-
International Journal of Molecular... Jan 2022Aluminum (Al) toxicity is the main factor limiting plant growth and the yield of cereal crops in acidic soils. Al-induced oxidative stress could lead to the excessive...
Aluminum (Al) toxicity is the main factor limiting plant growth and the yield of cereal crops in acidic soils. Al-induced oxidative stress could lead to the excessive accumulation of reactive oxygen species (ROS) and aldehydes in plants. Aldehyde dehydrogenase () genes, which play an important role in detoxification of aldehydes when exposed to abiotic stress, have been identified in most species. However, little is known about the function of this gene family in the response to Al stress. Here, we identified an gene in maize, , involved in protection against Al-induced oxidative stress. Al stress up-regulated expression in both the roots and leaves. The expression of only responded to Al toxicity but not to other stresses including low pH and other metals. The heterologous overexpression of in increased Al tolerance by promoting the ascorbate-glutathione cycle, increasing the transcript levels of antioxidant enzyme genes as well as the activities of their products, reducing MDA, and increasing free proline synthesis. The overexpression of also reduced Al accumulation in roots. Taken together, these findings suggest that participates in Al-induced oxidative stress and Al accumulation in roots, conferring Al tolerance in transgenic .
Topics: Adaptation, Physiological; Aldehyde Dehydrogenase; Aluminum; Amino Acid Sequence; Antioxidants; Arabidopsis; Ascorbate Peroxidases; Ascorbic Acid; Cloning, Molecular; Gene Expression Regulation, Plant; Genes, Plant; Glutathione; Glutathione Reductase; Hydrogen Peroxide; Lipid Peroxidation; Oxidative Stress; Phylogeny; Plant Leaves; Plant Roots; Plants, Genetically Modified; Proline; RNA, Messenger; Subcellular Fractions; Superoxides; Nicotiana; Zea mays
PubMed: 35008903
DOI: 10.3390/ijms23010477 -
Molecules (Basel, Switzerland) Jan 2021YqhD, an alcohol/aldehyde oxidoreductase, is an enzyme able to produce valuable bio-renewable fuels and fine chemicals from a broad range of starting materials. Herein,...
YqhD, an alcohol/aldehyde oxidoreductase, is an enzyme able to produce valuable bio-renewable fuels and fine chemicals from a broad range of starting materials. Herein, we report the first computational solution-phase structure-dynamics analysis of YqhD, shedding light on the effect of oxidized and reduced NADP/H cofactor binding on the conformational dynamics of the biocatalyst using molecular dynamics (MD) simulations. The cofactor oxidation states mainly influence the interdomain cleft region conformations of the YqhD monomers, involved in intricate cofactor binding and release. The ensemble of NADPH-bound monomers has a narrower average interdomain space resulting in more hydrogen bonds and rigid cofactor binding. NADP-bound YqhD fluctuates between open and closed conformations, while it was observed that NADPH-bound YqhD had slower opening/closing dynamics of the cofactor-binding cleft. In the light of enzyme kinetics and structural data, simulation findings have led us to postulate that the frequently sampled open conformation of the cofactor binding cleft with NADP leads to the more facile release of NADP while increased closed conformation sampling during NADPH binding enhances cofactor binding affinity and the aldehyde reductase activity of the enzyme.
Topics: Aldehyde Reductase; Binding Sites; Coenzymes; Escherichia coli; Escherichia coli Proteins; Hydrogen Bonding; Molecular Conformation; Molecular Docking Simulation; Molecular Dynamics Simulation; NADP; Protein Binding; Protein Interaction Domains and Motifs; Structure-Activity Relationship; Substrate Specificity
PubMed: 33430436
DOI: 10.3390/molecules26020270 -
The Journal of Biological Chemistry Nov 2019Oxidative modification of Cys residues by NO results in -nitrosylation, a ubiquitous post-translational modification and a primary mediator of redox-based cellular...
Oxidative modification of Cys residues by NO results in -nitrosylation, a ubiquitous post-translational modification and a primary mediator of redox-based cellular signaling. Steady-state levels of -nitrosylated proteins are largely determined by denitrosylase enzymes that couple NAD(P)H oxidation with reduction of -nitrosothiols, including protein and low-molecular-weight (LMW) -nitrosothiols (-nitroso-GSH (GSNO) and -nitroso-CoA (SNO-CoA)). SNO-CoA reductases require NADPH, whereas enzymatic reduction of GSNO can involve either NADH or NADPH. Notably, GSNO reductase (GSNOR, ) accounts for most NADH-dependent GSNOR activity, whereas NADPH-dependent GSNOR activity is largely unaccounted for (CBR1 mediates a minor portion). Here, we purified NADPH-coupled GSNOR activity from mammalian tissues and identified aldo-keto reductase family 1 member A1 (AKR1A1), the archetypal mammalian SNO-CoA reductase, as a primary mediator of NADPH-coupled GSNOR activity in these tissues. Kinetic analyses suggested an AKR1A1 substrate preference of SNO-CoA > GSNO. AKR1A1 deletion from murine tissues dramatically lowered NADPH-dependent GSNOR activity. Conversely, GSNOR-deficient mice had increased AKR1A1 activity, revealing potential cross-talk among GSNO-dependent denitrosylases. Molecular modeling and mutagenesis of AKR1A1 identified Arg-312 as a key residue mediating the specific interaction with GSNO; in contrast, substitution of the SNO-CoA-binding residue Lys-127 minimally affected the GSNO-reducing activity of AKR1A1. Together, these findings indicate that AKR1A1 is a multi-LMW-SNO reductase that can distinguish between and metabolize the two major LMW-SNO signaling molecules GSNO and SNO-CoA, allowing for wide-ranging control of protein -nitrosylation under both physiological and pathological conditions.
Topics: Aldehyde Oxidoreductases; Aldehyde Reductase; Animals; Coenzyme A; Humans; Kinetics; Mammals; Mice, Inbred C57BL; Mice, Knockout; NADP; Nitric Oxide; Oxidation-Reduction; S-Nitrosothiols; Signal Transduction
PubMed: 31649033
DOI: 10.1074/jbc.RA119.011067 -
Scientific Reports Apr 2022The role of aldose reductase (ALR2) in causing diabetic complications is well-studied, with overactivity of ALR2 in the hyperglycemic state leading to an accumulation of...
The role of aldose reductase (ALR2) in causing diabetic complications is well-studied, with overactivity of ALR2 in the hyperglycemic state leading to an accumulation of intracellular sorbitol, depletion of cytoplasmic NADPH and oxidative stress and causing a variety of different conditions including retinopathy, nephropathy, neuropathy and cardiovascular disorders. While previous efforts have sought to develop inhibitors of this enzyme in order to combat diabetic complications, non-selective inhibition of both ALR2 and the homologous enzyme aldehyde reductase (ALR1) has led to poor toxicity profiles, with no drugs targeting ALR2 currently approved for therapeutic use in the Western world. In the current study, we have synthesized a series of N-substituted thiosemicarbazones with added phenolic moieties, of which compound 3m displayed strong and selective ALR2 inhibitory activity in vitro (IC 1.18 µM) as well as promising antioxidant activity (75.95% free radical scavenging activity). The target binding modes of 3m were studied via molecular docking studies and stable interactions with ALR2 were inferred through molecular dynamics simulations. We thus report the N-substituted thiosemicarbazones as promising drug candidates for selective inhibition of ALR2 and possible treatment of diabetic complications.
Topics: Aldehyde Reductase; Diabetes Complications; Enzyme Inhibitors; Humans; Molecular Docking Simulation; Structure-Activity Relationship; Thiosemicarbazones
PubMed: 35388067
DOI: 10.1038/s41598-022-09658-z -
Antioxidants (Basel, Switzerland) May 2023Fanconi anemia (FA) is a rare genetic disease characterized by a dysfunctional DNA repair and an oxidative stress accumulation due to defective mitochondrial energy...
Fanconi anemia (FA) is a rare genetic disease characterized by a dysfunctional DNA repair and an oxidative stress accumulation due to defective mitochondrial energy metabolism, not counteracted by endogenous antioxidant defenses, which appear down-expressed compared to the control. Since the antioxidant response lack could depend on the hypoacetylation of genes coding for detoxifying enzymes, we treated lymphoblasts and fibroblasts mutated for the gene with some histone deacetylase inhibitors (HDACi), namely, valproic acid (VPA), beta-hydroxybutyrate (OHB), and EX527 (a Sirt1 inhibitor), under basal conditions and after hydrogen peroxide addition. The results show that VPA increased catalase and glutathione reductase expression and activity, corrected the metabolic defect, lowered lipid peroxidation, restored the mitochondrial fusion and fission balance, and improved mitomycin survival. In contrast, OHB, despite a slight increase in antioxidant enzyme expressions, exacerbated the metabolic defect, increasing oxidative stress production, probably because it also acts as an oxidative phosphorylation metabolite, while EX527 showed no effect. In conclusion, the data suggest that VPA could be a promising drug to modulate the gene expression in FA cells, confirming that the antioxidant response modulation plays a pivotal in FA pathogenesis as it acts on both oxidative stress levels and the mitochondrial metabolism and dynamics quality.
PubMed: 37237966
DOI: 10.3390/antiox12051100 -
RSC Advances Jan 2020To avoid random chemical linkage and achieve precisely directed immobilization, mutant enzymes were obtained and immobilized using an incorporated reactive nonstandard...
To avoid random chemical linkage and achieve precisely directed immobilization, mutant enzymes were obtained and immobilized using an incorporated reactive nonstandard amino acid (NSAA). For this purpose, aldehyde ketone reductase (AKR) was used as a model enzyme, and 110Y, 114Y, 143Y, 162Q and 189Q were each replaced with -azido-l-phenylalanine (pAzF). Then, the mutant AKR was coupled to the functionalized support by strain-promoted alkyne-azide cycloaddition (SPAAC). The effects of the incorporation number and site of NSAAs on the loading and thermal stability of the immobilized AKR were examined. The results show that the mutant enzymes presented better specific activity than the wild type, except for AKR-110Y, and AKR-114Y showed 1.16-fold higher activity than the wild type. Moreover, the half-life ( ) of the five-point immobilized AKR reached 106 h and 45 h, 13 and 7 times higher than that of the free enzyme at 30 °C and 60 °C, respectively. Comparison of these three types of enzymes shows that multi-point immobilization provides improved loading and thermal stability and facilitates one-step purification. We expect this platform to facilitate a fundamental understanding of precisely oriented and controllable covalent immobilization and enable bio-manufacturing paradigms for fine chemicals and pharmaceuticals.
PubMed: 35496112
DOI: 10.1039/c9ra09067c -
Bio-protocol Apr 2020Microbial production of alkanes employing synthetic biology tools has gained tremendous attention owing to the high energy density and similarity of alkanes to existing...
Microbial production of alkanes employing synthetic biology tools has gained tremendous attention owing to the high energy density and similarity of alkanes to existing petroleum fuels. One of the most commonly studied pathways includes the production of alkanes by AAR (acyl-ACP (acyl carrier protein) reductase)-ADO (aldehyde deformylating oxygenase) pathway. Here, the intermediates of fatty acid synthesis pathway are used as substrate by the AAR enzyme to make fatty aldehyde, which is then deformylated by ADO to make linear chain alkane. However, the variation in substrate availability to the first enzyme of the pathway, , AAR, via fatty acid synthesis pathway and low turnover of the ADO enzyme make calculation of yields and titers under conditions extremely difficult. assay employing external addition of defined substrates for ADO enzyme into the medium helps to monitor the influx of substrate hence providing a more accurate measurement of the product yields. In this protocol, we include a detailed guide for implementing the assay for monitoring alkane production in .
PubMed: 33659559
DOI: 10.21769/BioProtoc.3593 -
Frontiers in Plant Science 2022The plant is well known for its production of artemisinin, a sesquiterpene lactone that is an effective antimalarial compound. Although remarkable progress has been...
The plant is well known for its production of artemisinin, a sesquiterpene lactone that is an effective antimalarial compound. Although remarkable progress has been made toward understanding artemisinin biosynthesis, the effect of MADS-box family transcription factors on artemisinin biosynthesis is still poorly understood. In this study, we identified a MADS transcription factor, AaSEP4, that was predominantly expressed in trichome. AaSEP4 acts as a nuclear-localized transcriptional activator activating the expression of . Dual-luciferase and Yeast one-hybrid assays revealed that AaSEP4 directly bound to the CArG motif in the promoter region of . Overexpression of in significantly induced the expression of and four artemisinin biosynthesis genes, including , , and . Furthermore, the results of high-performance liquid chromatography (HPLC) showed that the artemisinin content was significantly increased in the overexpressed plants. In addition, RT-qPCR results showed that was induced by methyl jasmonic acid (MeJA) treatment. Taken together, these results explicitly demonstrate that AaSEP4 is a positive regulator of artemisinin biosynthesis, which can be used in the development of high-artemisinin yielding varieties.
PubMed: 36119604
DOI: 10.3389/fpls.2022.982317 -
Lipids in Health and Disease Jun 2024Nonalcoholic steatohepatitis (NASH) is a prevalent chronic liver condition. However, the potential therapeutic benefits and underlying mechanism of nicotinate-curcumin...
BACKGROUND
Nonalcoholic steatohepatitis (NASH) is a prevalent chronic liver condition. However, the potential therapeutic benefits and underlying mechanism of nicotinate-curcumin (NC) in the treatment of NASH remain uncertain.
METHODS
A rat model of NASH induced by a high-fat and high-fructose diet was treated with nicotinate-curcumin (NC, 20, 40 mg·kg), curcumin (Cur, 40 mg·kg) and metformin (Met, 50 mg·kg) for a duration of 4 weeks. The interaction between NASH, Cur and Aldo-Keto reductase family 1 member B10 (AKR1B10) was filter and analyzed using network pharmacology. The interaction of Cur, NC and AKR1B10 was analyzed using molecular docking techniques, and the binding energy of Cur and NC with AKR1B10 was compared. HepG2 cells were induced by Ox-LDL (25 µg·ml, 24 h) in high glucose medium. NC (20µM, 40µM), Cur (40µM) Met (150µM) and epalrestat (Epa, 75µM) were administered individually. The activities of ALT, AST, ALP and the levels of LDL, HDL, TG, TC and FFA in serum were quantified using a chemiluminescence assay. Based on the changes in the above indicators, score according to NAS standards. The activities of Acetyl-CoA and Malonyl-CoA were measured using an ELISA assay. And the expression and cellular localization of AKR1B10 and Acetyl-CoA carboxylase (ACCα) in HepG2 cells were detected by Western blotting and immunofluorescence.
RESULTS
The results of the animal experiments demonstrated that NASH rat model induced by a high-fat and high-fructose diet exhibited pronounced dysfunction in liver function and lipid metabolism. Additionally, there was a significant increase in serum levels of FFA and TG, as well as elevated expression of AKR1B10 and ACCα, and heightened activity of Acetyl-CoA and Malonyl-CoA in liver tissue. The administration of NC showed to enhance liver function in rats with NASH, leading to reductions in ALT, AST and ALP levels, and decrease in blood lipid and significant inhibition of FFA and TG synthesis in the liver. Network pharmacological analysis identified AKR1B10 and ACCα as potential targets for NASH treatment. Molecular docking studies revealed that both Cur and NC are capable of binding to AKR1B10, with NC exhibiting a stronger binding energy to AKR1B10. Western blot analysis demonstrated an upregulation in the expression of AKR1B10 and ACCα in the liver tissue of NASH rats, accompanied by elevated Acetyl-CoA and Malonyl-CoA activity, and increased levels of FFA and TG. The results of the HepG2 cell experiments induced by Ox-LDL suggest that NC significantly inhibited the expression and co-localization of AKR1B10 and ACCα, while also reduced levels of TC and LDL-C and increased level of HDL-C. These effects are accompanied by a decrease in the activities of ACCα and Malonyl-CoA, and levels of FFA and TG. Furthermore, the impact of NC appears to be more pronounced compared to Cur.
CONCLUSION
NC could effectively treat NASH and improve liver function and lipid metabolism disorder. The mechanism of NC is related to the inhibition of AKR1B10/ACCα pathway and FFA/TG synthesis of liver.
Topics: Curcumin; Non-alcoholic Fatty Liver Disease; Animals; Humans; Hep G2 Cells; Aldo-Keto Reductases; Rats; Male; Triglycerides; Acetyl-CoA Carboxylase; Aldehyde Reductase; Diet, High-Fat; Molecular Docking Simulation; Liver; Metformin; Rats, Sprague-Dawley; Disease Models, Animal; Rhodanine; Thiazolidines
PubMed: 38937844
DOI: 10.1186/s12944-024-02162-5 -
Scientific Reports Feb 2023Cocultures of engineered thermophilic bacteria can ferment lignocellulose without costly pretreatment or added enzymes, an ability that can be exploited for low cost...
Cocultures of engineered thermophilic bacteria can ferment lignocellulose without costly pretreatment or added enzymes, an ability that can be exploited for low cost biofuel production from renewable feedstocks. The hemicellulose-fermenting species Thermoanaerobacterium thermosaccharolyticum was engineered for high ethanol yield, but we found that the strains switched from growth-coupled production of ethanol to growth uncoupled production of acetate and 1,2-propanediol upon growth cessation, producing up to 6.7 g/L 1,2-propanediol from 60 g/L cellobiose. The unique capability of this species to make 1,2-propanediol from sugars was described decades ago, but the genes responsible were not identified. Here we deleted genes encoding methylglyoxal reductase, methylglyoxal synthase and glycerol dehydrogenase. Deletion of the latter two genes eliminated propanediol production. To understand how carbon flux is redirected in this species, we hypothesized that high ATP levels during growth cessation downregulate the activity of alcohol and aldehyde dehydrogenase activities. Measurements with cell free extracts show approximately twofold and tenfold inhibition of these activities by 10 mM ATP, supporting the hypothesized mechanism of metabolic redirection. This result may have implications for efforts to direct and maximize flux through alcohol dehydrogenase in other species.
Topics: Propylene Glycol; Ethanol; Propylene Glycols; Adenosine Triphosphate; Fermentation
PubMed: 36765076
DOI: 10.1038/s41598-023-29220-9