Purification, structural characterization, and neuroprotective effect of 3,6-diisobutyl-2,5-piperazinedione from Halomonas pacifica CARE-V15 against okadaic acid-induced neurotoxicity in zebrafish model.

Halomonas pacifica CARE-V15 was isolated from the southeastern coast of India to determine its genome sequence. Secondary metabolite gene clusters were identified using an anti-SMASH server. The concentrated crude ethyl acetate extract was evaluated by GC-MS. The bioactive compound from the crude ethyl acetate extract was fractionated by gel column chromatography. HPLC was used to purify the 3,6-diisobutyl-2,5-piperazinedione (DIP), and the structure was determined using FTIR and NMR spectroscopy. Purified DIP was used in an in silico molecular docking analysis. Purified DIP exhibits a stronger affinity for antioxidant genes like glutathione peroxidase (GPx), glutathione-S-transferase (GST), and glutathione reductase (GSR). Using in silco molecular docking analysis, the protein-ligand binding affinities of GSR (-4.70 kcal/mol), GST (-5.27 kcal/mol), and GPx (-5.37 kcal/mol) were measured. The expression of antioxidant genes were investigated by qRT-PCR. The in vivo reactive oxygen species production, lipid peroxidation, and cell death levels were significantly (p ≤ 0.05) increased in OA-induced group, but all these levels were significantly (p ≤ 0.05) decreased in the purified DIP pretreated group. Purified DIP from halophilic bacteria could thus be a useful treatment for neurological disorders associated with oxidative stress.

Antioxidant Defenses Against Air Humidity Stress in Fruit Bodies of Auricularia heimuer (Agaricomycetes).

Air humidity is an important environmental factor restricting the fruit body growth of Auricularia heimuer. Low air humidity causes the fruit body to desiccate and enter dormancy. However, the survival mechanisms to low air humidity for fruit bodies before dormancy remain poorly understood. In the present study, we cultivated A. heimuer in a greenhouse and collected the fruit bodies at different air humidities (90%, 80%, 70%, 60%, and 50%) to determine the contents of malondialdehyde (MDA) and non-enzymatic antioxidants such as ascorbic acid (AsA) and glutathione (GSH); and the activities of enzymatic antioxidants including superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX), glutathione peroxidase (GPX) and glutathione reductase (GR). Results showed that the MDA contents tended to increase with decreasing relative air humidity. Relative air humidity below 90% caused membrane lipid peroxidation and oxidative stress (based on MDA contents) to the fruit body, which we named air humidity stress. In contrast to the control and with the degree of stress, the GSH contents and activities of SOD, CAT, GR, GPX, and APX tended to ascend, whereas AsA showed a declining trend; the POD activity only rose at 50%. The antioxidants favored the fruit body to alleviate oxidative damage and strengthened its tolerance to air humidity stress. The antioxidant defense system could be an important mechanism for the fruit body of A. heimuer in air humidity stress.

Understanding baseline levels of physiological stress tolerance from excessive exercise in a holobenthic octopus species, Octopus pallidus.

Cephalopods receive a great deal of attention due to their socioeconomically important fisheries and aquaculture industries as well their unique biological features. However, basic information about their physiological responses under stress conditions is lacking. This study investigated the impact of a simple stressor, exercise to exhaustion, on the activity levels of antioxidant enzymes and the concentrations of molecules involved in oxidative stress response in the pale octopus (Octopus pallidus). Eight biochemical assays were measured in the humoral (plasma) and cellular (hemocyte) components of O. pallidus haemolymph, the invertebrate analogue to vertebrate blood. Overall, exercise resulted in an increase in activity of plasma catalase (CAT) and glutathione-S-transferase (GST) and the decrease in activity of plasms glutathione reductase (GR). In the hemocytes, the exercise elicited a different response, with a reduction in the activity of superoxide dismutase (SOD), GR, and glutathione peroxidase (GPX) and a reduction in nitric oxide (NO) concentration. Malondialdehyde (MDA) activity was similar in the plasma and haemocytes in control and exercised treatments, indicating that exercise did not induce lipid peroxidation. These results provide an important baseline for understanding oxidative stress in octopus, with exercise to exhaustion serving as a simple stressor which will ultimately inform our ability to detect and understand physiological responses to more complex stressors.

Modification in the ascorbate-glutathione cycle leads to a better acclimation to high light in the rose Bengal resistant mutant of Nannochloropsis oceanica.

Understanding how to adapt outdoor cultures of Nannochloropsis oceanica to high light (HL) is vital for boosting productivity. The N. oceanica RB2 mutant, obtained via ethyl methanesulfonate mutagenesis, was chosen for its tolerance to Rose Bengal (RB), a singlet oxygen (1O2) generator. Compared to the wild type (WT), the RB2 mutant showed higher resilience to excess light conditions. Analyzing the ascorbate-glutathione cycle (AGC), involving ascorbate peroxidases (APX, EC 1.11.1.11), dehydroascorbate reductase (DHAR, EC 1.8.5.1), and glutathione reductase (GR, EC 1.8.1.7), in the RB2 mutant under HL stress provided valuable insights. At 250 µmol photon m-2 s-1 (HL), the WT strain displayed superoxide anion radicals (O2▪-) and hydrogen peroxide (H2O2) accumulation, increased lipid peroxidation, and cell death compared to normal light (NL) conditions (50 µmol photon m-2 s-1). The RB2 mutant didn't accumulate O2▪- and H2O2 after HL exposure, and exhibited increased APX, DHAR, and GR activities and transcript levels compared to WT and remained consistent after HL treatment. Although the RB2 mutant had a smaller ascorbate (AsA) pool than the WT, its ability to regenerate dehydroascorbate (DHA) increased post HL exposure, indicated by a higher AsA/DHA ratio. Additionally, under HL conditions, the RB2 mutant displayed an improved glutathione (GSH) regeneration rate (GSH/GSSG ratio) without changing the GSH pool size. Remarkably, H2O2 or menadione (a O2▪- donor) treatment induced cell death in the WT strain but not in the RB2 mutant. These findings emphasize the essential role of AGC in the RB2 mutant of Nannochloropsis in handling photo-oxidative stress.

The effects of morin and methotrexate on pentose phosphate pathway enzymes and GR/GST/TrxR enzyme activities: An in vivo and in silico study.

In this study, the mechanisms by which the enzymes glucose-6-phosphate dehydrogenase (G6PD), 6-phosphogluconate dehydrogenase (6PGD), glutathione reductase (GR), glutathione-S-transferase (GST), and thioredoxin reductase (TrxR) are inhibited by methotrexate (MTX) were investigated, as well as whether the antioxidant morin can mitigate or prevent these adverse effects in vivo and in silico. For 10 days, rats received oral doses of morin (50 and 100 mg/kg body weight). On the fifth day, a single intraperitoneal injection of MTX (20 mg/kg body weight) was administered to generate toxicity. Decreased activities of G6PD, 6PGD, GR, GST, and TrxR were associated with MTX-related toxicity while morin treatment increased the activity of the enzymes. The docking analysis indicated that H-bonds, pi-pi stacking, and pi-cation interactions were the dominant interactions in these enzyme-binding pockets. Furthermore, the docked poses of morin and MTX against GST were subjected to molecular dynamic simulations for 200 ns, to assess the stability of both complexes and also to predict key amino acid residues in the binding pockets throughout the simulation. The results of this study suggest that morin may be a viable means of alleviating the enzyme activities of important regulatory enzymes against MTX-induced toxicity.

Effects of acute exposure to environmentally realistic tebuconazole concentrations on stress responses of kidney and digestive gland of Lymnaea stagnalis.

This study aimed to investigate the effects of 24 and 72 h exposure to environmentally relevant concentrations of tebuconazole (TEB) (10, 100 and 500 µg/L) fungicide on the freshwater snail Lymnaea stagnalis. The focus was induction of oxidative stress, alteration of gene expressions and histopathological changes in the kidney and digestive gland. TEB treatment induced a time- and concentration-dependent increase in intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) levels, while the total antioxidant capacity (TAC) was decreased. The activities of glutathione peroxidase (GPx), glutathione reductase (GR), and catalase (CAT) also increased in a time- and concentration-dependent manner in both tissues. TEB exposure significantly increased the mRNA levels of CAT, GPx, GR, heat shock proteins HSP40 and HSP70. Histological analysis revealed nephrocyte degeneration and disrupted digestive cells. The study concludes that acute exposure to TEB induces oxidative stress, alters antioxidant defense mechanisms, and leads to histopathological changes in L. stagnalis.

A genome-scale metabolic model of parasitic whipworm.

Genome-scale metabolic models are widely used to enhance our understanding of metabolic features of organisms, host-pathogen interactions and to identify therapeutics for diseases. Here we present iTMU798, the genome-scale metabolic model of the mouse whipworm Trichuris muris. The model demonstrates the metabolic features of T. muris and allows the prediction of metabolic steps essential for its survival. Specifically, that Thioredoxin Reductase (TrxR) enzyme is essential, a prediction we validate in vitro with the drug auranofin. Furthermore, our observation that the T. muris genome lacks gsr-1 encoding Glutathione Reductase (GR) but has GR activity that can be inhibited by auranofin indicates a mechanism for the reduction of glutathione by the TrxR enzyme in T. muris. In addition, iTMU798 predicts seven essential amino acids that cannot be synthesised by T. muris, a prediction we validate for the amino acid tryptophan. Overall, iTMU798 is as a powerful tool to study not only the T. muris metabolism but also other Trichuris spp. in understanding host parasite interactions and the rationale design of new intervention strategies.

The role of salicylic acid on glutathione metabolism under endoplasmic reticulum stress in tomato.

The endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) are highly dependent on phytohormones such as salicylic acid (SA). In this study, the effect of SA supplementation and the lack of endogenous SA on glutathione metabolism were investigated under ER stress in wild-type (WT) and transgenic SA-deficient NahG tomato (Solanum lycopersicum L.) plants. The expression of the UPR marker gene SlBiP was dependent on SA levels and remained lower in NahG plants. Exogenous application of the chemical chaperone 4-phenylbutyrate (PBA) also reduced tunicamycin (Tm)-induced SlBiP transcript accumulation. At the same time, Tm-induced superoxide and hydrogen peroxide production were independent of SA, whereas the accumulation of reduced form of glutathione (GSH) and the oxidised glutathione (GSSG) was regulated by SA. Tm increased the activity of glutathione reductase (GR; EC 1.6.4.2) independently of SA, but the activities of dehydroascorbate reductase (DHAR; EC 1.8.5.1) and glutathione S-transferases (GSTs; EC 2.5.1.18) were increased by Tm in a SA-dependent manner. SlGR2, SlGGT and SlGSTT2 expression was activated in a SA-dependent way upon Tm. Although expression of SlGSH1, SlGSTF2, SlGSTU5 and SlGTT3 did not change upon Tm treatment in leaves, SlGR1 and SlDHAR2 transcription decreased. PBA significantly increased the expression of SlGR1, SlGR2, SlGSTT2, and SlGSTT3, which contributed to the amelioration of Tm-induced ER stress based on the changes in lipid peroxidation and cell viability. Malondialdehyde accumulation and electrolyte leakage were significantly higher in WT as compared to NahG tomato leaves under ER stress, further confirming the key role of SA in this process.

Cold acclimation alleviates photosynthetic inhibition and oxidative damage induced by cold stress in citrus seedlings.

Cold stress seriously inhibits plant growth and development, geographical distribution, and yield stability of plants. Cold acclimation (CA) is an important strategy for modulating cold stress, but the mechanism by which CA induces plant resistance to cold stress is still not clear. The purpose of this study was to investigate the effect of CA treatment on the cold resistance of citrus seedlings under cold stress treatment, and to use seedlings without CA treatment as the control (NA). The results revealed that CA treatment increased the content of photosynthetic pigments under cold stress, whereas cold stress greatly reduced the value of gas exchange parameters. CA treatment also promoted the activity of Rubisco and FBPase, as well as led to an upregulation of the transcription levels of photosynthetic related genes (rbcL and rbcS)，compared to the NA group without cold stress. In addition, cold stress profoundly reduced photochemical chemistry of photosystem II (PSII), especially the maximum quantum efficiency (Fv/Fm) in PSII. Conversely, CA treatment improved the chlorophyll a fluorescence parameters, thereby improving electron transfer efficiency. Moreover, under cold stress, CA treatment alleviated oxidative stress damage to cell membranes by inhibiting the concentration of H2O2 and MDA, enhancing the activities of superoxide dismutase (SOD), catalase (CAT), ascorbic acid peroxidase (APX) and glutathione reductase (GR), accompanied by an increase in the expression level of antioxidant enzyme genes (CuZnSOD1, CAT1, APX and GR). Additionally, CA also increased the contents of abscisic acid (ABA) and salicylic acid (SA) in plants under cold stress. Overall, we concluded that CA treatment suppressed the negative effects of cold stress by enhancing photosynthetic performance, antioxidant enzymes functions and plant hormones contents.

Nephroprotective effect of vitamin D Against Levofloxacin-induced renal injury: an observational study.

The pathogenesis of kidney damage involves complicated interactions between vascular endothelial and tubular cell destruction. Evidence has shown that vitamin D may have anti-inflammatory effects in several models of kidney damage. In this study, we evaluated the effects of synthetic vitamin D on levofloxacin-induced renal injury in rats. Forty-two white Albino rats were divided into six groups, with each group comprising seven rats. Group I served as the control (negative control) and received intraperitoneal injections of normal saline (0.5 ml) once daily for twenty-one days. Group II and Group III were treated with a single intraperitoneal dose of Levofloxacin (50 mg/kg/day) and (100 mg/kg/day), respectively, for 14 days (positive control groups). Group IV served as an additional negative control and received oral administration of vitamin D3 (500 IU/rat/day) for twenty-one days. In Group V, rats were orally administered vitamin D3 (500 IU/rat/day) for twenty-one days, and intraperitoneal injections of Levofloxacin (50 mg/kg/day) were administered on day 8 for 14 days. Group VI received oral vitamin D3 supplementation (500 IU/rat/day) for twenty-one days, followed by intraperitoneal injections of Levofloxacin (100 mg/kg/day) on day 8 for fourteen days. Blood samples were collected to measure creatinine, urea, malondialdehyde, glutathione reductase, and superoxide dismutase levels. Compared to the positive control group, vitamin D supplementation lowered creatinine, urea, and malondialdehyde levels, while increasing glutathione reductase and superoxide dismutase levels. Urea, creatinine, and malondialdehyde levels were significantly (p<0.05) higher in rats administered LFX 50mg and 100mg compared to rats given (LFX + vitamin D). The main findings of this study show that vitamin D reduces renal dysfunction, suggesting that vitamin D has antioxidant properties and may be used to prevent renal injury.

Identification of Streptococcus pneumoniae genes associated with hypothiocyanous acid tolerance through genome-wide screening.

Streptococcus pneumoniae is a commensal bacterium and invasive pathogen that causes millions of deaths worldwide. The pneumococcal vaccine offers limited protection, and the rise of antimicrobial resistance will make treatment increasingly challenging, emphasizing the need for new antipneumococcal strategies. One possibility is to target antioxidant defenses to render S. pneumoniae more susceptible to oxidants produced by the immune system. Human peroxidase enzymes will convert bacterial-derived hydrogen peroxide to hypothiocyanous acid (HOSCN) at sites of colonization and infection. Here, we used saturation transposon mutagenesis and deep sequencing to identify genes that enable S. pneumoniae to tolerate HOSCN. We identified 37 genes associated with S. pneumoniae HOSCN tolerance, including genes involved in metabolism, membrane transport, DNA repair, and oxidant detoxification. Single-gene deletion mutants of the identified antioxidant defense genes sodA, spxB, trxA, and ahpD were generated and their ability to survive HOSCN was assessed. With the exception of ΔahpD, all deletion mutants showed significantly greater sensitivity to HOSCN, validating the result of the genome-wide screen. The activity of hypothiocyanous acid reductase or glutathione reductase, known to be important for S. pneumoniae tolerance of HOSCN, was increased in three of the mutants, highlighting the compensatory potential of antioxidant systems. Double deletion of the gene encoding glutathione reductase and sodA sensitized the bacteria significantly more than single deletion. The HOSCN defense systems identified in this study may be viable targets for novel therapeutics against this deadly pathogen. IMPORTANCE Streptococcus pneumoniae is a human pathogen that causes pneumonia, bacteremia, and meningitis. Vaccination provides protection only against a quarter of the known S. pneumoniae serotypes, and the bacterium is rapidly becoming resistant to antibiotics. As such, new treatments are required. One strategy is to sensitize the bacteria to killing by the immune system. In this study, we performed a genome-wide screen to identify genes that help this bacterium resist oxidative stress exerted by the host at sites of colonization and infection. By identifying a number of critical pneumococcal defense mechanisms, our work provides novel targets for antimicrobial therapy.

Melatonin enhanced the heavy metal-stress tolerance of pepper by mitigating the oxidative damage and reducing the heavy metal accumulation.

Heavy metals (HMs), like vanadium (V), chromium (Cr), cadmium (Cd), and nickel (Ni) toxicity due to anthropogenic, impair plant growth and yield, which is a challenging issue for agricultural production. Melatonin (ME) is a stress mitigating molecule, which alleviates HM-induced phytotoxicity, but the possible underlying mechanism of ME functions under HMs' phytotoxicity is still unclear. Current study uncovered key mechanisms for ME-mediated HMs-stress tolerance in pepper. HMs toxicity greatly reduced growth by impeding leaf photosynthesis, root architecture system, and nutrient uptake. Conversely, ME supplementation markedly enhanced growth attributes, mineral nutrient uptake, photosynthetic efficiency, as measured by chlorophyll content, gas exchange elements, chlorophyll photosynthesis genes' upregulation, and reduced HMs accumulation. ME treatment showed a significant decline in the leaf/root V, Cr, Ni, and Cd concentration which was about 38.1/33.2%, 38.5/25.9%, 34.8/24.9%, and 26.6/25.1%, respectively, when compared with respective HM treatment. Furthermore, ME remarkably reduced the ROS (reactive oxygen species) accumulation, and reinstated the integrity of cellular membrane via activating antioxidant enzymes (SOD, superoxide dismutase; CAT, catalase; APX, ascorbate peroxidase; GR, glutathione reductase; POD, peroxidase; GST, glutathione S-transferase; DHAR, dehydroascorbate reductase; MDHAR, monodehydroascorbate reductase) and as well as regulating ascorbate-glutathione (AsA-GSH) cycle. Importantly, oxidative damage showed efficient alleviations through upregulating the genes related to key defense such as SOD, CAT, POD, GR, GST, APX, GPX, DHAR, and MDHAR; along with the genes related to ME biosynthesis. ME supplementation also enhanced the level of proline and secondary metabolites, and their encoding genes expression, which may control excessive H2O2 (hydrogen peroxide) production. Finally, ME supplementation enhanced the HM stress tolerance of pepper seedlings.

Redox perturbations in yeast cells lacking glutathione reductase.

Cellular redox homeostasis has a major effect on cell functions and its maintenance is supported by glutathione and protein thiols which serve as redox buffers in cells. The regulation of the glutathione biosynthetic pathway is a focus of a lot of scientific research. However, still little is known about how complex cellular networks influence glutathione homeostasis. In this work was used an experimental system based on an S. cerevisiae yeast mutant with a lack of the glutathione reductase enzyme and allyl alcohol as a precursor of acrolein inside the cell to determine the cellular processes influencing glutathione homeostasis. The absence of Glr1p slows down the growth rate of the cell population, especially in the presence of allyl alcohol, but does not lead to complete inhibition of the cell's reproductive capacity. It also amends the GSH/GSSG ratio and the share of NADPH and NADP+ in the total NADP(H) pool. The obtained results show that potential pathways involved in the maintenance of redox homeostasis are based from one side on de novo synthesis of GSH as indicated by increased activity of γ-GCS and increased expression of GSH1 gene in the Δglr1 mutant, from the other hand, on increased the level of NADPH. This is because the lower ratio of GSH/GSSG can be counterbalanced with the NADPH/NADP+ alternative system. The higher level of NADPH can be used by the thioredoxin system and other enzymes requiring NADPH to reduce cytosolic GSSG and maintain glutathione redox potential.

GR1 and NTRA involved in pollen tube growth in the stigma of Arabidopsis.

MAIN CONCLUSION: Arabidopsis GR1 and NTRA function in pollen tube penetrating the stigma into the transmitting tract during pollination. During pollination, recognition between pollen (tube) and stigma mediates the hydration and germination of pollen, as well as the growth of the pollen tube on the stigma. Arabidopsis glutathione reductase 1 (GR1) and NADPH-dependent thioredoxin reductase A (NTRA) are involved in regulating cell redox hemostasis. Both GR1 and NTRA are expressed in pollen, but their roles in pollen germination and the growth of the pollen tube need further investigation. In this study, we performed pollination experiments and found that the Arabidopsis gr1/ + ntra/- and gr1/- ntra/ + double mutation compromised the transmission of male gametophytes. Pollen morphology and viability of the mutants did not show obvious abnormalities. Additionally, the pollen hydration and germination of the double mutants on solid pollen germination medium were comparable to those of the wild type. However, the pollen tubes with gr1 ntra double mutation were unable to penetrate the stigma and enter the transmitting tract when they grew on the surface of the stigma. Our results indicate that GR1 and NTRA play a role in regulating the interaction between the pollen tube and the stigma during pollination.

Structure-Activity Relationship of Methyl 4-Aminobenzoate Derivatives as Being Drug Candidate Targeting Glutathione Related Enzymes: in Vitro and in Silico Approaches.

A thiol compound, glutathione, is essential for healthy cell defence against xenobiotics and oxidative stress. Glutathione reductase (GR) and glutathione S-transferase (GST) are two glutathione-related enzymes that function in the antioxidant and the detoxification systems. In this study, potential inhibitory effects of methyl 4-aminobenzoate derivatives on GR and GST were examined in vitro. GR and GST were isolated from human erythrocytes with 7.63 EU/mg protein and 5.66 EU/mg protein specific activity, respectively. It was found that compound 1 (methyl 4-amino-3-bromo-5-fluorobenzoate with Ki value of 0.325±0.012 µM) and compound 5 (methyl 4-amino-2-nitrobenzoate with Ki value of 92.41±22.26 µM) inhibited GR and GST stronger than other derivatives. Furthermore, a computer-aided method was used to predict the binding affinities of derivatives, ADME characteristics, and toxicities. Derivatives 4 (methyl 4-amino-2-bromobenzoate) and 6 (methyl 4-amino-2-chlorobenzoate) were estimated to have the lowest binding energies into GR and GST receptors, respectively according to results of in silico studies.

Descriptive Analysis of Transient-State Observations for Thioredoxin/Glutathione Reductase (Sec597Cys) from Schistosoma mansoni.

Thioredoxin/glutathione reductase from Schistosoma mansoni (SmTGR) catalyzes the reduction of both oxidized thioredoxin and glutathione with electrons from reduced nicotinamide adenine dinucleotide phosphate (NADPH). SmTGR is a drug target for the treatment of Schistosomiasis, an infection caused by Schistosoma platyhelminths residing in the blood vessels of the host. Schistosoma spp. are reliant on TGR enzymes as they lack catalase and so use reduced thioredoxin and glutathione to regenerate peroxiredoxins consumed in the detoxification of reactive oxygen species. SmTGR is a flavin adenine dinucleotide (FAD)-dependent enzyme, and we have used the flavin as a spectrophotometric reporter to observe the movement of electrons within the enzyme. The data show that NADPH fractionally reduces the active site flavin with an observed rate constant estimated in this study to be ∼3000 s-1. The flavin then reoxidizes by passing electrons at a similar rate to the proximal Cys159-Cys154 disulfide pair. The dissociation of NADP+ occurs with a rate of ∼180 s-1, which induces the deprotonation of Cys159, and this coincides with the accumulation of an intense FAD-thiolate charge transfer band. It is proposed that the electrons then pass to the Cys596-Cys597 disulfide pair of the associated subunit in the dimer with a net rate constant of ∼2 s-1. (Note: Cys597 is Sec597 in wild-type (WT) SmTGR.) From this position, the electrons can be passed to oxidized thioredoxin or further into the protein to reduce the Cys28-Cys31 disulfide pair of the originating subunit of the dimer. From the Cys28-Cys31 center, electrons can then pass to oxidized glutathione that has a binding site directly adjacent.

Purification and characterisation of glutathione reductase from scorpionfish (scorpaena porcus) and investigation of heavy metal ions inhibition.

In the current study, glutathione reductase was purified from Scorpion fish (Scorpaena porcus) liver tissue and the effects of heavy metal ions on the enzyme activity were determined. The purification process consisted of three stages; preparation of the homogenate, ammonium sulphate precipitation and affinity chromatography purification. At the end of these steps, the enzyme was purified 25.9-fold with a specific activity of 10.479 EU/mg and a yield of 28.3%. The optimum pH was found to be 6.5, optimum substrate concentration was 2 mM NADPH and optimum buffer was 300 mM KH2PO4. After purification, inhibition effects of Mn+2, Cd+2, Ni+2, and Cr3+, as heavy metal ions were investigated. IC50 values of the heavy metals were calculated as 2.4 µM, 30 µM, 135 µM and 206 µM, respectively.

Age-related changes in antioxidant defenses of the Mediterranean centipede Scolopendra cingulata (Chilopoda).

The activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), glutathione reductase (GR), and glutathione S-transferase (GST), as well as the concentrations of sulfhydryl (SH) groups and glutathione (GSH) were analyzed in five age classes of the Mediterranean centipede Scolopendra cingulata as follows: embryo, adolescens, maturus junior, maturus, and maturus senior. The data obtained showed the presence of SOD, CAT, GSH-Px, GR, GST, and SH groups in embryos. The transition from embryo to adolescens was accompanied by an increase in the activities of all studied enzymes, in response to the increased production of ROS due to the increased metabolic activity of the centipede associated with growth and development. Our results show that trends in antioxidant enzyme (AOE) activities were not uniform among adult age classes, suggesting that maturus junior, maturus, and maturus senior differentially respond and/or have different susceptibility to ROS. On the other hand, GSH concentration in embryos was undetectable, highest in adolescens and decreased in the latter part of life. Pearson correlation analysis in embryos showed that the activities of the AOEs were strongly and positively correlated with each other but negatively correlated with GSH and SH groups. At later age classes, SOD, CAT, GSH-Px, GR, GSH, and SH groups were no longer significantly correlated with GST. In the discriminant analysis, the variables that separated the age classes were GR, GST, SH groups, and body length. Body length was directly related to the age of individuals, clearly indicating that development/aging affects the regulation of antioxidant defense in this species.

Overexpression of CcFALDH from spider plant (Chlorophytum comosum) enhances the formaldehyde removing capacity of transgenic gloxinia (Sinningia speciosa)1.

Formaldehyde can cause leukemia and nasopharyngeal cancer in humans, and is a major indoor air pollutant. In this study, to improve the ability of flowering plants to purify formaldehyde, we cloned the CcFALDH gene encoding formaldehyde dehydrogenase (FALDH) from the spider plant (Chlorophytum comosum), which encodes 379 amino acids with the alcohol dehydrogenase (ADH) structural domain, and used it to transform the flowering plant gloxinia (Sinningia speciosa). The FALDH activity of transgenic gloxinia was 1.8-2.7 times that of wild-type (WT) with a considerable increase in formaldehyde stress tolerance. The activities of the antioxidant enzymes SOD, POD, and CAT of transgenic gloxinia were 1.5-2.0 times those of the WT under formaldehyde stress; H2O2, O2-, and MDA contents were markedly lower than those in WT. Liquid formaldehyde and gaseous formaldehyde were metabolized at 2.1-2.8 and 2.1-2.7 times higher rates in transgenic gloxinia than in WT. Our findings indicate that overexpression of CcFALDH can enhance the capacity of flowering plants to metabolize formaldehyde, which provides a new strategy to tackle the indoor formaldehyde pollution problem.

Overexpression of tomato glutathione reductase (SlGR) in transgenic tobacco enhances salt tolerance involving the S-nitrosylation of GR.

S-nitrosylation, a post-translational modification (PTM) dependent on nitric oxide, is essential for plant development and environmental responsiveness. However, the function of S-nitrosylation of glutathione reductase (GR) in tomato (SlGR) under NaCl stress is yet uncertain. In this study, sodium nitroprusside (SNP), an exogenous NO donor, alleviated the growth inhibition of tomato under NaCl treatment, particularly at 100 µM. Following NaCl treatment, the transcripts, enzyme activity, and S-nitrosylated level of GR were increased. In vitro, the SlGR protein was able to be S-nitrosylated by S-nitrosoglutathione (GSNO), significantly increasing the activity of GR. SlGR overexpression transgenic tobacco plants exhibited enhanced germination rate, fresh weight, and increased root length in comparison to wild-type (WT) seedlings. The accumulation of reactive oxygen species (ROS) was lower, whereas the expression and activities of GR, ascorbate peroxidase (APX), superoxide dismutase (SOD), and catalase (CAT); the ratio of ascorbic acid/dehydroascorbic acid (AsA/DHA), reduced glutathione/oxidized glutathione (GSH/GSSG), total soluble sugar and proline contents; and the expression of stress-related genes were higher in SlGR overexpression transgenic plants in comparison to the WT plants following NaCl treatment. The accumulation of NO and S-nitrosylated levels of GR in transgenic plants was higher in comparison to WT plants following NaCl treatment. These results indicated that S-nitrosylation of GR played a significant role in salt tolerance by regulating the oxidative state.