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NOX-like ROS production by glutathione reductase

ABSTRACT: Summary In organisms from bacteria to mammals, NADPH oxidase (NOX) catalyzes the production of beneficial reactive oxygen species (ROS) such as superoxide (O2−). However, our previous research implicated glutathione reductase (GR), a canonical antioxidant enzyme, as a source of extracellular superoxide in the marine diatom Thalassiosira oceanica. Here, we expressed and characterized the two GR isoforms of T. oceanica. Both coupled the oxidation of NADPH, the native electron donor, to oxygen reduction, giving rise to superoxide in the absence of glutathione disulfide, the native electron acceptor. Superoxide production by ToGR1 exhibited similar kinetics as representative NOX enzymes, and inhibition assays agreed with prior organismal studies, supporting a physiological role. ToGR is similar to GR from human, yeast, and bacteria, suggesting that NOX-like ROS production by GR could be widespread. Yet unlike NOX, GR-mediated ROS production is independent of iron, which may provide an advantageous way of making ROS under micronutrient stress. Graphical abstract Highlights • Glutathione reductase expressed from a microbial phototroph is found to produce ROS• This promiscuous reaction shows similar kinetics and inhibition as NOX-derived ROS• The GR pathway of ROS production has cellular benefits under physiological stress• GR may function similarly to NOX in other taxa, providing metabolic versatility Earth sciences; Oceanography; Microbiology; Biocatalysis; Bioengineering

SUBMITTER: Diaz J

PROVIDER: S-EPMC9519596 | biostudies-literature | 2022 Sep

REPOSITORIES: biostudies-literature

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c-Jun N terminal kinase modulates NOX-4 derived ROS production and myofibroblasts differentiation in human breast stromal cells.

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Synergistic effect of elevated glucose levels with SARS-CoV-2 spike protein induced NOX-dependent ROS production in endothelial cells.

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The IRAK-ERK-p67phox-Nox-2 axis mediates TLR4, 2-induced ROS production for IL-1β transcription and processing in monocytes.

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Cystatin SN inhibits auranofin-induced cell death by autophagic induction and ROS regulation via glutathione reductase activity in colorectal cancer.

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Ebselen exerts antifungal activity by regulating glutathione (GSH) and reactive oxygen species (ROS) production in fungal cells.

Project description:BackgroundEbselen, an organoselenium compound and a clinically safe molecule has been reported to possess potent antifungal activity, but its antifungal mechanism of action and in vivo antifungal activity remain unclear.MethodsThe antifungal effect of ebselen was tested against Candida albicans, C. glabrata, C. tropicalis, C. parapsilosis, Cryptococcus neoformans, and C. gattii clinical isolates. Chemogenomic profiling and biochemical assays were employed to identify the antifungal target of ebselen. Ebselen's antifungal activity in vivo was investigated in a Caenorhabditis elegans animal model.ResultsEbselen exhibits potent antifungal activity against both Candida spp. and Cryptococcus spp., at concentrations ranging from 0.5 to 2μg/ml. Ebselen rapidly eradicates a high fungal inoculum within 2h of treatment. Investigation of the drug's antifungal mechanism of action indicates that ebselen depletes intracellular glutathione (GSH) levels, leading to increased production of reactive oxygen species (ROS), and thereby disturbs the redox homeostasis in fungal cells. Examination of ebselen's in vivo antifungal activity in two Caenorhabditis elegans models of infection demonstrate that ebselen is superior to conventional antifungal drugs (fluconazole, flucytosine and amphotericin) in reducing Candida and Cryptococcus fungal load.ConclusionEbselen possesses potent antifungal activity against clinically relevant isolates of both Candida and Cryptococcus by regulating GSH and ROS production. The potent in vivo antifungal activity of ebselen supports further investigation for repurposing it for use as an antifungal agent.General significanceThe present study shows that ebselen targets glutathione and also support that glutathione as a potential target for antifungal drug development.

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Formaldehyde production from isoprene oxidation across NOx regimes.

Project description:The chemical link between isoprene and formaldehyde (HCHO) is a strong, non-linear function of NOx (= NO + NO2). This relationship is a linchpin for top-down isoprene emission inventory verification from orbital HCHO column observations. It is also a benchmark for overall photochemical mechanism performance with regard to VOC oxidation. Using a comprehensive suite of airborne in situ observations over the Southeast U.S., we quantify HCHO production across the urban-rural spectrum. Analysis of isoprene and its major first-generation oxidation products allows us to define both a "prompt" yield of HCHO (molecules of HCHO produced per molecule of freshly-emitted isoprene) and the background HCHO mixing ratio (from oxidation of longer-lived hydrocarbons). Over the range of observed NOx values (roughly 0.1 - 2 ppbv), the prompt yield increases by a factor of 3 (from 0.3 to 0.9 ppbv ppbv-1), while background HCHO increases by a factor of 2 (from 1.6 to 3.3 ppbv). We apply the same method to evaluate the performance of both a global chemical transport model (AM3) and a measurement-constrained 0-D steady state box model. Both models reproduce the NOx dependence of the prompt HCHO yield, illustrating that models with updated isoprene oxidation mechanisms can adequately capture the link between HCHO and recent isoprene emissions. On the other hand, both models under-estimate background HCHO mixing ratios, suggesting missing HCHO precursors, inadequate representation of later-generation isoprene degradation and/or under-estimated hydroxyl radical concentrations. Detailed process rates from the box model simulation demonstrate a 3-fold increase in HCHO production across the range of observed NOx values, driven by a 100% increase in OH and a 40% increase in branching of organic peroxy radical reactions to produce HCHO.

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Exploring the Potential of Glutathione Reductase Overexpression to Improve Tellurium Nanoparticle Production in <i>Escherichia coli</i>.

Project description:Tellurium nanoparticles (TeNPs) are emerging as valuable materials in various technological and biomedical applications due to their unique physicochemical properties. In general, TeNPs are prepared using chemical methods based on a redox reaction in which strong reducing agents are employed which are often toxic and harmful to the environment. Biological biosynthesis provides a green strategy for substituting the commonly used reducing chemical agents with microorganisms or enzymes. Among the enzymes noted as key players in microbial tellurite reduction, glutathione reductase (GR) has been identified; however, its specific role in enhancing TeNP biosynthesis has yet to be fully elucidated. In this study, we aimed to evaluate the impact of GR overexpression on TeNP production in Escherichia coli (E. coli). For this purpose, four GR enzymes from different microorganisms identified as tellurite resistant were heterogeneously expressed and purified from E. coli. The kinetic parameters for NADPH and oxidized glutathione (GSSG), the native substrates of GR, were determined to evaluate their TR activity under saturated NADPH concentrations. The GR from Pseudomonas lini strain BNF22 presented the highest catalytic efficiency for NADPH and exhibited greater TR activity. This enzyme was overexpressed in E. coli MG1655 (DE3) and cells overexpressing GR increased the yield of TeNPs in those cells, presenting an increased elemental cell tellurium composition. Our results provide valuable insights for the development of engineered E. coli as a platform for TeNP biosynthesis. Using microorganisms as a green strategy for TeNP production, the results of this study highlight the enzymatic mechanisms underlying the role of GR in the biosynthesis of TeNPs.

| S-EPMC11855334 | biostudies-literature

Electrophiles modulate glutathione reductase activity via alkylation and upregulation of glutathione biosynthesis.

Project description:Cells evolved robust homeostatic mechanisms to protect against oxidation or alkylation by electrophilic species. Glutathione (GSH) is the most abundant intracellular thiol, protects cellular components from oxidation and is maintained in a reduced state by glutathione reductase (GR). Nitro oleic acid (NO2-OA) is an electrophilic fatty acid formed under digestive and inflammatory conditions that both reacts with GSH and induces its synthesis upon activation of Nrf2 signaling. The effects of NO2-OA on intracellular GSH homeostasis were evaluated. In addition to upregulation of GSH biosynthesis, we observed that NO2-OA increased intracellular GSSG in an oxidative stress-independent manner. NO2-OA directly inhibited GR in vitro by covalent modification of the catalytic Cys61, with kon of (3.45 ± 0.04) × 103 M-1 s-1, koff of (4.4 ± 0.4) × 10-4 s-1, and Keq of (1.3 ± 0.1) × 10-7 M. Akin to NO2-OA, the electrophilic Nrf2 activators bardoxolone-imidazole (CDDO-Im), bardoxolone-methyl (CDDO-Me) and dimethyl fumarate (DMF) also upregulated GSH biosynthesis while promoting GSSG accumulation, but without directly inhibiting GR activity. In vitro assays in which GR was treated with increasing GSH concentrations and GSH depletion experiments in cells revealed that GR activity is finely regulated via product inhibition, an observation further supported by theoretical (kinetic modeling of cellular GSSG:GSH levels) approaches. Together, these results describe two independent mechanisms by which electrophiles modulate the GSH/GSSG couple, and provide a novel conceptual framework to interpret experimentally determined values of GSH and GSSG.

| S-EPMC6348771 | biostudies-literature

Ectopic suicide inhibition of thioredoxin glutathione reductase.

Project description:Selective suicide inhibitors represent a seductively attractive approach for inactivation of therapeutically relevant enzymes since they are generally devoid of off-target toxicity in vivo. While most suicide inhibitors are converted to reactive species at enzyme active sites, theoretically bioactivation can also occur in ectopic (secondary) sites that have no known function. Here, we report an example of such an "ectopic suicide inhibition", an unprecedented bioactivation mechanism of a suicide inhibitor carried out by a non-catalytic site of thioredoxin glutathione reductase (TGR). TGR is a promising drug target to treat schistosomiasis, a devastating human parasitic disease. Utilizing hits selected from a high throughput screening campaign, time-resolved X-ray crystallography, molecular dynamics, mass spectrometry, molecular modeling, protein mutagenesis and functional studies, we find that 2-naphtholmethylamino derivatives bound to this novel ectopic site of Schistosoma mansoni (Sm)TGR are transformed to covalent modifiers and react with its mobile selenocysteine-containing C-terminal arm. In particular, one 2-naphtholmethylamino compound is able to specifically induce the pro-oxidant activity in the inhibited enzyme. Since some 2-naphtholmethylamino analogues show worm killing activity and the ectopic site is not conserved in human orthologues, a general approach to development of novel and selective anti-parasitic therapeutics against schistosoma is proposed.

| S-EPMC7583042 | biostudies-literature

Cytosolic glutathione reductase (OsGR2) regulates plant development and salt stress tolerance in rice

Project description:Rice genome contains three genes that encode for glutathione reductase (GR) viz., OsGR1, 2 and 3. GR is an important component of the anti-oxidative machinery of plant cells. GR2 down-regulated plants were produced by RNAi mediated down-regulation of GR2 (GR2-Ri). GR2-Ri plants were significantly smaller and have significantly lower grain yield (grain number per panicle) compared to WT(Wild type), under control conditions. RNA-Seq analysis of panicles (differentiation stage) identified genes known to affect grain size to be differentially regulated in GR2-Ri compared to WT, respectively, under control conditions.

2024-08-30 | E-MTAB-7172 | biostudies-arrayexpress

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