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Sample records for gamma-ifn-inducible-lysosomal thiol reductase

  1. The Thiol Reductase Activity of YUCCA6 Mediates Delayed Leaf Senescence by Regulating Genes Involved in Auxin Redistribution.

    Cha, Joon-Yung; Kim, Mi R; Jung, In J; Kang, Sun B; Park, Hee J; Kim, Min G; Yun, Dae-Jin; Kim, Woe-Yeon

    2016-01-01

    Auxin, a phytohormone that affects almost every aspect of plant growth and development, is biosynthesized from tryptophan via the tryptamine, indole-3-acetamide, indole-3-pyruvic acid, and indole-3-acetaldoxime pathways. YUCCAs (YUCs), flavin monooxygenase enzymes, catalyze the conversion of indole-3-pyruvic acid (IPA) to the auxin (indole acetic acid). Arabidopsis thaliana YUC6 also exhibits thiol-reductase and chaperone activity in vitro; these activities require the highly conserved Cys-85 and are essential for scavenging of toxic reactive oxygen species (ROS) in the drought tolerance response. Here, we examined whether the YUC6 thiol reductase activity also participates in the delay in senescence observed in YUC6-overexpressing (YUC6-OX) plants. YUC6 overexpression delays leaf senescence in natural and dark-induced senescence conditions by reducing the expression of SENESCENCE-ASSOCIATED GENE 12 (SAG12). ROS accumulation normally occurs during senescence, but was not observed in the leaves of YUC6-OX plants; however, ROS accumulation was observed in YUC6-OX(C85S) plants, which overexpress a mutant YUC6 that lacks thiol reductase activity. We also found that YUC6-OX plants, but not YUC6-OX(C85S) plants, show upregulation of three genes encoding NADPH-dependent thioredoxin reductases (NTRA, NTRB, and NTRC), and GAMMA-GLUTAMYLCYSTEINE SYNTHETASE 1 (GSH1), encoding an enzyme involved in redox signaling. We further determined that excess ROS accumulation caused by methyl viologen treatment or decreased glutathione levels caused by buthionine sulfoximine treatment can decrease the levels of auxin efflux proteins such as PIN2-4. The expression of PINs is also reduced in YUC6-OX plants. These findings suggest that the thiol reductase activity of YUC6 may play an essential role in delaying senescence via the activation of genes involved in redox signaling and auxin availability. PMID:27242830

  2. Thioredoxin Reductase Is Essential for Thiol/Disulfide Redox Control and Oxidative Stress Survival of the Anaerobe Bacteroides fragilis▿ †

    Rocha, Edson R.; Tzianabos, Arthur O; Smith, C. Jeffrey

    2007-01-01

    Results of this study showed that the anaerobic, opportunistic pathogen Bacteroides fragilis lacks the glutathione/glutaredoxin redox system and possesses an extensive number of putative thioredoxin (Trx) orthologs. Analysis of the genome sequence revealed six Trx orthologs and an absence of genes required for synthesis of glutathione and glutaredoxins. In addition, it was shown that the thioredoxin reductase (TrxB)/Trx system is the major or sole redox system for thiol/disulfide cellular hom...

  3. Crystal structure of the YffB protein from Pseudomonas aeruginosa suggests a glutathione-dependent thiol reductase function

    Dauter Zbigniew

    2004-03-01

    Full Text Available Abstract Background The yffB (PA3664 gene of Pseudomonas aeruginosa encodes an uncharacterized protein of 13 kDa molecular weight with a marginal sequence similarity to arsenate reductase from Escherichia coli. The crystal structure determination of YffB was undertaken as part of a structural genomics effort in order to assist with the functional assignment of the protein. Results The structure was determined at 1.0 Å resolution by single-wavelength anomalous diffraction. The fold is very similar to that of arsenate reductase, which is an extension of the thioredoxin fold. Conclusion Given the conservation of the functionally important residues and the ability to bind glutathione, YffB is likely to function as a GSH-dependent thiol reductase.

  4. Location of the redox-active thiols of ribonucleotide reductase: sequences similarity between the Escherichia coli and Lactobacillus leichmannii enzymes

    The redox-active thiols of Escherichia coli ribonucleoside diphosphate reductase and of Lactobacillus leichmannii ribonucleoside triphosphate reductase have been located by a procedure involving (1) prereduction of enzyme with dithiothreitol, (2) specific oxidation of the redox-active thiols by treatment with substrate in the absence of exogenous reductant, (3) alkylation of other thiols with iodoacetamide, and (4) reduction of the disulfides with dithiothreitol and alkylation with [1-14C]iodoacetamide. The dithiothreitol-reduce E. coli B1 subunit is able to convert 3 equiv of CDP to dCDP and is labeled with 5.4 equiv of 14C. Sequencing of tryptic peptides shows that 2.8 equiv of 14C is on cysteines-752 and -757 at the C-terminus of B1, while 1.0-1.5 equiv of 14C is on cysteines-222 and -227. It thus appears that two sets of redox-active dithiols are involved in substrate reduction. The L. leichmannii reductase is able to convert 1.1 equiv of CTP to dCTP and is labeled with 2.1 equiv of 14C. Sequencing of tryptic peptides shows that 1.4 equiv of 14C is located on the two cysteines of C-E-G-G-A-C-P-I-K. This peptide shows remarkable and unexpected similarity to the thiol-containing region of the C-terminal peptide of E. coli B1, C-E-S-G-A-C-K-I

  5. The first echinoderm gamma-interferon-inducible lysosomal thiol reductase (GILT) identified from sea cucumber (Stichopus monotuberculatus).

    Ren, Chunhua; Chen, Ting; Jiang, Xiao; Luo, Xing; Wang, Yanhong; Hu, Chaoqun

    2015-01-01

    Gamma-interferon-inducible lysosomal thiol reductase (GILT) has been described as a key enzyme that facilitating the processing and presentation of major histocompatibility complex class II-restricted antigen in mammals. In this study, the first echinoderm GILT named StmGILT was identified from sea cucumber (Stichopus monotuberculatus). The StmGILT cDNA is 1529 bp in length, containing a 5'-untranslated region (UTR) of 87 bp, a 3'-UTR of 674 bp and an open reading frame (ORF) of 768 bp that encoding a protein of 255 amino acids with a deduced molecular weight of 27.82 kDa and a predicted isoelectric point of 4.73. The putative StmGILT protein possesses all the main characteristics of known GILT proteins, including a signature sequence, a reductase active site CXXC, twelve conserved cysteines, and two potential N-linked glycosylation sites. For the gene structure, StmGILT contains four exons separated by three introns. In the promoter region of StmGILT gene, an NF-κB binding site and an IFN-γ activation site were found. The thiol reductase activity of recombinant StmGILT protein was also demonstrated in this study. In addition, the highest level of mRNA expression was noticed in coelomocytes of S. monotuberculatus. In in vitro experiments performed in coelomocytes, the expression of StmGILT mRNA was significantly up-regulated by lipopolysaccharides (LPS), inactivated bacteria or polyriboinosinic polyribocytidylic acid [poly (I:C)] challenge, suggested that the sea cucumber GILT might play critical roles in the innate immune defending against bacterial and viral infections. PMID:25449705

  6. Increase in thiol oxidative stress via glutathione reductase inhibition as a novel approach to enhance cancer sensitivity to X-ray irradiation.

    Zhao, Yong; Seefeldt, Teresa; Chen, Wei; Carlson, Laura; Stoebner, Adam; Hanson, Sarah; Foll, Ryan; Matthees, Duane P; Palakurthi, Srinath; Guan, Xiangming

    2009-07-15

    Depletion of the reduced form of glutathione (GSH) has been extensively studied for its effect on sensitizing cancer to radiation. However, little is known about the effects of thiol oxidative stress created through an increase in glutathione disulfide (GSSG) on cancer sensitivity to radiation. In this study, an increase in GSSG was effectively created using 2-acetylamino-3-[4-(2-acetylamino-2-carboxyethylsulfanylthiocarbonylamino)phenylthiocarbamoylsulfanyl]propionic acid (2-AAPA), an irreversible glutathione reductase (GR) inhibitor. Our results demonstrate that the GSSG increase significantly enhanced cancer sensitivity to X-ray irradiation in four human cancer cell lines (A431, MCF7, NCI-H226, and OVCAR-3). When cells were pretreated with 2-AAPA followed by X-ray irradiation, the IC(50) values for X-ray irradiation of A431, MCF7, NCI-H226, and OVCAR-3 cells were reduced, from 24.2 +/- 2.8, 42.5 +/- 3.0, 43.0 +/- 3.6, and 27.8+/-3.5 Gy to 6.75 +/- 0.9, 8.1 +/- 1.1, 6.75 +/- 1.0, and 12.1 +/- 1.7 Gy, respectively. The synergistic effects observed from the combination of X-rays plus 2-AAPA were comparable to those from the combination of X-rays plus buthionine sulfoximine, a reference compound known to increase cancer sensitivity to radiation. The synergistic effect was correlated with an increase in cell thiol oxidative stress, which was reflected by a five-to sixfold increase in GSSG and 25% increase in total disulfides. No change in GSH or total thiols was observed as a result of GR inhibition. PMID:19397999

  7. Increase in Thiol Oxidative Stress via Glutathione Reductase Inhibition as a Novel Approach to Enhance Cancer Sensitivity to X-Ray Irradiation

    Zhao, Yong; Seefeldt, Teresa; Chen, Wei; Carlson, Laura; Stoebner, Adam; Hanson, Sarah; Foll, Ryan; Matthees, Duane P.; Palakurthi, Srinath; Guan, Xiangming

    2009-01-01

    Depletion of reduced form glutathione (GSH) has been extensively studied for its effect on sensitizing cancer to radiation. However, little is known about the effect of thiol oxidative stress created through an increase in glutathione disulfide (GSSG) on cancer sensitivity to radiation. In this study, an increase in GSSG was effectively created by 2-acetylamino-3-[4-(2-acetylamino-2-carboxyethylsulfanylthiocarbonylamino)phenylthiocarbamoylsulfanyl]propionic acid (2-AAPA), an irreversible glut...

  8. Thiol biochemistry of prokaryotes

    Fahey, Robert C.

    1986-01-01

    The present studies have shown that GSH metabolism arose in the purple bacteria and cyanobacteria where it functions to protect against oxygen toxicity. Evidence was obtained indicating that GSH metabolism was incorporated into eucaryotes via the endosymbiosis giving rise to mitochrondria and chloroplasts. Aerobic bacteria lacking GSH utilize other thiols for apparently similar functions, the thiol being coenzyme A in Gram positive bacteria and chi-glutamylcysteine in the halobacteria. The thiol biochemistry of prokaryotes is thus seen to be much more highly diversified than that of eucaryotes and much remains to be learned about this subject.

  9. Cell surface thiol isomerases may explain the platelet-selective action of S-nitrosoglutathione

    Xiao, Fang; Gordge, Michael P

    2011-01-01

    S-nitrosoglutathione (GSNO) at low concentration inhibits platelet aggregation without causing vasodilation, suggesting platelet-selective nitric oxide delivery. The mechanism of this selectivity is unknown, but may involve cell surface thiol isomerases, in particular protein disulphide isomerase (csPDI) (EC 5.3.4.1). We have now compared csPDI expression and activity on platelets, endothelial cells and vascular smooth muscle cells, and the dependence on thiol reductase activity of these cell...

  10. Thiols and radiosensitivity

    The role played by non-protein (NPSH) and protein sulfhydryls (PSH) in hypoxic and aerated cell radiosensitivity was investigated using human skin fibroblasts derived from patients affected with 5-oxoprolinuria. These cells have lowered levels of the enzyme GSH-synthetase which results in a decreased concentration of glutathione. Six cell lines were studied; GM3877 and GM3878, SR and SUR from a single family and OB and AB from a French family. Only GM3877, with GSH levels of 0.6 nmoles/mg protein and NPSH levels of 4 nmoles/mg protein, was found to exhibit a reduced OER of 1.8. Experiments are now in progress to investigate the effect of depleting thiol levels with the ∫-glutamyl cysteine synthetase inhibitor DL Buthionine-SR-sulfoximine to determine if the OER is further reduced, especially in the cell line which already has a lowered OER. The results are discussed with a view toward developing a model which takes into account the role of thiols and DNA repair processes in the resistance of hypoxic cells to ionizing radiation

  11. Composite materials having thiol groups

    A composite material having thiol groups comprises a rigid support material. The composite material may comprise a deformable gel (eg agarose) having thiol groups retained within the pore structure of a porous rigid support material (e.g. Kieselghur). The particles of composite material are used to prepare a radioactive gold isotope from a mercury 'parent' isotope. (author)

  12. Histochemical Localization of Glutathione Dependent NBT-Reductase in Mouse Skin

    2001-01-01

    Objective Localization of the glutathione dependent Nitroblue tetrazolium (NBT) reductase in fresh frozen sections of mouse skin and possible dependence of NBT reductase on tissue thiol levels has been investigated. Methods The fresh frozen tissue sections (8m thickness) were prepared and incubated in medium containing NBT, reduced glutathione (GSH) and phosphate buffer. The staining for GSH was performed with mercury orange. Results  The activity of the NBT-reductase in mouse skin has been found to be localized in the areas rich in glutathione and actively proliferating area of the skin. Conclusion The activity of the NBT-reductase seems to be dependent on the glutathione contents.

  13. Cell surface thiol isomerases may explain the platelet-selective action of S-nitrosoglutathione.

    Xiao, Fang; Gordge, Michael P

    2011-10-30

    S-nitrosoglutathione (GSNO) at low concentration inhibits platelet aggregation without causing vasodilation, suggesting platelet-selective nitric oxide delivery. The mechanism of this selectivity is unknown, but may involve cell surface thiol isomerases, in particular protein disulphide isomerase (csPDI) (EC 5.3.4.1). We have now compared csPDI expression and activity on platelets, endothelial cells and vascular smooth muscle cells, and the dependence on thiol reductase activity of these cell types for NO uptake from GSNO. csPDI expression was measured by flow cytometry and its reductase activity using the pseudosubstrate dieosin glutathione disulphide. This activity assay was adapted and validated for 96-well plate format. Flow cytometry revealed csPDI on all three cell types, but percentage positivity of expression was higher on platelets than on vascular cells. Consistent with this, thiol isomerase-related reductase activity was higher on platelets (Pionomycin) increased csPDI activity on both platelets and smooth muscle cells, but not on endothelium. Intracellular NO delivery from GSNO was greater in platelets than in vascular cells (Pselective actions of GSNO and help define its antithrombotic potential. PMID:21642008

  14. Thiol redox homeostasis in neurodegenerative disease

    Gethin J. McBean

    2015-08-01

    Full Text Available This review provides an overview of the biochemistry of thiol redox couples and the significance of thiol redox homeostasis in neurodegenerative disease. The discussion is centred on cysteine/cystine redox balance, the significance of the xc− cystine–glutamate exchanger and the association between protein thiol redox balance and neurodegeneration, with particular reference to Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis and glaucoma. The role of thiol disulphide oxidoreductases in providing neuroprotection is also discussed.

  15. Thiol/Disulfide system plays a crucial role in redox protection in the acidophilic iron-oxidizing bacterium Leptospirillum ferriphilum.

    Javiera Norambuena

    Full Text Available Thiol/disulfide systems are involved in the maintenance of the redox status of proteins and other molecules that contain thiol/disulfide groups. Leptospirillum ferriphilum DSM14647, an acidophilic bacterium that uses Fe(2+ as electron donor, and withstands very high concentrations of iron and other redox active metals, is a good model to study how acidophiles preserve the thiol/disulfide balance. We studied the composition of thiol/disulfide systems and their role in the oxidative stress response in this extremophile bacterium. Bioinformatic analysis using genomic data and enzymatic assays using protein extracts from cells grown under oxidative stress revealed that the major thiol/disulfide system from L. ferriphilum are a cytoplasmic thioredoxin system (composed by thioredoxins Trx and thioredoxin reductase TR, periplasmic thiol oxidation system (DsbA/DsbB and a c-type cytochrome maturation system (DsbD/DsbE. Upon exposure of L. ferriphilum to reactive oxygen species (ROS-generating compounds, transcriptional activation of the genes encoding Trxs and the TR enzyme, which results in an increase of the corresponding activity, was observed. Altogether these data suggest that the thioredoxin-based thiol/disulfide system plays an important role in redox protection of L. ferriphilum favoring the survival of this microorganism under extreme environmental oxidative conditions.

  16. Thiol isomerases in thrombus formation

    Furie, Bruce; Flaumenhaft, Robert

    2014-01-01

    Protein disulfide isomerase, ERp5 and ERp57, among perhaps other thiol isomerases, are important for the initiation of thrombus formation. Using the laser injury thrombosis model in mice to induce in vivo arterial thrombus formation, it was shown that thrombus formation is associated with PDI secretion by platelets, that inhibition of PDI blocked platelet thrombus formation and fibrin generation, and that endothelial cell activation leads to PDI secretion. Similar results using this and other...

  17. The role of glutathione reductase and related enzymes on cellular redox homoeostasis network.

    Couto, Narciso; Wood, Jennifer; Barber, Jill

    2016-06-01

    In this review article we examine the role of glutathione reductase in the regulation, modulation and maintenance of cellular redox homoeostasis. Glutathione reductase is responsible for maintaining the supply of reduced glutathione; one of the most abundant reducing thiols in the majority of cells. In its reduced form, glutathione plays key roles in the cellular control of reactive oxygen species. Reactive oxygen species act as intracellular and extracellular signalling molecules and complex cross talk between levels of reactive oxygen species, levels of oxidised and reduced glutathione and other thiols, and antioxidant enzymes such as glutathione reductase determine the most suitable conditions for redox control within a cell or for activation of programmed cell death. Additionally, we discuss the translation and expression of glutathione reductase in a number of organisms including yeast and humans. In yeast and human cells, a single gene expresses more than one form of glutathione reductase, destined for residence in the cytoplasm or for translocation to different organelles; in plants, however, two genes encoding this protein have been described. In general, insects and kinetoplastids (a group of protozoa, including Plasmodia and Trypanosoma) do not express glutathione reductase or glutathione biosynthetic enzymes. Instead, they express either the thioredoxin system or the trypanothione system. The thioredoxin system is also present in organisms that have the glutathione system and there may be overlapping functions with cross-talk between the two systems. Finally we evaluate therapeutic targets to overcome oxidative stress associated cellular disorders. PMID:26923386

  18. The effect of copper and gallium compounds on ribonucleotide reductase

    Narasimhan, J.

    1992-01-01

    The mode of action of copper complexes (CuL and CuKTS) and gallium compounds (gallium nitrate and citrate) in cytotoxicity was studied. The effects of these agents on the enzyme ribonucleotide reductase was investigated by monitoring the tyrosyl free radical present in the active site of the enzyme through electron spin resonance (ESR) spectroscopy. Ribonucleotide reductase, a key enzyme in cellular proliferation, consists of two subunits. M1, a dimer of molecular weight 170,000 contains the substrate and effector binding sites. M2, a dimer of molecular weight 88,000, contains non-heme iron and tyrosyl free radical essential for the activity of the enzyme. In studies using copper complexes, the cellular oxidative chemistry was examined by ESR studies on adduct formation with membranes, and oxidation of thiols. Membrane thiols were oxidized through the reduction of the ESR signal of the thiol adduct and the analysis of sulfhydryl content. Using the radiolabel [sup 59]Fe, the inhibitory action of copper thiosemicarbazones on cellular iron uptake was shown. The inhibitory action of CuL on ribonucleotide reductase was shown by the quenching of the tyrosyl free radical on the M2 subunit. The hypothesis that gallium directly interacts with the M2 subunit of the enzyme and displaces the iron from it was proven. The tyrosyl free radical signal from cell lysates was inhibited by the direct addition of gallium compounds. Gallium content in the cells was measured by a fluorimetric method, to ensure the presence of sufficient amounts of gallium to compete with the iron in the M2 subunit. The enzyme activity, measured by the conversion of [sup 14]C-CDP to the labeled deoxy CDP, was inhibited by the addition of gallium nitrate in a cell free assay system. The immunoprecipitation studies of the [sup 59]Fe labeled M2 protein using the monoclonal antibody directed against this subunit suggested that gallium releases iron from the M2 subunit.

  19. Histochemical Localization of Glutathione Dependent NBT—Reductase in Mouse Skin

    YOESHWERSHUKLA

    2001-01-01

    Objective:Localization of the glutathione dependent Nitroblue tetrazolium(NBT) reductase in fresh frozen sections of mouse skin and possible dependence of NBT reductase on tissue thiol levels has been investigated.Methods:The fresh frozen tissue sections(8m thickness)were prepared and incuated in medium containing NBT,reduced glutathione(GSH) and Phosphate uffer,The staining for GSH was performed with mercury orange.Results:The activity of the NBT-reductase in mouse skin has een found to be localized in the areas rich in glutatione and actively proliferating area of the skin.Conclusion:The activity of the NBT-reductase seems to be dependent on the glutatione contents.

  20. Thiol-sensitive promoters of Escherichia coli.

    Javor, G T; Stringer, C D; Ryu, J.

    1988-01-01

    Mu dX(lac) insertion mutants of Escherichia coli CSH50 in which the expression of the lacZ gene was sensitive to the presence of exogenous 1-thioglycerol or dithiothreitol were isolated. Both stimulatory and inhibitory mutants were found. The existence of several thiol-sensitive promoters suggests that exogenous thiols may provoke global stress responses in E. coli.

  1. Entamoeba thiol-based redox metabolism: A potential target for drug development.

    Jeelani, Ghulam; Nozaki, Tomoyoshi

    2016-01-01

    Amebiasis is an intestinal infection widespread throughout the world caused by the human pathogen Entamoeba histolytica. Metronidazole has been a drug of choice against amebiasis for decades despite its low efficacy against asymptomatic cyst carriers and emergence of resistance in other protozoa with similar anaerobic metabolism. Therefore, identification and characterization of specific targets is urgently needed to design new therapeutics for improved treatment against amebiasis. Toward this goal, thiol-dependent redox metabolism is of particular interest. The thiol-dependent redox metabolism in E. histolytica consists of proteins including peroxiredoxin, rubrerythrin, Fe-superoxide dismutase, flavodiiron proteins, NADPH: flavin oxidoreductase, and amino acids including l-cysteine, S-methyl-l-cysteine, and thioprolines (thiazolidine-4-carboxylic acids). E. histolytica completely lacks glutathione and its metabolism, and l-cysteine is the major intracellular low molecular mass thiol. Moreover, this parasite possesses a functional thioredoxin system consisting of thioredoxin and thioredoxin reductase, which is a ubiquitous oxidoreductase system with antioxidant and redox regulatory roles. In this review, we summarize and highlight the thiol-based redox metabolism and its control mechanisms in E. histolytica, in particular, the features of the system unique to E. histolytica, and its potential use for drug development against amebiasis. PMID:26775086

  2. Reaction of alkylcobalamins with thiols

    Carbon-13 NMR spectroscopy and phosphorus-31 NMR spectroscopy have been used to study the reaction of several alkylcobalamins with 2-mercaptoethanol. At alkaline pH, when the thiol is deprotonated, the alkyl-transfer reactions involve a nucleophilic attack of the thiolate anion on the Co-methylene carbon of the cobalamins, yielding alkyl thioethers and cob(II)alamin. In these nucleophilic displacement reactions cob(I)alamin is presumably formed as an intermediate. The higher alkylcobalamins react more slowly than methylcobalamin. The lower reactivity of ethyl- and propylcobalamin is probably the basis of the inhibition of the corrinoid-dependent methyl-transfer systems by propyl iodide. The transfer of the upper nucleoside ligand of adenosylcobalamin to 2-mercaptoethanol is a very slow process; S-adenosylmercaptoethanol and cob(II)alamin are the final products of the reaction. The dealkylation of (carboxymethyl)cobalamin is a much more facile reaction. At alkaline pH S-(carboxymethyl)mercaptoethanol and cob(II)alamin are produced, while at pH values below 8 the carbon-cobalt bond is cleaved reductively to acetate and cob(II)alamin. The reductive cleavage of the carbon-cobalt bond of (carboxymethyl)cobalamin by 2-mercaptoethanol is extremely fast when the cobalamin is in the base-off form. Because the authors have been unable to detect trans coordination of 2-mercaptoethanol, they favor a mechanism that involves a hydride attack on the Co-methylene carbon of (carboxymethyl) rather than a trans attack of the thiol on the cobalt atom

  3. The Effects of Acrolein on Peroxiredoxins, Thioredoxins, and Thioredoxin Reductase in Human Bronchial Epithelial Cells

    Myers, Charles R.; Myers, Judith M.

    2008-01-01

    Inhalation is a common form of exposure to acrolein, a toxic reactive volatile aldehyde that is a ubiquitous environmental pollutant. Bronchial epithelial cells would be directly exposed to inhaled acrolein. The thioredoxin (Trx) system is essential for the maintenance of cellular thiol redox balance, and is critical for cell survival. Normally, thioredoxin reductase (TrxR) maintains the cytosolic (Trx1) and mitochondrial (Trx2) thioredoxins in the reduced state, and the thioredoxins keep the...

  4. Induction of Glutathione Synthesis and Glutathione Reductase Activity by Abiotic Stresses in Maize and Wheat

    Gabor Kocsy; Gabriella Szalai; Gabor Galiba

    2002-01-01

    The effect of different abiotic stresses (extreme temperatures and osmotic stress) on the synthesis of glutathione and hydroxymethylglutathione, on the ratio of the reduced to oxidised forms of these thiols (GSH/GSSG, hmGSH/hmGSSG), and on the glutathione reductase (GR) activity was studied in maize and wheat genotypes having different sensitivity to low temperature stress. Cold treatment induced a greater increase in total glutathione (TG) content and in GR activity in tolerant genotypes of ...

  5. Evaluation and Control of Thiol-ene/Thiol-epoxy Hybrid Networks

    Carioscia, Jacquelyn A.; Stansbury, Jeffrey W.; Bowman, Christopher N.

    2007-01-01

    The development of thiol-ene/thiol-epoxy hybrid networks offers the advantage of tailorable polymerization kinetics while producing a highly crosslinked, high Tg polymer that has significantly reduced shrinkage stress. Stoichiometric mixtures of pentaerythritol tetra(3-mercaptopropionate) (PETMP)/triallyl-1,3,5-triazine-2,4,6-trione (TATATO) (thiol-ene, mixture 1) and PETMP/bisphenol a diglycidyl ether (BADGE) (thiol-epoxy, mixture 2) were prepared and hybrid mixtures of 75/25, 50/50, 25/75, ...

  6. Thiol-reactivity of the fungicide maneb

    James R. Roede

    2014-01-01

    Full Text Available Maneb (MB is a manganese-containing ethylene bis-dithiocarbamate fungicide that is implicated as an environmental risk factor for Parkinson's disease, especially in combination with paraquat (PQ. Dithiocarbamates inhibit aldehyde dehydrogenases, but the relationship of this to the combined toxicity of MB + PQ is unclear because PQ is an oxidant and MB activates Nrf2 and increases cellular GSH without apparent oxidative stress. The present research investigated the direct reactivity of MB with protein thiols using recombinant thioredoxin-1 (Trx1 as a model protein. The results show that MB causes stoichiometric loss of protein thiols, reversibly dimerizes the protein and inhibits its enzymatic activity. MB reacted at similar rates with low-molecular weight, thiol-containing chemicals. Together, the data suggest that MB can potentiate neurotoxicity of multiple agents by disrupting protein thiol functions in a manner analogous to that caused by oxidative stress, but without GSH depletion.

  7. Thiol-reactivity of the fungicide maneb.

    Roede, James R; Jones, Dean P

    2014-01-01

    Maneb (MB) is a manganese-containing ethylene bis-dithiocarbamate fungicide that is implicated as an environmental risk factor for Parkinson's disease, especially in combination with paraquat (PQ). Dithiocarbamates inhibit aldehyde dehydrogenases, but the relationship of this to the combined toxicity of MB + PQ is unclear because PQ is an oxidant and MB activates Nrf2 and increases cellular GSH without apparent oxidative stress. The present research investigated the direct reactivity of MB with protein thiols using recombinant thioredoxin-1 (Trx1) as a model protein. The results show that MB causes stoichiometric loss of protein thiols, reversibly dimerizes the protein and inhibits its enzymatic activity. MB reacted at similar rates with low-molecular weight, thiol-containing chemicals. Together, the data suggest that MB can potentiate neurotoxicity of multiple agents by disrupting protein thiol functions in a manner analogous to that caused by oxidative stress, but without GSH depletion. PMID:24936438

  8. Hypochlorite-induced oxidation of thiols

    Davies, Michael Jonathan; Hawkins, C L

    2000-01-01

    -molecular-weight thiols such as reduced glutathione (GSH), and sulfur-containing amino acids in proteins, are major targets for HOCl. Radicals have not generally been implicated as intermediates in thiol oxidation by HOCl, though there is considerable literature evidence for the involvement of radicals in the metal ion......-, thermal- or UV light-catalysed decomposition of sulfenyl or sulfonyl chlorides which are postulated intermediates in thiol oxidation. In this study we show that thiyl radicals are generated on reaction of a number of low-molecular-weight thiols with HOCl. With sub-stoichiometric amounts of HOCl, relative...... for the involvement of sulfenyl chlorides (RSCl) in the formation of these radicals, and studies with an authentic sulfenyl chloride have demonstrated that this compound readily decomposes in thermal-, metal-ion- or light-catalysed reactions to give thiyl radicals. The formation of thiyl radicals on oxidation...

  9. Coenzyme A disulfide reductase, the primary low molecular weight disulfide reductase from Staphylococcus aureus. Purification and characterization of the native enzyme.

    delCardayre, S B; Stock, K P; Newton, G L; Fahey, R C; Davies, J E

    1998-03-01

    The human pathogen Staphylococcus aureus does not utilize the glutathione thiol/disulfide redox system employed by eukaryotes and many bacteria. Instead, this organism produces CoA as its major low molecular weight thiol. We report the identification and purification of the disulfide reductase component of this thiol/disulfide redox system. Coenzyme A disulfide reductase (CoADR) catalyzes the specific reduction of CoA disulfide by NADPH. CoADR has a pH optimum of 7.5-8.0 and is a dimer of identical subunits of Mr 49,000 each. The visible absorbance spectrum is indicative of a flavoprotein with a lambdamax = 452 nm. The liberated flavin from thermally denatured enzyme was identified as flavin adenine dinucleotide. Steady-state kinetic analysis revealed that CoADR catalyzes the reduction of CoA disulfide by NADPH at pH 7.8 with a Km for NADPH of 2 muM and for CoA disulfide of 11 muM. In addition to CoA disulfide CoADR reduces 4,4'-diphosphopantethine but has no measurable ability to reduce oxidized glutathione, cystine, pantethine, or H2O2. CoADR demonstrates a sequential kinetic mechanism and employs a single active site cysteine residue that forms a stable mixed disulfide with CoA during catalysis. These data suggest that S. aureus employs a thiol/disulfide redox system based on CoA/CoA-disulfide and CoADR, an unorthodox new member of the pyridine nucleotide-disulfide reductase superfamily. PMID:9488707

  10. Genomics and X-ray microanalysis indicate that Ca2+ and thiols mediate the aggregation and adhesion of Xylella fastidiosa

    Leite B.

    2002-01-01

    Full Text Available The availability of the genome sequence of the bacterial plant pathogen Xylella fastidiosa, the causal agent of citrus variegated chlorosis, is accelerating important investigations concerning its pathogenicity. Plant vessel occlusion is critical for symptom development. The objective of the present study was to search for information that would help to explain the adhesion of X. fastidiosa cells to the xylem. Scanning electron microscopy revealed that adhesion may occur without the fastidium gum, an exopolysaccharide produced by X. fastidiosa, and X-ray microanalysis demonstrated the presence of elemental sulfur both in cells grown in vitro and in cells found inside plant vessels, indicating that the sulfur signal is generated by the pathogen surface. Calcium and magnesium peaks were detected in association with sulfur in occluded vessels. We propose an explanation for the adhesion and aggregation process. Thiol groups, maintained by the enzyme peptide methionine sulfoxide reductase, could be active on the surface of the bacteria and appear to promote cell-cell aggregation by forming disulfide bonds with thiol groups on the surface of adjacent cells. The enzyme methionine sulfoxide reductase has been shown to be an auxiliary component in the adhesiveness of some human pathogens. The negative charge conferred by the ionized thiol group could of itself constitute a mechanism of adhesion by allowing the formation of divalent cation bridges between the negatively charged bacteria and predominantly negatively charged xylem walls.

  11. Molecular study of DNA radioprotection by thiols

    Savoye, C.; Charlier, M.; Spotheim-Maurizot, M. [Centre National de la Recherche Scientifique (CNRS), 45 - Orleans-la-Source (France); Swenberg, C. [ACRD, Armed Forces Radiobiology Research Institute, Bethesda (United States); Sabattier, R. [Centre Hospitalier Regional d`Orleans, 45 (France)

    1997-03-01

    Polyamines (PA) are natural components of mammalian cells, essential for growth processes. Since a decrease of the cellular level of PA increases the effect of radiotherapy on tumour cells, we have supposed that PA may act as DNA radioprotectors. The search of non-toxic agents that protect specifically normal cells led to the discovery of the agent WR-2721 used now in cancer cancer therapy under the name Ethyol (Amifostine) and of the agent WR-151327. Both have a chemical structure close to that of natural PA. The main radioprotective metabolites of these agents are the thiols WR>61065 and WR-151326. We have compared here here the protective effects of these thiols to those of another simpler thiol, the cysteamine, and of a related PA, the putrescine, on the number and location of fast neutrons-induced DNA strand breaks. (authors)

  12. Glutathione reductase: solvent equilibrium and kinetic isotope effects

    Glutathione reductase catalyzes the NADPH-dependent reduction of oxidized glutathione (GSSG). The kinetic mechanism is ping-pong, and we have investigated the rate-limiting nature of proton-transfer steps in the reactions catalyzed by the spinach, yeast, and human erythrocyte glutathione reductases using a combination of alternate substrate and solvent kinetic isotope effects. With NADPH or GSSG as the variable substrate, at a fixed, saturating concentration of the other substrate, solvent kinetic isotope effects were observed on V but not V/K. Plots of Vm vs mole fraction of D2O (proton inventories) were linear in both cases for the yeast, spinach, and human erythrocyte enzymes. When solvent kinetic isotope effect studies were performed with DTNB instead of GSSG as an alternate substrate, a solvent kinetic isotope effect of 1.0 was observed. Solvent kinetic isotope effect measurements were also performed on the asymmetric disulfides GSSNB and GSSNP by using human erythrocyte glutathione reductase. The Km values for GSSNB and GSSNP were 70 microM and 13 microM, respectively, and V values were 62 and 57% of the one calculated for GSSG, respectively. Both of these substrates yield solvent kinetic isotope effects greater than 1.0 on both V and V/K and linear proton inventories, indicating that a single proton-transfer step is still rate limiting. These data are discussed in relationship to the chemical mechanism of GSSG reduction and the identity of the proton-transfer step whose rate is sensitive to solvent isotopic composition. Finally, the solvent equilibrium isotope effect measured with yeast glutathione reductase is 4.98, which allows us to calculate a fractionation factor for the thiol moiety of GSH of 0.456

  13. Human brain aldehyde reductases: relationship to succinic semialdehyde reductase and aldose reductase.

    Hoffman, P L; Wermuth, B; von Wartburg, J P

    1980-08-01

    Human brain contains multiple forms of aldehyde-reducing enzymes. One major form (AR3), as previously shown, has properties that indicate its identity with NADPH-dependent aldehyde reductase isolated from brain and other organs of various species; i.e., low molecular weight, use of NADPH as the preferred cofactor, and sensitivity to inhibition by barbiturates. A second form of aldehyde reductase ("SSA reductase") specifically reduces succinic semialdehyde (SSA) to produce gamma-hydroxybutyrate. This enzyme form has a higher molecular weight than AR3, and uses NADH as well as NADPH as cofactor. SSA reductase was not inhibited by pyrazole, oxalate, or barbiturates, and the only effective inhibitor found was the flavonoid quercetine. Although AR3 can also reduce SSA, the relative specificity of SSA reductase may enhance its in vivo role. A third form of human brain aldehyde reductase, AR2, appears to be comparable to aldose reductases characterized in several species, on the basis of its activity pattern with various sugar aldehydes and its response to characteristic inhibitors and activators, as well as kinetic parameters. This enzyme is also the most active in reducing the aldehyde derivatives of biogenic amines. These studies suggest that the various forms of human brain aldehyde reductases may have specific physiological functions. PMID:6778961

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

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

    2009-01-01

    Thiol redox state (TRS) is an important parameter to reflect intracellular oxidative stress and is associated with various normal and abnormal biochemical processes. Agents that can be used to increase intracellular TRS will be valuable tools in TRS-related research. Glutathione reductase (GR) is a critical enzyme in the homeostasis of TRS. The enzyme catalyzes the reduction of GSSG to GSH to maintain a high GSH:GSSG ratio. Inhibition of the enzyme can be used to incre...

  15. Microfluidic devices using thiol-ene polymers

    Bou, Simon J. M. C.; Ellis, Amanda V.

    2013-12-01

    Here, a new polymeric microfluidic platform using off-stoichiometric thiol-ene (OSTE) polymers was developed. Thiolene polymers were chosen as they afford rapid UV curing, low volume shrinkage and optical transparency for use in microfluidic devices. Three different off-stoichiometric thiol-ene polymers with 30% excess allyl, 50% excess thiol and a 90% excess thiol (OSTE Allyl-30, OSTE-50 and OSTE-90, respectively) were fabricated. Attenuated reflectance Fourier transform infrared (ATR-FTIR) spectroscopy and solid-state cross polarisation-magic angle spinning (CP-MAS) nuclear magnetic resonance (NMR) spectroscopy confirmed which functional groups (thiol or allyl) were present in excess in the OSTE polymers. The polymers were shown to have a more hydrophilic surface (water contact angle of 65°+/- 3) compared to polydimethylsiloxane (water contact angle of 105° +/- 5). Testing of the mechanical properties showed the glass transition temperatures to be 15.09 °C, 43.15 °C and, 57.48 °C for OSTE-90, OSTE Allyl-30 and, OSTE-50, respectively. The storage modulus was shown to be less than 10 MPa for the OSTE-90 polymer and approximately 1750 MPa for the OSTE Allyl-30 and OSTE-50 polymers. The polymers were then utilised to fabricate microfluidic devices via soft lithography practices and devices sealed using a one-step UV lamination "click" reaction technique. Finally, gold nanoparticles were used to form gold films on the OSTE-90 and OSTE-50 polymers as potential electrodes. Atomic force microscopy and sheet resistances were used to characterise the films.

  16. Functional and Structural Characterization of a Thiol Peroxidase from Mycobacterium tuberculosis

    Rho,B.; Hung, L.; Holton, J.; Vigil, D.; Kim, S.; Park, M.; Terwilliger, T.; Pedelacq, j.

    2006-01-01

    A thiol peroxidase (Tpx) from Mycobacterium tuberculosis was functionally analyzed. The enzyme shows NADPH-linked peroxidase activity using a thioredoxin-thioredoxin reductase system as electron donor, and anti-oxidant activity in a thiol-dependent metal-catalyzed oxidation system. It reduces H{sub 2}O{sub 2}, t-butyl hydroperoxide, and cumene hydroperoxide, and is inhibited by sulfhydryl reagents. Mutational studies revealed that the peroxidatic (Cys60) and resolving (Cys93) cysteine residues are critical amino acids for catalytic activity. The X-ray structure determined to a resolution of 1.75 Angstroms shows a thioredoxin fold similar to that of other peroxiredoxin family members. Superposition with structural homologues in oxidized and reduced forms indicates that the M. tuberculosis Tpx is a member of the atypical two-Cys peroxiredoxin family. In addition, the short distance that separates the Ca atoms of Cys60 and Cys93 and the location of these cysteine residues in unstructured regions may indicate that the M. tuberculosis enzyme is oxidized, though the side-chain of Cys60 is poorly visible. It is solely in the reduced Streptococcus pneumoniae Tpx structure that both residues are part of two distinct helical segments. The M. tuberculosis Tpx is dimeric both in solution and in the crystal structure. Amino acid residues from both monomers delineate the active site pocket.

  17. Quantifying the global cellular thiol-disulfide status

    Hansen, Rosa E; Roth, Doris; Winther, Jakob R

    2009-01-01

    It is widely accepted that the redox status of protein thiols is of central importance to protein structure and folding and that glutathione is an important low-molecular-mass redox regulator. However, the total cellular pools of thiols and disulfides and their relative abundance have never been...... determined. In this study, we have assembled a global picture of the cellular thiol-disulfide status in cultured mammalian cells. We have quantified the absolute levels of protein thiols, protein disulfides, and glutathionylated protein (PSSG) in all cellular protein, including membrane proteins. These data...... cell types. However, when cells are exposed to a sublethal dose of the thiol-specific oxidant diamide, PSSG levels increase to >15% of all protein cysteine. Glutathione is typically characterized as the "cellular redox buffer"; nevertheless, our data show that protein thiols represent a larger active...

  18. Tunable degradation of maleimide-thiol adducts in reducing environments

    Baldwin, Aaron D.; Kiick, Kristi L.

    2011-01-01

    Addition chemistries are widely used in preparing biological conjugates, and in particular, maleimide-thiol adducts have been widely employed. Here we show that the resulting succinimide thioether formed by a Michael type addition of a thiol to N-ethylmaleimide (NEM), generally accepted as stable, can in fact undergo retro and exchange reactions in the presence of other thiol compounds at physiological pH and temperature, offering a novel strategy for controlled release. Model studies (1H NMR...

  19. Copper complexation by thiol compounds in estuarine waters

    Laglera, L.M.; van den Berg, C.M.G.

    2003-01-01

    The stability of copper complexes with thiol substances in estuarine waters was determined for the first time using a new procedure based on cathodic stripping voltammetry (CSV). The free thiol concentration was monitored during titrations with copper in the presence of a competing ligand salicylaldoxime (SA); concentrations of copper-complexing ligands and conditional stability constants were determined simultaneously but independently. The decrease in the free thiol concentration with incre...

  20. Protein Thiols as an Indication of Oxidative Stress

    Yousef Rezaei Chianeh; Krishnananda Prabhu

    2014-01-01

    Thiol is an organic compound that contain sulphhydryl group that have a critical role in preventing any involvement of oxidative stress in the cell. These defensive functions are generally considered to be carried out by the low molecular weight thiol glutathione and by cysteine residues in the active sites of proteins such as thioredoxin and peroxiredoxin. In addition, there are thiols exposed on protein surfaces that are not directly involved with protein function, although they can intera...

  1. Thiol-containing polymeric embedding materials for nanoskiving

    Mays, Robin L.; Pourhossein, Parisa; Savithri, Dhanalekshmi; Genzer, Jan; Chiechi, Ryan C.; Dickey, Michael D.

    2013-01-01

    This paper describes the characterization of new embedding resins for nanoskiving (ultramicrotomy) that contain thiols. Nanoskiving is a technique to produce nanoscale structures using an ultramicrotome to section thin films of materials (e. g., gold) embedded in polymer. Epoxies are used typically as embedding resins for microtomy. Epoxies, however, do not adhere well to gold or other smooth metallic structures that are used commonly for nanoskiving. Thiol-ene and thiol-epoxy polymers provid...

  2. Designed Chemical Intervention with Thiols for Prophylactic Contraception

    Monika Sharma; Lokesh Kumar; Ashish Jain; Vikas Verma; Vikas Sharma; Bhavana Kushwaha; Nand Lal; Lalit Kumar; Tara Rawat; Dwivedi, Anil K.; Jagdamba P Maikhuri; Sharma, Vishnu L.; Gopal Gupta

    2013-01-01

    Unlike somatic cells, sperm have several-fold more available-thiols that are susceptible to redox-active agents. The present study explains the mechanism behind the instant sperm-immobilizing and trichomonacidal activities of pyrrolidinium pyrrolidine-1-carbodithioate (PPC), a novel thiol agent rationally created for prophylactic contraception by minor chemical modifications of some known thiol drugs. PPC, and its three derivatives (with potential active-site blocked by alkylation), were synt...

  3. Thiol groups of gizzard myosin heavy chains

    Bailin, G.

    1986-05-01

    Proteolysis of phosphorylated and /sup 3/H-labeled dinitrophenylated chicken gizzard myosin with trypsin released major fragments of M/sub r/ 25,000, 50,000 and 66,000 in a 1:1 ratio. They contained 57% of the dinitrophenyl (N/sub 2/ph) group bound to thiols of the heavy chains; 28% of the label was bound to the light chains. The fragments of M/sub r/ 25,000 and M/sub r/ 66,000 were dinitrophenylated predominantly when the K/sup +/-ATPase activity was inhibited. Thiolysis of phosphorylated and dinitrophenylated myosin with 2-mercaptoethanol removed 60% and 25% of the N/sub 2/ph group from the N-terminal and M/sub r/ 66,000 fragments of the heavy chain, respectively, when 48% of the K/sup +/-ATPase activity was restored. Papain proteolysis of the tryptic digest of modified myosin released a C-terminal segment from the fragment of M/sub r/ 66,000 and it contained most of the remaining label. Proteolysis of /sup 3/H-labeled dinitrophenylated myosin alone resulted in the same digestion pattern but less of the label was bound to the heavy chain fragments. In this case, restoration of enzymic activity occurred in thiolyzed dinitrophenylated myosin when the N/sub 2/ph group was removed from the light chains, predominantly. Conformational changes in gizzard myosin, mediated by phosphorylation, altered the reactivity of the thiols in specific fragments of the heavy chain. Thiol groups of the N- and C-terminal heavy chain regions are involved in maintaining the ATPase activity of myosin.

  4. Genetics Home Reference: sepiapterin reductase deficiency

    Skip to main content Your Guide to Understanding Genetic Conditions Enable Javascript for addthis links to activate. ... Conditions Genes Chromosomes & mtDNA Resources Help Me Understand Genetics Home Health Conditions sepiapterin reductase deficiency sepiapterin reductase ...

  5. Thiol-ene-based monolithic microreactors

    Novotný, Jakub; Lafleur, J. P.; Kutter, J. P.

    Brno : Institute of Analytical Chemistry AS CR, 2014 - (Foret, F.; Křenková, J.; Drobníková, I.; Guttman, A.; Klepárník, K.), s. 53-55 ISBN 978-80-904959-2-0. [CECE 2014. International Interdisciplinary Meeting on Bioanalysis /11./. Brno (CZ), 20.10.2014-22.10.2014] Institutional support: RVO:68081715 Keywords : thiol-ene * monolith * enzyme immobilization Subject RIV: CB - Analytical Chemistry, Separation http://www.ce-ce.org/CECE2014/CECE%202014%20proceedings_full.pdf

  6. Studies on alterations of the 86-rubidium efflux from rat pancreatic islets caused by thiol and thiol oxidants

    The following findings were revealed by this study: 1) Oxidation-reduction (redox) of the intracellular system of glutathione influences the potassium efflux by way of an increase in the 86-rubidium efflux brought about by the oxidation of intracellular thiols. 2) The 86-rubidium efflux is not subject to change by oxidation of extracellular thiols located in the membrane, nor can it in any way be influenced by reduced glutathione of exogenous origin. 3) The potassium efflux from rat pancreatic islets, being generally known to trigger the electric activities of the beta-cell, is controlled by the oxidation-reduction of intracellular thiols rather than by that of extracellular thiols. (TRV)

  7. Serum paraoxonase activity and protein thiols in patients with hyperlipidemia

    Mungli Prakash; Jeevan K Shetty; Sudeshna Tripathy; Pannuri Vikram; Manish Verma

    2009-01-01

    Objective: In the present study we evaluated the paraoxonase activity and protein thiols level in south Indian population with newly diagnosed hyperlipidemia. Methods: The study was conducted on 55 newly diagnosed hyperlipidemic pa-tients and 57 healthy controls. Serum paraoxonase activity and protein thiols were estimated by spectrophotometeric method and lipid profile by enzymatic kinetic assay method. Results: Serum paraoxonase activity, protein thiols and high density lipoprotein levels were low and total cholesterol, triglycerides and low density lipoprutein levels were high in patients with hyperlipidemia compared to healthy controls ( P < 0.01 ). Serum paranxonase activity correlated positively with protein thiols and high density lipoprotein (P<0.01). Conclusion: Decreased paraoxonase activity and protein thiols were found in patients with hyperlipi-demia. This may indicate the susceptibility of this population to accelerated atherogenesis and protein oxidation.

  8. Quantification of protein-derived thiols during atmosphere-controlled brewing in laboratory scale

    Murmann, Anne Nordmark; Andersen, Preben; Mauch, Alexander;

    2016-01-01

    . Fermentation caused an increase in free thiols, and the balance between free and total thiols was shifted toward a higher degree of free thiols. This was explained by either a reducing effect of fermentation or secretion of thiol-containing compounds from yeast. The efficiency of sulfite to reduce reversibly...

  9. Surface functionalized thiol-ene waveguides for fluorescence biosensing in microfluidic devices

    Feidenhans'l, Nikolaj Agentoft; Lafleur, Josiane P.; Jensen, Thomas Glasdam; Kutter, Jörg Peter

    -ene waveguides were fabricated from 40% excess thiol thiol-ene to ensure the presence of thiol functional groups at the surface of the waveguide. Biotin alkyne was photografted at specific locations using a photomask, directly at the interface between the microfluidic channel and the thiol-ene waveguide prior to...

  10. Thiol-based redox signaling in the nitrogen-fixing symbiosis

    Pierre eFrendo

    2013-09-01

    Full Text Available In nitrogen poor soils legumes establish a symbiotic interaction with rhizobia that results in the formation of root nodules. These are unique plant organs where bacteria differentiate into bacteroids, which express the nitrogenase enzyme complex that reduces atmospheric N2 to ammonia. Nodule metabolism requires a tight control of the concentrations of reactive oxygen and nitrogen species (RONS so that they can perform useful signaling roles while avoiding nitro-oxidative damage. In nodules a thiol-dependent regulatory network that senses, transmits and responds to redox changes is starting to be elucidated. A combination of enzymatic, immunological, pharmacological and molecular analyses has allowed to conclude that glutathione and its legume-specific homolog, homoglutathione, are abundant in meristematic and infected cells, their spatio-temporally distribution is correlated with the corresponding (homoglutathione synthetase activities, and are crucial for nodule development and function. Glutathione is at high concentrations in the bacteroids and at moderate amounts in the mitochondria, cytosol and nuclei. Less information is available on other components of the network. The expression of multiple isoforms of glutathione peroxidases, peroxiredoxins, thioredoxins, glutaredoxins and NADPH-thioredoxin reductases has been detected in nodule cells using antibodies and proteomics. Peroxiredoxins and thioredoxins are essential to regulate and in some cases to detoxify RONS in nodules. Further research is necessary to clarify the regulation of the expression and activity of thiol redox-active proteins in response to abiotic, biotic and developmental cues, their interactions with downstream targets by disulfide-exchange reactions, and their participation in signaling cascades. The availability of mutants and transgenic lines will be crucial to facilitate systematic investigations into the function of the various proteins in the legume

  11. Fatty acyl-CoA reductase

    Reiser, Steven E.; Somerville, Chris R.

    1998-12-01

    The present invention relates to bacterial enzymes, in particular to an acyl-CoA reductase and a gene encoding an acyl-CoA reductase, the amino acid and nucleic acid sequences corresponding to the reductase polypeptide and gene, respectively, and to methods of obtaining such enzymes, amino acid sequences and nucleic acid sequences. The invention also relates to the use of such sequences to provide transgenic host cells capable of producing fatty alcohols and fatty aldehydes.

  12. Molecular modeling, structural analysis and identification of ligand binding sites of trypanothione reductase from Leishmania mexicana

    Ozal Mutlu

    2013-01-01

    Background & objectives: Trypanothione reductase (TR) is a member of FAD-dependent NADPH oxidoreductase protein family and it is a key enzyme which connects the NADPH and the thiol-based redox system. Inhibition studies indicate that TR is an essential enzyme for parasite survival. Therefore, it is an attractive target enzyme for novel drug candidates. There is no structural model for TR of Leishmania mexicana (LmTR) in the protein databases. In this work, 3D structure of TR from L. mexicana ...

  13. Mechanism-based inhibition of a mutant Escherichia coli ribonucleotide reductase (cysteine-225----serine) by its substrate CDP.

    Mao, S. S.; Johnston, M I; Bollinger, J M; Stubbe, J.

    1989-01-01

    The B1 subunit of Escherichia coli ribonucleotide reductase (EC 1.17.4.1) has been overexpressed using the pT7-5/pGP1-2 system developed by Tabor and Richardson [Tabor, S. & Richardson, C. (1985) Proc. Natl. Acad. Sci. USA 82, 1074-1078]. This method has allowed the preparation of two mutant B1 subunits in which two of the four thiols postulated to be within the active site of the enzyme, Cys-225 and Cys-759, have been changed to serines. Incubation of the [Ser225]B1 mutant with the B2 subuni...

  14. Community Analysis of a Mercury Hot Spring Supports Occurrence of Domain-Specific Forms of Mercuric Reductase

    Simbahan, Jessica; Kurth, Elizabeth; Schelert, James; Dillman, Amanda; Moriyama, Etsuko; Jovanovich, Stevan; Blum, Paul

    2005-01-01

    Mercury is a redox-active heavy metal that reacts with active thiols and depletes cellular antioxidants. Active resistance to the mercuric ion is a widely distributed trait among bacteria and results from the action of mercuric reductase (MerA). Protein phylogenetic analysis of MerA in bacteria indicated the occurrence of a second distinctive form of MerA among the archaea, which lacked an N-terminal metal recruitment domain and a C-terminal active tyrosine. To assess the distribution of the ...

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

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

    2016-07-01

    Glutathione is the most abundant thiol in the vast majority of organisms and is maintained in its reduced form by the flavoenzyme glutathione reductase. In this work, we describe the genetic and functional analysis of the Caenorhabditis elegans gsr-1 gene that encodes the only glutathione reductase protein in this model organism. By using green fluorescent protein reporters we demonstrate that gsr-1 produces two GSR-1 isoforms, one located in the cytoplasm and one in the mitochondria. gsr-1 loss of function mutants display a fully penetrant embryonic lethal phenotype characterized by a progressive and robust cell division delay accompanied by an aberrant distribution of interphasic chromatin in the periphery of the cell nucleus. Maternally expressed GSR-1 is sufficient to support embryonic development but these animals are short-lived, sensitized to chemical stress, have increased mitochondrial fragmentation and lower mitochondrial DNA content. Furthermore, the embryonic lethality of gsr-1 worms is prevented by restoring GSR-1 activity in the cytoplasm but not in mitochondria. Given the fact that the thioredoxin redox systems are dispensable in C. elegans, our data support a prominent role of the glutathione reductase/glutathione pathway in maintaining redox homeostasis in the nematode. PMID:27117030

  16. Facially amphiphilic thiol capped gold and silver nanoparticles

    Bhat, Shreedhar; Maitra, Uday

    2008-01-01

    A series of bile acid-derived facially amphiphilic thiols have been used to cap sliver and gold nanoparticles. The self-assembling properties of these steroid-capped nanoparticles have been investigated and reported in this article.

  17. Intracellular thiols: involvement in drug metabolism and radiation response

    Nitro compunds are activated by coupled enzyme reactions to oxygen reactive intermediates leading to the formation of peroxide, under aerobic conditions, and to the depletion of thiols, under anaerobic conditions. Some nitro compounds as substrates for glutathione-S-transferase, show peroxide production without prior thiol removal. Other drugs reacting spontaneouly with glutathione also produce peroxide. Glutathione plays an important role in the metabolism of the nitrocompounds either by directly reacting with them or their reduced intermediates such as the nitroso, nitro and hydroxyl radical. In the case of misonidazole, protection against their cytotoxic effects can be achieved by the addition of exogenous thiols such as glutathione or cysteamine. Results indicate that oxygen and peroxide electrodes provide convenient means for measuring the products of metabolic activation of nitro compounds. Mechanisms are proposed whereby protein, nonprotein and glutathione thiols can interact with drug radicals or with DNA radicals. 60 references, 14 figures, 5 tables

  18. Facially amphiphilic thiol capped gold and silver nanoparticles

    Shreedhar Bhata; Uday Maitra

    2008-11-01

    A series of bile acid-derived facially amphiphilic thiols have been used to cap sliver and gold nanoparticles. The self-assembling properties of these steroid-capped nanoparticles have been investigated and reported in this article.

  19. Reversible inactivation of CO dehydrogenase with thiol compounds

    Highlights: • Rather large thiols (e.g. coenzyme A) can reach the active site of CO dehydrogenase. • CO- and H2-oxidizing activity of CO dehydrogenase is inhibited by thiols. • Inhibition by thiols was reversed by CO or upon lowering the thiol concentration. • Thiols coordinate the Cu ion in the [CuSMo(=O)OH] active site as a third ligand. - Abstract: Carbon monoxide dehydrogenase (CO dehydrogenase) from Oligotropha carboxidovorans is a structurally characterized member of the molybdenum hydroxylase enzyme family. It catalyzes the oxidation of CO (CO + H2O → CO2 + 2e− + 2H+) which proceeds at a unique [CuSMo(=O)OH] metal cluster. Because of changing activities of CO dehydrogenase, particularly in subcellular fractions, we speculated whether the enzyme would be subject to regulation by thiols (RSH). Here we establish inhibition of CO dehydrogenase by thiols and report the corresponding Ki-values (mM): L-cysteine (5.2), D-cysteine (9.7), N-acetyl-L-cysteine (8.2), D,L-homocysteine (25.8), L-cysteine–glycine (2.0), dithiothreitol (4.1), coenzyme A (8.3), and 2-mercaptoethanol (9.3). Inhibition of the enzyme was reversed by CO or upon lowering the thiol concentration. Electron paramagnetic resonance spectroscopy (EPR) and X-ray absorption spectroscopy (XAS) of thiol-inhibited CO dehydrogenase revealed a bimetallic site in which the RSH coordinates to the Cu-ion as a third ligand ([MoVI(=O)OH(2)SCuI(SR)S-Cys]) leaving the redox state of the Cu(I) and the Mo(VI) unchanged. Collectively, our findings establish a regulation of CO dehydrogenase activity by thiols in vitro. They also corroborate the hypothesis that CO interacts with the Cu-ion first. The result that thiol compounds much larger than CO can freely travel through the substrate channel leading to the bimetallic cluster challenges previous concepts involving chaperone function and is of importance for an understanding how the sulfuration step in the assembly of the bimetallic cluster might proceed

  20. Reversible inactivation of CO dehydrogenase with thiol compounds

    Kreß, Oliver [Department of Microbiology, University of Bayreuth, 95440 Bayreuth (Germany); Gnida, Manuel [Department of Chemistry, University of Paderborn, 33098 Paderborn (Germany); Pelzmann, Astrid M. [Department of Microbiology, University of Bayreuth, 95440 Bayreuth (Germany); Marx, Christian [Institute of Biochemistry and Biophysics, Friedrich-Schiller-University of Jena, 07745 Jena (Germany); Meyer-Klaucke, Wolfram [Department of Chemistry, University of Paderborn, 33098 Paderborn (Germany); Meyer, Ortwin, E-mail: Ortwin.Meyer@uni-bayreuth.de [Department of Microbiology, University of Bayreuth, 95440 Bayreuth (Germany)

    2014-05-09

    Highlights: • Rather large thiols (e.g. coenzyme A) can reach the active site of CO dehydrogenase. • CO- and H{sub 2}-oxidizing activity of CO dehydrogenase is inhibited by thiols. • Inhibition by thiols was reversed by CO or upon lowering the thiol concentration. • Thiols coordinate the Cu ion in the [CuSMo(=O)OH] active site as a third ligand. - Abstract: Carbon monoxide dehydrogenase (CO dehydrogenase) from Oligotropha carboxidovorans is a structurally characterized member of the molybdenum hydroxylase enzyme family. It catalyzes the oxidation of CO (CO + H{sub 2}O → CO{sub 2} + 2e{sup −} + 2H{sup +}) which proceeds at a unique [CuSMo(=O)OH] metal cluster. Because of changing activities of CO dehydrogenase, particularly in subcellular fractions, we speculated whether the enzyme would be subject to regulation by thiols (RSH). Here we establish inhibition of CO dehydrogenase by thiols and report the corresponding K{sub i}-values (mM): L-cysteine (5.2), D-cysteine (9.7), N-acetyl-L-cysteine (8.2), D,L-homocysteine (25.8), L-cysteine–glycine (2.0), dithiothreitol (4.1), coenzyme A (8.3), and 2-mercaptoethanol (9.3). Inhibition of the enzyme was reversed by CO or upon lowering the thiol concentration. Electron paramagnetic resonance spectroscopy (EPR) and X-ray absorption spectroscopy (XAS) of thiol-inhibited CO dehydrogenase revealed a bimetallic site in which the RSH coordinates to the Cu-ion as a third ligand ([Mo{sup VI}(=O)OH{sub (2)}SCu{sup I}(SR)S-Cys]) leaving the redox state of the Cu(I) and the Mo(VI) unchanged. Collectively, our findings establish a regulation of CO dehydrogenase activity by thiols in vitro. They also corroborate the hypothesis that CO interacts with the Cu-ion first. The result that thiol compounds much larger than CO can freely travel through the substrate channel leading to the bimetallic cluster challenges previous concepts involving chaperone function and is of importance for an understanding how the sulfuration step in

  1. Oxidative Thiol Modifications in Pro- and Eukaryotic Organisms

    Brandes, Nicolas

    2010-01-01

    Cystein spielt eine wichtige Rolle in der Biochemie vieler Proteine. Aufgrund der Redox-Eigenschaften und der hohen Reaktivität der freien Thiol-Gruppe sowie dessen Fähigkeit Metallionen zu koordinieren, ist Cystein oft Bestandteil von katalytischen Zentren vieler Enzyme. Zudem lassen sich Cysteine durch reaktive Sauerstoff- und Stickstoffspezies leicht reversibel oxidativ modifizieren. In den letzten Jahren wurde gezeigt, dass Proteine redox-bedingte Thiol-Modifikationen nutzen, um Veränderu...

  2. A maize gene encoding an NADPH binding enzyme highly homologous to isoflavone reductases is activated in response to sulfur starvation.

    Petrucco, S; Bolchi, A; Foroni, C; Percudani, R; Rossi, G L; Ottonello, S

    1996-01-01

    we isolated a novel gene that is selectively induced both in roots and shoots in response to sulfur starvation. This gene encodes a cytosolic, monomeric protein of 33 kD that selectively binds NADPH. The predicted polypeptide is highly homologous ( > 70%) to leguminous isoflavone reductases (IFRs), but the maize protein (IRL for isoflavone reductase-like) belongs to a novel family of proteins present in a variety of plants. Anti-IRL antibodies specifically recognize IFR polypeptides, yet the maize protein is unable to use various isoflavonoids as substrates. IRL expression is correlated closely to glutathione availability: it is persistently induced in seedlings whose glutathione content is about fourfold lower than controls, and it is down-regulated rapidly when control levels of glutathione are restored. This glutathione-dependent regulation indicates that maize IRL may play a crucial role in the establishment of a thiol-independent response to oxidative stress under glutathione shortage conditions. PMID:8597660

  3. Short communication: characterization of soluble thiols in bovine milk.

    Niero, G; De Marchi, M; Masi, A; Penasa, M; Cassandro, M

    2015-09-01

    Antioxidants are molecules essential for the maintenance of cell homeostasis and their intake through the diet has positive effects on human health. Among antioxidants, low-molecular-weight (LMW) thiols represent an important class of compounds. The aim of this study was to identify LMW thiols in bovine milk. A total of 96 individual milk samples from Brown Swiss, Holstein-Friesian, Alpine Grey, and Simmental cattle breeds were collected in 8 herds. The LMW thiols were extracted from the soluble fraction of milk and, following a derivatization protocol, they were separated by reverse phase HPLC and detected fluorimetrically. Six thiol species were detected and 2, glutathione (GSH) and cysteine-glycine (Cys-Gly), were identified and quantified. Regardless of the breed, the average concentration of Cys-Gly in milk was greater than that of GSH. Overall, milk from dual-purpose breeds (Simmental and Alpine Grey) was richer in LMW thiols than milk from dairy cows (Holstein-Friesian and Brown Swiss). Glutathione and Cys-Gly, closely linked metabolically, were strongly correlated. Pearson correlations of Cys-Gly with protein and casein contents were moderately low, and no relationship was found between GSH and milk chemical composition. Future research should focus on the identification of all detected LMW thiol species. PMID:26188581

  4. Protein Thiols as an Indication of Oxidative Stress

    Yousef Rezaei Chianeh

    2014-06-01

    Full Text Available Thiol is an organic compound that contain sulphhydryl group that have a critical role in preventing any involvement of oxidative stress in the cell. These defensive functions are generally considered to be carried out by the low molecular weight thiol glutathione and by cysteine residues in the active sites of proteins such as thioredoxin and peroxiredoxin. In addition, there are thiols exposed on protein surfaces that are not directly involved with protein function, although they can interact with the intracellular environment.The process of protection of the cell against an oxidative damage occur by thiol and cystein residue that has a low molecular weight. These residue are present in the active sites of a protein like, peroxiredoxin and thioredoxin. Apart from intracellular antioxidant defense mechanism by protein thiol, there are presence of thiol in outer surface of protein that are not involved with the function of protein, even though they can interact with intracellular part of the cell. [Archives Medical Review Journal 2014; 23(3.000: 443-456

  5. Interfacial thiol-ene photoclick reactions for forming multilayer hydrogels.

    Shih, Han; Fraser, Andrew K; Lin, Chien-Chi

    2013-03-13

    Interfacial visible light-mediated thiol-ene photoclick reactions were developed for preparing step-growth hydrogels with multilayer structures. The effect of a noncleavage type photoinitiator eosin-Y on visible-light-mediated thiol-ene photopolymerization was first characterized using in situ photorheometry, gel fraction, and equilibrium swelling ratio. Next, spectrophotometric properties of eosin-Y in the presence of various relevant macromer species were evaluated using ultraviolet-visible light (UV-vis) spectrometry. It was determined that eosin-Y was able to reinitiate the thiol-ene photoclick reaction, even after light exposure. Because of its small molecular weight, most eosin-Y molecules readily leached out from the hydrogels. The diffusion of residual eosin-Y from preformed hydrogels was exploited for fabricating multilayer step-growth hydrogels. Interfacial hydrogel coating was formed via the same visible-light-mediated gelation mechanism without adding fresh initiator. The thickness of the thiol-ene gel coating could be easily controlled by adjusting visible light exposure time, eosin-Y concentration initially loaded in the core gel, or macromer concentration in the coating solution. The major benefits of this interfacial thiol-ene coating system include its simplicity and cytocompatibility. The formation of thiol-ene hydrogels and coatings neither requires nor generates any cytotoxic components. This new gelation chemistry may have great utilities in controlled release of multiple sensitive growth factors and encapsulation of multiple cell types for tissue regeneration. PMID:23384151

  6. Designed Chemical Intervention with Thiols for Prophylactic Contraception.

    Monika Sharma

    Full Text Available Unlike somatic cells, sperm have several-fold more available-thiols that are susceptible to redox-active agents. The present study explains the mechanism behind the instant sperm-immobilizing and trichomonacidal activities of pyrrolidinium pyrrolidine-1-carbodithioate (PPC, a novel thiol agent rationally created for prophylactic contraception by minor chemical modifications of some known thiol drugs. PPC, and its three derivatives (with potential active-site blocked by alkylation, were synthesized and evaluated against live human sperm and metronidazole-susceptible and resistant Trichomonas vaginalis, in vitro. Sperm hexokinase activity was evaluated by coupled enzyme assay. PPC irreversibly immobilized 100% human sperm in ∼30 seconds and totally eliminated Trichomonas vaginalis more efficiently than nonoxynol-9 and metronidazole. It significantly inhibited (P<0.001 thiol-sensitive sperm hexokinase. However, the molecule completely lost all its biological activities once its thiol group was blocked by alkylation. PPC was subsequently formulated into a mucoadhesive vaginal film using GRaS excipients and evaluated for spermicidal and microbicidal activities (in vitro, and contraceptive efficacy in rabbits. PPC remained fully active in quick-dissolving, mucoadhesive vaginal-film formulation, and these PPC-films significantly reduced pregnancy and fertility rates in rabbits. The films released ∼90% of PPC in simulated vaginal fluid (pH 4.2 at 37°C in 5 minutes, in vitro. We have thus discovered a common target (reactive thiols on chiefly-anaerobic, redox-sensitive cells like sperm and Trichomonas, which is susceptible to designed chemical interference for prophylactic contraception. The active thiol in PPC inactivates sperm and Trichomonas via interference with crucial sulfhydryl-disulfide based reactions, e.g. hexokinase activation in human sperm. In comparison to non-specific surfactant action of OTC spermicide nonoxynol-9, the action of

  7. The impact of thiol peroxidases on redox regulation.

    Flohé, Leopold

    2016-01-01

    The biology of glutathione peroxidases and peroxiredoxins is reviewed with emphasis on their role in metabolic regulation. Apart from their obvious function in balancing oxidative challenge, these thiol peroxidases are not only implicated in orchestrating the adaptive response to oxidative stress, but also in regulating signaling triggered by hormones, growth factors and cytokines. The mechanisms presently discussed comprise dampening of redox-sensitive regulatory processes by elimination of hydroperoxides, suppression of lipoxygenase activity, committing suicide to save H2O2 for signaling, direct binding to receptors or regulatory proteins in a peroxidase activity-independent manner, or acting as sensors for hydroperoxides and as transducers of oxidant signals. The various mechanistic proposals are discussed in the light of kinetic data, which unfortunately are scarce. Taking into account pivotal criteria of a meaningful regulatory circuit, kinetic plausibility and specificity, the mechanistic concepts implying a direct sensor/transducer function of the thiol peroxidases appear most appealing. With rate constants for the reaction with hydroperoxide of 10(5)-10(8) M(-1) s(-1), thiol peroxidases are qualified as kinetically preferred hydroperoxide sensors, and the ability of the oxidized enzymes to react with defined protein thiols lends specificity to the transduction process. The versatility of thiol peroxidases, however, allows multiple ways of interaction with regulatory pathways. PMID:26291534

  8. Protection by thiols against poisoning by radiomimetic agents. Chapter 8

    A review is presented of reports of studies aimed at detecting a protective effect of thiols against radiomimetic alkylating agents such as those used in cancer therapy (nitrogen mustards (HN2), sarcolysine, busulfan, etc.). Protection by thiols against alkylating agents has been observed in mammals, plant cells, bacteria, isolated mammalian cells and in model systems. The lack of correlation between the protective power of various thiols against radiomimetic agents and ionizing radiations indicates that different mechanisms are involved. Studies have been made of the toxicity of the protector and the competition factor, increased excretion of detoxication products of alkylating agents, decreased alkylation of DNA and RNA both in vivo and in vitro, the protection of hematopoietic tissues, tumours and the adrenal cortex, and the modification of the effects of nitrosoalkylamines, carbon tetrachloride and fungistatics by thiols. The restriction of DNA alkylation by the competitive removal of radiomimetic agents is thought to account for the protective effect of thiols against radiomimetic agents. (U.K.)

  9. Designed Chemical Intervention with Thiols for Prophylactic Contraception.

    Sharma, Monika; Kumar, Lokesh; Jain, Ashish; Verma, Vikas; Sharma, Vikas; Kushwaha, Bhavana; Lal, Nand; Kumar, Lalit; Rawat, Tara; Dwivedi, Anil K; Maikhuri, Jagdamba P; Sharma, Vishnu L; Gupta, Gopal

    2013-01-01

    Unlike somatic cells, sperm have several-fold more available-thiols that are susceptible to redox-active agents. The present study explains the mechanism behind the instant sperm-immobilizing and trichomonacidal activities of pyrrolidinium pyrrolidine-1-carbodithioate (PPC), a novel thiol agent rationally created for prophylactic contraception by minor chemical modifications of some known thiol drugs. PPC, and its three derivatives (with potential active-site blocked by alkylation), were synthesized and evaluated against live human sperm and metronidazole-susceptible and resistant Trichomonas vaginalis, in vitro. Sperm hexokinase activity was evaluated by coupled enzyme assay. PPC irreversibly immobilized 100% human sperm in ∼30 seconds and totally eliminated Trichomonas vaginalis more efficiently than nonoxynol-9 and metronidazole. It significantly inhibited (Pnonoxynol-9, the action of thiol-active PPC is apparently much more specific, potent and safe. PPC presents a proof-of-concept for prophylactic contraception via manipulation of thiols in vagina for selective targeting of sperm and Trichomonas, and qualifies as a promising lead for the development of dually protective vaginal-contraceptive. PMID:23826278

  10. Thiol and non-thiol antioxidants effect radiation damage expressed by IEC-6 cells

    Full text: The epithelial lining of the GI mucosal surface is traditionally viewed as a passive barrier serving a largely protective function. Recently, enterocytes have been seen to function as sensitive indicators of oxidative stress, with defined responses to shifts in the oxidative balance. Radiation damage, long recognized as the result of the generation of reactive oxygen species, would theoretically be modulated by the presence of radical scavenging anti-oxidants. Cultures of IEC-6 cells, a model for the gastrointestinal epithelium were found to have lower numbers of adherent cells in response either n-acetyl cysteine (NAC) or l-ascorbate in the medium. In both cases the response was dose dependent, with inhibition in response to l-ascorbate well established by 48 hours. However, in contrast to the thiol antioxidant NAC where a late recovery was observed at high dose, no statistically significant increase in adherent cell numbers were observed with high dose l-ascorbate, and adherent cell numbers actually fell off with time. Exposure to antioxidants potentiated the damage from x-ray irradiation further reducing cell numbers. While we have previously shown that this damage was not associated with mitotic delay or inhibition of proliferation, the mechanism of this response remains undefined. Non-adherent cells were found to increase with dose in the presence of antioxidants with those cells having morphology consistent with apoptotic cells including nuclear condensation, and blabbing. Annexin V cells increased in the non-adherent cell layer, but the numbers did not seem to account for the severe reduction in cell numbers observed. It has been suggested that a p53 mutation alters the response to oxidative damage in these cells via a thiol containing motif sensitive to the cellular glutathione pool resulting in an automatic signal to release from the basement membrane, however it does not explain the similarity in effects to ascorbate

  11. Effect of γ-irradiation on thiol compounds in grapefruit

    The effect of 60Co γ-irradiation on thiol compounds in grapefruit was investigated. Thiols were separated by HPLC and measured with a fluorescence detector. Reduced glutathione (GSH), cysteine (CySH), cysteinylglycine (CySGly), and a number of unknown peaks were observed in unirradiated grapefruit. GSH was the main thiol at an average concentration of 143.3 μM. GSH content exponentially decreased with increased radiation doses, and after 100 krad only 80% of the original remained. The G value based on the result of 100 krad was 0.29. Authentic GSH in water or citrate buffer (pH 3) was converted mainly to its oxidized form (GSSG) with γ-irradiated grapefruits showed no equivalent increase, however

  12. Resistivity of thiol-modified gold thin films

    In this work, we study the effect of thiol self assembled monolayers on the electrical resistivity of metallic thin films. The analysis is based on the Fuchs–Sondheimer–Lucas theory and on electrical transport measurements. We determined resistivity change due to dodecanethiol adsorption on gold thin films. For this purpose, we controlled the deposition and annealing temperatures of the films to change the surface topography and to diminish the effect of electron grain boundary scattering. Results show that the electrical response to the absorption of thiols strongly depends on the initial topography of the surface. - Highlights: • We study the effect of self assembled monolayers on the resistivity of thin films. • Fuchs–Sondheimer theory reproduces the resistivity increase due to thiol deposition. • We determined resistivity change due to dodecanethiol deposition on gold thin films. • The electrical response strongly depends on the substrate surface topography

  13. Effect of gamma irradiation on thiol compounds in grapefruit

    The effect of 60Co γ-irradiation on thiol compounds in grapefruit was investigated. Thiols were separated by HPLC and measured with a fluorescence detector. Reduced glutathione (GSH), cysteine (CySH), cysteinylglycine (CySGly), and a number of unknown peaks were observed in unirradiated grapefruit. GSH was the main thiol at an average concentration of 143.3 μM. GSH content exponentially decreased with increased radiation doses, and after 100 krad only 80% of the original remained. The G value based on the result of 100 krad was 0.29. Authentic GSH in water or citrate buffer (pH 3) was converted mainly to its oxidized form (GSSG) with γ-irradiation. GSSG in irradiated grapefruits showed no equivalent increase, however. (author)

  14. Interaction of thiols and non-thiol ·OH scavengers in the modification of radiation-induced DNA damage

    Oxygen has long been known to sensitize cells to the lethal effects of ionizing radiation and is widely believed to do so by the fixation of potentially reversible radical damage to cellular DNA. A number of studies have suggested that this widely observed oxygen enhancement of cell killing requires the presence of reduced thiols. Published in vitro studies of the modification of DNA damage by glutathione or other thiols have generally shown peak oxygen enhancement ratios (OERs) much higher than those observed for cell killing. However, these studies measured loss of DNA transforming activity or induction of single-strand DNA breaks (SSBs), related endpoints which are not thought to represent lethal lesions, rather than double-strand breaks (DSBs), which are generally believed to be the dominant lethal lesions from ionizing radiation. In addition, non-thiol scavengers of OH radicals were not generally present. There is also evidence that, in addition to their protective effects, some non-thiol ·OH scavengers can produce radicals which are damaging to DNA under anoxic conditions. In the present investigation, the authors have adapted a previously used in vitro model system to simultaneously investigate the effects on radiation-induced single- and double-strand DNA breaks of various combinations of glutathione and glycerol, a widely used non-thiol scavenger, in the presence and absence of oxygen

  15. Identification, Characterization, and Classification of Genes Encoding Perchlorate Reductase

    Bender, Kelly S.; Shang, Ching; Chakraborty, Romy; Belchik, Sara M.; Coates, John D.; Achenbach, Laurie A.

    2005-01-01

    The reduction of perchlorate to chlorite, the first enzymatic step in the bacterial reduction of perchlorate, is catalyzed by perchlorate reductase. The genes encoding perchlorate reductase (pcrABCD) in two Dechloromonas species were characterized. Sequence analysis of the pcrAB gene products revealed similarity to α- and β-subunits of microbial nitrate reductase, selenate reductase, dimethyl sulfide dehydrogenase, ethylbenzene dehydrogenase, and chlorate reductase, all of which are type II m...

  16. 5-Furan-2yl[1,3,4]oxadiazole-2-thiol, 5-Furan-2yl-4H [1,2,4] triazole-3-thiol and Their Thiol-Thione Tautomerism

    A. Cansız

    2005-02-01

    Full Text Available 5-Furan-2-yl[1,3,4]oxadiazole-2-thiol (Ia and 5-furan-2-yl-4H-[1,2,4]-triazole-3-thiol (Ib were synthesized from furan-2-carboxylic acid hydrazide. Mannich basesand methyl derivatives were then prepared. The structures of the synthesized compoundswere confirmed by elemental analyses, IR and 1H-NMR spectra. Their thiol-thione tautomericequilibrium is described.

  17. Sepiapterin Reductase Deficiency: Mimic of Cerebral Palsy

    J Gordon Millichap

    2012-01-01

    Researchers at University of California at San Diego, and 22 other US national and international centers studied the clinical, biochemical, and molecular findings in a cohort of 38 patients with sepiapterin reductase deficiency (SRD).

  18. Naphthalimide Scaffold Provides Versatile Platform for Selective Thiol Sensing and Protein Labeling.

    Zhou, Pengcheng; Yao, Juan; Hu, Guodong; Fang, Jianguo

    2016-04-15

    Reversible thiol modifications are fundamental of cellular redox regulation. Specific thiol detection, including thiol sensing and protein thiols labeling, is critical to study such modifications. We reported the discovery of 4-methylsulfonyl-N-n-butyl-1,8-naphthalimide (MSBN), a highly selective fluorogenic probe for thiols based on the 1,8-naphthalimide scaffold. Thiols react with MSBN nearly quantitatively via nucleophilic aromatic substitution to replace the methylsulfonyl group and restore the quenched fluorescence (>100-fold increase). MSBN was employed to selectively image thiols in live cells and specifically label protein thiols with a turn-on signal to determine diverse reversible protein thiol modifications. In addition, we introduced a bulky group into the MSBN as a mass tag to create a probe MSBN-TPP, which readily discriminates the reduced thioredoxin from the oxidized one. The specific reaction of MSBN with thiols and the easy manipulation of the naphthalimide unit enable MSBN a versatile scaffold in developing novel probes for thiol-based protein bioconjugation and studying various thiol modifications. PMID:26813105

  19. Visible light-initiated interfacial thiol-norbornene photopolymerization for forming islet surface conformal coating

    Shih, Han; Mirmira, Raghavendra G.; Lin, Chien-Chi

    2014-01-01

    A cytocompatible visible light-mediated interfacial thiol-norbornene photopolymerization scheme was developed for creating hydrogel conformal coating on pancreatic islets. The step-growth thiol-norbornene reaction affords high consistency and tunability in gel coating thickness. Furthermore, isolated islets coated with thiol-norbornene gel maintained their viability and function in vitro.

  20. Multiple aldehyde reductases of human brain.

    Hoffman, P L; Wermuth, B; von Wartburg, J P

    1980-01-01

    Human brain contains four forms of aldehyde reducing enzymes. One major activity, designated AR3, has properties indicating its identity with the NADPH-dependent aldehyde reductase, EC 1.1.1.2. The other major form of human brain enzyme, AR1, which is also NADPH-dependent, reduces both aldehyde and ketone-containing substrates, including vitamin K3 (menadione) and daunorubicin, a cancer chemotherapeutic agent. This enzyme is very sensitive to inhibition by the flavonoids quercitrin and quercetine, and may be analogous to a daunorubicin reductase previously described in liver of other species. One minor form of human brain aldehyde reductase, AR2, demonstrates substrate specificity and inhibitor sensitivity which suggest its similarity to aldose reductases found in lens and other tissues of many species. This enzyme, which can also use NADH as cofactor to some extent, is the most active in reducing the aldehyde derivatives of the biogenic amines. The fourth human brain enzyme ("SSA reductase") differs from the other forms in its ability to use NADH as well as or better than NADPH as cofactor, and in its molecular weight, which is nearly twice that of the other forms. It is quite specific for succinic semialdehyde (SSA) as substrate, and was found to be significantly inhibited only by quercetine and quercitrin. AR3 can also reduce SSA, and both enzymes may contribute to the production of gamma-hydroxybutyric acid in vivo. These results indicate that the human brain aldehyde reductases can play relatively specific physiologic roles. PMID:7424738

  1. Preparation of Novel Hydrolyzing Urethane Modified Thiol-Ene Networks

    Bridget S. Confait

    2011-10-01

    Full Text Available Novel tetra-functional hydrolyzing monomers were prepared from the reaction of TEOS and select alkene-containing alcohols, ethylene glycol vinyl ether or 2-allyloxy ethanol, and combined with trimethylolpropane tris(3-mercaptopropionate (tri-thiol in a thiol-ene “click” polymerization reaction to produce clear, colorless thiol-ene networks using both radiation and thermal-cure techniques. These networks were characterized for various mechanical characteristics, and found to posses Tg’s (DSC, hardness, tack, and thermal stability (TGA consistent with their molecular structures. A new ene-modified urethane oligomer was prepared based on the aliphatic polyisocyanate Desmodur® N 3600 and added to the thiol-ene hydrolyzable network series in increasing amounts, creating a phase-segregated material having two Tg’s. An increase in water absorption in the ene-modified urethane formulations leading to a simultaneous increase in the rate of hydrolysis was supported by TGA data, film hardness measurements, and an NMR study of closely related networks. This phenomenon was attributed to the additional hydrogen bonding elements and polar functionality brought to the film with the addition of the urethane segment. SEM was utilized for visual analysis of topographical changes in the film’s surface upon hydrolysis and provides support for surface-driven erosion. Coatings prepared in this study are intended for use as hydrolyzing networks for marine coatings to protect against ship fouling.

  2. Arsenate reduction: thiol cascade chemistry with convergent evolution.

    Messens, Joris; Silver, Simon

    2006-09-01

    The frequent abundance of arsenic in the environment has guided the evolution of enzymes for the reduction of arsenate. The arsenate reductases (ArsC) from different sources have unrelated sequences and structural folds, and can be divided into different classes on the basis of their structures, reduction mechanisms and the locations of catalytic cysteine residues. The thioredoxin-coupled arsenate reductase class is represented by Staphylococcus aureus pI258 ArsC and Bacillus subtilis ArsC. The ArsC from Escherichia coli plasmid R773 and the eukaryotic ACR2p reductase from Saccharomyces cerevisiae represent two distinct glutaredoxin-linked ArsC classes. All are small cytoplasmic redox enzymes that reduce arsenate to arsenite by the sequential involvement of three different thiolate nucleophiles that function as a redox cascade. In contrast, the ArrAB complex is a bacterial heterodimeric periplasmic or a surface-anchored arsenate reductase that functions as a terminal electron acceptor and transfers electrons from the membrane respiratory chain to arsenate. Finally, the less well documented arsenate reductase activity of the monomeric arsenic(III) methylase, which is an S-adenosylmethionine (AdoMet)-dependent methyltransferase. After each oxidative methylation cycle and before the next methylation step, As(V) is reduced to As(III). Methylation by this enzyme is also considered an arsenic-resistance mechanism for bacteria, fungi and mammals. PMID:16905151

  3. Structural and mechanistic insights on nitrate reductases.

    Coelho, Catarina; Romão, Maria João

    2015-12-01

    Nitrate reductases (NR) belong to the DMSO reductase family of Mo-containing enzymes and perform key roles in the metabolism of the nitrogen cycle, reducing nitrate to nitrite. Due to variable cell location, structure and function, they have been divided into periplasmic (Nap), cytoplasmic, and membrane-bound (Nar) nitrate reductases. The first crystal structure obtained for a NR was that of the monomeric NapA from Desulfovibrio desulfuricans in 1999. Since then several new crystal structures were solved providing novel insights that led to the revision of the commonly accepted reaction mechanism for periplasmic nitrate reductases. The two crystal structures available for the NarGHI protein are from the same organism (Escherichia coli) and the combination with electrochemical and spectroscopic studies also lead to the proposal of a reaction mechanism for this group of enzymes. Here we present an overview on the current advances in structural and functional aspects of bacterial nitrate reductases, focusing on the mechanistic implications drawn from the crystallographic data. PMID:26362109

  4. Respiratory arsenate reductase as a bidirectional enzyme

    Richey, C.; Chovanec, P.; Hoeft, S.E.; Oremland, R.S.; Basu, P.; Stolz, J.F.

    2009-01-01

    The haloalkaliphilic bacterium Alkalilimnicola ehrlichii is capable of anaerobic chemolithoautotrophic growth by coupling the oxidation of arsenite (As(III)) to the reduction of nitrate and carbon dioxide. Analysis of its complete genome indicates that it lacks a conventional arsenite oxidase (Aox), but instead possesses two operons that each encode a putative respiratory arsenate reductase (Arr). Here we show that one homolog is expressed under chemolithoautotrophic conditions and exhibits both arsenite oxidase and arsenate reductase activity. We also demonstrate that Arr from two arsenate respiring bacteria, Alkaliphilus oremlandii and Shewanella sp. strain ANA-3, is also biochemically reversible. Thus Arr can function as a reductase or oxidase. Its physiological role in a specific organism, however, may depend on the electron potentials of the molybdenum center and [Fe–S] clusters, additional subunits, or constitution of the electron transfer chain. This versatility further underscores the ubiquity and antiquity of microbial arsenic metabolism.

  5. Respiratory arsenate reductase as a bidirectional enzyme

    The haloalkaliphilic bacterium Alkalilimnicola ehrlichii is capable of anaerobic chemolithoautotrophic growth by coupling the oxidation of arsenite (As(III)) to the reduction of nitrate and carbon dioxide. Analysis of its complete genome indicates that it lacks a conventional arsenite oxidase (Aox), but instead possesses two operons that each encode a putative respiratory arsenate reductase (Arr). Here we show that one homolog is expressed under chemolithoautotrophic conditions and exhibits both arsenite oxidase and arsenate reductase activity. We also demonstrate that Arr from two arsenate respiring bacteria, Alkaliphilus oremlandii and Shewanella sp. strain ANA-3, is also biochemically reversible. Thus Arr can function as a reductase or oxidase. Its physiological role in a specific organism, however, may depend on the electron potentials of the molybdenum center and [Fe-S] clusters, additional subunits, or constitution of the electron transfer chain. This versatility further underscores the ubiquity and antiquity of microbial arsenic metabolism.

  6. Radical Scavenging Efficacy of Thiol Capped Silver Nanoparticles

    Kumudini Chandraker; Sandeep Kumar Vaishanav; Rekha Nagwanshi; Manmohan L Satnami

    2015-12-01

    Radical scavenging efficacy of L-cysteine (L-Cys), glutathione (GSH) and thioctic acid (TA) in the presence of silver nanoparticles (AgNPs) were determined by 1,1-diphenyl 2-picryl hydrazil (DPPH), nitric oxide (NO) and hydroxyl (OH) radicals as spectrophotometric assay. The hydrogen peroxide (H2O2) scavenging efficacy has been determined by titration method. Ascorbic acid has been used as standard for all radical scavenging efficacies. In general, antioxidant activity decreases in the presence of AgNPs. The covalent interactions of thiols (-SH) were found to be a key factor for the decreases in scavenging activity. The effect of thiol concentrations has been discussed. The size and shape of the nanoparticles and AgNP-SR interactions have been characterized through Transmission Electron Microscopy (TEM) and Fourier Transform Infrared (FTIR) spectroscopy, respectively.

  7. Water electrolyte promoted oxidation of functional thiol groups.

    Lauwers, K; Breynaert, E; Rombouts, I; Delcour, J A; Kirschhock, C E A

    2016-04-15

    The formation of disulfide bonds is of the utmost importance for a wide range of food products with gluten or globular proteins as functional agents. Here, the impact of mineral electrolyte composition of aqueous solutions on thiol oxidation kinetics was studied, using glutathione (GSH) and cysteine (CYS) as model systems. Interestingly, the oxidation rate of both compounds into their corresponding disulfides was significantly higher in common tap water than in ultrapure water. The systematic study of different electrolyte components showed that especially CaCl2 improved the oxidation rate of GSH. However, this effect was not observed for CYS, which indicated a strong impact of the local chemical environment on thiol oxidation kinetics. PMID:26675862

  8. EUV negative-resist based on thiol-yne system

    Shirai, Masamitsu; Maki, Koichi; Okamura, Haruyuki; Kaneyama, Koji; Itani, Toshiro

    2011-04-01

    Non-conventional chemically amplified (CA) negative resist for EUV lithography was studied. We have designed negative-tone EUV resist based on thiol-yne stepwise radical reactions. OH groups of poly(4-hydroxystyrene) (PHS) were modified with functional units bearing C-C triple bond structure. Resist was formulated as a mixture of modified-PHS, multifunctional thiol compound, and photoradical generator. The present resist was developable with standard 2.38 wt% TMAH aq. solution. Photo-sensitivity of the resist was obtained on irradiation at 254 nm and 13.5 nm. The resist was highly sensitive to EUV exposure. The sensitivity and the contrast were affected by the structure of modified-PHS and process conditions.

  9. Functional graphene by thiol-ene click chemistry.

    Luong, Nguyen Dang; Sinh, Le Hoang; Johansson, Leena-Sisko; Campell, Joseph; Seppälä, Jukka

    2015-02-16

    Thiol-ene click reaction was successfully employed for chemical modification of graphene oxide (GO) by one-step synthesis. Herein, 2,2-azobis(2-methylpropionitrile) (AIBN) was used as thermal catalyst and cysteamine hydrochloride (HS-(CH2 )2 -NH2 HCl) was used as thiol-containing compound, which is incorporated to GO surface upon reaction with the C=C bonds. The hydrochloride acts as protecting group for the amine, which is finally eliminated by adding sodium hydroxide. The modified GO contains both S- and N-containing groups (NS-GO). We found that NS-GO sheets form good dispersion in water, ethanol, and ethylene glycol. These graphene dispersions can be processed into functionalized graphene film. Besides, it was demonstrated that NS-GO was proved to be an excellent host matrix for platinum nanoparticles. The developed method paves a new way for graphene modification and its functional nanocomposites. PMID:25580698

  10. Functional Conducting Polymers via Thiol-ene Chemistry

    Martin, David C.; Feldman, Kathleen E.

    2012-01-01

    We demonstrate here that thiol-ene chemistry can be used to provide side-chain functionalized monomers based on 3,4-propylenedioxythiophene (ProDOT) containing ionic, neutral, hydrophobic, and hydrophilic side chains. All reactions gave high yields and purification could generally be accomplished through precipitation. These monomers were polymerized either chemically or electro-chemically to give soluble materials or conductive films, respectively. This strategy provides for facile tuning of...

  11. Investigation of thiol derivatized gold nanoparticle sensors for gas analysis

    Stephens, Jared S.

    Analysis of volatile organic compounds (VOCs) in air and exhaled breath by sensor array is a very useful testing technique. It can provide non-invasive, fast, inexpensive testing for many diseases. Breath analysis has been very successful in identifying cancer and other diseases by using a chemiresistor sensor or array with gold nanoparticles to detect biomarkers. Acetone is a biomarker for diabetes and having a portable testing device could help to monitor diabetic and therapeutic progress. An advantage to this testing method is it is conducted at room temperature instead of 200 degrees Celsius. 3. The objective of this research is to determine the effect of thiol derivatized gold nanoparticles based on sensor(s) detection of VOCs. The VOCs to be tested are acetone, ethanol, and a mixture of acetone and ethanol. Each chip is tested under all three VOCs and three concentration levels (0.1, 1, and 5.0 ppm). VOC samples are used to test the sensors' ability to detect and differentiate VOCs. Sensors (also referred to as a chip) are prepared using several types of thiol derivatized gold nanoparticles. The factors are: thiol compound and molar volume loading of the thiol in synthesis. The average resistance results are used to determine the VOC selectivity of the sensors tested. The results show a trend of increasing resistance as VOC concentration is increased relative to dry air; which is used as baseline for VOCs. Several sensors show a high selectivity to one or more VOCs. Overall the 57 micromoles of 4-methoxy-toluenethiol sensor shows the strongest selectivity for VOCs tested. 3. Gerfen, Kurt. 2012. Detection of Acetone in Air Using Silver Ion Exchanged ZSM-5 and Zinc Oxide Sensing Films. Master of Science thesis, University of Louisville.

  12. Water electrolyte promoted oxidation of functional thiol groups

    Lauwers, Karl; Breynaert, Eric; Rombouts, Ine; Delcour, Jan; Kirschhock, Christine

    2016-01-01

    The formation of disulfide bonds is of the utmost importance for a wide range of food products with gluten or globular proteins as functional agents. Here, the impact of mineral electrolyte composition of aqueous solutions on thiol oxidation kinetics was studied, using glutathione (GSH) and cysteine (CYS) as model systems. Interestingly, the oxidation rate of both compounds into their corresponding disulfides was significantly higher in common tap water than in ultrapure water. The systematic...

  13. Trichuris suis: thiol protease activity from adult worms.

    Hill, D E; Sakanari, J A

    1997-01-01

    Trichuris suis, the whipworm of swine, causes anemia, weight loss, anorexia, mucohemorrhagic diarrhea, and death in heavy infections. A zinc metalloprotease has been suggested to play a role in the severe enteric pathology associated with infection and the infiltration of opportunistic bacteria into deeper tissues in the swine colon. In this study, a thiol protease from gut extracts of adult T. suis and from excretory/secretory components (E/S) of adult worms was characterized using fluorogenic peptide substrates and protein substrate gels. The protease cleaved the fluorogenic substrate Z-Phe-Arg-AMC, and this cleavage was completely inhibited by the thiol protease inhibitors E-64, leupeptin, Z-Phe-Ala-CH2F, and Z-Phe-Arg-CH2F. Gelatin substrate gels and fluorescence assays using both the gut and the stichosome extracts and E/S revealed enhanced activity when 2 mM dithiothreitol or 5 mM cysteine was included in the incubation buffer, and optimal activity was seen over a pH range of 5.5 to 8.5. Incubation of gut extracts or E/S material with inhibitors of aspartic, serine, or metalloproteases had no effect on the cleavage of Z-Phe-Arg-AMC. Thiol protease activity was found in extracts of gut tissue but not in the extracts of stichocytes of adult worms. N-terminal amino acid sequencing of the protease revealed sequence homologies with cathepsin B-like thiol protease identified from parasitic and free-living nematodes. PMID:9024202

  14. Isoquinoline-mediated S-vinylation and N-vinylation of benzo[d]oxazole-2-thiol and benzo[d]thiazole-2-thiol

    Issa Yavari; Samira Nasiri-Gheidari; Anvar Mirzaei

    2012-01-01

    An effective route to S-vinylated andN-vinylated benzo[d]oxazole-2(3H)-thiones and benzo[d]thiazole-2(3H)-thionesis described via reaction ofacetylenic esters and benzo[d] oxazole-2-thiol and benzo [d]thiazole-2-thiol in the presence of 15 mol% of isoquinoline.

  15. 21 CFR 864.7375 - Glutathione reductase assay.

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Glutathione reductase assay. 864.7375 Section 864... reductase assay. (a) Identification. A glutathione reductase assay is a device used to determine the... fluorescence and photometry. The results of this assay are used in the diagnosis of liver disease,...

  16. Characterization of the chlorate reductase from Pseudomonas chloritidismutans

    Wolterink, A.F.W.M.; Schiltz, E.; Hagedoorn, P.L.; Hagen, W.R.; Kengen, S.W.M.; Stams, A.J.M.

    2003-01-01

    A chlorate reductase has been purified from the chlorate-reducing strain Pseudomonas chloritidismutans. Comparison with the periplasmic (per)chlorate reductase of strain GR-1 showed that the cytoplasmic chlorate reductase of P. chloritidismutans reduced only chlorate and bromate. Differences were al

  17. Decreased antimony uptake and overexpression of genes of thiol metabolism are associated with drug resistance in a canine isolate of Leishmania infantum.

    Gómez Pérez, Verónica; García-Hernandez, Raquel; Corpas-López, Victoriano; Tomás, Ana M; Martín-Sanchez, Joaquina; Castanys, Santiago; Gamarro, Francisco

    2016-08-01

    Visceral leishmaniasis (VL) caused by the protozoan parasite Leishmania infantum, is one of the most important zoonotic diseases affecting dogs and humans in the Mediterranean area. The presence of infected dogs as the main reservoir host of L. infantum is regarded as the most significant risk for potential human infection. We have studied the susceptibility profile to antimony and other anti-leishmania drugs (amphotericin B, miltefosine, paromomycin) in Leishmania infantum isolates extracted from a dog before and after two therapeutic interventions with meglumine antimoniate (subcutaneous Glucantime(®), 100 mg/kg/day for 28 days). After the therapeutic intervention, these parasites were significantly less susceptible to antimony than pretreatment isolate, presenting a resistance index of 6-fold to Sb(III) for promastigotes and >3-fold to Sb(III) and 3-fold to Sb(V) for intracellular amastigotes. The susceptibility profile of this resistant L. infantum line is related to a decreased antimony uptake due to lower aquaglyceroporin-1 expression levels. Additionally, other mechanisms including an increase in thiols and overexpression of enzymes involved in thiol metabolism, such as ornithine decarboxylase, trypanothione reductase, mitochondrial tryparedoxin and mitochondrial tryparedoxin peroxidase, could contribute to the resistance as antimony detoxification mechanisms. A major contribution of this study in a canine L. infantum isolate is to find an antimony-resistant mechanism similar to that previously described in other human clinical isolates. PMID:27317865

  18. Operation of trans-thylakoid thiol-metabolizing pathways in photosynthesis

    Karamoko, Mohamed; Gabilly, Stéphane T.; Hamel, Patrice P.

    2013-01-01

    Thiol oxidation to disulfides and the reverse reaction, i.e., disulfide reduction to free thiols, are under the control of catalysts in vivo. Enzymatically assisted thiol-disulfide chemistry is required for the biogenesis of all energy-transducing membrane systems. However, until recently, this had only been demonstrated for the bacterial plasma membrane. Long considered to be vacant, the thylakoid lumen has now moved to the forefront of photosynthesis research with the realization that its p...

  19. Mercuric reductase activity and evidence of broad-spectrum mercury resistance among clinical isolates of rapidly growing mycobacteria

    Resistance to mercury was evaluated in 356 rapidly growing mycobacteria belonging to eight taxonomic groups. Resistance to inorganic Hg2+ ranged from 0% among the unnamed third biovariant complex of Mycobacterium fortuitum to 83% among M. chelonae-like organisms. With cell extracts and 203Hg(NO3)2 as the substrate, mercuric reductase (HgRe) activity was demonstrable in six of eight taxonomic groups. HgRe activity was inducible and required NADPH or NADH and a thiol donor for optimai activity. Species with HgRe activity were also resistant to organomercurial compounds, including phenylmercuric acetate. Attempts at intraspecies and intragenus transfer of HgRe activity by conjugation or transformation were unsuccessful. Mercury resistance is common in rapidly growing mycobacteria and appears to function via the same inducible enzyme systems already defined in other bacterial species. This system offers potential as a strain marker for epidemiologic investigations and for studying genetic systems in rapidly growing mycobacteria

  20. Ferredoxin-thioredoxin reductase: a catalytically active dithiol group links photoreduced ferredoxin to thioredoxin functional in photosynthetic enzyme regulation

    The mechanism by which the ferredoxin-thioredoxin system activates the target enzyme, NADP-malate dehydrogenase, was investigated by analyzing the sulfhydryl status of individual protein components with [14C]iodoacetate and monobromobimane. The data indicate that ferredoxin-thioredoxin reductase (FTR)--an iron-sulfur enzyme present in oxygenic photosynthetic organisms--is the first member of a thiol chain that links light to enzyme regulation. FTR possesses a catalytically active dithiol group localized on the 13 kDa (similar) subunit, that occurs in all species investigated and accepts reducing equivalents from photoreduced ferredoxin and transfers them stoichiometrically to the disulfide form of thioredoxin m. The reduced thioredoxin m, in turn, reduces NADP-malate dehydrogenase, thereby converting it from an inactive (S-S) to an active (SH) form. The means by which FTR is able to combine electrons (from photoreduced ferredoxin) with protons (from the medium) to reduce its active disulfide group remains to be determined

  1. Crystal Growth of Thiol-Stabilized Gold Nanoparticles by Heat-Induced Coalescence

    Moon SookYoung

    2010-01-01

    Full Text Available Abstract A monolayer of dodecanethiol-stabilized gold nanoparticles changed into two-dimensional and three-dimensional self-organized structures by annealing at 323 K. Subsequent crystal growth of gold nanoparticles occurred. Thiol molecules, although chemisorbed, form relatively unstable bonds with the gold surface; a few thiols desorbed from the surface and oxidized to disulfides at 323 K, because the interaction energy between thiol macromolecules is larger than that between a thiol and a nanoparticle. The gold nanoparticles approached each other and grew into large single or twinned crystals because of the van der Waals attraction and the heat generated by the exothermic formation of disulfides.

  2. A Two-Photon Fluorescent Probe for Lysosomal Thiols in Live Cells and Tissues

    Fan, Jiangli; Han, Zhichao; Kang, Yao; Peng, Xiaojun

    2016-01-01

    Lysosome-specific fluorescent probes are exclusive to elucidate the functions of lysosomal thiols. Moreover, two-photon microscopy offers advantages of less phototoxicity, better three dimensional spatial localization, deeper penetration depth and lower self-absorption. However, such fluorescent probes for thiols are still rare. In this work, an efficient two-photon fluorophore 1,8-naphthalimide-based probe conjugating a 2,4-dinitrobenzenesulfonyl chloride and morpholine was designed and synthesized, which exhibited high selectivity and sensitivity towards lysosomal thiols by turn-on fluorescence method quantitatively and was successfully applied to the imaging of thiols in live cells and tissues by two-photon microscopy. PMID:26794434

  3. A Two-Photon Fluorescent Probe for Lysosomal Thiols in Live Cells and Tissues

    Fan, Jiangli; Han, Zhichao; Kang, Yao; Peng, Xiaojun

    2016-01-01

    Lysosome-specific fluorescent probes are exclusive to elucidate the functions of lysosomal thiols. Moreover, two-photon microscopy offers advantages of less phototoxicity, better three dimensional spatial localization, deeper penetration depth and lower self-absorption. However, such fluorescent probes for thiols are still rare. In this work, an efficient two-photon fluorophore 1,8-naphthalimide-based probe conjugating a 2,4-dinitrobenzenesulfonyl chloride and morpholine was designed and synthesized, which exhibited high selectivity and sensitivity towards lysosomal thiols by turn-on fluorescence method quantitatively and was successfully applied to the imaging of thiols in live cells and tissues by two-photon microscopy.

  4. Transsulfuration pathway thiols and methylated arginines: the Hunter Community Study.

    Arduino A Mangoni

    Full Text Available BACKGROUND: Serum homocysteine, when studied singly, has been reported to be positively associated both with the endogenous nitric oxide synthase inhibitor asymmetric dimethylarginine [ADMA, via inhibition of dimethylarginine dimethylaminohydrolase (DDAH activity] and with symmetric dimethylarginine (SDMA. We investigated combined associations between transsulfuration pathway thiols, including homocysteine, and serum ADMA and SDMA concentrations at population level. METHODS: Data on clinical and demographic characteristics, medication exposure, C-reactive protein, serum ADMA and SDMA (LC-MS/MS, and thiols (homocysteine, cysteine, taurine, glutamylcysteine, total glutathione, and cysteinylglycine; capillary electrophoresis were collected from a sample of the Hunter Community Study on human ageing [n = 498, median age (IQR = 64 (60-70 years]. RESULTS: REGRESSION ANALYSIS SHOWED THAT: a age (P = 0.001, gender (P = 0.03, lower estimated glomerular filtration rate (eGFR, P = 0.08, body mass index (P = 0.008, treatment with beta-blockers (P = 0.03, homocysteine (P = 0.02, and glutamylcysteine (P = 0.003 were independently associated with higher ADMA concentrations; and b age (P = 0.001, absence of diabetes (P = 0.001, lower body mass index (P = 0.01, lower eGFR (P<0.001, cysteine (P = 0.007, and glutamylcysteine (P < 0.001 were independently associated with higher SDMA concentrations. No significant associations were observed between methylated arginines and either glutathione or taurine concentrations. CONCLUSIONS: After adjusting for clinical, demographic, biochemical, and pharmacological confounders the combined assessment of transsulfuration pathway thiols shows that glutamylcysteine has the strongest and positive independent associations with ADMA and SDMA. Whether this reflects a direct effect of glutamylcysteine on DDAH activity (for ADMA and/or cationic amino acid transport requires further investigations.

  5. Functional Conducting Polymers via Thiol-ene Chemistry

    David C. Martin

    2012-08-01

    Full Text Available We demonstrate here that thiol-ene chemistry can be used to provide side-chain functionalized monomers based on 3,4-propylenedioxythiophene (ProDOT containing ionic, neutral, hydrophobic, and hydrophilic side chains. All reactions gave high yields and purification could generally be accomplished through precipitation. These monomers were polymerized either chemically or electro-chemically to give soluble materials or conductive films, respectively. This strategy provides for facile tuning of the solubility, film surface chemistry, and film morphology of this class of conducting polymers.

  6. "Oxygen Sensing" by Na,K-ATPase: These Miraculous Thiols.

    Bogdanova, Anna; Petrushanko, Irina Y; Hernansanz-Agustín, Pablo; Martínez-Ruiz, Antonio

    2016-01-01

    Control over the Na,K-ATPase function plays a central role in adaptation of the organisms to hypoxic and anoxic conditions. As the enzyme itself does not possess O2 binding sites its "oxygen-sensitivity" is mediated by a variety of redox-sensitive modifications including S-glutathionylation, S-nitrosylation, and redox-sensitive phosphorylation. This is an overview of the current knowledge on the plethora of molecular mechanisms tuning the activity of the ATP-consuming Na,K-ATPase to the cellular metabolic activity. Recent findings suggest that oxygen-derived free radicals and H2O2, NO, and oxidized glutathione are the signaling messengers that make the Na,K-ATPase "oxygen-sensitive." This very ancient signaling pathway targeting thiols of all three subunits of the Na,K-ATPase as well as redox-sensitive kinases sustains the enzyme activity at the "optimal" level avoiding terminal ATP depletion and maintaining the transmembrane ion gradients in cells of anoxia-tolerant species. We acknowledge the complexity of the underlying processes as we characterize the sources of reactive oxygen and nitrogen species production in hypoxic cells, and identify their targets, the reactive thiol groups which, upon modification, impact the enzyme activity. Structured accordingly, this review presents a summary on (i) the sources of free radical production in hypoxic cells, (ii) localization of regulatory thiols within the Na,K-ATPase and the role reversible thiol modifications play in responses of the enzyme to a variety of stimuli (hypoxia, receptors' activation) (iii) redox-sensitive regulatory phosphorylation, and (iv) the role of fine modulation of the Na,K-ATPase function in survival success under hypoxic conditions. The co-authors attempted to cover all the contradictions and standing hypotheses in the field and propose the possible future developments in this dynamic area of research, the importance of which is hard to overestimate. Better understanding of the processes

  7. Study of Highly Selective and Efficient Thiol Derivatization using Selenium Reagents by Mass Spectrometry

    Xu, Kehua; Zhang, Yun W.; Tang, Bo; Laskin, Julia; Roach, Patrick J.; Chen, Hao

    2010-08-15

    Biological thiols are critical physiological components and their detection often involves derivatization. This paper reports a systemic mass spectrometry (MS) investigation of the cleavage of Se-N bond by thiol to form a new Se-S bond, the new selenium chemistry for thiol labeling. Our data shows that the reaction is highly selective, rapid, reversible and efficient. For instance, among twenty amino acids, only cysteine was found to be reactive with Se-N containing reagents and the reaction takes place in seconds. By adding dithiothreitol (DTT), the newly formed Se-S bond of peptides/proteins can be reduced back to free thiol. The high selectivity and excellent reversibility of the reaction provide potential of using this chemistry for selective identification of thiol compounds or enriching and purifying thiol peptides/proteins. In addition, the derivatized thiol peptides have interesting dissociation behavior, which is tunable using different selenium reagents. For example, by introducing an adjacent nucleophilic group into the selenium reagent in the case of using ebselen, the reaction product of ebselen with glutathione (GSH) is easy to lose the selenium tag upon collision-induced dissociation (CID), which is useful to "fish out" those peptides containing free cysteine residues by precursor ion scan. By contrast, the selenium tag of N-(phenylseleno) phthalimide reagent can be stable and survive in CID process, which would be of value in pinpointing thiol location using a top-down proteomic approach. Also, the high conversion yield of the reaction allows the counting of total number of thiol in proteins. We believe that ebselen or N-(phenylseleno) phthalimide as tagging thiol-protein reagents will have important applications in both qualitative and quantitative analysis of different thiol-proteins derived from living cells by MS method.

  8. Electrodeposition of gold templated by patterned thiol monolayers

    She, Zhe; Di Falco, Andrea; Hähner, Georg; Buck, Manfred

    2016-06-01

    The electrochemical deposition of Au onto Au substrates modified by self-assembled monolayers (SAMs) was studied by linear sweep voltammetry (LSV), atomic force microscopy (AFM) and scanning electron microscopy (SEM). Patterned SAMs exhibiting electrochemical contrast were prepared by two different methods. One used microcontact printing (μCP) to generate a binary SAM of ω-(4‧-methyl-biphenyl-4-yl)-propane thiol (CH3-C6H4-C6H4-(CH2)3-SH, MBP3) and octadecane thiol (CH3(CH2)17SH, ODT). Templated by the SAM, a gold microelectrode structure was electrodeposited featuring a line 15 μm wide and 3 mm long. After transfer to an epoxy substrate the structure proved to be electrically conductive across the full length. The other patterning method applied electron beam lithography (EBL) where electrochemical contrast was achieved by crosslinking molecules in a single component SAM of MBP3. An electron dose above 250 mC/cm2 results in a high deposition contrast. The choice of parameters for the deposition/lift-off process is found to be more critical for Au compared to Cu studied previously. The origin of the differences and implications for nanoscale patterning are discussed.

  9. Regeneration of thiol-functionalized mesostructured silica adsorbents of mercury

    Arencibia, Amaya; Aguado, José; Arsuaga, Jesús M.

    2010-06-01

    The regeneration of thiol-functionalized SBA-15 adsorbents of mercury is presented in this article. The influence of temperature and pH on the adsorption process was studied. The effect due to the presence of complexing agents in aqueous solution on the desorption step was also evaluated. Hg(II) maximum adsorption capacities at different temperatures ranging from 20 °C to 60 °C were obtained and it was found that temperature does not affect the adsorption process. Mercury adsorption capacity was also determined in the presence of HNO 3 and HCl up to 3 M concentration. The comparison of the results showed that whereas hydrochloric acid exhibits an appreciable capacity to regenerate the thiol-functionalized SBA-15 adsorbent, the nitric acid results inefficient. The difference was attributed to the mercury complexing ability of chloride anion. Four complexing compounds, KBr, KSCN, (NH 2) 2CS, and HBr were tested for desorbing mercury in regeneration experiments. All agents were able to remove significant amounts of adsorbed mercury, being hydrobromic acid the complexing compound that yields the best results.

  10. Direct thiol-ene photocoating of polyorganosiloxane microparticles.

    Kuttner, Christian; Maier, Petra C; Kunert, Carmen; Schlaad, Helmut; Fery, Andreas

    2013-12-31

    This work presents the modification of polyorganosiloxane microparticles by surface-initiated thiol-ene photochemistry. By this photocoating, we prepared different core/shell particles with a polymeric shell within narrow size distributions (PDI = 0.041-0.12). As core particle, we used highly monodisperse spherical polyorganosiloxane particles prepared from (3-mercaptopropyl)trimethoxysilane (MPTMS) with a radius of 0.49 μm. We utilize the high surface coverage of mercaptopropyl functions to generate surface-localized radicals upon irradiation with UVA-light without additional photoinitiator. The continuous generation of radicals was followed by a dye degradation experiment (UV/vis spectroscopy). Surface-localized radicals were used as copolymer anchoring sites ("grafting-onto" deposition of different PB-b-PS diblock copolymers) and polymerization initiators ("grafting-from" polymerization of PS). Photocoated particles were characterized for their morphology (SEM, TEM), size, and size distribution (DLS). For PS-coated particles, the polymer content (up to 24% in 24 h) was controlled by the polymerization time upon UVA exposure. The coating thickness was evaluated by thermogravimetric analysis (TGA) using a simple analytical core/shell model. Raman spectroscopy was applied to directly follow the time-dependent consumption of thiols by photoinitiation. PMID:24320891

  11. Molecular modeling, structural analysis and identification of ligand binding sites of trypanothione reductase from Leishmania mexicana

    Ozal Mutlu

    2013-01-01

    Full Text Available Background & objectives: Trypanothione reductase (TR is a member of FAD-dependent NADPH oxidoreductase protein family and it is a key enzyme which connects the NADPH and the thiol-based redox system. Inhibition studies indicate that TR is an essential enzyme for parasite survival. Therefore, it is an attractive target enzyme for novel drug candidates. There is no structural model for TR of Leishmania mexicana (LmTR in the protein databases. In this work, 3D structure of TR from L. mexicana was identified by template-based in silico homology modeling method, resultant model was validated, structurally analyzed and possible ligand binding pockets were identified. Methods: For computational molecular modeling study, firstly, template was identified by BLAST search against PDB database. Multiple alignments were achieved by ClustalW2. Molecular modeling of LmTR was done and possible drug targeting sites were identified. Refinement of the model was done by performing local energy minimization for backbone, hydrogen and side chains. Model was validated by web-based servers. Results: A reliable 3D model for TR from L. mexicana was modeled by using L. infantum trypanothione reductase (LiTR as a template. RMSD results according to C-alpha, visible atoms and backbone were 0.809 Å, 0.732 Å and 0.728 Å respectively. Ramachandran plot indicates that model shows an acceptable stereochemistry. Conclusion: Modeled structure of LmTR shows high similarity with LiTR based on overall structural features like domains and folding patterns. Predicted structure will provide a source for the further docking studies of various peptide-based inhibitors.

  12. A fluorescent probe which allows highly specific thiol labeling at low pH

    Nielsen, Jonas W.; Jensen, Kristine Steen; Hansen, Rosa E.; Gotfredsen, Charlotte Held; Winther, Jakob R.

    2012-01-01

    Determination of the thiol-disulfide status in biological systems is challenging as redox pools are easily perturbed during sample preparation. This is particularly pertinent under neutral to mildly alkaline conditions typically required for alkylation of thiols. Here we describe the synthesis an...

  13. EFFECTS OF ATMOSPHERIC H2S ON THIOL COMPOSITION OF CROP PLANTS

    BUWALDA, F; DE KOK, LJ; Stulen, I.

    1993-01-01

    Exposure of crop plants to H2S resulted in an increase in thiol level and a change in the composition of the thiol pool. Non-leguminous species accumulated cysteine and glutathione in the light, whereas in the dark, substantial amounts of gamma-glutamyl-cysteine were also detected. In leguminous spe

  14. Antioxidant generation and regeneration in lipid bilayers: the amazing case of lipophilic thiosulfinates and hydrophilic thiols.

    Zheng, Feng; Pratt, Derek A

    2013-09-25

    We demonstrate that the garlic-derived chemopreventive agent allicin and the related anamu-derived petivericin are poor radical-trapping antioxidants in lipid bilayers, but that the in situ reaction of a lipophilic analog and a hydrophilic thiol yields an extremely potent radical-trapping antioxidant that can be recycled in the presence of excess thiol. PMID:23938951

  15. Lithium BINOL Phosphate Catalyzed Desymmetrization of meso-Epoxides with Aromatic Thiols

    Ingle, Gajendrasingh; Mormino, Michael G.; Antilla, Jon C.

    2014-01-01

    A highly enantioselective method for desymmetrization of meso-epoxides using thiols is reported. This is the first example of epoxide activation achieved using metal BINOL phosphates. The reaction has a broad scope in terms of epoxide substrates and aromatic thiol nucleophiles. The resulting β-hydroxyl sulfides are obtained in excellent yield and enantioselectivity.

  16. Rapid photochemical surface patterning of proteins in thiol-ene based microfluidic devices

    Lafleur, Josiane P.; Kwapiszewski, Radoslaw; Jensen, Thomas G.; Kutter, Jörg P.

    " and "ene" monomers present in the microfluidic chip bulk material provides a simple and efficient way of tuning the chip's surface chemistry. Here, thiol-ene chips displaying an excess of functional thiol groups at their surfaces are functionalized with biotin and streptavidin in a controlled fashion...

  17. Rapid photochemical surface patterning of proteins in thiol-ene based microfluidic devices

    Lafleur, Josiane P.; Kwapiszewski, Radoslaw; Jensen, Thomas Glasdam; Kutter, Jörg Peter

    ” and “ene” monomers present in the microfluidic chip bulk material provides a simple and efficient way of tuning the chip’s surface chemistry. Here, thiol-ene chips displaying an excess of functional thiol groups at their surfaces are functionalized with biotin and streptavidin in a controlled fashion...

  18. Mechanism-based inhibition of a mutant Escherichia coli ribonucleotide reductase (cysteine-225 → serine) by its substrate CDP

    The B1 subunit of Escherichia coli ribonucleotide reductase has been overexpressed using the pT7-5/pGP1-2 system developed by Tabor and Richardson. This method has allowed the preparation of two mutant B1 subunits in which two of the four thiols postulated to be within the active site of the enzyme, Cys-225 and Cys-759, have been changed to serines. Incubation of the [Ser225]B1 mutant with the B2 subunit, [U-14C]CDP, and the allosteric effector ATP results in production of cytosine, destruction of the tyrosyl radical in B2, radiolabeling of the protein, and cleavage of the B1 subunit into two pieces of 26 and 61.5 kDa. This process is independent of the identity of reductant. The [Ser759]B1 mutant reduces CDP in the presence of thioredoxin/thioredoxin reductase at 7.7% the rate of wild-type B1. When dithiothreitol is utilized as reductant, however, the rate of CDP reduction with [Ser759]B1 is identical to that observed with wild type

  19. Thiol-independent activity of a cholesterol-binding enterohemolysin produced by enteropathogenic Escherichia coli

    Figueirêdo P.M.S.

    2003-01-01

    Full Text Available Enterohemolysin produced by Escherichia coli associated with infant diarrhea showed characteristics similar to those of thiol-activated hemolysins produced by Gram-positive bacteria, including inactivation by cholesterol, lytic activity towards eukaryotic cells and thermoinstability. However, enterohemolysin activity was not inactivated by oxidation or by SH group-blocking agents (1 mM HgCl2, 1 mM iodoacetic acid and the hemolysin (100 µg/ml was not lethal to mice, in contrast to the lethality of the thiol-activated hemolysin family to animals. Earlier reports showed that intravenous injection of partially purified streptolysin O preparations (0.2 µg was rapidly lethal to mice. These results suggest that E. coli enterohemolysin is not a thiol-activated hemolysin, despite its ability to bind cholesterol, probably due to the absence of free thiol-group(s that characterize the active form of the thiol-activated hemolysin molecule.

  20. SCREENING OF HMG CO A REDUCTASE INHIBITOR PRODUCING MARINE ACTINOMYCETES

    SRINU, PHANI BHUSHAN,MOGES, SRILAKSHMI, SANKAR, PRABHAKAR,LAKSHMINARAYANA

    2013-01-01

    The objective of the present study was screening of 3-hydroxy-3- methyl glutaryl Co A (HMG CoA) reductase inhibitor producing marine actinomycetes. A total of 65 morphologically different actinomycetes were screened for HMG CoA reductase inhibitor production in a two stage submerged fermentation and evaluated for HMG CoA reductase inhibitor activity by agar diffusion and thin layer chromatography technique using lovostatin as a control. Among 65 marine Actinomycete strains, only one strain pr...

  1. Characterization of the chlorate reductase from Pseudomonas chloritidismutans

    Wolterink, A.F.W.M.; Schiltz, E; Hagedoorn, P.L.; Hagen, W.R.; Kengen, S.W.M.; Stams, A.J.M.

    2003-01-01

    A chlorate reductase has been purified from the chlorate-reducing strain Pseudomonas chloritidismutans. Comparison with the periplasmic (per)chlorate reductase of strain GR-1 showed that the cytoplasmic chlorate reductase of P. chloritidismutans reduced only chlorate and bromate. Differences were also found in N-terminal sequences, molecular weight, and subunit composition. Metal analysis and electron paramagnetic resonance measurements showed the presence of iron and molybdenum, which are al...

  2. Characterization of the Chlorate Reductase from Pseudomonas chloritidismutans

    2003-01-01

    A chlorate reductase has been purified from the chlorate-reducing strain Pseudomonas chloritidismutans. Comparison with the periplasmic (per)chlorate reductase of strain GR-1 showed that the cytoplasmic chlorate reductase of P. chloritidismutans reduced only chlorate and bromate. Differences were also found in N-terminal sequences, molecular weight, and subunit composition. Metal analysis and electron paramagnetic resonance measurements showed the presence of iron and molybdenum, which are al...

  3. HPLC analysis of nonprotein thiols in planktonic diatoms: Pool size, redox state and response to copper and cadmium exposure

    Rijstenbil, J.W.; Wijnholds, J.A.

    1996-01-01

    A sensitive method was developed to analyze low molecular weight thiols involved in metal homeostasis and detoxification in phytoplankton. The aims of this study were to (1) separate and measure all relevant thiols in a single HPLC run; (2) measure redox states of the thiols and (3) identify specifi

  4. Thiol-Disulfide Exchange between Glutaredoxin and Glutathione

    Iversen, Rasmus; Andersen, Peter Anders; Jensen, Kristine Steen;

    2010-01-01

    Glutaredoxins are ubiquitous thiol-disulfide oxidoreductases which catalyze the reduction of glutathione-protein mixed disulfides. Belonging to the thioredoxin family, they contain a conserved active site CXXC motif. The N-proximal active site cysteine can form a mixed disulfide with glutathione or...... an intramolecular disulfide with the C-proximal cysteine. The C-proximal cysteine is not known to be involved in the catalytic mechanism. The stability of the mixed disulfide with glutathione has been investigated in detail using a mutant variant of yeast glutaredoxin 1, in which the C......-proximal active site cysteine has been replaced with serine. The exchange reaction between the reduced protein and oxidized glutathione leading to formation of the mixed disulfide could readily be monitored by isothermal titration calorimetry (ITC) due to the enthalpic contributions from the noncovalent...

  5. Towards thiol functionalization of vanadium pentoxide nanotubes using gold nanoparticles

    Template-directed synthesis is a promising route to realize vanadate-based 1-D nanostructures, an example of which is the formation of vanadium pentoxide nanotubes and associated nanostructures. In this work, we report the interchange of long-chained alkyl amines with alkyl thiols. This reaction was followed using gold nanoparticles prepared by the Chemical Liquid Deposition (CLD) method with an average diameter of ∼0.9nm and a stability of ∼85 days. V2O5 nanotubes (VOx-NTs) with lengths of ∼2μm and internal hollow diameters of 20-100nm were synthesized and functionalized in a Au-acetone colloid with a nominal concentration of ∼4x10-3mol dm-3. The interchange reaction with dodecylamine is found only to occur in polar solvents and incorporation of the gold nanoparticles is not observed in the presence of n-decane

  6. Induction of Glutathione Synthesis and Glutathione Reductase Activity by Abiotic Stresses in Maize and Wheat

    Gábor Kocsy

    2002-01-01

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

  7. Functional thioredoxin reductase from pathogenic and free-living Leptospira spp.

    Sasoni, Natalia; Iglesias, Alberto A; Guerrero, Sergio A; Arias, Diego G

    2016-08-01

    Low molecular mass thiols and antioxidant enzymes have essential functions to detoxify reactive oxygen and nitrogen species maintaining cellular redox balance. The metabolic pathways for redox homeostasis in pathogenic (Leptospira interrogans) and free-living (Leptospira biflexa) leptospires species were not functionally characterized. We performed biochemical studies on recombinantly produced proteins to in depth analyze kinetic and structural properties of thioredoxin reductase (LinTrxR) and thioredoxin (LinTrx) from L. interrogans, and two TrxRs (LbiTrxR1 and LbiTrxR2) from L. biflexa. All the TrxRs were characterized as homodimeric flavoproteins, with LinTrxR and LbiTrxR1 catalyzing the NADPH dependent reduction of LinTrx and DTNB. The thioredoxin system from L. interrogans was able to use glutathione disulfide, lipoamide disulfide, cystine and bis-γ-glutamyl cysteine and homologous peroxiredoxin as substrates. Classic TrxR activity of LinTrxR2 had not been evidenced in vitro, but recombinant Escherichia coli cells overexpressing LbiTrxR2 showed high tolerance to oxidative stress. The enzymatic systems herein characterized could play a key role for the maintenance of redox homeostasis and the function of defense mechanisms against reactive oxidant species in Leptospira spp. Our results contribute to the general knowledge about redox biochemistry in these bacteria, positioning TrxR as a critical molecular target for the development of new anti-leptospiral drugs. PMID:27178006

  8. Mercury Binding Sites in Thiol-Functionalized Mesostructured Silica

    Thiol-functionalized mesostructured silica with anhydrous compositions of (SiO2)1-x(LSiO1.5)x, where L is a mercaptopropyl group and x is the fraction of functionalized framework silicon centers, are effective trapping agents for the removal of mercuric(II) ions from water. In the present work, we investigate the mercury-binding mechanism for representative thiol-functionalized mesostructures by atomic pair distribution function (PDF) analysis of synchrotron X-ray powder diffraction data and by Raman spectroscopy. The mesostructures with wormhole framework structures and compositions corresponding to x = 0.30 and 0.50 were prepared by direct assembly methods in the presence of a structure-directing amine porogen. PDF analyses of five mercury-loaded compositions with Hg/S ratios of 0.50-1.30 provided evidence for the bridging of thiolate sulfur atoms to two metal ion centers and the formation of chain structures on the pore surfaces. We find no evidence for Hg-O bonds and can rule out oxygen coordination of the mercury at greater than the 10% level. The relative intensities of the PDF peaks corresponding to Hg-S and Hg-Hg atomic pairs indicate that the mercury centers cluster on the functionalized surfaces by virtue of thiolate bridging, regardless of the overall mercury loading. However, the Raman results indicate that the complexation of mercury centers by thiolate depends on the mercury loading. At low mercury loadings (Hg/S (le) 0.5), the dominant species is an electrically neutral complex in which mercury most likely is tetrahedrally coordinated to bridging thiolate ligands, as in Hg(SBut)2. At higher loadings (Hg/S 1.0-1.3), mercury complex cations predominate, as evidenced by the presence of charge-balancing anions (nitrate) on the surface. This cationic form of bound mercury is assigned a linear coordination to two bridging thiolate ligands.

  9. Structure and mechanism of dimethylsulfoxide reductase, a molybdopterin-containing enzyme of DMSO reductase family

    Full text: Apart from nitrogenase, enzymes containing molybdenum are members of a superfamily, the molybdopterin-containing enzymes. Most of these enzymes catalyse an oxygen atom transfer and two electron transfer reaction. During catalysis the Mo at the active site cycles between the Mo(VI) and Mo(IV) states. The DMSO reductase family of molybdopterin-containing enzymes all contain a bis(molybdopterin guanine dinucleotide)Mo cofactor and over thirty examples have now been described. Over the last five years crystal structures of dimethylsulfoxide (DMSO) reductase and four other enzymes of the DMSO reductase family have revealed that enzymes of this family have a similar tertiary structure. The Mo atom at the active site is coordinated by four thiolate ligands provided by the dithiolene side chains of the two MGD molecules of the bis(MGD)Mo cofactor as well as a ligand provided by an amino acid side chain. In addition, an oxygen atom in the form of an oxo, hydroxo or aqua group is also coordinated to the Mo atom. In the case of dimethylsulfoxide reductase X-ray crystallography of the product-reduced species and Raman spectroscopy has demonstrated that the enzyme contains a single exchangeable oxo group that is H-bonded to W116

  10. Structure and function of NADPH-cytochrome P450 reductase and nitric oxide synthase reductase domain

    NADPH-cytochrome P450 reductase (CPR) and the nitric oxide synthase (NOS) reductase domains are members of the FAD-FMN family of proteins. The FAD accepts two reducing equivalents from NADPH (dehydrogenase flavin) and FMN acts as a one-electron carrier (flavodoxin-type flavin) for the transfer from NADPH to the heme protein, in which the FMNH ·/FMNH2 couple donates electrons to cytochrome P450 at constant oxidation-reduction potential. Although the interflavin electron transfer between FAD and FMN is not strictly regulated in CPR, electron transfer is activated in neuronal NOS reductase domain upon binding calmodulin (CaM), in which the CaM-bound activated form can function by a similar mechanism to that of CPR. The oxygenated form and spin state of substrate-bound cytochrome P450 in perfused rat liver are also discussed in terms of stepwise one-electron transfer from CPR. This review provides a historical perspective of the microsomal mixed-function oxidases including CPR and P450. In addition, a new model for the redox-linked conformational changes during the catalytic cycle for both CPR and NOS reductase domain is also discussed

  11. Thiol redox biology of trypanosomatids and potential targets for chemotherapy.

    Leroux, Alejandro E; Krauth-Siegel, R Luise

    2016-01-01

    Trypanosomatids are the causative agents of African sleeping sickness, Chagas' disease, and the different forms of leishmaniasis. This family of protozoan parasite possesses a trypanothione-based redox metabolism that provides the reducing equivalents for various vital processes such as the biosynthesis of DNA precursors and the detoxification of hydroperoxides. Almost all enzymes of the redox pathway proved to be essential and therefore fulfil one crucial prerequisite for a putative drug target. Trypanothione synthetase and trypanothione reductase are present in all trypanosomatids but absent from the mammalian host which, in addition to the essentiality, renders them highly specific. The chemotherapy research on both enzymes is further supported by the availability of high-throughput screening techniques and crystal structures. In this review we focus on the recent advances and limitations in the development of lead compounds targeting trypanothione synthetase and trypanothione reductase. We present an overview of the available inhibitors and discuss future perspectives including other components of the parasite-specific redox pathway. PMID:26592324

  12. Genetic identification of a respiratory arsenate reductase

    Saltikov, Chad W.; Newman, Dianne K.

    2003-01-01

    For more than a decade, it has been recognized that arsenate [H2AsO41-; As(V)] can be used by microorganisms as a terminal electron acceptor in anaerobic respiration. Given the toxicity of arsenic, the mechanistic basis of this process is intriguing, as is its evolutionary origin. Here we show that a two-gene cluster (arrAB; arsenate respiratory reduction) in the bacterium Shewanella sp. strain ANA-3 specifically confers respiratory As(V) reductase activity. Mutants with in-frame deletions of...

  13. Influence of N,N-dimethylaniline on the association of phenobarbital-induced cytochrome P-450 and NADPH-cytochrome c(P-450) reductase in a reconstituted rabbit liver microsomal enzyme system.

    Hlavica, P; Golly, I; Wolf, J

    1987-09-01

    N,N-Dimethylaniline when added to reaction mixtures provokes deviation from Michaelis-Menten law of the interaction kinetics of NADPH-cytochrome c(P-450) reductase (NADPH:ferrihaemoprotein oxidoreductase, EC 1.6.2.4) with highly purified phenobarbital-induced rabbit liver microsomal cytochrome P-450 (P-450LM2). This phenomenon is not associated with the low-to-high spin transition in the iron-coordination sphere of the haemoprotein, as elicited by the arylamine. Substrate-triggered departure from linearity of the kinetics is abolished by inclusion into the assay media of p-chloromercuribenzoate, hinting at a vital role in the process of thiols. Similarly, the parabolic progress curve (nH = 1.7) is transformed to a straight line (nH = 1.01) when the N-terminal reductase-binding domain in the P-450LM2 molecule is selectively blocked through covalent attachment of fluorescein isothiocyanate (FITC); such a modification does not alter the affinity of the haemoprotein for the amine substrate. Steady-state fluorescence polarization measurements reveal that N,N-dimethylaniline perturbs the motional properties of the fluorophore-bearing reductase-binding region, suggesting the induction of a conformational change. Summarizing these results, the data possibly indicate N,N-dimethylaniline-induced cooperativity in the association of reductase with P-450LM2. PMID:3113486

  14. Determination of thiol functional groups on bacteria and natural organic matter in environmental systems

    Anandha Rao, Balaji [ORNL; Lin, Hui [ORNL; Liang, Liyuan [ORNL; Gu, Baohua [ORNL

    2014-01-01

    Organic thiols (R-SH) are known to react and form complexes with some toxic soft metals such as mercury (Hg) in both biotic and abiotic systems. However, a clear understanding of these interactions is currently limited because quantifying thiols in environmental matrices is difficult due to their low abundance, susceptibility to oxidation, and measurement interference by non-thiol compounds in samples. Here, we report a fluorescence-labeling method using a maleimide containing probe, ThioGlo-1 (TG-1), to determine total thiols directly on bacterial cells and natural organic matter (NOM). We systematically evaluated the optimal thiol labeling conditions and interference from organic compounds such as disulfide, methionine, thiourea, and amine, and inorganic ions such as Na+, K+, Ca2+, Fe2+, Cl-, SO42-, HCO3-, and SCN-, and found that the method is highly sensitive and selective. Only relatively high levels of sulfide (S2-) and sulfite (SO32-) significantly interfere with the thiol analysis. The method was successful in determining thiols in a bacterium Geobacter sulfurreducens PCA and its mutants in a phosphate buffered saline solution. The measured value of ~2.1 104 thiols cell-1 (or ~0.07 mol g-1 wet cells) is in good agreement with that observed during reactions between Hg and PCA cells. Using the standard addition, we determined the total thiols of two reference NOM samples, the reduced Elliot soil humic acid and Suwanee River NOM, to be 3.6 and 0.7 mol g-1, respectively, consistent with those obtained based on their reactions with Hg.

  15. Der Thiol:Disulfid-Redox Metabolismus und der Blaulichtrezeptor Lmo0799 von Listeria monocytogenes

    Ondrusch, Nicolai

    2010-01-01

    Der Thiol-Redox-Metabolismus, der in allen lebenden Zellen zu finden ist, wirkt oxidativem Stress entgegen. Des Weiteren dient er auch der Aufrechterhaltung der intrazellulären Thiol:Disulfid-Balance, die wiederum für die Funktion vieler Proteine essentiell ist. Auch stellt er Reduktionsäquivalente für die Produktion von Desoxyribonucleotiden für die DNA-Synthese bereit und hilft oxidierte Proteine zu reparieren. Der Thiol:Disulfid-Redox-Metabolismus (TDRM) unterscheidet sich von anderen meta...

  16. Quinoline-2-thiol Derivatives as Fluorescent Sensors for Metals, pH and HNO

    Naphtali A. O’Connor

    2014-06-01

    Full Text Available A tautomeric equilibrium exists for quinoline-2-thiol and quinoline-2(1H-thione. Quantum mechanical calculations predict the thione is the major tautomer and this is confirmed by the absorption spectra. The utility of quinolone-2-thiol/quinoline-2(1H-thione as a chromophore for developing fluorescent sensors is explored. No fluorescence is observed when excited at absorption maxima, however a fluorescence increase is observed when exposed to HNO, a molecule of import as a cardiovascular therapeutic. Alkylated quinoline-2-thiol derivatives are found to be fluorescent and show a reduction in fluorescence when exposed to metals and changes in pH.

  17. Thiazolidine-Protected β-Thiol Asparagine: Applications in One-Pot Ligation-Desulfurization Chemistry.

    Sayers, Jessica; Thompson, Robert E; Perry, Kristen J; Malins, Lara R; Payne, Richard J

    2015-10-01

    The synthesis of a β-thiol asparagine derivative bearing a novel (2,4,6-trimethoxyphenyl)thiazolidine protecting group is described. The efficient incorporation of the amino acid into the N-termini of peptides is demonstrated as well as the utility of the β-thiol asparagine moiety for rapid ligation reactions with peptide thioesters. The streamlined synthesis of native peptide products could be accomplished using a one-pot radical desulfurization of the β-thiol auxiliary following the ligation event. The utility of the amino acid is highlighted in the efficient one-pot assembly of the HIV entry inhibitor enfuvirtide. PMID:26398220

  18. Synthesis of coordination compounds of copper(II) and 1.2.4-triazole thiol-5

    Present work is devoted to synthesis of coordination compounds of copper(II) and 1.2.4-triazole thiol-5. New coordination compounds of copper(II) and 1.2.4-triazole thiol-5 in aqueous-ethanol medium at molar ration Cu:L from 1:1 till 1:4, where L-1.2.4-triazole thiol-5 are synthesized. Composition and structure of synthesized coordination compounds are defined by means of chemical analysis, infrared spectroscopy, and X-ray phase analysis methods.

  19. Membrane-associated chromate reductase activity from Enterobacter cloacae.

    P. C. Wang; Mori, T.; Toda, K.; Ohtake, H

    1990-01-01

    Washed cells of Enterobacter cloacae HO1 reduced hexavalent chromium (chromate: CrO4(2-) anaerobically. Chromate reductase activity was preferentially associated with the membrane fraction of the cells. Right-side-out membrane vesicles prepared from E. cloacae cells showed high chromate reductase activities when ascorbate-reduced phenazine methosulfate was added as an electron donor.

  20. Canopy and seasonal profiles of nitrate reductase in soybeans

    Harper, J.E.; Hageman, R.H.

    1972-01-01

    Nitrate reductase activity of soybeans (Glycine max L. Merr.) was evaluated in soil plots and outdoor hydroponic gravel culture systems throughout the growing season. Nitrate reductase profiles within the plant canopy were also established. Mean activity per gram fresh weight per hour of the entire plant canopy was highest in the seedling stage while total activity (activity per gram fresh weight per hour times the total leaf weight) reached a maximum when plants were in the full bloom to midpod fill stage. Nitrate reductase activity per gram fresh weight per hour was highest in the uppermost leaf just prior to full expansion and declined with leaf positions lower in the canopy. Total nitrate reductase activity per leaf was also highest in the uppermost fully expanded leaf during early growth stages. Maximum total activity shifted to leaf positions lower in the plant canopy with later growth stages. Nitrate reductase activity of soybeans grown in hydroponic systems was significantly higher than activity of adjacent soil grown plants at later growth stages, which suggested that under normal field conditions the potential for nitrate utilization may not be realized. Nitrate reductase activity per gram fresh weight per hour and nitrate content were positively correlated over the growing season with plants grown in either soil or solution culture. Computations based upon the nitrate reductase assay of plants grown in hydroponics indicated that from 1.7 to 1.8 grams N could have been supplied to the plant via the nitrate reductase process. 11 references, 9 figures, 3 tables.

  1. Preparation and Preliminary Dielectric Characterization of Structured C60-Thiol-Ene Polymer Nanocomposites Assembled Using the Thiol-Ene Click Reaction

    Hanaa M. Ahmed

    2015-11-01

    Full Text Available Fullerene-containing materials have the ability to store and release electrical energy. Therefore, fullerenes may ultimately find use in high-voltage equipment devices or as super capacitors for high electric energy storage due to this ease of manipulating their excellent dielectric properties and their high volume resistivity. A series of structured fullerene (C60 polymer nanocomposites were assembled using the thiol-ene click reaction, between alkyl thiols and allyl functionalized C60 derivatives. The resulting high-density C60-urethane-thiol-ene (C60-Thiol-Ene networks possessed excellent mechanical properties. These novel networks were characterized using standard techniques, including infrared spectroscopy (FTIR, differential scanning calorimetry (DSC, dynamic mechanical analysis (DMA, and thermal gravimetric analysis (TGA. The dielectric spectra for the prepared samples were determined over a broad frequency range at room temperature using a broadband dielectric spectrometer and a semiconductor characterization system. The changes in thermo-mechanical and electrical properties of these novel fullerene-thiol-ene composite films were measured as a function of the C60 content, and samples characterized by high dielectric permittivity and low dielectric loss were produced. In this process, variations in chemical composition of the networks were correlated to performance characteristics.

  2. Atmospheric reactivity of alcohols, thiols and fluoroalcohols with chlorine atoms

    Garzon Ruiz, Andres

    Alcohols, thiols and fluoroalcohols are volatile organic compounds (VOCs) which are emitted to the atmosphere from both natural (vegetation, oceans, volcanoes, etc.) and anthropogenic sources (fuels, solvents, wastewater, incinerators, refrigerants, etc.). These pollutants can be eliminated from the troposphere by deposition on the terrestrial surface, direct photolysis or reaction with different tropospheric oxidants. Reactions of VOCs with tropospheric oxidants are involved in the well-known atmospheric phenomenon of photochemical smog or the production of tropospheric ozone. The oxidation of these VOCs in the troposphere is mainly initiated by reaction with OH radicals during the daytime and with NO radicals at night. However, in recent years, the oxidation by chlorine atoms (Cl) has gained great importance in the study of atmospheric reactions because they may exert some influence in the boundary layer, particularly in marine and coastal environments. In general, Cl atoms are much more reactive species than OH and NO; radicals and therefore low concentrations of Cl may compete with OH and NO3 in hydrocarbon oxidation processes. The main source of tropospheric Cl atoms is believed to be the photolysis of chlorine-containing molecules generated by heterogeneous reactions of sea salt aerosols. It has also been proposed that Cl atoms, produced in the photolysis of Cl2 emitted from industrial processes, may enhance hydrocarbon oxidation rates and ozone production in urban environments. In this work, a kinetic, theoretical and mechanistic study of the reaction of several alcohols, thiols, and fluoroalcohols with Cl atoms has been carried out. Pulsed laser photolysis-fluorescence resonance (PLP-RF) technique was used for the kinetic study as a function of temperature and pressure. An environmental chamber-Fourier transform infrared (FTIR) system was also employed in the kinetic studies. Tropospheric lifetimes of these pollutants were estimated using obtained kinetic

  3. AFM-assisted fabrication of thiol SAM pattern with alternating quantified surface potential

    Simons Janet

    2011-01-01

    Full Text Available Abstract Thiol self-assembled monolayers (SAMs are widely used in many nano- and bio-technology applications. We report a new approach to create and characterize a thiol SAMs micropattern with alternating charges on a flat gold-coated substrate using atomic force microscopy (AFM and Kelvin probe force microscopy (KPFM. We produced SAMs-patterns made of alternating positively charged, negatively charged, and hydrophobic-terminated thiols by an automated AFM-assisted manipulation, or nanografting. We show that these thiol patterns possess only small topographical differences as revealed by AFM, and distinguished differences in surface potential (20-50 mV, revealed by KPFM. The pattern can be helpful in the development of biosensor technologies, specifically for selective binding of biomolecules based on charge and hydrophobicity, and serve as a model for creating surfaces with quantified alternating surface potential distribution.

  4. Thiol-activated serine proteinases from nymphal hemolymph of the African migratory locust, Locusta migratoria migratorioides.

    Hanzon, Jacob; Smirnoff, Patricia; Applebaum, Shalom W; Mattoo, Autar K; Birk, Yehudith

    2003-02-01

    Two unique serine proteinase isoenzymes (LmHP-1 and LmHP-2) were isolated from the hemolymph of African migratory locust (Locusta migratoria migratorioides) nymphs. Both have a molecular mass of about 23 kDa and are activated by thiol-reducing agents. PMSF abolishes enzymes activity only after thiol activation, while the cysteine proteinase inhibitors E-64, iodoacetamide, and heavy metals fail to inhibit the thiol-activated enzymes. The N-terminal sequence was determined for the more-abundant LmHP-2 isoenzyme. It exhibits partial homology to that of other insect serine proteinases and similar substrate specificity and inhibition by the synthetic and protein trypsin inhibitors pABA, TLCK, BBI, and STI. The locust trypsins LmHP-1 and LmHP-2 constitute a new category of serine proteases wherein the active site of the enzyme is exposed by thiol activation without cleavage of peptide bonds. PMID:12559979

  5. Performance comparison of acrylic and thiol-acrylic resins in two-photon polymerization.

    Jiang, Lijia; Xiong, Wei; Zhou, Yushen; Liu, Ying; Huang, Xi; Li, Dawei; Baldacchini, Tommaso; Jiang, Lan; Lu, Yongfeng

    2016-06-13

    Microfabrication by two-photon polymerization is investigated using resins based on thiol-ene chemistry. In particular, resins containing different amounts of a tetrafunctional acrylic monomer and a tetrafunctional thiol molecule are used to create complex microstructures. We observe the enhancement of several characteristics of two-photon polymerization when using thiol-acrylic resins. Specifically, microfabrication is carried out using higher writing velocities and it produces stronger polymeric microstructures. Furthermore, the amount of shrinkage typically observed in the production of three-dimensional microstructures is reduced also. By means of microspectrometry, we confirm that the thiol-acrylate mixture in TPP resins promote monomer conversion inducing a higher degree of cross-linked network formation. PMID:27410383

  6. Application of metal-modified silica gels as sorbents for alkane-thiols

    In order to elaborate high effective method of purification of industry wastes from alkane-thiols the sorption properties of silica gels modified by ions of heavy metals are studied. The optimal conditions of sorption are defined.

  7. Degradable thiol-acrylate hydrogels as tunable matrices for three-dimensional hepatic culture.

    Hao, Yiting; Lin, Chien-Chi

    2014-11-01

    A degradable poly(ethylene glycol)-diacrylate (PEGDA) hydrogel system was developed using simple macromer formulations and visible light initiated thiol-acrylate photopolymerization. In addition to PEGDA, other components in this gelation system include eosin-Y as a photo-sensitizer, bi-functional thiol (dithiothreitol, DTT) as a dual-purpose co-initiator and cross-linker, and N-vinylpyrrolidone (NVP) as a co-monomer. Gelation was achieved through a mixed-mode step-chain growth polymerization mechanism under bright visible light exposure. Increasing photo-sensitizer or NVP concentrations accelerated photo-crosslinking and increased final gel stiffness. Increasing bi-functional thiol content in the prepolymer solution only increased gel stiffness to some degree. As the concentration of thiol surpassed certain range, thiol-mediated chain-transfer events caused thiol-acrylate gels to form with lower degree of cross-linking. Pendant peptide, such as integrin ligand RGDS, was more effectively immobilized in the network via a thiol-acrylate reaction (using thiol-bearing peptide Ac-CRGDS. Underline indicates cross-linkable motif) than through homo-polymerization of acrylated peptide (e.g., acryl-RGDS). The incorporation of pendant peptide comes with the expense of a lower degree of gel cross-linking, which was rectified by increasing co-monomer NVP content. Without the use of any readily degradable macromer, these visible light initiated mixed-mode cross-linked hydrogels degraded hydrolytically due to the formation of thiol-ether-ester bonds following thiol-acrylate reactions. An exponential growth relationship was identified between the hydrolytic degradation rate and bifunctional thiol content in the prepolymer solution. Finally, we evaluated the cytocompatibility of these mixed-mode cross-linked degradable hydrogels using in situ encapsulation of hepatocellular carcinoma Huh7 cells. Encapsulated Huh7 cells remained alive and proliferated as time to form cell clusters

  8. Expression, purification, crystallization and initial X-ray diffraction analysis of thiol peroxidase from Yersinia pseudotuberculosis

    Recombinant thiol peroxidase from Y. pseudotuberculosis has been purified and crystallized in three crystal forms. Thiol peroxidase is an atypical 2-Cys peroxiredoxin that reduces alkyl hydroperoxides. Wild-type and C61S mutant protein have been recombinantly expressed in Escherichia coli and purified using nickel-affinity chromatography. Initial crystallization trials yielded three crystal forms in three different space groups (P21, P64 and P212121) both in the presence and the absence of DTT

  9. Visible light cured thiol-vinyl hydrogels with tunable degradation for 3D cell culture

    Hao, Yiting; Shih, Han; Muňoz, Zachary; Kemp, Arika; Lin, Chien-Chi

    2013-01-01

    We report here a synthetically simple yet highly tunable and diverse visible light mediated thiol-vinyl gelation system for fabricating cell-instructive hydrogels. Gelation was achieved via a mixed-mode step-and-chain-growth photopolymerization using functionalized 4-arm poly(ethylene glycol) as backbone macromer, eosin-Y as photosensitizer, and di-thiol containing molecule as dual purpose co-initiator/cross-linker. N-vinylpyrrolidone (NVP) was used to accelerate gelation kinetics and to adju...

  10. Optimization of Optical Properties of Polycarbonate Film with Thiol Gold-Nanoparticles

    Claudio Larosa; Enrico Stura; Roberto Eggenhöffner; Claudio Nicolini

    2009-01-01

    A new nanostructured composite film based on thiol gold nanoparticles dispersed in polycarbonate and prepared by evaporating a solution of 1-dodecanthiol gold nanoparticles and polycarbonate was developed for applications as optical lenses. Lenses with superior mechanical properties, coloring and UV ray absorption and with the same transparency as the matrix were obtained. The supporting highly transparent polycarbonate matrix and the chloroform solution of thiol gold nanoparticles, 3 nm mean...

  11. Thiol Modification of Psyllium Husk Mucilage and Evaluation of Its Mucoadhesive Applications

    Meenakshi Bhatia; Munish Ahuja

    2013-01-01

    Thiol functionalization of psyllium was carried out to enhance its mucoadhesive potential. Thiolation of psyllium was achieved by esterification with thioglycolic acid. Thiolation was observed to change the surface morphology of psyllium from fibrous to granular and result in a slight increase in the crystallinity and swelling. Thiolated psyllium was found to contain 3.282 m moles of thiol groups/g of the polymer. Mucoadhesive applications of thiolated psylium were explored by formulating ge...

  12. Endoplasmic Reticulum Thiol Oxidase Deficiency Leads to Ascorbic Acid Depletion and Noncanonical Scurvy in Mice

    Zito, Ester; Hansen, Henning Gram; Yeo, Giles S.H.; Fujii, Junichi; Ron, David

    2012-01-01

    Summary Endoplasmic reticulum (ER) thiol oxidases initiate a disulfide relay to oxidatively fold secreted proteins. We found that combined loss-of-function mutations in genes encoding the ER thiol oxidases ERO1α, ERO1β, and PRDX4 compromised the extracellular matrix in mice and interfered with the intracellular maturation of procollagen. These severe abnormalities were associated with an unexpectedly modest delay in disulfide bond formation in secreted proteins but a profound, 5-fold lower pr...

  13. A Versatile Synthetic Extracellular Matrix Mimic via Thiol-Norbornene Photopolymerization

    Fairbanks, Benjamin D.; Schwartz, Michael P.; Halevi, Alexandra E.; Nuttelman, Charles R.; Bowman, Christopher N.; Anseth, Kristi S.

    2009-01-01

    Synthetic hydrogels with engineered, cell-mediated degradation sites are an important category of biomimetic materials. Here, hydrogels are synthesized by a step-growth reaction mechanism via a radically mediated thiol-norbornene (thiol-ene) photopolymerization. This reaction combines the advantages of ideal, homogeneous polymer network formation, facile incorporation of peptides without post-synthetic modification, and spatial and temporal control over the network evolution into a single sys...

  14. Interfacial thiol-ene photo-click reactions for forming multilayer hydrogels

    Shih, Han; Fraser, Andrew K.; Lin, Chien-Chi

    2013-01-01

    Interfacial visible light-mediated thiol-ene photo-click reactions were developed for preparing step-growth hydrogels with multilayer structures. The effect of a non-cleavage type photoinitiator eosin-Y on visible light-mediated thiol-ene photopolymerization was first characterized using in situ photo-rheometry, gel fraction, and equilibrium swelling ratio. Next, spectrophotometric properties of eosin-Y in the presence of various relevant macromer species were evaluated using UV/Vis spectrome...

  15. Oxidative stress and decreased thiol level in patients with migraine: cross-sectional study.

    Eren, Yasemin; Dirik, Ebru; Neşelioğlu, Salim; Erel, Özcan

    2015-12-01

    Although migraine is a neurological disorder known since long, its physiopathology remains unclear. Recent studies suggest that migraine is associated with oxidative stress; however, they report divergent results. The aim of the present study was to evaluate total antioxidant status (TAS), total oxidant status (TOS), oxidative stress index (OSI), and serum thiol level in migraine patients with or without aura. The study group consisted of 141 migraine patients. The control group included 70 healthy subjects. TAS, TOS, OSI were evaluated using a method developed by Erel. Serum thiol level was measured using the Hu method. No difference was found in TAS, TOS, OSI between the patients and controls. The level of thiol was significantly lower in patients than in controls. Negative correlations were detected between thiol level and Migraine Disability Assessment score in patients. Although TAS, TOS, and OSI were similar to those of the control group, serum thiol level, an important marker of antioxidant capacity, was significantly lower in migraines compared with controls, and caused more serious disability. Novel treatment approaches may be developed based on these data, and compounds containing thiol, such as alpha lipoic acid and N-acetyl cysteine, may be used in prophylaxis. PMID:25595415

  16. Purification, Characterization, and Effect of Thiol Compounds on Activity of the Erwinia carotovora L-Asparaginase

    Suchita C. Warangkar

    2010-01-01

    Full Text Available L-asparaginase was extracted from Erwinia carotovora and purified by ammonium sulfate fractionation (60–70%, Sephadex G-100, CM cellulose, and DEAE sephadex chromatography. The apparent Mr of enzyme under nondenaturing and denaturing conditions was 150 kDa and 37±0.5 kDa, respectively. L-asparaginase activity was studied in presence of thiols, namely, L-cystine (Cys, L-methionine (Met, N-acetyl cysteine (NAC, and reduced glutathione (GSH. Kinetic parameters in presence of thiols (10–400 M showed an increase in Vmax values (2000, 2223, 2380, 2500, and control 1666.7 moles mg−1min−1 and a decrease in K values (0.086, 0.076, 0.062, 0.055 and control 0.098 mM indicating nonessential mode of activation. KA values displayed propensity to bind thiols. A decrease in Vmax/K ratio in concentration plots showed inverse relationship between free thiol groups (NAC and GSH and bound thiol group (Cys and Met. Enzyme activity was enhanced in presence of thiol protecting reagents like dithiothreitol (DTT, 2-mercaptoethanol (2-ME, and GSH, but inhibited by p-chloromercurybenzoate (PCMB and iodoacetamide (IA.

  17. Interfacial thiol-ene photo-click reactions for forming multilayer hydrogels

    Shih, Han; Fraser, Andrew K.; Lin, Chien-Chi

    2014-01-01

    Interfacial visible light-mediated thiol-ene photo-click reactions were developed for preparing step-growth hydrogels with multilayer structures. The effect of a non-cleavage type photoinitiator eosin-Y on visible light-mediated thiol-ene photopolymerization was first characterized using in situ photo-rheometry, gel fraction, and equilibrium swelling ratio. Next, spectrophotometric properties of eosin-Y in the presence of various relevant macromer species were evaluated using UV/Vis spectrometry. It was determined that eosin-Y was able to re-initiate thiol-ene photo-click reaction even after light exposure. Due to its small molecular weight, most eosin-Y molecules readily leached out from the hydrogels. The diffusion of residual eosin-Y from pre-formed hydrogels was exploited for fabricating multilayer step-growth hydrogels. Interfacial hydrogel coating was formed via the same visible light-mediated gelation mechanism without adding fresh initiator. The thickness of the thiol-ene gel coating could be easily controlled by adjusting visible light exposure time, eosin-Y concentration initially loaded in the core gel, or macromer concentration in the coating solution. The major benefits of this interfacial thiol-ene coating system include its simplicity and cytocompatibility. The formation of thiol-ene hydrogels and coatings neither requires nor generates any cytotoxic components. This new gelation chemistry may have great utilities in controlled release of multiple sensitive growth factors and encapsulation of multiple cell types for tissue regeneration. PMID:23384151

  18. Human erythrocyte thiol methyltransferase: radiochemical microassay and biochemical properties

    A radiochemical microassay for the measurement of thiol methyltransferase (TMT) activity in human red blood cell (RBC) membranes has been developed. Both 2-mercaptoethanol and dithiothreitol were used as substrates for the enzyme. The pH optimum of the reaction was approximately 9.0 when glycine-NaOH was used as a buffer. The apparent Michaelis-Menten (Ksub(M)) value for the methyl donor for the reaction, S-adenosyl-L-methionine, was 43 μmol/l. Human RBC TMT activity was neither activated nor inhibited by Ca2+, Mg2+, or tropolone, but the enzyme was inhibited by SKF 525A and by reagents that react with sulfhydryl grcups. The mean TMT activity in blood from 289 randomly selected adult white subjects was 10.93 +- 3.22 units per mg protein (mean +- S.D.). The activity was the same in samples from men and women. The results of experiments in which TMT activity was measured in mixtures of RBC membranes with relatively ''low'' and relatively ''high'' activities provided no evidence that individual variations in the enzyme activity were due to variations in endogenous TMT activators or inhibitors. (Auth.)

  19. Oligomerization of Indole Derivatives with Incorporation of Thiols

    Jarl E.S. Wikberg

    2008-08-01

    Full Text Available Abstract: Two molecules of indole derivative, e.g. indole-5-carboxylic acid, reacted with one molecule of thiol, e.g. 1,2-ethanedithiol, in the presence of trifluoroacetic acid to yield adducts such as 3-[2-(2-amino-5-carboxyphenyl-1-(2-mercaptoethylthioethyl]-1Hindole-5-carboxylic acid. Parallel formation of dimers, such as 2,3-dihydro-1H,1'H-2,3'-biindole-5,5'-dicarboxylic acid and trimers, such as 3,3'-[2-(2-amino-5-carboxyphenyl ethane-1,1-diyl]bis(1H-indole-5-carboxylic acid of the indole derivatives was also observed. Reaction of a mixture of indole and indole-5-carboxylic acid with 2-phenylethanethiol proceeded in a regioselective way, affording 3-[2-(2-aminophenyl-1-(phenethylthioethyl]-1H-indole-5-carboxylic acid. An additional product of this reaction was 3-[2-(2-aminophenyl-1-(phenethylthioethyl]-2,3-dihydro-1H,1'H-2,3'-biindole-5'-carboxylic acid, which upon standing in DMSO-d6 solution gave 3-[2-(2-aminophenyl-1-(phenethylthioethyl]-1H,1'H-2,3'-biindole-5'-carboxylic acid. Structures of all compounds were elucidated by NMR, and a mechanism for their formation was suggested.

  20. Cytokinin Determines Thiol-Mediated Arsenic Tolerance and Accumulation.

    Mohan, Thotegowdanapalya C; Castrillo, Gabriel; Navarro, Cristina; Zarco-Fernández, Sonia; Ramireddy, Eswarayya; Mateo, Cristian; Zamarreño, Angel M; Paz-Ares, Javier; Muñoz, Riansares; García-Mina, Jose M; Hernández, Luis E; Schmülling, Thomas; Leyva, Antonio

    2016-06-01

    The presence of arsenic in soil and water is a constant threat to plant growth in many regions of the world. Phytohormones act in the integration of growth control and stress response, but their role in plant responses to arsenic remains to be elucidated. Here, we show that arsenate [As(V)], the most prevalent arsenic chemical species in nature, causes severe depletion of endogenous cytokinins (CKs) in the model plant Arabidopsis (Arabidopsis thaliana). We found that CK signaling mutants and transgenic plants with reduced endogenous CK levels showed an As(V)-tolerant phenotype. Our data indicate that in CK-depleted plants exposed to As(V), transcript levels of As(V)/phosphate-transporters were similar or even higher than in wild-type plants. In contrast, CK depletion provoked the coordinated activation of As(V) tolerance mechanisms, leading to the accumulation of thiol compounds such as phytochelatins and glutathione, which are essential for arsenic sequestration. Transgenic CK-deficient Arabidopsis and tobacco lines show a marked increase in arsenic accumulation. Our findings indicate that CK is an important regulatory factor in plant adaptation to arsenic stress. PMID:27208271

  1. Solution structure of an arsenate reductase-related protein, YffB, from Brucella melitensis, the etiological agent responsible for brucellosis

    B. melitensis is a NIAID Category B microorganism that is responsible for brucellosis and is a potential agent for biological warfare. Here, the solution structure of the 116-residue arsenate reductase-related protein Bm-YffB (BR0369) from this organism is reported. Brucella melitensis is the etiological agent responsible for brucellosis. Present in the B. melitensis genome is a 116-residue protein related to arsenate reductases (Bm-YffB; BR0369). Arsenate reductases (ArsC) convert arsenate ion (H2AsO4−), a compound that is toxic to bacteria, to arsenite ion (AsO2−), a product that may be efficiently exported out of the cell. Consequently, Bm-YffB is a potential drug target because if arsenate reduction is the protein’s major biological function then disabling the cell’s ability to reduce arsenate would make these cells more sensitive to the deleterious effects of arsenate. Size-exclusion chromatography and NMR spectroscopy indicate that Bm-YffB is a monomer in solution. The solution structure of Bm-YffB shows that the protein consists of two domains: a four-stranded mixed β-sheet flanked by two α-helices on one side and an α-helical bundle. The α/β domain is characteristic of the fold of thioredoxin-like proteins and the overall structure is generally similar to those of known arsenate reductases despite the marginal sequence similarity. Chemical shift perturbation studies with 15N-labeled Bm-YffB show that the protein binds reduced glutathione at a site adjacent to a region similar to the HX3CX3R catalytic sequence motif that is important for arsenic detoxification activity in the classical arsenate-reductase family of proteins. The latter observation supports the hypothesis that the ArsC-YffB family of proteins may function as glutathione-dependent thiol reductases. However, comparison of the structure of Bm-YffB with the structures of proteins from the classical ArsC family suggest that the mechanism and possibly the function of Bm-YffB and other

  2. Molecular modeling of the reductase domain to elucidate the reaction mechanism of reduction of peptidyl thioester into its corresponding alcohol in non-ribosomal peptide synthetases

    Lee Gwang

    2010-01-01

    Full Text Available Abstract Background Nonribosomal peptide synthetases (NRPSs are multienzymatic, multidomain megasynthases involved in the biosynthesis of pharmaceutically important nonribosomal peptides. The peptaibol synthetase from Trichoderma virens (TPS is an important member of the NRPS family that exhibits antifungal properties. The majority of the NRPSs terminate peptide synthesis with the thioesterase (TE domain, which either hydrolyzes the thioester linkage, releasing the free peptic acid, or catalyzes the intramolecular macrocyclization to produce a macrolactone product. TPS is an important NRPS that does not encompass a TE domain, but rather a reductase domain (R domain to release the mature peptide product reductively with the aid of a NADPH cofactor. However, the catalytic mechanism of the reductase domain has not yet been elucidated. Results We present here a three-dimensional (3D model of the reductase domain based on the crystal structure of vestitone reductase (VR. VR belongs to the short-chain dehydrogenase/reductase (SDR superfamily and is responsible for the nicotinamide dinucleotide phosphate (NADPH-dependent reduction of the substrate into its corresponding secondary alcohol product. The binding sites of the probable linear substrates, alamethicin, trichotoxin, antiamoebin I, chrysopermin C and gramicidin, were identified within the modeled R domain using multiple docking approaches. The docking results of the ligand in the active site of the R domain showed that reductase side chains have a high affinity towards ligand binding, while the thioester oxygen of each substrate forms a hydrogen bond with the OH group of Tyr176 and the thiol group of the substrate is closer to the Glu220. The modeling and docking studies revealed the reaction mechanism of reduction of thioester into a primary alcohol. Conclusion Peptaibol biosynthesis incorporates a single R domain, which appears to catalyze the four-electron reduction reaction of a peptidyl

  3. Thioredoxin f1 and NADPH-Dependent Thioredoxin Reductase C Have Overlapping Functions in Regulating Photosynthetic Metabolism and Plant Growth in Response to Varying Light Conditions

    Thormählen, Ina; Meitzel, Tobias; Groysman, Julia; Öchsner, Alexandra Bianca; von Roepenack-Lahaye, Edda; Naranjo, Belén; Cejudo, Francisco J; Geigenberger, Peter

    2015-01-01

    . In Arabidopsis (Arabidopsis thaliana) mutants, combined, but not single, deficiencies of Trx f1 and NTRC led to severe growth inhibition and perturbed light acclimation, accompanied by strong impairments of Calvin-Benson cycle activity and starch accumulation. Light activation of key enzymes of these...... pathways, fructose-1,6-bisphosphatase and ADP-glucose pyrophosphorylase, was almost completely abolished. The subsequent increase in NADPH-NADP(+) and ATP-ADP ratios led to increased nitrogen assimilation, NADP-malate dehydrogenase activation, and light vulnerability of photosystem I core proteins. In an......Two different thiol redox systems exist in plant chloroplasts, the ferredoxin-thioredoxin (Trx) system, which depends on ferredoxin reduced by the photosynthetic electron transport chain and, thus, on light, and the NADPH-dependent Trx reductase C (NTRC) system, which relies on NADPH and thus may...

  4. Structure and expression of human dihydropteridine reductase

    Dihydropteridine reductase catalyzes the NADH-mediated reduction of quinonoid dihydrobiopterin and is an essential component of the pterindependent aromatic amino acid hydroxylating systems. A cDNA for human DHPR was isolated from a human liver cDNA library in the vector λgt11 using a monospecific antibody against sheep DHPR. The nucleic acid sequence and amino acid sequence of human DHPR were determined from a full-length clone. A 112 amino acid sequence of sheep DHPR was obtained by sequencing purified sheep DHPR. This sequence is highly homologous to the predicted amino acid sequence of the human protein. Gene transfer of the recombinant human DHPR into COS cells leads to expression of DHPR enzymatic activity. These results indicate that the cDNA clone identified by antibody screening is an authentic and full-length cDNA for human DHPR

  5. Nitrite Reductase Activity in Engineered Azurin Variants.

    Berry, Steven M; Strange, Jacob N; Bladholm, Erika L; Khatiwada, Balabhadra; Hedstrom, Christine G; Sauer, Alexandra M

    2016-05-01

    Nitrite reductase (NiR) activity was examined in a series of dicopper P.a. azurin variants in which a surface binding copper site was added through site-directed mutagenesis. Four variants were synthesized with copper binding motifs inspired by the catalytic type 2 copper binding sites found in the native noncoupled dinuclear copper enzymes nitrite reductase and peptidylglycine α-hydroxylating monooxygenase. The four azurin variants, denoted Az-NiR, Az-NiR3His, Az-PHM, and Az-PHM3His, maintained the azurin electron transfer copper center, with the second designed copper site located over 13 Å away and consisting of mutations Asn10His,Gln14Asp,Asn16His-azurin, Asn10His,Gln14His,Asn16His-azurin, Gln8Met,Gln14His,Asn16His-azurin, and Gln8His,Gln14His,Asn16His-azurin, respectively. UV-visible absorption spectroscopy, EPR spectroscopy, and electrochemistry of the sites demonstrate copper binding as well as interaction with small exogenous ligands. The nitrite reduction activity of the variants was determined, including the catalytic Michaelis-Menten parameters. The variants showed activity (0.34-0.59 min(-1)) that was slower than that of native NiRs but comparable to that of other model systems. There were small variations in activity of the four variants that correlated with the number of histidines in the added copper site. Catalysis was found to be reversible, with nitrite produced from NO. Reactions starting with reduced azurin variants demonstrated that electrons from both copper centers were used to reduce nitrite, although steady-state catalysis required the T2 copper center and did not require the T1 center. Finally, experiments separating rates of enzyme reduction from rates of reoxidation by nitrite demonstrated that the reaction with nitrite was rate limiting during catalysis. PMID:27055058

  6. The arsenic hyperaccumulating Pteris vittata expresses two arsenate reductases

    Patrizia Cesaro; Chiara Cattaneo; Elisa Bona; Graziella Berta; Maria Cavaletto

    2015-01-01

    Enzymatic reduction of arsenate to arsenite is the first known step in arsenate metabolism in all organisms. Although the presence of one mRNA arsenate reductase (PvACR2) has been characterized in gametophytes of P. vittata, no arsenate reductase protein has been directly observed in this arsenic hyperaccumulating fern, yet. In order to assess the possible presence of arsenate reductase in P. vittata, two recombinant proteins, ACR2-His6 and Trx-His6-S-Pv2.5–8 were prepared in Escherichia coli...

  7. Small-scale polymer structures enabled by thiol-ene copolymer systems

    Kasprzak, Scott Edward

    2009-12-01

    The research described herein is aimed at exploring the thermomechanical properties of thiol-ene polymers in bulk form, investigating the ability of thiol-ene polymers to behave desirably as photolithographic media, and providing the first characterization of the mechanical properties of two-photon stereolithography-produced polymer structures. The thiol-ene polymerization reaction itself is well-characterized and described in the literature, but the thermomechanical properties of thiol-ene and thiol-ene/acrylate polymers still require more rigorous study. Understanding the behavior of thiol-ene networks is a crucial step towards their expanded use in bulk form, and particularly in specialized applications such as shape memory devices. Additionally, the thiol-ene polymerization reaction mechanism exhibits unique properties which make these polymers well suited to photolithography, overcoming the typical dichotomy of current materials which either exhibit excellent photolithographic behavior or have controllable properties. Finally, before two-photon stereolithography can create mechanisms and devices which can serve any mechanically functional role, the mechanical properties of the polymers they produce must be quantitatively characterized, which is complicated by the extremely small scale at which these structures are produced. As such, mechanical characterization to date has been strictly qualitative. Fourier transfer infrared spectroscopy revealed functional group conversion information and sol-fraction testing revealed the presence of unconverted monomer and impurities, while dynamic mechanical analysis (DMA) and tensile testing revealed the thermomechanical responses of the systems. Nanoindentation was employed to characterize the mechanical properties of micrometer-scale polymer structures produced by two-photon stereolithography. Optical and electron microscopy were exploited to provide both quantitative and qualitative evaluations of thiol-ene/acrylate and

  8. A Study of Functional Polymer Colloids Prepared Using Thiol-Ene/Yne Click Chemistry

    Durham, Olivia Z.

    This project demonstrates the first instance of thiol-ene chemistry as the polymerization method for the production of polymer colloids in two-phase heterogeneous suspensions, miniemulsions, and emulsions. This work was also expanded to thiol-yne chemistry for the production of polymer particles containing increased crosslinking density. The utility of thiol-ene and thiol-yne chemistries for polymerization and polymer modification is well established in bulk systems. These reactions are considered 'click' reactions, which can be defined as processes that are both facile and simple, offering high yields with nearly 100% conversion, no side products, easy product separation, compatibility with a diverse variety of commercially available starting materials, and orthogonality with other chemistries. In addition, thiol-ene and thiol-yne chemistry follow a step-growth mechanism for the development of highly uniform polymer networks, where polymer growth is dependent on the coupling of functional groups. These step-growth polymerization systems are in stark contrast to the chain-growth mechanisms of acrylic and styrenic monomers that have dominated the field of conventional heterogeneous polymerizations. Preliminary studies evaluated the mechanism of particle production in suspension and miniemulsion systems. Monomer droplets were compared to the final polymer particles to confirm that particle growth occurred through the polymerization of monomer droplets. Additional parameters examined include homogenization energy (mechanical mixing), diluent species and concentration, and monomer content. These reactions were conducted using photoinitiation to yield particles in a matter of minutes with diameters in the size range of several microns to hundreds of microns in suspensions or submicron particles in miniemulsions. Improved control over the particle size and size distribution was examined through variation of reaction parameters. In addition, a method of seeded suspension

  9. Synthesis of alkenyl sulfides through the iron-catalyzed cross-coupling reaction of vinyl halides with thiols.

    Lin, Yun-Yung; Wang, Yu-Jen; Lin, Che-Hung; Cheng, Jun-Hao; Lee, Chin-Fa

    2012-07-20

    We report here the iron-catalyzed cross-coupling reaction of alkyl vinyl halides with thiols. While many works are devoted to the coupling of thiols with alkyl vinyl iodides, interestingly, the known S-vinylation of vinyl bromides and chlorides is limited to 1-(2-bromovinyl)benzene and 1-(2-chlorovinyl)benzene. Investigation on the coupling reaction of challenging alkyl vinyl bromides and chlorides with thiols is rare. Since the coupling of 1-(2-bromovinyl)benzene and 1-(2-chlorovinyl)benzene with thiols can be performed in the absence of any catalyst, here we focus on the coupling of thiols with alkyl vinyl halides. This system is generally reactive for alkyl vinyl iodides and bromides to provide the products in good yields. 1-(Chloromethylidene)-4-tert-butyl-cyclohexane was also coupled with thiols, giving the targets in moderate yields. PMID:22708836

  10. Covalent thiol adducts arising from reactive intermediates of cocaine biotransformation.

    Schneider, Kevin J; DeCaprio, Anthony P

    2013-11-18

    Exposure to cocaine results in the depletion of hepatocellular glutathione and macromolecular protein binding in humans. Such cocaine-induced responses have generally been attributed to oxidative stress and reactive metabolites resulting from oxidative activation of the cocaine tropane nitrogen. However, little conclusive data exists on the mechanistic pathways leading to protein modification or the structure and specificity of cocaine-derived adduction products. We now report a previously uncharacterized route of cocaine bioactivation leading to the covalent adduction of biological thiols, including cysteine and glutathione. Incubation of cocaine with biological nucleophiles in an in vitro biotransformation system containing human liver microsomes identified a monooxygenase-mediated event leading to the oxidation of, and subsequent sulfhydryl addition to, the cocaine aryl moiety. Adduct structures were confirmed using ultra-high performance liquid chromatography coupled to high resolution, high mass accuracy mass spectrometry. Examination of assays containing transgenic bactosomes expressing single human cytochrome P450 isoforms determined the role of P450s 1A2, 2C19, and 2D6 in the oxidation process resulting in adduct formation. P450-catalyzed aryl epoxide formation and subsequent attack by free nucleophilic moieties is consistent with the resulting adduct structures, mechanisms of formation, and the empirical observation of multiple structural and stereo isomers. Analogous adduction mechanisms were maintained across all sulfhydryl-containing nucleophile models examined; N-acetylcysteine, glutathione, and a synthetic cysteine-containing hexapeptide. Predictive in silico calculations of molecular reactivity and electrophilicity/nucleophilicity were compared to the results of in vitro assay incubations in order to better understand the adduction process using the principles of hard and soft acid and base (HSAB) theory. This study elucidated a novel metabolic

  11. Regulation of ribonucleotide reductase by Spd1 involves multiple mechanisms

    Nestoras, Konstantinos; Mohammed, Asma Hadi; Schreurs, Ann-Sofie; Fleck, Oliver; Watson, Adam T; Poitelea, Marius; O'Shea, Charlotte; Chahwan, Charly; Holmberg, Christian; Kragelund, Birthe B; Nielsen, Olaf; Osborne, Mark; Carr, Antony M; Liu, Cong

    2010-01-01

    The correct levels of deoxyribonucleotide triphosphates and their relative abundance are important to maintain genomic integrity. Ribonucleotide reductase (RNR) regulation is complex and multifaceted. RNR is regulated allosterically by two nucleotide-binding sites, by transcriptional control, and...

  12. Structural features of the ribonucleotide reductase of Aujeszky's disease virus.

    Kaliman, A V; Boldogköi, Z; Fodor, I

    1994-01-01

    A gene construct of the Aujeszky's disease virus (ADV) genome was prepared and the DNA fragment encoding the ribonucleotide reductase was structurally characterized. We determined the entire DNA sequence of two adjacent open reading frames of the ribonucleotide reductase genes with the intergenic sequence of nine base pairs. From the sequence analysis we predict that Aujeszky's disease virus encodes a ribonucleotide reductase which comprises two polypeptides--large and small subunits, with sizes of 835 and 303 amino acids, respectively. Nucleotide and amino acid sequences of the large and small subunits of the Aujeszky's disease virus ribonucleotide reductase have been compared with that of other herpesviruses, and structural features of both proteins have been characterized. PMID:7810419

  13. Electrochemistry behavior of endogenous thiols on fluorine doped tin oxide electrodes

    Rojas, Luciana; Molero, Leonard; Tapia, Ricardo A.; Rio, Rodrigo del; Valle, M. Angelica del; Antilen, Monica [Departamento de Quimica Inorganica, Facultad de Quimica, Pontificia Universidad Catolica de Chile, Av Vicuna Mackenna 4860, Casilla 306, Correo 22, Macul, Santiago (Chile); Armijo, Francisco, E-mail: jarmijom@uc.cl [Departamento de Quimica Inorganica, Facultad de Quimica, Pontificia Universidad Catolica de Chile, Av Vicuna Mackenna 4860, Casilla 306, Correo 22, Macul, Santiago (Chile)

    2011-10-01

    Highlights: > The first time that fluorine doped tin oxide electrodes are used for the electrooxidation of endogenous thiols. > Low potentials of electrooxidation were obtained for the different thiols. > The electrochemical behavior of thiols depends on the pH and the ionic electroactive species, the electrooxidation proceeds for a process of adsorption of electroactive species on FTO and high values the heterogeneous electron tranfer rate constant of the reaction were obtained. - Abstract: In this work the electrochemical behavior of different thiols on fluorine doped tin oxide (FTO) electrodes is reported. To this end, the mechanism of electrochemical oxidation of glutathione (GSH), cysteine (Cys), homocysteine (HCys) and acetyl-cysteine (ACys) at different pH was investigated. FTO showed electroactivity for the oxidation of the first three thiols at pH between 2.0 and 4.0, but under these conditions no acetyl-cysteine oxidation was observed on FTO. Voltammetric studies of the electro-oxidation of GSH, Cys and HCys showed peaks at about 0.35, 0.29, and 0.28 V at optimum pH 2.4, 2.8 and 3.4, respectively. In addition, this study demonstrated that GSH, Cys and HCys oxidation occurs when the zwitterion is the electro-active species that interact by adsorption on FTO electrodes. The overall reaction involves 4e{sup -}/4H{sup +} and 2e{sup -}/2H{sup +}, respectively, for HCys and for GSH and Cys and high heterogeneous electron transfer rate constants. Besides, the use of FTO for the determination of different thiols was evaluated. Experimental square wave voltammetry shows a linear current vs. concentrations response between 0.1 and 1.0 mM was found for HCys and GSH, indicating that these FTO electrodes are promising candidates for the efficient electrochemical determination of these endogenous thiols.

  14. Aldose reductase inhibitory activity and antioxidant capacity of pomegranate extracts

    Karasu, Çimen; CUMAOĞLU, Ahmet; Gürpinar, Ali Rifat; Kartal, Murat; Kovacikova, Lucia; Milackova, Ivana; Stefek, Milan

    2012-01-01

    The pomegranate, Punica granatum L., has been the subject of current interest as a medicinal agent with wide-ranging therapeutic indications. In the present study, pomegranate ethanolic seed and hull extracts were tested, in comparison with a commercial sample, for the inhibition of aldose reductase, an enzyme involved in the etiology of diabetic complications. In vitro inhibition of rat lens aldose reductase was determined by a conventional method. Pomegranate ethanolic hull extract and comm...

  15. Evaluation of Thiol Raman Activities and pKa Values Using Internally Referenced Raman-Based pH Titration.

    Suwandaratne, Nuwanthi; Hu, Juan; Siriwardana, Kumudu; Gadogbe, Manuel; Zhang, Dongmao

    2016-04-01

    Thiols, including organothiol and thiol-containing biomolecules, are among the most important classes of chemicals that are used broadly in organic synthesis, biological chemistry, and nanosciences. Thiol pKa values are key indicators of thiol reactivity and functionality. Reported herein is an internally referenced Raman-based pH titration method that enables reliable quantification of thiol pKa values for both mono- and dithiols in water. The degree of thiol ionization is monitored directly using the peak intensity of the S-H stretching feature in the 2600 cm(-1) region relative to an internal reference peak as a function of the titration solution's pH. The thiol pKa values and Raman activity relative to its internal reference were then determined by curve fitting the experimental data with equations derived on the basis of the Henderson-Hasselbalch equation. Using this Raman titration method, we determined for the first time the first and second thiol pKa values for 1,2-benzenedithiol in water. This Raman-based method is convenient to implement, and its underlying theory is easy to follow. It should therefore have broad application for thiol pKa determinations and verification. PMID:26956547

  16. Facile homolytic CS bond breaking: the reaction of 2-propene-1-thiol on Mo(110)

    Wiegand, B. C.; Friend, C. M.; Uvdal, P.; Napier, M. E.

    1996-06-01

    Investigations of 2-propene-1-thiol on Mo(110) show that the facility for thiol desulfurization correlates with the homolytic CS bond strength of the thiol. A substantial amount of CS bond scission occurs upon adsorption of 2-propene-1-thiol on Mo(110) at 120 K based on X-ray photoelectron data. 2-Propene-1-thiol has an extremely weak CS bond because of the resonance stabilization of the allyl radical. Propene, the product of 2-propene-1-thiol hydrogenolysis, evolves into the gas phase in the range 160-250 K; the propene evolution being controlled by the rate of desorption. Vibrational (electron energy loss) spectra are consistent with formation of 2-propene-1-thiolate via SH bond cleavage, as well as hydrocarbon fragments at low temperature. A large fraction of the thiolate desulfurizes at 120 K to afford propene. A second hydrocarbon species is formed in the range 160-200 K, characterized by a low-frequency CH stretch mode at 2770 cm -1, which reacts further below 250 K. The CC bond in the thiol and in the propene product leads to a greater degree of non-selective reaction than is anticipated from a comparison of saturated thiols. Only in the order of 50% of the thiol reacts to afford propene, despite the facility for CS bond scission.

  17. An overview on 5alpha-reductase inhibitors.

    Aggarwal, Saurabh; Thareja, Suresh; Verma, Abhilasha; Bhardwaj, Tilak Raj; Kumar, Manoj

    2010-02-01

    Benign prostatic hyperplasia (BPH) is the noncancerous proliferation of the prostate gland associated with benign prostatic obstruction and lower urinary tract symptoms (LUTS) such as frequency, hesitancy, urgency, etc. Its prevalence increases with age affecting around 70% by the age of 70 years. High activity of 5alpha-reductase enzyme in humans results in excessive dihydrotestosterone levels in peripheral tissues and hence suppression of androgen action by 5alpha-reductase inhibitors is a logical treatment for BPH as they inhibit the conversion of testosterone to dihydrotestosterone. Finasteride (13) was the first steroidal 5alpha-reductase inhibitor approved by U.S. Food and Drug Administration (USFDA). In human it decreases the prostatic DHT level by 70-90% and reduces the prostatic size. Dutasteride (27) another related analogue has been approved in 2002. Unlike Finasteride, Dutasteride is a competitive inhibitor of both 5alpha-reductase type I and type II isozymes, reduced DHT levels >90% following 1 year of oral administration. A number of classes of non-steroidal inhibitors of 5alpha-reductase have also been synthesized generally by removing one or more rings from the azasteroidal structure or by an early non-steroidal lead (ONO-3805) (261). In this review all categories of inhibitors of 5alpha-reductase have been covered. PMID:19879888

  18. Regulative roles of glutathione reductase and four glutaredoxins in glutathione redox, antioxidant activity, and iron homeostasis of Beauveria bassiana.

    Zhang, Long-Bin; Tang, Li; Ying, Sheng-Hua; Feng, Ming-Guang

    2016-07-01

    Multiple glutaredoxins (Grx) and glutathione reductase (Glr) are vital for the thiol-disulfide redox system in budding yeast but generally unexplored in filamentous fungi. Here we characterized the Beauveria bassiana redox system comprising dithiol Grx1, monothiol Grx2-4, Grx-like Grx5, and Glr orthologue. Each grx or glr deletion was compensated by increased transcripts of some other grx genes in normal cultures. Particularly, grx3 compensated the absence of grx1, grx2, grx5, or glr under oxidative stress while its absence was compensated only by undeletable grx4 under normal conditions but by most of other undeleted grx and glr genes in response to menadione. Consequently, the redox state was disturbed in Δglr more than in Δgrx3 but not in Δgrx1/2/5. Superoxide dismutases were more active in normal Δgrx1-3 cultures but less in Δgrx5 or Δglr response to menadione. Total catalase activity increased differentially in all the mutant cultures stressed with or without H2O2 while total peroxidase activity decreased more in the normal or H2O2-stressed culture of Δglr than of Δgrx3. Among the mutants, Δgrx3 showed slightly increased sensitivity to menadione or H2O2; Δglr exhibited greater sensitivity to thiol-oxidizing diamide than thiol-reducing 1-chloro-2,4-dinitrobenzene as well as increased sensitivity to the two oxidants. Intriguingly, all the mutants grew slower in a Fe(3+)-inclusive medium perhaps due to elevated transcripts of two Fe(3+) transporter genes. More or fewer phenotypes linked with biocontrol potential were altered in four deletion mutants excluding Δgrx5. All the changes were restored by targeted gene complementation. Overall, Grx3 played more critical role than other Grx homologues in the Glr-dependent redox system of the fungal entomopathogen. PMID:26969041

  19. Pore surface engineering in a zirconium metal–organic framework via thiol-ene reaction

    Gui, Bo; Hu, Guiping; Zhou, Tailin; Wang, Cheng, E-mail: chengwang@whu.edu.cn

    2015-03-15

    A porous olefin-functionalized Zr(IV)-based metal–organic framework, denoted as UiO-68-allyl, has been constructed. Our results clearly demonstrated that the surface of UiO-68-allyl could be decorated with organic molecule (ethanethiol) via thiol-ene reaction. More importantly, the crystallinity of the framework were maintained during the post-synthetic modification process. However, the microporosity of the framework is retained but the surface area decreased, due to the grafting of ethylthio groups into the pores. From our studies, we can conclude that the strategy of post-synthetic modification of UiO-68-allyl via thiol-ene reaction may be general. Furthermore, we may anchor other desired functional group onto the pore walls in Zr-MOFs via thiol-ene reaction, enabling more potential applications. - graphical abstract: In this manuscript, we reported the post-synthetic modification of an olefin-functionalized Zr(IV)-based metal–organic framework via thiol-ene reaction. - Highlights: • A porous olefin-functionalized Zr(IV)-based metal–organic framework has been constructed. • The surface of olefin-functionalized Zr-MOF could be decorated with organic molecules via thiol-ene reaction. • The crystallinity and permanent porosity of the framework were maintained during the post-synthetic modification process.

  20. Proteinase from germinating bean cotyledons. Evidence for involvement of a thiol group in catalysis.

    Csoma, C; Polgár, L

    1984-09-15

    To degrade storage proteins germinating seeds synthesize proteinases de novo that can be inhibited by thiol-blocking reagents [Baumgartner & Chrispeels (1977) Eur. J. Biochem. 77, 223-233]. We have elaborated a procedure for isolation of such a proteinase from the cotyledons of Phaseolus vulgaris. The purification procedure involved fractionation of the cotyledon homogenate with acetone and with (NH4)2SO4 and successive chromatographies on DEAE-cellulose, activated thiol-Sepharose Sepharose and Sephacryl S-200. The purified enzyme has an Mr of 23,400, proved to be highly specific for the asparagine side chain and blocking of its thiol group resulted in loss of the catalytic activity. The chemical properties of the thiol group of the bean enzyme were investigated by acylation with t-butyloxycarbonyl-L-asparagine p-nitro-phenyl ester and by alkylations with iodoacetamide and iodoacetate. Deviations from normal pH-rate profile were observed, which indicated that the thiol group is not a simple functional group, but constitutes a part of an interactive system at the active site. The pKa value for acylation and the magnitude of the rate constant for alkylation with iodoacetate revealed that the bean proteinase possesses some properties not shared by papain and the other cysteine proteinases studied to date. PMID:6385962

  1. Radiosensitization by misonidazole during recovery of cellular thiols following depletion by BSO or DEM

    V79 cells have been depleted of their endogenous thiols by treatment with 100 microM BSO for 16-18 hr, or 0.5 mM DEM for 1 hr. The recovery of cellular thiols after removal of the drugs was determined by h.p.l.c. or flow cytometry and the sensitizer enhancement ratio for 100 microM misonidazole was measured as a function of time after removal of the drugs. The SER of 1.2 for control (hypoxic) cells increased to 1.8 for BSO treated (hypoxic) cells and 2.2 for DEM treated ones, when thiol levels were below 10% of controls. The SER and thiol levels returned to control values within 5 hr of removing DEM. After BSO there was little change during the first 5 hr and then a gradual return to normal values by 24 hrs. The major fall in the SER after removal of the drugs occurred as the cellular thiols increased to 60% of control values

  2. Characterization of photopolymerizable nanoparticle-(thiol-ene) polymer composites for volume holographic recording at 404 nm

    Kawana, Masaru; Takahashi, Jun-ichiro; Yasui, Satoru; Tomita, Yasuo

    2014-05-01

    We report on volume holographic recording in thiol-ene based nanoparticle-polymer composites (NPCs) at a wavelength of 404 nm by using a highly coherent blue diode laser. We study the photopolymerization dynamics of two types of thiol-ene based NPCs doped with different blue-sensitive initiator/sensitizer systems (Darocur ® TPO and Irgacure ® 784/BzO2) at various doping concentrations. We also characterize a volume holographic grating recorded in these two types of thiol-ene based NPCs. Such material characterization includes the refractive index modulation, the material recording sensitivity and polymerization shrinkage. It is shown that Darocur R _ TPO provides larger refractive index modulation and higher material recording sensitivity than those with Irgacure ® 784/BzO2 but these two blue-sensitive initiator/sensitizer systems amount to meeting the requirements of the refractive index modulation and the material recording sensitivity for holographic data storage. However, it is found that shrinkage reduction of a volume grating recorded in these two types of thiol-ene based NPCs at 404 nm is not as effective as the same thiol-ene based NPC doped with Irgacure ® 784/BzO2 at 532 nm.

  3. Pore surface engineering in a zirconium metal–organic framework via thiol-ene reaction

    A porous olefin-functionalized Zr(IV)-based metal–organic framework, denoted as UiO-68-allyl, has been constructed. Our results clearly demonstrated that the surface of UiO-68-allyl could be decorated with organic molecule (ethanethiol) via thiol-ene reaction. More importantly, the crystallinity of the framework were maintained during the post-synthetic modification process. However, the microporosity of the framework is retained but the surface area decreased, due to the grafting of ethylthio groups into the pores. From our studies, we can conclude that the strategy of post-synthetic modification of UiO-68-allyl via thiol-ene reaction may be general. Furthermore, we may anchor other desired functional group onto the pore walls in Zr-MOFs via thiol-ene reaction, enabling more potential applications. - graphical abstract: In this manuscript, we reported the post-synthetic modification of an olefin-functionalized Zr(IV)-based metal–organic framework via thiol-ene reaction. - Highlights: • A porous olefin-functionalized Zr(IV)-based metal–organic framework has been constructed. • The surface of olefin-functionalized Zr-MOF could be decorated with organic molecules via thiol-ene reaction. • The crystallinity and permanent porosity of the framework were maintained during the post-synthetic modification process

  4. Binding of Fidarestat Stereoisomers with Aldose Reductase

    Dae-Sil Lee

    2006-11-01

    Full Text Available The stereospecificity in binding to aldose reductase (ALR2 of two fidarestat {6-fluoro-2',5'-dioxospiro[chroman-4,4'-imidazolidine]-2-carboxamide} stereoisomers [(2S,4Sand (2R,4S] has been investigated by means of molecular dynamics simulations using freeenergy integration techniques. The difference in the free energy of binding was found to be2.0 ± 1.7 kJ/mol in favour of the (2S,4S-form, in agreement with the experimentalinhibition data. The relative mobilities of the fidarestats complexed with ALR2 indicate alarger entropic penalty for hydrophobic binding of (2R,4S-fidarestat compared to (2S,4S-fidarestat, partially explaining its lower binding affinity. The two stereoisomers differmainly in the orientation of the carbamoyl moiety with respect to the active site and rotationof the bond joining the carbamoyl substituent to the ring. The detailed structural andenergetic insights obtained from out simulations allow for a better understanding of thefactors determining stereospecific inhibitor-ALR2 binding in the EPF charges model.

  5. Aldose reductase inhibitory compounds from Xanthium strumarium.

    Yoon, Ha Na; Lee, Min Young; Kim, Jin-Kyu; Suh, Hong-Won; Lim, Soon Sung

    2013-09-01

    As part of our ongoing search for natural sources of therapeutic and preventive agents for diabetic complications, we evaluated the inhibitory effects of components of the fruit of Xanthium strumarium (X. strumarium) on aldose reductase (AR) and galactitol formation in rat lenses with high levels of glucose. To identify the bioactive components of X. strumarium, 7 caffeoylquinic acids and 3 phenolic compounds were isolated and their chemical structures were elucidated on the basis of spectroscopic evidence and comparison with published data. The abilities of 10 X. strumarium-derived components to counteract diabetic complications were investigated by means of inhibitory assays with rat lens AR (rAR) and recombinant human AR (rhAR). From the 10 isolated compounds, methyl-3,5-di-O-caffeoylquinate showed the most potent inhibition, with IC₅₀ values of 0.30 and 0.67 μM for rAR and rhAR, respectively. In the kinetic analyses using Lineweaver-Burk plots of 1/velocity and 1/substrate, methyl-3,5-di-O-caffeoylquinate showed competitive inhibition of rhAR. Furthermore, methyl-3,5-di-O-caffeoylquinate inhibited galactitol formation in the rat lens and in erythrocytes incubated with a high concentration of glucose, indicating that this compound may be effective in preventing diabetic complications. PMID:23604720

  6. Aldose reductase mediates retinal microglia activation.

    Chang, Kun-Che; Shieh, Biehuoy; Petrash, J Mark

    2016-04-29

    Retinal microglia (RMG) are one of the major immune cells in charge of surveillance of inflammatory responses in the eye. In the absence of an inflammatory stimulus, RMG reside predominately in the ganglion layer and inner or outer plexiform layers. However, under stress RMG become activated and migrate into the inner nuclear layer (INL) or outer nuclear layer (ONL). Activated RMG in cell culture secrete pro-inflammatory cytokines in a manner sensitive to downregulation by aldose reductase inhibitors. In this study, we utilized CX3CR1(GFP) mice carrying AR mutant alleles to evaluate the role of AR on RMG activation and migration in vivo. When tested on an AR(WT) background, IP injection of LPS induced RMG activation and migration into the INL and ONL. However, this phenomenon was largely prevented by AR inhibitors or in AR null mice, or was exacerbated in transgenic mice that over-express AR. LPS-induced increases in ocular levels of TNF-α and CX3CL-1 in WT mice were substantially lower in AR null mice or were reduced by AR inhibitor treatment. These studies demonstrate that AR expression in RMG may contribute to the proinflammatory phenotypes common to various eye diseases such as uveitis and diabetic retinopathy. PMID:27033597

  7. Modulation of cell surface protein free thiols: a potential novel mechanism of action of the sesquiterpene lactone parthenolide.

    Jolanta Skalska

    Full Text Available BACKGROUND: There has been much interest in targeting intracellular redox pathways as a therapeutic approach for cancer. Given recent data to suggest that the redox status of extracellular protein thiol groups (i.e. exofacial thiols effects cell behavior, we hypothesized that redox active anti-cancer agents would modulate exofacial protein thiols. METHODOLOGY/PRINCIPAL FINDINGS: To test this hypothesis, we used the sesquiterpene lactone parthenolide, a known anti-cancer agent. Using flow cytometry, and western blotting to label free thiols with Alexa Fluor 633 C(5 maleimide dye and N-(biotinoyl-N-(iodoacetyl ethylendiamine (BIAM, respectively, we show that parthenolide decreases the level of free exofacial thiols on Granta mantle lymphoma cells. In addition, we used immuno-precipitation techniques to identify the central redox regulator thioredoxin, as one of the surface protein thiol targets modified by parthenolide. To examine the functional role of parthenolide induced surface protein thiol modification, we pretreated Granta cells with cell impermeable glutathione (GSH, prior to exposure to parthenolide, and showed that GSH pretreatment; (a inhibited the interaction of parthenolide with exofacial thiols; (b inhibited parthenolide mediated activation of JNK and inhibition of NFkappaB, two well established mechanisms of parthenolide activity and; (c blocked the cytotoxic activity of parthenolide. That GSH had no effect on the parthenolide induced generation of intracellular reactive oxygen species supports the fact that GSH had no effect on intracellular redox. Together these data support the likelihood that GSH inhibits the effect of parthenolide on JNK, NFkappaB and cell death through its direct inhibition of parthenolide's modulation of exofacial thiols. CONCLUSIONS/SIGNIFICANCE: Based on these data, we postulate that one component of parthenolide's anti-lymphoma activity derives from its ability to modify the redox state of critical

  8. Competitive reduction of perferrylmyoglobin radicals by protein thiols and plant phenols.

    Jongberg, Sisse; Lund, Marianne N; Skibsted, Leif H; Davies, Michael J

    2014-11-19

    Radical transfer from perferrylmyoglobin to other target species (myofibrillar proteins, MPI) and bovine serum albumin (BSA), extracts from green tea (GTE), maté (ME), and rosemary (RE), and three phenolic compounds, catechin, caffeic acid, and carnosic acid) was investigated by electron paramagnetic resonance (EPR) spectroscopy to determine the concentrations of plant extracts required to protect against protein oxidation. Blocking of MPI thiol groups by N-ethylmaleimide was found to reduce the rate of reaction of MPI with perferrylmyoglobin radicals, signifying the importance of protein thiols as radical scavengers. GTE had the highest phenolic content of the three extracts and was most effective as a radical scavenger. IC50 values indicated that the molar ratio between phenols in plant extract and MPI thiols needs to be >15 in order to obtain efficient protection against protein-to-protein radical transfer in meat. Caffeic acid was found most effective among the plant phenols. PMID:25343706

  9. Enhancement in the Glass Transition Temperature in Latent Thiol-Epoxy Click Cured Thermosets

    Dailyn Guzmán

    2015-04-01

    Full Text Available Tri and tetrafunctional thiol were used as curing agent for diglycidyl ether of bisphenol A (DGEBA catalyzed by a commercially available amine precursor, LC-80. Triglycidyl isocianurate (TGIC was added in different proportions to the mixture to increase rigidity and glass transition temperature (Tg. The cooperative effect of increasing functionality of thiol and the presence of TGIC in the formulation leads to an increased Tg without affecting thermal stability. The kinetics of the curing of mixtures was studied by calorimetry under isothermal and non-isothermal conditions. The latent characteristics of the formulations containing amine precursors were investigated by rheometry and calorimetry. The increase in the functionality of the thiol produces a slight decrease in the storage lifetime of the mixture. The materials obtained with tetrathiol as curing agent showed the highest values of Young’s modulus and Tg.

  10. Diamond surface functionalization with biomimicry – Amine surface tether and thiol moiety for electrochemical sensors

    Sund, James B., E-mail: jim@jamessund.com [Department of Electrical and Computer Engineering, Duke University, Durham, NC (United States); Causey, Corey P. [Departments of Chemistry and Biochemistry, Duke University, Durham, NC (United States); Wolter, Scott D. [Department of Physics, Elon University, Elon, NC 27244 (United States); Parker, Charles B., E-mail: charles.parker@duke.edu [Department of Electrical and Computer Engineering, Duke University, Durham, NC (United States); Stoner, Brian R. [Department of Electrical and Computer Engineering, Duke University, Durham, NC (United States); Research Triangle Institute (RTI) International, Research Triangle Park, NC (United States); Toone, Eric J. [Departments of Chemistry and Biochemistry, Duke University, Durham, NC (United States); Glass, Jeffrey T. [Department of Electrical and Computer Engineering, Duke University, Durham, NC (United States)

    2014-05-01

    Highlights: • Diamond surfaces were functionalized with organic molecules using a novel approach. • Used biomimicry to select a molecule to bind NO, similar to the human body. • Molecular orbital theory predicted the molecule-analyte oxidation behavior. • A thiol moiety was attached to an amine surface tether on the diamond surface. • XPS analysis verified each surface functionalization step. - Abstract: The surface of conducting diamond was functionalized with a terminal thiol group that is capable of binding and detecting nitrogen–oxygen species. The functionalization process employed multiple steps starting with doped diamond films grown by plasma enhanced chemical vapor deposition followed by hydrogen termination and photochemical attachment of a chemically protected amine alkene. The surface tether was deprotected to reveal the amine functionality, which enabled the tether to be extended with surface chemistry to add a terminal thiol moiety for electrochemical sensing applications. Each step of the process was validated using X-ray photoelectron spectroscopy analysis.

  11. UV-Induced Tetrazole-Thiol Reaction for Polymer Conjugation and Surface Functionalization.

    Feng, Wenqian; Li, Linxian; Yang, Chengwu; Welle, Alexander; Trapp, Oliver; Levkin, Pavel A

    2015-07-20

    A UV-induced 1,3-dipolar nucleophilic addition of tetrazoles to thiols is described. Under UV irradiation the reaction proceeds rapidly at room temperature, with high yields, without a catalyst, and in both polar protic and aprotic solvents, including water. This UV-induced tetrazole-thiol reaction was successfully applied for the synthesis of small molecules, protein modification, and rapid and facile polymer-polymer conjugation. The reaction has also been demonstrated for the formation of micropatterns by site-selective surface functionalization. Superhydrophobic-hydrophilic micropatterns were successfully created by sequential modifications of a tetrazole-modified porous polymer surface with hydrophobic and hydrophilic thiols. A biotin-functionalized surface could be fabricated in aqueous solutions under long-wavelength UV irradiation. PMID:26059870

  12. Thiol peptides induction in the seagrass Thalassia testudinum (Banks ex Koenig) in response to cadmium exposure

    Alvarez-Legorreta, Teresa [Departamento de Recursos del Mar, CINVESTAV-IPN, Unidad Merida, Apdo. Postal 73-Cordemex, Merida, Yucatan 97310 (Mexico); Mendoza-Cozatl, David; Moreno-Sanchez, Rafael [Departamento de Bioquimica, Instituto Nacional de Cardiologia, Mexico D.F. 14080 (Mexico); Gold-Bouchot, Gerardo [Departamento de Recursos del Mar, CINVESTAV-IPN, Unidad Merida, Apdo. Postal 73-Cordemex, Merida, Yucatan 97310 (Mexico)], E-mail: gold@mda.cinvestav.mx

    2008-01-20

    Trace metal accumulation and thiol compounds synthesis as induced by cadmium exposure was studied in the seagrass Thalassia testudinum. Shoots were exposed for 24, 48, 96 and 144 h to several CdCl{sub 2} concentrations (0, 30, 50 and 70 {mu}M). Levels of cadmium, cysteine, glutathione (GSH), {gamma}-glutamylcysteine ({gamma}-EC), and phytochelatin-like peptides were determined in green blades, live sheaths and root/rhizomes tissues. Metal accumulation was dependent on Cd concentration and type of tissue, with green blades showing the highest content followed by live sheaths and root/rhizomes. All tissues experienced an increase in thiol-containing compounds as a response to cadmium exposure. Live sheaths showed the highest levels of cysteine, GSH and {gamma}-EC. This is the first report of induction of thiol peptides, presumably phytochelatins, by a trace metal in a sea grass species.

  13. Isolation of mammalian cell variants with enhanced endogenous thiol content at low survival levels following irradiation

    Approximately half of a group of Chinese hamster V79 cell clones isolated from radiation survivors at low surviving fractions had significantly higher endogenous levels of non-protein and protein thiols than unirradiated cells. A similar group of cell lines cloned from unirradiated cells had thiol levels in the same range as the original unirradiated population. In some cases, clones isolated following irradiation are also more resistant to misonidazole toxicity and to radiation. This phenotype can persist through many cell generations for weeks or months of continuous growth; however, in many clones with altered phenotypes isolated following irradiation, reversion to the same phenotype as that of unirradiated populations has been observed. Induction of elevated thiol levels in tumours by radiotherapy could reduce both efficacy of the radiation itself and of radiation-modifying or chemotherapeutic drugs given in combination with radiation. (author)

  14. Diamond surface functionalization with biomimicry – Amine surface tether and thiol moiety for electrochemical sensors

    Highlights: • Diamond surfaces were functionalized with organic molecules using a novel approach. • Used biomimicry to select a molecule to bind NO, similar to the human body. • Molecular orbital theory predicted the molecule-analyte oxidation behavior. • A thiol moiety was attached to an amine surface tether on the diamond surface. • XPS analysis verified each surface functionalization step. - Abstract: The surface of conducting diamond was functionalized with a terminal thiol group that is capable of binding and detecting nitrogen–oxygen species. The functionalization process employed multiple steps starting with doped diamond films grown by plasma enhanced chemical vapor deposition followed by hydrogen termination and photochemical attachment of a chemically protected amine alkene. The surface tether was deprotected to reveal the amine functionality, which enabled the tether to be extended with surface chemistry to add a terminal thiol moiety for electrochemical sensing applications. Each step of the process was validated using X-ray photoelectron spectroscopy analysis

  15. Mutations in the RAM network confer resistance to the thiol oxidant 4,4'-dipyridyl disulfide

    López-Mirabal, H Reynaldo; Winther, Jakob R; Thorsen, Michael; Kielland-Brandt, Morten C

    2008-01-01

    Thiol oxidants are expected to have multiple effects in living cells. Hence, mutations giving resistance to such agents are likely to reveal important targets and/or mechanisms influencing the cellular capacity to withstand thiol oxidation. A screen for mutants resistant to the thiol......-specific oxidant dipyridyl disulfide (DPS) yielded tao3-516, which is impaired in the function of the RAM signaling network protein Tao3/Pag1p. We suggest that the DPS-resistance of the tao3-516 mutant might be due to deficient cell-cycle-regulated production of the chitinase Cts1p, which functions in post...... might relate to bypass for abnormal septum-associated protein sorting. The broad resistance toward oxidants (DPS, diamide and H(2)O(2)) of the Deltacts1 strain links cell wall function to the resistance to oxidative stress and suggests the existence of targets that are common for these oxidants....

  16. Thiol peptides induction in the seagrass Thalassia testudinum (Banks ex Koenig) in response to cadmium exposure

    Trace metal accumulation and thiol compounds synthesis as induced by cadmium exposure was studied in the seagrass Thalassia testudinum. Shoots were exposed for 24, 48, 96 and 144 h to several CdCl2 concentrations (0, 30, 50 and 70 μM). Levels of cadmium, cysteine, glutathione (GSH), γ-glutamylcysteine (γ-EC), and phytochelatin-like peptides were determined in green blades, live sheaths and root/rhizomes tissues. Metal accumulation was dependent on Cd concentration and type of tissue, with green blades showing the highest content followed by live sheaths and root/rhizomes. All tissues experienced an increase in thiol-containing compounds as a response to cadmium exposure. Live sheaths showed the highest levels of cysteine, GSH and γ-EC. This is the first report of induction of thiol peptides, presumably phytochelatins, by a trace metal in a sea grass species

  17. Characterization of volume holographic recording in photopolymerizable nanoparticle-(thiol-ene) polymer composites at 404 nm

    Kawana, Masaru; Takahashi, Jun-ichiro; Yasui, Satoru; Tomita, Yasuo

    2015-02-01

    We report on the photopolymerization dynamics and the volume holographic recording properties of a thiol-ene based nanoparticle-polymer composite (NPC) doped with a blue-sensitive photoinitiator, Darocur® TPO, by using a highly coherent blue diode laser operating at a wavelength of 404 nm. Our study indicates that volume gratings recorded in the NPC amount to meeting the material requirements of refractive index modulation and material recording sensitivity for holographic data storage media. It is also found that polymerization shrinkage of recorded NPC gratings is higher than that of the same thiol-ene based NPC with a green (523 nm)-sensitive photoinitiator, Irgacure® 784/BzO2. We attribute such a difference in shrinkage to the photopolymerization dynamics at these recording wavelengths. We show that this shrinkage increase at 404 nm can be mitigated to some extent by controlling the thiol-ene stoichiometry in the NPC.

  18. Transcripts of Anthocyanidin Reductase and Leucoanthocyanidin Reductase and Measurement of Catechin and Epicatechin in Tartary Buckwheat

    Yeon Bok Kim

    2014-01-01

    Full Text Available Anthocyanidin reductase (ANR and leucoanthocyanidin reductase (LAR play an important role in the monomeric units biosynthesis of proanthocyanidins (PAs such as catechin and epicatechin in several plants. The aim of this study was to clone ANR and LAR genes involved in PAs biosynthesis and examine the expression of these two genes in different organs under different growth conditions in two tartary buckwheat cultivars, Hokkai T8 and T10. Gene expression was carried out by quantitative real-time RT-PCR, and catechin and epicatechin content was analyzed by high performance liquid chromatography. The expression pattern of ANR and LAR did not match the accumulation pattern of PAs in different organs of two cultivars. Epicatechin content was the highest in the flowers of both cultivars and it was affected by light in only Hokkai T8 sprouts. ANR and LAR levels in tartary buckwheat might be regulated by different mechanisms for catechin and epicatechin biosynthesis under light and dark conditions.

  19. Equine 5α-reductase activity and expression in epididymis.

    Corbin, C J; Legacki, E L; Ball, B A; Scoggin, K E; Stanley, S D; Conley, A J

    2016-10-01

    The 5α-reductase enzymes play an important role during male sexual differentiation, and in pregnant females, especially equine species where maintenance relies on 5α-reduced progesterone, 5α-dihydroprogesterone (DHP). Epididymis expresses 5α-reductases but was not studied elaborately in horses. Epididymis from younger and older postpubertal stallions was divided into caput, corpus and cauda and examined for 5α-reductase activity and expression of type 1 and 2 isoforms by quantitative real-time polymerase chain reaction (qPCR). Metabolism of progesterone and testosterone to DHP and dihydrotestosterone (DHT), respectively, by epididymal microsomal protein was examined by thin-layer chromatography and verified by liquid chromatography tandem mass spectrometry (LC-MS/MS). Relative inhibitory potencies of finasteride and dutasteride toward equine 5α-reductase activity were investigated. Pregnenolone was investigated as an additional potential substrate for 5α-reductase, suggested previously from in vivo studies in mares but never directly examined. No regional gradient of 5α-reductase expression was observed by either enzyme activity or transcript analysis. Results of PCR experiments suggested that type 1 isoform predominates in equine epididymis. Primers for the type 2 isoform were unable to amplify product from any samples examined. Progesterone and testosterone were readily reduced to DHP and DHT, and activity was effectively inhibited by both inhibitors. Using epididymis as an enzyme source, no experimental evidence was obtained supporting the notion that pregnenolone could be directly metabolized by equine 5α-reductases as has been suggested by previous investigators speculating on alternative metabolic pathways leading to DHP synthesis in placenta during equine pregnancies. PMID:27466384

  20. Stretching of BDT-gold molecular junctions: Thiol or thiolate termination?

    Souza, Amaury De Melo

    2014-01-01

    It is often assumed that the hydrogen atoms in the thiol groups of a benzene-1,4-dithiol dissociate when Au-benzene-1,4-dithiol-Au junctions are formed. We demonstrate, by stability and transport property calculations, that this assumption cannot be made. We show that the dissociative adsorption of methanethiol and benzene-1,4-dithiol molecules on a flat Au(111) surface is energetically unfavorable and that the activation barrier for this reaction is as high as 1 eV. For the molecule in the junction, our results show, for all electrode geometries studied, that the thiol junctions are energetically more stable than their thiolate counterparts. Due to the fact that density functional theory (DFT) within the local density approximation (LDA) underestimates the energy difference between the lowest unoccupied molecular orbital and the highest occupied molecular orbital by several electron-volts, and that it does not capture the renormalization of the energy levels due to the image charge effect, the conductance of the Au-benzene-1,4-dithiol-Au junctions is overestimated. After taking into account corrections due to image charge effects by means of constrained-DFT calculations and electrostatic classical models, we apply a scissor operator to correct the DFT energy level positions, and calculate the transport properties of the thiol and thiolate molecular junctions as a function of the electrode separation. For the thiol junctions, we show that the conductance decreases as the electrode separation increases, whereas the opposite trend is found for the thiolate junctions. Both behaviors have been observed in experiments, therefore pointing to the possible coexistence of both thiol and thiolate junctions. Moreover, the corrected conductance values, for both thiol and thiolate, are up to two orders of magnitude smaller than those calculated with DFT-LDA. This brings the theoretical results in quantitatively good agreement with experimental data.

  1. Radiosensitization by oxygen and radioprotection by Thiols: Experimental evaluation of a theoretical model

    The general purpose of the investigations presented in this thesis was a further elucidation of the mechanism by which oxygen sensitizes cells and tissues to radiation. Recently, the 'X model' was put forward as an explanation for the oxygen effect. The model postulated that oxygen and cellular thiols compete for radiation induced target radicals, oxygen fixing and thiols repairing the radiation damage, the latter, however, with the reservation that only a certain proportion of the damage is reparable. The study summarized in the thesis was performed with the aim of testing the validity of the model by a comparative analysis of the theoretical predictions with experimental observations. Human fibroblasts and Chinese hamster cells in culture were used as experimental cell material. The thiol level of the cells varied either due to a genetic damage, or to treatment with buthionine sulphoximine (BSO) which decreased the thiol content, or to treatment with dithiothreitol or N-acetylcysteine which increased it. Exposures to radiation were made in severe hypoxia or in the presence of oxygen in varying concentrations. The yield of DNA breaks, expressing the effect of the initial radical reactions and, in a few cases, clonogenic survival, expressing also the effect of additional superimposed enzymatic repair processes, were chosen as measure of the radiation response. Four different predictions of the X model were tested experimentally. In all cases a good general agreement between the pattern of the radiation response, predicted by the X model, and the experimental observations was found. In contrast, major differences were noted between the experimental data and the prediction of an alternative model in which, as opposed to the X model, radiation induced radical damage is assumed to be reparable by thiols in full. The validity of the X model, indicated by these investigations suggests that, in general two types of damage are produced by radiation, only one of which can

  2. Importance of thiols in the repair mechanisms of DNA containing AP (apurinic or apyrimidinic) sites.

    Bailly, V; Verly, W G

    1988-01-01

    Addition of thiol compounds containing an anionic group to the 3'-terminal unsaturated sugar of the 5' fragment obtained from an oligonucleotide containing an AP site cleaved by beta-elimination, can be followed by gel electrophoresis. The technique enables to distinguish between two mechanisms of cleavage of the C3'-O-P bond 3' to an AP site: hydrolysis or beta-elimination. Addition of thiols to the double-bond of the 3'-terminal sugar resulting from beta-elimination prevents a subsequent de...

  3. Gold Nanoparticles Protected with Thiol-Derivatized Amphiphilic Poly( -caprolactone)-b-poly(acrylic acid)

    Javakhishvili, Irakli; Hvilsted, Søren

    2008-01-01

    Amphiphilic poly(c-caprolactone)-b-poly(acrylic acid) (HS-PCL-b-PAA) bearing thiol functionality at the PCL terminal has been synthesized by a combination of ring-opening polymerization (ROP) of c-caprolactone (c-CL), esterification of hydroxy chain end with protected mercaptoacetic acid, subsequ......Amphiphilic poly(c-caprolactone)-b-poly(acrylic acid) (HS-PCL-b-PAA) bearing thiol functionality at the PCL terminal has been synthesized by a combination of ring-opening polymerization (ROP) of c-caprolactone (c-CL), esterification of hydroxy chain end with protected mercaptoacetic acid...

  4. Straightforward synthesis of deuterated precursors to demonstrate the biogenesis of aromatic thiols in wine.

    Roland, Aurélie; Schneider, Rémi; Razungles, Alain; Le Guernevé, Christine; Cavelier, Florine

    2010-10-13

    Straightforward synthesis of labeled S-3-(hexan-1-ol)-glutathione and S-4-(4-methylpentan-2-one)-glutathione has been developed through a conjugate addition optimization study. Sauvignon blanc fermentation experiments with the [(2)H(10)] S-4-(4-methylpentan-2-one)-glutathione used as a tracer released the corresponding deuterated thiol, thus proving the direct relationship with the 4-mercapto-4-methylpentan-2-one under enological conditions. The conversion yield of such transformation was estimated to be close to 0.3%, opening an avenue for additional study on varietal thiol biogenesis. PMID:20825191

  5. NMR study of 2-amino-cyclo-hexane thiols N-substituted

    Considering that thiols and mainly β-aminoacids are been successful in research aiming the discovering of new substances with protective actions against the radiation effects on living being, 2-amine-cyclo-hexane thiols N-alkyl-substituted were synthesized and submitted to to biological tests. The reactions were planned to favor the SN 2 mechanism, in order to obtain the respective trans isomers from each β-aminothiol (nonpolar medium; solvent: benzene or hexane). This work presents the results obtained in the NMR study of four by products of this class

  6. Kinetics and Thioredoxin Specificity of Thiol Modulation of the Chloroplast H+-ATPase

    Schwarz, Oliver; Schürmann, Peter; Strotmann, Heinrich

    2007-01-01

    The kinetics of thiol modulation of the chloroplast H+-ATPase (CF0CF1) in membrana were analyzed by employing thioredoxins that were kept reduced by 0.1 mM dithiothreitol. The kinetics of thiol modulation depend on the extent of the proton gradient. The process is an exponential function of the thioredoxin concentration and reaction time and can be described by an irreversible second order reaction. The results indicate that the formation of the complex between thioredoxin and CF0CF1 is slow ...

  7. Diamond surface functionalization with biomimicry - Amine surface tether and thiol moiety for electrochemical sensors

    Sund, James B.; Causey, Corey P.; Wolter, Scott D.; Parker, Charles B.; Stoner, Brian R.; Toone, Eric J.; Glass, Jeffrey T.

    2014-05-01

    The surface of conducting diamond was functionalized with a terminal thiol group that is capable of binding and detecting nitrogen-oxygen species. The functionalization process employed multiple steps starting with doped diamond films grown by plasma enhanced chemical vapor deposition followed by hydrogen termination and photochemical attachment of a chemically protected amine alkene. The surface tether was deprotected to reveal the amine functionality, which enabled the tether to be extended with surface chemistry to add a terminal thiol moiety for electrochemical sensing applications. Each step of the process was validated using X-ray photoelectron spectroscopy analysis.

  8. Mechanism of misonidazole linked cytotoxicity and altered radiation response: role of cellular thiols

    The effectiveness of misonidazole as a hypoxic radiosensitizer of mammalian cells is increased by prolonged exposure of hypoxic cells to the drug. It was found that drug intermediates might react with endogenous non-protein thiols (NPSH). These thiols function to protect the cell against deleterious intermediates that could otherwise attach and modify critical macromolecules such as DNA, RNA and protein. This paper presents studies on the effects of misonidazole, as well as newly developed hypoxic cell radiosensitizers, in an attempt to (1) identify the alterations in the NPSH, and (2) elucidate the role that the changes in NPSH play in cytotoxic and radiosensitizing effects of nitro compounds

  9. Thiol-Disulfide Interchange in the Tocinoic Acid/Glutathione System During Freezing and Drying

    THING, METTE; Zhang, Jun; LAURENCE, JENNIFER; Topp, Elizabeth M.

    2010-01-01

    Thiol-disulfide interchange (“disulfide scrambling”) is a common mechanism of covalent aggregation for protein drugs. Using tocinoic acid (cyclo-S-Cys-Tyr-Ile-Gln-Asn-Cys-(S); TA(ox)) and glutathione (γGlu-Cys-Gly; GSH), our previous work demonstrated that thiol/disulfide interchange is affected by lyophilization in a manner consistent with irreversible and regioselective loss of TA(ox) (Zhang et al., 2009, J Pharm Sci 98/9: 3312–3318). Here, we explore the contributions of stages of the lyop...

  10. Metal triflate-mediated coupling of allylgermanes with thiols: a facile route to thiogermanes.

    Kuciński, K; Pawluć, P; Hreczycho, G

    2015-06-28

    A novel coupling reaction of thiols with (2-methylallyl)germanes catalyzed by metal triflates has been developed. This reaction provides a direct and efficient method to afford thiogermanes and opens a valuable and general synthetic route for the Ge-S cross-coupling with the elimination of isobutylene as a single by-product. Scandium(iii) triflate demonstrates the highest catalytic activity among the tested triflates. All reactions were carried out under extremely mild conditions to give thiogermanes in excellent yields. This Ge-S coupling reaction shows high generality for the variety of thiols. PMID:26006777

  11. Poly(ethylene glycol)-based thiol-ene hydrogel coatings: curing chemistry, aqueous stability, and potential marine antifouling applications

    Lundberg, P.; Bruin, A.; Klijnstra, J.W.; Nyström, A.M.; Johansson, M.; Malkoch, M.; Hult, A.

    2010-01-01

    Photocured thiol-ene hydrogel coatings based on poly(ethylene glycol) (PEG) were investigated for marine antifouling purposes. By varying the PEG length, vinylic end-group, and thiol cross-linker, a library of hydrogel coatings with different structural composition was efficiently accomplished, with

  12. Rapid and simple preparation of thiol-ene emulsion-templated monoliths and their application as enzymatic microreactors

    Lafleur, Josiane P; Senkbeil, Silja; Novotny, Jakub;

    2015-01-01

    electron microscopy showed that the methanol-based emulsion templating process resulted in a network of highly interconnected and regular thiol-ene beads anchored solidly inside thiol-ene microchannels. Surface area measurements indicate that the monoliths are macroporous, with no or little micro- or...

  13. Volume holographic recording in photopolymerizable nanocomposite materials based on radical-mediated thiol-yne step-growth polymerizations

    Mitsube, Ken; Nishimura, Yuki; Takayama, Shingo; Nagaya, Kohta; Tomita, Yasuo

    2013-05-01

    We propose the use of radical-mediated thiol-yne step-growth photopolymerizations for volume holographic recording in NPC films to overcome the drawback of low crosslinking densities but retain the advantage of low shrinkage in the thiol-ene photopolymerizations. The thiol-yne photopolymerization mechanism is different from the thiol-ene photopolymeriztions in the sense that each alkyne functional group can react consecutively with two thiol functional groups. We show that thiol-yne based NPC films dispersed with silica nanoparticles give the saturated refractive index change as large as 0.008 and the material recording sensitivity as high as 2005 cm/J at a wavelength of 532 nm, larger than the minimum acceptable values of 0.005 and 500 cm/J, respectively, for holographic data storage. We also show that the shrinkage of a recorded hologram can be as low as that of thiol-ene based NPC films and that the thermal stability is improved better. In addition, we demonstrate digital data page recording in thiol-yne based NPC films, showing a low symbol error rate and a high signal-to-noise ratio to be 2.8×10-4 and 8, respectively.

  14. Induction of Thioredoxin Reductase 1 by Korean Red Ginseng Water Extract Regulates Cytoprotective Effects on Human Endothelial Cells

    Hye Rim Park

    2015-01-01

    Full Text Available Korean Red Ginseng is a popular herbal medicine and is widely used in many food products. KRG has biological benefits related to vascular diseases including diabetes, hypertension, atherosclerosis, and other cardiac diseases and KRG has antioxidant and anti-hyperlipidemic actions. KRG decreases the level of oxidative stress and suppresses proinflammatory cytokines and cell adhesion molecules, thus protecting endothelial dysfunction. Mammalian Thioredoxin reductase 1 is an NADPH-dependent selenoprotein, essential for antioxidant defense and DNA synthesis and repair, that regulates the redox system by modulating redox-sensitive transcription factors and thiol-containing proteins. Here, we show that KRG water extract increases the expression of TrxR1 in human umbilical vein endothelial cells via the p38 and PKC-δ signaling pathways. The induction of TrxR1 expression by KRG was confirmed by Western blot analysis and reverse transcription polymerase chain reaction. However, the increase in TrxR1 expression was abolished by specific silencing of the p38 and PKC-δ genes. In addition, we demonstrated that auranofin, a TrxR1 inhibitor, weakens the protective effect of KRG against H2O2-induced cell death as measured by the terminal transferase dUTP nick end labeling assay. These results suggest that KRG may have protective effects in vascular diseases by upregulating TrxR1 in endothelial cells, thereby inhibiting the generation of reactive oxygen species and cell death.

  15. Ferredoxin-thioredoxin reductase: a catalytically active dithiol group links photoreduced ferredoxin to thioredoxin functional in photosynthetic enzyme regulation

    Droux, M.; Miginiac-Maslow, M.; Jacquot, J.P.; Gadal, P.; Crawford, N.A.; Kosower, N.S.; Buchanan, B.B.

    1987-07-01

    The mechanism by which the ferredoxin-thioredoxin system activates the target enzyme, NADP-malate dehydrogenase, was investigated by analyzing the sulfhydryl status of individual protein components with (/sup 14/C)iodoacetate and monobromobimane. The data indicate that ferredoxin-thioredoxin reductase (FTR)--an iron-sulfur enzyme present in oxygenic photosynthetic organisms--is the first member of a thiol chain that links light to enzyme regulation. FTR possesses a catalytically active dithiol group localized on the 13 kDa (similar) subunit, that occurs in all species investigated and accepts reducing equivalents from photoreduced ferredoxin and transfers them stoichiometrically to the disulfide form of thioredoxin m. The reduced thioredoxin m, in turn, reduces NADP-malate dehydrogenase, thereby converting it from an inactive (S-S) to an active (SH) form. The means by which FTR is able to combine electrons (from photoreduced ferredoxin) with protons (from the medium) to reduce its active disulfide group remains to be determined.

  16. The role of biliverdin reductase in colorectal cancer

    In recent years, the effects of biliverdin and bilirubin have been studied extensively, and an inhibitory effect of bile pigments in cancer progression has been proposed. In this study we focused on the effects of biliverdin reductase, the enzyme that converts biliverdin to bilirubin, in colorectal cancer. For in vitro experiments we used a human colorectal carcinoma cell line and transfected it with an expression construct of shRNA specific for biliverdin reductase, to create cells with stable knock-down of enzyme expression. Cell proliferation was analyzed using the CASY model TT cell counting device. Western blot protein analysis was performed to study intracellular signaling cascades. Samples of human colorectal cancer were analyzed using immunohistochemistry. We were able to confirm the antiproliferative effects of bile pigments on cancer cells in vitro. However, this effect was attenuated in biliverdin reductase knock down cells. ERK and Akt activation seen under biliverdin and bilirubin treatment was also reduced in biliverdin reductase deficient cells. Immunohistochemical analysis of tumor samples from patients with colorectal cancer showed elevated biliverdin reductase levels. High enzyme expression was associated with lower overall and disease free patient survival. We conclude that BVR is required for bile pigment mediated effects regarding cancer cell proliferation and modulation of intracellular signaling cascades. The role of BVR overexpression in vivo and its exact influence on cancer progression and patient survival need to be further investigated. (author)

  17. Formation of Underbrushes on thiolated Poly (ethylene glycol) PEG monolayers by Oligoethylene glycol (OEG) terminated Alkane Thiols on Gold

    Lokanathan, Arcot R.

    2011-01-01

    Adding underbrushes of oligoethylene glycol (OEG) to monolayers of long chain PEG molecules on a surface is one of the strategies [1] in designing a suitable platform for antifouling purpose, where it is possible to have high graft density and molecular conformational freedom[4] simultaneously......, there by maximal retention of activity of covalently immobilised antifouling enzyme [2] on PEG surfaces along with resistance to protein adsorption[3]. Here we present some our studies on the addition of OEG thiol molecules over a self assembled monolayer of PEG thiol on gold. The kinetics of addition...... of OEG thiol to monolayers of PEG thiol was followed using X- ray photoelectron spectroscopy (XPS), which indicated the time point of maximum graft density and beyond this time point there was predominant desorption of OEG thiol as indicated by the C/O ratio. The initial increase in graft density was...

  18. Intramolecular electron transfer in Pseudomonas aeruginosa cd(1) nitrite reductase

    Farver, Ole; Brunori, Maurizio; Cutruzzolà, Francesca;

    2009-01-01

    The cd(1) nitrite reductases, which catalyze the reduction of nitrite to nitric oxide, are homodimers of 60 kDa subunits, each containing one heme-c and one heme-d(1). Heme-c is the electron entry site, whereas heme-d(1) constitutes the catalytic center. The 3D structure of Pseudomonas aeruginosa...... nitrite reductase has been determined in both fully oxidized and reduced states. Intramolecular electron transfer (ET), between c and d(1) hemes is an essential step in the catalytic cycle. In earlier studies of the Pseudomonas stutzeri enzyme, we observed that a marked negative cooperativity is...... controlling this internal ET step. In this study we have investigated the internal ET in the wild-type and His369Ala mutant of P. aeruginosa nitrite reductases and have observed similar cooperativity to that of the Pseudomonas stutzeri enzyme. Heme-c was initially reduced, in an essentially diffusion...

  19. Expression and site-directed mutagenesis of human dihydrofolate reductase

    A procaryotic high-level expression vector for human dihydrofolate reductase has been constructed and the protein characterized as a first step toward structure-function studies of this enzyme. A vector bearing the tac promoter, four synthetic oligodeoxynucleotides, and a restriction fragment from the dihydrofolate reductase cDNA were ligated in a manner which optimized the transcriptional and translational frequency of the enzyme mRNA. The reductase, comprising ca. 17% of the total soluble protein in the host bacteria, was purified to apparent homogeneity as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and characterized by amino acid composition, partial amino acid sequence, and steady-sate kinetic analysis. This expression vector has been used as a template for double-stranded plasmid DNA site-specific mutagenesis. Functional studies on a Cys-6 → Ser-6 mutant enzyme support the contention that Cys-6 is obligatory for organomercurial activation of human dihydrofolate reductase. The Ser-6 mutant enzyme was not activated to any extent following a 24-h incubation with p-(hydroxymercuri)benzoate and nicotinamide adenine dinucleotide phosphate (reduced) (NADPH), whereas the k/sub cat/ for Cys-6 reductase increased 2-fold under identical conditions. The specific activities of the Cys-6 and Ser-6 enzymes were virtually identical as determined by methotrexate titration as were the K/sub m/ values for both dihydrofolate and NADPH. The Ser-6 mutant showed a decreased temperature stability and was more sensitive to inactivation by α-chymotrypsin when compared to the wild-type enzyme. These results suggest that the Ser-6 mutant reductase is conformationally altered relative to the Cys-6 native enzyme

  20. Aldo keto reductases 1B in endocrinology and metabolism

    AntoineMartinez

    2012-08-01

    Full Text Available The aldose reductase (human AKR1B1/mouse Akr1b3 has been the focus of many research because of its role in diabetic complications. The starting point of these alterations is the massive entry of glucose in polyol pathway where it is converted into sorbitol by this enzyme. However, the issue of aldose reductase function in non-diabetic condition remains unresolved. Aldose reductase-like enzymes (AKR1B10, Akr1b7 and Akr1b8 are highly related isoforms often co-expressed with bona fide aldose reductase, making functional analysis of one or the other isoform a challenging task. AKR1B/Akr1b members share at least 65% protein identity and the general ability to reduce many redundant substrates such as aldehydes provided from lipid peroxidation, steroids and their by-products and xenobiotics in vitro. Based on these properties, AKR1B/Akr1b are generally considered as detoxifying enzymes. Considering that divergences should be more informative than similarities to help understanding their physiological functions, we chose to review specific hallmarks of each human/mouse isoforms by focusing on tissue distribution and specific mechanisms of gene regulation. Indeed, although the aldose reductase shows ubiquitous expression, aldose reductase-like proteins exhibit tissue-specific patterns of expression. We focused on 3 organs where certain isoforms are enriched, the adrenal gland, enterohepatic and adipose tissues and tried to connect recent enzymatic and regulation data with endocrine and metabolic functions of these organs. We presented recent mouse models showing unsuspected physiological functions in the regulation of glucido-lipidic metabolism and adipose tissue homeostasis. Beyond the widely accepted idea that AKR1B/Akr1b are detoxification enzymes, these recent reports provide growing evidences that they are able to modify or generate signal molecules. This conceptually shifts this class of enzymes from unenviable status of scavenger to upper class of

  1. Nanoparticles under the light: click functionalization by photochemical thiol-yne reaction, towards double click functionalization.

    Demay-Drouhard, Paul; Nehlig, Emilie; Hardouin, Julie; Motte, Laurence; Guénin, Erwann

    2013-06-24

    A light click away: The first application of the thiol-yne reaction to nanoparticle functionalization is described (see figure). This metal-free click chemistry approach is compatible with the addition of various molecules at the surface and can be combined with CuAAC methodology to perform chemoselective double functionalization. PMID:23744751

  2. Mercury and non-protein thiol compounds in the seagrass Posidonia oceanica.

    Ferrat, Lila; Gnassia-Barelli, Mauricette; Pergent-Martini, Christine; Roméo, Michèle

    2003-01-01

    Mercury concentrations, non-protein thiol levels and the enzyme activities of glutathione-S-transferase (GST) were measured in the blades and sheaths of the marine phanerogam Posidonia oceanica. The seagrass was collected in January and June and at three sites: the Bay of Rosignano (Italy) known for its mercury contamination, the north of the Lérins islands (Bay of Cannes, France), the Bay of Tonnara (Corsica, France). The two latter sites are considered as free of any known industrial inputs. Mercury concentrations and GST activities in both tissues were always higher in samples from Rosignano, particularly in June. Non-protein thiol levels were significantly higher in the blades than in the sheaths of P. oceanica from Tonnara and Lérins. In contrast, at Rosignano, the sheaths presented a significantly higher non-protein thiol concentration than the blades, particularly in June. Levels in the sheaths appeared to increase with the degree of pollution. Western Blot performed on sheaths of P. oceanica collected in June at Rosignano and Lérins revealed a characteristic band of GSTs at 31 kDa, proving the presence of the GST enzyme in this tissue. Mercury seemed to exert an influence upon non-protein thiol metabolism, including GST induction, in P. oceanica collected from the NW Mediterranean. PMID:12524027

  3. Quantitative interpretation of the transition voltages in gold-poly(phenylene) thiol-gold molecular junctions

    Wu, Kunlin

    2013-01-01

    The transition voltage of three different asymmetric Au/poly(phenylene) thiol/Au molecular junctions in which the central molecule is either benzene thiol, biphenyl thiol, or terphenyl thiol is investigated by first-principles quantum transport simulations. For all the junctions, the calculated transition voltage at positive polarity is in quantitative agreement with the experimental values and shows weak dependence on alterations of the Au-phenyl contact. When compared to the strong coupling at the Au-S contact, which dominates the alignment of various molecular orbitals with respect to the electrode Fermi level, the coupling at the Au-phenyl contact produces only a weak perturbation. Therefore, variations of the Au-phenyl contact can only have a minor influence on the transition voltage. These findings not only provide an explanation to the uniformity in the transition voltages found for π-conjugated molecules measured with different experimental methods, but also demonstrate the advantage of transition voltage spectroscopy as a tool for determining the positions of molecular levels in molecular devices. © 2013 AIP Publishing LLC.

  4. Endothelium-dependent vasorelaxation in inhibited by in vivo depletion of vascular thiol levels

    Laursen, J B; Boesgaard, S; Trautner, S; Rubin, I; Poulsen, H E; Aldershvile, J

    2001-01-01

    ) affected by BSO (BSO: 4415 +/- 123, control: 4105 +/- 455 counts/mg; n = 6) in rat aorta. It is concluded that in vivo thiol depletion results in endothelial dysfunction and a reduced receptor-mediated vascular relaxation. This effect is caused by reduced endothelial NO formation....

  5. Gold Nanoparticles Protected with Thiol-Derivatized Amphiphilic Poly(epsilon-caprolactone)-b-poly(acrylic acid)

    Javakhishvili, Irakli; Hvilsted, Søren

    2009-01-01

    Amphiphilic poly(epsilon-caprolactone)-b-poly(acrylic acid) (HS-PCL-b-PAA) with a thiol functionality in the PCL terminal has been prepared in a novel synthetic cascade. Initially, living anionic ring-opening polymerization (ROP) of epsilon-caprolactone (epsilon-CL) employing the difunctional...

  6. Influence cadmium ions on the synthesis of thiol compounds for flax

    Olga Krystofova; Václav Diopan; Jiří Baloun; Vojtech Adam; Rene Kizek

    2010-01-01

    Evaluation of the effectiveness of phytoremediation technologies isvery difficult. One way to quickly and inexpensively identifyphytoremediation potential of plants is found easily detectablemarker. In our study, we examined the content of thiol compoundsin plants, of Flax effects of various concentrations of cadmium ions.

  7. Passivation of copper surfaces for selective-area ALD using a thiol self-assembled monolayer

    Färm, Elina; Vehkamäki, Marko; Ritala, Mikko; Leskelä, Markku

    2012-07-01

    Self-assembled monolayers (SAMs) of 1-dodecanethiol (CH3(CH2)11SH) were prepared from the vapor phase and used as a passivation layer for selective-area ALD. Thiol SAMs have commonly been prepared by immersing the substrates into a solution containing alkyl thiols. Formation of SAMs from the vapor phase, however, has advantages compared to liquid phase preparation. Passivation of surface can be done as a part of the ALD process forming a SAM first and then continuing with the common ALD process. SAMs can also be applied to three-dimensional structures relying on chemical selectivity of the thiol SAM formation. For example in the copper damascene process the thiol SAMs should form only on the copper surface but not on the insulators. In this study, the SAMs were prepared by placing the substrate and the alkylthiol to the reaction chamber and heating the system to the temperature of 73 °C. Preparation time varied from 0.5 to 24 h. Passivation properties of SAMs were tested with ALD iridium and polyimide processes. Iridium was deposited at 250  ° C for 500 cycles and polyimide at 160  ° C for 20 cycles.

  8. Inhibition of a biological sulfide oxidation under haloalkaline conditions by thiols and diorgano polysulfanes

    Roman, Pawel; Lipińska, Joanna; Bijmans, Martijn F.M.; Sorokin, Dimitry Y.; Keesman, Karel J.; Janssen, Albert J.H.

    2016-01-01

    A novel approach has been developed for the simultaneous description of reaction kinetics to describe the formation of polysulfide and sulfate anions from the biological oxidation of hydrogen sulfide (H2S) using a quick, sulfide-dependent respiration test. Next to H2S, thiol

  9. Biotechnological removal of H2S and thiols from sour gas streams under haloalkaline conditions

    Roman, P.

    2016-01-01

    Biotechnological removal of H2S and thiols from sour gas streams under haloalkaline conditions Paweł Roman Abstract Biological removal of H2S from sour gas streams became popular in recent years because of high process efficiency and low operational costs. To expand

  10. GC/MS DETERMINATION OF 1-P-MENTHEN-8-THIOL IN GRAPEFRUIT JUICE

    P-menthene-8-thiol is a very potent aroma compound found in grapefruit juice. It currently is not measured commercially because it is present in only trace quantities and due to the fact that it requires an expensive specialized detector. However, in this report, it is shown that utilizing a stand...

  11. Development of ionic gels using thiol-based monomers in ionic liquid

    Ahmed, Kumkum; Naga, Naofumi; Kawakami, Masaru; Furukawa, Hidemitsu

    2016-04-01

    Ionic gels (IGs) using ionic liquids (ILs) can propose diverse applications in the field of optics, sensors and separation have opened wide prospects in materials science. ILs have attracted remarkable interest for gel polymer electrolytes and batteries based on their useful properties such as non-volatility, non-flammability, a wide electrochemical window, high thermal stability and a high ionic conductivity. The formation of gel in IL media makes it possible to immobilize ILs within organic or inorganic matrices and to take advantage of their unique properties in the solid state, thus eliminating some shortcomings related to shaping and risk of leakage. In this work for the first time we used multifunctional thiol monomers having uniform structure and good compatibility with the IL of our interest. Therefore we focused on developing thiol monomer-based IGs using multifunctional thiol monomers and acrylate crosslinkers utilizing thiol-ene reaction between monomer and crosslinking molecules in an IL medium and characterize their physico-chemical properties like thermal, conductive, mechanical properties etc.. This work has been focused mainly to improve the mechanical strength of IGs and make prospects of IGs in tribology and lubricants.

  12. Association of bilirubin and protein thiols in relation to copper and ceruloplasmin in hyperbilirubinemic patients

    Mungli Prakash; Jeevan K Shetty; Roshan D'Souza; Suhasa Upadhya; Vijay Kumar

    2009-01-01

    Objective:Bilirubin is a double edged sword in biological system,acting as a toxic molecule and cytoprotecrant.Unconjugated bilirubin is proved to show antioxidant activity in vitro and in vivo.In the current work we tried to know the relationship between both conjugated and unconjugated bilirubin with copper and protein thiols in patients with hyperbilirnbinemia.Methods:Study was conducted on 56 hyperbilirubinemic cases and 56 healthy controls.Serum copper,ceruloplasmin,protein thiols,total bilirubin,conjugated and unconjugated bilirubin,unconjugated bilimbin/albumin ratio,total protein,albumin,AST,ALT and ALP were estimated.Results:There was significant increase in serum copper,total bilirubin,conjugated and unconjugated bilimbin.uriconjugated bilirubin/albumin ratio,AST,ALT,and ALP,and decrease in serum ceruloplasmin,protein thiols,total protein,and albumin in hyperbilimbinemic cases when compared to healthy controls.Conjugated bilimbin correlated positively with liver enzymes AST and ALP,and negatively with protein thials,total protein and albumin.Unconjugated bilirubin correlated positively with ALT.Protein thiols correlated negatively with copper and positively with ceruloplasmin,and also correlated negativelv with liver enzymes like AST,ALT and ALP,and positively with total protein and albumin.Conclusion:Combination of elevated levels of trace elements like copper and availability of reducing agent like bilimbin may prove deleterious by generating free radicals.

  13. Synthesis of Novel Fluorescence reagents and Their Application in Determination of Thiol Compounds

    LIANG Shu-Cai; WANG Hong; ZHANG Hua-Shan; HU Xian-Ming

    2003-01-01

    @@ The identification and determination of thiol compounds is essential both for the clinical diagnosis and the con trol of diseases because alteration of their concentrations in biological systems are somehow responsive for some diseases such as myocardial infarction, diabetes, peripheral vascular and so on. [1

  14. Thiols as biomarkers of heavy metal tolerance in the aquatic macrophytes of Middle Urals, Russia.

    Borisova, Galina; Chukina, Nadezda; Maleva, Maria; Kumar, Adarsh; Prasad, M N V

    2016-10-01

    Aquatic macrophytes, viz. Sagittaria sagittifolia L., Lemna gibba L., Elodea canadensis Michx., Batrachium trichophyllum (Chaix.) Bosch., Ceratophyllum demersum L. and Potamogeton sp. (P. perfoliatus L., P. alpinus Balb., P. crispus L., P. berchtoldii Fieber, P. friesii Rupr., P. pectinatus L.) were collected from 11 sites for determining their metal accumulation and thiols content. Cu(2+), Ni(2+), Mn(2+), Zn(2+), and Fe(3+) exceeded maximum permissible concentrations in chosen sites. Significant transfer of metals from water to leaves is observed in the order of Ni(2+) < Cu(2+) < Zn(2+) < Fe(3+) < Mn(2+). The maximum variation of bioconcentration factor was noticed for manganese. The accumulation of heavy metals in leaves was correlated with non-protein and protein thiols, confirming their important role in metal tolerance. The largest contribution was provided by Cu(2+) (on the average r = 0.88, p < 0.05), which obviously can be explained as an important role of these ions in thiols synthesis. Increased synthesis of thiols in the leaves allows the usage of SH-containing compounds as biomarkers of metal tolerance. Considering accumulation of metals and tolerance, B. trichophyllum, C. demersum and L. gibba are the most suitable species for phytoremediation of highly multimetal contamination, while E. canadensis and some species of Potamageton are suitable for moderately metal-polluted sites. PMID:27167595

  15. Influence cadmium ions on the synthesis of thiol compounds for flax

    Olga Krystofova

    2010-12-01

    Full Text Available Evaluation of the effectiveness of phytoremediation technologies isvery difficult. One way to quickly and inexpensively identifyphytoremediation potential of plants is found easily detectablemarker. In our study, we examined the content of thiol compoundsin plants, of Flax effects of various concentrations of cadmium ions.

  16. Thimerosal Exposure and the Role of Sulfation Chemistry and Thiol Availability in Autism

    Mark R. Geier

    2013-08-01

    Full Text Available Autism spectrum disorder (ASD is a neurological disorder in which a significant number of the children experience a developmental regression characterized by a loss of previously acquired skills and abilities. Typically reported are losses of verbal, nonverbal, and social abilities. Several recent studies suggest that children diagnosed with an ASD have abnormal sulfation chemistry, limited thiol availability, and decreased glutathione (GSH reserve capacity, resulting in a compromised oxidation/reduction (redox and detoxification capacity. Research indicates that the availability of thiols, particularly GSH, can influence the effects of thimerosal (TM and other mercury (Hg compounds. TM is an organomercurial compound (49.55% Hg by weight that has been, and continues to be, used as a preservative in many childhood vaccines, particularly in developing countries. Thiol-modulating mechanisms affecting the cytotoxicity of TM have been identified. Importantly, the emergence of ASD symptoms post-6 months of age temporally follows the administration of many childhood vaccines. The purpose of the present critical review is provide mechanistic insight regarding how limited thiol availability, abnormal sulfation chemistry, and decreased GSH reserve capacity in children with an ASD could make them more susceptible to the toxic effects of TM routinely administered as part of mandated childhood immunization schedules.

  17. Investigations of thiol-modified phenol derivatives for the use in thiol–ene photopolymerizations

    Sebastian Reinelt

    2014-07-01

    Full Text Available Thiol–ene photopolymerizations gain a growing interest in academic research. Coatings and dental restoratives are interesting applications for thiol–ene photopolymerizations due to their unique features. In most studies the relative flexible and hydrophilic ester derivative, namely pentaerythritoltetra(3-mercaptopropionate (PETMP, is investigated as the thiol component. Thus, in the present study we are encouraged to investigate the performance of more hydrophobic ester-free thiol-modified bis- and trisphenol derivatives in thiol–ene photopolymerizations. For this, six different thiol-modified bis- and trisphenol derivatives exhibiting four to six thiol groups are synthesized via the radical addition of thioacetic acid to suitable allyl-modified precursors and subsequent hydrolysis. Compared to PETMP better flexural strength and modulus of elasticity are achievable in thiol–ene photopolymerizations employing 1,3,5-triallyl-1,3,5-triazine-2,4,6-trione (TATATO as the ene derivative. Especially, after storage in water, the flexural strength and modulus of elasticity is twice as high compared to the PETMP reference system.

  18. Synthesis and Microstructural Investigations of Organometallic Pd(II Thiol-Gold Nanoparticles Hybrids

    Cervellino Antonio

    2008-01-01

    Full Text Available Abstract In this work the synthesis and characterization of gold nanoparticles functionalized by a novel thiol-organometallic complex containing Pd(II centers is presented. Pd(II thiol,trans, trans-[dithiolate-dibis(tributylphosphinedipalladium(II-4,4′-diethynylbiphenyl] was synthesized and linked to Au nanoparticles by the chemical reduction of a metal salt precursor. The new hybrid made of organometallic Pd(II thiol-gold nanoparticles, shows through a single S bridge a direct link between Pd(II and Au nanoparticles. The size-control of the Au nanoparticles (diameter range 2–10 nm was achieved by choosing the suitable AuCl4 −/thiol molar ratio. The size, strain, shape, and crystalline structure of these functionalized nanoparticles were determined by a full-pattern X-ray powder diffraction analysis, high-resolution TEM, and X-ray photoelectron spectroscopy. Photoluminescence spectroscopy measurements of the hybrid system show emission peaks at 418 and 440 nm. The hybrid was exposed to gaseous NO x with the aim to evaluate the suitability for applications in sensor devices; XPS measurements permitted to ascertain and investigate the hybrid –gas interaction.

  19. The intramolecular electron transfer between copper sites of nitrite reductase

    Farver, O; Eady, R R; Abraham, Z H;

    1998-01-01

    The intramolecular electron transfer (ET) between the type 1 Cu(I) and the type 2 Cu(II) sites of Alcaligenes xylosoxidans dissimilatory nitrite reductase (AxNiR) has been studied in order to compare it with the analogous process taking place in ascorbate oxidase (AO). This internal process is......(I) and the trinuclear copper centre in ascorbate oxidase, and the characteristics of the internal ET processes of these enzymes are compared. The data are consistent with the faster ET observed in nitrite reductase arising from a more advantageous entropy of activation when compared with ascorbate...

  20. Effect of vanadium on nitrate reductase activity in tomato leaves

    J. Buczek

    2015-01-01

    The activity of nitrate reductase in cell-free extracts from tomato leaves is completely inhibited by 100 μM NaVO3 or VOCl2. In experiments in vivo vanadium ions inhibit the activity of the enzyme in 50 to 60 per cent. Addition of l mM vanadium to the medium on which tomato seedlings are grown causes after 24 h almost complete inhibition of nitrate reductase activity in cell-free extracts of the enzyme. Inhibition with vanadium may be abolished in experiments in vitro if the extract is treate...

  1. Transdermal thiol-acrylate polyethylene glycol hydrogel synthesis using near infrared light

    Chung, Solchan; Lee, Hwangjae; Kim, Hyung-Seok; Kim, Min-Gon; Lee, Luke P.; Lee, Jae Young

    2016-07-01

    Light-induced polymerization has been widely applied for hydrogel synthesis, which conventionally involves the use of ultraviolet or visible light to activate a photoinitiator for polymerization. However, with these light sources, transdermal gelation is not efficient and feasible due to their substantial interactions with biological systems, and thus a high power is required. In this study, we used biocompatible and tissue-penetrating near infrared (NIR) light to remotely trigger a thiol-acrylate reaction for efficient in vivo gelation with good controllability. Our gelation system includes gold nanorods as a photothermal agent, a thermal initiator, diacrylate polyethylene glycol (PEG), and thiolated PEG. Irradiation with a low-power NIR laser (0.3 W cm-2) could induce gelation via a mixed-mode reaction with a small increase in temperature (~5 °C) under the optimized conditions. We also achieved successful transdermal gelation via the NIR-assisted photothermal thiol-acryl reactions. This new type of NIR-assisted thiol-acrylate polymerization provides new opportunities for in situ hydrogel formation for injectable hydrogels and delivery of drugs/cells for various biomedical applications.Light-induced polymerization has been widely applied for hydrogel synthesis, which conventionally involves the use of ultraviolet or visible light to activate a photoinitiator for polymerization. However, with these light sources, transdermal gelation is not efficient and feasible due to their substantial interactions with biological systems, and thus a high power is required. In this study, we used biocompatible and tissue-penetrating near infrared (NIR) light to remotely trigger a thiol-acrylate reaction for efficient in vivo gelation with good controllability. Our gelation system includes gold nanorods as a photothermal agent, a thermal initiator, diacrylate polyethylene glycol (PEG), and thiolated PEG. Irradiation with a low-power NIR laser (0.3 W cm-2) could induce gelation

  2. Artificial electron donors for nitrate and nitrite reductases usable as mediators in amperometric biosensors

    Strehlitz, B. (Umweltforschungszentrum Leipzig-Halle GmbH, Leipzig (Germany)); Gruendig, B. (Institut fuer Chemo- und Biosensorik, Muenster-Roxel (Germany)); Vorlop, K.D. (Bundesforschungsanstalt fuer Landwirtschaft, Braunschweig (Germany). Inst. fuer Technologie); Bartholmes, P. (Witten-Herdecke Univ., Witten (Germany). Inst. fuer Biochemie); Kotte, H. (Umweltforschungszentrum Leipzig-Halle GmbH, Leipzig (Germany)); Stottmeister, U. (Umweltforschungszentrum Leipzig-Halle GmbH, Leipzig (Germany))

    1994-07-01

    Various nitrate and nitrite reductases are capable of accepting electrons from artificial donors. Combining these redox active donors with an amperometric redox electrode which is covered with an immobilized layer of such a nitrate or nitrite reductase, new enzyme sensors can be created for the detection of nitrate or nitrite, respectively. A range of suitable electron donors for nitrate reductases and nitrite reductase from different sources have been selected and characterized by electrochemical methods. (orig.)

  3. Acetate- and thiol-capped monodisperse ruthenium nanoparticles: XPS, XAS, and HRTEM studies.

    Chakroune, Nassira; Viau, Guillaume; Ammar, Souad; Poul, Laurence; Veautier, Delphine; Chehimi, Mohamed M; Mangeney, Claire; Villain, Françoise; Fiévet, Fernand

    2005-07-19

    Monodisperse ruthenium nanoparticles were prepared by reduction of RuCl3 in 1,2-propanediol. The mean particle size was controlled by appropriate choice of the reduction temperature and the acetate ion concentration. Colloidal solutions in toluene were obtained by coating the metal particles with dodecanethiol. High-resolution transmission electron microscopy (HRTEM), X-ray photoelectron spectroscopy (XPS), and X-ray absorption spectroscopy (XANES and EXAFS for the Ru K-absorption edge) were performed on particles of two different diameters, 2 and 4 nm, and in different environments, polyol/acetate or thiol. For particles stored in polyol/acetate XPS studies revealed superficial oxidation limited to one monolayer and a surface coating containing mostly acetate ions. Analysis of the EXAFS spectra showed both oxygen and ruthenium atoms around the ruthenium atoms with a Ru-Ru coordination number N smaller than the bulk value, as expected for fine particles. In the case of 2 nm acetate-capped particles N is consistent with particles made up of a metallic core and an oxidized monolayer. For 2 nm thiol-coated particles, a Ru-S bond was evidenced by XPS and XAS. For the 4 nm particles XANES and XPS studies showed that most of the ruthenium atoms are in the zerovalent state. Nevertheless, in both cases, when capped with thiol, the Ru-Ru coordination number inferred from EXAFS is much smaller than for particles of the same size stored in polyol. This is attributed to a structural disorganization of the particles by thiol chemisorption. HRTEM studies confirm the marked dependence of the structural properties of the ruthenium particles on their chemical environment; they show the acetate-coated particles to be single crystals, whereas the thiol-coated particles appear to be polycrystalline. PMID:16008388

  4. Cooperative functions of manganese and thiol redox system against oxidative stress in human spermatozoa

    Amrit Kaur Bansal

    2009-01-01

    Full Text Available Aims: In this study, the effects of 0.1 mM Mn 2+ on thiol components (total thiols [TSH], glutathione reduced [GSH], glutathione oxidized [GSSG] and redox ratio [GSH/ GSSG] have been determined in human spermatozoa. Settings and Design: The subjects of the study were healthy males having more than 75% motility and 80 x 10 6 sperms/mL. Materials and Methods: Fresh semen was suspended in phosphate-buffered saline (PBS (pH 7.2 and this suspension was divided into eight equal fractions. All fractions, control (containing PBS and experimental (treated/untreated with [ferrous ascorbate, FeAA - 200 FeSO 4 μM, 1000 μM ascorbic acid, nicotine (0.5 mM and FeAA + nicotine], supplemented/unsupplemented with Mn 2+ [0.1 mM], were incubated for 2 h at 378C. These fractions were assessed for determining the thiol components. Statistical Analysis: The data were statistically analyzed by Students " t" test. Results and Conclusions: Ferrous ascorbate, nicotine and ferrous ascorbate + nicotine induced oxidative stress and decreased GSH and redox ratio (GSH/GSSG ratio but increased the TSH and GSSG levels. Mn 2+ supplementation improved TSH, GSH and redox ratio (GSH/GSSG but decreased the GSSG level under normal and oxidative stress conditions. Thiol groups serve as defense mechanisms of sperm cells to fight against oxidative stress induced by stress inducers such as ferrous ascorbate, nicotine and their combination (ferrous ascorbate + nicotine. In addition, Mn 2+ supplementation maintains the thiol level by reducing oxidative stress.

  5. Inhibition of Albendazole and Oxfendazole on the Activity of Fumaric Reductase in Cysticercus cellulosae

    GAO Xue-jun; LI Qing-zhang; LI Xia

    2004-01-01

    The activity of fumaric reductase in Cysticercus cellulosae tissue homogenate with albendazole and oxfendazole individually was detected. Results showed that the two kinds of drugs both could inhabite the activity of fumaric reductase. The results indicate that the mechanism of action of benzimidazole carbamate drugs is probably inhabiting the complex of fumaric reductase noncompetently, thus lead to the exhaostion of energy and death.

  6. The design of redox active thiol peroxidase mimics: Dihydrolipoic acid recognition correlates with cytotoxicity and prooxidant action.

    Zadehvakili, B; McNeill, S M; Fawcett, J P; Giles, G I

    2016-03-15

    Redox active molecules containing organoselenium or organotellurium groups catalyse the oxidation of cellular thiols by hydrogen peroxide and are currently being developed as therapeutic agents. Potentially these synthetic thiol peroxidase (TPx) mimics can protect cells from oxidative stress by catalysing the reduction of reactive oxygen species by the cellular thiol glutathione, an activity which mimics the function of the antioxidant enzyme glutathione peroxidase. Alternatively they can act as prooxidants by catalysing the oxidation of essential thiol species within the cell. However the structure-activity relationships which determine the choice of thiol substrate, and hence the overall antioxidant or prooxidant outcome of drug administration, remain unknown. We report the first study that relates the pharmacological properties of TPx mimics with their solubility and catalytic activity using different thiol substrates. We used a series of structurally related compounds PhMCnH2n+1 (M=Se, Te; n=4-7) and investigated their ability to catalyse the oxidation of the cellular thiols glutathione and dihydrolipoic acid by hydrogen peroxide. The resulting rate constants (kobs) were then related to compound cytotoxicity and antioxidant versus prooxidant action in A549 cancer cells. The results show that the dihydrolipoic acid kobs values correlate with both cytotoxicity and prooxidant function. This enabled us to define a relationship, IC50=10+280e(-5(DHLA)(kobs)()), which allows the prediction of TPx mimic cytotoxicity. In contrast, hydrophobicity and glutathione kobs were unrelated to the compounds' redox pharmacology. PMID:26801688

  7. Reaction Mechanisms of Metals with Hydrogen Sulfide and Thiols in Model Wine. Part 1: Copper-Catalyzed Oxidation.

    Kreitman, Gal Y; Danilewicz, John C; Jeffery, David W; Elias, Ryan J

    2016-05-25

    Sulfidic off-odors as a result of hydrogen sulfide (H2S) and low-molecular-weight thiols are commonly encountered in wine production. These odors are usually removed by the process of Cu(II) fining, a process that remains poorly understood. The present study aims to elucidate the underlying mechanisms by which Cu(II) interacts with H2S and thiol compounds (RSH) under wine-like conditions. Copper complex formation was monitored along with H2S, thiol, oxygen, and acetaldehyde concentrations after the addition of Cu(II) (50 or 100 μM) to air-saturated model wine solutions containing H2S, cysteine, 6-sulfanylhexan-1-ol, or 3-sulfanylhexan-1-ol (300 μM each). The presence of H2S and thiols in excess to Cu(II) led to the rapid formation of ∼1.4:1 H2S/Cu and ∼2:1 thiol/Cu complexes, resulting in the oxidation of H2S and thiols and reduction of Cu(II) to Cu(I), which reacted with oxygen. H2S was observed to initially oxidize rather than form insoluble copper sulfide. The proposed reaction mechanisms provide insight into the extent to which H2S can be selectively removed in the presence of thiols in wine. PMID:27133282

  8. Determination of thiol metabolites in human urine by stable isotope labeling in combination with pseudo-targeted mass spectrometry analysis

    Liu, Ping; Qi, Chu-Bo; Zhu, Quan-Fei; Yuan, Bi-Feng; Feng, Yu-Qi

    2016-02-01

    Precursor ion scan and multiple reaction monitoring scan (MRM) are two typical scan modes in mass spectrometry analysis. Here, we developed a strategy by combining stable isotope labeling (IL) with liquid chromatography-mass spectrometry (LC-MS) under double precursor ion scan (DPI) and MRM for analysis of thiols in 5 types of human cancer urine. Firstly, the IL-LC-DPI-MS method was applied for non-targeted profiling of thiols from cancer samples. Compared to traditional full scan mode, the DPI method significantly improved identification selectivity and accuracy. 103 thiol candidates were discovered in all cancers and 6 thiols were identified by their standards. It is worth noting that pantetheine, for the first time, was identified in human urine. Secondly, the IL-LC-MRM-MS method was developed for relative quantification of thiols in cancers compared to healthy controls. All the MRM transitions of light and heavy labeled thiols were acquired from urines by using DPI method. Compared to DPI method, the sensitivity of MRM improved by 2.1-11.3 folds. In addition, the concentration of homocysteine, γ-glutamylcysteine and pantetheine enhanced more than two folds in cancer patients compared to healthy controls. Taken together, the method demonstrated to be a promising strategy for identification and comprehensive quantification of thiols in human urines.

  9. Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels

    M. Ryan Smith

    2016-08-01

    Full Text Available Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP, decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231 breast adenocarcinoma cells up to 6 days after an initial 24 h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10 µM of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC protein levels, although other protein levels were

  10. Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels.

    Smith, M Ryan; Vayalil, Praveen K; Zhou, Fen; Benavides, Gloria A; Beggs, Reena R; Golzarian, Hafez; Nijampatnam, Bhavitavya; Oliver, Patsy G; Smith, Robin A J; Murphy, Michael P; Velu, Sadanandan E; Landar, Aimee

    2016-08-01

    Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP), decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231) breast adenocarcinoma cells up to 6 days after an initial 24h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR) in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10µM) of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC) protein levels, although other protein levels were unaffected. This study