible involvement of other TNF-{alpha}-induced biological effects. Inhibition of glutathione synthesis by L-buthionine-(S,R)-sulfoximine (BSO) resulted in down-regulations of the TNF-{alpha}-induced adhesion molecule expression and monocyte-endothelial monolayermore » binding. BSO attenuated the TNF-{alpha}-induced nuclear factor-kappaB (NF-{kappa}B) activation, however, with no detectable effect on AP-1 and its related mitogen-activated protein kinases (MAPKs). Deletion of an AP-1 binding site in intercellular adhesion molecule-1 (ICAM-1) promoter totally abolished its constitutive promoter activity and its responsiveness to TNF-{alpha}. Inhibition of ERK, JNK, or NF-{kappa}B attenuates TNF-{alpha}-induced ICAM-1 promoter activation and monocyte-endothelial monolayer binding. Our study indicates that TNF-{alpha} induces adhesion molecule expression and monocyte-endothelial monolayer binding mainly via activation of NF-{kappa}B in a glutathione-sensitive manner. We also demonstrated that intracellular glutathione does not modulate the activation of MAPKs and/or their downstream AP-1 induced by lower TNF-{alpha}. Although AP-1 activation by the lower TNF-{alpha} was not detected in our systems, we could not rule out the possible involvement of transiently activated MAPKs/AP-1 in the regulation of TNF-{alpha}-induced adhesion molecule expression.« less

Glutathione S-transferase P1 (GSTP1) directly influences platinum drug chemosensitivity in ovarian tumour cell lines.

PubMed

Sawers, L; Ferguson, M J; Ihrig, B R; Young, H C; Chakravarty, P; Wolf, C R; Smith, G

2014-09-09

Chemotherapy response in ovarian cancer patients is frequently compromised by drug resistance, possibly due to altered drug metabolism. Platinum drugs are metabolised by glutathione S-transferase P1 (GSTP1), which is abundantly, but variably expressed in ovarian tumours. We have created novel ovarian tumour cell line models to investigate the extent to which differential GSTP1 expression influences chemosensitivity. Glutathione S-transferase P1 was stably deleted in A2780 and expression significantly reduced in cisplatin-resistant A2780DPP cells using Mission shRNA constructs, and MTT assays used to compare chemosensitivity to chemotherapy drugs used to treat ovarian cancer. Differentially expressed genes in GSTP1 knockdown cells were identified by Illumina HT-12 expression arrays and qRT-PCR analysis, and altered pathways predicted by MetaCore (GeneGo) analysis. Cell cycle changes were assessed by FACS analysis of PI-labelled cells and invasion and migration compared in quantitative Boyden chamber-based assays. Glutathione S-transferase P1 knockdown selectively influenced cisplatin and carboplatin chemosensitivity (2.3- and 4.83-fold change in IC50, respectively). Cell cycle progression was unaffected, but cell invasion and migration was significantly reduced. We identified several novel GSTP1 target genes and candidate platinum chemotherapy response biomarkers. Glutathione S-transferase P1 has an important role in cisplatin and carboplatin metabolism in ovarian cancer cells. Inter-tumour differences in GSTP1 expression may therefore influence response to platinum-based chemotherapy in ovarian cancer patients.

Poly(vinylpyrrolidone) supported copper nanoclusters: glutathione enhanced blue photoluminescence for application in phosphor converted light emitting devices

NASA Astrophysics Data System (ADS)

Wang, Zhenguang; Susha, Andrei S.; Chen, Bingkun; Reckmeier, Claas; Tomanec, Ondrej; Zboril, Radek; Zhong, Haizheng; Rogach, Andrey L.

2016-03-01

Poly(vinylpyrrolidone) supported Cu nanoclusters were synthesized by reduction of Cu(ii) ions with ascorbic acid in water, and initially showed blue photoluminescence with a quantum yield of 8%. An enhancement of the emission quantum yield has been achieved by treatment of Cu clusters with different electron-rich ligands, with the most pronounced effect (photoluminescence quantum yield of 27%) achieved with glutathione. The bright blue emission of glutathione treated Cu NCs is fully preserved in the solid state powder, which has been combined with commercial green and red phosphors to fabricate down-conversion white light emitting diodes with a high colour rendering index of 92.Poly(vinylpyrrolidone) supported Cu nanoclusters were synthesized by reduction of Cu(ii) ions with ascorbic acid in water, and initially showed blue photoluminescence with a quantum yield of 8%. An enhancement of the emission quantum yield has been achieved by treatment of Cu clusters with different electron-rich ligands, with the most pronounced effect (photoluminescence quantum yield of 27%) achieved with glutathione. The bright blue emission of glutathione treated Cu NCs is fully preserved in the solid state powder, which has been combined with commercial green and red phosphors to fabricate down-conversion white light emitting diodes with a high colour rendering index of 92. Electronic supplementary information (ESI) available: The optical spectra of control experiments for Cu NC synthesis, optimization of the reaction conditions, and spectra for LEDs chips and blue LEDs. See DOI: 10.1039/c6nr00806b

Glutathione depletion triggers actin cytoskeleton changes via actin-binding proteins.

PubMed

Zepeta-Flores, Nahum; Valverde, Mahara; Lopez-Saavedra, Alejandro; Rojas, Emilio

2018-06-04

The importance of glutathione (GSH) in alternative cellular roles to the canonically proposed, were analyzed in a model unable to synthesize GSH. Gene expression analysis shows that the regulation of the actin cytoskeleton pathway is strongly impacted by the absence of GSH. To test this hypothesis, we evaluate the effect of GSH depletion via buthionine sulfoximine (5 and 12.5 mM) in human neuroblastoma MSN cells. In the present study, 70% of GSH reduction did not induce reactive oxygen species, lipoperoxidation, or cytotoxicity, which enabled us to evaluate the effect of glutathione in the absence of oxidative stress. The cells with decreasing GSH levels acquired morphology changes that depended on the actin cytoskeleton and not on tubulin. We evaluated the expression of three actin-binding proteins: thymosin β4, profilin and gelsolin, showing a reduced expression, both at gene and protein levels at 24 hours of treatment; however, this suppression disappears after 48 hours of treatment. These changes were sufficient to trigger the co-localization of the three proteins towards cytoplasmic projections. Our data confirm that a decrease in GSH in the absence of oxidative stress can transiently inhibit the actin binding proteins and that this stimulus is sufficient to induce changes in cellular morphology via the actin cytoskeleton.

Single-Cell Force Spectroscopy of Probiotic Bacteria

PubMed Central