Reductase System Research Articles

Early Steps of the Biosynthesis of the Anticancer Antibiotic Pleurotin.

Pleurotin is a meroterpenoid specialized metabolite made by the fungus Hohenbuehelia grisea, and it is a lead anticancer molecule due to its irreversible inhibition of the thioredoxin-thioredoxin reductase system. Total synthesis of pleurotin has been achieved, including through a stereoselective route; however, its biosynthesis has not been characterized. In this study, we used isotope-labeled precursor feeding to show that the nonterpenoid quinone ring of pleurotin and its congeners is derived from phenylalanine. We sequenced the genome of H. grisea and used comparative transcriptomics to identify putative genes involved in pleurotin biosynthesis. We heterologously expressed a UbiA-like prenyltransferase from H. grisea that led to the accumulation of the first predicted pleurotin biosynthetic intermediate, 3-farnesyl-4-hydroxybenzoic acid. This work sets the foundation to fully elucidate the biosynthesis of pleurotin and its congeners, with long-term potential to optimize their production for therapeutic use and engineer the pathway toward the biosynthesis of valuable analogues.

Characterization of thioredoxin-thioredoxin reductase system in Filifactor alocis.

Filifactor alocis is a newly appreciated member of the periodontal community with a strong periodontal disease correlation. Little is known about the survival mechanisms by which F.alocis copes with oxidative stress and establishes the infection within the local inflammatory microenvironment of the periodontal pocket. The aim of this study is to investigate if F.alocis putative peroxiredoxin/AhpC protein FA768 may constitute an alkyl hydroperoxide reductase system utilizing putative thioredoxin reductase protein FA608, and putative thioredoxin/glutaredoxin homolog FA1411/FA455. FA768, FA608, FA1411 and FA455 proteins from F.alocis were expressed and purified from Escherichia coli. Insulin and 5,5-dithio-bis-2-nitrobenzoic acid (DTNB) reduction assays were performed to determine if purified FA1411 and FA455 proteins could be a substrate for FA608. The peroxidase activity of FA768 was examined by measuring its ability to reduce hydrogen peroxide (H2O2) with FA608 and FA1411/FA455 provided as the reducing systems. Further, the hydroperoxide substrate specificity of FA768 was analyzed by monitoring the NADPH oxidation in the presence of different peroxides, including H2O2, cumyl hydroperoxide (CHP), and tert-butyl hydroperoxide (t-BHP). In this study, we have demonstrated the existence of a functioning thioredoxin-dependent alkyl hydroperoxide system in F.alocis. This system is comprised of a thioredoxin reductase (FA608), a thioredoxin/glutaredoxin homolog (FA1411/FA455), and a typical 2-cysteine peroxiredoxin/AhpC (FA768). FA608, together with FA1411/FA455, can function as a thioredoxin reductase system to reduce insulin, DTNB, and FA768. FA455 is a glutaredoxin-like protein with thioredoxin functions in F.alocis. Both the FA768/FA608/FA1411 and FA768/FA608/FA455 reductase systems were NADPH-dependent and exhibited specificity for broad hydroperoxide substrates H2O2, CHP, and t-BHP. This is the first study of a thioredoxin dependent alkyl hydroperoxide system from a periodontal pathogen. This system is proposed to protect F.alocis against oxidative stress due to the likely absence of a catalase or an additional peroxiredoxin homolog.

Activation of cellular antioxidative stress and migration activities by purified components from immortalized stem cells from human exfoliated deciduous teeth

Although stem cell-based regenerative medicine has been extensively studied, it remains difficult to reconstruct three dimensional tissues and organs in combination with vascular systems in vitro. One clinically successful therapy is transplantation of mesenchymal stem cells (MSC) into patients with graft versus host disease. However, transplanted cells are immediately damaged and destroyed because of innate immune reactions provoked by thrombogenic inflammation, and patients need to take immunosuppressive drugs for the immunological regulation of allogeneic cells. This reduces the benefits of stem cell transplantation. Therefore, alternative therapies are more realistic options for clinical use. In this study, we aimed to take advantage of the therapeutic efficacy of MSC and use multiple cytokines released from MSC, that is, stem cells from human exfoliated deciduous teeth (SHEDs). Here, we purified components from conditioned media of immortalized SHED (IM-SHED-CM) and evaluated the activities of intracellular dehydrogenase, cell migration, and antioxidative stress by studying the cells. The immortalization of SHED could make the stable supply of CM possible. We found that the fractionated component of 50–100 kD from IM-SHED-CM had higher efficacy than the original IM-SHED-CM in terms of intracellular dehydrogenase and cell migration in which intracellular signal transduction was activated via receptor tyrosine kinases, and the glutathione peroxidase and reductase system was highly active. Although antioxidative stress activities in the fractionated component of 50–100 kD had slightly lower than that of original IM-SHE-CM, the fraction still had the activity. Thus, the use of fractionated components of 50–100 kD from IM-SHED-CM could be an alternative choice for MSC transplantation because the purified components from CM could maintain the effect of cytokines from SHED.

Inhibition of zinc ions in sulfur-driven autotrophic denitrification process: What is the behavior of extracellular polymeric substances?

Sulfur-driven autotrophic denitrification (SAD) process is a cost-effective and sustainable method for nitrogen removal from wastewater. However, a higher concentration of zinc ions (Zn(II)) flowing into wastewater treatment plants poses a potential threat to the SAD process. This study examined that a half maximal inhibitory concentration (IC50) of Zn(II) was 7 mg·L−1 in the SAD process. Additionally, the addition of 20 mg·L−1 Zn(II) resulted in a severe accumulation of nitrite to 150.20 ± 6.00 mg·L−1 when the initial concentration of nitrate was 500 mg·L−1. Moreover, the activities of nitrate reductase, nitrite reductase, dehydrogenase and electron transport system were significantly inhibited under Zn(II) stress. The addition of Zn(II) inhibited EPS secretion and worsened electrochemical properties. The result was attributed to the spontaneous binding between EPS and Zn(II), with a ΔG of −17.50 KJ·mol−1 and a binding constant of 1.77 × 104 M−1, respectively. Meanwhile, the protein, fulvic acid, and humic-like substances occurred static quenching after Zn(II) addition, with -OH and -C=O groups providing binding sites. The binding sequence was fulvic acid→protein→humic acid and -OH → -C=O. Zn(II) also reduced the content of α-helix, which was unfavorable for electron transfer. Additionally, the Zn(II) loosened protein structure, resulting in a 50 % decrease in α-helix/(β-sheet+random coil). This study reveals the effect of Zn(II) on the SAD process and enhances our understanding of EPS behavior under metal ions stress.

The architecture of redox microdomains: Cascading gradients and peroxiredoxins’ redox-oligomeric coupling integrate redox signaling and antioxidant protection

In the cytosol of human cells under low oxidative loads, hydrogen peroxide is confined to microdomains around its supply sites, due to its fast consumption by peroxiredoxins. So are the sulfenic and disulfide forms of the 2-Cys peroxiredoxins, according to a previous theoretical analysis [Travasso et al., Redox Biology 15 (2017) 297]. Here, an extended reaction-diffusion model that for the first time considers the differential properties of human peroxiredoxins 1 and 2 and the thioredoxin redox cycle predicts important new aspects of the dynamics of redox microdomains. The peroxiredoxin 1 sulfenates and disulfides are more localized than the corresponding peroxiredoxin 2 forms, due to the former peroxiredoxin's faster resolution step. The thioredoxin disulfides are also localized. As the H2O2 supply rate (vsup) approaches and then surpasses the maximal rate of the thioredoxin/thioredoxin reductase system (V), these concentration gradients become shallower, and then vanish. At low vsup the peroxiredoxin concentration determines the H2O2 concentrations and gradient length scale, but as vsup approaches V, the thioredoxin reductase activity gains influence. A differential mobility of peroxiredoxin disulfide dimers vs. reduced decamers enhances the redox polarity of the cytosol: as vsup approaches V, reduced decamers are preferentially retained far from H2O2 sources, attenuating the local H2O2 buildup. Substantial total protein concentration gradients of both peroxiredoxins emerge under these conditions, and the concentration of reduced peroxiredoxin 1 far from the H2O2 sources even increases with vsup. Altogether, the properties of 2-Cys peroxiredoxins and thioredoxin are such that localized H2O2 supply induces a redox and functional polarization between source-proximal regions (redox microdomains) that facilitate peroxiredoxin-mediated signaling and distal regions that maximize antioxidant protection.

Analysis of critical residues for peroxygenation and improved peroxygenase activity via in situ H2O2 generation in CYP105D18.

Limited numbers of CYPs have been reported to work naturally as peroxygenases. The peroxide shunt pathway can be efficiently used as an alternative for the NAD(P)H and reductase systems, particularly in high hydrogen peroxide (H2O2) resistance CYPs. We reported the structural and biochemical features of CYP105D18 peroxygenase for its high H2O2 tolerance capacity. Q348 was a crucial residue for the stability of CYP105D18 during the exposure to H2O2. In addition, the role of the hydrophilic amino acid T239 from the I helix for peroxygenation and regiospecificity toward testosterone was investigated. Interestingly, T239E differs in product formation from wild type, catalyzing testosterone to androstenedione in the presence of H2O2. The other variant, T239A, worked with the Pdx/Pdr system and was unable to catalyze testosterone conversion in the presence of H2O2, suggesting the transformation of peroxygenase into monooxygenase. CYP105D18 supported the alternative method of H2O2 used for the catalysis of testosterone. The use of the same concentration of urea hydrogen peroxide adducts in place of direct H2O2 was more efficient for 2β-hydroxytestosterone conversion. Furthermore, in situ H2O2 generation using GOx/glucose system enhanced the catalytic efficiency (kcat/Km) for wild type and F184A by 1.3- and 1.9-fold, respectively, compared to direct use of H2O2 The engineering of CYP105D18, its improved peroxygenase activity, and alteration in the product oxidation facilitate CYP105D18 as a potential candidate for biotechnological applications.

Cardiac nitric oxide scavenging: role of myoglobin and mitochondria.

Vascular production of nitric oxide (NO) regulates vascular tone. However, highly permeable NO entering the cardiomyocyte would profoundly impact metabolism and signalling without scavenging mechanisms. The purpose of this study was to establish mechanisms of cardiac NO scavenging. Quantitative optical studies of normoxic working hearts demonstrated that micromolar NO concentrations did not alter mitochondria redox state or respiration despite detecting NO oxidation of oxymyoglobin to metmyoglobin. These data are consistent with proposals that the myoglobin/myoglobin reductase (Mb/MbR) system is the major NO scavenging site. However, kinetic studies in intact hearts reveal a minor role (∼9%) for the Mb/MbR system in NO scavenging. In vitro, oxygenated mitochondria studies confirm that micromolar concentrations of NO bind cytochrome oxidase (COX) and inhibit respiration. Mitochondria had a very high capacity for NO scavenging, importantly, independent of NO binding to COX. NO is also known to quickly react with reactive oxygen species (ROS) in vitro. Stimulation of NO scavenging with antimycin and its inhibition by substrate depletion are consistent with NO interacting with ROS generated in Complex I or III under aerobic conditions. Extrapolating these in vitro data to the intact heart supports the hypothesis that mitochondria are a major site of cardiac NO scavenging. KEY POINTS: Cardiomyocyte scavenging of vascular nitric oxide (NO) is critical in maintaining normal cardiac function. Myoglobin redox cycling via myoglobin reductase has been proposed as a major NO scavenging site in the heart. Non-invasive optical spectroscopy was used to monitor the effect of NO on mitochondria and myoglobin redox state in intact beating heart and isolated mitochondria. These non-invasive studies reveal myoglobin/myoglobin reductase plays a minor role in cardiac NO scavenging. A high capacity for NO scavenging by heart mitochondria was demonstrated, independent of cytochrome oxidase binding but dependent on oxygen and high redox potentials consistent with generation of reactive oxygen species.

Hypoxia-responsive calixarene-grafted self-assembled peptide hydrogel for inflammation suppression in ischemic stroke

Hypoxia is a significant characteristic of ischemic stroke. Recently, development of the pathology of tissue injury and deployment of new hypoxia-responsive technologies provide further opportunity for the application of hypoxia-responsive materials beyond cancer. To fully take advantage of hypoxic pathological feature of stroke and make advancement of stroke therapy, a hypoxia-responsive self-assembled peptide hydrogel was prepared for inflammation suppression in ischemic stroke treatment, utilizing azocalixarene (CA) as a hypoxia-responsive controlled release drug carrier. After induction of PBS, the peptide hydrogel was fabricated with brain-similar rheological modulus and shear-thinning property. The hypoxia-responsive performance of the peptide hydrogel was validated by three hypoxia & reductase systems including sodium hydrosulfite (SDT), rat liver microsomes and oxygen-glucose deprivation model (OGD) to achieve the responsive release of Cyanine 5-dimethyl (CY5-DM) or Fingolimod (FTY720) in vitro. After local administration of the hypoxia-responsive self-assembled peptide hydrogel, the improved motor function, reduced infarct volume and alleviated inflammation were achieved, revealing the effective stroke therapy in vivo.

CBSX2 is required for the efficient oxidation of chloroplast redox-regulated enzymes in darkness.

Thiol/disulfide-based redox regulation in plant chloroplasts is essential for controlling the activity of target proteins in response to light signals. One of the examples of such a role in chloroplasts is the activity of the chloroplast ATP synthase (CFoCF1), which is regulated by the redox state of the CF1γ subunit and involves two cysteines in its central domain. To investigate the mechanism underlying the oxidation of CF1γ and other chloroplast redox-regulated enzymes in the dark, we characterized the Arabidopsis cbsx2 mutant, which was isolated based on its altered NPQ (non-photochemical quenching) induction upon illumination. Whereas in dark-adapted WT plants CF1γ was completely oxidized, a small amount of CF1γ remained in the reduced state in cbsx2 under the same conditions. In this mutant, reduction of CF1γ was not affected in the light, but its oxidation was less efficient during a transition from light to darkness. The redox states of the Calvin cycle enzymes FBPase and SBPase in cbsx2 were similar to those of CF1γ during light/dark transitions. Affinity purification and subsequent analysis by mass spectrometry showed that the components of the ferredoxin-thioredoxin reductase/thioredoxin (FTR-Trx) and NADPH-dependent thioredoxin reductase (NTRC) systems as well as several 2-Cys peroxiredoxins (Prxs) can be co-purified with CBSX2. In addition to the thioredoxins, yeast two-hybrid analysis showed that CBSX2 also interacts with NTRC. Taken together, our results suggest that CBSX2 participates in the oxidation of the chloroplast redox-regulated enzymes in darkness, probably through regulation of the activity of chloroplast redox systems in vivo.

Compromise of mitochondrial thiol reductase systems by MitoCDNB magnifies immune cell responses to inflammatory stimuli

Compromise of mitochondrial thiol reductase systems by MitoCDNB magnifies immune cell responses to inflammatory stimuli

Glutathionylation of pyruvate dehydrogenase complex E2 and inflammatory cytokine production during acute inflammation are magnified by mitochondrial oxidative stress

Lipopolysaccharide (LPS) is a known inducer of inflammatory signaling which triggers generation of reactive oxygen species (ROS) and cell death in responsive cells like THP-1 promonocytes and freshly isolated human monocytes. A key LPS-responsive metabolic pivot point is the 9 MDa mitochondrial pyruvate dehydrogenase complex (PDC), which provides pyruvate dehydrogenase (E1), lipoamide-linked transacetylase (E2) and lipoamide dehydrogenase (E3) activities to produce acetyl-CoA from pyruvate. While phosphorylation-dependent decreases in PDC activity following LPS treatment or sepsis have been deeply investigated, redox-linked processes have received less attention. Data presented here demonstrate that LPS-induced reversible oxidation within PDC occurs in PDCE2 in both THP-1 cells and primary human monocytes. Knockout of PDCE2 by CRISPR and expression of FLAG-tagged PDCE2 in THP-1 cells demonstrated that LPS-induced glutathionylation is associated with wild type PDCE2 but not mutant protein lacking the lipoamide-linking lysine residues. Moreover, the mitochondrially-targeted electrophile MitoCDNB, which impairs both glutathione- and thioredoxin-based reductase systems, elevates ROS similar to LPS but does not cause PDCE2 glutathionylation. However, LPS and MitoCDNB together are highly synergistic for PDCE2 glutathionylation, ROS production, and cell death. Surprisingly, the two treatments together had differential effects on cytokine production; pro-inflammatory IL-1β production was enhanced by the co-treatment, while IL-10, an important anti-inflammatory cytokine, dropped precipitously compared to LPS treatment alone. This new information may expand opportunities to understand and modulate PDC redox status and activity and improve the outcomes of pathological inflammation.

Disruption of Bacterial Thiol-Dependent Redox Homeostasis by Magnolol and Honokiol as an Antibacterial Strategy.

Some traditional Chinese medicines (TCMs) possess various redox-regulation properties, but whether the redox regulation contributes to antibacterial mechanisms is not known. Here, ginger juice processed Magnoliae officinalis cortex (GMOC) was found to show strong antibacterial activities against some Gram-positive bacteria, but not Gram-negative bacteria including E. coli, while the redox-related transcription factor oxyR deficient E. coli mutant was sensitive to GMOC. In addition, GMOC and its main ingredients, magnolol and honokiol, exhibited inhibitory effects on the bacterial thioredoxin (Trx) system, a major thiol-dependent disulfide reductase system in bacteria. The effects of magnolol and honokiol on cellular redox homeostasis were further verified by elevation of the intracellular ROS levels. The therapeutic efficacies of GMOC, magnolol and honokiol were further verified in S. aureus-caused mild and acute peritonitis mouse models. Treatments with GMOC, magnolol and honokiol significantly reduced the bacterial load, and effectively protected the mice from S. aureus-caused peritonitis infections. Meanwhile, magnolol and honokiol produced synergistic effects when used in combination with several classic antibiotics. These results strongly suggest that some TCMs may exert their therapeutic effects via targeting the bacterial thiol-dependent redox system.

The Thioredoxin System in Edwardsiella piscicida Contributes to Oxidative Stress Tolerance, Motility, and Virulence.

Edwardsiella piscicida is an important fish pathogen that causes substantial economic losses. In order to understand its pathogenic mechanism, additional new virulence factors need to be identified. The bacterial thioredoxin system is a major disulfide reductase system, but its function is largely unknown in E. piscicida. In this study, we investigated the roles of the thioredoxin system in E. piscicida (named TrxBEp, TrxAEp, and TrxCEp, respectively) by constructing a correspondingly markerless in-frame mutant strain: ΔtrxB, ΔtrxA, and ΔtrxC, respectively. We found that (i) TrxBEp is confirmed as an intracellular protein, which is different from the prediction made by the Protter illustration; (ii) compared to the wild-type strain, ΔtrxB exhibits resistance against H2O2 stress but high sensitivity to thiol-specific diamide stress, while ΔtrxA and ΔtrxC are moderately sensitive to both H2O2 and diamide conditions; (iii) the deletions of trxBEp, trxAEp, and trxCEp damage E. piscicida's flagella formation and motility, and trxBEp plays a decisive role; (iv) deletions of trxBEp, trxAEp, and trxCEp substantially abate bacterial resistance against host serum, especially trxBEp deletion; (v) trxAEp and trxCEp, but not trxBEp, are involved in bacterial survival and replication in phagocytes; (vi) the thioredoxin system participates in bacterial dissemination in host immune tissues. These findings indicate that the thioredoxin system of E. piscicida plays an important role in stress resistance and virulence, which provides insight into the pathogenic mechanism of E. piscicida.

Ferroptosis-modulating small molecules for targeting drug-resistant cancer: Challenges and opportunities in manipulating redox signaling.

Ferroptosis is an iron-dependent cell death program that is characterized by excessive lipid peroxidation. Triggering ferroptosis has been proposed as a promising strategy to fight cancer and overcome drug resistance in antitumor therapy. Understanding the molecular interactions and structural features of ferroptosis-inducing compounds might therefore open the door to efficient pharmacological strategies against aggressive, metastatic, and therapy-resistant cancer. We here summarize the molecular mechanisms and structural requirements of ferroptosis-inducing small molecules that target central players in ferroptosis. Focus is placed on (i) glutathione peroxidase (GPX) 4, the only GPX isoenzyme that detoxifies complex membrane-bound lipid hydroperoxides, (ii) the cystine/glutamate antiporter system Xc - that is central for glutathione regeneration, (iii) the redox-protective transcription factor nuclear factor erythroid 2-related factor (NRF2), and (iv) GPX4 repression in combination with induced heme degradation via heme oxygenase-1. We deduce common features for efficient ferroptotic activity and highlight challenges in drug development. Moreover, we critically discuss the potential of natural products as ferroptosis-inducing lead structures and provide a comprehensive overview of structurally diverse biogenic and bioinspired small molecules that trigger ferroptosis via iron oxidation, inhibition of the thioredoxin/thioredoxin reductase system or less defined modes of action.

Development of vitamin B12 dependency in Saccharomyces cerevisiae.

For decades, the industrial vitamin B12 (cobalamin) production has been based on bacterial producer strains. Due to limited methods for strain optimization and difficult strain handling, the desire for new vitamin B12-producing hosts has risen. As a vitamin B12-independent organism with a big toolbox for genomic engineering and easy-to-handle cultivation conditions, Saccharomyces cerevisiae has high potential for heterologous vitamin B12 production. However, the B12 synthesis pathway is long and complex. To be able to easily engineer and evolve B12-producing recombinant yeast cells, we have developed an S. cerevisiae strain whose growth is dependent on vitamin B12. For this, the B12-independent methionine synthase Met6 of yeast was replaced by a B12-dependent methionine synthase MetH from Escherichia coli. Adaptive laboratory evolution, RT-qPCR, and overexpression experiments show that additional high-level expression of a bacterial flavodoxin/ferredoxin-NADP+ reductase (Fpr-FldA) system is essential for in vivo reactivation of MetH activity and growth. Growth of MetH-containing yeast cells on methionine-free media is only possible with the addition of adenosylcobalamin or methylcobalamin. A heterologous vitamin B12 transport system turned out to be not necessary for the uptake of cobalamins. This strain should be a powerful chassis to engineer B12-producing yeast cells.

A Novel TrxR1 Inhibitor Regulates NK and CD8+ T Cell Infiltration and Cytotoxicity, Enhancing the Efficacy of Anti-PD-1 Immunotherapy against Hepatocarcinoma.

Hepatocellular carcinoma (HCC) has the third highest cancer-related mortality rate globally. The immunosuppressive microenvironment of HCC limits effective treatment options. HCC cells and associated microenvironmental factors suppress NK and T cell infiltration and cytotoxic activities. The abnormal number or function of NK and T cells leads to a lack of immune surveillance. Recently, immunotherapy targeting PD-1 and PD-L1 has been shown to activate functionally exhausted cytotoxic immune cells in some solid tumors. However, the response rate and therapeutic efficacy against solid tumors with little lymphocyte infiltration are limited, especially for HCC. Therefore, new targets and therapeutics that induce tumor cell apoptosis and overcome the problem of depletion of immune cells, thereby inhibiting the immune escape of HCC cells, are urgently required. Butaselen (2-bis[2-(1,2-benzisothiazol-2(2H)-ketone)]butane), an organic molecule containing selenium, is a new type of thioredoxin reductase inhibitor. In this study, we found that butaselen promoted NK and T cell activity and infiltration in the tumor microenvironment in HCC-bearing mice by enhancing the expression of CXCR3, NKG2D, and their respective ligands. When used alone, it can significantly inhibit tumor growth and exert a synergistic effect in combination with PD-1 blockade. We suggested the role of the thioredoxin reductase system in the regulation of the tumor immunosuppressive microenvironment and developed a new effective therapeutic molecule for HCC, revealing the mechanism of butaselen in inhibiting tumor cell immune escape.

Systematic Optimization of HPO-CPR to Boost (+)-Nootkatone Synthesis in Engineered Saccharomyces cerevisiae.

As an important and expensive natural sesquiterpene compound in grapefruit, the interest in (+)-nootkatone is stimulated by its strong grapefruit-like odor and physiological activities, which induce efforts for its microbial production. However, the low catalytic efficiency of the cytochrome P450-P450 reductase (HPO-CPR) system is the main challenge. We developed a high-throughput screening (HTS) method using the principle of the color reaction between carbonyl compounds and 2,4-dinitrophenylhydrazine (DNPH), which could rapidly screen the activity of candidate HPO mutants. After optimizing the pairing of HPO and CPR and through semirational design, the optimal mutant HPO_M18 with catalytic performance 2.54 times that of the initial was obtained. An encouraging (+)-nootkatone titer of 2.39 g/L was achieved through two-stage fed-batch fermentation after metabolic engineering and endoplasmic reticulum engineering, representing the highest titer reported to date. Our findings lay the foundation for the development of an economically viable bioprocess for (+)-nootkatone.

Bacterial redox response factors in the management of environmental oxidative stress.

Bacteria evolved to survive in the available environmental chemosphere via several cellular mechanisms. A rich pool of antioxidants and stress regulators plays a significant role in the survival of bacteria in unfavorable environmental conditions. Most of the microbes exhibit resistant phenomena in toxic environment niches. Naturally, bacteria possess efficient thioredoxin reductase, glutaredoxin, and peroxiredoxin redox systems to handle environmental oxidative stress. Further, an array of transcriptional regulators senses the oxidative stress conditions. Transcription regulators, such as OxyR, SoxRS, PerR, UspA, SsrB, MarA, OhrR, SarZ, etc., sense and transduce bacterial oxidative stress responses. The redox-sensitive transcription regulators continuously recycle the utilized antioxidant enzymes during oxidative stress. These regulators promote the expression of antioxidant enzymes such as superoxide dismutase, catalase, and peroxides that overcome oxidative insults. Therefore, the transcriptional regulations maintain steady-state activities of antioxidant enzymes representing the resistance against host cell/environmental oxidative insults. Further, the redox system provides reducing equivalents to synthesize biomolecules, thereby contributing to cellular repair mechanisms. The inactive transcriptional regulators in the undisturbed cells are activated by oxidative stress. The oxidized transcriptional regulators modulate the expression of antioxidant and cellular repair enzymes to survive in extreme environmental conditions. Therefore, targeting these antioxidant systems and response regulators could alter cellular redox homeostasis. This review presents the mechanisms of different redox systems that favor bacterial survival in extreme environmental oxidative stress conditions.

S-Denitrosylation: A Crosstalk between Glutathione and Redoxin Systems.

S-nitrosylation of proteins occurs as a consequence of the derivatization of cysteine thiols with nitric oxide (NO) and is often associated with diseases and protein malfunction. Aberrant S-nitrosylation, in addition to other genetic and epigenetic factors, has gained rapid importance as a prime cause of various metabolic, respiratory, and cardiac disorders, with a major emphasis on cancer and neurodegeneration. The S-nitrosoproteome, a term used to collectively refer to the diverse and dynamic repertoire of S-nitrosylated proteins, is relatively less explored in the field of redox biochemistry, in contrast to other covalently modified versions of the same set of proteins. Advancing research is gradually unveiling the enormous clinical importance of S-nitrosylation in the etiology of diseases and is opening up new avenues of prompt diagnosis that harness this phenomenon. Ever since the discovery of the two robust and highly conserved S-nitrosoglutathione reductase and thioredoxin systems as candidate denitrosylases, years of rampant speculation centered around the identification of specific substrates and other candidate denitrosylases, subcellular localization of both substrates and denitrosylases, the position of susceptible thiols, mechanisms of S-denitrosylation under basal and stimulus-dependent conditions, impact on protein conformation and function, and extrapolating these findings towards the understanding of diseases, aging and the development of novel therapeutic strategies. However, newer insights in the ever-expanding field of redox biology reveal distinct gaps in exploring the crucial crosstalk between the redoxins/major denitrosylase systems. Clarifying the importance of the functional overlap of the glutaredoxin, glutathione, and thioredoxin systems and examining their complementary functions as denitrosylases and antioxidant enzymatic defense systems are essential prerequisites for devising a rationale that could aid in predicting the extent of cell survival under high oxidative/nitrosative stress while taking into account the existence of the alternative and compensatory regulatory mechanisms. This review thus attempts to highlight major gaps in our understanding of the robust cellular redox regulation system, which is upheld by the concerted efforts of various denitrosylases and antioxidants.

The Thioredoxin System of Mammalian Cells and Its Modulators.

Oxidative stress involves the increased production and accumulation of free radicals, peroxides, and other metabolites that are collectively termed reactive oxygen species (ROS), which are produced as by-products of aerobic respiration. ROS play a significant role in cell homeostasis through redox signaling and are capable of eliciting damage to macromolecules. Multiple antioxidant defense systems have evolved to prevent dangerous ROS accumulation in the body, with the glutathione and thioredoxin/thioredoxin reductase (Trx/TrxR) systems being the most important. The Trx/TrxR system has been used as a target to treat cancer through the thiol–disulfide exchange reaction mechanism that results in the reduction of a wide range of target proteins and the generation of oxidized Trx. The TrxR maintains reduced Trx levels using NADPH as a co-substrate; therefore, the system efficiently maintains cell homeostasis. Being a master regulator of oxidation–reduction processes, the Trx-dependent system is associated with cell proliferation and survival. Herein, we review the structure and catalytic properties of the Trx/TrxR system, its role in cellular signaling in connection with other redox systems, and the factors that modulate the Trx system.