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

  1. Functional requirements for the lysosomal thiol reductase GILT in MHC class II-restricted antigen processing.

    Science.gov (United States)

    Hastings, K Taraszka; Lackman, Rebecca L; Cresswell, Peter

    2006-12-15

    Ag processing and presentation via MHC class II is essential for activation of CD4(+) T lymphocytes. gamma-IFN-inducible lysosomal thiol reductase (GILT) is present in the MHC class II loading compartment and has been shown to facilitate class II Ag processing and recall responses to Ags containing disulfide bonds such as hen egg lysozyme (HEL). Reduction of proteins within the MHC class II loading compartment is hypothesized to expose residues for class II binding and protease trimming. In vitro analysis has shown that the active site of GILT involves Cys(46) and Cys(49), present in a CXXC motif that shares similarity with the thioredoxin family. To define the functional requirements for GILT in MHC class II Ag processing, a GILT-deficient murine B cell lymphoma line was generated and stably transduced with wild-type and cysteine mutants of GILT. Intracellular flow cytometric, immunoblotting, and immunofluorescence analyses demonstrated that wild-type and mutant GILT were expressed and maintained lysosomal localization. Transduction with wild-type GILT reconstituted MHC class II processing of a GILT-dependent HEL epitope. Mutation of either Cys(46) or Cys(49) abrogated MHC class II processing of a GILT-dependent HEL epitope. In addition, biochemical analysis of these mutants suggested that the active site facilitates processing of precursor GILT to the mature form. Precursor forms of GILT-bearing mutations in Cys(200) or Cys(211), previously found to display thiol reductase activity in vitro, could not mediate Ag processing. These studies demonstrate that the thiol reductase activity of GILT is its essential function in MHC class II-restricted Ag processing. PMID:17142755

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

    International Nuclear Information System (INIS)

    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

  3. Molecular and biological characterization of interferon-?-inducible-lysosomal thiol reductase gene in zebrafish (Danio rerio).

    Science.gov (United States)

    Cui, Xian-wei; Ji, Chen-bo; Cao, Xin-guo; Fu, Zi-yi; Zhang, Shuang-quan; Guo, Xi-rong

    2012-11-01

    In mammals, interferon-?-inducible-lysosomal thiol reductase (GILT) has been demonstrated to play a key role in the processing and presentation of MHC class II-restricted antigen (Ag) by catalyzing disulfide bond reduction, thus unfolding native protein Ag and facilitating subsequent cleavage by proteases. Here, we reported the cloning of a GILT gene homologue from zebrafish (zGILT), a tropical freshwater fish. The full-length cDNA of zGILT gene is 768 nucleotides (nt) encoding a protein of 255 amino acids (aa), with a putative molecular weight of 28.33 kDa. The deduced protein is highly homologous to that of fish and mammalian GILTs and shares 57.1% sequence identity to that of Atlantic salmon and 55.7-21.6% sequence identity to that of various mammals. The deduced protein possesses all the main features characteristic of known GILT proteins including the signature sequence CQHGX2ECX2NX4C spanning residues 117-132, CXXC motif at residues 72-75, one potential sites for N-linked glycosylation at residual positions 54. The zGILT expression is obviously up-regulated in spleen and kidney after immunization with LPS although it also is constitutively expressed in heart, liver, muscle and intestine, suggesting that zGILT may be involved in the immune response to bacterial challenge. The soluble recombinant protein was successfully purified using Ni-nitrilotriacetic acid resin. Recombinant His-zsGILT appeared on SDS-PAGE in the ranges of their estimated size of 18.94-kDa. After purification, further study revealed that zsGILT was capable of catalyzing the reduction of the interchain disulfide bonds intact IgG. These results will allow for further investigation to unravel the role of this key enzyme in class II MHC-restricted antigen processing and to use zebrafish as an in vivo model for related studies. PMID:22982335

  4. Molecular characterization and expression analysis of interferon-gamma-inducible lysosomal thiol reductase (GILT) gene from pearl oyster Pinctada fucata.

    Science.gov (United States)

    Zhang, Dianchang; Pan, Dequan; Cui, Shuge; Su, Tianfeng; Qiu, Lihua; Zhu, Caiyan; Jiang, Shigui

    2010-09-01

    Interferon-gamma-inducible lysosomal thiol reductase (GILT) is an important thiol reductase, involved in class, MHC-restricted antigen processing by catalyzing disulfide bond reduction in mammals. Herein, we describe the identification and characterization of pearl oyster Pinctada fucata GILT (designated as poGILT). The poGILT cDNA was 1273bp long and consisted of a 5'-untranslated region (UTR) of 24bp, a 3'-UTR of 484bp with two cytokine RNA instability motifs (ATTTA), and an open reading frame (ORF) of 765bp encoding a polypeptide of 254 amino acids with an estimated molecular mass of 28.9kDa and a theoretical isoelectric point of 7.4. The N-terminus of the poGILT was found to have a putative signal peptide with a cleavage site amino acid position at 19-20. SMART analysis showed that the poGILT contained a GILT active-site C(69)PDC(72) motif and a GILT signature motif C(115)QHGKEECIGNLIETC(130). Homology analysis of the deduced amino acid sequence of the poGILT with other known GILT sequences by MatGAT software revealed that the poGILT shared 42.9-67.3% similarity and 22.9-49.8% identity to the other known GILT sequences. The expression level of poGILT mRNA was higher in digestive gland, moderate in adductor muscle, gills, gonad, intestine and mantle, and lower in hemocytes. The poGILT mRNA expression was significantly up-regulated in gill and digestive gland after LPS or V. alginolyticus stimulation, respectively. These results suggested that the poGILT was a constitutively expressed acute-phase protein, the expression of which can be enhanced after LPS or V. algrinolyticus stimulation, perhaps involved in the innate immune response of pearl oyster. PMID:20444427

  5. Molecular cloning, expression and functional characterization of interferon-?-inducible lysosomal thiol reductase (GILT) gene from mandarin fish (Siniperca chuatsi).

    Science.gov (United States)

    Song, Jinyun; Liu, Hongzhen; Ma, Lei; Ma, Li; Gao, Cuixiang; Zhang, Shuangquan

    2014-06-01

    Interferon-?-inducible lysosomal thiol reductase (GILT) plays a key role in the processing and presentation of MHC class II-restricted antigen (Ag) by catalyzing disulfide bond reduction, thus unfolding native protein Ag and facilitating subsequent cleavage by proteases. For this important function in the immune system, we cloned a GILT gene homologue from mandarin fish (designated mGILT), a kind of precious freshwater fish with high market value. Through reverse transcription PCR and rapid amplification of cDNA ends (RACE) strategies, we obtained the full-length cDNA of mGILT, which consists of 1008bp with a 771bp open reading frame, encoding a protein of 256 amino acids, with a putative molecular weight of 28.47kDa. The deduced protein possesses the typical structural features of known GILT proteins, including an active-site motif, a GILT signature sequence, and 6 conserved cysteines. The result of real-time quantitative PCR showed that mGILT mRNA was expressed in a tissue-specific manner. In addition, the expression of mGILT mRNA was obviously up-regulated in splenocytes and kidney after induction with lipopolysaccharide (LPS). Recombinant mGILT fused with His6 tag was efficiently expressed in Escherichia coli BL21 (DE3) and purified using Ni-nitrilotriacetic acid resin. Further study revealed that mGILT exhibit thiol reductase activity on IgG substrate. These results suggest mGILT is highly likely to play a role in the immune responses in mandarin fish. PMID:24698993

  6. Cloning and expression analysis of interferon-gamma-inducible-lysosomal thiol reductase gene in large yellow croaker (Pseudosciaena crocea).

    Science.gov (United States)

    Zheng, Wenbiao; Chen, Xinhua

    2006-05-01

    In mammals, interferon-gamma-inducible-lysosomal thiol reductase (GILT) has been demonstrated to play a key role in the processing and presentation of MHC class II-restricted antigen (Ag) by catalyzing disulfide bond reduction, thus unfolding native protein Ag and facilitating subsequent cleavage by proteases. Here, we reported the cloning of a GILT gene homologue from the spleen of large yellow croaker, a marine fish (LycGILT). The full-length cDNA of LycGILT gene is 1033 nucleotides (nt) encoding a protein of 256 amino acids (aa), with a putative molecular weight of 28.9 kDa. The deduced protein is highly homologous to that of mammalian and zebrafish GILTs and shares 54.1% sequence identity to that of zebrafish and 43.2-39.2% sequence identity to that of various mammals. The deduced LycGILT possesses the typical structural feature of mammalian GILT, including an active-site CXXC motif, a GILT signature sequence CQHGX2ECX2NX4C, and other six cysteines responsible for the formation of disulfide bonds in the C-terminus. Genomic analysis revealed that LycGILT gene, spanning a 3159nt fragment, contained seven exons interrupted by six introns and exhibited a similar exon-intron organization to human and mouse GILT genes except for a slightly more compact intron arrangement. The LycGILT expression is obviously up-regulated in spleen and kidney after immunization with inactivated trivalent bacterial vaccine consisting of Vibrio alginolyticus, V. paraphaemolyticus, and Aeromonas hydrophila although it also is constitutively expressed in liver, gills, brain, and heart, suggesting that LycGILT may be involved in the immune response to bacterial challenge in large yellow croaker. A search of NCBI sequence data with LycGILT cDNA identified a pufferfish (fugu rubrides) GILT homologue cDNA and its genomic DNA sequence, where two putative interferon-gamma activation sites (GAS) were found within the promoter region. This provided evidence that a fish GILT homologue like mammalian GILT, may also be regulated by interferon-gamma (IFN-gamma) through the JAK-STAT signal pathway. These results indicate that the bony fish GILT is a functional homologue of mammalian GILT. PMID:16478632

  7. Identification of interferon-?-inducible-lysosomal thiol reductase (GILT) gene in goldfish (Carassius auratus) and its immune response to LPS challenge.

    Science.gov (United States)

    Li, Jian Feng; Li, Jian; Wang, Zhi Guo; Liu, Hong Zhen; Zhao, You Long; Zhang, Jin Xi; Zhang, Shuang Quan; Liu, Jun Ping

    2015-02-01

    The interferon-?-inducible lysosomal thiol reductase (GILT) has been demonstrated to play an important role in the processing and presentation of MHC class II restricted antigen (Ag) by catalyzing disulfide bond reduction. In this study, we cloned a GILT gene homolog from goldfish (designated gGILT), a kind of precious freshwater fish with high market value. The open reading frame of gGILT consists of 756 bases encoding a protein of 251 amino acids with an estimated molecular mass of 27.8 kDa and a theoretical isoelectric point of 5.24. The deduced protein possesses the typical structural features of known GILT proteins, including an active-site motif, a GILT signature sequence, and 10 conserved cysteines. RT-PCR results showed that gGILT and gIFN-? (goldfish IFN-?) mRNA were expressed in a tissue-specific manner and obviously up-regulated in splenocytes and the cells from head kidney after induction with LPS. Recombinant gGILT fused with His6 tag was efficiently expressed in Escherichia coli BL21 (DE3) and purified by Ni-NTA affinity chromatography. Further study revealed that gGILT was capable of catalyzing the reduction of the interchain disulfide bonds from intact IgG. This study shows that gGILT may be involved in the immune response to bacteria challenge and maintain first line of innate immune defense at basal level in goldfish. It also provides the basis for investigating on the role of GILT using goldfish as an animal model. PMID:25447639

  8. Thiol biochemistry of prokaryotes

    Science.gov (United States)

    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. Electroanalysis of Plant Thiols

    Directory of Open Access Journals (Sweden)

    Rene Kizek

    2007-06-01

    Full Text Available Due to unique physico-chemical properties of –SH moiety thiols comprise widegroup of biologically important compounds. A review devoted to biological functions ofglutathione and phytochelatins with literature survey of methods used to analysis of thesecompounds and their interactions with cadmium(II ions and Murashige-Skoog medium ispresented. For these purposes electrochemical techniques are used. Moreover, we revealedthe effect of three different cadmium concentrations (0, 10 and 100 μM on cadmiumuptake and thiols content in maize plants during 192 hours long experiments usingdifferential pulse anodic stripping voltammetry to detect cadmium(II ions and highperformance liquid chromatography with electrochemical detection to determineglutathione. Cadmium concentration determined in tissues of the plants cultivated innutrient solution containing 10 μM Cd was very low up to 96 hours long exposition andthen the concentration of Cd markedly increased. On the contrary, the addition of 100 μMCd caused an immediate sharp increase in all maize plant parts to 96 hours Cd expositionbut subsequently the Cd concentration increased more slowly. A high performance liquidchromatography with electrochemical detection was used for glutathione determination intreated maize plants after 96 and 192 hours of treatment. The highest total content of glutathione per one plant was 6 μg (96 h, 10 μM Cd in comparison with non-treated plant (control where glutathione content was 1.5 μg. It can be concluded that electrochemical techniques have proved to be useful to analyse plant thiols.

  10. Rapid and thiol-specific high-throughput assay for simultaneous relative quantification of total thiols, protein thiols, and nonprotein thiols in cells.

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    Yang, Yang; Guan, Xiangming

    2015-01-01

    Thiol groups in biological molecules play a significant role in various physiological functions and pathological conditions. Thiols are divided into two major groups: protein thiols and nonprotein thiols. Numerous methods have been reported for thiol assays. Most of these methods have been developed for glutathione, the principal nonprotein thiol, despite the fact that cellular protein thiols are more abundant than glutathione. Further, these methods usually involve a process of biological sample preparation followed by a separation method, and they are time-consuming. We reported previously a series of thiol-specific fluorogenic benzofurazan sulfides. These nonfluorescent benzofurazan sulfides react rapidly and specifically with a thiol to form a strong fluorescent thiol adduct. The rapid reaction, thiol-specific and fluorogenic nature of the sulfides successfully yielded an application of one of the sulfides for relative quantitation of total thiols in live cells through fluorescence microscopy. In this work, we employed the same compound to develop the first high-throughput method for simultaneous monitoring of protein thiols, nonprotein thiols, and total thiols in cells in a 96-well plate on a fluorescence microplate reader at ?(ex) = 430 nm and ?(em) = 520 nm, respectively. The method is rapid and sensitive, and has been validated by an HPLC thiol assay method. The method can detect thiols with cell concentrations as low as 500 cells/well. We also demonstrated that the method can readily monitor changes in cellular thiol levels. Although the method cannot provide an absolute quantification for thiols because fluorescence intensity of different thiol adducts varies, it provides an accurate measurement of relative quantification, relative to the control. The method will be a valuable tool in thiol-related biomedical/pharmaceutical research. PMID:25423115

  11. Polythioethers by thiol-ene click polyaddition of ?,?-alkylene thiols.

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    Deubel, Frank; Bretzler, Victor; Holzner, Richard; Helbich, Tobias; Nuyken, Oskar; Rieger, Bernhard; Jordan, Rainer

    2013-06-25

    The straightforward synthesis of a series of poly(thioether)s by photoinduced thiol-ene click polyaddition of ?,?-alkylene thiols is reported. It is found that linear and telechelic poly(thioether)s can be directly obtained from ?,?-alkylene thiols with, for example, alkyl chain length of m = 1,2,3, and 9. The reaction proceeds without additives such as (radical) initiators or metal compounds and can simply be carried out by UV-irradiation of the bulk monomer or monomer solution. Ex situ kinetic studies reveal that the reaction proceeds by a typical a step-growth polyaddition mechanism. As the homologue series of poly(thioether)s are now synthetically accessible, new direct pathways to tailored poly(alkyl sulphoxide)s and poly(alkyl sulfone)s are now possible. PMID:23649794

  12. Quantification of thiols and disulfides

    DEFF Research Database (Denmark)

    Winther, Jakob R.; Thorpe, Colin

    2014-01-01

    Disulfide bond formation is a key posttranslational modification, with implications for structure, function and stability of numerous proteins. While disulfide bond formation is a necessary and essential process for many proteins, it is deleterious and disruptive for others. Cells go to great lengths to regulate thiol-disulfide bond homeostasis, typically with several, apparently redundant, systems working in parallel. Dissecting the extent of oxidation and reduction of disulfides is an ongoing challenge due, in part, to the facility of thiol/disulfide exchange reactions.

  13. Inactivation of Penicillins by Thiol Broth

    OpenAIRE

    Murray, Patrick R.; Niles, Ann C.

    1982-01-01

    Thiol broth with sodium polyanetholesulfonate inactivated penicillin G, carbenicillin, nafcillin, oxacillin, and gentamicin, but had no effect on cephalothin, cefoxitin, clindamycin, chloramphenicol, erythromycin, and tetracycline. Only Thiol broth was capable of this inactivation, which was not influenced by the presence of blood.

  14. Enzymatic reduction of alloxan by thioredoxin and NADPH-thioredoxin reductase.

    OpenAIRE

    Holmgren, A.; Lyckeborg, C.

    1980-01-01

    Alloxan reacts with certain sulfhydryl groups either by chemical modification or reduction to dialuric acid. The effects of the drug on NADPH-thioredoxin oxidoreductase, EC 1.6.4.5] and thioredoxin-(SH)2, a ubiquitous thiol-dependent disulfide reductase system, are described. Alloxan was a direct substrate for a nearly homogenous preparation of calf thymus NADPH-thioredoxin reductase with an apparent Km of 330 microM and a Kcat of 1000 min-1 at pH 7.0 and 25 degrees C. Alloxan was not a subst...

  15. Platyhelminth Mitochondrial and Cytosolic Redox Homeostasis Is Controlled by a Single Thioredoxin Glutathione Reductase and Dependent on Selenium and Glutathione*S?

    OpenAIRE

    Bonilla, Mariana; Denicola, Ana; Novoselov, Sergey V.; Turanov, Anton A.; Protasio, Anna; Izmendi, Darwin; Gladyshev, Vadim N.; Salinas, Gustavo

    2008-01-01

    Platyhelminth parasites are a major health problem in developing countries. In contrast to their mammalian hosts, platyhelminth thiol-disulfide redox homeostasis relies on linked thioredoxin-glutathione systems, which are fully dependent on thioredoxin-glutathione reductase (TGR), a promising drug target. TGR is a homodimeric enzyme comprising a glutaredoxin domain and thioredoxin reductase (TR) domains with a C-terminal redox center containing selenocysteine (Sec). In...

  16. Thiol-based redox switches in prokaryotes.

    Science.gov (United States)

    Hillion, Melanie; Antelmann, Haike

    2015-05-01

    Bacteria encounter reactive oxygen species (ROS) as a consequence of the aerobic life or as an oxidative burst of activated neutrophils during infections. In addition, bacteria are exposed to other redox-active compounds, including hypochloric acid (HOCl) and reactive electrophilic species (RES) such as quinones and aldehydes. These reactive species often target the thiol groups of cysteines in proteins and lead to thiol-disulfide switches in redox-sensing regulators to activate specific detoxification pathways and to restore the redox balance. Here, we review bacterial thiol-based redox sensors that specifically sense ROS, RES and HOCl via thiol-based mechanisms and regulate gene transcription in Gram-positive model bacteria and in human pathogens, such as Staphylococcus aureus and Mycobacterium tuberculosis. We also pay particular attention to emerging widely conserved HOCl-specific redox regulators that have been recently characterized in Escherichia coli. Different mechanisms are used to sense and respond to ROS, RES and HOCl by 1-Cys-type and 2-Cys-type thiol-based redox sensors that include versatile thiol-disulfide switches (OxyR, OhrR, HypR, YodB, NemR, RclR, Spx, RsrA/RshA) or alternative Cys phosphorylations (SarZ, MgrA, SarA), thiol-S-alkylation (QsrR), His-oxidation (PerR) and methionine oxidation (HypT). In pathogenic bacteria, these redox-sensing regulators are often important virulence regulators and required for adapation to the host immune defense. PMID:25720121

  17. Cytoplasmic glutathione redox status determines survival upon exposure to the thiol-oxidant 4,4'-dipyridyl disulfide.

    DEFF Research Database (Denmark)

    Lpez-Mirabal, H Reynaldo; Thorsen, Michael

    2007-01-01

    Dipyridyl disulfide (DPS) is a highly reactive thiol oxidant that functions as electron acceptor in thiol-disulfide exchange reactions. DPS is very toxic to yeasts, impairing growth at low micromolar concentrations. The genes TRX2 (thioredoxin), SOD1 (superoxide dismutase), GSH1 (gamma-glutamyl-cysteine synthetase) and, particularly, GLR1 (glutathione reductase) are required for survival on DPS. DPS is uniquely thiol-specific, and we found that the cellular mechanisms for DPS detoxification differ substantially from that of the commonly used thiol oxidant diamide. In contrast to this oxidant, the full antioxidant pools of glutathione (GSH) and thioredoxin are required for resistance to DPS. We found that DPS-sensitive mutants display increases in the disulfide form of GSH (GSSG) during DPS exposure that roughly correlate with their more oxidizing GSH redox potential in the cytosol and their degree of DPS sensitivity. DPS seems to induce a specific disulfide stress, where an increase in the cytoplasmic/nuclearGSSG/GSH ratio results in putative DPS target(s) becoming sensitive to DPS. Udgivelsesdato: 2007-May

  18. Endothelium-dependent vasorelaxation in inhibited by in vivo depletion of vascular thiol levels : role of endothelial nitric oxide synthase

    DEFF Research Database (Denmark)

    Laursen, J B; Boesgaard, S

    2001-01-01

    Thiols like glutathione may serve as reducing cofactors in the production of nitric oxide (NO) and protect NO from inactivation by radical oxygen species. Depletion of thiol compounds reduces NO-mediated vascular effects in vitro and in vivo. The mechanisms underlying these actions are not clear, but may involve decreased synthesis of NO and/or increased degradation of NO. This study investigates the effect of glutathione depletion on the response to NO-mediated vasodilation induced by acetylcholine (Ach, 10 micrograms/kg), endothelial NO synthase (eNOS) activity and potential markers of vascular superoxide anion (O2.-) production in conscious chronically catheterized rats. Thiol depletion induced by buthionine sulfoximine (BSO, 1 g i.p. within 24 h) decreased the hypotensive effect of Ach by 30% (MAP reduction before BSO 27 +/- 3 mmHg, 19 +/- 3 mmHg after BSO, (mean +/- SEM), p < .05, n = 8). The impaired effect of Ach was associated with a significant reduction in eNOS activity (control: 7.7 +/- 0.8, BSO: 3.9 +/- 0.4 pmol/min/mg protein (p < .05), n = 6). In contrast, neither NADH/NADPH driven membrane-associated oxidases nor lucigenin reductase activity were significantly (p < .05) 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.

  19. Molecular study of DNA radioprotection by thiols

    Energy Technology Data Exchange (ETDEWEB)

    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)

  20. Thiol modification in kidney: Implications for radioprotection

    International Nuclear Information System (INIS)

    Kidney radiosensitivity is frequently a dose-limiting factor when planning abdominal radiation therapy. In addition to its clinical importance, the kidney has unusual transport and metabolic activity such as high gamma-glutamyl transpeptidase (Meister Methods in Enzymol, vol. 113, pg. 571-585, 1985), making it an ideal organ to study thiol modification. Although glutahione has been most extensively investigated, attention to other thiols may be also be of interest, since some thiols may be more effective as radioprotectors. Glutathione, glutathione ester, and cysteine concentrations in the kidney can be modified. The authors believe is cysteine is an attractive agent for kidney radioprotection. The data is used in conjunction with results from radiochemical studies with model systems to design strategies for kidney radioprotection

  1. Total Thiols: Biomedical Importance And Their Alteration In Various Disorders

    Directory of Open Access Journals (Sweden)

    Mungli Prakash

    2009-09-01

    Full Text Available Thiols are the organic compounds that contain a sulphydryl group. Among all the antioxidants that are available in the body, thiols constitute the major portion of the total body antioxidants and they play a significant role in defense against reactive oxygen species. Total thiols composed of both intracellular and extracellular thiols either in the free form as oxidized or reduced glutathione, or thiols bound to proteins. Among the thiols that are bound to proteins, albumin makes the major portion of the protein bound thiols, which binds to sufhydryl group at its cysteine-34 portion. Apart from their role in defense against free radicals, thiols share significant role in detoxification, signal transduction, apoptosis and various other functions at molecular level. The thiol status in the body can be assessed easily by determining the serum levels of thiols. Decreased levels of thiols has been noted in various medical disorders including chronic renal failure and other disorders related to kidney, cardiovascular disorders, stroke and other neurological disorders, diabetes mellitus, alcoholic cirrhosis and various other disorders. Therapy using thiols has been under investigation for certain disorders.

  2. Thiol-ene-based monolithic microreactors.

    Czech Academy of Sciences Publication Activity Database

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

    Brno : Institute of Analytical Chemistry AS CR, 2014 - (Foret, F.; K?enkov, J.; Drobnkov, I.; Guttman, A.; Kleprnk, 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

  3. Thiol-derivatized minihepcidins retain biological activity.

    Science.gov (United States)

    Fung, Eileen; Chua, Kristine; Ganz, Tomas; Nemeth, Elizabeta; Ruchala, Piotr

    2015-02-15

    Minihepcidins are small peptides that mimic biological activity of the iron-regulatory hormone hepcidin. Structurally, they contain thiol-free-cysteine residue in position 7 which is crucial for their bioactivity. Nonetheless, free sulfhydryl group is not desirable in pharmaceutical entities as it may lead to dermatological side effects. Moreover free thiol moiety is quite reactive and depending on conditions/reagents may be alkylated and/or oxidized giving various Cys-derivatives: S-alkyl cysteines, sulfoxides, sulfones, disulfides, cysteinesulfinic and cysteic acids. To limit such reactivity and maintain bioactivity of minihepcidin(s) we used thiol-protection strategy based on activated vinyl thioethers. Novel S-protected analogs of physiologically active minihepcidin PR73 were synthesized and tested in vitro showing activity comparable to parental molecule. The most active compound, PR73SH was also tested in vivo showing activity profile analogous to PR73. Collectively, our findings suggest that S-vinyl-derivatization of minihepcidin(s) may be a suitable approach in the development of physiologically active agonists of hepcidin. PMID:25599838

  4. Respiratory substrates regulate S-nitrosylation of mitochondrial proteins through a thiol-dependent pathway.

    Science.gov (United States)

    Chang, Allen H K; Sancheti, Harsh; Garcia, Jerome; Kaplowitz, Neil; Cadenas, Enrique; Han, Derick

    2014-05-19

    S-Nitrosylation is a reversible post-translational modification on cysteinyl thiols that can modulate the function of redox-sensitive proteins. The S-nitrosylation of mitochondrial proteins has been shown to regulate various mitochondrial activities involved in energy-transducing systems and mitochondrion-driven apoptosis. In isolated rat brain mitochondria, we demonstrate that mitochondrial protein S-nitrosylation is regulated by respiratory substrates (glutamate/malate) through a thiol-dependent pathway. Mitochondrial proteins become susceptible to S-nitrosoglutathione (GSNO)-induced S-nitrosylation in mitochondria with an oxidized environment (low glutathione (GSH), NADH, and NADPH, and high GSSG, NAD(+), and NADP(+)) caused by isolation of mitochondria using a discontinuous Percoll gradient. Activation of mitochondrial respiration by respiratory substrates leads to increased NAD(P)H and GSH levels, which in turn reduces mitochondrial S-nitrosylated proteins. 1-Chloro-2,4-dinitrobenzene (CDNB), which depletes mitochondrial GSH and inhibits the thioredoxin-thioredoxin reductase system, prevented the denitrosylation of mitochondrial proteins caused by respiratory substrate treatment. Using biotin-switch coupled with LC-MS/MS, several mitochondrial proteins were identified as targets of S-nitrosylation including adenine nucleotide translocase (ANT) and voltage-dependent anion channel (VDAC), important components of the mitochondria permeability transition pore (MPTP), as well as ATP synthase. The S-nitrosylation of ATP synthase by GSNO was found to inhibit its activity. These findings emphasize the importance of respiratory substrates in regulating S-nitrosylation through a thiol-dependent (GSH and/or thioredoxin) pathway, with implications for mitochondrial bioenergetics and mitochondrion-driven apoptosis. PMID:24716714

  5. Thiol-disulfide oxidoreductases in Bacillus subtilis

    OpenAIRE

    Dorenbos, Ronald,

    2004-01-01

    An important function in protein folding is the formation of disulfide bonds. In different compartments of bacteria specific oxido reductases are responsible for this function. Thanks to the identification of all genes from various model organisms the understanding of the function of the various members of this family has increased dramatically. Zie: Summary

  6. NADP(+)-dependent dehydrogenase activity of carbonyl reductase on glutathionylhydroxynonanal as a new pathway for hydroxynonenal detoxification.

    Science.gov (United States)

    Moschini, Roberta; Peroni, Eleonora; Rotondo, Rossella; Renzone, Giovanni; Melck, Dominique; Cappiello, Mario; Srebot, Massimo; Napolitano, Elio; Motta, Andrea; Scaloni, Andrea; Mura, Umberto; Del-Corso, Antonella

    2015-06-01

    An NADP(+)-dependent dehydrogenase activity on 3-glutathionyl-4-hydroxynonanal (GSHNE) was purified to electrophoretic homogeneity from a line of human astrocytoma cells (ADF). Proteomic analysis identified this enzymatic activity as associated with carbonyl reductase 1 (EC 1.1.1.184). The enzyme is highly efficient at catalyzing the oxidation of GSHNE (KM 33M, kcat 405min(-1)), as it is practically inactive toward trans-4-hydroxy-2-nonenal (HNE) and other HNE-adducted thiol-containing amino acid derivatives. Combined mass spectrometry and nuclear magnetic resonance spectroscopy analysis of the reaction products revealed that carbonyl reductase oxidizes the hydroxyl group of GSHNE in its hemiacetal form, with the formation of the corresponding 3-glutathionylnonanoic-?-lactone. The relevance of this new reaction catalyzed by carbonyl reductase 1 is discussed in terms of HNE detoxification and the recovery of reducing power. PMID:25680283

  7. Designed Chemical Intervention with Thiols for Prophylactic Contraception

    OpenAIRE

    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 synt...

  8. Thiol-Based Redox Switches in Eukaryotic Proteins

    OpenAIRE

    Brandes, Nicolas; Schmitt, Sebastian; Jakob, Ursula

    2009-01-01

    For many years, oxidative thiol modifications in cytosolic proteins were largely disregarded as in vitro artifacts, and considered unlikely to play significant roles within the reducing environment of the cell. Recent developments in in vivo thiol trapping technology combined with mass spectrometric analysis have now provided convincing evidence that thiol-based redox switches are used as molecular tools in many proteins to regulate their activity in response to reactive oxygen and nitrogen s...

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

    OpenAIRE

    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...

  10. Enrichment of O-GlcNAc-modified peptides using novel thiol-alkyne and thiol-disulfide exchange.

    Science.gov (United States)

    Tsumoto, Hiroki; Ogasawara, Daisuke; Hashii, Noritaka; Suzuki, Takayoshi; Akimoto, Yoshihiro; Endo, Tamao; Miura, Yuri

    2015-07-01

    We have developed a selective method for the enrichment of O-linked ?-N-acetylglucosamine (O-GlcNAc)-modified peptides, which uses a newly synthesized thiol-alkyne and a thiol-disulfide exchange. First, O-GlcNAc-modified peptides were enzymatically labeled with an azide-containing GalNAc analog. Then, the azide moiety was reacted with thiol-alkyne through a copper(I)-catalyzed azide-alkyne cycloaddition. The thiol-modified peptides were enriched with thiol-reactive resin through a thiol-disulfide exchange. At least 500fmol of O-GlcNAc-modified peptides was selectively isolated from ?-crystallin tryptic peptides and detected by mass spectrometry. This novel enrichment strategy could be used for O-GlcNAcome analysis of biological samples. PMID:25980911

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

    International Nuclear Information System (INIS)

    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)

  12. Chirality sensing using Ag(+)-thiol coordination polymers.

    Science.gov (United States)

    Zhang, Qian; Hong, Yuan; Chen, Na; Tao, Dan-Dan; Li, Zhao; Jiang, Yun-Bao

    2015-05-11

    We report here chirality sensing using achiral Ag(+)-thiol coordination polymers as the framework which becomes CD active upon interaction with chiral species, taking p-mercaptophenylboronic acid as a thiol ligand that bears a binding group for monosaccharides. PMID:25766879

  13. Genetics Home Reference: 5-alpha reductase deficiency

    Science.gov (United States)

    ... the Dominican Republic, Papua New Guinea, Turkey, and Egypt. What genes are related to 5-alpha reductase ... Read more about the SRD5A2 gene. How do people inherit 5-alpha reductase deficiency? This condition is ...

  14. Purification and kinetic analysis of cytosolic and mitochondrial thioredoxin glutathione reductase extracted from Taenia solium cysticerci.

    Science.gov (United States)

    Plancarte, Agustin; Nava, Gabriela

    2015-02-01

    Thioredoxin glutathione reductases (TGRs) (EC 1.8.1.9) were purified to homogeneity from the cytosolic (cTsTGR) and mitochondrial (mTsTGR) fractions of Taenia solium, the agent responsible for neurocysticercosis, one of the major central nervous system parasitic diseases in humans. TsTGRs had a relative molecular weight of 132,000, while the corresponding value per subunit obtained under denaturing conditions, was of 62,000. Specific activities for thioredoxin reductase and glutathione reductase substrates for both TGRs explored were in the range or lower than values obtained for other platyhelminths and mammalian TGRs. cTsTGR and mTsTGR also showed hydroperoxide reductase activity using hydroperoxide as substrate. Km(DTNB) and Kcat(DTNB) values for cTsTGR and mTsTGR (88?M and 1.9?s(-1); 45?M and 12.6?s(-1), respectively) and Km(GSSG) and Kcat(GSSG) values for cTsTGR and mTsTGR (6.3?M and 0.96?s(-1); 4?M and 1.62?s(-1), respectively) were similar to or lower than those reported for mammalian TGRs. Mass spectrometry analysis showed that 12 peptides from cTsTGR and seven from mTsTGR were a match for gi|29825896 thioredoxin glutathione reductase [Echinococcus granulosus], confirming that both enzymes are TGRs. Both T. solium TGRs were inhibited by the gold compound auranofin, a selective inhibitor of thiol-dependent flavoreductases (I???=?3.25, 2.29?nM for DTNB and GSSG substrates, respectively for cTsTGR; I???=?5.6, 25.4?nM for mTsTGR toward the same substrates in the described order). Glutathione reductase activity of cTsTGR and mTsTGR exhibited hysteretic behavior with moderate to high concentrations of GSSG; this result was not observed either with thioredoxin, DTNB or NADPH. However, the observed hysteretic kinetics was suppressed with increasing amounts of both parasitic TGRs. These data suggest the existence of an effective substitute which may account for the lack of the detoxification enzymes glutathione reductase and thioredoxin reductase in T. solium, as has been described for very few other platyhelminths. PMID:25541385

  15. The thiol pool in human plasma: The central contribution of albumin to redox processes

    OpenAIRE

    Turell, Luca; Radi, Rafael; Alvarez, Beatriz

    2013-01-01

    The plasma compartment has particular features regarding the nature and concentration of low and high molecular weight thiols and oxidized derivatives. Plasma is relatively poor in thiol-based antioxidants; thiols are in lower concentrations than in cells and mostly oxidized. The different thiol-disulfide pairs are not in equilibrium and the steady-state concentrations of total thiols as well as reduced versus oxidized ratios are maintained by kinetic barriers, including the rates of reaction...

  16. Evidence for the participation of Cys558 and Cys559 at the active site of mercuric reductase

    International Nuclear Information System (INIS)

    Mercuric reductase, with FAD and a reducible disulfide at the active site, catalyzes the two-electron reduction of Hg(II) by NADPH. Addition of reducing equivalents rapidly produces a spectrally distinct EH2 form of the enzyme containing oxidized FAD and reduced active site thiols. Formation of EH2 has previously been reported to require only 2 electrons for reduction of the active site disulfide. The authors present results of anaerobic titrations of mercuric reductase with NADPH and dithionite showing that the equilibrium conversion of oxidized enzyme to EH2 actually requires 2 equiv of reducing agent or 4 electrons. Kinetic studies conducted both at 4 degree C and at 25 degree C indicate that reduction of the active site occurs rapidly, as previously reported; this is followed by a slower reduction of another redox group via reaction with the active site. [14C]Iodoacetamide labeling experiments demonstrate that the C-terminal residues, Cys558 and Cys559, are involved in this disulfide. The fluorescence, but not the absorbance, of the enzyme-bound FAD was found to be highly dependent on the redox state of the C-terminal thiols. Thus, Eox with Cys558 and Cys559 as thiols exhibits less than 50% of the fluorescence of Eox where these residues are present as a disulfide, indicating that the thiols remain intimately associated with the active site. Initial velociiated with the active site. Initial velocity measurements show that the auxiliary disulfide must be reduced before catalytic Hg(II) reduction can occur, consistent with the report of a preactivation phenomenon with NADPH or cysteine. A modified minimal catalytic mechanism is proposed as well as several chemical mechanisms for the Hg(II) reduction step

  17. Hydrophillic Thiols in the Porewaters of Coastal Marine Sediments

    Science.gov (United States)

    Kalnejais, L.; Martin, W. R.; Moffett, J. W.; Nelson, R. K.; Francois, R.

    2003-12-01

    Porewater profiles of thiolic compounds have been determined at a strongly reducing site in Boston Harbor, Massachusetts. Derivatization with monobromobimane followed by reverse phase high performance liquid chromatography and fluorescence detection have been used to detect at least six thiols in the porewaters. Maximum individual thiol concentrations of up to 10 ? M (glutathione equivalents) were either coincident with the sulfide maximum or showed an increase with depth. The profiles of at least two thiols indicate a flux to the overlying water. Samples from a benthic chamber have detected glutathione and several of the unidentified thiols in the oxygenated water overlying these sediments. Silver, copper, and lead porewater profiles have also been determined so that the importance of the metal-RS complexation in porewaters can be considered.

  18. Glutation. The thiol in intracellular is a good radioprotector?

    International Nuclear Information System (INIS)

    Since 1988 there is a changing opinion about the Glutation's role in intracellular radiation protection. We review the thiols mechanism of radiation protection with special attention to GSH; and present the new features about this topic. (Author) 125 ref

  19. DNA microarrays on silicon surfaces through thiol-ene chemistry

    OpenAIRE

    Escorihuela Fuentes, Jorge; Bauls Polo, M Jos; Puchades Pla, Rosa; Maquieira Catala, ngel

    2012-01-01

    The potential of thiol-ene chemistry as a selective strategy to functionalize silicon materials for DNA microarraying is demonstrated and applied to discriminate genetic variations. 2012 The Royal Society of Chemistry.

  20. Induction Curing of Thiol-acrylate and Thiolene Composite Systems

    OpenAIRE

    Ye, Sheng; Cramer, Neil B.; Stevens, Blake E.; Sani, Robert L.; Bowman, Christopher N

    2011-01-01

    Induction curing is demonstrated as a novel type of in situ radiation curing that maintains most of the advantages of photocuring while eliminating the restriction of light accessibility. Induction curing is utilized to polymerize opaque composites comprised of thiol-acrylate and thiol-ene resins, nanoscale magnetic particles, and carbon nanotubes. Nanoscale magnetic particles are dispersed in the resin and upon exposure to the magnetic field, these particles lead to induction heating that ra...

  1. ORIGINAL ARTICLE: Protein Thiols in Essential Hypertension Patients

    Directory of Open Access Journals (Sweden)

    Jagadish. B. Ingin

    2012-01-01

    Full Text Available Background: Essential hypertension is one of the most prevalent diseases of the world and is an unequivocal risk factor for cardiovascular morbidity and mortality. Several previous studies have shown that increased free radical activity is suggested to play an important role in the lipid peroxidation and protein oxidationof cellular structures causing cell injury and is implicated in the pathogenesis of vascular disease and essential hypertension is one ofthem. The thiols groups on proteins are known to play a major role in maintaining the antioxidant status of the body. Aims and Objectives: The current study was designed to find out the levels of such protein bound thiols in essential hypertension patients. Materials and Methods: The study was conducted on 45 essential hypertension patients and 25 healthy subjects without any underlying medical disorder as controls. Patients were divided into three groups according to the 2003 European Society of Hypertension-European Society of Cardiology (ESH/ ESC guidelines (grade I, II,III. Serum and urine protein thiols were determined by spectrophotometric methods using dithionitrobenzoic acid (DTNB andcorrelated with blood pressure (BP. Results: There was a significant decrease in serum protein thiols in essential hypertensive patients compared to healthy controls (p<0.01 and correlated positively with grade II and grade III essential hypertension. In urine samples there was a significant increase in protein and a decrease in protein bound thiols (p<0.01 which also correlated positively with grade II and grade III essential hypertension. Conclusions: Protein bound thiols; the major antioxidants in the body are decreased in essential hypertensive patients. Due to increased consumption of protein bound thiols in such oxidative environment, there was a significant decrease in protein bound thiols in urine.

  2. Protein Thiols as an Indication of Oxidative Stress

    Directory of Open Access Journals (Sweden)

    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

  3. Quantifying Reversible Oxidation of Protein Thiols in Photosynthetic Organisms

    Science.gov (United States)

    Slade, William O.; Werth, Emily G.; McConnell, Evan W.; Alvarez, Sophie; Hicks, Leslie M.

    2015-04-01

    Photosynthetic organisms use dynamic post-translational modifications to survive and adapt, which include reversible oxidative modifications of protein thiols that regulate protein structure, function, and activity. Efforts to quantify thiol modifications on a global scale have relied upon peptide derivatization, typically using isobaric tags such as TMT, ICAT, or iTRAQ that are more expensive, less accurate, and provide less proteome coverage than label-free approachessuggesting the need for improved experimental designs for studies requiring maximal coverage and precision. Herein, we present the coverage and precision of resin-assisted thiol enrichment coupled to label-free quantitation for the characterization of reversible oxidative modifications on protein thiols. Using C. reinhardtii and Arabidopsis as model systems for algae and plants, we quantified 3662 and 1641 unique cysteinyl peptides, respectively, with median coefficient of variation (CV) of 13% and 16%. Further, our method is extendable for the detection of protein abundance changes and stoichiometries of cysteine oxidation. Finally, we demonstrate proof-of-principle for our method, and reveal that exogenous hydrogen peroxide treatment regulates the C. reinhardtii redox proteome by increasing or decreasing the level of oxidation of 501 or 67 peptides, respectively. As protein activity and function is controlled by oxidative modifications on protein thiols, resin-assisted thiol enrichment coupled to label-free quantitation can reveal how intracellular and environmental stimuli affect plant survival and fitness through oxidative stress.

  4. All in the family: aldose reductase and closely related aldo-keto reductases.

    Science.gov (United States)

    Petrash, J M

    2004-04-01

    Aldose reductase catalyzes the first step in the polyol pathway and is thought to be involved in the pathogenesis of diabetic complications. In addition to polyol synthesis, aldose reductase may have multiple other activities that intersect with signal processing and oxidative defense mechanisms. Multiple aldose reductase-like proteins have been discovered to have structures and catalytic properties that broadly overlap those of aldose reductase. This chapter will summarize new insights into properties and functions of aldose reductase and closely related members of the aldo-keto reductase enzyme superfamily. PMID:15094999

  5. Comparison of concentration changes in nonprotein SH-groups, reduced glutathione and glutathione reductase activity in some radioprotection methods

    International Nuclear Information System (INIS)

    The concentration dynamics of nonprotein SH-groups, reduced glutathione, and glutathione reductase activity were compared in different rat tissues in the course of the first hour following intraperitoneal administration of radioprotectively effective doses of cystamine or of a combination of cystamine with mexamine. The results obtained confirmed the participation of endogenous reduced glutathione in the radioprotective effect of cystamine in the rat organism, showing, however, that the increase in the concentration of reduced glutathione alone does not explain the total concentration increase of nonprotein thiols after the administration of radioprotective agents. With respect to the increase in the concentration of both SH-groups and reduced glutathione, the observed reduction in glutathione reductase activity may be due to the action of the regulatory mechanism. (author)

  6. Direct UV-induced functionalization of surface hydroxy groups by thiol-ol chemistry.

    Science.gov (United States)

    Li, Linxian; Li, Junsheng; Du, Xin; Welle, Alexander; Grunze, Michael; Trapp, Oliver; Levkin, Pavel A

    2014-04-01

    A novel UV-initiated surface modification method for the direct functionalization of surface hydroxy groups with thiol-containing molecules (termed "thiol-ol" modification) is described. This method is based on the oxidative conjugation of thiols to hydroxy groups. We demonstrate that different thiol-containing molecules, such as fluorophores, thiol-terminated poly(ethylene glycol) (PEG-SH), and a cysteine-containing peptide, can be attached onto the surface of porous poly(2-hydroxyethyl methacrylate-co-ethylene dimethacrylate). Direct functionalization of other hydroxy-group-bearing surfaces, fabrication of micropatterns, and double patterning have been also demonstrated using the thiol-ol method. PMID:24595963

  7. Fatty acyl-CoA reductase

    Energy Technology Data Exchange (ETDEWEB)

    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.

  8. 21 CFR 864.7375 - Glutathione reductase assay.

    Science.gov (United States)

    2010-04-01

    ...2010-04-01 false Glutathione reductase assay. 864.7375 Section 864.7375 Food...Packages 864.7375 Glutathione reductase assay. (a) Identification. A glutathione reductase assay is a device used to determine the...

  9. Organized thiol functional groups in mesoporous core shell colloids

    International Nuclear Information System (INIS)

    The co-condensation in situ of tetraethoxysilane (TEOS) and mercaptopropyltrimethoxysilane (MPTMS) using cetyltrimethylammonium bromide (CTAB) as a template results in the synthesis of multilayered mesoporous structured SiO2 colloids with onion-like chemical environments. Thiol groups were anchored to an inner selected SiO2 porous layer in a bilayered core shell particle producing different chemical regions inside the colloidal layered structure. X-Ray Photoelectron Spectroscopy (XPS) shows a preferential anchoring of the SH groups in the double layer shell system, while porosimetry and simple chemical modifications confirm that pores are accessible. We can envision the synthesis of interesting colloidal objects with defined chemical environments with highly controlled properties. - Graphical abstract: Mesoporous core shell SiO2 colloids with organized thiol groups. Highlights: ? Double shell mesoporous silica colloids templated with CTAB. ? Sequential deposition of mesoporous SiO2 layers with different chemistries. ? XPS shows the selective functionalization of mesoporous layers with thiol groups.

  10. Distribution, adaptation and physiological meaning of thiols from vertebrate hemoglobins.

    Science.gov (United States)

    Reischl, Evaldo; Dafre, Alcir Luiz; Franco, Jeferson Luis; Wilhelm Filho, Danilo

    2007-01-01

    In the present review, the sequences of hemoglobins (Hb) of 267 adult vertebrate species belonging to eight major vertebrate taxa are examined for the presence and location of cysteinyl residues in an attempt at correlation with their ecophysiology. Essentially, all vertebrates have surface cysteinyl residues in Hb molecules whereby their thiol groups may become highly reactive. Thiol-rich Hbs may display eight or more thiols per tetramer. In vertebrates so far examined, the cysteinyl residues occur in 44 different sequence positions in alpha chains and 41 positions in beta chains. Most of them are conservatively located and occur in only a few positions in Teleostei, Aves and Mammalia, whereas they are dispersed in Amphibia. The internal cysteinyl residue alpha104 is ubiquitous in vertebrates. Residue beta93 is highly conserved in reptiles, birds and mammals. The number of cysteine residues per tetramer with solvent access varies in vertebrates, mammalians and bony fish having the lowest number of external residues, whereas nearly all external cysteine residues in Aves and Lepidosauria are of the surface crevice type. In cartilaginous fish, amphibians, Crocodylidae and fresh water turtles, a substantial portion of the solvent accessible thiols are of the totally external type. Recent evidence shows that some Hb thiol groups are highly reactive and undergo extensive and reversible S-thiolation, and that they may be implicated in interorgan redox equilibrium processes. Participation of thiol groups in nitric oxide ((*)NO) metabolism has also been proved. The evidence argues for a new physiologically relevant role for Hb via involvement in free radical and antioxidant metabolism. PMID:17368111

  11. Effect of thiol group on the curing process of alkaline developable photo-resists

    International Nuclear Information System (INIS)

    Photosensitivity of a conventional radical photo-initiator in an alkaline developable photoresist is boosted by substitution with a thiol group. Evidence is presented that the thiol group acts via chain transfer mechanism

  12. Structural features of the aldose reductase and aldehyde reductase inhibitor-binding sites.

    Science.gov (United States)

    El-Kabbani, O; Wilson, D K; Petrash, M; Quiocho, F A

    1998-09-29

    The three-dimensional structures of aldose reductase and aldehyde reductase, members of the aldo-keto reductase superfamily, are composed of similar alpha/beta TIM-barrels. However, examination of the structures reveals that the inhibitor-binding site of aldose reductase differs from that of aldehyde reductase due to the participation of non-conserved residues in its formation. This information will be useful in the design of inhibitors to prevent or delay diabetic retinopathy. A review of the structures of the inhibitor-binding sites is presented. PMID:9756955

  13. Controlling Topological Entanglement in Engineered Protein Hydrogels with a Variety of Thiol Coupling Chemistries

    OpenAIRE

    BradleyOlsen

    2014-01-01

    Topological entanglements between polymer chains are achieved in associating protein hydrogels through the synthesis of high molecular weight proteins via chain extension using a variety of thiol coupling chemistries, including disulfide formation, thiol-maleimide, thiol-bromomaleimide and thiol-ene. Coupling of cysteines via disulfide formation results in the most pronounced entanglement effect in hydrogels, while other chemistries provide versatile means of changing the extent of entanglem...

  14. 5-Furan-2yl[1,3,4]oxadiazole-2-thiol, 5-Furan-2yl-4H [1,2,4] triazole-3-thiol and Their Thiol-Thione Tautomerism

    Directory of Open Access Journals (Sweden)

    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.

  15. Surface functionalized thiol-ene waveguides for ?uorescence biosensing in micro?uidic devices

    DEFF Research Database (Denmark)

    Feidenhans'l, Nikolaj Agentoft; Lafleur, Josiane P.

    2013-01-01

    Thiol-ene polymers possess physical, optical, and chemical characteristics thatmake them ideal substrates for the fabrication of optofluidic devices. In this work, thiol-ene polymers are used to simultaneously create microfluidic channels and optical waveguides in one simple moulding step. The reactive functional groups present at the surface of the thiol-ene polymer are subsequently used for the rapid, one step, site-specific functionalization of the waveguide with biological recognition molecules. It was found that while the bulk properties and chemical surface properties of thiol-ene materials vary considerably with variations in stoichiometric composition, their optical properties remain mostly unchanged with an average refractive index value of 1.566 0.008 for thiol-ene substrates encompassing a range from 150% excess ene to 90% excess thiol. Microfluidic chips featuring thiol-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 conjugation with fluorescently labeled streptavidin. Fluorescence excitation was achieved by launching light through the thiol-ene waveguide, revealing bright fluorescent patterns along the channel/waveguide interface

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

    DEFF Research Database (Denmark)

    Feidenhans'l, Nikolaj A; Lafleur, Josiane P.

    2014-01-01

    Thiol-ene polymers possess physical, optical, and chemical characteristics that make them ideal substrates for the fabrication of optofluidic devices. In this work, thiol-ene polymers are used to simultaneously create microfluidic channels and optical waveguides in one simple moulding step. The reactive functional groups present at the surface of the thiol-ene polymer are subsequently used for the rapid, one step, site-specific functionalization of the waveguide with biological recognition molecules. It was found that while the bulk properties and chemical surface properties of thiol-ene materials vary considerably with variations in stoichiometric composition, their optical properties remain mostly unchanged with an average refractive index value of 1.566 0.008 for thiol-ene substrates encompassing a range from 150% excess ene to 90% excess thiol. Microfluidic chips featuring thiol-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 conjugation with fluorescently labeled streptavidin. Fluorescence excitation was achieved by launching light through the thiol-ene waveguide, revealing bright fluorescent patterns along the channel/waveguide interface.

  17. Applications of amino thiols as radioprotectors in mice

    International Nuclear Information System (INIS)

    Amino ethyl urea (AET) and mercapto ethyl amine or cysteamine (MEA) compounds containing - SH and - N H2 groups were tested in their capacity of increasing the survival of 9 Gy 60 Co irradiated mice (4.55 Gy/min). In spite of AET has the thiol radical covered by an urea molecule its effectiveness depends on the physiological pH in order to liberate a free thiol, forming the mercapto ethyl guanidine. The animals were injected intraperitoneally with 300 mg / kg AET or 125 mg/kg MEA dissolved in 0.85% Na Cl, 30 minutes before irradiation being the survival and weight curves registered during 30 days. peritoneal cell counts were made 2 hours, 3 and 6 days after irradiation. Both compounds showed a radioprotective action confirming data from the literature. Among possible mechanisms about the action of amino thiols as radioprotectors, it is suggested: free radicals scavengers of radical formed by water radiolysis; protection through O2 remotion; alterations in cellular protein thiols; increase of the radiation damage or alterations in the activity of the enzymes involved in the reduction of oxidative damage like GSH-peroxidase or transferase. (author)

  18. Oxidative stability of (-)-epigallocatechin gallate in the presence of thiols.

    Science.gov (United States)

    Unnadkat, Nausheel R; Elias, Ryan J

    2012-10-31

    Polyphenols are attractive ingredients due to their purported health benefits, but their addition to foods is limited by their chemical instability, as they are rapidly oxidized under many conditions. This oxidation not only compromises the potential biological activity of the phenolic compound, but can also affect the chemical stability of the surrounding food matrix. Polyphenols bearing catechol or gallate groups, when oxidized to their benzoquinone forms, are strong electrophiles capable of reacting with nucleophilic thiols via 1,4-Michael addition reactions. These reactions are known to proceed in foods during processing and storage, and can profoundly affect the quality and biological efficacy of polyphenols when they are added as functional food ingredients. The stability of (-)-epigallocatechin gallate (EGCG) in the presence of three thiol-containing species [cysteine (Cys), glutathione (GSH), 3-mercaptohexan-1-ol (3SH)] was followed under both neutral and acidic conditions. Both Cys and GSH increased the rate of EGCG oxidation at pH 4. At pH 7, only Cys was found to increase the rate of EGCG oxidation. On the basis of these results, the reactivity of thiols toward EGCG follows the trend: Cys > GSH > 3SH, which is consistent with observed thiol-quinone adduct formation rates. Contrary to the results observed for Cys and GSH, 3SH was observed to inhibit EGCG oxidation. PMID:23035942

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

    DEFF Research Database (Denmark)

    Lafleur, Josiane P.; Kwapiszewski, Radoslaw

    2013-01-01

    The suitable optical properties of thiolene polymers combined with the ease of modifying their surface for the attachment of recognition molecules make them ideal candidates in many biochip applications. This paper reports the rapid one-step photochemical surface patterning of biomolecules in microfluidic thiolene chips. This work focuses on thiolene substrates featuring an excess of thiol groups at their surface. The thiolene stoichiometric composition can be varied to precisely control the number of surface thiol groups available for surface modification up to an average surface density of 136 ! 17 SH nm"2. Biotin alkyne was patterned directly inside thiolene microchannels prior to conjugation with fluorescently labelled streptavidin. The surface bound conjugates were detected by evanescent waveinduced fluorescence (EWIF), demonstrating the success of the grafting procedure and its potential for biochip applications.

  20. Functional studies of aldo-keto reductases in Saccharomyces cerevisiae*

    OpenAIRE

    CHANG Qing; Griest, Terry A.; Harter, Theresa M.; PETRASH, J. MARK

    2006-01-01

    We utilized the budding yeast Saccharomyces cerevisiae as a model to systematically explore physiological roles for yeast and mammalian aldo-keto reductases. Six open reading frames encoding putative aldo-keto reductases were identified when the yeast genome was queried against the sequence for human aldose reductase, the prototypical mammalian aldo-keto reductase. Recombinant proteins produced from five of these yeast open reading frames demonstrated NADPH-dependent reductase activity with a...

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

    OpenAIRE

    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 ?- and ?-subunits of microbial nitrate reductase, selenate reductase, dimethyl sulfide dehydrogenase, ethylbenzene dehydrogenase, and chlorate reductase, all of which are type II m...

  2. Aldo-Keto Reductases in the Eye

    OpenAIRE

    G. Bhanuprakesh Reddy; Mark Petrash, J.; Theresa Harter; Ross Arjun Varma; Philip Ruzycki; Suryanarayana Palla; Shun Ping Huang

    2010-01-01

    Aldose reductase (AKR1B1) is an NADPH-dependent aldo-keto reductase best known as the rate-limiting enzyme of the polyol pathway. Accelerated glucose metabolism through this pathway has been implicated in diabetic cataract and retinopathy. Some human tissues contain AKR1B1 as well as AKR1B10, a closely related member of the aldo-keto reductase gene superfamily. This opens the possibility that AKR1B10 may also contribute to diabetic complications. The goal of the current study was to character...

  3. Adsorption of Dissolved Metals in the Berkeley Pit using Thiol-Functionalized Self-Assembled Monolayers on Mesoporous Supports (Thiol-SAMMS)

    International Nuclear Information System (INIS)

    The Berkeley Pit in Butte, Montana, is heavily contaminated with dissolved metals. Adsorption and extraction of these metals can be accomplished through the use of a selective adsorbent. For this research, the adsorbent used was thiol-functionalized Self-Assembled Monolayers on Mesoporous Supports (thiol-SAMMS), which was developed at Pacific Northwest National Laboratory (PNNL). Thiol-SAMMS selectively binds to numerous types of dissolved metals. The objective of this research was to evaluate the loading and kinetics of aluminum, beryllium, copper, and zinc on thiol-SAMMS. For the loading tests, a series of Berkeley Pit water to thiol-SAMMS ratios (mL:g) were tested. These ratios were 1000:1, 500:1, 100:1, and 50:1. Berkeley Pit water is acidic (pH ? 2.5). This can affect the performance of SAMMS materials. Therefore, the effect of pH was evaluated by conducting parallel series of loading tests wherein the Berkeley Pit water was neutralized before or after addition of thiol-SAMMS, and a series of kinetics tests wherein the Berkeley Pit water was neutralized before addition of thiol-SAMMS for the first test and was not neutralized for the second test. For the kinetics tests, one Berkeley Pit water to thiol-SAMMS ratio was tested, which was 2000:1. The results of the loading and kinetics tests suggest that a significant decrease in dissolved metal concentration at Berkeley Pit could be realized through neutralization of Berkeley Pit water. Thiol-SAMMS technology has y Pit water. Thiol-SAMMS technology has a limited application under the highly acidic conditions posed by the Berkeley Pit. However, thiol-SAMMS could provide a secondary remedial technique which would complete the remedial system and remove dissolved metals from the Berkeley Pit to below drinking water standards.

  4. Induction Curing of Thiol-acrylate and Thiolene Composite Systems

    Science.gov (United States)

    Ye, Sheng; Cramer, Neil B.; Stevens, Blake E.; Sani, Robert L.; Bowman, Christopher N.

    2011-01-01

    Induction curing is demonstrated as a novel type of in situ radiation curing that maintains most of the advantages of photocuring while eliminating the restriction of light accessibility. Induction curing is utilized to polymerize opaque composites comprised of thiol-acrylate and thiol-ene resins, nanoscale magnetic particles, and carbon nanotubes. Nanoscale magnetic particles are dispersed in the resin and upon exposure to the magnetic field, these particles lead to induction heating that rapidly initiates the polymerization. Heat transfer profiles and reaction kinetics of the samples are modeled during the reactions with varying induction heater power, species concentration, species type and sample thickness, and the model is compared with the experimental results. Thiol-ene polymerizations achieved full conversion between 1.5 minutes and 1 hour, depending on the field intensity and the composition, with the maximum reaction temperature decreasing from 146 87 C when the induction heater power was decreased from 8 3 kW. The polymerization reactions of the thiol-acrylate system were demonstrated to achieve full conversion between 0.6 and 30 minutes with maximum temperatures from 139 to 86 C. The experimental behavior was characterized and the temperature profile modeled for the thiol-acrylate composite comprised of sub100nm nickel particles and induction heater power in the range of 32 to 20 kW. A 9C average deviation was observed between the modeling and experimental results for the maximum temperature rise. The model also was utilized to predict reaction temperatures and kinetics for systems with varying thermal initiator concentration, initiator half-life, monomer molecular weight and temperature gradients in samples with varying thickness, thereby demonstrating that induction curing represents a designable and tunable polymerization method. Finally, induction curing was utilized to cure thiol-acrylate systems containing carbon nanotubes where 1 wt% carbon nanotubes resulted in systems where the storage modulus increased from 17.6 0.2 to 21.6 0.1 MPa and an electrical conductivity that increased from <10?7 to 0.33 0.5 S/m. PMID:21765552

  5. Periplasmic nitrate reductases: structural and spectroscopic studies

    OpenAIRE

    Javier Gonzalez, Pablo

    2006-01-01

    Nitrate reduction occurs in the cell in order to incorporate nitrogen into biomolecules(assimilatory ammonification), as the final electron acceptor when bacteria are grown in anaerobic conditions (denitrification/dissimilatory ammonification) and to eliminate energy excess generated by the cell metabolism (dissimilatory ammonification). Nitrate reductases are enzimes that catalize the conversion of nitrate to nitrite. Most nitrate reductases are mononuclear molybdenum-containing enzymes t...

  6. Functional graphene by thiol-ene click chemistry.

    Science.gov (United States)

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

  7. Stabilising cysteinyl thiol oxidation and nitrosation for proteomic analysis.

    Science.gov (United States)

    Ratnayake, Shibani; Dias, Irundika H K; Lattman, Eric; Griffiths, Helen R

    2013-10-30

    Oxidation and S-nitrosylation of cysteinyl thiols (Cys-SH) to sulfenic (Cys-SOH), sulfinic (Cys-SO2H), sulfonic acids (Cys-SO3H), disulphides and S-nitrosothiols are suggested as important post-translational modifications that can activate or deactivate the function of many proteins. Non-enzymatic post-translational modifications to cysteinyl thiols have been implicated in a wide variety of physiological and pathophysiological states but have been difficult to monitor in a physiological setting because of a lack of experimental tools. The purpose of this review is to bring together the approaches that have been developed for stably trapping cysteine either in its reduced or oxidised forms for enrichment and or subsequent mass spectrometric analysis. These tools are providing insight into potential targets for post-translational modifications to cysteine modification in vivo. This article is part of a Special Issue entitled: Special Issue: Posttranslational Protein modifications in biology and Medicine. PMID:23796488

  8. Thiol-ene/methacrylate systems for mechanical damping

    Science.gov (United States)

    McNair, Olivia; Senyurt, Askim; Wei, Huanyu; Gould, Trent; Piland, Scott; Hoyle, Charles; Savin, Daniel

    2010-03-01

    Ternary thiol-ene-methacrylate (TEMA) networks as materials for mechanical energy damping are unique to the sports world. Using a photoinitiation process, TEMA systems are formed via an initial thiol-ene step-growth mechanism along with traditional radical polymerization of acrylate and ene monomers. Final networks have two-part morphologies: acrylate homopolymer sectors imbedded in a multi-component mesh. Several (TEMA) systems have been synthesized and analyzed via thermal and mechanical probing. Initial studies on these ternary systems have shown excellent properties compared to traditional ethylene vinyl alcohol (EVA) copolymers. For example, PEMA networks exhibit glass transition temperatures 33 K higher than EVA, resulting in improved damping at room temperature. This research will help develop relationships between tan delta, glass transition and their effects on mechanical energy damping for ternary (TEMA) systems.

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

    Science.gov (United States)

    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.

  10. Measuring the Poise of Thiol/Disulfide Couples in vivo

    OpenAIRE

    Dean P. Jones; Liang, Yongliang

    2009-01-01

    The reduction potentials (Eh) for the redox couples GSH/GSSG and cysteine/cystine (Cys/CySS) in plasma are useful indicators of systemic oxidative stress and other medically relevant physiological states. This paper describes a sensitive method for determining plasma levels of GSH, GSSG, Cys and CySS used to calculate the in vivo Eh values. The method uses iodoacetate to alkylate free thiols; derivatization with dansyl chloride to fluorescently tag amino groups; and HPLC and fluorescence to s...

  11. Odorant polyfunctional thiols issued from bottle beer fermentation

    OpenAIRE

    Nizet, Sabrina; Gros, Jacques; Collin, Sonia; XII Weurman flavour research symposium

    2011-01-01

    Bottle refermentation which imparts beer effervescence and resistance against infection and oxidation is also known to improve flavor profile and stability. By this process, some stale off-flavors exhaled by aldehydes (trans-2-nonenal, 3-methylthiopropionaldehyde, 3- methylbutanal ..) are reduced into alcohols (1, 2). Unfortunately, yeast esterases can also strongly affect the beer fruity character by hydrolyzing isoamyl acetate, ethyl hexanoate and ethyl octanoate (1, 2). Thiols are known to...

  12. Chiral liquid crystalline thiols for preparation of polybutadiene diols.

    Czech Academy of Sciences Publication Activity Database

    Bubnov, Alexej; Kapar, Miroslav; Sedlkov, Zde?ka; Ilavsk, Michal

    2008-01-01

    Ro?. 35, ?. 5 (2008), s. 653-660. ISSN 0267-8292 R&D Projects: GA ?R GA202/05/0431; GA Mk OC 175; GA AV ?R IAA4112401; GA AV ?R IAA100100710 Institutional research plan: CEZ:AV0Z10100520; CEZ:AV0Z40500505 Keywords : monomers * liquid crystal * polybutadienes * chiral thiol Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 1.132, year: 2008

  13. Contrasting bonding behavior of thiol molecules on carbon fullerene structures

    International Nuclear Information System (INIS)

    We have performed semiempirical as well as ab initio density-functional theory (DFT) calculations at T=0 to analyze the equilibrium configurations and electronic properties of spheroidal C60 as well as of cylindrical armchair (5,5) and (8,8) fullerenes passivated with SCH3 and S(CH2)2CH3 thiols. Our structural results reveal that the lowest-energy configurations of the adsorbates strongly depend on their chain length and on the structure of the underlying substrate. In the low-coverage regime, both SCH3 and S(CH2)2CH3 molecules prefer to organize into a molecular cluster on one side of the C60 surface, providing thus a less protective organic coating for the carbon structure. However, with increasing the number of adsorbed thiols, a transition to a more uniform distribution is obtained, which actually takes place for six and eight adsorbed molecules when using S(CH2)2CH3 and SCH3 chains, respectively. In contrast, for the tubelike arrangements at the low-coverage regime, a quasi-one-dimensional zigzag organization of the adsorbates along the tubes is always preferred. The sulfur-fullerene bond is considerably strong and is at the origin of outward and lateral displacements of the carbon atoms, leading to the stabilization of three-membered rings on the surface (spheroidal structures) as well as to sizable nonuniform uctures) as well as to sizable nonuniform radial deformations (cylindrical configurations). The electronic spectrum of our thiol-passivated fullerenes shows strong variations in the energy difference between the highest occupied and lowest unoccupied molecular orbitals as a function of the number and distribution of adsorbed thiols, opening thus the possibility to manipulate the transport properties of these compounds by means of selective adsorption mechanisms

  14. Functional Conducting Polymers via Thiol-ene Chemistry

    OpenAIRE

    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...

  15. Activities of Bismuth Thiols against Staphylococci and Staphylococcal Biofilms

    OpenAIRE

    Domenico, Philip; Baldassarri, Lucilla; Schoch, Paul E.; Kaehler, Kristina; Sasatsu, Masanori; Cunha, Burke A.

    2001-01-01

    Indwelling medical devices are associated with infectious complications. Incorporating antimicrobials into indwelling materials may reduce bacterial colonization. Bismuth thiols are antibiofilm agents with up to 1,000-fold-greater antibacterial activity than other bismuth salts. Staphylococci are particularly sensitive, as determined by agar diffusion and broth dilution susceptibility testing. Bismuth-ethanedithiol inhibited 10 methicillin-resistant Staphylococcus epidermidis strains at 0.9 t...

  16. Functionalization of Polymer Nanoparticles by Thiol-Ene Addition

    OpenAIRE

    Korthals, Brigitte; Morant-Minana, Maria Carmen; Schmid, Marius; Mecking, Stefan

    2010-01-01

    Syndiotactic 1,2-polybutadiene nanoparticles (volume average diameter 13 nm) were functionalized in aqueous dispersion by free radical mercaptan addition. By appropriate choice of both watersoluble radical initiator and mercaptan concentration, nanoparticles with a degree of functionalization of up to 85% were prepared using 3-mercaptopropionic acid methyl ester and 3-mercaptopropionic acid. Only a minor portion of double bonds formed cycles instead of the desired thiol-ene addition products....

  17. Analytical detection of biological thiols in a microchip capillary channel.

    Science.gov (United States)

    Chand, Rohit; Jha, Sandeep Kumar; Islam, Kamrul; Han, Dawoon; Shin, Ik-Soo; Kim, Yong-Sang

    2013-02-15

    Sulfur-containing amino acids, such as cysteine and homocysteine play crucial roles in biological systems for the diagnosis of medical states. In this regard, this paper deals with separation, aliquot and detection of amino thiols on a microchip capillary electrophoresis with electrochemical detection in an inverted double Y-shaped microchannel. Unlike the conventional capillary electrophoresis, the modified microchannel design helps in storing the separated thiols in different reservoirs for further analysis, if required; and also eliminates the need of electrodes regeneration. The device was fabricated using conventional photolithographic technique which consisted of gold microelectrodes on a soda lime glass wafer and microchannels in PDMS mold. Multiple detections were performed using in-house fabricated dual potentiostat. Based on amperometric detection, cysteine and homocysteine were analyzed in 105 s and 120 s, respectively after diverting in branched channels. Repeated experiments proved the good reproducibility of the device. The device produced a linear response for both cysteine and homocysteine in electrochemical analysis. To prove the practicality of device, we also analyzed cysteine and homocysteine in real blood samples without any pre-treatment. Upon calculation, the device showed a very low limit of detection of 0.05 ?M. The modified microchip design shall find a broad range of analytical applications involving assays of thiols and other biological compounds. PMID:22940195

  18. Aminoethylation in model peptides reveals conditions for maximizing thiol specificity.

    Science.gov (United States)

    Hopkins, Christopher E; Hernandez, Gonzalo; Lee, Jonathan P; Tolan, Dean R

    2005-11-15

    Control of pH in aminoethylation reactions is critical for maintaining high selectivity towards cysteine modification. Measurement of aminoethylation rate constants by liquid chromatography mass spectrometry demonstrates reaction selectivity of cysteine>amino-terminus>histidine. Lysine and methionine were not reactive at the conditions used. For thiol modification, the acid/base property of the gamma-thialysine residue measured by NMR results in a 1.15 decrease in pK(a) (relative to a lysine residue). NMR confirms ethylene imine is the reactive intermediate for alkylation of peptide nucleophiles with bromoethylamine. Conversion of bromoethylamine into ethylene imine prior to exposure to the target thiol, provides a reagent that promotes selectivity by allowing precise control of reaction pH. Reaction selectivity plots of relative aminoethylation rates for cysteine, histidine, and N-terminus imine demonstrate increasing alkaline conditions favors thiol modification. When applied to protein modification, the conversion of bromoethylamine into ethylene imine and buffering at alkaline pH will allow optimal cysteine residue aminoethylation. PMID:16229814

  19. Thiol switches in redox regulation of chloroplasts: balancing redox state, metabolism and oxidative stress.

    Science.gov (United States)

    Dietz, Karl-Josef; Hell, Rdiger

    2015-05-01

    In photosynthesizing chloroplasts, rapidly changing energy input, intermediate generation of strong reductants as well as oxidants and multiple participating physicochemical processes and pathways, call for efficient regulation. Coupling redox information to protein function via thiol modifications offers a powerful mechanism to activate, down-regulate and coordinate interdependent processes. Efficient thiol switching of target proteins involves the thiol-disulfide redox regulatory network, which is highly elaborated in chloroplasts. This review addresses the features of this network. Its conditional function depends on specificity of reduction and oxidation reactions and pathways, thiol redox buffering, but also formation of heterogeneous milieus by microdomains, metabolite gradients and macromolecular assemblies. One major player is glutathione. Its synthesis and function is under feedback redox control. The number of thiol-controlled processes and involved thiol switched proteins is steadily increasing, e.g., in tetrapyrrole biosynthesis, plastid transcription and plastid translation. Thus chloroplasts utilize an intricate and versatile redox regulatory network for intraorganellar and retrograde communication. PMID:25741945

  20. Thiol protection in membrane protein purifications: A study with phage holins

    OpenAIRE

    Dewey, Jill S.; Struck, Douglas K.; Young, Ry

    2009-01-01

    The lambda holin, or S105, is a small cytoplasmic membrane protein that controls the timing of host lysis. Using thiol-specific reagents, we determined that the single cysteine residue within S105 was heterogeneously modified during membrane extraction and subsequent immobilized metal ion chromatography. Here we describe the use of a specific and reversible thiol reagent, 2,2? dithiodipyridine, to generate purified protein with its cysteine residues in the native thiol state. The 2,2? dit...

  1. Fabrication and bonding of thiol-ene-based microfluidic devices : Technical Note

    DEFF Research Database (Denmark)

    Sikanen, Tiina M; Lafleur, Josiane P.

    2013-01-01

    In this work, the bonding strength of microchips fabricated by thiol-ene free-radical polymerization was characterized in detail by varying the monomeric thiol/allyl composition from the stoichiometric ratio (1:1) up to 100% excess of thiol (2:1) or allyl (1:2) functional groups. Four different thiol-ene to thiol-ene bonding combinations were tested by bonding: (i) two stoichiometric layers, (ii) two layers bearing complementary excess of thiols and allyls, (iii) two layers both bearing excess of thiols, or (iv) two layers both bearing excess of allyls. The results showed that the stiffness of the cross-linked polymer plays the most crucial role regarding the bonding strength. The most rigid polymer layers were obtained by using the stoichiometric composition or an excess of allyls, and thus, the bonding combinations (i) and (iv) withstood the highest pressures (up to the cut-off value of 6 bar). On the other hand, excess of thiol monomers yielded more elastic polymer layers and thus decreased the pressure tolerance for bonding combinations (ii) and (iii). By using monomers with more thiol groups (e.g. tetrathiol versus trithiol), a higher cross-linking ratio, and thus, greater stiffness was obtained. Surface characterization by infrared spectroscopy confirmed that the changes in the monomeric thiol/allyl composition were also reflected in the surface chemistry. The flexibility of being able to bond different types of thiol-enes together allows for tuning of the surface chemistry to yield the desired properties for each application. Here, a capillary electrophoresis separation is performed to demonstrate the attractive properties of stoichiometric thiol-ene microchips.

  2. The platelet-surface thiol isomerase enzyme ERp57 modulates platelet function

    OpenAIRE

    Holbrook, L-M; Sasikumar, P; Stanley, R G; SIMMONDS, A D; BICKNELL, A B; Gibbins, J M

    2012-01-01

    Background: Thiol isomerases are a family of endoplasmic reticulum enzymes which orchestrate redox-based modifications of protein disulphide bonds. Previous studies have identified important roles for the thiol isomerases PDI and ERp5 in the regulation of normal platelet function. Aim: Recently, we demonstrated the presence of a further five thiol isomerases at the platelet surface. In this report we aim to report the role of one of these enzymes ERp57 in the regulation of platelet functi...

  3. Turn-on fluorescence probe integrated polymer nanoparticles for sensing biological thiol molecules

    OpenAIRE

    Ang, Chung Yen; Tan, Si Yu; Lu, Yunpeng; Bai, Linyi; Li, Menghuan; Li, Peizhou; Zhang, Quan; Selvan, Subramanian Tamil; Zhao, Yanli

    2014-01-01

    A turn-on thiol-responsive fluorescence probe was synthesized and integrated into polymeric nanoparticles for sensing intracellular thiols. There is a photo-induced electron transfer process in the off state of the probe, and this process is terminated upon the reaction with thiol compounds. Configuration interaction singles (CIS) calculation was performed to confirm the mechanism of this process. A series of sensing studies were carried out, showing that the probe-integrated nanopartic...

  4. A First-Principles Study of CdSe Nanoclusters Capped by Thiol Ligands

    OpenAIRE

    Wu, Shanshan; McGuigan, Michael; Tiano, Amanda L.; Wong, Stanislaus S.; Glimm, James G.

    2013-01-01

    A first-principles study of small CdnSen quantum dots (QDs) (n = 6, 13, and 33) has been performed for the study of QD-sensitized solar cells. We assessed the effects of the passivating thiol-radical ligands on the optimized structure, the energy gap, and on the absorption spectrum. The simplest thiol, methanethiol, and four other thiol type ligands, namely - cysteine (Cys), mercaptopropionic acid (MPA), and their reduced-chain analogues, were investigated. We have come to t...

  5. Characterization of chitosan thiolation and application to thiol quantification onto nanoparticle surface.

    Science.gov (United States)

    Bravo-Osuna, I; Teutonico, D; Arpicco, S; Vauthier, C; Ponchel, G

    2007-08-01

    The objective of the present work was to establish a simple and appropriated method for the quantification of thiol groups standing on the surface of core-shell nanoparticles elaborated with poly(isobutyl cyanoacrylates) and thiolated chitosan. A critical analysis of the widely used Ellman's method for the determination of thiol groups in various compounds was made. The reduced solubility of the thiolated polymer at the optimal pH of the Ellman's assay (pH 8-8.5) made difficult the accessibility of the Ellman's reagent to thiol groups in the cross-linked polymer. Furthermore, the lack of stability of the Ellman's reaction with time lead to the conclusion that the Ellman's method was of limited value to evaluate thiol groups in thiolated polymers like thiolated chitosan. An alternative and very simple thiol quantification method was developed on the bases of the classical iodine titration. The new method allowed the determination of thiol groups in small amount of samples at acidic pH, and the monitoring of the thiol determination kinetic with time. It was successfully applied to the quantification of active thiol groups on the surface of poly(isobutyl cyanoacrylates) nanoparticles coated with thiol chitosan. PMID:17592748

  6. The thiol pool in human plasma: the central contribution of albumin to redox processes.

    Science.gov (United States)

    Turell, Luca; Radi, Rafael; Alvarez, Beatriz

    2013-12-01

    The plasma compartment has particular features regarding the nature and concentration of low and high molecular weight thiols and oxidized derivatives. Plasma is relatively poor in thiol-based antioxidants; thiols are in lower concentrations than in cells and mostly oxidized. The different thiol-disulfide pairs are not in equilibrium and the steady-state concentrations of total thiols as well as reduced versus oxidized ratios are maintained by kinetic barriers, including the rates of reactions and transport processes. The single thiol of human serum albumin (HSA-SH) is the most abundant plasma thiol. It is an important target for oxidants and electrophiles due to its reactivity with a wide variety of species and its relatively high concentration. A relatively stable sulfenic (HSA-SO3H) acid can be formed in albumin exposed to oxidants. Plasma increases in mixed disulfides (HSA-SSR) or in sulfinic (HSA-SO2H) and sulfonic (HSA-SO3H) acids are associated with different pathologies and may constitute biomarkers of the antioxidant role of the albumin thiol. In this work we provide a critical review of the plasma thiol pool with a focus on human serum albumin. PMID:23747983

  7. Impact of thiol and amine functionalization on photoluminescence properties of ZnO films

    International Nuclear Information System (INIS)

    In the present study, we have investigated surface functionalization of ZnO films with dodecanethiol (Thiol) and trioctylamine (amine) by X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM), contact angle (CA) and photoluminescence (PL) measurements. The chemical bondings of thiol and amine with ZnO have been confirmed via the formation of ZnS and ZnN bonds by XPS measurements. AFM measurements on ZnO films before and after surface functionalization with thiol and amine provide evidence for the successful functionalization of thiol and amine on ZnO surfaces without any island formation. The CA measurements on ZnO films before and after surface functionalization with thiol and amine show the hydrophobic nature. PL measurements of thiol and amine functionalized ZnO show enhancements of UV emission and quenching of visible emission. The enhanced UV emissions in thiol and amine functionalized ZnO films suggest that the surface defects such as oxygen vacancies are passivated by thiol and amine functionalization. -- Highlights: ? Surface functionalization is a new approach to reduce surface dependent non-radiative process. ? Oxygen vacancies are passivated on surface functionalization. ? Thiol and amine functionalized ZnO show enhancements of UV emission

  8. The Corynebacterium glutamicum mycothiol peroxidase is a reactive oxygen species-scavenging enzyme that shows promiscuity in thiol redox control.

    Science.gov (United States)

    Pedre, Brandn; Van Molle, Inge; Villadangos, Almudena F; Wahni, Khadija; Vertommen, Didier; Turell, Luca; Erdogan, Huriye; Mateos, Luis M; Messens, Joris

    2015-06-01

    Cysteine glutathione peroxidases (CysGPxs) control oxidative stress levels by reducing hydroperoxides at the expense of cysteine thiol (-SH) oxidation, and the recovery of their peroxidatic activity is generally accomplished by thioredoxin (Trx). Corynebacterium glutamicum mycothiol peroxidase (Mpx) is a member of the CysGPx family. We discovered that its recycling is controlled by both the Trx and the mycothiol (MSH) pathway. After H2 O2 reduction, a sulfenic acid (-SOH) is formed on the peroxidatic cysteine (Cys36), which then reacts with the resolving cysteine (Cys79), forming an intramolecular disulfide (S-S), which is reduced by Trx. Alternatively, the sulfenic acid reacts with MSH and forms a mixed disulfide. Mycoredoxin 1 (Mrx1) reduces the mixed disulfide, in which Mrx1 acts in combination with MSH and mycothiol disulfide reductase as a biological relevant monothiol reducing system. Remarkably, Trx can also take over the role of Mrx1 and reduce the Mpx-MSH mixed disulfide using a dithiol mechanism. Furthermore, Mpx is important for cellular survival under H2 O2 stress, and its gene expression is clearly induced upon H2 O2 challenge. These findings add a new dimension to the redox control and the functioning of CysGPxs in general. PMID:25766783

  9. Curcumin analogs as potent aldose reductase inhibitors.

    Science.gov (United States)

    Du, Zhi-Yun; Bao, Ya-Dan; Liu, Zhong; Qiao, Wei; Ma, Lin; Huang, Zhi-Shu; Gu, Lian-Quan; Chan, Albert S C

    2006-03-01

    In the present study, curcuminoids isolated from curcuma longa were demonstrated to possess inhibitory activities on bovine lens aldose reductase. In order to find more potent aldose reductase inhibitor, curcumin analogs were synthesized and evaluated for their ability to inhibit bovine lens aldose reductase enzyme. The results indicated that the compounds with tetrahydroxyl groups, 2,6-bis(3,4-dihydroxybenzylidene)cyclohexanone (A(2)), 2,5-bis(3,4-dihydroxybenzylidene)cyclopentanone (B(2)), 1,5-bis(3,4-dihydroxyphenyl)-1,4-pentadiene-3-one (C(2)), and 3,5-bis(3,4-dihydroxybenzylidene)-4-piperidone (D(2)) showed remarkably potent inhibitory effects on aldose reductase with IC(50) of 2.9 microM, 2.6 microM, 3.4 microM, and 4.9 microM, respectively. The structure-activity relationship revealed that the curcumin analogs with ortho-dihydroxyl groups could form a more tight affinity with aldose reductase to exert more potential inhibitory activities. PMID:16528793

  10. Evaluation of Nitrate Reductase Activity in Rhizobium japonicum

    OpenAIRE

    Streeter, John G.; DeVine, Paul J.

    1983-01-01

    Nitrate reductase activity was evaluated by four approaches, using four strains of Rhizobium japonicum and 11 chlorate-resistant mutants of the four strains. It was concluded that in vitro assays with bacteria or bacteroids provide the most simple and reliable assessment of the presence or absence of nitrate reductase. Nitrite reductase activity with methyl viologen and dithionite was found, but the enzyme activity does not confound the assay of nitrate reductase.

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

    International Nuclear Information System (INIS)

    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

  12. Respiratory arsenate reductase as a bidirectional enzyme

    Science.gov (United States)

    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 [FeS] clusters, additional subunits, or constitution of the electron transfer chain. This versatility further underscores the ubiquity and antiquity of microbial arsenic metabolism.

  13. Functional Conducting Polymers via Thiol-ene Chemistry

    Directory of Open Access Journals (Sweden)

    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.

  14. Polysulfides Link H2S to Protein Thiol Oxidation

    OpenAIRE

    Greiner, Romy; Plinks, Zoltn; Bsell, Katrin; Becher, Drte; Antelmann, Haike; Nagy, Pter; Dick, Tobias P

    2013-01-01

    Aims: Hydrogen sulfide (H2S) is suggested to act as a gaseous signaling molecule in a variety of physiological processes. Its molecular mechanism of action was proposed to involve protein S-sulfhydration, that is, conversion of cysteinyl thiolates (Cys-S?) to persulfides (Cys-S-S?). A central and unresolved question is how H2Sthat is, a molecule with sulfur in its lowest possible oxidation state (?2)can lead to oxidative thiol modifications. Results: Using the lipid phosphatase PT...

  15. Role of the Dinitrogenase Reductase Arginine 101 Residue in Dinitrogenase Reductase ADP-Ribosyltransferase Binding, NAD Binding, and Cleavage

    OpenAIRE

    Ma, Yan; Ludden, Paul W.

    2001-01-01

    Dinitrogenase reductase is posttranslationally regulated by dinitrogenase reductase ADP-ribosyltransferase (DRAT) via ADP-ribosylation of the arginine 101 residue in some bacteria. Rhodospirillum rubrum strains in which the arginine 101 of dinitrogenase reductase was replaced by tyrosine, phenylalanine, or leucine were constructed by site-directed mutagenesis of the nifH gene. The strain containing the R101F form of dinitrogenase reductase retains 91%, the strain containing the R101Y form ret...

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

    Science.gov (United States)

    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

  17. Structure of Coenzyme A-Disulfide Reductase from Staphylococcus aureus at 1.54 Angstrom Resolution

    Energy Technology Data Exchange (ETDEWEB)

    Mallett,T.; Wallen, J.; Karplus, P.; Sakai, H.; Tsukihara, T.; Claiborne, A.

    2006-01-01

    Coenzyme A (CoASH) replaces glutathione as the major low molecular weight thiol in Staphylococcus aureus; it is maintained in the reduced state by coenzyme A-disulfide reductase (CoADR), a homodimeric enzyme similar to NADH peroxidase but containing a novel Cys43-SSCoA redox center. The crystal structure of S. aureus CoADR has been solved using multiwavelength anomalous dispersion data and refined at a resolution of 1.54 {angstrom}. The resulting electron density maps define the Cys43-SSCoA disulfide conformation, with Cys43-S{gamma} located at the flavin si face, 3.2 {angstrom} from FAD-C4aF, and the CoAS- moiety lying in an extended conformation within a cleft at the dimer interface. A well-ordered chloride ion is positioned adjacent to the Cys43-SSCoA disulfide and receives a hydrogen bond from Tyr361'-OH of the complementary subunit, suggesting a role for Tyr361' as an acid-base catalyst during the reduction of CoAS-disulfide. Tyr419'-OH is located 3.2 {angstrom} from Tyr361'-OH as well and, based on its conservation in known functional CoADRs, also appears to be important for activity. Identification of residues involved in recognition of the CoAS-disulfide substrate and in formation and stabilization of the Cys43-SSCoA redox center has allowed development of a CoAS-binding motif. Bioinformatics analyses indicate that CoADR enzymes are broadly distributed in both bacterial and archaeal kingdoms, suggesting an even broader significance for the CoASH/CoAS-disulfide redox system in prokaryotic thiol/disulfide homeostasis.

  18. Direct thiol-ene photocoating of polyorganosiloxane microparticles.

    Science.gov (United States)

    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 ?m. 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

  19. Thiol/Disulfide System Plays a Crucial Role in Redox Protection in the Acidophilic Iron-Oxidizing Bacterium Leptospirillum ferriphilum

    OpenAIRE

    Norambuena, Javiera; Flores, Rodrigo; Ca?rdenas, Juan P.; Quatrini, Raquel; Cha?vez, Renato; Levica?n, Gloria

    2012-01-01

    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 Fe2+ 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 respo...

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

    Directory of Open Access Journals (Sweden)

    Figueirdo 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 g/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 g 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.

  1. Effect of thiol compounds on bleomycin-induced DNA and chromosome damage in human cells.

    Science.gov (United States)

    Mira, Anabela; Gimenez, Esteban M; Bolzn, Alejandro D; Bianchi, Martha S; Lpez-Larraza, Daniel M

    2013-01-01

    Non-protein thiols are considered radioprotectors, preventing DNA damage by ionizing radiation. As bleomycin (BLM) is a radiomimetic agent it was proposed that thiols may prevent DNA damage produced by this antibiotic. However, results obtained with thiols and BLM-combined treatments in living cells are contradictory. The goal of this work was to assess the influence of five non-protein thiols of different electrical charge and chemical composition, on the DNA damage, DNA repair, chromosomal aberrations and cell killing induced by BLM. We found that, at the chromosomal level and cell killing, Glutathione, ?-Mercaptoethanol and cysteine showed a protective effect, while ditiothreitol and cysteamine increased them, whereas at the DNA level all thiols potentiated the DNA damage induced by BLM, most probably due to a reactivation of the BLM complex. PMID:23428061

  2. The influence of nonprotein thiols on DNA damage induced by bleomycin in single human cells.

    Science.gov (United States)

    Mira, Anabela; Gili, Juan A; Lopez-Larraza, Daniel M

    2013-01-01

    Nonprotein thiols are considered radioprotectors, preventing DNA damage by ionizing radiation. Because bleomycin (BLM) is a radiomimetic agent, it was proposed that thiols may prevent DNA damage produced by this antibiotic. However, results obtained with treatments combining thiols and BLM in living cells are contradictory. The goal of this study was to analyze the DNA damage induced by BLM and the influence of 3 nonprotein thiols of different electrical charges and chemical compositions at the level of single cells (comet assay). We also studied the morphological signs of apoptosis produced by BLM in these same conditions. We found that all thiols potentiated DNA damage induced by BLM, most probably by reactivating the BLM complex once it generated free radicals. Cysteamine (positive) potentiated BLM action the most, glutathione (negative) potentiated this antibiotic the least, whereas cysteine had an intermediate effect compared with the other two. PMID:24266408

  3. Alkyl thiols as a sulfur precursor for the preparation of monodisperse metal sulfide nanostructures

    International Nuclear Information System (INIS)

    We demonstrate a new method for preparing metal sulfide nanocrystals using alkyl thiols as a sulfur precursor. Alkyl thiols have many advantages for practical synthesis because they are miscible with most organic solvents and very stable under an air atmosphere. CdS nanocrystals were made with CdO and thiols with different alkyl chains such as n-octanethiol and octadecanethiol. They exhibited uniform size, highly crystalline structure and a sharp photoluminescence spectrum. Also, CdSe/CdS core/shell nanocrystals can be prepared by single injection of a mixture consisting of alkyl thiol and Se in trioctylphosphine to a Cd precursor. A reaction scheme is proposed as alkyl thiols react with the metal precursor to form stable metal thiolate intermediates during the initial period of reaction, and the thiolate decomposes slowly to form homogeneous nuclei

  4. Probing the thiol-gold planar interface by spin polarized tunneling

    International Nuclear Information System (INIS)

    Reports of induced magnetism at thiol-gold interface have generated considerable recent interest. In these studies, the sample magnetization was generally measured by superconducting quantum interference device magnetometry which has limitation in determining surface and interface magnetism. In this work, we have fabricated planar tunnel junctions incorporating a thiol-gold interface. An observed room temperature humidity effect together with low temperature inelastic electron tunneling spectroscopy confirmed the existence of a thiol-gold interface in the organic-inorganic hybrid heterostructure. Spin polarized tunneling measurements were performed to probe the spin polarization at the thiol-gold interface; however, the obtained spin polarized tunneling spectra indicate no measurable spin polarization at the thiol-gold interface

  5. Isolation and Characterization of cDNAs Encoding Leucoanthocyanidin Reductase and Anthocyanidin Reductase from Populus trichocarpa

    OpenAIRE

    Lijun WANG; Jiang, Yuanzhong; Yuan, Li; Lu, Wanxiang; Yang, Li; Karim, Abdul; Luo, Keming

    2013-01-01

    Proanthocyanidins (PAs) contribute to poplar defense mechanisms against biotic and abiotic stresses. Transcripts of PA biosynthetic genes accumulated rapidly in response to infection by the fungus Marssonina brunnea f.sp. multigermtubi, treatments of salicylic acid (SA) and wounding, resulting in PA accumulation in poplar leaves. Anthocyanidin reductase (ANR) and leucoanthocyanidin reductase (LAR) are two key enzymes of the PA biosynthesis that produce the main subunits: (+)-catechin and (?)-...

  6. Functional studies of aldo-keto reductases in Saccharomyces cerevisiae*

    Science.gov (United States)

    Chang, Qing; Griest, Terry A.; Harter, Theresa M.; Petrash, J. Mark

    2007-01-01

    SUMMARY We utilized the budding yeast Saccharomyces cerevisiae as a model to systematically explore physiological roles for yeast and mammalian aldo-keto reductases. Six open reading frames encoding putative aldo-keto reductases were identified when the yeast genome was queried against the sequence for human aldose reductase, the prototypical mammalian aldo-keto reductase. Recombinant proteins produced from five of these yeast open reading frames demonstrated NADPH-dependent reductase activity with a variety of aldehyde and ketone substrates. A triple aldo-keto reductase null mutant strain demonstrated a glucose-dependent heat shock phenotype which could be rescued by ectopic expression of human aldose reductase. Catalytically-inactive mutants of human or yeast aldo-keto reductases failed to effect a rescue of the heat shock phenotype, suggesting that the phenotype results from either an accumulation of one or more unmetabolized aldo-keto reductase substrates or a synthetic deficiency of aldo-keto reductase products generated in response to heat shock stress. These results suggest that multiple aldo-keto reductases fulfill functionally redundant roles in the stress response in yeast. PMID:17140678

  7. Functional studies of aldo-keto reductases in Saccharomyces cerevisiae.

    Science.gov (United States)

    Chang, Qing; Griest, Terry A; Harter, Theresa M; Petrash, J Mark

    2007-03-01

    We utilized the budding yeast Saccharomyces cerevisiae as a model to systematically explore physiological roles for yeast and mammalian aldo-keto reductases. Six open reading frames encoding putative aldo-keto reductases were identified when the yeast genome was queried against the sequence for human aldose reductase, the prototypical mammalian aldo-keto reductase. Recombinant proteins produced from five of these yeast open reading frames demonstrated NADPH-dependent reductase activity with a variety of aldehyde and ketone substrates. A triple aldo-keto reductase null mutant strain demonstrated a glucose-dependent heat shock phenotype which could be rescued by ectopic expression of human aldose reductase. Catalytically-inactive mutants of human or yeast aldo-keto reductases failed to effect a rescue of the heat shock phenotype, suggesting that the phenotype results from either an accumulation of one or more unmetabolized aldo-keto reductase substrates or a synthetic deficiency of aldo-keto reductase products generated in response to heat shock stress. These results suggest that multiple aldo-keto reductases fulfill functionally redundant roles in the stress response in yeast. PMID:17140678

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

    Directory of Open Access Journals (Sweden)

    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 , 0.732 and 0.728 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.

  9. Thiol synthesis and arsenic hyperaccumulation in Pteris vittata (Chinese brake fern)

    International Nuclear Information System (INIS)

    Pteris vittata (Chinese brake fern) has potential for phytoremediation of As-contaminated sites. In this study, the synthesis of total thiols and acid-soluble thiols in P. vittata was investigated under arsenic exposure. The strong and positive correlation between As concentration and acid-soluble thiols in plant leaflets suggests that acid-soluble thiols may play a role in As detoxification. A major As-induced thiol was purified and characterized. A molecular ion (M+1) of 540 m/z suggests that the thiol was a phytochelatin (PC) with two base units (PC2). However, the ratios of acid-soluble thiols to As in leaflets exposed to As ranged from 0.012 to 0.026, suggesting that only a very small part of As is complexed by PC2. PCs could play a minor detoxification role in this hyperaccumulator. A PC-independent mechanism appears to be mainly involved in As tolerance, while PC-dependent detoxification seems to be a supplement

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

    International Nuclear Information System (INIS)

    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 ?2?m 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

  11. Preparation and comparison between different thiol-protected Au nanoparticles

    International Nuclear Information System (INIS)

    We report on the synthesis of monolayer-protected gold cluster (MPCs) in aquatic phase under different conditions. Three thiols, i.e., 2-mercaptoethanesulfonic acid (MPS), 3-mercaptopropionic acid (MPA), and cystamine (CYS) have been used as the protecting monolayer. Furthermore, we studied the effect of varying the ratio between the concentrations of the protecting layer, the gold ions, and the reducing agent in the preparation solution. Comparison was based on measuring the average diameter, stability, and size distribution of the MPCs. Best results were obtained for MPS-based MPCs and that the optimum molar concentration ratio between the protecting layer, the gold ions, and the reducing agent was 1:1:2.

  12. Mercury binding sites in thiol-functionalized mesostructured silica.

    Science.gov (United States)

    Billinge, Simon J L; McKimmy, Emily J; Shatnawi, Mouath; Kim, HyunJeong; Petkov, Valeri; Wermeille, Didier; Pinnavaia, Thomas J

    2005-06-15

    Thiol-functionalized mesostructured silica with anhydrous compositions of (SiO(2))(1)(-)(x)()(LSiO(1.5))(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 < or = 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(SBu(t))(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. PMID:15941284

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  14. Oxidative desorption of thiols as a route to controlled formation of binary self assembled monolayer surfaces

    International Nuclear Information System (INIS)

    Binary self-assembled monolayers have been prepared by an alternative route to the conventional approach where a surface is exposed to a solution of both thiols at open circuit. A particular composition can be prepared with relative ease and the method is particularly useful for binary monolayers comprising two thiols with very different chain lengths since the problem of the longer chain thiol displacing the shorter over time is avoided. This is achieved by first preparing a full monolayer of the longer thiol, then oxidative desorption is carried out using cyclic voltammetry to remove a known fraction (as measured from the charge associated with gold oxide reduction) of the original SAM. Experiments with several thiols of chain lengths from 9 to 16 carbon atoms show that a considerable degree of control may be exerted over the rate at which the thiol is removed, using factors such as the potential limit, scan rate, pH and electrolyte polarity. The amount of thiol removed can thus also be controlled in a similar fashion. Once the oxidative desorption has taken place to the required degree, the vacancies created can be filled by exposure to a solution of a second thiol, differing in either chain length, termination or both. Deposition of thiols was carried out both under potential control (for longer chains) and by simple adsorption (for shorter chains). Several binary SAMs were prepared using this method and characterised using simple cyclic voltammetry with redox probes and selective reductive desorption. This method of preparation of binary SAMS may provide an alternative to both conventional methods using solution exposure and to methods that employ reductive desorption to expose selected facets on a polycrystalline surface. It may also be used to create partially blocked electrodes

  15. Mutagenesis of the redox-active disulfide in mercuric ion reductase: Catalysis by mutant enzymes restricted to flavin redox chemistry

    International Nuclear Information System (INIS)

    Mercuric reductase, a flavoenzyme that possesses a redox-active cystine, Cys135Cys140, catalyzes the reduction of Hg(II) to Hg(0) by NADPH. As a probe of mechanism, the authors have constructed mutants lacking a redox-active disulfide by eliminating Cys135 (Ala135Cys140), Cys14 (Cys135Ala140), or both (Ala135Ala140). Additionally, they have made double mutants that lack Cys135 (Ala135Cys139Cys140) or Cys140 (Cys135Cys139Ala140) but introduce a new Cys in place of Gly139 with the aim of constructing dithiol pairs in the active site that do not form a redox-active disulfide. The resulting mutant enzymes all lack redox-active disulfides and are hence restricted to FAD/FADH2 redox chemistry. Each mutant enzyme possesses unique physical and spectroscopic properties that reflect subtle differences in the FAD microenvironment. Preliminary evidence for the Ala135Cys139Cys14 mutant enzyme suggests that this protein forms a disulfide between the two adjacent Cys residues. Hg(II) titration experiments that correlate the extent of charge-transfer quenching with Hg(II) binding indicate that the Ala135Cys140 protein binds Hg(II) with substantially less avidity than does the wild-type enzyme. All mutant mercuric the wild-type enzyme. All mutant mercuric reductases catalyze transhydrogenation and oxygen reduction reactions through obligatory reduced flavin intermediates at rates comparable to or greater than that of the wild-type enzyme. In multiple-turnover assays which monitored the production of Hg(0), two of the mutant enzymes were observed to proceed through at least 30 turnovers at rates ca. 1000-fold slower than that of wild-type mercuric reductase. They conclude that the Cys135 and Cys140 thiols serve as Hg(II) ligands that orient the Hg(II) for subsequent reduction by a reduced flavin intermediate

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

    Directory of Open Access Journals (Sweden)

    Naphtali A. OConnor

    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. The intracellular redox stress caused by hexavalent chromium is selective for proteins that have key roles in cell survival and thiol redox control

    International Nuclear Information System (INIS)

    Hexavalent chromium [Cr(VI)] compounds (e.g. chromates) are strong oxidants that readily enter cells where they are reduced to reactive Cr intermediates that can directly oxidize some cell components and can promote the generation of reactive oxygen and nitrogen species. Inhalation is a major route of exposure which directly exposes the bronchial epithelium. Previous studies with non-cancerous human bronchial epithelial cells (BEAS-2B) demonstrated that Cr(VI) treatment results in the irreversible inhibition of thioredoxin reductase (TrxR) and the oxidation of thioredoxins (Trx) and peroxiredoxins (Prx). The mitochondrial Trx/Prx system is somewhat more sensitive to Cr(VI) than the cytosolic Trx/Prx system, and other redox-sensitive mitochondrial functions are subsequently affected including electron transport complexes I and II. Studies reported here show that Cr(VI) does not cause indiscriminant thiol oxidation, and that the Trx/Prx system is among the most sensitive of cellular protein thiols. Trx/Prx oxidation is not unique to BEAS-2B cells, as it was also observed in primary human bronchial epithelial cells. Increasing the intracellular levels of ascorbate, an endogenous Cr(VI) reductant, did not alter the effects on TrxR, Trx, or Prx. The peroxynitrite scavenger MnTBAP did not protect TrxR, Trx, Prx, or the electron transport chain from the effects of Cr(VI), implying that peroxynitrite is not required for these effects. Nitration of tyrosine residues of TrxR was not observed following Cr(VI) treatment, further ruling out peroxynitrite as a significant contributor to the irreversible inhibition of TrxR. Cr(VI) treatments that disrupt the TrxR/Trx/Prx system did not cause detectable mitochondrial DNA damage. Overall, the redox stress that results from Cr(VI) exposure shows selectivity for key proteins which are known to be important for redox signaling, antioxidant defense, and cell survival.

  18. Regulation of human methylenetetrahydrofolate reductase by phosphorylation

    OpenAIRE

    Yamada, Kazuhiro; Strahler, John R.; Andrews, Philip C; Matthews, Rowena G

    2005-01-01

    Methylenetetrahydrofolate reductase (MTHFR) catalyzes the reduction of methylenetetrahydrofolate to methyltetrahydrofolate, the methyl donor for the conversion of homocysteine to methionine. Regulation of MTHFR activity is crucial for maintaining cellular concentrations of methionine and S-adenosylmethionine (AdoMet). Purified recombinant human MTHFR expressed in insect cells is multiply phosphorylated on an N-terminal extension of the protein that contains a highly conserved serine-rich regi...

  19. Methylenetetrahydrofolate reductase polymorphism and pre-eclampsia.

    OpenAIRE

    Sohda, S.; Arinami, T.; Hamada, H.; Yamada, N.; Hamaguchi, H.; Kubo, T.

    1997-01-01

    A common missense mutation in the methylenetetrahydrofolate reductase (MTHFR) gene, a C to T substitution at nucleotide 677, is responsible for reduced MTHFR activity and associated with modestly increased plasma homocysteine concentrations. Since underlying maternal vascular disease increases the risk of pre-eclampsia, we had the working hypothesis that pre-eclampsia patients would have an increased T677 allele frequency compared with controls. The MTHFR genotypes were determined in 67 pre-e...

  20. Studies toward novel peptidomimetic inhibitors of thioredoxin-thioredoxin reductase system.

    Science.gov (United States)

    K?ossowski, Szymon; Muchowicz, Angelika; Firczuk, Ma?gorzata; Swiech, Marta; Redzej, Adam; Golab, Jakub; Ostaszewski, Ryszard

    2012-01-12

    Thioredoxins (Trx) are ubiquitous multifunctional low-molecular weight proteins that together with thioredoxin reductases (TrxR) participate in the maintenance of protein thiol homeostasis in NADPH-dependent reactions. An increasing number of data reveal that the Trx-TrxR system is an attractive target for anticancer therapies. In this work, we have elaborated a new and simple synthetic approach employing Ugi reaction to synthesize several new inhibitors of this system. The influence of various electrophilic fragments of this new class of compounds on the inhibition of the Trx-TrxR system was evaluated. As a result, a new compound 19a (SK053), which inhibits the activity of the Trx-TrxR system and exhibits antitumor activity, was obtained. Biologic analyses revealed that 19a inhibits induction of NF-?B and AP-1 and decreases H(2)O(2) scavenging capacity in tumor cells. Altogether, we show that 19a is a novel potential antitumor peptidomimetic inhibitor that can be used as a starting compound for further optimization. PMID:22128876

  1. Characterization of the chlorate reductase from Pseudomonas chloritidismutans

    OpenAIRE

    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. SCREENING OF HMG CO A REDUCTASE INHIBITOR PRODUCING MARINE ACTINOMYCETES

    OpenAIRE

    Srinu, Phani Bhushan

    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...

  3. Distribution and immunological characterization of microbial aldehyde reductases.

    Science.gov (United States)

    Kataoka, M; Shimizu, S; Yamada, H

    1992-01-01

    The distribution of microbial aldo-keto reductases was examined and their immunochemical characterization was performed. p-Nitrobenzaldehyde, pyridine-3-aldehyde and ethyl 4-chloro-3-oxobutanoate reductase activities were found to be widely distributed in a variety of microorganisms. In immunodiffusion studies, most yeasts belonging to the genera Sporobolomyces, Sporidiobolus and Rhodotorula formed precipitin bands with anti-Sporobolomyces salmonicolor aldehyde reductase serum. Furthermore, the results of immunotitration experiments suggested that Sporobolomyces salmonicolor AKU 4429 contains other enzyme(s) which can reduce p-nitrobenzaldehyde, pyridine-3-aldehyde and/or ethyl 4-chloro-3-oxobutanoate, and which are inactivated by anti-Sporobolomyces salmonicolor aldehyde reductase serum. PMID:1510561

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

    Directory of Open Access Journals (Sweden)

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

    2013-09-01

    Full Text Available 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 produced HMG Co A reductase inhibitor

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

    Directory of Open Access Journals (Sweden)

    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.

  6. Controlling Topological Entanglement in Engineered Protein Hydrogels with a Variety of Thiol Coupling Chemistries

    Directory of Open Access Journals (Sweden)

    BradleyOlsen

    2014-05-01

    Full Text Available Topological entanglements between polymer chains are achieved in associating protein hydrogels through the synthesis of high molecular weight proteins via chain extension using a variety of thiol coupling chemistries, including disulfide formation, thiol-maleimide, thiol-bromomaleimide and thiol-ene. Coupling of cysteines via disulfide formation results in the most pronounced entanglement effect in hydrogels, while other chemistries provide versatile means of changing the extent of entanglement, achieving faster chain extension, and providing a facile method of controlling the network hierarchy and incorporating stimuli responsivities. The addition of trifunctional coupling agents causes incomplete crosslinking and introduces branching architecture to the protein molecules. The high-frequency plateau modulus and the entanglement plateau modulus can be tuned by changing the ratio of difunctional chain extender to the trifunctional branching unit. Therefore, these chain extension reactions show promise in delicately controlling the relaxation and mechanical properties of engineered protein hydrogels in ways that complement their design through genetic engineering.

  7. Interactions of oxaliplatin with the cytoplasmic thiol containing ligand glutathione.

    Science.gov (United States)

    Shoeib, Tamer; Sharp, Barry L

    2012-12-01

    The complexation of the Pt-based anti-cancer drug oxaliplatin (OxPt) with biological ligands other than DNA is believed to be a major cellular sink for the drug reducing its therapeutic effect and acting as a potential cause of toxicity. In this paper, linear ion trap electrospray ionization mass spectrometry was employed to study the interaction of oxaliplatin with the cytoplasmic thiol containing tripeptide ligand ?-l-glutamyl-l-cysteinyl-glycine (GSH) this being the most abundant low-molecular-weight thiol containing molecule in human cells. Evidence of protonated dimers and multimers of oxaliplatin, protonated multimers of glutathione as well as several different combinations of these protonated species is presented. Most species observed were unambiguously assigned and compared to their theoretical isotopic patterns. Fragmentation of the collisionally-activated protonated complex of glutathione with oxaliplatin [GSH + OxPt + H](+) resulted in the formation of several species. No experimental evidence for [GSH + H](+) formation from the [OxPt + GSH + H](+) precursor was observed. Density functional calculations at B3LYP/LANL2DZ were used to obtain structural information and relative free energies of different isomers of the observed precursor [OxPt + GSH + H](+) both in the gas phase and in solution as well as to probe its fragmentation, highlighting mechanisms that account for all the experimental results. Data are presented to show several binding modes between electron rich sites such as S, N, O centers of GSH and the Pt metal of oxaliplatin. Calculations were also employed to obtain proton affinities and free energies of key reactions. The proton affinities of GSH and OxPt at 298 K were calculated to be 255.3 and 233.5 kcal mol(-1) respectively. The enthalpy and free energy, based on the most thermodynamically favored conformers of the reactants and products, for the addition reaction [Pt(dach)](2+) + [GSH - H](-) ? [GSH - H + Pt(dach)](+) (where dach represents diaminocyclohexane) in the gas phase at 298 K were determined to be -311.3 and -290.2 kcal mol(-1) respectively. Similarly, the enthalpy of the gas phase reaction [Pt(dach)](2+) + GSH ? [GSH + Pt(dach)](2+) at 298 K was determined to be -169.2 kcal mol(-1). PMID:23128601

  8. Yeast genes required for conversion of grape precursors to varietal thiols in wine.

    Science.gov (United States)

    Santiago, Margarita; Gardner, Richard C

    2015-08-01

    Three varietal thiols are important for the tropical fruit aromas of Sauvignon blanc: 4-mercapto-4-methylpentan-2-one (4MMP), 3-mercaptohexanol (3MH) and its acetylated derivative 3-mercaptohexyl acetate (3MHA). These thiols are produced by yeast during fermentation from precursors in grape juice. Here we identify genes in Saccharomyces cerevisiae that are required for the transport and cleavage of two thiol precursors: cysteine-4MMP and glutathione-3MH. A full-length copy of IRC7 is absolutely required for the cleavage of both precursors in the tested strains; the deleted form of the enzyme found in most yeast strains is incapable of converting these compounds into detectable thiols. By using strains that overexpress full-length IRC7, we further show that the glutathione transporter OPT1 and the transpeptidase CIS2 are also required for conversion of glut-3MH to its varietal thiol. No transporter for cys-4MMP was identified: a strain deleted for all nine known cysteine transport genes was still capable of converting cys-4MMP to its varietal thiol, and was also able to take up cysteine at high concentrations. Based on these results, we conclude that cysteine and glutathione precursors make a relatively minor contribution to 3MH production from most grape juices. PMID:26038341

  9. XANES spectroscopy studies of Cr(VI) reduction by thiols in organosulfur compounds and humic substances.

    Science.gov (United States)

    Szulczewski, M D; Helmke, P A; Bleam, W F

    2001-03-15

    The reduction of Cr(VI) by the thiol-containing compounds cysteine and glutathione and by reduced sulfur in humic substances was monitored with sulfur and chromium X-ray absorption near-edge structure (XANES) spectroscopy in chromium-contaminated soils. Reaction of humic acids with Cr(VI) resulted in a reduction of the peak area of thiols and an increase in the peak area of disulfides in the sulfur XANES spectra. Analysis of the sulfur XANES spectra in various systems indicates that the reduction of Cr(VI) by humic substances involves a thiol/disulfide redox couple analogous to that of the Cr(VI) reduction by the simple thiol-containing compounds cysteine and glutathione. A fraction of the hexavalent chromium present in industrially-contaminated soils was not reducible by thiols. Reduction of Cr(VI) to Cr(III) in soils by thiols has little effect on the pH of the system in contrast to the pH decrease resulting from reduction by Fe(II). PMID:11347925

  10. "Turn-on" fluorescence probe integrated polymer nanoparticles for sensing biological thiol molecules.

    Science.gov (United States)

    Ang, Chung Yen; Tan, Si Yu; Lu, Yunpeng; Bai, Linyi; Li, Menghuan; Li, Peizhou; Zhang, Quan; Selvan, Subramanian Tamil; Zhao, Yanli

    2014-01-01

    A "turn-on" thiol-responsive fluorescence probe was synthesized and integrated into polymeric nanoparticles for sensing intracellular thiols. There is a photo-induced electron transfer process in the off state of the probe, and this process is terminated upon the reaction with thiol compounds. Configuration interaction singles (CIS) calculation was performed to confirm the mechanism of this process. A series of sensing studies were carried out, showing that the probe-integrated nanoparticles were highly selective towards biological thiol compounds over non-thiolated amino acids. Kinetic studies were also performed to investigate the relative reaction rate between the probe and the thiolated amino acids. Subsequently, the Gibbs free energy of the reactions was explored by means of the electrochemical method. Finally, the detection system was employed for sensing intracellular thiols in cancer cells, and the sensing selectivity could be further enhanced with the use of a cancer cell-targeting ligand in the nanoparticles. This development paves a path for the sensing and detection of biological thiols, serving as a potential diagnostic tool in the future. PMID:25394758

  11. Glutathione-S-Transferase and Thiol Stress in patients with acute renal failure

    Directory of Open Access Journals (Sweden)

    Mungli Prakash

    2010-07-01

    Full Text Available Introduction: Tubular damage is common finding in acute renal failure (ARF. Various etiologies have been put forth to explain the tubular damage in ARF, one important mechanism among them is oxidative damage to renal tubules. Several biomolecules including low-molecular weight peptides and enzymes in urine have been proposed as early markers of renal failure. Current study has been undertaken to study the thiol stress and glutathione-S-transferase (GST levels in ARF patients. Method: 58 ARF patients and 55 healthy controls were selected based on inclusion and exclusion criteria. Serum thiols, GST, malanoldehyde (MDA and urine thiols were determined by spectrophotometer based methods. Results: Serum thiols and urine thiols were significantly decreased (p<0.0001, and serum GST and MDA levels were significantly increased (p<0.0001 in ARF patients compared to healthy controls. Serum GST and MDA correlated positively in ARF cases (r2 = 0.6938, p<0.0001. Conclusion: There is significant thiol stress and increased lipid peroxidation in ARF patients which leads to tubular cell membrane damage and release of GST into blood stream and into urine. This may be possible mechanism for the increased presence of GST in urine (enzymuria found in other studies.

  12. Adlayers of dimannoside thiols on gold: surface chemical analysis.

    Science.gov (United States)

    Dietrich, Paul M; Horlacher, Tim; Girard-Lauriault, Pierre-Luc; Gross, Thomas; Lippitz, Andreas; Min, Hyegeun; Wirth, Thomas; Castelli, Riccardo; Seeberger, Peter H; Unger, Wolfgang E S

    2011-04-19

    Carbohydrate films on gold based on dimannoside thiols (DMT) were prepared, and a complementary surface chemical analysis was performed in detail by X-ray photoelectron spectroscopy (XPS), time-of-flight secondary ion mass spectrometry (ToF-SIMS), near-edge X-ray absorption fine structure (NEXAFS), FT-IR, and contact angle measurements in order to verify formation of ?-carbohydrate-functionalized alkylthiol films. XPS (C 1s, O 1s, and S 2p) reveals information on carbohydrate specific alkoxy (C-O) and acetal moieties (O-C-O) as well as thiolate species attached to gold. Angle-resolved synchrotron XPS was used for chemical speciation at ultimate surface sensitivity. Angle-resolved XPS analysis suggests the presence of an excess top layer composed of unbound sulfur components combined with alkyl moieties. Further support for DMT attachment on Au is given by ToF-SIMS and FT-IR analysis. Carbon and oxygen K-edge NEXAFS spectra were interpreted by applying the building block model supported by comparison to data of 1-undecanethiol, poly(vinyl alcohol), and polyoxymethylene. No linear dichroism effect was observed in the angle-resolved C K-edge NEXAFS. PMID:21417247

  13. Surface Eroding, Semicrystalline Polyanhydrides via Thiol-Ene "Click" Photopolymerization.

    Science.gov (United States)

    Poetz, Katie L; Mohammed, Halimatu S; Shipp, Devon A

    2015-05-11

    Surface eroding and semicrystalline polyanhydrides, with tunable erosion times and drug delivery pharmacokinetics largely dictated by erosion, are produced easily with thiol-ene "click" polymerization. This strategy yields both linear and cross-linked network polyanhydrides that are readily and fully cured within minutes using photoinitiation, can contain up to 60% crystallinity, and have tensile moduli up to 25 MPa for the compositions studied. Since they readily undergo hydrolysis and exhibit the oft-preferred surface erosion mechanism, they may be particularly useful in drug delivery applications. The polyanhydrides were degraded under pseudophysiological conditions and cylindrical samples (10 mm diameter 5 mm height) were completely degraded within ?10 days, with the mass-time profile being linear for much of this time after a ?24 h induction period. Drug release studies, using lidocaine as a model, showed pharmacokinetics that displayed a muted burst release in the early stages of erosion, but then a delayed release profile that is closely correlated to the erosion kinetics. Furthermore, cytotoxicity studies of the linear and cross-linked semicrystalline polyanhydrides, and degradation products, against fibroblast cells indicate that the materials have good cytocompatibility. Overall, cells treated with up to 2500 mg/L of the semicrystalline polyanhydrides and degradation products show >90% human dermal fibroblast adult (HDFa) cell viability indicative of good cytocompatibility. PMID:25867183

  14. Tunable thiolepoxy shape memory polymer foams

    Science.gov (United States)

    Ellson, Gregory; Di Prima, Matthew; Ware, Taylor; Tang, Xiling; Voit, Walter

    2015-05-01

    Shape memory polymers (SMPs) are uniquely suited to a number of applications due to their shape storage and recovery abilities and the wide range of available chemistries. However, many of the desired performance properties are tied to the polymer chemistry which can make optimization difficult. The use of foaming techniques is one way to tune mechanical response of an SMP without changing the polymer chemistry. In this work, a novel thiolepoxy SMP was foamed using glass microspheres (40 and 50% by volume Q-Cel 6019), using expandable polymer microspheres (1% 930 DU 120), and by a chemical blowing agent (1% XOP-341). Each approach created SMP foam with a differing density and microstructure from the others. Thermal and thermomechanical analysis was performed to observe the behavioral difference between the foaming techniques and to confirm that the glass transition (Tg) was relatively unchanged near 50 C while the glassy modulus varied from 19.1 to 345 MPa and the rubbery modulus varied from 0.04 to 2.2 MPa. The compressive behavior of the foams was characterized through static compression testing at different temperatures, and cyclic compression testing at Tg. Constrained shape recovery testing showed a range of peak recovery stress from 5 MPa for the syntactic Q-Cel foams to ?0.1 MPa for the chemically blown XOP-341 foam. These results showed that multiple foaming approaches can be used with a novel SMP to vary the mechanical response independent of Tg and polymer chemistry.

  15. Modular degradable hydrogels based on thiol-reactive oxanorbornadiene linkers.

    Science.gov (United States)

    Higginson, Cody J; Kim, Seung Yeon; Pelez-Fernndez, Miguel; Fernndez-Nieves, Alberto; Finn, M G

    2015-04-22

    Oxanorbornadiene dicarboxylate (OND) reagents are potent Michael acceptors, the adducts of which undergo fragmentation by retro-Diels-Alder reaction at rates that vary with the substitution pattern on the OND moiety. Rapid conjugate addition between thiol-terminated tetravalent PEG and multivalent ONDs yielded self-supporting hydrogels within 1 min at physiological temperature and pH. Erosion of representative hydrogel formulations occurred with predictable and pH-independent rates on the order of minutes to weeks. These materials could be made non-degradable by epoxidation of the OND linkers without slowing gelation. Hydrogels prepared with OND linkers of equal valence had comparable physical properties, as determined by equilibrium swelling behavior, indicating similar internal network structure. Diffusion and release of entrained cargo varied with both the rate of degradation of PEG-OND hydrogels and the hydrodynamic radius of the entrained species. These results highlight the utility of OND linkers in the preparation of degradable network materials with potential applications in sustained release. PMID:25871459

  16. A new reagent for stable thiol-specific conjugation.

    Science.gov (United States)

    Badescu, George; Bryant, Penny; Swierkosz, Julia; Khayrzad, Farzad; Pawlisz, Estera; Farys, Monika; Cong, Yuehua; Muroni, Maurizio; Rumpf, Norbert; Brocchini, Steve; Godwin, Antony

    2014-03-19

    Many clinically used protein therapeutics are modified to increase their efficacy. Example modifications include the conjugation of cytotoxic drugs to monoclonal antibodies or poly(ethylene glycol) (PEG) to proteins and peptides. Monothiol-specific conjugation can be efficient and is often accomplished using maleimide-based reagents. However, maleimide derived conjugates are known to be susceptible to exchange reactions with endogenous proteins. To address this limitation in stability, we have developed PEG-mono-sulfone 3, which is a latently reactive, monothiol selective conjugation reagent. Comparative reactions with PEG-maleimide and other common thiol-selective PEGylation reagents including vinyl sulfone, acrylate, and halo-acetamides show that PEG-mono-sulfone 3 undergoes more efficient conjugation under mild reaction conditions. Due to the latent reactivity of PEG-mono-sulfone 3, its reactivity can be tailored and, once conjugated, the electron-withdrawing ketone is easily reduced under mild conditions to prevent undesirable deconjugation and exchange reactions from occurring. We describe a comparative stability study demonstrating a PEG-maleimide conjugate to be more labile to deconjugation than the corresponding conjugate obtained using PEG-mono-sulfone 3. PMID:24512057

  17. Human carbonyl reductase 4 is a mitochondrial NADPH-dependent quinone reductase.

    Science.gov (United States)

    Endo, Satoshi; Matsunaga, Toshiyuki; Kitade, Yukio; Ohno, Satoshi; Tajima, Kazuo; El-Kabbani, Ossama; Hara, Akira

    2008-12-26

    A protein encoded in the gene Cbr4 on human chromosome 4q32.3 belongs to the short-chain dehydrogenase/reductase family. Contrary to the functional annotation as carbonyl reductase 4 (CBR4), we show that the recombinant tetrameric protein, composed of 25-kDa subunits, exhibits NADPH-dependent reductase activity for o- and p-quinones, but not for other aldehydes and ketones. The enzyme was insensitive to dicumarol and quercetin, potent inhibitors of cytosolic quinone reductases. The 25-kDa CBR4 was detected in human liver, kidney and cell lines on Western blotting using anti-CBR4 antibodies. The overexpression of CBR4 in bovine endothelial cells reveals that the enzyme has a non-cleavable mitochondrial targeting signal. We further demonstrate that the in vitro quinone reduction by CBR4 generates superoxide through the redox cycling, and suggest that the enzyme may be involved in the induction of apoptosis by cytotoxic 9,10-phenanthrenequinone. PMID:19000905

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

    International Nuclear Information System (INIS)

    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

  19. A functional nitric oxide reductase model

    OpenAIRE

    Collman, James P.; Yang, Ying; Dey, Abhishek; Decre?au, Richard A.; Ghosh, Somdatta; Ohta, Takehiro; Solomon, Edward I.

    2008-01-01

    A functional heme/nonheme nitric oxide reductase (NOR) model is presented. The fully reduced diiron compound reacts with two equivalents of NO leading to the formation of one equivalent of N2O and the bis-ferric product. NO binds to both heme Fe and nonheme Fe complexes forming individual ferrous nitrosyl species. The mixed-valence species with an oxidized heme and a reduced nonheme FeB does not show NO reduction activity. These results are consistent with a so-called trans mechanism fo...

  20. Biliverdin reductase: a target for cancer therapy?

    OpenAIRE

    Gibbs, Peter E. M; Miralem, Tihomir; Maines, Mahin. D.

    2015-01-01

    Biliverdin reductase (BVR) is a multifunctional protein that is the primary source of the potent antioxidant, bilirubin. BVR regulates activities/functions in the insulin/IGF-1/IRK/PI3K/MAPK pathways. Activation of certain kinases in these pathways is/are hallmark(s) of cancerous cells. The protein is a scaffold/bridge and intracellular transporter of kinases that regulate growth and proliferation of cells, including PKCs, ERK and Akt, and their targets including NF-?B, Elk1, HO-1, and iNOS. ...

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  2. The mechanism of Hg2+ toxicity in cultured human oral fibroblasts: the involvement of cellular thiols.

    Science.gov (United States)

    Liu, Y; Cotgreave, I; Atzori, L; Grafstrm, R C

    1992-11-30

    To study amalgam-related toxicity in a primary target cell type, human oral fibroblasts were grown in a low-serum medium containing 1.25% fetal bovine serum and exposed to Hg2+, a corrosion product of amalgam. A 1-h exposure to various concentrations of Hg2+ resulted in a dose-dependent loss of colony forming efficiency. Removal of the low-molecular-weight thiol cysteine from the medium increased the toxicity of Hg2+ almost 50-fold in comparison with complete medium or medium without fetal bovine serum. Accordingly, fetal bovine serum was not found to contain detectable levels of low-molecular-weight thiols. The levels of cellular free protein thiols were shown to be depleted Hg2+ at significantly lower concentrations of the metal ion than those required to decrease the levels of the major cellular low-molecular weight thiol glutathione. These decreases were dependent on the exposure conditions, i.e. the presence of serum and thiols, in a manner similar to the effect on colony forming efficiency. Other functions commonly related to cell viability, including the accumulation of the vital dye neutral red, the cytosolic retention of deoxyglucose and the mitochondrial reduction of tetrazolium were also inhibited by Hg2+, albeit at higher concentrations. Finally, the depletion of cellular glutathione, by pre-exposure of the cells to the glutathione synthesis inhibitor buthionine sulfoximine, somewhat increased the toxicity of Hg2+ and potentiated the depletion of protein thiols. Taken together, the toxicity of Hg2+ in human oral fibroblasts was demonstrated in several assays of which colony forming efficiency was the most sensitive, cell killing by this agent was related to its high affinity for protein thiols, whereas glutathione showed a significant, but limited, ability to protect the cells from Hg2+ toxicity. PMID:1458551

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

    International Nuclear Information System (INIS)

    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-/4H+ and 2e-/2H+, respectively, for HCys and for GSH and Cys and high heterogeneous electron transfer rate conrogeneous 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.

  4. Functionalization of embedded thiol-ene waveguides for evanescent wave-induced fluorescence detection in a microfluidic device

    DEFF Research Database (Denmark)

    Feidenhans, Nikolaj A.; Jensen, Thomas Glasdam

    2013-01-01

    We demonstrate the use of functional surface groups inherently present on off-stoichiometric thiol-ene polymers, for site-specific immobilization of biomolecules and detection by evanescent wave-induced fluorescence. An optofluidic chip featuring an embedded thiol-ene waveguide was selectively functionalized with biotin using photografting. The biotin was used for immobilization of fluorescently labelled streptavidin, and experiments revealed a linear correlation between streptavidin concentration and fluorescent intensity. To further demonstrate the attractiveness of using thiol-ene for optofluidic devices, the optical properties of thiol-ene was evaluated by determining the transparency and refractive index of the cured polymer.

  5. Quantification of protein thiols and dithiols in the picomolar range using sodium borohydride and 4,4'-dithiodipyridine.

    DEFF Research Database (Denmark)

    Hansen, Rosa E; stergaard, Henrik

    2007-01-01

    Experimental determination of the number of thiols in a protein requires methodology that combines high sensitivity and reproducibility with low intrinsic thiol oxidation disposition. In detection of disulfide bonds, it is also necessary to efficiently reduce disulfides and to quantify the liberated thiols. Ellman's reagent (5,5'-dithiobis-[2-nitrobenzoic acid], DTNB) is the most widely used reagent for quantification of protein thiols, whereas dithiothreitol (DTT) is commonly used for disulfide reduction. DTNB suffers from a relatively low sensitivity, whereas DTT reduction is inconvenient because the reagent must be removed before thiol quantification. Furthermore, both reagents require a reaction pH > 7.0 where oxidation by ambient molecular oxygen is significant. Here we describe a quick and highly sensitive assay for protein thiol and dithiol quantification using the reducing agent sodium borohydride and the thiol reagent 4,4'-dithiodipyridine (4-DPS). Because borohydride is efficiently destroyed by the addition of acid, the complete reduction and quantification can be performed conveniently in one tube without desalting steps. Furthermore, the use of reverse-phase high-performance liquid chromatography for the thiol quantification by 4-DPS reduces the detection limit to the picomolar range (equivalent to 1 microg of a 50-kDa protein containing 1 thiol) while at the same time maintaining low pH throughout the procedure. Udgivelsesdato: 2007-Apr-1

  6. [The dynamic indices of the thiol-disulfide system and of the electrocardiogram under conditions of surgical stress].

    Science.gov (United States)

    Afanas'eva, I V; Goncharova, L L; Petrash, V V; Pokrovskaia, L A

    1998-01-01

    Changes of the thiol-disulfide balance and electrocardiographic indices were studied in ischemic heart disease patients undergoing operation stress. It was shown that the operative procedure resulted in a shift of the redox state to oxidation in the thiol-disulfide system, especially in its protein part. The revealed correlation between the value of the thiol-disulfide ratio and ECG changes attest to the significance of the state of the blood thiol-disulfide system for the assessment of the degree of ischemic alterations in the myocardium. PMID:9825447

  7. Functionalization of embedded thiol-ene waveguides for evanescent wave induced fluorescence detection in a microfluidic device

    DEFF Research Database (Denmark)

    Feidenhans'l, Nikolaj Agentoft; Jensen, Thomas Glasdam

    2013-01-01

    We demonstrate the use of functional surface groups inherently present on off-stoichiometric thiol?ene polymers, for site-specific immobilization of biomolecules and detection by evanescent wave-induced fluorescence. An optofluidic chip featuring an embedded thiol?ene waveguide was selectively functionalized with biotin using photografting. The biotin was used for immobilization of fluorescently labelled streptavidin, and experiments revealed a linear correlation between streptavidin concentration and fluorescent intensity. To further demonstrate the attractiveness of using thiol?ene for optofluidic devices, the optical properties of thiol?ene was evaluated by determining the transparency and refractive index of the cured polymer.

  8. NADH-Ferricyanide Reductase of Leaf Plasma Membranes : Partial Purification and Immunological Relation to Potato Tuber Microsomal NADH-Ferricyanide Reductase and Spinach Leaf NADH-Nitrate Reductase

    OpenAIRE

    Askerlund, Per; Laurent, Pascal; Nakagawa, Hiroki; Kader, Jean-Claude

    1991-01-01

    Plasma membranes obtained by two-phase partitioning of microsomal fractions from spinach (Spinacea oleracea L. cv Medania) and sugar beet leaves (Beta vulgaris L.) contained relatively high NADH-ferricyanide reductase and NADH-nitrate reductase (NR; EC 1.6.6.1) activities. Both of these activities were latent. To investigate whether these activities were due to the same enzyme, plasma membrane polypeptides were separated with SDS-PAGE and analyzed with immunoblotting methods. Antibodies raise...

  9. Oxidation of the albumin thiol to sulfenic acid and its implications in the intravascular compartment

    Scientific Electronic Library Online (English)

    L., Turell; S., Carballal; H., Botti; R., Radi; B., Alvarez.

    2009-04-01

    Full Text Available Human serum albumin (HSA) is the most abundant protein in the intravascular compartment. It possesses a single thiol, Cys34, which constitutes ~80% of the total thiols in plasma. This thiol is able to scavenge plasma oxidants. A central intermediate in this potential antioxidant activity of human se [...] rum albumin is sulfenic acid (HSA-SOH). Work from our laboratories has demonstrated the formation of a relatively stable sulfenic acid in albumin through complementary spectrophotometric and mass spectrometric approaches. Recently, we have been able to obtain quantitative data that allowed us to measure the rate constants of sulfenic acid reactions with molecules of analytical and biological interest. Kinetic considerations led us to conclude that the most likely fate for sulfenic acid formed in the plasma environment is the reaction with low molecular weight thiols to form mixed disulfides, a reversible modification that is actually observed in ~25% of circulating albumin. Another possible fate for sulfenic acid is further oxidation to sulfinic and sulfonic acids. These irreversible modifications are also detected in the circulation. Oxidized forms of albumin are increased in different pathophysiological conditions and sulfenic acid lies in a mechanistic junction, relating oxidizing species to final thiol oxidation products.

  10. Influence of thiol stress on oxidative phosphorylation and generation of ROS in Streptomyces coelicolor

    Directory of Open Access Journals (Sweden)

    Hemendra J. Vekaria

    2010-11-01

    Full Text Available Thiols play very important role in the intracellular redox homeostasis. Imbalance in the redox status leads to changes in the intracellular metabolism including respiration. Thiol stress, a reductive type of stress can also cause redox imbalance. When Gram-positive bacterium Strep- tomyces coelicolor was exposed to thiol stress, catalaseA was induced. Induction of catalaseA is the consequence of elevation of ROS (reactive oxygen species. The two major sources of reactive oxygen species are Fenton reaction and slippage of electrons from electron transport chain during respiration. Hence, the effect of thiol stress was checked on the rate of oxidative phosphorylation in S. coelicolor. We found correlation in the increase of oxidative phosphorylation rate and the generation of ROS, subsequently leading to induction of catalase. It was observed that thiol stress does not affect the functionality of the individual complexes of the ETC, but still there was an increase in the overall respiration, which may lead to generation of more ROS leading to induction of catalase.

  11. Fabrication of nano-pillar with sub-100nm resolution based on thiol-ene

    Science.gov (United States)

    Zhang, Man; Deng, Qiling; Shi, Lifang; Pang, Hui; Cao, Axiu; Hu, Song

    2014-11-01

    This paper demonstrates an approach to fabricate nano-pillar based on thiol-ene via soft-lithography. The template is anodic aluminum oxygen (AAO) with ordered nano-holes with the diameter of 90nm.The nano-pillar consists of rigid thiol-ene features on an elastic poly(dimethylsiloxane) (PDMS) support. It is capable of patterning both flat and curved substrate. The thiol-ene is a new green UV-curable polymer material, including a number of advantages such as rapid UV-curing in the natural environment, low-cost, high resolution, and regulative performance characteristic. Here, we fabricated a two-layer structure, which included rigid thiol-ene nano-pillar with sub-100nm resolution and soft PDMS substrate. The experiment results show that this approach can be used to fabricate high-resolution features and the thiol-ene is an excellent imprint material. The fabrication technique in this paper is simple, low-cost, high-resolution and easy to high throughput, which has broad application prospects in the preparation of nanostructures.

  12. The aldo-keto reductases (AKRs): Overview.

    Science.gov (United States)

    Penning, Trevor M

    2015-06-01

    The aldo-keto reductase (AKR) protein superfamily contains >190 members that fall into 16 families and are found in all phyla. These enzymes reduce carbonyl substrates such as: sugar aldehydes; keto-steroids, keto-prostaglandins, retinals, quinones, and lipid peroxidation by-products. Exceptions include the reduction of steroid double bonds catalyzed by AKR1D enzymes (5?-reductases); and the oxidation of proximate carcinogen trans-dihydrodiol polycyclic aromatic hydrocarbons; while the ?-subunits of potassium gated ion channels (AKR6 family) control Kv channel opening. AKRs are usually 37kDa monomers, have an (?/?)8-barrel motif, display large loops at the back of the barrel which govern substrate specificity, and have a conserved cofactor binding domain. AKRs catalyze an ordered bi bi kinetic mechanism in which NAD(P)H cofactor binds first and leaves last. In enzymes that favor NADPH, the rate of release of NADP(+) is governed by a slow isomerization step which places an upper limit on kcat. AKRs retain a conserved catalytic tetrad consisting of Tyr55, Asp50, Lys84, and His117 (AKR1C9 numbering). There is conservation of the catalytic mechanism with short-chain dehydrogenases/reductases (SDRs) even though they show different protein folds. There are 15 human AKRs of these AKR1B1, AKR1C1-1C3, AKR1D1, and AKR1B10 have been implicated in diabetic complications, steroid hormone dependent malignancies, bile acid deficiency and defects in retinoic acid signaling, respectively. Inhibitor programs exist world-wide to target each of these enzymes to treat the aforementioned disorders. Inherited mutations in AKR1C and AKR1D1 enzymes are implicated in defects in the development of male genitalia and bile acid deficiency, respectively, and occur in evolutionarily conserved amino acids. The human AKRs have a large number of nsSNPs and splice variants, but in many instances functional genomics is lacking. AKRs and their variants are now poised to be interrogated using modern genomic and informatics approaches to determine their association with human health and disease. PMID:25304492

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

    Energy Technology Data Exchange (ETDEWEB)

    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 nitrogenoxygen 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.

  14. Low temperature synthesis of thiol-functionalized CdTe nanoclusters with different tellurium contents

    Energy Technology Data Exchange (ETDEWEB)

    Sathyamoorthy, R.; Sudhagar, P.; Kumar, R.S.; Sathyadevan, T.M. [PG and Research Department of Physics, Kongunadu Arts and Science College, Coimbatore 641029, Tamilnadu (India)

    2010-01-15

    In recent years thiol-functionalized cadmium telluride nanoparticles act as a core layers for obtaining high photovoltaic efficiency in conjugated polymer/inorganic nanocrystal solar cells and also provide easy tuning of quantum size effect by varying thiol groups and processing temperature. In this work, thiol functionalized CdTe nanoclusters were prepared for different tellurium concentrations at low temperature and its impact has been discussed. Structural parameters estimated by X-ray diffraction revealed hexagonal phases at lower tellurium concentration and cubic phase at higher tellurium concentrations. Surface morphology of the samples analyzed by transmission electron microscopy, exhibited cluster type of morphology. Optical band gap energy estimated by diffused electron microscopy showed an increase with Te content and band edge emission has been observed in the photoluminescence spectra of CdTe nanoclusters around 620 nm. (copyright 2010 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim) (orig.)

  15. A selective colorimetric chemosensor for thiols based on intramolecular charge transfer mechanism

    International Nuclear Information System (INIS)

    Compound 1 as an electron donor-acceptor compound with N,N-dimethylaniline and quinone units was designed for a highly selective colorimetric determination of thiol-containing amino acids and peptides, by making use of the unique reactivity of thiol towards quinone. Compound 1 shows a strong intramolecular charge transfer (ICT) band around 582 nm; but, it decreased after addition of either cysteine (Cys) or glutathione (GSH). Moreover, the ICT band intensity at 582 nm decreased linearly with the increasing concentrations of Cys or GSH. The interference from other amino acids can be neglected. Therefore, compound 1 can be employed as a selective colorimetric visual chemosensor for thiol-containing amino acids and peptides

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

    Science.gov (United States)

    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

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

    International Nuclear Information System (INIS)

    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 ?M). Levels of cadmium, cysteine, glutathione (GSH), ?-glutamylcysteine (?-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 ?-EC. This is the first report of induction of thiol peptides, presumably phytochelatins, by a trace metal in a sea grass species

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

    Science.gov (United States)

    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), mat (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

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

    Directory of Open Access Journals (Sweden)

    Dailyn Guzmn

    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 Youngs modulus and Tg.

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

    Science.gov (United States)

    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.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

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

    International Nuclear Information System (INIS)

    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 nitrogenoxygen 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

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

    DEFF Research Database (Denmark)

    Jongberg, Sisse; Lametsch, Marianne Lund

    2014-01-01

    Radical transfer from perferrylmyoglobin to other target species (myofibrillar proteins, MPI) and bovine serum albumin (BSA), extracts from green tea (GTE), mat (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.

  4. Metabolic coupling of two small-molecule thiols programs the biosynthesis of lincomycin A.

    Science.gov (United States)

    Zhao, Qunfei; Wang, Min; Xu, Dongxiao; Zhang, Qinglin; Liu, Wen

    2015-02-01

    Low-molecular-mass thiols in organisms are well known for their redox-relevant role in protection against various endogenous and exogenous stresses. In eukaryotes and Gram-negative bacteria, the primary thiol is glutathione (GSH), a cysteinyl-containing tripeptide. In contrast, mycothiol (MSH), a cysteinyl pseudo-disaccharide, is dominant in Gram-positive actinobacteria, including antibiotic-producing actinomycetes and pathogenic mycobacteria. MSH is equivalent to GSH, either as a cofactor or as a substrate, in numerous biochemical processes, most of which have not been characterized, largely due to the dearth of information concerning MSH-dependent proteins. Actinomycetes are able to produce another thiol, ergothioneine (EGT), a histidine betaine derivative that is widely assimilated by plants and animals for variable physiological activities. The involvement of EGT in enzymatic reactions, however, lacks any precedent. Here we report that the unprecedented coupling of two bacterial thiols, MSH and EGT, has a constructive role in the biosynthesis of lincomycin A, a sulfur-containing lincosamide (C8 sugar) antibiotic that has been widely used for half a century to treat Gram-positive bacterial infections. EGT acts as a carrier to template the molecular assembly, and MSH is the sulfur donor for lincomycin maturation after thiol exchange. These thiols function through two unusual S-glycosylations that program lincosamide transfer, activation and modification, providing the first paradigm for EGT-associated biochemical processes and for the poorly understood MSH-dependent biotransformations, a newly described model that is potentially common in the incorporation of sulfur, an element essential for life and ubiquitous in living systems. PMID:25607359

  5. Engineering volatile thiol release in Saccharomyces cerevisiae for improved wine aroma.

    Science.gov (United States)

    Swiegers, Jan H; Capone, Dimitra L; Pardon, Kevin H; Elsey, Gordon M; Sefton, Mark A; Francis, I Leigh; Pretorius, Isak S

    2007-07-01

    Volatile thiols, such as 4-mercapto-4-methylpentan-2-one (4MMP), 3-mercaptohexan-1-ol (3MH) and 3-mercaptohexyl acetate (3MHA), are among the most potent aroma compounds found in wine and can have a significant effect on wine quality and consumer preferences. At optimal concentrations in wine, these compounds impart flavours of passionfruit, grapefruit, gooseberry, blackcurrant, lychee, guava and box hedge. The enzymatic release of aromatic thiols from grape-derived, non-volatile cysteinylated precursors (Cys-4MMP and Cys-3MH) and the further modification thereof (conversion of 3MH into 3MHA) during fermentation, enhance the varietal characters of wines such as Sauvignon Blanc. Wine yeast strains have limited and varying capacities to produce aroma-enhancing thiols from their non-volatile counterparts in grape juice. Even under optimal fermentation conditions, the most efficient thiol-releasing Saccharomyces cerevisiae wine strain known realizes less than 5% of the thiol-related flavour potential of grape juice. The objective of this study was to develop a wine yeast able to unleash the untapped thiol aromas in grape juice during winemaking. To achieve this goal, the Escherichia coli tnaA gene, encoding a tryptophanase with strong cysteine-beta-lyase activity, was cloned and overexpressed in a commercial wine yeast strain under the control of the regulatory sequences of the yeast phosphoglycerate kinase I gene (PGK1). This modified strain expressing carbon-sulphur lyase activity released up to 25 times more 4MMP and 3MH in model ferments than the control host strain. Wines produced with the engineered strain displayed an intense passionfruit aroma. This yeast offers the potential to enhance the varietal aromas of wines to predetermined market specifications. PMID:17492802

  6. Redox regulation of Rac1 by thiol oxidation.

    Science.gov (United States)

    Hobbs, G Aaron; Mitchell, Lauren E; Arrington, Megan E; Gunawardena, Harsha P; DeCristo, Molly J; Loeser, Richard F; Chen, Xian; Cox, Adrienne D; Campbell, Sharon L

    2015-02-01

    The Rac1 GTPase is an essential and ubiquitous protein that signals through numerous pathways to control critical cellular processes, including cell growth, morphology, and motility. Rac1 deletion is embryonic lethal, and its dysregulation or mutation can promote cancer, arthritis, cardiovascular disease, and neurological disorders. Rac1 activity is highly regulated by modulatory proteins and posttranslational modifications. Whereas much attention has been devoted to guanine nucleotide exchange factors that act on Rac1 to promote GTP loading and Rac1 activation, cellular oxidants may also regulate Rac1 activation by promoting guanine nucleotide exchange. Herein, we show that Rac1 contains a redox-sensitive cysteine (Cys(18)) that can be selectively oxidized at physiological pH because of its lowered pKa. Consistent with these observations, we show that Rac1 is glutathiolated in primary chondrocytes. Oxidation of Cys(18) by glutathione greatly perturbs Rac1 guanine nucleotide binding and promotes nucleotide exchange. As aspartate substitutions have been previously used to mimic cysteine oxidation, we characterized the biochemical properties of Rac1(C18D). We also evaluated Rac1(C18S) as a redox-insensitive variant and found that it retains structural and biochemical properties similar to those of Rac1(WT) but is resistant to thiol oxidation. In addition, Rac1(C18D), but not Rac1(C18S), shows greatly enhanced nucleotide exchange, similar to that observed for Rac1 oxidation by glutathione. We employed Rac1(C18D) in cell-based studies to assess whether this fast-cycling variant, which mimics Rac1 oxidation by glutathione, affects Rac1 activity and function. Expression of Rac1(C18D) in Swiss 3T3 cells showed greatly enhanced GTP-bound Rac1 relative to Rac1(WT) and the redox-insensitive Rac1(C18S) variant. Moreover, expression of Rac1(C18D) in HEK-293T cells greatly promoted lamellipodia formation. Our results suggest that Rac1 oxidation at Cys(18) is a novel posttranslational modification that upregulates Rac1 activity. PMID:25289457

  7. Studies of Aqueous U(IV) Complexation under Thiol-rich Conditions

    International Nuclear Information System (INIS)

    Organic thiol compounds and hydrogen sulfide (H2S) are electron donors and metabolic products of sulfate reducing bacteria. In addition, they are among redox potential (Eh) determinants of groundwater systems due to their redox characteristics. The low values of acid dissociation constants for .SH (pKa, 7-9) compared to those of aliphatic or phenolic .OH, impart greater anionic and metal-binding properties to the molecules. Recently, we demonstrated that a thiol compound (i. e., thiosalicylate) enhances the solubility of U(VI) at higher pH levels (2 nanoparticles may explain the observed solubility increase

  8. Phosphine-mediated one-pot thiol-ene "click" approach to polymer-protein conjugates.

    Science.gov (United States)

    Jones, Mathew W; Mantovani, Giuseppe; Ryan, Sinead M; Wang, Xuexuan; Brayden, David J; Haddleton, David M

    2009-09-21

    We employ water-soluble organic phosphines as key reagents in a one-pot synthetic protocol where a (poly)peptide disulfide bridge is first reduced followed by subsequent reaction of the two thiols in situ with poly(monomethoxy ethylene glycol)(meth)acrylates (p(mPEG)(M)A); we use salmon calcitonin (sCT) as a disulfide bridge-containing peptide, which contains a disulfide bridge-Cys1-Cys7-that can be reduced to give two sulfhydryl units available for thiol functionalisation; bioactivity is retained. PMID:19707643

  9. Synthesis and Biological Evaluation of Novel Benzothiazole-2-thiol Derivatives as Potential Anticancer Agents

    OpenAIRE

    Luo-Ting Yu; You-Zhi Xu; Yu-Quan Wei; Mei Deng; Ting-Hong Ye; Yong Xia; Zhao Wang (Texas A-M); Xuan-Hong Shi

    2012-01-01

    A series of novel benzothiazole-2-thiol derivatives were synthesized and their structures determined by 1H-NMR, 13C-NMR and HRMS (ESI). The effects of all compounds on a panel of different types of human cancer cell lines were investigated. Among them, pyridinyl-2-amine linked benzothiazole-2-thiol compounds 7d, 7e, 7f and 7i exhibited potent and broad-spectrum inhibitory activities. Compound 7e displayed the most potent anticancer activity on SKRB-3 (IC50 = 1.2 nM), SW620 (IC50 = 4.3 nM), A5...

  10. On the reaction of thiols with primary radiation lesions in bacteriophage

    International Nuclear Information System (INIS)

    In order to answer the question in which form, RSH of RS-, thiols react with the target responsible for reproductive cell death a study was made of the pH dependence (4.7-8.2) of the radioprotective effect of mercaptoethanol and cysteamine on the bacteriophage T4 under the anoxic conditions. It was shown that the protective effect does not depend upon pH. Since the concentration of RSH, within the studied range of pH values, remains virtually invariable, and RS- concentration sharply changes, the obtained results indicate that the RSH is the form in which thiols react with primary radiation damages to the phage

  11. Thiol activated prodrugs of sulfur dioxide (SO2) as MRSA inhibitors.

    Science.gov (United States)

    Pardeshi, Kundansingh A; Malwal, Satish R; Banerjee, Ankita; Lahiri, Surobhi; Rangarajan, Radha; Chakrapani, Harinath

    2015-07-01

    Drug resistant infections are becoming common worldwide and new strategies for drug development are necessary. Here, we report the synthesis and evaluation of 2,4-dinitrophenylsulfonamides, which are donors of sulfur dioxide (SO2), a reactive sulfur species, as methicillin-resistant Staphylococcus aureus (MRSA) inhibitors. N-(3-Methoxyphenyl)-2,4-dinitro-N-(prop-2-yn-1-yl)benzenesulfonamide (5e) was found to have excellent in vitro MRSA inhibitory potency. This compound is cell permeable and treatment of MRSA cells with 5e depleted intracellular thiols and enhanced oxidative species both results consistent with a mechanism involving thiol activation to produce SO2. PMID:25981687

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

    Science.gov (United States)

    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.

  13. Voltammetry and Electrocatalysis of Achrornobacter Xylosoxidans Copper Nitrite Reductase on Functionalized Au(111)-Electrode Surfaces

    DEFF Research Database (Denmark)

    Welinder, Anna C.; Zhang, Jingdong

    2007-01-01

    A long-standing issue in protein film voltammetry (PFV), particularly electrocatalytic voltammetry of redox enzyme monolayers, is the variability of protein adsorption modes, reflected in distributions of catalytic activity of the adsorbed protein/enzyme molecules. Use of well-defined, atomically planar electrode surfaces is a step towards the resolution of this central issue. We report here the voltammetry of copper nitrite reductase (CNiR, Achromobacter xylosoxidons) on Au(111)-electrode surfaces modified by monolayers of a broad variety of thiol-based linker molecules. These represent positively charged and electrostatically neutral, hydrophobic and hydrophilic, aliphatic and aromatic, and variable-length micro-environments, as well as their combinations. Optimal conditions for enzyme function seems to be a combination of hydrophobic and hydrophilic surface linker properties, which can lead to close to complete non-catalytic monolayer interfacial electron transfer function and electrocatalysis with activity approaching enzyme activity in homogeneous solution. Thiophenol (combined hydrophobic stacking and interdispersed water molecules), 4-methyl-thiophenol (hydrophobic and water molecules), and 3- and 4-aminothiophenol(hydrophilic, hydrophobic) offer the overall most efficient micro-environments. Subtle differences with even small structural linker differences, however, lead to widely different electrocatalytic properties, strikingly illuminated by the (omega-mercaptoamines. CuNiR thus shows highly efficient, close to ideal reversible electrocatalytic voltammetry on cysteamine-covered Au(111)-electrode surfaces, most likely due to two cysteamine orientations previously disclosed by in situ scanning tunnelling microscopy. Such a dual orientation exposes both a hydrophobic and a positively charged, hydrophilic surface feature. In contrast, the higher cysteamine homologues expose only the hydrophilic component with no electrocatalytic activity on these surfaces. These results offer a basis for rational surface design in forthcoming biological electrocatalysis useful both fundamentally and in novel biosensor technology.

  14. Species-Specific Differences in Translational Regulation of Dihydrofolate Reductase

    OpenAIRE

    Hsieh, Yi-ching; Skacel, Nancy E.; Bansal, Nitu; Scotto, Kathleen W.; Banerjee, Debabrata; Bertino, Joseph R.; Abali, Emine Ercikan

    2009-01-01

    We have observed that rodent cell lines (mouse, hamster) contain approximately 10 times the levels of dihydrofolate reductase as human cell lines, yet the sensitivity to methotrexate (ED50), the folate antagonist that targets this enzyme, is similar. Our previous studies showed that dihydrofolate reductase protein levels increased after methotrexate exposure, and we proposed that this increase was due to the relief of feedback inhibition of translation as a consequence of methotrexate binding...

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

    OpenAIRE

    Karasu, imen; Ahmet CUMAO?LU; Grpinar, 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...

  16. INHIBITION OF TYPE I 5?-REDUCTASE BY MEDICINAL PLANT EXTRACTS

    Directory of Open Access Journals (Sweden)

    Patil Vijaya

    2011-03-01

    Full Text Available Type I 5?-reductase has been implicated in skin disorders such as acne, hirsutism and male pattern baldness and its inhibition offers a potential treatment for these disorders. The aim of this study was to investigate the inhibition of type I 5?-reductase activity by extracts from Indian medicinal plants. Plant extracts were screened and selected based on their ability to inhibit Propionibacterium acnes and Staphylococcus epidermidis. Since type I 5?-reductase metabolises testosterone to ?4-androstene-3, 17-dione, the activity of enzyme was determined using RIA for testosterone and ?4-androstene-3, 17-dione. It was found that methanolic extract of Embelia ribes was a potent inhibitor of type I 5?-reductase (IC50:100?g/mL. Extracts of Vitex negundo, Terminalia chebula, and Terminalia bellerica which also inhibited type I 5?-reductase (IC50: 200-390 ?g /mL. Therefore herbal formulation of these plant extracts may be used in the treatment of skin disorders involving type I 5?-reductase.

  17. Carbon-carbon double-bond reductases in nature.

    Science.gov (United States)

    Huang, Minmin; Hu, Haihong; Ma, Li; Zhou, Quan; Yu, Lushan; Zeng, Su

    2014-08-01

    Reduction of C = C bonds by reductases, found in a variety of microorganisms (e.g. yeasts, bacteria, and lower fungi), animals, and plants has applications in the production of metabolites that include pharmacologically active drugs and other chemicals. Therefore, the reductase enzymes that mediate this transformation have become important therapeutic targets and biotechnological tools. These reductases are broad-spectrum, in that, they can act on isolation/conjugation C = C-bond compounds, ?,?-unsaturated carbonyl compounds, carboxylic acids, acid derivatives, and nitro compounds. In addition, several mutations in the reductase gene have been identified, some associated with diseases. Several of these reductases have been cloned and/or purified, and studies to further characterize them and determine their structure in order to identify potential industrial biocatalysts are still in progress. In this study, crucial reductases for bioreduction of C = C bonds have been reviewed with emphasis on their principal substrates and effective inhibitors, their distribution, genetic polymorphisms, and implications in human disease and treatment. PMID:24750117

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

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Kunlin; Bai, Meilin; Hou, Shimin, E-mail: smhou@pku.edu.cn [Key Laboratory for the Physics and Chemistry of Nanodevices, Department of Electronics, Peking University, Beijing 100871 (China); Sanvito, Stefano [School of Physics, AMBER and CRANN Institute, Trinity College, Dublin 2 (Ireland)

    2013-11-21

    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 ?-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.

  19. OXIDATION OF THIOLS USING K2S2O8 IN IONIC LIQUID

    OpenAIRE

    Hajipour, Abdol R.; Mostafavi, Majid; Ruoho, Arnold E.

    2009-01-01

    A green, straightforward and novel method for oxidation of thiols to the corresponding disulfides is reported using K2S2O8 in the ionic liquid 1-butyl-3-methylimidazolium bromide [(bmim)Br] at 6570 C. The corresponding disulfides were obtained in excellent yield and short reaction time.

  20. Analysis of thiols by microchip capillary electrophoresis for in situ planetary investigations.

    Science.gov (United States)

    Mora, Maria F; Stockton, Amanda M; Willis, Peter A

    2013-01-01

    The detection of thiols on extraterrestrial bodies could provide evidence for life, as well as a host of potential prebiological or abiological processes. Here, we report a novel protocol to analyze organic thiols by microchip CE with LIF detection. Thiols were labeled with Pacific Blue C5 maleimide and analyzed by MEKC. The separation buffer consisted of 15 mM tetraborate pH 9.2 and 25 mM SDS. The optimized method provided LODs ranging from 1.4 to 15 nM. The method was validated using samples collected from geothermal pools at Hot Creek Gorge, California, which were found to contain 2-propanethiol and 1-butanethiol in the nanomolar concentration range. These samples serve as chemical analogues to material potentially present in the reducing environment of primitive Earth and also at sulfurous regions of Mars. Hence, the protocol developed here enables highly sensitive thiol analysis in samples with complexity comparable to that expected in astrobiologically relevant extraterrestrial settings. This new protocol could be readily added to the existing suite of microfluidic chemical analyses developed for in situ planetary exploration; all that is required is the incorporation of two new reagents to the payload of an existing instrument concept. PMID:23161601

  1. Sytematic Study of the Adsorption of Thiol Molecules on Noble-Metal Nanoparticles

    Science.gov (United States)

    Barron, H.; Hidalgo, F.; Fernandez-Seivane, L.; Noguez, C.; Lopez-Lozano, X.

    2012-03-01

    The study of the interaction between nanoparticles and different types of ligands has been intensively investigated in the last years due to the potential contribution of their properties to the nanotechnology device design. These properties have opened new research fields like plasmonics, with interesting applications in optics, electronics, biophysics, medicine, pharmacology and materials science. Self-assembly monolayers have been thoroughly studied at experimental and theoretical level on extended (111) gold and silver surfaces. However, nanoparticle and molecule properties after the adsorption are still not well understood due to the different factors involved in this process such as the adsorption sites, size and element type of the nanoparticle. In this work we have performed a systematic study of the adsorption of methyl-thiol molecules on Au55 and Ag55 clusters through density functional theory calculations with the SIESTA code. Different adsorption modes of the methyl-thiol molecule on Au55 and Ag55 were considered. In general, for both type of nanoparticles, the methyl-thiol molecule prefers to be adsorbed on the Bridge sites. These results provide valuable information of the structural and electronic properties of methyl-thiol passivated Au and Ag nanoparticles.

  2. Synthesis of low molecular weight thiols in response to Cd exposure in Thlaspi caerulescens.

    Science.gov (United States)

    Hernndez-Allica, J; Garbisu, C; Becerril, J M; Barrutia, O; Garca-Plazaola, J I; Zhao, F J; Mcgrath, S P

    2006-07-01

    In this study, we investigated the accumulation of phytochelatins (PCs) and other low molecular weight (LMW) thiols in response to Cd exposure in two contrasting ecotypes differing in Cd accumulation. Using a root elongation test, we found that the highly accumulating ecotype Ganges was more tolerant to Cd than the low Cd-accumulation ecotype Prayon. L-buthionine-(S,R)-sulphoximine (BSO), a potent inhibitor of the gamma-glutamylcysteine synthetase gamma-ECS) (an enzyme involved in the PC biosynthetic pathway), increased the Cd sensitivity of Prayon, but had no effect on Ganges. Although PC accumulation increased in response to Cd exposure, no significant differences were observed between the two ecotypes. Cd exposure induced a dose-dependent accumulation of both Cys and a still unidentified LMW thiol in roots of both ecotypes. Root accumulation of Cys and this thiol was higher in Ganges than in Prayon; the ecotypic differences were more pronounced when the plants were treated with BSO. These findings suggest that PCs do not contribute to the Cd hypertolerance displayed by the Ganges ecotype of Thlaspi caerulescens, whereas Cys and other LMW thiols might be involved. PMID:17080963

  3. Tritium isotopic exchange between hydrogen sulfide and methyl thiol in gas phase

    International Nuclear Information System (INIS)

    The tritium exchange reaction between HTS and CH3SH was carried out. The degree of tritium exchange was estimated. Its dependence on time of reaction is shown. The dependence of the rate of exchange on the concentrations of methyl thiol and hydrogen sulfide is presented too. (Z.R.)

  4. Enantiospecific synthesis of [2.2]paracyclophane-4-thiol and derivatives

    Directory of Open Access Journals (Sweden)

    Gareth J. Rowlands

    2009-03-01

    Full Text Available This paper describes a simple route to enantiomerically enriched [2.2]paracyclophane-4-thiol via the stereospecific introduction of a chiral sulfoxide to the [2.2]paracyclophane skeleton. The first synthesis of an enantiomerically enriched planar chiral benzothiazole is also reported.

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

    Directory of Open Access Journals (Sweden)

    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.

  6. Phosphorylation and degradation of HMG CoA reductase.

    Science.gov (United States)

    Miller, S J; Parker, R A; Gibson, D M

    1989-01-01

    3-Hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase is the limiting enzyme step in cholesterol formation in mammalian liver and other tissues. It is a glycoprotein of 97,000 daltons embedded in the endoplasmic reticulum with a long cytoplasmic extension that is the site of catalytic conversion of HMG CoA to mevalonate. The enzyme is subject to both long-term (induction/repression; degradation) and short-term control (reversible phosphorylation) mediated by endocrine signaling (insulin, glucagon) and through negative feedback by metabolic products of mevalonate (e.g., cholesterol). The catalytic capacity of microsomal reductase falls rapidly in the presence of several protein kinases (reductase kinase, protein kinase-C, calmodulin-dependent protein kinase). Activity is restored with various protein phosphatases. Increased phosphorylation of reductase in intact cells after addition of glucagon or mevalonate is followed by enhanced degradation of the enzyme. In an in vitro model system, phosphorylated, native microsomal reductase is more rapidly cleaved by the calcium-dependent, neutral protease calpain than the dephosphorylated from of reductase. Our present research which centers on the mechanism of the in vitro model system is reviewed. Calpain in the presence of Ca2+ cleaves the cytosolic domain of phosphorylated 97 kDa reductase at two points giving rise to two fragments of nearly the same size that appear as a 52-56,000 dalton doublet by electrophoresis and immunoblotting. In the same system native reductase labeled with [gamma-32P]ATP generates a doublet with 32P solely in the upper (heavier) band. This indicates that serine phosphorylation sites lie between the two calpain cleavage loci. These are positioned in the "linker" region of the long carboxy-terminal cytosolic domain near the membrane. This segment possesses five invariant serine residues and two PEST sequences (constellations of proline, glutamate, serine and threonine) that are characteristic of proteins with short half-lives. If phosphorylation of HMG CoA reductase is confined to the linker region, we must look to this domain in order to interpret the resulting conformational changes that markedly influence reductase catalytic activity and prepare the enzyme for degradation. PMID:2624176

  7. Aerobic nitric oxide-induced thiol nitrosation in the presence and absence of magnesium cations.

    Science.gov (United States)

    Kolesnik, Bernd; Heine, Christian L; Schmidt, Renate; Schmidt, Kurt; Mayer, Bernd; Gorren, Antonius C F

    2014-11-01

    Although different routes for the S-nitrosation of cysteinyl residues have been proposed, the main in vivo pathway is unknown. We recently demonstrated that direct (as opposed to autoxidation-mediated) aerobic nitrosation of glutathione is surprisingly efficient, especially in the presence of Mg(2+). In the present study we investigated this reaction in greater detail. From the rates of NO decay and the yields of nitrosoglutathione (GSNO) we estimated values for the apparent rate constants of 8.9 0.4 and 0.55 0.06 M(-1)s(-1) in the presence and absence of Mg(2+). The maximum yield of GSNO was close to 100% in the presence of Mg(2+) but only about half as high in its absence. From this observation we conclude that, in the absence of Mg(2+), nitrosation starts by formation of a complex between NO and O2, which then reacts with the thiol. Omission of superoxide dismutase (SOD) reduced by half the GSNO yield in the absence of Mg(2+), demonstrating O2(-) formation. The reaction in the presence of Mg(2+) seems to involve formation of a Mg(2+)glutathione (GSH) complex. SOD did not affect Mg(2+)-stimulated nitrosation, suggesting that no O2(-) is formed in that reaction. Replacing GSH with other thiols revealed that reaction rates increased with the pKa of the thiol, suggesting that the nucleophilicity of the thiol is crucial for the reaction, but that the thiol need not be deprotonated. We propose that in cells Mg(2+)-stimulated NO/O2-induced nitrosothiol formation may be a physiologically relevant reaction. PMID:25236749

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

    Directory of Open Access Journals (Sweden)

    Bansal Amrit

    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 ?M, 1000 ?M 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.

  9. Thiol-disulfide redox equilibria of glutathione metaboloma compounds investigated by tandem mass spectrometry.

    Science.gov (United States)

    Rubino, Federico M; Pitton, Marco; Caneva, Enrico; Pappini, Marco; Colombi, Antonio

    2008-12-01

    The thiol group of cysteine plays a pivotal role in structural and functional biology. We use mass spectrometry to study glutathione-related homo- and heterodimeric disulfides, aiming at understanding the factors affecting the redox potentials of different disulfide/thiol pairs. Several electrospray ionization (ESI)-protonated disulfides of cysteamine, cysteine, penicillamine, N-acetylcysteine, N-acetylpenicillamine, gammaGluCySH, HSCyGly, and glutathione were analyzed on a triple quadrupole instrument to measure their energy-resolved tandem mass spectra. Fission of the disulfide bond yields RSH*H(+) and RS(+) ions. The logarithm of the intensity ratio of the RS(+)/RSH*H(+) fragments in homodimeric disulfides is proportional to the normal reduction potential of their RSSR/RSH pairs determined by nuclear magnetic resonance (NMR) in solution, the more reducing ones yielding the higher ratios. Also in some R(1)S-SR(2) disulfides, the ratio of the intensities of the RSH + H(+) and RS(+) ions of each participating thiol shows a linear relationship with the Nernst equation potential difference of the corresponding redox pairs. This behavior allows us to measure the redox potentials of some disulfide/thiol pairs by using different thiol-reducing probes of known oxidoreductive potential as reference. To assist understanding of the fission mechanism of the disulfide bond, the fragments tentatively identified as 'sulfenium' were themselves fragmented; accurate mass measurement of the resulting second-generation fragments demonstrated a loss of thioformaldehyde, thus supporting the assigned structure of this elusive intermediate of the oxidative stress pathway. Understanding this fragmentation process allows us to employ this technique with larger molecules to measure by mass spectrometry the micro-redox properties of different disulfide bonds in peptides with catalytic and signaling biological activity. PMID:19003853

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

    DEFF Research Database (Denmark)

    Lafleur, Josiane P; Senkbeil, Silja

    2015-01-01

    A novel, rapid and simple method for the preparation of emulsion-templated monoliths in microfluidic channels based on thiol-ene chemistry is presented. The method allows monolith synthesis and anchoring inside thiol-ene microchannels in a single photoinitiated step. Characterization by scanning 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 mesopores. As a demonstration, galactose oxidase and peptide-N-glycosidase F (PNGase F) were immobilized at the surface of the synthesized thiol-ene monoliths via two different mechanisms. First, cysteine groups on the protein surface were used for reversible covalent linkage to free thiol functional groups on the monoliths. Second, covalent linkage was achieved via free primary amino groups on the protein surface by means of thiol-ene click chemistry and l-ascorbic acid linkage. Thus prepared galactose oxidase and PNGase F microreactors demonstrated good enzymatic activity in a galactose assay and the deglycosilation of ribonuclease B, respectively.

  11. Role of free thiol groups in the biological activities of stonustoxin, a lethal factor from stonefish (Synanceja horrida) venom.

    Science.gov (United States)

    Khoo, H E; Chen, D; Yuen, R

    1998-03-01

    Stonustoxin (SNTX) is a two-subunit protein purified from the venom of a stonefish, Synanceia horrida. It has potent lethal activity and is also a membrane pore-forming cytolysin. The role of thiol groups in the biological activities of SNTX was investigated. Both the hemolytic and lethal activities of SNTX were potentiated by the reducing agent, dithiothreitol (DTT). The hemolytic activity of SNTX was sensitive to the modification of thiol groups by 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB). The loss of haemolytic activity correlated with the number of thiol groups that were titrated with DTNB. Thiol modification of SNTX with DTNB also inhibited its lethality. These inhibitory effects of thiol modification could be reversed by reduction with DTT. It was also found that the haemolytic activity of SNTX could not be inhibited by cholesterol. These observations indicate that free thiol groups play an important role in the haemolytic activity and lethality of SNTX but unlike other thiol-activated cytolysins, SNTX was not inhibited by cholesterol. Thus, SNTX may represent a new class of cytolytic toxin. PMID:9637366

  12. B Cells Induce Tolerance by Presenting Endogenous Peptide-IgG on MHC Class II Molecules via an IFN-?-Inducible Lysosomal Thiol Reductase-Dependent Pathway1

    OpenAIRE

    Su, Yan; Carey, Gregory; Mari?, Maja; Scott, David W.

    2008-01-01

    We have previously demonstrated that splenic B cells, transduced with peptide-IgG fusion proteins, are efficient tolerogenic APCs in vivo. Specific hyporesponsiveness to epitopes encoded in the peptide-IgG fusion protein has been achieved to over one dozen Ags, and clinical efficacy has been established in animal models for several autoimmune diseases and hemophilia. Previous studies also demonstrated that tolerance in this system requires MHC class II expression by the transduced B cells. Ye...

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

    DEFF Research Database (Denmark)

    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 reflected in the superior resistance towards non specific adsorption of proteins as shown by N 1s signal. We also performedprotein adsorption studies using quartz crystal microbalance (QCM-D). Studies involving addition of alkane thiol instead of OEG terminating alkane thiol showed the importance of OEG part of the molecule in superior resistance towards protein adsorption. The surfaces with underbrushes were imaged using atomic force microscopy (AFM) to detect any changes in mechanical properties of PEG thiol covered surfaces upon addition of OEG thiol. References: 1. Katsumi Uchida, Yuki Hoshino, Atsushi Tamura, Keitaro Yoshimoto, Shuji Kojima and Keichiro Yamashita, Ichiro Yamanaka, Hidenori Otsuka, Kazunori Kataoka, Yukio Nagasaki, Biointerphases. 2007, 2, 4, 126. 2. L. Selan, F. Berluti, C. Passariello, M. R. Comodiballanti, M. C. Thaller, Antimicrobial agents and chemotherapy, 1993, 37, 12, 2618. 3. Susan J. Sofia, V. Premnath, and Edward W. Merrill, Macromolecules, 1998, 31, 15, 5059. 4. Hidenori Otsuka, Yukio Nagasaki, and Kazunori Kataoka, Langmuir, 2004, 20, 26, 11285

  14. Labelling by 3H-N-ethylmaleimide of diamide-oxidized thiol groups in sheep red blood cell (SRBC) membranes

    International Nuclear Information System (INIS)

    Exposure of SRBC to the thiol oxidant diamide activates K:Cl cotransport, reversed upon metabolic restoration of cellular glutathione suggesting redox control of the K:Cl cotransporter, as well as by subsequent exposure to dithiothreitol (DTT). The thiols crucial for activation may be either on the transporter or on a membrane or cytoplasmic regulator. To test this hypothesis, the authors attempted to label with 3H-N-ethylmaleimide (3H-NEM) the thiols protected by diamide oxidation and reduced subsequently by DTT. SRBC were first treated with a diamide concentration activating K:Cl cotransport, followed by a second exposure to unlabeled (cold) NEM to block any non-oxidized thiol, and then hemolyzed to obtain white ghosts. The ghosts were again treated with cold NEM and after reduction by DTT exposed to 3H-NEM with and without cold NEM. Saturation labelling by 3H-NEM of diamide protected groups occurred in the range of CTT concentrations inactivating the diamide-stimulated K:Cl cotransport. Saturation labelling with 3H-NEM occurred at about 25?M NEM suggesting a Ki of less than 10?M NEM. The number of diamide protected thiols was about 5-10,000/cell membrane. At 100?M 3H-NEM, SRBC not treated with diamide possess at least 100,000 thiols cell and this number is likely to rise by tenfold at higher NEM concentrations. Thus, diamide protected about 1/1,000 of the membrane thiols in both genetically low anbrane thiols in both genetically low and high K SRBC, assayed under conditions where K:Cl cotransport is activated in intact cells. Therefore, at least some of the thiols crucial for potential regulation of K:Cl cotransport reside within the plasma membrane

  15. Periplasmic methacrylate reductase activity in Wolinella succinogenes.

    Science.gov (United States)

    Gross, R; Simon, J; Krger, A

    2001-10-01

    The cell homogenate and the soluble cell fraction of Wolinella succinogenes grown with formate and fumarate catalyzed the oxidation of benzyl viologen radical by methacrylate [apparent Km=0.23 mM, Vmax=1.0 U (mg cell protein) -1] or acrylate [apparent Km=0.50 mM, Vmax=0.77 U (mg cell protein) -1]. Crotonate did not serve as an oxidant. A mutant of W. succinogenes lacking the fccABC operon was unable to catalyze methacrylate or acrylate reduction. In contrast, the inactivation of fccC alone had no effect on these activities. Methacrylate reduction by benzyl viologen radical was not catalyzed by fumarate reductase isolated from the membrane of W. succinogenes. Cells grown with formate and fumarate did not catalyze methacrylate reduction by formate, and W. succinogenes did not grow with formate and methacrylate as catabolic substrates. The results suggest that the reduction of methacrylate or acrylate by benzyl viologen radical is most likely catalyzed either by the periplasmic flavoprotein FccA or by a complex consisting of FccA and the predicted c-type cytochrome FccB. The metabolic function of the fccABC operon remains unknown. PMID:11685377

  16. Immunoquantitation of aldose reductase in human tissues

    International Nuclear Information System (INIS)

    Rabbit antibodies raised against bovine kidney aldose reductase (ALR2) were shown to be monospecific for human ALR2 by Western blot analysis of human muscle homogenates. The human enzyme was detected, by reaction with the antiserum (alpha-BKALR2), in homogenates of adrenal gland, muscle, lens, brain, testes, kidney, and placenta, but not in erythrocytes or leukocytes. The amount of enzyme in each tissue was determined by densitometric analysis of autoradiographs of Western blots probed with alpha-BKALR2 and [125I]protein A. Standard curves of radiographic intensity versus amount of purified human muscle ALR2 were linear in the 20 to 200-ng range; a similar sensitivity was seen in tissue homogenates containing up to 675 micrograms total protein. The results presented here for the ALR2 level in human tissues (adrenal greater than muscle greater than lens approximately brain approximately testes greater than kidney approximately placenta) are in agreement with literature values for those tissues from which the enzyme has previously been purified. A notable exception was the absence of detectable ALR2 in human erythrocytes. A quantitative comparison of immunoradiographic response showed that bovine kidney ALR2 was about sevenfold more reactive with a alpha-BKALR2 compared to the human muscle enzyme

  17. Increased 5?-reductase activity in idiopathic hirsutism

    International Nuclear Information System (INIS)

    In vitro, genital skin 5?-reductase activity (5?-RA) was measured in ten hirsute women with normal androgen levels (idiopathic hirsutism (IH)) and in ten hirsute women with elevated androgen levels (polycystic ovary syndrome (PCO)) in order to determine the influence of secreted androgens on 5?-RA. In vitro 5?-RA was assessed by incubations of skin with 14C-testosterone (T) for 2 hours, after which steroids were separated and the radioactivity of dihydrotestosterone (DHT) and 5?-androstane 3?-17?-estradiol (3?-diol) in specific eluates were determined. All androgens were normal in IH with the exception of higher levels of 3?-diol glucuronide which were similar to the levels of PCO. The conversion ratio (CR) of T to DHT in IH and PCO were similar, yet significantly greater than the CR of control subjects. The CR of T to 3?-diol in IH and PCO were similar, yet higher than in control subjects. Serum androgens showed no correlation with 5?-RA, while the CR of T to DHT showed a significant positive correlation with the Ferriman and Gallwey score. The increased 5?-RA in IH appears to be independent of serum androgen levels and is, therefore, an inherent abnormality. The term idiopathic is a misnomer, because hirsutism in these patients may be explained on the basis of increased skin 5?-RA

  18. Binding of Fidarestat Stereoisomers with Aldose Reductase

    Directory of Open Access Journals (Sweden)

    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.

  19. Redox regulation of surface protein thiols: identification of integrin alpha-4 as a molecular target by using redox proteomics

    OpenAIRE

    GIANAZZA, ELISABETTA

    2003-01-01

    Thiols affect a variety of cell functions, an effect known as redox regulation. We show here that treatment (1-2 h) of cells with 0.1-5 mM N-acetyl-L-cysteine (NAC) increases surface protein thiol expression in human peripheral blood mononuclear cells. This effect is not associated with changes in cellular glutathione (GSH) and is also observed with a non-GSH precursor thiol N-acetyl-D-cysteine or with GSH itself, which is not cell-permeable, suggesting a direct reducing action. NAC did not a...

  20. Redox regulation of surface protein thiols: Identification of integrin ?-4 as a molecular target by using redox proteomics

    OpenAIRE

    Laragione, Teresa; Bonetto, Valentina; Casoni, Filippo; Massignan, Tania; Bianchi, Giancarlo; GIANAZZA, ELISABETTA; Ghezzi, Pietro

    2003-01-01

    Thiols affect a variety of cell functions, an effect known as redox regulation. We show here that treatment (12 h) of cells with 0.15 mM N-acetyl-l-cysteine (NAC) increases surface protein thiol expression in human peripheral blood mononuclear cells. This effect is not associated with changes in cellular glutathione (GSH) and is also observed with a non-GSH precursor thiol N-acetyl-d-cysteine or with GSH itself, which is not cell-permeable, suggesting a direct reducing action. NAC did n...

  1. Characterization of a Thermolabile Sulfite Reductase from Salmonella pullorum1

    Science.gov (United States)

    Hoeksema, Walter D.; Schoenhard, Delbert E.

    1971-01-01

    The biochemical basis for sulfite accumulation by sulfate-using revertants of Salmonella pullorum was determined. All of the sulfate-using mutants isolated from wild-type S. pullorum accumulated sulfite when grown at 37 but not at 25 C. The specific activity of reduced nicotinamide adenine dinucleotide (NADPH)-dependent sulfite reductase (H 2S-NADP oxidoreductase, EC 1.8.1.2) and of reduced methyl viologen (MVH)-dependent sulfite reductase (H 2S-MV oxidoreductase), in extracts prepared from cells incubated at 37 C, declined as the incubation period lengthened. However, the specific activity of both reductases from cells incubated at 25 C did not decline. Thermolability of cell-free NADPH-dependent sulfite reductase from cells of S. pullorum incubated at 37 C was greater than the lability of this enzyme either from cells of S. typhimurium incubated at 37 C or from cells of S. pullorum incubated at 25 C. Cells cultured at 37 C continued to accumulate sulfite when the incubation temperature was shifted to 25 C; cells cultured at 25 C and shifted to 37 C accumulated no sulfite, whereas these cells shifted to 41 C accumulated sulfite. It was concluded that the configuration of the sulfite reductase of S. pullorum strain 618 is a function of the incubation temperature at which synthesis occurs. PMID:5122801

  2. The role of biliverdin reductase in colorectal cancer

    International Nuclear Information System (INIS)

    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 a. The role of BVR overexpression in vivo and its exact influence on cancer progression and patient survival need to be further investigated. (author)

  3. Aldose Reductase Inhibition Prevents Endotoxin-Induced Inflammatory Responses in Retinal Microglia

    OpenAIRE

    Chang, Kun-Che; Ponder, Jessica; LaBarbera, Daniel V.; PETRASH, J. MARK

    2014-01-01

    Retinal microglia contribute to the production of pro-inflammatory cytokines in the eye. We show that downregulation of aldose reductase in microglia inhibits response to endotoxin, suggesting that aldose reductase inhibitors may be useful against retinal inflammatory disease.

  4. Pseudo-constitutivity of nitrate-responsive genes in nitrate reductase mutants

    OpenAIRE

    Schinko, Thorsten; Gallmetzer, Andreas; Amillis, Sotiris; Strauss, Joseph

    2013-01-01

    ? Constitutive phenotype in nitrate-reductase mutants depends on nitrate transporters. ? Intracellular nitrate derives from media components. ? Internal nitrate generation from nitric oxide. ? Nitrate transporters are functional in cells lacking nitrate reductase.

  5. Roles for cytosolic NADPH redox in regulating pulmonary artery relaxation by thiol oxidation-elicited subunit dimerization of protein kinase G1?.

    Science.gov (United States)

    Neo, Boon Hwa; Patel, Dhara; Kandhi, Sharath; Wolin, Michael S

    2013-08-01

    The activity of glucose-6-phosphate dehydrogenase (G6PD) appears to control a vascular smooth muscle relaxing mechanism regulated through cytosolic NADPH oxidation. Since our recent studies suggest that thiol oxidation-elicited dimerization of the 1? form of protein kinase G (PKG1?) contributes to the relaxation of isolated endothelium-removed bovine pulmonary arteries (BPA) to peroxide and responses to hypoxia, we investigated whether cytosolic NADPH oxidation promoted relaxation by PKG1? dimerization. Relaxation of BPA to G6PD inhibitors 6-aminonicotinamide (6-AN) and epiandrosterone (studied under hypoxia to minimize basal levels of NADPH oxidation and PKG1? dimerization) was associated with increased PKG1? dimerization and PKG-mediated vasodilator-stimulated phosphoprotein (VASP) phosphorylation. Depletion of PKG1? by small inhibitory RNA (siRNA) inhibited relaxation of BPA to 6-AN and attenuated the increase in VASP phosphorylation. Relaxation to 6-AN did not appear to be altered by depletion of soluble guanylate cyclase (sGC). Depletion of G6PD, thioredoxin-1 (Trx-1), and Trx reductase-1 (TrxR-1) in BPA with siRNA increased PKG1? dimerization and VASP phosphorylation and inhibited force generation under aerobic and hypoxic conditions. Depletion of TrxR-1 with siRNA inhibited the effects of 6-AN and enhanced similar responses to peroxide. Peroxiredoxin-1 depletion by siRNA inhibited PKG dimerization to peroxide, but it did not alter PKG dimerization under hypoxia or the stimulation of dimerization by 6-AN. Thus regulation of cytosolic NADPH redox by G6PD appears to control PKG1? dimerization in BPA through its influence on Trx-1 redox regulation by the NADPH dependence of TrxR-1. NADPH regulation of PKG dimerization may contribute to vascular responses to hypoxia that are associated with changes in NADPH redox. PMID:23709600

  6. The interplay between thiol-compounds against chromium (VI) in the freshwater green alga Monoraphidium convolutum: Toxicology, photosynthesis, and oxidative stress at a glance

    International Nuclear Information System (INIS)

    In this paper, the multifaceted Cr(VI) toxicity over the freshwater green alga Monoraphidium convolutum was assessed by concomitantly monitoring thiol-dependent redox balances, photosynthesis activity and growth-survival scores. Control group showed exponential growth rate at (5.78 0.29) division/day until 8th day with linear increasing chlorophyll a/protein ratios (CHLa/PROT) throughout the period. Cultures of M. convolutum were exposed for 5 days to Cr(VI) concentrations from 0 up to 100 mg/L showing that CHLa/PROT ratios were sensibly affected, in agreement to the calculated LC50,48h (5.38 0.72) mg/L from the concentration-response curve of cell mortality after 48 h. Regarding photosynthesis effects, Cr(VI) concentrations >1.0 mg/L showed significant increases in short-term (after 2 h) electron transfer rates (ETR) and quantum yields of photosystem II (?PSII), followed by subsequent decline of both parameters after 48 and 72 h. Biochemical analyses showed that maximal GSH concentrations in algal cultures were observed upon 1 mg Cr(VI)/L and higher dichromate concentrations dramatically increased the activity of antioxidant GSH-dependent enzymes ascorbate peroxidase and glutathione reductase. However, no variation was observed in the cellular GSH levels, whereas GSSG and lipid peroxidation indexes abruptly increased upon 10 mg Cr(VI)/L exposure. Altogether, plant physiology, photosynthesis and biochemical data suggest that the GSH-dependemical data suggest that the GSH-dependent antioxidant system is capable to sustain M. convolutum viability through efficient photosynthesis activity and adequate antioxidant responses up to Cr(VI) concentrations of 1.0 mg/L, when redox unbalances were first evidenced.

  7. Cytochrome c biogenesis in Campylobacter jejuni requires cytochrome c6 (CccA; Cj1153) to maintain apocytochrome cysteine thiols in a reduced state for haem attachment.

    Science.gov (United States)

    Liu, Yang-Wei; Kelly, David J

    2015-06-01

    The microaerophilic food-borne pathogen Campylobacter jejuni uses complex cytochrome-rich respiratory chains for growth and host colonisation. Cytochrome c biogenesis requires haem ligation to reduced apocytochrome cysteines, catalysed by the cytochrome c synthase, CcsBA. While ccsBA could not be deleted, we showed that the thiol reductase DsbD and the CcsX homologue Cj1207 are involved in, but not essential for, cytochromes c biogenesis. Mutant phenotypic analyses and biochemical studies with purified proteins revealed that the mono-haem c-type cytochromes Cj1153 (CccA) and Cj1020 (CccB) and the di-haem Cj0037 (CccC) are electron donors to the cb-oxidase (CcoNOQP), with CccC being more efficient than CccA. Remarkably, cccA deletion or site-directed mutagenesis resulted in an almost complete loss of all other c-type cytochromes. Cytochrome c structural and biogenesis genes were still transcribed in the cccA deletion mutant and the quinol oxidase genes (cioAB) were up-regulated. Cytochrome c production could be rescued in this mutant by growth with exogenous dithiothreitol or L-cysteine, suggesting that in the absence of CccA, apocytochrome c haem binding motifs become oxidised, preventing haem attachment. Our results identify CccA, the most abundant periplasmic c-type cytochrome in C.?jejuni, as a novel and unexpected protein required for cytochrome c biogenesis in this pathogen. PMID:25825009

  8. The interplay between thiol-compounds against chromium (VI) in the freshwater green alga Monoraphidium convolutum: Toxicology, photosynthesis, and oxidative stress at a glance

    Energy Technology Data Exchange (ETDEWEB)

    Takami, R. [Postgraduate Program in Environmental Chemistry, CBS, Universidade Cruzeiro do Sul, 08060070, Sao Paulo, SP (Brazil); Almeida, J.V. [Department of Biochemistry, Instituto de Quimica, Universidade de Sao Paulo (IQ-USP), Sao Paulo, SP (Brazil); Vardaris, C.V. [Postgraduate Program in Environmental Chemistry, CBS, Universidade Cruzeiro do Sul, 08060070, Sao Paulo, SP (Brazil); Colepicolo, P. [Department of Biochemistry, Instituto de Quimica, Universidade de Sao Paulo (IQ-USP), Sao Paulo, SP (Brazil); Barros, M.P., E-mail: marcelo.barros@cruzeirodosul.edu.br [Postgraduate Program in Environmental Chemistry, CBS, Universidade Cruzeiro do Sul, 08060070, Sao Paulo, SP (Brazil)

    2012-08-15

    In this paper, the multifaceted Cr(VI) toxicity over the freshwater green alga Monoraphidium convolutum was assessed by concomitantly monitoring thiol-dependent redox balances, photosynthesis activity and growth-survival scores. Control group showed exponential growth rate at (5.78 {+-} 0.29) division/day until 8th day with linear increasing chlorophyll a/protein ratios (CHLa/PROT) throughout the period. Cultures of M. convolutum were exposed for 5 days to Cr(VI) concentrations from 0 up to 100 mg/L showing that CHLa/PROT ratios were sensibly affected, in agreement to the calculated LC{sub 50,48h} (5.38 {+-} 0.72) mg/L from the concentration-response curve of cell mortality after 48 h. Regarding photosynthesis effects, Cr(VI) concentrations >1.0 mg/L showed significant increases in short-term (after 2 h) electron transfer rates (ETR) and quantum yields of photosystem II ({Phi}{sub PSII}), followed by subsequent decline of both parameters after 48 and 72 h. Biochemical analyses showed that maximal GSH concentrations in algal cultures were observed upon 1 mg Cr(VI)/L and higher dichromate concentrations dramatically increased the activity of antioxidant GSH-dependent enzymes ascorbate peroxidase and glutathione reductase. However, no variation was observed in the cellular GSH levels, whereas GSSG and lipid peroxidation indexes abruptly increased upon 10 mg Cr(VI)/L exposure. Altogether, plant physiology, photosynthesis and biochemical data suggest that the GSH-dependent antioxidant system is capable to sustain M. convolutum viability through efficient photosynthesis activity and adequate antioxidant responses up to Cr(VI) concentrations of 1.0 mg/L, when redox unbalances were first evidenced.

  9. Identification and selective inhibition of an isozyme of steroid 5 alpha-reductase in human scalp.

    Science.gov (United States)

    Harris, G; Azzolina, B; Baginsky, W; Cimis, G; Rasmusson, G H; Tolman, R L; Raetz, C R; Ellsworth, K

    1992-11-15

    Steroid 5 alpha-reductase (EC 1.3.1.22) catalyzes the reduction of testosterone to dihydrotestosterone. The 5 alpha-reductase found in human scalp has been compared with the enzyme found in prostate. The scalp reductase has a broad pH optimum centered at pH 7.0. This is distinctly different from the pH optimum of 5.5 observed with the prostatic form of the enzyme. These enzymes also differ in the Km for testosterone, which is 25-fold higher for the scalp reductase. The most significant difference between the two enzymes is their affinity for inhibitors. Two 4-azasteroids and a 3-carboxyandrostadiene are potent inhibitors of the prostatic reductase but are weak inhibitors of the scalp reductase. In contrast, several N-4-methylazasteroids are good inhibitors of the scalp reductase. These findings support a proposal that different isozymes of 5 alpha-reductase may exist in scalp and prostate. The scalp reductase was also compared to 5 alpha-reductase 1, one of the two enzymes recently cloned from human prostate [Andersson, S. & Russell, D. W. (1990) Proc. Natl. Acad. Sci. USA 87, 3640-3644; and Andersson, S., Berman, D. M., Jenkins, E. P. & Russell, D. W. (1991) Nature (London) 354, 159-161]. The characteristics of the cloned reductase 1 are comparable to those of the scalp reductase. PMID:1438277

  10. Aldo keto reductases 1B in endocrinology and metabolism

    Directory of Open Access Journals (Sweden)

    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 messengers.

  11. A study of oxidative stress, thiol proteins and role of vitamin E supplementation in chronic obstructive pulmonary disease (COPD

    Directory of Open Access Journals (Sweden)

    Anita M. Raut

    2013-04-01

    Full Text Available Background: Lipid peroxide plays an important role in inflammatory lung disease. Increased epithelial permeability produced by cigarette smoke is likely to be mediated through depletion of thiol proteins. Imbalance between oxidants and thiol proteins is also an established fact in these patients. Materials & methods: In the present study 30 healthy non-smokers were served as controls and 20 patients with stable COPD were included. Their base line clinical examination, Malondialdehyde (MDA as an oxidant, alpha tocopherol and erythrocyte superoxide dismutase (SOD as an antioxidants and thiol proteins levels were measured. All above parameters were repeated after 12 weeks of supplementation with 400 IU of vitamin E daily. Results: We observed that the mean malondialdehyde levels in these patients at base line were high (p<0.001 than Control Plasma alpha-tocopherol, SOD and thiol proteins levels were low (p<0.001 in the patients compared to controls. Exogenous vitamin E (400 IU twice daily Supplementation did not bring about any significant change in plasma Erythrocyte Superoxide Dismutase and vitamin E. But slight increase in the plasma thiol proteins levels was seen. The present study shows that initially the plasma lipid peroxide (MDA levels were high antioxidant (alpha- tocopherol, SOD and thiol proteins were low in patients with COPD. Exogenous supplementation with vitamin E increases slightly thiol proteins levels and brings down the levels of MDA showing attenuation of further damage. Conclusion: Our study confirmed the existence of oxidative stress and and the augmentation of antioxidant defenses as shown by slight increase in thiol proteins level. The antioxidant therapy is adjunct in lung disease patients and opens a promising field in prevention of oxidative stress related complications in these patients.

  12. A Dendritic Thioester Hydrogel Based on Thiol-Thioester Exchange as a Dissolvable Sealant System for Wound Closure

    OpenAIRE

    Ghobril, Cynthia; Charoen, Kristie; Rodriguez, Edward K.; Nazarian, Ara; Grinstaff, Mark W.

    2013-01-01

    A dissolvable dendritic thioester hydrogel based on thiol-thioester exchange for wound closure is reported. The hydrogel sealant adheres strongly to tissues, closes an ex vivo vein puncture, and withstands high pressures placed on a wound. The hydrogel sealant can be completely washed off upon exposure to thiolates based on thiol-thioester exchange and allow gradual wound re-exposure during definitive surgical care.

  13. Modification of porous silicon rugate filters through thiol-yne photochemistry

    Energy Technology Data Exchange (ETDEWEB)

    Soeriyadi, Alexander H., E-mail: alexander.soeriyadi@unsw.edu.au; Zhu, Ying, E-mail: alexander.soeriyadi@unsw.edu.au; Gooding, J. Justin, E-mail: justin.gooding@unsw.edu.au [Australian Centre for Nanomedicine and School of Chemistry, University of New South Wales, Sydney 2052 (Australia); Reece, Peter [School of Physics, University of New South Wales, Sydney 2052 (Australia)

    2014-02-24

    Porous silicon (PSi) has a considerable potential as biosensor platform. In particular, the ability to modify the surface chemistry of porous silicon is of interest. Here we present a generic method to modify the surface of porous silicon through thiol-yne photochemistry initiated by a radical initiator. Firstly, a freshly etched porous silicon substrate is modified through thermal hydrosilylation with 1,8-nonadiyne to passivate the surface and introduce alkyne functionalities. The alkyne functional surface could then be further reacted with thiol species in the presence of a radical initiator and UV light. Functionalization of the PSi rugate filter is followed with optical reflectivity measurements as well as high resolution X-ray photoelectron spectroscopy (XPS)

  14. Field effect on digestive ripening of thiol-capped gold nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Meng-Lin; Peng, J. S.; Lee, Sanboh, E-mail: sblee@mx.nthu.edu.tw [Department of Materials Science and Engineering, National Tsing Hua University, Hsinchu 30013, Taiwan (China); Yang, Fuqian [Department of Chemical and Materials Engineering, University of Kentucky, Lexington, Kentucky 40506 (United States)

    2014-02-07

    We studied the digestive ripening of thiol-capped gold nanoparticles under simultaneous action of electric field and reflux heating in a silicone oil bath at 130?C, using transmission electron microscopy. Observation revealed that a polydispersed gold nanoparticle system reached the state of nearly monodispersity under the action of an electric field and the thiol-capped gold nanoparticles carried negative charges. The electric field caused the increase of the particle size for the nearly monodispersed gold nanoparticle system. The self-assembly of the nearly monodisperse gold nanoparticles under the action of an electric field of a high field intensity was observed. The gold nanoparticles tended to form self-assembled nanostructures of six-fold symmetry. This study provides a new route for system engineering to control the particle size of metallic nanoparticles by electric field and digestive ripening.

  15. Tritium isotopic exchange between hydrogen sulfide and thiols in gas phase

    International Nuclear Information System (INIS)

    It has been established that the rate constant of tritium isotopic exchange between hydrogen sulfide and thiols within the temperature range 293-333K is given by k[dm3.mole-1.s-1]=p.exp[-6000[J.mole-1]/RT] where p are (6+-1.7)x10-2, (4.6+-0.5)x10-2, (3.8+-0.7)x10-2, (7.1+-1.2)x10-3 for methyl, ethyl, n-propyl, i-propyl thiol, respectively. The total order of the reaction is equal to 2, and the partial orders in respect to H2S and thol are equal to 1. The mechanism of the isotopic exchange is discussed. (author)

  16. Synthesis and Biological Evaluation of Novel Benzothiazole-2-thiol Derivatives as Potential Anticancer Agents

    Directory of Open Access Journals (Sweden)

    Luo-Ting Yu

    2012-03-01

    Full Text Available A series of novel benzothiazole-2-thiol derivatives were synthesized and their structures determined by 1H-NMR, 13C-NMR and HRMS (ESI. The effects of all compounds on a panel of different types of human cancer cell lines were investigated. Among them, pyridinyl-2-amine linked benzothiazole-2-thiol compounds 7d, 7e, 7f and 7i exhibited potent and broad-spectrum inhibitory activities. Compound 7e displayed the most potent anticancer activity on SKRB-3 (IC50 = 1.2 nM, SW620 (IC50 = 4.3 nM, A549 (IC50 = 44 nM and HepG2 (IC50 = 48 nM and was found to induce apoptosis in HepG2 cancer cells.

  17. Reactive superhydrophobic surface and its photoinduced disulfide-ene and thiol-ene (bio)functionalization.

    Science.gov (United States)

    Li, Junsheng; Li, Linxian; Du, Xin; Feng, Wenqian; Welle, Alexander; Trapp, Oliver; Grunze, Michael; Hirtz, Michael; Levkin, Pavel A

    2015-01-14

    Reactive superhydrophobic surfaces are highly promising for biotechnological, analytical, sensor, or diagnostic applications but are difficult to realize due to their chemical inertness. In this communication, we report on a photoactive, inscribable, nonwettable, and transparent surface (PAINTS), prepared by polycondensation of trichlorovinylsilane to form thin transparent reactive porous nanofilament on a solid substrate. The PAINTS shows superhydrophobicity and can be conveniently functionalized with the photoclick thiol-ene reaction. In addition, we show for the first time that the PAINTS bearing vinyl groups can be easily modified with disulfides under UV irradiation. The effect of superhydrophobicity of PAINTS on the formation of high-resolution surface patterns has been investigated. The developed reactive superhydrophobic coating can find applications for surface biofunctionalization using abundant thiol or disulfide bearing biomolecules, such as peptides, proteins, or antibodies. PMID:25486338

  18. Magnetically controlled bioelectrocatalytic system based on ferrocene-tagged magnetic nanoparticles by thiol-ene reaction

    International Nuclear Information System (INIS)

    A simple and versatile method for the introduction of electrochemical moieties onto the surface of Fe3O4 nanoparticles has been developed based on UV-induced thiol-ene click chemistry. Thiol-terminated Fe3O4 nanoparticles were synthesized and further reacted with vinylferrocence under 365 nm UV. The functionalized magnetic nanoparticles were characterized using a powder X-ray diffractometer (XRD), transmission electron microscope (TEM), Fourier transform infrared spectroscope (FTIR), and vibrating sample magnetometer (VSM). The resulting nanocomposites possess of magnetism and electrochemical activity. Based on the superparamagnetism of Fe3O4 nanoparticles and the electrocatalytic activity of ferrocene, a recyclable, magneto-controlled bioelectrocatalytic system for glucose oxidation is developed. The switching of the biocatalytic activity and recyclable usage of the ferrocene functionalized nanoparticles by means of the external magnet could provide a simple, green and convenient strategy for bioelectrosensing.

  19. Highly stretchable thermoset fibers and nonwovens using thiol-ene photopolymerization.

    Science.gov (United States)

    Shanmuganathan, Kadhiravan; Elliot, Steven M; Lane, Austin P; Ellison, Christopher J

    2014-08-27

    In this report, we describe the preparation and characterization of a new class of thermoset fibers with high elongation and elastic recovery. Integrating UV-activated thiol-ene photopolymerization and electrospinning, we demonstrate an environmentally friendly single step approach to convert small monomeric precursor molecules into highly elastic fibers and nonwoven mats. The fibers were derived by in situ photopolymerization of a trifunctional vinyl ether monomer and a tetrafunctional thiol. Although thermosets often offer good chemical and thermal stability, these fibers also have a high average elongation at break of 62%. The elastomeric nature of these vinyl-ether based fibers can be partly attributed to their subambient Tg and partly to the cross-link density, monomer structure, and resulting network homogeneity. Nonwoven mats of these fibers were also stretchable and exhibited a much higher elongation at break of about 85%. These thermoset stretchable fibers could have potential applications as textile, biomedical, hot chemical filtration, and composite materials. PMID:25075754

  20. Patterned Hydrophilization of Nanoporous 1,2?PB by Thiol?ene Photochemistry

    DEFF Research Database (Denmark)

    Berthold, Anton; Sagar, Kaushal Shashikant

    2011-01-01

    We present an efficient method for functionalizing the large polymerair interface of a gyroid nanoporous polymer. The hydrophilicity of nanoporous cross?linked 1,2?polybutadiene is tuned by thiol?ene photo?grafting of mercaptosuccinic acid or sodium 2?mercaptoethanesulfonate. The reaction is monitored by FT?IR, UVVis, contact angle, and gravimetry. Overall quantum yields are calculated for the two thiol?ene click reactions in nano?confinement, neatly revealing their chain?like nature. Topdown photolithographic patterning is demonstrated, realizing hydrophilic nanoporous corridors exclusively hosting water. The presented approach can be relevant for many applications where, e.g., high control and contrast in hydrophilicity, chemical functionality or refractive index are needed.

  1. Pleiotropic effects of the HMG-CoA reductase inhibitors

    Directory of Open Access Journals (Sweden)

    Mihos CG

    2011-04-01

    Full Text Available Christos G Mihos, Orlando SantanaColumbia University Division of Cardiology, Mount Sinai Heart Institute, Miami Beach, FL, USAAbstract: The HMG-CoA reductase inhibitors (statins are used extensively in the treatment of hyperlipidemia. They have also demonstrated a benefit in a variety of other disease processes. These secondary actions are known as pleiotropic effects. Our paper serves as a focused and updated discussion on the pleiotropy of statins and emphasizes the importance of randomized placebo-controlled trials to further elucidate this interesting phenomenon.Keywords: HMG-CoA inhibitors, pleiotropic, reductase, review, statins

  2. A new turn-on chemosensor for bio-thiols based on the nanoaggregates of a tetraphenylethene-coumarin fluorophore

    Science.gov (United States)

    Lou, Xiaoding; Zhao, Zujin; Hong, Yuning; Dong, Chao; Min, Xuehong; Zhuang, Yuan; Xu, Xuemei; Jia, Yongmei; Xia, Fan; Tang, Ben Zhong

    2014-11-01

    In this work, a tetraphenylethene-coumarin hybrid fluorophore (TPE-Cou) that contains a Schiff base form is designed and synthesized. A combination of plentiful optical properties and chemical reactivity towards thiols allows TPE-Cou to work as an excellent turn-on probe of thiols with a wide linear range, revealing the great potential of this dye as a quantitative fluorescence indicator. By means of NMR and optical spectrum analyses, a mechanistic picture at the molecular level has been drawn to illustrate how this dye works as a bio-thiol-sensitive fluorescent probe.In this work, a tetraphenylethene-coumarin hybrid fluorophore (TPE-Cou) that contains a Schiff base form is designed and synthesized. A combination of plentiful optical properties and chemical reactivity towards thiols allows TPE-Cou to work as an excellent turn-on probe of thiols with a wide linear range, revealing the great potential of this dye as a quantitative fluorescence indicator. By means of NMR and optical spectrum analyses, a mechanistic picture at the molecular level has been drawn to illustrate how this dye works as a bio-thiol-sensitive fluorescent probe. Electronic supplementary information (ESI) available: Details of the experimental procedure and analytical data are provided. See DOI: 10.1039/c4nr04593a

  3. Assemblies of thiol-capped nanocrystals as functional units for use in nanotechnology

    OpenAIRE

    Gaponik, Nikolai

    2013-01-01

    This work summarizes results of about 10 years of the authors own activities in the field of aqueous synthesis and handling of thiol-capped semiconductor nanocrystals. As this field has also been explored by hundreds of other scientists, I have endeavoured to do my utmost to provide a short but comprehensive overview presenting my own results as part of the knowledge framework jointly created by multiple colleagues and collaborators from all over the world. This habilitation thesis consist...

  4. Characterization of thiol-functionalised silica films deposited on electrode surfaces

    OpenAIRE

    Ivana Cesarino; der Tadeu Gomes Cavalheiro

    2008-01-01

    Thiol-functionalised silica films were deposited on various electrode surfaces (gold, platinum, glassy carbon) by spin-coating sol-gel mixtures in the presence of a surfactant template. Film formation occurred by evaporation induced self-assembly (EISA) involving the hydrolysis and (co)condensation of silane and organosilane precursors on the electrode surface. The characterization of such material was performed by IR spectroscopy, thermogravimetry (TG), elemental analysis (EA), atomic force ...

  5. Thiol-mediated multiple mechanisms centered on selenodiglutathione determine selenium cytotoxicity against MCF-7 cancer cells.

    Science.gov (United States)

    Tobe, Takao; Ueda, Koji; Ando, Motozumi; Okamoto, Yoshinori; Kojima, Nakao

    2015-06-01

    Selenium (Se) is an essential antioxidative micronutrient but can exert cancer-selective cytotoxicity if the nutritional levels are too high. Selenodiglutathione (GSSeSG) is a primary Se metabolite conjugated with two glutathione (GSH) moieties. GSSeSG has been suggested to be an important molecule for cytotoxicity. Here, we propose the underlying mechanisms for the potent cytotoxicity of GSSeSG: cellular intake; reductive metabolism; production of reactive oxygen species; oxidative damage to DNA; apoptosis induction. GSSeSG rather than selenite decreased cell viability and induced apoptosis accompanied by increases in intracellular Se contents. Therefore, GSSeSG-specific cytotoxicity may be ascribed to its preferable incorporation. Base oxidation and strand fragmentation in genomic DNA preceded cell death, suggesting that oxidative stress (including DNA damage) is crucial for GSSeSG cytotoxicity. Strand breaks of purified DNA were caused by the coexistence of GSSeSG and thiols (GSH, cysteine, homocysteine), but not the oxidized form or non-thiol reductants. This implies the important role of intracellular thiols in the mechanism of Se toxicity. GSH-assisted DNA strand breaks were inhibited by specific scavengers for hydrogen peroxide or hydroxyl radicals. The GSSeSG metabolite selenide induced some DNA strand breaks without GSH, whereas elemental Se did so only with GSH. These observations suggest involvement of Fenton-type reaction in the absence of transition metals and reactivation of inert elemental Se. Overall, our results suggest that chemical interactions between Se and the sulfur of thiols are crucial for the toxicity mechanisms of Se. PMID:25783495

  6. Requirements for heme and thiols for the nonenzymatic modification of nitrotyrosine

    OpenAIRE

    Balabanli, Barbaros; KAMISAKI, YOSHINORI; Martin, Emil; Murad, Ferid

    1999-01-01

    Peroxynitrite-dependent formation of nitrotyrosine has been associated with inactivation of various enzymes and proteins possessing functionally important tyrosines. We have previously reported an enzymatic activity modifying the nitrotyrosine residues in nitrated proteins. Here we are describing a nonenzymatic reduction of nitrotyrosine to aminotyrosine, which depends on heme and thiols. Various heme-containing proteins can mediate the reaction, although the reaction also is catalyzed by hem...

  7. Sequential processing of lysosomal acid phosphatase by a cytoplasmic thiol proteinase and a lysosomal aspartyl proteinase.

    OpenAIRE

    Gottschalk, S.; Waheed, A.; Schmidt, B.; Laidler, P.; Von Figura, K.

    1989-01-01

    BHK cells expressing human lysosomal acid phosphatase (LAP) transport LAP to lysosomes as an integral membrane protein. In lysosomes LAP is released from the membrane by proteolytic processing, which involves at least two cleavages at the C terminus of LAP. The first cleavage is catalysed by a thiol proteinase at the outside of the lysosomal membrane and removes the bulk of the cytoplasmic tail of LAP. The second cleavage is catalysed by an aspartyl proteinase inside the lysosomes and release...

  8. Immobilization of Enzymes via Microcontact Printing and Thiol-Ene Click Chemistry.

    Science.gov (United States)

    Buhl, Moritz; Vonhren, Benjamin; Ravoo, Bart Jan

    2015-06-17

    This Communication describes a bioconjugation method for the generation of enzyme microarrays on surfaces using photochemical thiol-ene chemistry in combination with microcontact printing. Glucose oxidase and lactase were readily immobilized (i.e., printing time 2 min) on alkene terminated self-assembled monolayers on glass as demonstrated by X-ray photoelectron spectroscopy and fluorescence microscopy. Furthermore, the activity of both immobilized enzymes was confirmed in single enzyme as well as cascade transformations. PMID:26030726

  9. Metal-Free Iodine-Catalyzed Direct Arylthiation of Substituted Anilines with Thiols.

    Science.gov (United States)

    Yang, Daoshan; Yan, Kelu; Wei, Wei; Zhao, Jing; Zhang, Mengqi; Sheng, Xuguang; Li, Guoqing; Lu, Shenglei; Wang, Hua

    2015-06-19

    Iodine-catalyzed direct arylthiation of substituted anilines for the synthesis of various diaryl sulfides has been developed under metal- and solvent-free conditions. The present method uses readily available thiols as the arylthiation reagents, and environmentally friendly and inexpensive I2 as the catalyst. Importantly, no base or ligand was necessary. Such a simple, efficient, and economical transformation provides an attractive approach to various diaryl sulfides in good to excellent yields. PMID:26030066

  10. Surface plasmon resonance and magnetism of thiol-capped gold nanoparticles

    OpenAIRE

    Guerrero, Estefani?a; Mun?oz-ma?rquez, Miguel A?ngel; Garci?a, M. A.; Crespo, Patricia; Ferna?ndez-pinel, Enrique; Hernando, Antonio; Ferna?ndez-camacho, A.

    2008-01-01

    Surface plasmon resonance measurements and magnetic characterization studies have been carried out for two types of thiol-capped gold nanoparticles (NPs) with similar diameters between 2.0 and 2.5 nm and different organic molecules linked to the sulfur atom: dodecanethiol and tiopronin. In addition, Au NPs capped with tetraoctyl ammonium bromide have also been included in the investigation since such capping molecules weakly interact with the gold surface atoms and, therefore, this system can...

  11. Labeling Thiols on Proteins, Living Cells, and Tissues with Enhanced Emission Induced by FRET

    OpenAIRE

    Yuan, Yue; Wang, Xijun; Mei, Bin; Zhang, Dongxin; Tang, Anming; An, Linna; He, Xiaoxiao; Jiang, Jun; Liang, Gaolin

    2013-01-01

    Using N-(2-Aminoethyl)maleimide-cysteine(StBu) (Mal-Cys) as a medium, protein thiols were converted into N-terminal cysteines. After a biocompatible condensation reaction between the N-terminal cysteine and fluorescent probe 2-cyanobenzothiazole-Gly-Gly-Gly-fluorescein isothiocyanate (CBT-GGG-FITC), a new fluorogenic structure Luciferin-GGG-FITC was obtained. The latter exhibits near one order of magnitude (7 folds) enhanced fluorescence emission compared to the precursor moiety due to fluore...

  12. Mitochondrial Sulfide Detoxification Requires a Functional Isoform O-Acetylserine(thiol)lyase C in Arabidopsis thaliana

    OpenAIRE

    lvarez, Consolacin; Garca, Irene; Romero, Luis C.; Gotor, Cecilia

    2012-01-01

    In non-cyanogenic species, the main source of cyanide derives from ethylene and camalexin biosyntheses. In mitochondria, cyanide is a potent inhibitor of the cytochrome c oxidase and is metabolised by the ?-Cyanoalanine synthase CYS-C1, catalysing the conversion of cysteine and cyanide to hydrogen sulfide and ?- cyanoalanine. The hydrogen sulfide released also inhibits the cytochrome c oxidase and needs to be detoxified by the O-acetylserine(thiol)lyase mitochondrial isoform, OAS-C, which c...

  13. Michael addition of thiols, carbon nucleophiles and amines to dehydroamino acid and dehydropeptide derivatives

    OpenAIRE

    Ferreira, Paula M.T.; Maia, Hernni L. S.; Monteiro, Lus S.; Sacramento, Joana

    2001-01-01

    Michael additions of nitrogen heterocycles, thiols, carbon nucleophiles and amines to dehydroalanine derivatives, including a glycyldehydroalanine peptide, were performed in fair to good yields. Dehydroaminobutyric acid derivatives reacted only with the stronger nucleophiles but in considerably lower yields and often no reaction was observed with the corresponding dehydrophenylalanine derivatives. When a tosyl group was bonded to the nitrogen atom of the dehydroamino acid, in some cases the a...

  14. Ruthenium(III Chloride Catalyzed Acylation of Alcohols, Phenols, and Thiols in Room Temperature Ionic Liquids

    Directory of Open Access Journals (Sweden)

    Mingzhong Cai

    2009-09-01

    Full Text Available Ruthenium(III chloride-catalyzed acylation of a variety of alcohols, phenols, and thiols was achieved in high yields under mild conditions (room temperature in the ionic liquid 1-butyl-3-methylimidazolium hexafluorophosphate ([bmim][PF6]. The ionic liquid and ruthenium catalyst can be recycled at least 10 times. Our system not only solves the basic problem of ruthenium catalyst reuse, but also avoids the use of volatile acetonitrile as solvent.

  15. Ruthenium(III) Chloride Catalyzed Acylation of Alcohols, Phenols, and Thiols in Room Temperature Ionic Liquids

    OpenAIRE

    Mingzhong Cai; Pingping Wang; Wenyan Hao; Zhiwen Xi

    2009-01-01

    Ruthenium(III) chloride-catalyzed acylation of a variety of alcohols, phenols, and thiols was achieved in high yields under mild conditions (room temperature) in the ionic liquid 1-butyl-3-methylimidazolium hexafluorophosphate ([bmim][PF6]). The ionic liquid and ruthenium catalyst can be recycled at least 10 times. Our system not only solves the basic problem of ruthenium catalyst reuse, but also avoids the use of volatile acetonitrile as solvent.

  16. Carbocysteine restores steroid sensitivity by targeting histone deacetylase 2 in a thiol/GSH-dependent manner.

    Science.gov (United States)

    Song, Yun; Lu, Hao-Zhong; Xu, Jian-Rong; Wang, Xiao-Lin; Zhou, Wei; Hou, Li-Na; Zhu, Liang; Yu, Zhi-Hua; Chen, Hong-Zhuan; Cui, Yong-Yao

    2015-01-01

    Steroid insensitivity is commonly observed in patients with chronic obstructive pulmonary disease. Here, we report the effects and mechanisms of carbocysteine (S-CMC), a mucolytic agent, in cellular and animal models of oxidative stress-mediated steroid insensitivity. The following results were obtained: oxidative stress induced higher levels of interleukin-8 (IL-8) and tumor necrosis factor alpha (TNF-?), which are insensitive to dexamethasone (DEX). The failure of DEX was improved by the addition of S-CMC by increasing histone deacetylase 2 (HDAC2) expression/activity. S-CMC also counteracted the oxidative stress-induced increase in reactive oxygen species (ROS) levels and decreases in glutathione (GSH) levels and superoxide dismutase (SOD) activity. Moreover, oxidative stress-induced events were decreased by the thiol-reducing agent dithiothreitol (DTT), enhanced by the thiol-oxidizing agent diamide, and the ability of DEX was strengthened by DTT. In addition, the oxidative stress-induced decrease in HDAC2 activity was counteracted by S-CMC by increasing thiol/GSH levels, which exhibited a direct interaction with HDAC2. S-CMC treatment increased HDAC2 recruitment and suppressed H4 acetylation of the IL-8 promoter, and this effect was further ablated by addition of buthionine sulfoximine, a specific inhibitor of GSH synthesis. Our results indicate that S-CMC restored steroid sensitivity by increasing HDAC2 expression/activity in a thiol/GSH-dependent manner and suggest that S-CMC may be useful in a combination therapy with glucocorticoids for treatment of steroid-insensitive pulmonary diseases. PMID:25500537

  17. Iron and thiols as two major players in carcinogenesis: friends or foes?

    OpenAIRE

    ShinyaToyokuni

    2014-01-01

    Iron is the most abundant metal in the human body and mainly works as a cofactor for proteins such as hemoglobin and various enzymes. No independent life forms on earth can survive without iron. However, excess iron is intimately associated with carcinogenesis by increasing oxidative stress via its catalytic activity to generate hydroxyl radicals. Biomolecules with redox-active sulfhydryl function(s) (thiol compounds) are necessary for the maintenance of mildly reductive cellular environme...

  18. A Novel Strategy for Global Analysis of the Dynamic Thiol Redox Proteome*

    OpenAIRE

    Martnez-Acedo, Pablo; Nez, Estefana; Gmez, Francisco J. Snchez; Moreno, Margoth; Ramos, Elena; Izquierdo-lvarez, Alicia; Mir-Casas, Elisabet; Mesa, Raquel; Rodriguez, Patricia; Martnez-Ruiz, Antonio; Dorado, David Garcia; Lamas, Santiago; Vzquez, Jess

    2012-01-01

    Nitroxidative stress in cells occurs mainly through the action of reactive nitrogen and oxygen species (RNOS) on protein thiol groups. Reactive nitrogen and oxygen species-mediated protein modifications are associated with pathophysiological states, but can also convey physiological signals. Identification of Cys residues that are modified by oxidative stimuli still poses technical challenges and these changes have never been statistically analyzed from a proteome-wide perspective. Here we sh...

  19. Antiviral effects of a thiol protease inhibitor on foot-and-mouth disease virus.

    OpenAIRE

    Kleina, L G; Grubman, M J

    1992-01-01

    The thiol protease inhibitor E-64 specifically blocks autocatalytic activity of the leader protease of foot-and-mouth disease virus (FMDV) and interferes with cleavage of the structural protein precursor in an in vitro translation assay programmed with virion RNA. Experiments with FMDV-infected cells and E-64 or a membrane-permeable analog, E-64d, have confirmed these results and demonstrated interference in virus assembly, causing a reduction in virus yield. In addition, there is a lag in th...

  20. Convenient Test for Screening Metallo-?-Lactamase-Producing Gram-Negative Bacteria by Using Thiol Compounds

    OpenAIRE

    Arakawa, Yoshichika; Shibata, Naohiro; Shibayama, Keigo; Kurokawa, Hiroshi; Yagi, Tetsuya; Fujiwara, Hiroshi; Goto, Masafumi

    2000-01-01

    A simple disk diffusion test was constructed for detection of IMP-1-type metallo-?-lactamase-producing gram-negative bacteria. Two Kirby-Bauer disks containing ceftazidime (CAZ) and a filter disk containing a metallo-?-lactamase inhibitor were used in this test. Several IMP-1 inhibitors such as thiol compounds including 2-mercaptopropionic acid, heavy metal salts, and EDTA were evaluated for this test. Two CAZ disks were placed on a Mueller-Hinton agar plate on which a bacterial suspension ...

  1. Quantification of Polyfunctional Thiols in Wine by HS-SPME-GC-MS Following Extractive Alkylation.

    Science.gov (United States)

    Musumeci, Lauren E; Ryona, Imelda; Pan, Bruce S; Loscos, Natalia; Feng, Hui; Cleary, Michael T; Sacks, Gavin L

    2015-01-01

    Analyses of key odorous polyfunctional volatile thiols in wines (3-mercaptohexanol (3-MH), 3-mercaptohexylacetate (3-MHA), and 4-mercapto-4-methyl-2-pentanone (4-MMP)) are challenging due to their high reactivity and ultra-trace concentrations, especially when using conventional gas-chromatography electron impact mass spectrometry (GC-EI-MS). We describe a method in which thiols are converted to pentafluorobenzyl (PFB) derivatives by extractive alkylation and the organic layer is evaporated prior to headspace solid phase microextraction (HS-SPME) and GC-EI-MS analysis. Optimal parameters were determined by response surface area modeling. The addition of NaCl solution to the dried SPME vials prior to extraction resulted in up to less than fivefold improvement in detection limits. Using 40 mL wine samples, limits of detection for 4-MMP, 3-MH, and 3-MHA were 0.9 ng/L, 1 ng/L, and 17 ng/L, respectively. Good recovery (90%-109%) and precision (5%-11% RSD) were achieved in wine matrices. The new method was used to survey polyfunctional thiol concentrations in 61 commercial California and New York State wines produced from V. vinifera (Riesling, Gewrztraminer, Cabernet Sauvignon, Sauvignon blanc and non-varietal ros wines), V. labruscana (Niagara), and Vitis spp. (Cayuga White). Mean 4-MMP concentrations in New York Niagara (17 ng/L) were not significantly different from concentrations in Sauvignon blanc, but were significantly higher than 4-MMP in other varietal wines. PMID:26154886

  2. Investigation into the Effect of Molds in Grasses on Their Content of Low Molecular Mass Thiols

    Directory of Open Access Journals (Sweden)

    Adam Nawrath

    2012-10-01

    Full Text Available The aim of this study was to investigate the effect of molds on levels of low molecular mass thiols in grasses. For this purpose, the three grass species Lolium perenne, Festulolium pabulare and Festulolium braunii were cultivated and sampled during four months, from June to September. The same species were also grown under controlled conditions. High-performance liquid chromatography with electrochemical detection was used for quantification of cysteine, reduced (GSH and oxidized (GSSG glutathione, and phytochelatins (PC2, PC3, PC4 and PC5. Data were statistically processed and analyzed. Thiols were present in all examined grass species. The effect of fungicide treatments applied under field conditions on the content of the evaluated thiols was shown to be insignificant. Species influenced (p < 0.05 PC3 and GSSG content. F. pabulare, an intergeneric hybrid of drought- and fungi-resistant Festuca arundinacea, was comparable in PC3 content with L. perenne and F. braunii under field conditions. Under controlled conditions, however, F. pabulare had higher (p < 0.05 PC3 content than did L. perenne and F. braunii. Under field conditions, differences between the evaluated species were recorded only in GSSG content, but only sampling in June was significant. F. pabulare had higher (p < 0.05 GSSG content in June than did L. perenne and F. braunii.

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

    DEFF Research Database (Denmark)

    Javakhishvili, Irakli; Hvilsted, Sren

    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 initiator, 2-hydroxyethyl 2-bromoisobutyrate, followed by esterification with 2,4-dinitrophenyl- or 4-monomethoxytrityl-protected mercaptoacetic acids (Prot-), provided well-defined PCL macroinitiators capped with protected thiols. The macroinitiators allowed atom transfer radical polymerization (ATRP) of tent-butyl acrylate (tBA) in a controlled fashion by use of NiBr2(PPh3)(2) catalyst to produce Prot-PCL-b-PtBA with narrow polydispersities (1.17-1.39). Subsequent mild deprotection protocols provided HS-PCL-b-PAA. Reduction of a gold salt in the presence of this macroligand under thiol-deficient conditions afforded stable, aggregation-free nanoparticles, as evidenced from UV-vis spectroscopy and transmission electron microscopy (TEM), the latter revealed nanoparticles with a mean diameter of 9.0 +/- 3.1 nm.

  4. Requirements for heme and thiols for the nonenzymatic modification of nitrotyrosine.

    Science.gov (United States)

    Balabanli, B; Kamisaki, Y; Martin, E; Murad, F

    1999-11-01

    Peroxynitrite-dependent formation of nitrotyrosine has been associated with inactivation of various enzymes and proteins possessing functionally important tyrosines. We have previously reported an enzymatic activity modifying the nitrotyrosine residues in nitrated proteins. Here we are describing a nonenzymatic reduction of nitrotyrosine to aminotyrosine, which depends on heme and thiols. Various heme-containing proteins can mediate the reaction, although the reaction also is catalyzed by heme. The reaction is most effective when vicinal thiols are used as reducing agents, although ascorbic acid also can replace thiols with lesser efficiency. The reaction could be inhibited by (z)-1-[2-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1- ium-1, but not other tested NO donors. HPLC with electrochemical detection analysis of the reaction identified aminotyrosine as the only reaction product. The reduction of nitrotyrosine was most effective at a pH close to physiological and was markedly decreased in acidic conditions. Various nitrophenol compounds also were modified in this reaction. Understanding the mechanism of this reaction could help define the enzymatic modification of nitrotyrosine-containing proteins. Furthermore, this also could assist in understanding the role of nitrotyrosine formation and reversal in the regulation of various proteins containing nitrotyrosine. It also could help define the role of nitric oxide and other reactive species in various disease states. PMID:10557286

  5. Enhanced electrochemical sensing of thiols based on cobalt phthalocyanine immobilized on nitrogen-doped graphene.

    Science.gov (United States)

    Xu, Huiying; Xiao, Jingjing; Liu, Baohong; Griveau, Sophie; Bedioui, Fethi

    2015-04-15

    A hybrid nanocomposite based on cobalt phthalocyanine (CoPc) immobilized on nitrogen-doped graphene (N-G) (N-G/CoPc) has been developed to modify glassy carbon electrode (GCE) for the sensitive detection of thiols. The nanocomposites were characterized by transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS) and electrochemical impedance spectroscopy (EIS). Cyclic voltammetric studies showed that cobalt phthalocyanine and nitrogen doped graphene have a synergic effect and significantly enhance the electrocatalytic activity of the modified electrode towards thiols oxidation compared with electrodes modified with solely CoPc or N-G. The electrochemical oxidation responses were studied and the reaction mechanisms were discussed. The sensors exhibited a wide linear response range from 1?? to 16mM and a low detection limit of 1?? for the determination of l-cysteine, reduced l-glutathione and 2-mercaptoethanesulfonic acid in alkaline aqueous solution. The proposed N-G/CoPc hybrids contribute to the construction of rapid, convenient and low-cost electrochemical sensors for sensitive detection of thiols. PMID:25497984

  6. Thiol-reactive compounds from garlic inhibit the epithelial sodium channel (ENaC).

    Science.gov (United States)

    Krumm, Patrick; Giraldez, Teresa; Alvarez de la Rosa, Diego; Clauss, Wolfgang G; Fronius, Martin; Althaus, Mike

    2012-07-01

    The epithelial sodium channel (ENaC) is a key factor in the transepithelial movement of sodium, and consequently salt and water homeostasis in various organs. Dysregulated activity of ENaC is associated with human diseases such as hypertension, the salt-wasting syndrome pseudohypoaldosteronism type 1, cystic fibrosis, pulmonary oedema or intestinal disorders. Therefore it is important to identify novel compounds that affect ENaC activity. This study investigated if garlic (Allium sativum) and its characteristic organosulfur compounds have impact on ENaCs. Human ENaCs were heterologously expressed in Xenopus oocytes and their activity was measured as transmembrane currents by the two-electrode voltage-clamp technique. The application of freshly prepared extract from 5g of fresh garlic (1% final concentration) decreased transmembrane currents of ENaC-expressing oocytes within 10 min. This effect was dose-dependent and irreversible. It was fully sensitive to the ENaC-inhibitor amiloride and was not apparent on native control oocytes. The effect of garlic was blocked by dithiothreitol and l-cysteine indicating involvement of thiol-reactive compounds. The garlic organosulsur compounds S-allylcysteine, alliin and diallyl sulfides had no effect on ENaC. By contrast, the thiol-reactive garlic compound allicin significantly inhibited ENaC to a similar extent as garlic extract. These data indicate that thiol-reactive compounds which are present in garlic inhibit ENaC. PMID:22668601

  7. Requirements for heme and thiols for the nonenzymatic modification of nitrotyrosine

    Science.gov (United States)

    Balabanli, Barbaros; Kamisaki, Yoshinori; Martin, Emil; Murad, Ferid

    1999-01-01

    Peroxynitrite-dependent formation of nitrotyrosine has been associated with inactivation of various enzymes and proteins possessing functionally important tyrosines. We have previously reported an enzymatic activity modifying the nitrotyrosine residues in nitrated proteins. Here we are describing a nonenzymatic reduction of nitrotyrosine to aminotyrosine, which depends on heme and thiols. Various heme-containing proteins can mediate the reaction, although the reaction also is catalyzed by heme. The reaction is most effective when vicinal thiols are used as reducing agents, although ascorbic acid also can replace thiols with lesser efficiency. The reaction could be inhibited by (z)-1-[2-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1, but not other tested NO donors. HPLC with electrochemical detection analysis of the reaction identified aminotyrosine as the only reaction product. The reduction of nitrotyrosine was most effective at a pH close to physiological and was markedly decreased in acidic conditions. Various nitrophenol compounds also were modified in this reaction. Understanding the mechanism of this reaction could help define the enzymatic modification of nitrotyrosine-containing proteins. Furthermore, this also could assist in understanding the role of nitrotyrosine formation and reversal in the regulation of various proteins containing nitrotyrosine. It also could help define the role of nitric oxide and other reactive species in various disease states. PMID:10557286

  8. Kandelia obovata (S., L.) Yong tolerance mechanisms to Cadmium: Subcellular distribution, chemical forms and thiol pools

    International Nuclear Information System (INIS)

    Highlights: ? Cadmium tolerance mechanisms of Kandelia obovata was investigated systematacially. ? Thiol pool can play roles in cadmium detoxification mechanisms. ? Increasing cadmium treatment strength caused proportional increase of cadmium uptake. ? More than half of cadmium was localized in cell walls, and lowest in membranes. ? Sodium chloride and acetic acid extractable fractions were dominant. - Abstract: In order to explore the detoxification mechanisms adopted by mangrove under cadmium (Cd) stress, we investigated the subcellular distribution and chemical forms of Cd, in addition to the change of the thiol pools in Kandelia obovata (S., L.) Yong, which were cultivated in sandy culture medium treated with sequential Cd solution. We found that Cd addition caused a proportional increase of Cd in the organs of K. obovata. The investigation of subcellular distribution verified that most of the Cd was localized in the cell wall, and the lowest was in the membrane. Results showed sodium chloride and acetic acid extractable Cd fractions were dominant. The contents of non-protein thiol compounds, Glutathione and phytochelatins in K. obovata were enhanced by the increasing strength of Cd treatment. Therefore, K. obovata can be defined as Cd tolerant plant, which base on cell wall compartmentalization, as well as protein and organic acids combination.

  9. The critical role of the cellular thiol homeostasis in cadmium perturbation of the lung extracellular matrix

    International Nuclear Information System (INIS)

    Cadmium (Cd) inhalation can result in emphysema. Cd exposure of rat lung fibroblasts (RFL6) enhanced levels of metal scavenging thiols, e.g., metallothionein (MT) and glutathione (GSH), and the heavy chain of ?-glutamylcysteine synthetase (?-GCS), a key enzyme for GSH biosynthesis, concomitant with downregulation of lysyl oxidase (LO), a copper-dependent enzyme for crosslinking collagen and elastin in the extracellular matrix (ECM). Cd downregulation of LO in treated cells was closely accompanied by suppression of synthesis of collagen, a major structure component of the lung ECM. Using rats intratracheally instilled with cadmium chloride (30 ?g, once a week) as an animal model, we further demonstrated that although 2-week Cd instillation induced a non-significant change in the lung LO activity and collagen synthesis, 4- and 6-week Cd instillation resulted in a steady decrease in the lung LO and collagen expression. The lung MT and total GSH levels were both upregulated upon the long-term Cd exposure. Emphysematous lesions were generated in lungs of 6-week Cd-dosed rats. Increases of cellular thiols by transfection of cells with MT-II expression vectors or treatment of cells with GSH monoethyl ester, a GSH delivery system, markedly inhibited LO mRNA levels and catalytic activities in the cell model. Thus, Cd upregulation of cellular thiols may be a critical cellular event facilitating downregulation of LO, a potential mechanism for Cd-induced emphysema.

  10. Thiol modified chitosan self-assembled monolayer platform for nucleic acid biosensor.

    Science.gov (United States)

    Mukherjee, Maumita Das; Solanki, Pratima R; Sumana, Gajjala; Manaka, Takaaki; Iwamoto, Mitsumasa; Malhotra, Bansi D

    2014-10-01

    A self-assembled monolayer (SAM) of thiol modified chitosan (SH-CHIT), with thioglycolic acid (TGA) as a modifier to bestow thiol groups, has been prepared onto gold (Au)-coated glass plates for fabrication of the nucleic acid biosensor. The chemical modification of CHIT via TGA has been evidenced by Fourier transform infrared spectroscopy (FT-IR) studies, and the biocompatibility studies reveal that CHIT retains its biocompatible nature after chemical modification. The electrochemical studies conducted onto SH-CHIT/Au electrode reveal that thiol modification in CHIT amino end enhances the electrochemical behavior indicating that it may be attributed to delocalization of electrons in CHIT skeleton that participates in the resonance process. The carboxyl group modified end of DNA probe has been immobilized onto SH-CHIT/Au electrode using N-ethyl-N'-(3-dimethylaminopropyl)carbodimide (EDC) and N-hydroxysuccinimide (NHS) chemistry for detection of complementary, one-base mismatch and non-complementary sequence using electrochemical and optical studies for Mycobacterium tuberculosis detection. It has been found that DNA-SH-CHIT/Au bioelectrode can specifically detect 0.01?M of target DNA concentration with sensitivity of 1.69??10(-6)A?M(-1). PMID:25205172

  11. A Novel Strategy for Global Analysis of the Dynamic Thiol Redox Proteome*

    Science.gov (United States)

    Martnez-Acedo, Pablo; Nez, Estefana; Gmez, Francisco J. Snchez; Moreno, Margoth; Ramos, Elena; Izquierdo-lvarez, Alicia; Mir-Casas, Elisabet; Mesa, Raquel; Rodriguez, Patricia; Martnez-Ruiz, Antonio; Dorado, David Garcia; Lamas, Santiago; Vzquez, Jess

    2012-01-01

    Nitroxidative stress in cells occurs mainly through the action of reactive nitrogen and oxygen species (RNOS) on protein thiol groups. Reactive nitrogen and oxygen species-mediated protein modifications are associated with pathophysiological states, but can also convey physiological signals. Identification of Cys residues that are modified by oxidative stimuli still poses technical challenges and these changes have never been statistically analyzed from a proteome-wide perspective. Here we show that GELSILOX, a method that combines a robust proteomics protocol with a new computational approach that analyzes variance at the peptide level, allows a simultaneous analysis of dynamic alterations in the redox state of Cys sites and of protein abundance. GELSILOX permits the characterization of the major endothelial redox targets of hydrogen peroxide in endothelial cells and reveals that hypoxia induces a significant increase in the status of oxidized thiols. GELSILOX also detected thiols that are redox-modified by ischemia-reperfusion in heart mitochondria and demonstrated that these alterations are abolished in ischemia-preconditioned animals. PMID:22647871

  12. A novel strategy for global analysis of the dynamic thiol redox proteome.

    Science.gov (United States)

    Martnez-Acedo, Pablo; Nez, Estefana; Gmez, Francisco J Snchez; Moreno, Margoth; Ramos, Elena; Izquierdo-lvarez, Alicia; Mir-Casas, Elisabet; Mesa, Raquel; Rodriguez, Patricia; Martnez-Ruiz, Antonio; Dorado, David Garcia; Lamas, Santiago; Vzquez, Jess

    2012-09-01

    Nitroxidative stress in cells occurs mainly through the action of reactive nitrogen and oxygen species (RNOS) on protein thiol groups. Reactive nitrogen and oxygen species-mediated protein modifications are associated with pathophysiological states, but can also convey physiological signals. Identification of Cys residues that are modified by oxidative stimuli still poses technical challenges and these changes have never been statistically analyzed from a proteome-wide perspective. Here we show that GELSILOX, a method that combines a robust proteomics protocol with a new computational approach that analyzes variance at the peptide level, allows a simultaneous analysis of dynamic alterations in the redox state of Cys sites and of protein abundance. GELSILOX permits the characterization of the major endothelial redox targets of hydrogen peroxide in endothelial cells and reveals that hypoxia induces a significant increase in the status of oxidized thiols. GELSILOX also detected thiols that are redox-modified by ischemia-reperfusion in heart mitochondria and demonstrated that these alterations are abolished in ischemia-preconditioned animals. PMID:22647871

  13. Brain PP2A is modified by thiol-disulfide exchange and intermolecular disulfide formation.

    Science.gov (United States)

    Foley, Timothy D; Kintner, Marissa E

    2005-05-20

    The regulation of protein phosphatase 2A (PP2A) activity by thiol-disulfide exchange and resulting formation of an intermolecular disulfide was examined following exposure of a rat brain soluble fraction to a biotinylated derivative of the model disulfide HPDP (HPDP-biotin) which would be expected to label reactive protein thiols with a disulfide-linked biotin. The results show that a low concentration (500 microM) of HPDP-biotin produced substantial inhibition of PP2A activity and promoted the binding of the catalytic subunit of PP2A to an immobilized avidin-affinity column. Both the inhibition of PP2A activity and the binding of PP2A to the avidin column were reversed by treatment with the disulfide reducing agent dithiothreitol (DTT). Furthermore, the specific activity of PP2A was up to 7-fold higher in the DTT-eluted fractions from the avidin-affinity column than in the soluble fraction. These findings demonstrate directly that PP2A is susceptible to reversible inhibitory modification by thiol-disulfide exchange and provide mechanistic support for the emerging view that PP2A is an oxidant-sensitive protein phosphatase. PMID:15823574

  14. Characterization of plasma thiol redox potential in a common marmoset model of aging

    Directory of Open Access Journals (Sweden)

    James R. Roede

    2013-01-01

    Full Text Available Due to its short lifespan, ease of use and age-related pathologies that mirror those observed in humans, the common marmoset (Callithrix jacchus is poised to become a standard nonhuman primate model of aging. Blood and extracellular fluid possess two major thiol-dependent redox nodes involving cysteine (Cys, cystine (CySS, glutathione (GSH and glutathione disulfide (GSSG. Alteration in these plasma redox nodes significantly affects cellular physiology, and oxidation of the plasma Cys/CySS redox potential (EhCySS is associated with aging and disease risk in humans. The purpose of this study was to determine age-related changes in plasma redox metabolites and corresponding redox potentials (Eh to further validate the marmoset as a nonhuman primate model of aging. We measured plasma thiol redox states in marmosets and used existing human data with multivariate adaptive regression splines (MARS to model the relationships between age and redox metabolites. A classification accuracy of 70.2% and an AUC of 0.703 were achieved using the MARS model built from the marmoset redox data to classify the human samples as young or old. These results show that common marmosets provide a useful model for thiol redox biology of aging.

  15. Ethyl propiolate derivatisation for the analysis of varietal thiols in wine.

    Science.gov (United States)

    Herbst-Johnstone, Mandy; Piano, Federico; Duhamel, Nina; Barker, David; Fedrizzi, Bruno

    2013-10-18

    Varietal thiols [3-mercaptohexan-1-ol (3MH), 3-mercaptohexyl acetate (3MHA) and 4-mercapto-4-methylpentan-2-one (4MMP)] have been extensively studied in the recent literature. Nonetheless the hardest obstacle for research focussing on this class of compounds is the lack of quick, user-friendly and sensitive analytical methods. The current paper presents the use of ethyl propriolate (ETP) as a novel derivatising agent to quantify varietal thiols and the first time quantification of the thiol-ETP adducts via gas chromatography-mass spectrometry. Method optimisation including choice of the best SPE cartridge, derivatisation pH and adducts stability is presented. Validation of the method via stable isotope dilution was carried out. Detection limits in both model wine (4MMP 7.2ng/L, 3MHA 40.0ng/L and 3MH 91.2ng/L) and white wine (4MMP 24.5ng/L, 3MHA 120.9ng/L and 3MH 194.6ng/L) for the novel ETP-based method were lower than those obtained with the p-HMB method. Finally, 14 New Zealand Sauvignon blanc were analysed with both the new method and the organo-mercury based procure: good correlations were obtained for 3MH and 3MHA. Detection limits obtained with the new methods, its rapidity and reproducibility make this protocol perfectly suitable for oenological purposes. PMID:24034138

  16. Pyridine Nucleotide Complexes with Bacillus anthracis Coenzyme A-Disulfide Reductase: A Structural Analysis of Dual NAD(P)H Specificity

    Energy Technology Data Exchange (ETDEWEB)

    Wallen,J.; Paige, C.; Mallett, T.; Karplus, P.; Claiborne, A.

    2008-01-01

    We have recently reported that CoASH is the major low-molecular weight thiol in Bacillus anthracis, and we have now characterized the kinetic and redox properties of the B. anthracis coenzyme A-disulfide reductase (CoADR, BACoADR) and determined the crystal structure at 2.30 Angstroms resolution. While the Staphylococcus aureus and Borrelia burgdorferi CoADRs exhibit strong preferences for NADPH and NADH, respectively, B. anthracis CoADR can use either pyridine nucleotide equally well. Sequence elements within the respective NAD(P)H-binding motifs correctly reflect the preferences for S. aureus and Bo. burgdorferi CoADRs, but leave questions as to how BACoADR can interact with both pyridine nucleotides. The structures of the NADH and NADPH complexes at ca. 2.3 Angstroms resolution reveal that a loop consisting of residues Glu180-Thr187 becomes ordered and changes conformation on NAD(P)H binding. NADH and NADPH interact with nearly identical conformations of this loop; the latter interaction, however, involves a novel binding mode in which the 2'-phosphate of NADPH points out toward solvent. In addition, the NAD(P)H-reduced BACoADR structures provide the first view of the reduced form (Cys42-SH/CoASH) of the Cys42-SSCoA redox center. The Cys42-SH side chain adopts a new conformation in which the conserved Tyr367'-OH and Tyr425'-OH interact with the nascent thiol(ate) on the flavin si-face. Kinetic data with Y367F, Y425F, and Y367, 425F BACoADR mutants indicate that Tyr425' is the primary proton donor in catalysis, with Tyr367' functioning as a cryptic alternate donor in the absence of Tyr425'.

  17. Metabolism of the Synthetic Progestogen Norethynodrel by Human Ketosteroid Reductases of the Aldo-Keto Reductase Superfamily

    OpenAIRE

    Jin, Yi; Duan, Ling; Chen, Mo; Penning, Trevor M.; Kloosterboer, Helenius J.

    2011-01-01

    Human ketosteroid reductases of the aldo-keto reductase (AKR) superfamily, i.e. AKR1C1-4, are implicated in the biotransformation of synthetic steroid hormones. Norethynodrel (NOR, 17?-ethynyl-17?-hydroxy-estra-5(10)-en-3-one), the progestin component of the first marketed oral contraceptive, is known to undergo rapid and extensive metabolism to 3?- and 3?-hydroxy metabolites. The ability of the four human AKR1C enzymes to catalyze the metabolism of NOR has now been characterized. AKR1C1 ...

  18. The novel disulfide reductase bis-gamma-glutamylcystine reductase and dihydrolipoamide dehydrogenase from Halobacterium halobium: purification by immobilized-metal-ion affinity chromatography and properties of the enzymes.

    OpenAIRE

    Sundquist, A. R.; Fahey, R. C.

    1988-01-01

    An NADPH-specific disulfide reductase that is active with bis-gamma-glutamylcystine has been purified 1,900-fold from Halobacterium halobium to yield a homogeneous preparation of the enzyme. Purification of this novel reductase, designated bis-gamma-glutamylcystine reductase (GCR), and purification of halobacterial dihydrolipoamide dehydrogenase (DLD) were accomplished with the aid of immobilized-metal-ion affinity chromatography in high-salt buffers. Chromatography of GCR on immobilized Cu2+...

  19. Chimeric 3-hydroxy-3-methylglutaryl coenzyme A reductase-dihydrofolate reductase genes display bidirectional expression and unidirectional regulation in stably transfected cells.

    OpenAIRE

    Abrams, J. M.; Schimke, R. T.

    1989-01-01

    We have constructed hybrid dihydrofolate reductase (DHFR) genes which are controlled by the sterol-responsive hamster 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase promoter. Stable transfection frequencies of these chimeric templates into a DHFR-deficient Chinese hamster cell line indicate that the HMG CoA reductase promoter fragment confers DHFR transformation irrespective of its orientation relative to a downstream murine DHFR cDNA. Sterol-regulated levels of DHFR RNA and protei...

  20. POTENTIOMETRIC RESPONSE OF A GRAPHITE ELECTRODE MODIFIED WITH COBALT PHTHALOCYANINE FOR THIOLS AND DISULFIDES

    Scientific Electronic Library Online (English)

    JOS H., ZAGAL; JAIME J.H., HENRIQUEZ.

    2000-06-01

    Full Text Available Se ha investigado la respuesta potenciomtrica the electrodos de grafito piroltico ordinario (OPG) modificados con ftalocianina de cobalto (Co-Pc) frente a los tioles (R-SH) 2-mecaptoetanol y L-cistena y los correspondientes disulfuros (R-SS-R). Al agregar pequeas cantidades de estos tioles a sol [...] uciones acuosas con pH en el rango 11-4, se obtienen respuestas en potencial estables dentro de unos pocos segundos despues de las adiciones. Grficas de potencial vs. log [R-SH] dan rectas para todos los casos, con pendientes cercanas a -0,060 V para concentraciones del tiol en el rango de 10-5 a 10-2 M. Estas medidas se efectuaron en presencia de aire u oxgeno. En atmsfera de nitrgeno, las pendientes de las grficas suben a -0,14 V. El potencial de circuito abierto de los electrodos OPG/Co-Pc result ser independiente de la concentracin de los disulfuros lo que indica que el potencial del electrodo modificado es slo sensible al tiol. Los electrodos de grafito sin modificar frente a los tioles dan respuestas en potencial irreproducibles, lo que muestra la importancia de la presencia Co-Pc para obtener respuestas potenciomtricas estables Abstract in english We have investigated the potentiometric response of ordinary pyrolytic graphite electrodes (OPG) modified with cobalt phthalocyanine (Co-Pc) for thiols (R-SH) 2-mercaptoethanol, L-cysteine and their corresponding disulfides (R-SS-R). Stable potentials are achieved after a few seconds of additions of [...] different amounts of thiols to aqueous solutions of pH values between 11 and 4. Plots of potential vs. log [R-SH] give straight lines for all cases with slopes ca. -0.060 V with concentrations of the thiol varying from 10-5 up to 10-2 M. These measurements were conducted in the presence of air or oxygen. Under nitrogen, the slopes increase to -0.14 V. The potential response of OPG/Co-Pc is independent of disulfide concentration, which shows that these modified electrodes are only sensitive to the thiol. Graphite electrodes without modification are not sensitive to the concentration of thiols in the solution and the potential response is erratic.

  1. POTENTIOMETRIC RESPONSE OF A GRAPHITE ELECTRODE MODIFIED WITH COBALT PHTHALOCYANINE FOR THIOLS AND DISULFIDES

    Directory of Open Access Journals (Sweden)

    JOS H. ZAGAL

    2000-06-01

    Full Text Available We have investigated the potentiometric response of ordinary pyrolytic graphite electrodes (OPG modified with cobalt phthalocyanine (Co-Pc for thiols (R-SH 2-mercaptoethanol, L-cysteine and their corresponding disulfides (R-SS-R. Stable potentials are achieved after a few seconds of additions of different amounts of thiols to aqueous solutions of pH values between 11 and 4. Plots of potential vs. log [R-SH] give straight lines for all cases with slopes ca. -0.060 V with concentrations of the thiol varying from 10-5 up to 10-2 M. These measurements were conducted in the presence of air or oxygen. Under nitrogen, the slopes increase to -0.14 V. The potential response of OPG/Co-Pc is independent of disulfide concentration, which shows that these modified electrodes are only sensitive to the thiol. Graphite electrodes without modification are not sensitive to the concentration of thiols in the solution and the potential response is erratic.Se ha investigado la respuesta potenciomtrica the electrodos de grafito piroltico ordinario (OPG modificados con ftalocianina de cobalto (Co-Pc frente a los tioles (R-SH 2-mecaptoetanol y L-cistena y los correspondientes disulfuros (R-SS-R. Al agregar pequeas cantidades de estos tioles a soluciones acuosas con pH en el rango 11-4, se obtienen respuestas en potencial estables dentro de unos pocos segundos despues de las adiciones. Grficas de potencial vs. log [R-SH] dan rectas para todos los casos, con pendientes cercanas a -0,060 V para concentraciones del tiol en el rango de 10-5 a 10-2 M. Estas medidas se efectuaron en presencia de aire u oxgeno. En atmsfera de nitrgeno, las pendientes de las grficas suben a -0,14 V. El potencial de circuito abierto de los electrodos OPG/Co-Pc result ser independiente de la concentracin de los disulfuros lo que indica que el potencial del electrodo modificado es slo sensible al tiol. Los electrodos de grafito sin modificar frente a los tioles dan respuestas en potencial irreproducibles, lo que muestra la importancia de la presencia Co-Pc para obtener respuestas potenciomtricas estables

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

    DEFF Research Database (Denmark)

    Javakhishvili, Irakli; Hvilsted, Sren

    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, subsequent chain-extension by atom transfer radical polymerization (ATRP) of tert-butyl acrylate (tBA), and final deprotection steps. ROP of c-CL initiated by 2-hydroxyethyl 2-bromoisobutyrate, and catalysed by tin octoate afforded Br-PCL-OH with the degree of polymerization of 30 and narrow molecular weight distribution (1.09). The hydroxy chain end of Br-PCL-OR was modified by reacting with a-(2,4-dinitrophenylthio)acetic acid or a-(4methoxytritylthio) acetic acid resulting in heterotelechelic PCL incorporating protected thiol and bromoester functionalities. It was then employed as macroinitiator in NiBr2(PPh3)2 catalysed ATRP of tBA. ATRP of tBA provided diblock copolymers with low polydispersity index (1.17-1.39) while preserving the protected thiol function. Sequential or simultaneous removal of 2,4-dinitrophenyl or 4-methoxytrityl and tert-butyl ester groups resulted in HS-PCL-b-PAA. The PCL backbone remained intact after mild deprotection protocols. Thus, reversible masking of thiol functionality allows facile fusion of the controlled polymerization techniques employing dual initiator strategy, and furnishes RS-PCL-bPAA with well-defined chain architecture which has been assessed by size exclusion chromatography (SEC), nuclear magnetic resonance eR NMR) and infrared (FT IR) spectroscopy. The capacity of the resulting block copolymer in preparation of monolayer-protected gold nanoparticles has been examined by reduction of a gold salt in the presence of this macroligand under thiol-deficient conditions. As a result stable, aggregation-free nanopaticles with moderate dispersity as estimated from UV-visible spectroscopy and transmission electron microscopy (TEM) data were obtained.

  3. Influence of extra-cellular and intra-cellular acting thiol oxidants on the 45calcium uptake by the islets of Langerhans of the rat

    International Nuclear Information System (INIS)

    The glucose-stimulated calcium uptake by the islets of Langerhans is dependent on the intra-cellular GSH/GSSG ratios. The inhibition of calcium uptake is not the consequence of a direct oxidation of membrane-fixed thiol groups. In contrast, direct oxidation of extra cellular thiols leads to an increase in calcium uptake when intra-cellular oxidation is simultaneously prevented. Since this effect only occurs at high intra-cellular GSH/GSSG ratios it can be assumed that the redox state of extra-cellular thiols is dependent on the redox state of the intra-cellular GSH/GSSG ratios. These findings support the theory that the oxidation of extra-cellular thiols by thiol oxidants leads to an increase in calcium uptake and that the extent of uptake is higher, the more the redox state of the extra-cellular thiols tends towards the reduced state prior to oxidation. (orig./MG)

  4. Feedback Regulation of 3-Hydroxy-3-Methylglutaryl Coenzyme A Reductase in Livers of Mice Treated with Mevinolin, a Competitive Inhibitor of the Reductase

    OpenAIRE

    Kita, Toru; Brown, Michael S.; Goldstein, Joseph L.

    1980-01-01

    Compactin (ML-236B) and the related compound, mevinolin, are competitive inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG CoA reductase), the rate-controlling enzyme in cholesterol synthesis. Previous studies have shown that administration of compactin to cultured cells elicits a compensatory increase in the amount of HMG CoA reductase in the cells. A similar increase in HMG CoA reductase has been reported in livers of rats and mice that have been treated with compactin. In ...

  5. Sucrose mimics the light induction of Arabidopsis nitrate reductase gene transcription

    DEFF Research Database (Denmark)

    Cheng, Chi-Lien; Acedo, Gregoria N

    1992-01-01

    Nitrate reductase, the first enzyme in nitrate assimilation, is located at the crossroad of two energy-consuming pathways: nitrate assimilation and carbon fixation. Light, which regulates the expression of many higher-plant carbon fixation genes, also regulates nitrate reductase gene expression. Located in the cytosol, nitrate reductase obtains its reductant not from photosynthesis but from carbohydrate catabolism. This relationship prompted us to investigate the indirect role that light might play, via photosynthesis, in the regulation of nitrate reductase gene expression. We show that sucrose can replace light in eliciting an increase of nitrate reductase mRNA accumulation in dark-adapted green Arabidopsis plants. We show further that sucrose alone is sufficient for the full expression of nitrate reductase genes in etiolated Arabidopsis plants. Finally, using a reporter gene, we show that a 2.7-kilobase region of 5' flanking sequence of the nitrate reductase gene is sufficient to confer the light or the sucrose response.

  6. Aldo-keto reductases as modulators of stress response.

    Science.gov (United States)

    Chang, Qing; Harter, Theresa M; Rikimaru, Loryn T; Petrash, J Mark

    2003-02-01

    Human aldose reductase (AKR1B1) has been implicated as a factor in the pathogenesis of diabetic complications. However, little is known about the physiological role of this enzyme or of related aldo-keto reductases in human tissues. In mammalian systems, a gene knock out approach is often employed as an experimental strategy to probe for gene function. However, in the murine system, phenotypic characterization of an aldose reductase (AKR1B3) knock out is likely to be complicated due to functional compensation by redundant AKRs including AKRs 1A (aldehyde reductase), 1B7 (FR-1) and 1B8 (MVDP). As an alternate strategy, we are examining the budding yeast Saccharomyces cerevisiae as a model system for a functional genomics study of AKRs. A distinct advantage of this system centers on the ability to readily ablate multiple targeted genes in a single strain. In addition to providing insights into functional redundancy, this system allows us to use a genetic approach to study possible effector pathways associated with one or more individual genes. Yeast open reading frames (ORFs) encoding AKRs with functional similarity to human aldose reductase (AKR1B1) were identified by BLAST analysis and were functionally validated by studies of recombinant proteins. By ablating three of the yeast AKR genes most functionally similar to AKR1B1, we have created a unique strain of S. cerevisiae that shows enhanced sensitivity to stress. Ongoing studies with oligonucleotide arrays show that the triple null strain has an altered transcription profile consistent with an enhanced stress response in comparison with the parental strain. These data indicate that AKR-null strains may provide new insights into signaling mechanisms involving this family of proteins. PMID:12604219

  7. Characterization of streptavidin binding to biotinylated, binary self-assembled thiol monolayers--influence of component ratio and solvent.

    Science.gov (United States)

    Seifert, Michael; Rinke, Matthias T; Galla, Hans-Joachim

    2010-05-01

    Many biosensor applications are based on streptavidin (SA) binding to partially biotinylated self-assembled thiol monolayers (SAMs). In our study, binary SAMs on gold were prepared from solutions containing 16-mercapto-1-hexadecanol (thiol I) and N-(8-biotinyl-3,6-dioxa-octanamidyl)-16-mercaptohexadecanamide (thiol II) in varying component ratios. Either chloroform or ethanol was used as solvent. After 24 h thiol incubation, SA was immobilized on the resulting SAMs using the strong SA-biotin interaction. The SA binding process was monitored by QCM-D (quartz crystal microbalance monitoring dissipation factor). It is shown that the Sauerbrey equation is valid to calculate the mass quantities of the immobilized SA layers. Under the chosen incubation conditions, marginal fractions of the biotinylated component II in chloroform ((n(I)/n(II))(solution) approximately = 1000) lead to SAMs which ensure a maximal SA binding quantity of m(Sauerbrey SA) approximately = 400 ng x cm(-2), being equivalent to a SA single-layer arrangement on the SAM surface. In case of incubations from ethanolic solutions, a complete SA layer formation needs significantly higher amounts of the biotinylated component II during SAM preparation ((n(I)/n(II))(solution) approximately = 50). X-ray photoelectron spectroscopy data show that the fraction of biotinylated thiol II in the SAM determines the amount of surface-bound SA. The SAM thiol ratio ((n(I)/n(II))(SAM)) not only depends on the corresponding component ratio in the incubation solution, but is also strongly influenced by the solvent. Using chloroform as solvent during SAM preparation significantly increased the fraction of biotinylated thiol II in the SAMs compared to ethanol. PMID:20158172

  8. The Polymorphisms in Methylenetetrahydrofolate Reductase, Methionine Synthase, Methionine Synthase Reductase, and the Risk of Colorectal Cancer

    Directory of Open Access Journals (Sweden)

    Daijun Zhou, Qiang Mei, Han Luo, Bo Tang, Peiwu Yu

    2012-01-01

    Full Text Available Polymorphisms in genes involved in folate metabolism may modulate the risk of colorectal cancer (CRC, but data from published studies are conflicting. The current meta-analysis was performed to address a more accurate estimation. A total of 41 (17,552 cases and 26,238 controls, 24(8,263 cases and 12,033 controls, 12(3,758 cases and 5,646 controls, and 13 (5,511 cases and 7,265 controls studies were finally included for the association between methylenetetrahydrofolate reductase (MTHFR C677T and A1289C, methione synthase reductase (MTRR A66G, methionine synthase (MTR A2756G polymorphisms and the risk of CRC, respectively. The data showed that the MTHFR 677T allele was significantly associated with reduced risk of CRC (OR = 0.93, 95%CI 0.90-0.96, while the MTRR 66G allele was significantly associated with increased risk of CRC (OR = 1.11, 95%CI 1.01-1.18. Sub-group analysis by ethnicity revealed that MTHFR C677T polymorphism was significantly associated with reduced risk of CRC in Asians (OR = 0.80, 95%CI 0.72-0.89 and Caucasians (OR = 0.84, 95%CI 0.76-0.93 in recessive genetic model, while the MTRR 66GG genotype was found to significantly increase the risk of CRC in Caucasians (GG vs. AA: OR = 1.18, 95%CI 1.03-1.36. No significant association was found between MTHFR A1298C and MTR A2756G polymorphisms and the risk of CRC. Cumulative meta-analysis showed no particular time trend existed in the summary estimate. Probability of publication bias was low across all comparisons illustrated by the funnel plots and Egger's test. Collectively, this meta-analysis suggested that MTHFR 677T allele might provide protection against CRC in worldwide populations, while MTRR 66G allele might increase the risk of CRC in Caucasians. Since potential confounders could not be ruled out completely, further studies were needed to confirm these results.

  9. Analysis of volatile thiols in alcoholic beverages by simultaneous derivatization/extraction and liquid chromatography-high resolution mass spectrometry.

    Science.gov (United States)

    Vichi, Stefania; Corts-Francisco, Nuria; Caixach, Josep

    2015-05-15

    A simultaneous derivatization/extraction method followed by liquid chromatography-electrospray-high resolution mass spectrometry for the determination of volatile thiols in hydroalcoholic matrixes was optimized and used to identify and quantify volatile thiols in wine and beer samples. The method was evaluated in terms of sensitivity, precision, accuracy and selectivity. The experimental LOQs of eleven thiols tested ranged between 0.01 ng/L and 10 ng/L. Intra-day relative standard deviation (RSD) was in general lower than 10% and inter-day RSD ranged between 10% and 30%. Recovery in the model and real matrixes ranged from 45% to 129%. The method was then applied for the analysis of four white wines and six beers. Five out of the eleven reference thiols were identified and quantified in the samples analyzed. The non-target approach, carried out by monitoring the diagnostic ion at m/z 275.9922 [C13H10ONSe](+) in the fragmentation spectrum, allowed detecting, in the same samples, fourteen non-target thiols. PMID:25577098

  10. Synthesis and Optical Properties of Thiol Functionalized CdSe/ZnS (Core/Shell) Quantum Dots by Ligand Exchange

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Huaping [ORNL; Hu, Michael Z. [ORNL; Shao, Lei [ORNL; Yu, Kui [SIMS, NRC of Canada; Dabestani, Reza T [ORNL; Zaman, Md. Badruz [SIMS, NRC of Canada; Liao, Dr. Shijun [South China University of Technology, Guangzhou, PR China

    2014-01-01

    The colloidal photoluminescent quantum dots (QDs) of CdSe (core) and CdSe/ZnS (core/shell) were synthesized at different temperatures with different growth periods. The optical properties (i.e., UV/Vis spectra and photoluminescent emission spectra) of the resulting QDs were investigated. The CdSe/ZnS QDs exhibited higher photoluminescent (PL) efficiency and stability than their corresponding CdSe core QDs. Ligand exchange with various thiol molecules was performed to replace the initial surface passivation ligands, that is, trioctylphosphine oxide (TOPO) and trioctylphosphine (TOP), and the optical properties of the surface-modified QDs were studied. The thiol ligand molecules used included 1,4-benzenedimethanethiol, 1,16-hexadecanedithiol, 1,11-undecanedithiol, 11-mercapto-1-undecanol, and 1,8 octanedithiol. After the thiol functionalization, the CdSe/ZnS QDs exhibited significantly enhanced PL efficiency and storage stability. Besides surface passivation effect, such enhanced performance of thiol-functionalized QDs could be due to self-assembly formation of dimer/trimer clusters, in which QDs are linked by dithiol molecules. Effects of ligand concentration, type of ligand, and heating on the thiol stabilization of QDs were also discussed.

  11. Facile construction of macroporous hybrid monoliths via thiol-methacrylate Michael addition click reaction for capillary liquid chromatography.

    Science.gov (United States)

    Lin, Hui; Ou, Junjie; Liu, Zhongshan; Wang, Hongwei; Dong, Jing; Zou, Hanfa

    2015-01-30

    A facile approach based on thiol-methacrylate Michael addition click reaction was developed for construction of porous hybrid monolithic materials. Three hybrid monoliths were prepared via thiol-methacrylate click polymerization by using methacrylate-polyhedral oligomeric silsesquioxane (POSS) (cage mixture, n=8, 10, 12, POSS-MA) and three multi-thiol crosslinkers, 1,6-hexanedithiol (HDT), trimethylolpropane tris(3-mercaptopropionate) (TPTM) and pentaerythritol tetrakis(3-mercaptopropionate) (PTM), respectively, in the presence of porogenic solvents (n-propanol and PEG 200) and a catalyst (dimethylphenylphosphine, DMPP). The obtained monoliths possessed high thermal and chemical stabilities. Besides, they all exhibited high column efficiencies and excellent separation abilities in capillary liquid chromatography (cLC). The highest column efficiency could reach ca. 195,000N/m for butylbenzene on the monolith prepared with POSS-MA and TPTM (monolith POSS-TPTM) in reversed-phase (RP) mode at 0.64mm/s. Good chromatographic performance were all achieved in the separations of polycyclic aromatic hydrocarbons (PAHs), phenols, anilines, EPA 610 as well as bovine serum albumin (BSA) digest. The high column efficiencies in the range of 51,400-117,000N/m (achieved on the monolith POSS-PTM in RP mode) convincingly demonstrated the high separation abilities of these thiol-methacrylate based hybrid monoliths. All the results demonstrated the feasibility of the phosphines catalyzed thiol-methacrylate Michael addition click reaction in fabrication of monolithic columns with high efficiency for cLC applications. PMID:25543303

  12. Activation of room temperature ferromagnetism in ZnO films by surface functionalization with thiol and amine

    International Nuclear Information System (INIS)

    Highlights: ? Room temperature ferromagnetism (RTFM) is observed in surface functionalized ZnO films. ? Surface functionalization is a new approach to make ZnO as ferromagnetic. ? The RTFM is attributed to the interaction between the adsorbates and the surface of ZnO. ? The oxygen vacancies are passivated upon surface functionalization. - Abstract: In this paper, we report the activation of room temperature ferromagnetism in ZnO films by surface functionalization with thiol and amine. The pure and surface functionalized ZnO films have been examined by X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), photoluminescence (PL) and vibrating sample magnetometer (VSM) measurements. XRD measurements show that all the films have single phase and (0 0 2) preferred orientation. The chemical bonding of ZnO with thiol and amine molecules has been confirmed by XPS measurements. The quenching of visible emission in PL spectra indicates that the surface defects are passivated by functionalization with thiol and amine. Surface functionalization of ZnO films with thiol and amine induces robust room temperature ferromagnetism in ZnO films as evidenced from VSM measurements. It is concluded that the observed ferromagnetic behavior in functionalized ZnO films is attributed to the different electronegativity of the atom in the thiol (or amine) and the surface of ZnO.

  13. Advantages and drawbacks of Thiol-ene based resins for 3D-printing

    Science.gov (United States)

    Leonards, Holger; Engelhardt, Sascha; Hoffmann, Andreas; Pongratz, Ludwig; Schriever, Sascha; Blsius, Jana; Wehner, Martin; Gillner, Arnold

    2015-03-01

    The technology of 3D printing is conquering the world and awakens the interest of many users in the most varying of applications. New formulation approaches for photo-sensitive thiol-ene resins in combination with various printing technologies, like stereolithography (SLA), projection based printing/digital light processing (DLP) or two-photon polymerization (TPP) are presented. Thiol-ene polymerizations are known for its fast and quantitative reaction and to form highly homogeneous polymer networks. As the resins are locally and temporally photo-curable the polymerization type is very promising for 3D-printing. By using suitable wavelengths, photoinitiator-free fabrication is feasible for single- and two photon induced polymerization. In this paper divinyl ethers of polyethylene glycols in combination with star-shaped tetrathiols were used to design a simple test-system for photo-curable thiol-ene resins. In order to control and improve curing depth and lateral resolution in 3D-polymerization processes, either additives in chemical formulation or process parameters can be changed. The achieved curing depth and resolution limits depend on the applied fabrication method. While two-/multiphoton induced lithography offers the possibility of micron- to sub-micron resolution it lacks in built-up speed. Hence single-photon polymerization is a fast alternative with optimization potential in sub-10-micron resolution. Absorber- and initiator free compositions were developed in order to avoid aging, yellowing and toxicity of resulting products. They can be cured with UV-laser radiation below 300 nm. The development at Fraunhofer ILT is focusing on new applications in the field of medical products and implants, technical products with respect to mechanical properties or optical properties of 3D-printed objects. Recent process results with model system (polyethylene glycol divinylether/ Pentaerithrytol tetrakis (3-mercaptopropionat), Raman measurements of polymer conversion and surface modifications using bifunctional crosslinkers are presented with advantages, drawbacks and a general outlook.

  14. Endogenous thiols and cellular radiosensitivity in heterogeneous murine mammary carcinoma cells in vitro

    International Nuclear Information System (INIS)

    The cellular radiation response of the 66 vs 67 cells is different between the lines in three different physiological states [i.e., proliferation (P), transition between P and Q/sub I/ cells (T), and nutrient-deprived quiescence (Q/sub I/)]. For example, the D/sub o/'s and D/sub q/'s of the 66 P vs Q/sub I/ cells is 109 +- 6 and 368 +- 58 vs 90 +- 7 and 150 +- 43 rad respectively, while the D/sub o/'s and D/sub q/'s of the 67 P vs Q/sub I/ cells is 90 +- 4 and 250 +- 31 vs 52 +- 3 and 188 +- 28 rad respectively. In an attempt to characterize the mechanisms underlying these radiation responses, the authors measured the cellular thiol levels (i.e., total TSH; protein, PSH; and nonprotein sulfhydrls, NPSH). The TSH and the PSH levels/sup +/ are both relatively constant amongst the physiological states within a cell line and not dramatically different between cell lines with a range of 6.5 - 10 for TSH and 5 - 8 for PSH. However, the NPSH values do show considerable variability with a low of 0.8 for 66 Q/sub I/ vs 3.0 for P cells and values of 2.5 vs 4.0 for the 67 Q/sub I/ vs P cells. These data strongly suggest that there is not a simple relationship between the endogeneous thiol levels and cellular radiosensitivity neither between the cell lines within a given physiological state nor amongst the various physiological states within a cell line. +All thiol levels are ?g SH/mg protein

  15. Iron and thiols as two major players in carcinogenesis: friends or foes?

    Directory of Open Access Journals (Sweden)

    ShinyaToyokuni

    2014-08-01

    Full Text Available Iron is the most abundant metal in the human body and mainly works as a cofactor for proteins such as hemoglobin and various enzymes. No independent life forms on earth can survive without iron. However, excess iron is intimately associated with carcinogenesis by increasing oxidative stress via its catalytic activity to generate hydroxyl radicals. Biomolecules with redox-active sulfhydryl function(s (thiol compounds are necessary for the maintenance of mildly reductive cellular environments to counteract oxidative stress, and for the execution of redox reactions for metabolism and detoxification. Involvement of glutathione S-transferase and thioredoxin has long attracted the attention of cancer researchers. Here, I update recent findings on the involvement of iron and thiol compounds during carcinogenesis and in cancer cells. It is now recognized that the cystine/glutamate transporter (antiporter is intimately associated with ferroptosis, an iron-dependent, non-apoptotic form of cell death, observed in cancer cells, and also with cancer stem cells; the former with transporter blockage but the latter with its stabilization. Excess iron in the presence of oxygen appears the most common known mutagen. Ironically, the persistent activation of antioxidant systems via genetic alterations in Nrf2 and Keap1 also contributes to carcinogenesis. Therefore, it is difficult to conclude the role of iron and thiol compounds as friends or foes, which depends on the quantity/distribution and induction/flexibility, respectively. Avoiding further mutation would be the most helpful strategy for cancer prevention, and myriad of efforts are being made to sort out the weaknesses of cancer cells.

  16. Thiols as Effective Capping Molecules to Synthesize High-Quality ZnO Nanocrystals

    International Nuclear Information System (INIS)

    In our present work, an alternative approach to synthesize Zinc oxide (ZnO) quantum dots (QDs) using simple aqueous route under ambient conditions is reported. ZnO QDs were synthesized using zinc nitrate as a precursor and different thiols as effective capping molecules. The synthesized powders were characterized by XRD and UV-Vis optical absorption. The XRD results indicated that as synthesized ZnO QDs had pure hexagonal wurtzite structure and 2-ME capped ZnO QDs gives smallest sized nanoparticles. All these nanoparticles emit clearly visible blue emission under UV lamp.

  17. Characterization of thiol-functionalised silica films deposited on electrode surfaces

    Directory of Open Access Journals (Sweden)

    Ivana Cesarino

    2008-12-01

    Full Text Available Thiol-functionalised silica films were deposited on various electrode surfaces (gold, platinum, glassy carbon by spin-coating sol-gel mixtures in the presence of a surfactant template. Film formation occurred by evaporation induced self-assembly (EISA involving the hydrolysis and (cocondensation of silane and organosilane precursors on the electrode surface. The characterization of such material was performed by IR spectroscopy, thermogravimetry (TG, elemental analysis (EA, atomic force microscopy (AFM, scanning electron microscopy (SEM and cyclic voltammetry (CV.

  18. Tricarbonyl-Rhenium Complexes of a Thiol-FunctionalizedAmphoteric Poly(amidoamine)

    OpenAIRE

    BISAZZA, AGNESE

    2009-01-01

    An amphoteric thiol-functionalized poly(amidoamine) nicknamed ISA23SH10% was synthesized. Rhenium complexes 1 and 2, containing 0.5 and 0.8 equiv of rhenium, respectively, were easily obtained by reacting ISA23SH10% with [Re(CO)3(H2O)3](CF3SO3) in aqueous solution at pH 5.5. Both ISA23SH10%, and its rhenium complexes were soluble in water under physiological conditions. The resultant solutions were stable, even in the presence of cysteine. Rhenium chelation occurred through the S ...

  19. Gold electrode modified by self-assembled monolayers of thiols to determine DNA sequences hybridization

    OpenAIRE

    Silva, Mzia M. S.; Cavalcanti, Igor T.; Barroso, M. Ftima; M. Goreti F. Sales; Dutra, Rosa Fireman

    2010-01-01

    The process of immobilization of biological molecules is one of the most important steps in the construction of a biosensor. In the case of DNA, the way it exposes its bases can result in electrochemical signals to acceptable levels. The use of self-assembled monolayer that allows a connection to the gold thiol group and DNA binding to an aldehydic ligand resulted in the possibility of determining DNA hybridization. Immobilized single strand of DNA (ssDNA) from calf thymus pre-for...

  20. Considering Hydrophobicity via Contact Angle Stability of Organic Thiols Measured with a Homemade Goniometer

    Science.gov (United States)

    Seraly, Mark; Ollander, Brooke; Statman, Ariel; Poynor, Adele

    2014-03-01

    When water meets an extended hydrophobic surface, an ultra-thin, low-density depletion layer is expected at the interface. Exactly how the depletion layer changes with change in hydrophobicity is still an open question. An accurate measure of contact angle is essential in determining how water meets a hydrophobic surface. Utilizing a homemade goniometer with ImageJ software we investigate the stability of self-assembled organic thiol monolayers, 1-octadecanethiol (ODT) and 11-mercaptoundecanoic acid (MUA). We report the changes in contact angle due to exposure to air, water, and ethanol. Other factors that affect contact angles were also considered in our investigation.

  1. Characterization of thiol-functionalised silica films deposited on electrode surfaces

    Scientific Electronic Library Online (English)

    Ivana, Cesarino; der Tadeu Gomes, Cavalheiro.

    2008-12-01

    Full Text Available Thiol-functionalised silica films were deposited on various electrode surfaces (gold, platinum, glassy carbon) by spin-coating sol-gel mixtures in the presence of a surfactant template. Film formation occurred by evaporation induced self-assembly (EISA) involving the hydrolysis and (co)condensation [...] of silane and organosilane precursors on the electrode surface. The characterization of such material was performed by IR spectroscopy, thermogravimetry (TG), elemental analysis (EA), atomic force microscopy (AFM), scanning electron microscopy (SEM) and cyclic voltammetry (CV).

  2. Biochemical Characterization of a Thiol-Activated, Oxidation Stable Keratinase from Bacillus pumilus KS12

    OpenAIRE

    Rinky Rajput; Richa Sharma; Rani Gupta

    2010-01-01

    An extracellular keratinase from Bacillus pumilus KS12 was purified by DEAE ion exchange chromatography. It was a 45?kDa monomer as determined by SDS PAGE analysis. It was found to be an alkaline, serine protease with pH and temperature optima of 10 and 60C?, respectively. It was thiol activated with two- and eight-fold enhancement in presence of 10 mM DTT and ?-mercaptoethanol, respectively. In addition, its activity was stimulated in the presence of various surfactants,...

  3. Redox Characteristics of Thiol Compounds Using Radicals Produced by Water Vapor Radio Frequency Discharge

    Science.gov (United States)

    Hayashi, Nobuya; Nakahigashi, Akari; Goto, Masaaki; Kitazaki, Satoshi; Koga, Kazunori; Shiratani, Masaharu

    2011-08-01

    The redox reaction between cystein and cystine is observed using radicals produced in water vapor plasma for the control of plant growth. Cystein is oxidized to cystine using the OH radical in the higher-pressure regime and cystine is reduced to cystein by the H radical generated in the lower-pressure regime. Also, the oxidative stress reaction of plants is observed when water vapor plasma is irradiated onto seeds of plants such as radish sprouts. The mechanism of the control of plant growth is explained by the change in thiol compound quantity of the plant cells induced by the radical reaction.

  4. Thiols as interfacial modifiers to enhance the performance and stability of perovskite solar cells.

    Science.gov (United States)

    Cao, Jing; Yin, Jun; Yuan, Shangfu; Zhao, Yun; Li, Jing; Zheng, Nanfeng

    2015-06-01

    Modifying the interfaces of CH3NH3PbI3 with TiO2 and hole transport layers using two different types of thiols leads to enhanced performance and stability of perovskite solar cells. The incorporation of HOOC-Ph-SH at the TiO2/perovskite interface facilitates electron transfer from perovskite to TiO2 and also alters the morphology of perovskite crystal growth to increase the power conversion efficiency. The modification of pentafluorobenzenethiol at the perovskite/hole transport layer interface improves the stability. PMID:25964159

  5. A novel bifunctional maleimido CHX-A'' chelator for conjugation to thiol-containing biomolecules.

    Science.gov (United States)

    Xu, Heng; Baidoo, Kwamena E; Wong, Karen J; Brechbiel, Martin W

    2008-04-15

    A novel bifunctional maleimido CHX-A'' DTPA chelator 5 was developed and conjugated to the monoclonal antibody trastuzumab (Herceptin) and subsequently radiolabeled with (111)In. The resulting (111)In labeled immunoconjugate 2 was demonstrated to bind to SKOV-3 ovarian cancer cells comparably to an isothiocyanato CHX-A'' DTPA modified native trastuzumab, 1. Through efficient thiol-maleimide chemistry, antibodies, peptides or other targeting vectors can now be modified with an established radioactive metal chelating agent CHX-A'' DTPA for imaging and/or therapies of cancer. PMID:18359632

  6. Thiol-disulfide organization in alliin lyase (alliinase) from garlic (Allium sativum)

    OpenAIRE

    Weiner, Lev; Shin, Irina; Shimon, Linda J. W.; Miron, Talia; Wilchek, Meir; Mirelman, David; Frolow, Felix; Rabinkov, Aharon

    2008-01-01

    Alliinase, an enzyme found in garlic, catalyzes the synthesis of the well-known chemically and therapeutically active compound allicin (diallyl thiosulfinate). The enzyme is a homodimeric glycoprotein that belongs to the fold-type I family of pyridoxal-5?-phosphate-dependent enzymes. There are 10 cysteine residues per alliinase monomer, eight of which form four disulfide bridges and two are free thiols. Cys368 and Cys376 form a SS bridge located near the C-terminal and plays an important...

  7. Thiol-Functionalized Gold-Nanoscale Organic Hybrid Materials-Attractive to Soft glasses

    Science.gov (United States)

    Agrawal, Akanksha; Archer, Lynden

    2014-03-01

    We report on the flow properties of self-suspended nanoparticles based on gold nanoparticles densely grafted with polyethylene glycol methyl ether thiol(PEG) chains. We studied the effect of temperature, olume fraction and polymer chain length on the transition from attractive glass to soft glassy flow behavior. Gold nanoparticles densely grafted with short PEG-thiol chains(MW 800,2kDa and 6kDa)are shown to form self-suspended systems over a range of polymer grafting densities and particle volume fractions, ?.Transmission electron and atomic force microscopy measurements reveal that the particles are uniformly dispersed. Oscillatory shear measurements performed on low ? systems show a two-step yielding behavior reflecting bond breaking and cage breaking transitions at the nanoscale; both characteristics of soft glassy materials dominated by attractive forces. With increased temperature a transition to one-step yielding and subsequently back to two-step yielding is observed. At high ? a single yielding transition and soft glassy flow behavior are observed. We employ SAXS, vibration spectroscopy, thermal analysis, and rheology to interrogate the configuration state of the tethered chains and particle-particle interactions in detail. We report on the flow properties of self-suspended nanoparticles based on gold nanoparticles densely grafted with polyethylene glycol methyl ether thiol(PEG) chains. We studied the effect of temperature, olume fraction and polymer chain length on the transition from attractive glass to soft glassy flow behavior. Gold nanoparticles densely grafted with short PEG-thiol chains(MW 800,2kDa and 6kDa)are shown to form self-suspended systems over a range of polymer grafting densities and particle volume fractions, ?.Transmission electron and atomic force microscopy measurements reveal that the particles are uniformly dispersed. Oscillatory shear measurements performed on low ? systems show a two-step yielding behavior reflecting bond breaking and cage breaking transitions at the nanoscale; both characteristics of soft glassy materials dominated by attractive forces. With increased temperature a transition to one-step yielding and subsequently back to two-step yielding is observed. At high ? a single yielding transition and soft glassy flow behavior are observed. We employ SAXS, vibration spectroscopy, thermal analysis, and rheology to interrogate the configuration state of the tethered chains and particle-particle interactions in detail. King Abdullah Univeristy of Science and Technology (KAUST), National Science Foundation, Cornell Center for Materials Research (CCMR), Advance Photon Source at Argonne National Lab.

  8. Tailoring the physical properties of thiol-capped PbS quantum dots by thermal annealing

    International Nuclear Information System (INIS)

    We show that the thermal annealing of thiol-capped PbS colloidal quantum dots provides a means of narrowing the nanoparticle size distribution, increasing the size of the quantum dots and facilitating their coalescence preferentially along the (100) crystallographic axes. We exploit these phenomena to tune the photoluminescence emission of an ensemble of dots and to narrow the optical linewidth to values that compare with those reported at room temperature for single PbS quantum dots. We probe the influence of annealing on the electronic properties of the quantum dots by temperature dependent studies of the photoluminescence and magneto-photoluminescence.

  9. From visible to white-light emission by siloxane-capped ZnO quantum dots upon interaction with thiols

    Science.gov (United States)

    Schejn, Aleksandra; Balan, Lavinia; Piatkowski, Dawid; Mackowski, Sebastian; Lulek, Janina; Schneider, Raphal

    2012-06-01

    The interaction of thiols (glutathione, cysteine, and cysteamine) with yellow-emitting siloxane-capped ZnO QDs was studied. A gradual enlargement of the PL emission band resulting in white-light emission was observed upon reaction with thiols, while the diameter (ca. 4 nm) and the crystallinity of the dots were not affected. The appearance of broad white-emission was accompanied by a decrease of the photoluminescence quantum yield from 16% to 5-6%. Generation of surface defect states through interaction of the thiols with Zn surface atoms of the dots provoking shrunk of the siloxane capping may be responsible of that broadband emission throughout most of the light spectrum.

  10. Preparing mono-dispersed liquid core PDMS microcapsules from thioleneepoxy-tailored flow-focusing microfluidic devices

    DEFF Research Database (Denmark)

    Mazurek, Piotr Stanislaw; Daugaard, Anders Egede

    2015-01-01

    An applied dual-cure system based on thiolene and thiolepoxy click chemistry reactions was proved to be an extremely effective and easy to use tool for preparing microfluidic chips, thereby allowing for precise control over material properties and providing the possibility of covalently bonding chip wafers. Different thioleneepoxy-based polymer compositions were tested with the help of DSC and ATR FTIR, in order to investigate their physical and chemical properties. Water contact angles were determined, thus verifying the high efficiency and selectivity of the chemical surface modification of compositions in relation to high hydrophilicity and hydrophobicity. An obtained microfluidic device was subsequently used in order to produce PDMS microcapsules of very narrow size distribution and which contained various common liquids, such as water and ethanol, as well as an ionic liquid 2-hydroxyethylammonium formate.

  11. Preparing mono-dispersed liquid core PDMS microcapsules from thiol-ene-epoxy-tailored flow-focusing microfluidic devices

    DEFF Research Database (Denmark)

    Mazurek, Piotr Stanislaw; Daugaard, Anders Egede

    2015-01-01

    An applied dual-cure system based on thiol-ene and thiol-epoxy "click chemistry" reactions was proved to be an extremely effective and easy to use tool for preparing microfluidic chips, thereby allowing for precise control over material properties and providing the possibility of covalently bonding chip wafers. Different thiol-ene-epoxy-based polymer compositions were tested with the help of DSC and ATR FTIR, in order to investigate their physical and chemical properties. Water contact angles were determined, thus verifying the high efficiency and selectivity of the chemical surface modification of compositions in relation to high hydrophilicity and hydrophobicity. An obtained microfluidic device was subsequently used in order to produce PDMS microcapsules of very narrow size distribution and which contained various common liquids, such as water and ethanol, as well as an ionic liquid 2-hydroxyethylammonium formate.

  12. Crystallization of purple nitrous oxide reductase from Pseudomonas stutzeri

    International Nuclear Information System (INIS)

    The physiologically active form of nitrous oxide reductase was isolated and crystallized under strict exclusion of dioxygen and diffraction data were collected from crystals belonging to two different space groups. Nitrous oxide reductase (N2OR) from Pseudomonas stutzeri catalyzes the final step in denitrification: the two-electron reduction of nitrous oxide to molecular dinitrogen. Crystals of the enzyme were grown under strict exclusion of dioxygen by sitting-drop vapour diffusion using 2R,3R-butanediol as a cryoprotectant. N2OR crystallized in either space group P1 or P65. Interestingly, the key determinant for the resulting space group was the crystallization temperature. Crystals belonging to space group P1 contained four 130 kDa dimers in the asymmetric unit, while crystals belonging to space group P65 contained a single dimer in the asymmetric unit. Diffraction data were collected to resolutions better than 2

  13. A second target of benzamide riboside: Dihydrofolate reductase

    OpenAIRE

    Roussel, Breton; Johnson-farley, Nadine; Kerrigan, John E.; Scotto, Kathleen W.; Banerjee, Debabrata; Felczak, Krzysztof; Pankiewicz, Krzysztof W.; Gounder, Murugesan; Lin, Hongxia; Abali, Emine Ercikan; Bertino, Joseph R.

    2012-01-01

    Dihydrofolate reductase (DHFR) is an essential enzyme involved in de novo purine and thymidine biosynthesis. For several decades, selective inhibition of DHFR has proven to be a potent therapeutic approach in the treatment of various cancers including acute lymphoblastic leukemia, non-Hodgkins lymphoma, osteogenic sarcoma, carcinoma of the breast, and head and neck cancer. Therapeutic success with DHFR inhibitor methotrexate (MTX) has been compromised in the clinic, which limits the succes...

  14. Intermediate hyperhomocysteinemia resulting from compound heterozygosity of methylenetetrahydrofolate reductase mutations.

    OpenAIRE

    S.S. Kang; Wong, P W; Bock, H G; Horwitz, A; Grix, A

    1991-01-01

    Four subjects with thermolabile methylenetetrahydrofolate reductase (MTHFR) were discovered among 16 "obligate" heterozygotes for severe MTHFR deficiency and their family members. All four subjects had less than 25% of normal mean MTHFR specific activity in lymphocyte extracts. Three of them with normal serum folate and cyanocobalamin had intermediate hyperhomocysteinemia, and one with high serum folate and cyanocobalamin had no excessive accumulation of serum homocysteine. The biochemical fe...

  15. Structural Basis for Substrate Specificity in Human Monomeric Carbonyl Reductases

    Science.gov (United States)

    El-Hawari, Yasser; Dunford, James E.; Kochan, Grazyna; Wsol, Vladimir; Martin, Hans-Joerg; Maser, Edmund; Oppermann, Udo

    2009-01-01

    Carbonyl reduction constitutes a phase I reaction for many xenobiotics and is carried out in mammals mainly by members of two protein families, namely aldo-keto reductases and short-chain dehydrogenases/reductases. In addition to their capacity to reduce xenobiotics, several of the enzymes act on endogenous compounds such as steroids or eicosanoids. One of the major carbonyl reducing enzymes found in humans is carbonyl reductase 1 (CBR1) with a very broad substrate spectrum. A paralog, carbonyl reductase 3 (CBR3) has about 70% sequence identity and has not been sufficiently characterized to date. Screening of a focused xenobiotic compound library revealed that CBR3 has narrower substrate specificity and acts on several orthoquinones, as well as isatin or the anticancer drug oracin. To further investigate structure-activity relationships between these enzymes we crystallized CBR3, performed substrate docking, site-directed mutagenesis and compared its kinetic features to CBR1. Despite high sequence similarities, the active sites differ in shape and surface properties. The data reveal that the differences in substrate specificity are largely due to a short segment of a substrate binding loop comprising critical residues Trp229/Pro230, Ala235/Asp236 as well as part of the active site formed by Met141/Gln142 in CBR1 and CBR3, respectively. The data suggest a minor role in xenobiotic metabolism for CBR3. Enhanced version This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1. PMID:19841672

  16. Binding of Natural and Synthetic Polyphenols to Human Dihydrofolate Reductase

    OpenAIRE

    Jos Neptuno Rodrguez-Lpez; Juan Cabezas-Herrera; Soledad Chazarra; Lus Snchez-del-Campo; Magal Sez-Ayala

    2009-01-01

    Dihydrofolate reductase (DHFR) is the subject of intensive investigation since it appears to be the primary target enzyme for antifolate drugs. Fluorescence quenching experiments show that the ester bond-containing tea polyphenols (-)-epigallocatechin gallate (EGCG) and (-)-epicatechin gallate (ECG) are potent inhibitors of DHFR with dissociation constants (KD) of 0.9 and 1.8 ?M, respectively, while polyphenols lacking the ester bound gallate moiety [e.g., (-)-epigallocatechin (EGC) and (-)-...

  17. 5?-reductase-2 Deficiencys Effect on Human Fertility

    OpenAIRE

    Kang, Hey-joo; Imperato-mcginley, Julianne; Zhu, Yuan-shan; Rosenwaks, Zev

    2014-01-01

    A most interesting and intriguing male disorder of sexual differentiation is due to 5?-reductase-2 isoenzyme deficiency. These males are born with ambiguous external genitalia due to a deficiency in their ability to catalyze the conversion of testosterone to dihydrotestosterone (DHT). DHT is a potent androgen responsible for differentiation of the urogenital sinus and genital tubercle into the external genitalia, urethra and prostate. Affected males are born with a clitoral-like phallus, bif...

  18. Targeting 5?-reductase for prostate cancer prevention and treatment

    OpenAIRE

    Nacusi, Lucas P.; Tindall, Donald J.

    2011-01-01

    Testosterone is the most abundant circulating androgen, and can be converted to dihydrotestosterone (DHT), a more potent androgen, by the 5?-reductase enzymes in target tissues. Current treatments for prostate cancer consist of reducing androgen levels by chemical or surgical castration or pure antiandrogen therapy that directly targets the androgen receptor (AR). Although these therapies reduce tumor burden and AR activity, the cancer inevitably recurs within 1830 months. An approach tar...

  19. ALDOSE REDUCTASE: New Insights for an Old Enzyme

    OpenAIRE

    Ramana, Kota V.

    2011-01-01

    In the past years aldose reductase (AKR1B1; AR) is thought to be involved in the pathogenesis of secondary diabetic complications such as retinopathy, neuropathy, nephropathy and cataractogenesis. Subsequently, a number of AR inhibitors have been developed and tested for diabetic complications. Although, these inhibitors have found to be safe for human use, they have not been successful at the clinical studies because of limited efficacy. Recently, the potential physiological role of AR has b...

  20. Structural basis of stereospecific reduction by quinuclidinone reductase

    OpenAIRE

    Takeshita, Daijiro; Kataoka, Michihiko; Miyakawa, Takuya; Miyazono, Ken-ichi; Kumashiro, Shoko; Nagai, Takahiro; Urano, Nobuyuki; Uzura, Atsuko; Nagata, Koji; Shimizu, Sakayu; Tanokura, Masaru

    2014-01-01

    Chiral molecule (R)-3-quinuclidinol, a valuable compound for the production of various pharmaceuticals, is efficiently synthesized from 3-quinuclidinone by using NADPH-dependent 3-quinuclidinone reductase (RrQR) from Rhodotorula rubra. Here, we report the crystal structure of RrQR and the structure-based mutational analysis. The enzyme forms a tetramer, in which the core of each protomer exhibits the ?/? Rossmann fold and contains one molecule of NADPH, whereas the characteristic substructure...

  1. Inhibition of Aldose Reductase by Gentiana lutea Extracts

    OpenAIRE

    Chandrasekhar Akileshwari; Puppala Muthenna; Branislav Nastasijevi?; Gordana Joksi?; Mark Petrash, J.; Geereddy Bhanuprakash Reddy

    2012-01-01

    Accumulation of intracellular sorbitol due to increased aldose reductase (ALR2) activity has been implicated in the development of various secondary complications of diabetes. Thus, ALR2 inhibition could be an effective strategy in the prevention or delay of certain diabetic complications. Gentiana lutea grows naturally in the central and southern areas of Europe. Its roots are commonly consumed as a beverage in some European countries and are also known to have medicinal properties. The wate...

  2. Aldo-keto reductase (AKR) superfamily: Genomics and annotation

    OpenAIRE

    Mindnich Rebekka D; Penning Trevor M

    2009-01-01

    Abstract Aldo-keto reductases (AKRs) are phase I metabolising enzymes that catalyse the reduced nicotinamide adenine dinucleotide (phosphate) (NAD(P)H)-dependent reduction of carbonyl groups to yield primary and secondary alcohols on a wide range of substrates, including aliphatic and aromatic aldehydes and ketones, ketoprostaglan-dins, ketosteroids and xenobiotics. In so doing they functionalise the carbonyl group for conjugation (phase II enzyme reactions). Although functionally diverse, AK...

  3. A Murine Model for Human Sepiapterin-Reductase Deficiency

    OpenAIRE

    Yang, Seungkyoung; Lee, Young Jae; KIM, JIN-MAN; Park, Sean; Peris, Joanna; Laipis, Philip; Park, Young Shik; Chung, Jae hoon; Oh, S. Paul

    2006-01-01

    Tetrahydrobiopterin (BH4) is an essential cofactor for several enzymes, including all three forms of nitric oxide synthases, the three aromatic hydroxylases, and glyceryl-ether mono-oxygenase. A proper level of BH4 is, therefore, necessary for the metabolism of phenylalanine and the production of nitric oxide, catecholamines, and serotonin. BH4 deficiency has been shown to be closely associated with diverse neurological psychiatric disorders. Sepiapterin reductase (SPR) is an enzyme that cata...

  4. Glutathione Reductase Is Essential for Host Defense against Bacterial Infection

    OpenAIRE

    Yan, Jing; Ralston, Melissa M.; Meng, Xiaomei; Bongiovanni, Kathleen D.; Jones, Amanda L.; Benndorf, Rainer; Nelin, Leif D.; Frazier, W. Joshua; Rogers, Lynette K.; Smith, Charles V.; Liu, Yusen

    2013-01-01

    Glutathione reductase (Gsr)1 catalyzes the reduction of glutathione disulfide to glutathione, a major cellular antioxidant. We have recently shown that Gsr is essential for host defense against the Gram-negative bacteria Escherichia coli in a mouse model of sepsis. While we have demonstrated that Gsr is required for sustaining the oxidative burst and the development of neutrophil extracellular traps, the role of Gsr in other phagocytic functions remains unclear. It is also unclear whether Gsr...

  5. Pleiotropic effects of the HMG-CoA reductase inhibitors

    OpenAIRE

    Cg, Mihos; Santana O

    2011-01-01

    Christos G Mihos, Orlando SantanaColumbia University Division of Cardiology, Mount Sinai Heart Institute, Miami Beach, FL, USAAbstract: The HMG-CoA reductase inhibitors (statins) are used extensively in the treatment of hyperlipidemia. They have also demonstrated a benefit in a variety of other disease processes. These secondary actions are known as pleiotropic effects. Our paper serves as a focused and updated discussion on the pleiotropy of statins and emphasizes the importance of randomize...

  6. Aldose Reductase-Deficient Mice Develop Nephrogenic Diabetes Insipidus

    OpenAIRE

    Ho, Horace T. B.; Chung, Sookja K.; Law, Janice W. S.; Ko, Ben C. B.; Tam, Sidney C. F.; Brooks, Heddwen L.; Knepper, Mark A.; Chung, Stephen S. M.

    2000-01-01

    Aldose reductase (ALR2) is thought to be involved in the pathogenesis of various diseases associated with diabetes mellitus, such as cataract, retinopathy, neuropathy, and nephropathy. However, its physiological functions are not well understood. We developed mice deficient in this enzyme and found that they had no apparent developmental or reproductive abnormality except that they drank and urinated significantly more than their wild-type littermates. These ALR2-deficient mice exhibited a pa...

  7. Active site modification of aldose reductase by nitric oxide donors.

    Science.gov (United States)

    Chandra, A; Srivastava, S; Petrash, J M; Bhatnagar, A; Srivastava, S K

    1997-09-01

    Nitric oxide (NO) donors sodium nitrosoprusside (SNP), S-nitroso-N-acetylpenicillamine (SNAP), and 3-morpholinosydnonemine (SIN-1) caused a time- and concentration-dependent loss of catalytic activity of recombinant human placental aldose reductase. Modification of the enzyme was prevented by NADPH and NADP and reversed partially by dithiothreitol (DTT) and sodium borohydride. The protection by NADPH was lost in the presence of both substrates (NADPH and glyceraldehyde), indicating that the enzyme becomes sensitive to inhibition by SNP during catalysis. Site-directed mutant form of the enzyme, in which active site cys-298 was substituted with serine (C298S) was not inactivated by NO donors, whereas, ARC80S and ARC303 were as sensitive as the wild type enzyme, indicating that inactivation of aldose reductase is due to modification of the active site at cys298. These results suggest that NO may be an endogenous regulator of aldose reductase, and consequently the polyol pathway of glucose metabolism; which has been implicated in the pathogenesis of secondary diabetic complications. PMID:9357961

  8. Nitrate reductase gene involvement in hexachlorobiphenyl dechlorination by Phanerochaete chrysosporium

    International Nuclear Information System (INIS)

    Polychlorobiphenyl (PCB) degradation usually occurs through reductive dechlorination under anaerobic conditions and phenolic ring cleavage under aerobic conditions. In this paper, we provide evidence of nitrate reductase (NaR) mediated dechlorination of hexachlorobiphenyl (PCB-153) in Phanerochaete chrysosporium under non-ligninolytic condition and the gene involved. The NaR enzyme and its cofactor, molybdenum (Mo), were found to mediate reductive dechlorination of PCBs even in aerobic condition. Tungsten (W), a competitive inhibitor of this enzyme, was found to suppress this dechlorination. Chlorine release assay provided further evidence of this nitrate reductase mediated dechlorination. Commercially available pure NaR enzyme from Aspergillus was used to confirm these results. Through homology search using TBLASTN program, NaR gene was identified, primers were designed and the RT-PCR product was sequenced. The NaR gene was then annotated in the P. chrysosporium genome (GenBank accession no. AY700576). This is the first report regarding the presence of nitrate reductase gene in this fungus with the explanation why this fungus can dechlorinate PCBs even in aerobic condition. These fungal inoculums are used commercially as pellets in sawdust for enhanced bioremediation of PCBs at the risk of depleting soil nitrates. Hence, the addition of nitrates to the pellets will reduce this risk as well as enhance its activity

  9. Cloning and sequence of the human adrenodoxin reductase gene

    International Nuclear Information System (INIS)

    Adrenodoxin reductase is a flavoprotein mediating electron transport to all mitochondrial forms of cytochrome P450. The authors cloned the human adrenodoxin reductase gene and characterized it by restriction endonuclease mapping and DNA sequencing. The entire gene is approximately 12 kilobases long and consists of 12 exons. The first exon encodes the first 26 of the 32 amino acids of the signal peptide, and the second exon encodes the remainder of signal peptide and the apparent FAD binding site. The remaining 10 exons are clustered in a region of only 4.3 kilobases, separated from the first two exons by a large intron of about 5.6 kilobases. Two forms of human adrenodoxin reductase mRNA, differing by the presence or absence of 18 bases in the middle of the sequence, arise from alternate splicing at the 5' end of exon 7. This alternately spliced region is directly adjacent to the NADPH binding site, which is entirely contained in exon 6. The immediate 5' flanking region lacks TATA and CAAT boxes; however, this region is rich in G+C and contains six copies of the sequence GGGCGGG, resembling promoter sequences of housekeeping genes. RNase protection experiments show that transcription is initiated from multiple sites in the 5' flanking region, located about 21-91 base pairs upstream from the AUG translational initiation codon

  10. Thioredoxin-thioredoxin reductase system of Streptomyces clavuligerus: sequences, expression, and organization of the genes.

    OpenAIRE

    Cohen, G.; Yanko, M.; Mislovati, M.; Argaman, A.; Schreiber, R.; Av-gay, Y.; Aharonowitz, Y.

    1993-01-01

    The genes that encode thioredoxin and thioredoxin reductase of Streptomyces clavuligerus were cloned, and their DNA sequences were determined. Previously, we showed that S. clavuligerus possesses a disulfide reductase with broad substrate specificity that biochemically resembles the thioredoxin oxidoreductase system and may play a role in the biosynthesis of beta-lactam antibiotics. It consists consists of two components, a 70-kDa NADPH-dependent flavoprotein disulfide reductase with two iden...

  11. Immunological comparison of the NADH:nitrate reductase from different cucumber tissues

    Directory of Open Access Journals (Sweden)

    Jolanta Marciniak

    2014-02-01

    Full Text Available Soluble nitrate reductase from cucumber roots (Cucumis sativus L. was isolated and purified with blue-Sepharose 4B. Specific antibodies against the NR protein were raised by immunization of a goat. Using polyclonal antibodies anti-NR properties of the nitrate reductase from various cucumber tissues were examined. Experiments showed difference in immuno-logical properties of nitrate reductase (NR from cotyledon roots and leaves.

  12. Immunological comparison of the NADH:nitrate reductase from different cucumber tissues

    OpenAIRE

    Jolanta Marciniak; Gra?yna K?obus; Jzef Buczek; Tadeusz Stefaniak; Jaros?aw Mazur

    1995-01-01

    Soluble nitrate reductase from cucumber roots (Cucumis sativus L.) was isolated and purified with blue-Sepharose 4B. Specific antibodies against the NR protein were raised by immunization of a goat. Using polyclonal antibodies anti-NR properties of the nitrate reductase from various cucumber tissues were examined. Experiments showed difference in immuno-logical properties of nitrate reductase (NR) from cotyledon roots and leaves.

  13. Inhibitory Activity of Some Plant Methanol Extracts on 3-Hydroxy-3-Methylglutaryl Coenzyme a Reductase

    OpenAIRE

    Shahouzehi, B.; Gholamhoseinian, A.; Sharifi-far, F.

    2010-01-01

    Beta-hydroxy-beta-methylglutaryl coenzyme a reductase (HMG CoA reductase) is the key enzyme in cholesterol biosynthesis. Inhibition of this enzyme reduces the synthesis of cholesterol and could be used in the management of coronary artery disease. Drugs used for the management of cholesterol biosynthesis have showed some side effects that are cause of new trends in the nature. This study was designed to find new HMG CoA reductase inhibitors from natural resources. One hundred plants were bota...

  14. Control of Cholesterol Synthesis through Regulated ER-Associated Degradation of HMG CoA Reductase

    OpenAIRE

    Jo, Youngah; Debose-boyd, Russell A.

    2010-01-01

    Multiple mechanisms for feedback control of cholesterol synthesis converge on the rate-limiting enzyme in the pathway, 3-hydroxy-3-methylglutaryl coenzyme A reductase. This complex feedback regulatory system is mediated by sterol and nonsterol metabolites of mevalonate, the immediate product of reductase activity. One mechanism for feedback control of reductase involves rapid degradation of the enzyme from membranes of the endoplasmic reticulum (ER). This degradation results from the accumula...

  15. Structure and reactivity of Trypanosoma brucei pteridine reductase: inhibition by the archetypal antifolate methotrexate

    OpenAIRE

    Dawson, Alice; Gibellini, Federica; Sienkiewicz, Natasha; Tulloch, Lindsay B; Fyfe, Paul K; McLuskey, Karen; Fairlamb, Alan H; Hunter, William N.

    2006-01-01

    The protozoan Trypanosoma brucei has a functional pteridine reductase (TbPTR1), an NADPH-dependent short-chain reductase that participates in the salvage of pterins, which are essential for parasite growth. PTR1 displays broad-spectrum activity with pterins and folates, provides a metabolic bypass for inhibition of the trypanosomatid dihydrofolate reductase and therefore compromises the use of antifolates for treatment of trypanosomiasis. Catalytic properties of recombinant TbPTR1 and inhibit...

  16. Thianthrene is a novel inhibitor of Leishmania donovani pteridine reductase 1 (PTR1)

    OpenAIRE

    Neeloo Singh; Prashant Singh; Ravishankar Ramachandran; Divya Dube; Jaspreet Kaur *1

    2012-01-01

    Pteridine reductase 1 (PTR1) from Leishmania donovani is a short chain reductase that catalyses the NADPH-dependent reduction of folates and pterins. It has gained attention as a therapeutic target because it acts as a metabolic bypass for dihydrofolate reductase (DHFR) targeting drugs and is thought to be responsible for the failure of conventional therapies against the trypanosomatids. In the present study, we report the identification of thianthrene as a potent inhibitor of L. donovani PTR...

  17. Gold nanoparticles decorated with oligo(ethylene glycol) thiols: surface charges and interactions with proteins in solution.

    OpenAIRE

    Schollbach, M; Zhang, F.(Department of Physics, University of Wisconsin, Madison, WI, United States of America); Roosen-Runge, F; Skoda, MW; Jacobs, RM; Schreiber, F

    2014-01-01

    We have studied oligo(ethylene glycol) (OEG) thiol self-assembled monolayer (SAM) coated gold nanoparticles (AuOEG) and their interactions with proteins in solutions using electrophoretic and dynamic light scattering (ELS and DLS). The results are compared with poly(ethylene glycol) (PEG) thiol coated AuNPs (AuPEG). We show that both AuOEG and AuPEG particles carry a low net negative charge and are very stable (remaining so for more than one year), but long-term aging or dialysis can reduce t...

  18. Synthesis of glycerin carbonate-based intermediates using thiol-ene chemistry and isocyanate free polyhydroxyurethanes therefrom

    OpenAIRE

    Benyahya, Sofia; Desroches, Myriam; Auvergne, Re?mi; Carlotti, Ste?phane; Caillol, Sylvain; Boutevin, Bernard

    2011-01-01

    A new synthesis of 4-[(prop-2-en-1-yloxy)methyl]-1,3-dioxolan-2-one (AGC) was performed by Williamson ether synthesis from 4-(hydroxymethyl)-1,3-dioxolan-2-one. Dicyclocarbonates were synthesized by UV thiol-ene coupling of allyl-cyclocarbonate with a 2,20-oxydiethanethiol. This photochemical thiol-ene reaction was carried out under air, with neither solvent nor photoinitiator. The products, obtained with high yield, were characterized by 1H NMR and FTIR analysis. The synthesized dicyclocarbo...

  19. Reaction of [3H]-taurine maleimide with platelet surface thiols

    International Nuclear Information System (INIS)

    Taurine Maleimide (2-maleimidoethanesulfonate, TM) was synthesized from [2-3H]-taurine and methoxycarbonylmaleimide (MCM). The yield of a 1 ?mol synthesis approached 100% (based on taurine) when MCM was used in 4-fold excess. The product (TM*) was purified by ion exchange chromatography. TM* reacted irreversibly with thiol groups on the surface of washed human platelets, leading to incorporation of radioactivity into platelet pellets. Incorporation was blocked by cysteine, mercuribenzenesulfonate (MBS), dithiobisnitrobenzoate, and N-ethylmaleimide, but not by taurine or by inhibitors of anion transport. Reaction of TM* with platelets showed the dependence on time and concentration characteristics of a bimolecular reaction. The number of reactive sites ranged from 1 to 5 x 105/platelet, and the apparent rate constant from 1 to 3 x 103/(M x min). TM was less effective than MBS as an inhibitor of platelet aggregation induced by several agents. TM had no effect on the uptake of serotonin, taurine, or phosphate by the platelets, processes which are sensitive to MBS. These differences, considered with the similarity in size and charge of TM and MBS, suggest that classes of thiols defined as exofacial by their accessibility to MBS can differ substantially in their reactivity with other impermeant reagents

  20. The Leishmania ATP-binding cassette protein PGPA is an intracellular metal-thiol transporter ATPase.

    Science.gov (United States)

    Lgar, D; Richard, D; Mukhopadhyay, R; Stierhof, Y D; Rosen, B P; Haimeur, A; Papadopoulou, B; Ouellette, M

    2001-07-13

    The Leishmania ATP-binding cassette (ABC) transporter PGPA is involved in metal resistance (arsenicals and antimony), although the exact mechanism by which PGPA confers resistance to antimony, the first line drug against Leishmania, is unknown. The results of co-transfection experiments, transport assays, and the use of inhibitors suggest that PGPA recognizes metals conjugated to glutathione or trypanothione, a glutathione-spermidine conjugate present in Leishmania. The HA epitope tag of the influenza hemagglutinin as well as the green fluorescent protein were fused at the COOH terminus of PGPA. Immunofluorescence, confocal, and electron microscopy studies of the fully functional tagged molecules clearly indicated that PGPA is localized in membranes that are close to the flagellar pocket, the site of endocytosis and exocytosis in this parasite. Subcellular fractionation of Leishmania tarentolae PGPAHA transfectants was performed to further characterize this ABC transporter. The basal PGPA ATPase activity was determined to be 115 nmol/mg/min. Transport experiments using radioactive arsenite-glutathione conjugates clearly showed that PGPA recognizes and actively transports thiol-metal conjugates. Overall, the results are consistent with PGPA being an intracellular ABC transporter that confers arsenite and antimonite resistance by sequestration of the metal-thiol conjugates. PMID:11306588

  1. Intracellular thiol levels and radioresistance: Studies with glutathione and glutathione mono ethyl ester

    International Nuclear Information System (INIS)

    Intracellular thiols such as glutathione (GSH) protect cells against free radicals formed during oxidative metabolism or from exposure to drugs or ionizing radiation. The role of intracellular GSH in the repair of radiation induced free radical damage was studied using GSH or its analog glutathione mono ethyl ester (GEE), which readily penetrates into the cell. Chinese hamster V79 cells with normal GSH levels were afforded equal protection under aerated and hypoxic conditions (DMF = 1.2 OER = 3.7) by both 10 mM GSH and GEE although GEE had raised interacellular GSH levels three-fold. Growth of V79 cells in cysteine free media resulted in undetectable levels of GSH and OER of 2.2 with no change in aerated survival. Restoration of intracellular GSH by 10 mM GEE resulted in an increase of the OER from 2.2. to 3.8 (DMF = 1.7). Only 14% of the intracellular GSH needs to be repleted to give an OER of 3.0. These experiments provide evidence that thiols do play a role in the oxygen effect and are present at levels in excess of what is necessary for maximal radioprotection

  2. Effect of silver nanomaterials on the activity of thiol-containing antioxidants.

    Science.gov (United States)

    Zhou, Yu-Ting; He, Weiwei; Lo, Y Martin; Hu, Xiaona; Wu, Xiaochun; Yin, Jun-Jie

    2013-08-14

    The use of nanomaterials in consumer products is rapidly expanding. In most studies, nanomaterials are examined as isolated ingredients. However, consumer products such as foods, cosmetics, and dietary supplements are complex chemical matrixes. Therefore, interactions between nanomaterials and other components of the product must be investigated to ensure the product's performance and safety. Silver nanomaterials are increasingly being used in food packaging as antimicrobial agents. Thiol-containing compounds, such as reduced glutathione (GSH), cysteine, and dihydrolipoic acid, are used as antioxidants in many consumer products. In the current study, we have investigated the interaction between silver nanomaterials and thiol-containing antioxidants. The selected Ag nanomaterials were Ag coated with citrate, Ag coated with poly(vinylpyrrolidone), and Au nanorods coated with Ag in a core/shell structure. We observed direct quenching of the 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) by all three Ag nanomaterials to varying degrees. The Ag nanomaterials also reduced the quenching of DPPH by GSH to varying degrees. In addition, we determined that the mixture of GSH and Au@Ag nanorods held at 37 C was less effective at quenching azo radical than at ambient temperature. Furthermore, we determined that Au@Ag nanorods significantly reduced the ability of GSH and cysteine to quench hydroxyl and superoxide radicals. The work presented here demonstrates the importance of examining the chemical interactions between nanomaterials used in products and physiologically important antioxidants. PMID:23889173

  3. Adsorption characteristics of self-assembled thiol and dithiol layer on gold

    International Nuclear Information System (INIS)

    Monolayers of functional proteins are important in many fields related to pure and applied biochemistry and biophysics. The formation of extended uniform protein monolayers by single- or multiple-step self-chemisorption depends on the quality of the functionalized gold surface. The optical and the electrical properties of the 1-nonanethiol and 1,9-nonanedithiol deposited on gold with the self-assembled technique were investigated. We use cyclic voltammetry and impedance spectroscopy to characterize the insulating properties of the two layers. The analysis of the impedance spectra in terms of equivalent circuit of the gold/electrolyte and gold/SAM/electrolyte interface allows defining the thickness of the two thiols and the percentage of coverage area. Atomic force microscopy, contact angle measurement and Fourier transform infra-red spectroscopy have been used for homogeneity, hydrophobic properties and molecular structure of the formed thiols layer, respectively. The measured thickness with impedance spectroscopy fit well the results found with atomic force microscopy

  4. Modulating the physicochemical and structural properties of gold-functionalized protein nanotubes through thiol surface modification.

    Science.gov (United States)

    Carreo-Fuentes, Liliana; Plascencia-Villa, Germn; Palomares, Laura A; Moya, Sergio E; Ramrez, Octavio T

    2014-12-16

    Biomolecules are advantageous scaffolds for the synthesis and ordering of metallic nanoparticles. Rotavirus VP6 nanotubes possess intrinsic affinity to metal ions, a property that has been exploited to synthesize gold nanoparticles over them. The resulting nanobiomaterials have unique properties useful for novel applications. However, the formed nanobiomaterials lack of colloidal stability and flocculate, limiting their functionality. Here we demonstrate that it is possible to synthesize thiol-protected gold nanoparticles over VP6 nanotubes, which resulted in soluble nanobiomaterials. With this strategy, it was possible to modulate the size, colloidal stability, and surface plasmon resonance of the synthesized nanoparticles by controlling the content of the thiolated ligands. Two types of water-soluble ligands were tested, a small linear ligand, sodium 3-mercapto-1-propanesulfonate (MPS), and a bulky ligand, 5-mercaptopentyl ?-D-glucopyranoside (GlcC5SH). The synthesized nanobiomaterials had a higher stability in suspension, as determined by Z-potential measurements. To the extent of our knowledge, this is the first time that a rational strategy is developed to modulate the particular properties of metal nanoparticles in situ synthesized over a protein bioscaffold through thiol coating, achieving a high spatial and structural organization of nanoparticles in a single integrative hybrid structure. PMID:25409000

  5. Thermal stabilization and plasticization of poly(vinyl chloride) by ester thiols: Update and current status

    Energy Technology Data Exchange (ETDEWEB)

    Starnes, William H. [Departments of Chemistry and Applied Science, College of William and Mary, P.O. Box 8795, Williamsburg, VA 23187-8795 (United States)]. E-mail: whstar@wm.edu; Du, Bin [Departments of Chemistry and Applied Science, College of William and Mary, P.O. Box 8795, Williamsburg, VA 23187-8795 (United States); Kim, Soungkyoo [Departments of Chemistry and Applied Science, College of William and Mary, P.O. Box 8795, Williamsburg, VA 23187-8795 (United States); Zaikov, Vadim G. [Departments of Chemistry and Applied Science, College of William and Mary, P.O. Box 8795, Williamsburg, VA 23187-8795 (United States); Ge, Xianlong [Departments of Chemistry and Applied Science, College of William and Mary, P.O. Box 8795, Williamsburg, VA 23187-8795 (United States); Culyba, Elizabeth K. [Departments of Chemistry and Applied Science, College of William and Mary, P.O. Box 8795, Williamsburg, VA 23187-8795 (United States)

    2006-03-15

    Poly(vinyl chloride) (PVC) is one of the most important medical plastics. Recently, however, the safety of flexible PVC containing the common plasticizer, di(2-ethylhexyl) phthalate, has been called into question. Widely used heat stabilizers for PVC that incorporate toxic heavy metals also have fallen into disfavor. In order to address these problems, we have synthesized and tested, as potential replacements, several organic thiols that contain one or more carboxylate ester functions and thus are highly compatible with the polymer. When introduced into PVC at high loading levels (e.g., 30-35 parts by weight), the ester thiols are extremely effective as heat stabilizers and also useful as primary plasticizers. When used at a low loading level (e.g., 3 parts by weight), they still are excellent heat stabilizers for both plasticized and rigid PVC. Importantly, their high potency is achieved in the absence of any costabilizers that incorporate heavy metals. Their syntheses are simple and straightforward, and their odors are not offensive, because their volatilities are low. Described here are some typical results obtained with this new additive technology, which was licensed for commercialization in 2005.

  6. Light-Driven Reversible Alignment Switching of Liquid Crystals Enabled by Azo Thiol Grafted Gold Nanoparticles.

    Science.gov (United States)

    Xue, Chenming; Xiang, Jie; Nemati, Hossein; Bisoyi, Hari Krishna; Gutierrez-Cuevas, Karla; Wang, Ling; Gao, Min; Zhou, Shuang; Yang, Deng-Ke; Lavrentovich, Oleg D; Urbas, Augustine; Li, Quan

    2015-06-22

    Stimuli-directed alignment control of liquid crystals (LCs) with desired molecular orientation is currently in the limelight for the development of smart functional materials and devices. Here, photoresponsive azo thiol (AzoSH) was grafted onto gold nanoparticles (GNPs). The resulting hybrid GNPs were able to homogeneously mix with a commercially available nematic LC host, as evidenced by Cryo-TEM. Interestingly, the LC nanocomposites were found to undergo reversible alignment transition upon light irradiation as a consequence of the trans-cis photoisomerization of the azo groups on the GNP surface. LC molecules in either planar or bare glass cells were able to change their alignment to vertical upon UV irradiation, while the vertically aligned LC molecules returned to the planar or random orientation under visible irradiation. Neither the azo thiol molecules nor the unfunctionalized GNPs alone promoted the alignment of the LC molecules in the system upon light irradiation. The photoinduced vertical alignment without applied electric or magnetic field was very stable over time and with respect to temperature. Furthermore, an optically switchable device based on the photostimulated reversible alignment control of LCs was demonstrated. PMID:26097118

  7. Ambient STM Study of Long Chain Hydrophobic Thiol SAMs on HOPG and Au-capped HOPG

    Science.gov (United States)

    Maurer, Mackenzie; Bowers, Alexis; Senevirathne, Indrajith

    2015-03-01

    Thiol-based self-assembled monolayers (SAM) surfaces are ubiquitous in many device applications including sensor engineering. The conductivity characteristics and surface molecular structure and orientation of these SAMs are important as physiochemical properties are dependent on the surface arrangement. This study attempts to quantify and model long chain -R terminated (hydrophobic) 1-dodecanethiol on both Highly Oriented Pyrolytic Graphite (HOPG) and thermally annealed Au thin films capped on HOPG substrates. Study uses Scanning Tunneling Microscopy (STM), contact angle measurements and reflective spectroscopy to assess the SAM layered surface. 5mM concentrated solutions of 1-dodecanethiol dissolved in 200 proof ethanol were prepared for the self-assembly process. These solutions were used in developing SAMs on freshly cleaved HOPG. Au thin layers were sputter deposited on HOPG and subsequently annealed. Data indicated Au deposition changes the surface consistency. Uniqueness of this study is the ambient conditions under which data was obtained. Surface structure, consistency and possible thiol molecular arrangement of the SAM layer will be discussed.

  8. Addition of thiols to phenylselenoalkynes using KF/Alumina under solvent-free conditions

    Scientific Electronic Library Online (English)

    Renata G, Lara; Elton L, Borges; Eder J, Lenardo; Diego, Alves; Raquel G, Jacob; Gelson, Perin.

    2125-21-01

    Full Text Available Um mtodo simples e eficiente foi desenvolvido para a hidrotiolao de fenilselenoalquinos utilizando KF/Al2O3 em meio livre de solvente. O mtodo geral e permite a preparao seletiva de (Z)-1-fenilseleno-2-organotio-1-alquenos a partir de tiis aromticos e alifticos em rendimentos de razoveis [...] a bons. A presena do grupamento organosselnio direcionou a regioqumica da adio do tiol. O sistema cataltico pode ser reutilizado mais quatro vezes sem tratamento prvio Abstract in english We present herein the results of a simple and efficient protocol for the hydrothiolation of phenylselenoalkynes promoted by KF/Al2O3 using solvent-free conditions. This improved method furnishes selectively the corresponding (Z)-1-phenylseleno-2-organylthio-1-alkenes in reasonable to good yields sta [...] rting from selenoalkynes and aliphatic or aromatic thiols. The presence of the phenylselenium group in the alkyne directed the regiochemistry of the thiol addition. The catalytic system can be reused up to 4 times without previous treatment.

  9. Photocured thiol-ene based optical fluorescence sensor for determination of gold(III)

    International Nuclear Information System (INIS)

    Graphical abstract: -- Highlights: Photopolymerized fluorescence sensor for Au(III) analysis has been developed. Preparation of polymeric sensor is simple and quick. Fluorescence sensor used for analysis of Au(III) in real samples. -- Abstract: This study describes the preparation and the characterization of a new thiol-ene based polymeric fluorescence sensor by photo initiated polymerization of trimethylolpropane tris(3-mercaptopropionate), 2-hydroxyethylacrylate, and 2,4,6-triallyloxy-1,3,5-triazine which are used as monomers and also a photo initiator (2,2-dimethoxy-2-phenylacetophenone) for its usage as optical sensor for gold ions. The thiol-ene based polymeric membrane sensor was characterized by using attenuated total reflectance-fourier transform infrared spectroscopy (ATR-FTIR) and scanning electron microscopy (SEM). The response characteristics of the sensors including dynamic range, pH effect, response time, and the effect of foreign ions were investigated. Fluorescence spectra showed that the excitation/emission maxima of the membrane were at 379/425 nm, respectively

  10. Detection of thiol-based redox switch processes in parasites - facts and future.

    Science.gov (United States)

    Rahbari, Mahsa; Diederich, Kathrin; Becker, Katja; Krauth-Siegel, R Luise; Jortzik, Esther

    2015-05-01

    Malaria and African trypanosomiasis are tropical diseases caused by the protozoa Plasmodium and Trypanosoma, respectively. The parasites undergo complex life cycles in the mammalian host and insect vector, during which they are exposed to oxidative and nitrosative challenges induced by the host immune system and endogenous processes. Attacking the parasite's redox metabolism is a target mechanism of several known antiparasitic drugs and a promising approach to novel drug development. Apart from this aspect, oxidation of cysteine residues plays a key role in protein-protein interaction, metabolic responses to redox events, and signaling. Understanding the role and dynamics of reactive oxygen species and thiol switches in regulating cellular redox homeostasis is crucial for both basic and applied biomedical approaches. Numerous techniques have therefore been established to detect redox changes in parasites including biochemical methods, fluorescent dyes, and genetically encoded probes. In this review, we aim to give an insight into the characteristics of redox networks in the pathogens Plasmodium and Trypanosoma, including a comprehensive overview of the consequences of specific deletions of redox-associated genes. Furthermore, we summarize mechanisms and detection methods of thiol switches in both parasites and discuss their specificity and sensitivity. PMID:25741735

  11. Photocured thiol-ene based optical fluorescence sensor for determination of gold(III)

    Energy Technology Data Exchange (ETDEWEB)

    ubuk, Soner, E-mail: sonercubuk@marmara.edu.tr; Kahraman, Memet Vezir; Yetimo?lu, Ece Kk; Kenan, Sibel

    2014-02-17

    Graphical abstract: -- Highlights: Photopolymerized fluorescence sensor for Au(III) analysis has been developed. Preparation of polymeric sensor is simple and quick. Fluorescence sensor used for analysis of Au(III) in real samples. -- Abstract: This study describes the preparation and the characterization of a new thiol-ene based polymeric fluorescence sensor by photo initiated polymerization of trimethylolpropane tris(3-mercaptopropionate), 2-hydroxyethylacrylate, and 2,4,6-triallyloxy-1,3,5-triazine which are used as monomers and also a photo initiator (2,2-dimethoxy-2-phenylacetophenone) for its usage as optical sensor for gold ions. The thiol-ene based polymeric membrane sensor was characterized by using attenuated total reflectance-fourier transform infrared spectroscopy (ATR-FTIR) and scanning electron microscopy (SEM). The response characteristics of the sensors including dynamic range, pH effect, response time, and the effect of foreign ions were investigated. Fluorescence spectra showed that the excitation/emission maxima of the membrane were at 379/425 nm, respectively.

  12. Monitoring of Saccharomyces cerevisiae cell proliferation on thiol-modified planar gold microelectrodes using impedance spectroscopy

    DEFF Research Database (Denmark)

    Heiskanen, Arto; Spegel, Christer F

    2008-01-01

    An impedance spectroscopic study of the interaction between thiol-modified Au electrodes and Saccharomyces cerevisiae of strain EBY44 revealed that the cells formed an integral part of the interface, modulating the capacitive properties until a complete monolayer was obtained, whereas the charge transfer resistance (R-ct) to the redox process of [Fe(CN)6](3-14-) showed a linear relationship to the number of cells even beyond the monolayer coverage. R,, showed strong pH dependence upon increasing the pH of the utilized buffer to 7.2. Upon addition of S. cerevisiae cells at pH 7.2, the obtained value of R,, showed over 560% increase with respect to the value obtained on the same thiol-modified electrode without cells. It was demonstrated that real-time monitoring of S. cerevisiae proliferation, with frequency-normalized imaginary admittance (real capacitance) as the indicator, was possible using a miniaturized culture system, ECIS Cultureware, with integrated planar cysteamine-modified Au microelectrodes. A monolayer coverage was reached after 20-28 h of cultivation, observed as an similar to 15% decrease in the real capacitance of the system.

  13. pH-sensitive photoluminescence for aqueous thiol-capped CdTe nanocrystals

    International Nuclear Information System (INIS)

    pH-sensitive photoluminescence (PL) is an important property of aqueous nanocrystals (NCs) towards NCs-based intelligent applications. Previous works mainly focused on the effect of pH during NC growth process on PL of the aqueous NCs. The effect of pH during application process on PL of as-prepared NCs is still not fully understood. In this work, we brought out a general mechanism for the pH-sensitive PL behaviors of as-prepared aqueous CdTe NCs capped by aqueous thiol ligands, such as carboxylic-acid-terminated 3-mercaptopropionic acid (MPA) and thioglycolic acid (TGA) ligands, hydroxyl-terminated 1-thioglycerol (TG) ligands and amine-terminated 2-mercaptoethylamine (MA) ligands. A major contribution of this work is finding the key role of ligand terminal groups in the diffuse process of free Cd-ligand complexes toward NCs. This terminal group effect is the main reason for PL alteration of NCs during pH adjustment process. Besides the terminal group effect, PL of aqueous NCs is also affected by the aggregation effect, the thiol group effect and the counter ion effect. These effects make different contributions to PL of NCs at different pH ranges. By using this mechanism, we successfully explained the complex pH-sensitive PL behaviors of MPA, TGA, TG and MA-capped CdTe NCs.

  14. Identification and selective inhibition of an isozyme of steroid 5 alpha-reductase in human scalp.

    OpenAIRE

    Harris, G.; Azzolina, B; Baginsky, W.; Cimis, G; Rasmusson, G H; Tolman, R L; Raetz, C R; Ellsworth, K

    1992-01-01

    Steroid 5 alpha-reductase (EC 1.3.1.22) catalyzes the reduction of testosterone to dihydrotestosterone. The 5 alpha-reductase found in human scalp has been compared with the enzyme found in prostate. The scalp reductase has a broad pH optimum centered at pH 7.0. This is distinctly different from the pH optimum of 5.5 observed with the prostatic form of the enzyme. These enzymes also differ in the Km for testosterone, which is 25-fold higher for the scalp reductase. The most significant differ...

  15. Developmental and metabolic regulation of the Drosophila melanogaster 3-hydroxy-3-methylglutaryl coenzyme A reductase.

    OpenAIRE

    Gertler, F. B.; Chiu, C. Y.; Richter-mann, L.; Chin, D. J.

    1988-01-01

    The enzyme 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase in Drosophila melanogaster synthesizes mevalonate for the production of nonsterol isoprenoids, which are essential for growth and differentiation. To understand the regulation and developmental role of HMG CoA reductase, we cloned the D. melanogaster HMG CoA reductase gene. The nucleotide sequence of the Drosophila HMG CoA reductase was determined from genomic and cDNA clones. A 2,748-base-pair open reading frame encoded a p...

  16. Analysis of nitrate reductase mRNA expression and nitrate reductase activity in response to nitrogen supply

    Directory of Open Access Journals (Sweden)

    Gholamreza Kavoosi

    2014-06-01

    Full Text Available Nitrate is one of the major sources of nitrogen for the growth of plants. It is taken up by plant roots and transported to the leaves where it is reduced to nitrite in the. The main objective of this research was to investigate stimulatory effects of sodium nitrate, potassium nitrate, ammonia and urea on the production/generation of the nitrate reductase mRNA in Triticum aestivum plants. The plants were grown in standard nutrient solution for 21 days and then starved in a media without nitrate for seven days. Starved plants were stimulated with various concentrations of sodium nitrate, potassium nitrate, ammonia and urea, and the expression of nitrate reductase mRNA was analyzed by real-time PCR. Our results indicated that starvation caused significant decrease in the production of nitrate reductase mRNA in the plant leaf. Sodium and potassium nitrate were capable of restoring the production of nitrate mRNA in a dose-dependent manner, since 50 mM of each produced the highest level of the mRNA. The stimulatory effect of potassium nitrate was higher than sodium nitrate, while ammonia and urea did not show such activity. At low concentrations, sodium nitrate and potassium nitrate caused significant increase in the nitrate/nitrite mRNA production, whereas high concentrations of these salts suppressed the expression of this gene considerably.

  17. Removal of lead(II ions from aqueous solutions using cashew nut shell liquid-templated thiol-silica materials

    Directory of Open Access Journals (Sweden)

    J. E. G. Mdoe

    2014-09-01

    Full Text Available A range of thiol-silica composites were prepared using cashew nut shell liquid (CNSL or one of its phenolic constituents, cardanol, as templates. The procedure involved formation of a CNSL or cardanol emulsion in a water-ethanol system into which (3-mercaptopropyl-trimethoxysilane and tetraethyl orthosilicate were simultaneously added at various ratios. The reaction mixture was aged at room temperature for 18 h followed by a Soxhlet extraction of the template and drying. The materials were characterized by diffuse reflectance Fourier transform infrared, nitrogen physisorption, scanning electron microscopy and acid titration. Results indicated that indeed the thiol-silica composites were successfully prepared, with thiol group loadings ranging from 1.6-2.5 mmol/g. The materials were tested for lead(II adsorption, and results showed that they had maximum adsorption capacities up to 66.7 mg/g, depending on the thiol group loading and type of template used in preparing the adsorbent. DOI: http://dx.doi.org/10.4314/bcse.v28i3.5

  18. Improved electronic coupling in hybrid organic-inorganic nanocomposites employing thiol-functionalized P3HT and bismuth sulfide nanocrystals.

    Science.gov (United States)

    Martinez, L; Higuchi, S; MacLachlan, A J; Stavrinadis, A; Miller, N C; Diedenhofen, S L; Bernechea, M; Sweetnam, S; Nelson, J; Haque, S A; Tajima, K; Konstantatos, G

    2014-09-01

    In this study, we employ a thiol-functionalized polymer (P3HT-SH) as a leverage to tailor the nanomorphology and electronic coupling in polymer-nanocrystal composites for hybrid solar cells. The presence of the thiol functional group allows for a highly crystalline semiconducting polymer film at low thiol content and allows for improved nanomorphologies in hybrid organic-inorganic systems when employing non-toxic bismuth sulfide nanocrystals. The exciton dissociation efficiency and carrier dynamics at this hybrid heterojunction are investigated through photoluminescence quenching and transient absorption spectroscopy measurements, revealing a larger degree of polaron formation when P3HT-SH is employed, suggesting an increased electronic interaction between the metal chalcogenide nanocrystals and the thiol-functionalized P3HT. The fabricated photovoltaic devices show 15% higher power conversion efficiencies as a result of the improved nanomorphology and better charge transfer mechanism together with the higher open circuit voltages arising from the deeper energy levels of P3HT-SH. PMID:25029606

  19. An electron spin resonance investigation of the structure and formation of sulfinyl radicals: Reaction of peroxyl radicals with thiols

    International Nuclear Information System (INIS)

    In this work we present an electron spin resonance investigation of the irradiation of the alkyl mercaptans methyl, n-butyl, and tert-butyl mercaptan and the radioprotective thiols cysteamine and dithiothreitol in a number of aqueous and organic matrices in the present of oxygen. Matrix peroxyl radicals (ROO*) are formed after the irradiation of organic matrices in the presence of oxygen at 77 K. Upon annealing, these react with added thiols to form sulfinyl radicals (RSO*). Evidence for a thiol peroxyl radical (RSOO*) intermediate is found. Hyperfine couplings and g values are reported for the sulfinyl radicals formed from the five thiols investigated. The incorporation of 17O-labeled oxygen into the RSO* radical confirms that molecular oxygen is the source of the oxygen atom in the radical. The isotropic and parallel anisotropic 17O couplings indicate slightly less than 0.5 spin density on the oxygen. The couplings and spin densities are compared to those predicted from ab inito molecular orbital calculations for CH3SO*. Calculations for the sulfur-peroxyl intermediate, CH3SOO*, predict that it will have similar ESR parameters as those predicted for the carbon-centered peroxyl radical, CH3OO*

  20. A pyrene maleimide with a flexible linker for sampling of longer inter-thiol distances by excimer formation.

    Science.gov (United States)

    Niwayama, Satomi; Kassar, Abdullatif S; Zhao, Tian; Sutton, Roger B; Altenberg, Guillermo A

    2011-01-01

    Pyrene-containing compounds are commonly used in a number of fluorescence-based applications because they can form excited-state dimers (excimers) by stacking interaction between excited-state and ground-state monomers. Their usefulness arises from the facts that excimer formation requires close proximity between the pyrenes and that the excimer emission spectrum is very different from that of the monomers. One of many applications is to assess proximity between specific sites of macromolecules labeled with pyrenes. This has been done using pyrene maleimide, a reagent that reacts with reduced thiols of cysteines, but its use for structural studies of proteins has been rather limited. This is because the introduction of two cysteines at sufficiently close distance from each other to obtain excimer fluorescence upon labeling with pyrene maleimide requires detailed knowledge of the protein structure or extensive site-directed mutagenesis trials. We synthesized and tested a new compound with a 4-carbon methylene linker placed between the maleimide and the pyrene (pyrene-4-maleimide), with the aim of increasing the sampling distance for excimer formation and making the use of excimer fluorescence simpler and more widespread. We tested the new compound on thiol-modified oligonucleotides and showed that it can detect proximity between thiols beyond the reach of pyrene maleimide. Based on its spectroscopic and chemical properties, we suggest that pyrene-4-maleimide is an excellent probe to assess proximities between cysteines in proteins and thiols in other macromolecules, as well as to follow conformational changes. PMID:22028936

  1. In situ generated hypoiodous acid in an efficient and heterogeneous catalytic system for the homo-oxidative coupling of thiols

    Directory of Open Access Journals (Sweden)

    Ghorbani-Choghamarani Arash

    2013-01-01

    Full Text Available Supported hydrogen peroxide on polyvinylpolypyrrolidone (PVPH2O2, silica sulfuric acid (SiO2-OSO3H and catalytic amounts of potassium iodide (KI has been developed as a heterogeneous medium for the rapid oxidative coupling of thiols into symmetrical homodisulfides. This oxidizing system proceeds under extremely mild conditions and gives no other oxidized side products.

  2. Structure and physical properties of liquid-crystalline polybutadiene-diols modified with thiols containing mesogenic group.

    Czech Academy of Sciences Publication Activity Database

    Ilavsk, Michal; Sedlkov, Zde?ka; Sp?v?ek, Ji?; Jigounov, A.

    Prague : Consortium for Research of Nanostructured and Crosslinked Polymeric Materials , 2005. s. 37. [Workshop Nanofun-Poly on Chemistry, Processing Structure, Properties and Applications of Nanostructured Polymers and Nanocomposites, Life-Cycle Engineering, Gender Issues /3./. 11.11.2005-12.11.2005, Prague] Institutional research plan: CEZ:AV0Z40500505 Keywords : thiols * Liquid crystalline polybutadiene diols Subject RIV: CD - Macromolecular Chemistry

  3. First-principles many-body calculations of electronic conduction in thiol- and amine-linked molecules

    OpenAIRE

    Strange, M.; Rostgaard, C.; Hakkinen, H.; Thygesen, K. S.

    2011-01-01

    The electronic conductance of a benzene molecule connected to gold electrodes via thiol, thiolate, and amino anchoring groups is calculated using nonequilibrium Green functions in combination with the fully selfconsistent GW approximation. The calculated conductance of benzenedithiol and benzenediamine is five times lower than predicted by standard density functional theory (DFT) in very good agreement with experiments. In contrast, the widely studied benzenedithiolate struc...

  4. Antibiofouling hybrid dendritic Boltorn/star PEG thiol-ene cross-linked networks.

    Science.gov (United States)

    Bartels, Jeremy W; Imbesi, Philip M; Finlay, John A; Fidge, Christopher; Ma, Jun; Seppala, Jonathan E; Nystrom, Andreas M; Mackay, Michael E; Callow, James A; Callow, Maureen E; Wooley, Karen L

    2011-06-01

    A series of thiol-ene generated amphiphilic cross-linked networks was prepared by reaction of alkene-modified Boltorn polyesters (Boltorn-ene) with varying weight percent of 4-armed poly(ethylene glycol) (PEG) tetrathiol (0-25 wt%) and varying equivalents of pentaerythritol tetrakis(3-mercaptopropionate) (PETMP) (0-64 wt%). These materials were designed to present complex surface topographies and morphologies, with heterogeneity of surface composition and properties and robust mechanical properties, to serve as nontoxic antibiofouling coatings that are amenable to large-scale production for application in the marine environment. Therefore, a two-dimensional matrix of materials compositions was prepared to study the physical and mechanical properties, over which the compositions spanned from 0 to 25 wt% PEG tetrathiol and 0-64 wt% PETMP (the overall thiol/alkene (SH/ene) ratios ranged from 0.00 to 1.00 equiv), with both cross-linker weight percentages calculated with respect to the weight of Boltorn-ene. The Boltorn-ene components were prepared through the esterification of commercially available Boltorn H30 with 3-butenoic acid. The subsequent cross-linking of the Boltorn-PEG-PETMP films was monitored using IR spectroscopy, where it was found that near-complete consumption of both thiol and alkene groups occurred when the stoichiometry was ca. 48 wt% PETMP (0.75 equiv SH/ene, independent of PEG amount). The thermal properties of the films showed an increase in T(g) with an increase in 4-armed PEG-tetrathiol wt%, regardless of the PETMP concentration. Investigation of the bulk mechanical properties in dry and wet states found that the Young's modulus was the greatest at 48 wt% PETMP (0.75 equiv of SH/ene). The ultimate tensile strength increased when PETMP was constant and the PEG concentration was increased. The Young's modulus was slightly lower for wet films at constant PEG or constant PETMP amounts, than for the dry samples. The nanoscopic surface features were probed using atomic force microscopy (AFM), where it was observed that the surface of the amphiphilic films became increasingly rough with increasing PEG wt%. On the basis of the physicochemical data from the diverse sample matrix, a focused compositional profile was then investigated further to determine the antifouling performance of the cross-linked Boltorn-PEG-PETMP networks. For these studies, a low, constant PETMP concentration of 16 wt% was maintained with variation in the PEG wt% (0-35 wt%). Antifouling and fouling-release activities were tested against the marine alga Ulva. Spore settlement densities were low on these films, compared to that on standards of polydimethylsiloxane and glass. PMID:21644572

  5. Crystallization and preliminary X-ray crystallographic analysis of Sulfolobus solfataricus thioredoxin reductase

    OpenAIRE

    Ruggiero, Alessia; Ruocco, Maria Rosaria; Grimaldi, Pasquale; Arcari, Paolo; Masullo, Mariorosario; Zagari, Adriana; Vitagliano, Luigi

    2005-01-01

    The thioredoxin reductase isolated from S. solfataricus has been crystallized. Diffraction data have been collected from the wild type and from the NADP-bound form of the enzyme to 1.80 and 1.95?, respectively. The structure of the thioredoxin reductase has been solved using MAD diffraction data.

  6. Evaluation of 5?-reductase inhibitory activity of certain herbs useful as antiandrogens.

    Science.gov (United States)

    Nahata, A; Dixit, V K

    2014-08-01

    This study demonstrates 5?-reductase inhibitory activity of certain herbs useful in the management of androgenic disorders. Ganoderma lucidum (Curtis) P. Karst (GL), Urtica dioica Linn. (UD), Caesalpinia bonducella Fleming. (CB), Tribulus terrestris Linn. (TT), Pedalium murex Linn. (PM), Sphaeranthus indicus Linn. (SI), Cuscuta reflexa Roxb. (CR), Citrullus colocynthis Schrad. (CC), Benincasa hispida Cogn. (BH), Phyllanthus niruri Linn. (PN) and Echinops echinatus Linn. (EE) were included in the study. Petroleum ether, ethanol and aqueous extracts of these herbs were tested for their 5?-reductase inhibitory activity against the standard 5?-reductase inhibitor, finasteride. A biochemical method to determine the activity of 5?-reductase was used to evaluate the inhibition of different extracts to the enzyme. The optical density (OD) value of each sample was measured continuously with ultraviolet spectrophotometer for the reason that the substrate NADPH has a specific absorbance at 340 nm. As the enzyme 5?-reductase uses NADPH as a substrate, so in the presence of 5?-reductase inhibitor, the NADPH concentration will increase with the function of time. This method thus implicates the activity of 5?-reductase. The method proved to be extremely useful to screen the herbs for their 5?-reductase inhibitory potential. GL, UD, BH, SI and CR came out to be promising candidates for further exploring their antiandrogenic properties. PMID:23710567

  7. biliverdin. Is there a role for biliverdin reductase?

    Directory of Open Access Journals (Sweden)

    AndreasDaiber

    2012-03-01

    Full Text Available Reactive oxygen species (ROS and signaling events are involved in the pathogenesis of endothelial dysfunction and represent a major contribution to vascular regulation. Molecular signaling is highly dependent on reactive oxygen species. But depending on the amount of ROS production it might have toxic or protective effects. Despite a large number of negative outcomes in large clinical trials (e.g. HOPE, HOPE-TOO, antioxidant molecules and agents are important players to influence the critical balance between production and elimination of RONS. However, chronic systemic antioxidant therapy lacks clinical efficacy, probably by interfering with important physiological redox signaling pathways. Therefore, it may be a much more promising attempt to induce intrinsic antioxidant pathways in order to increase the antioxidants not systemically but at the place of oxidative stress and complications. Among others, heme oxygenase (HO has been shown to be important for attenuating the overall production of ROS in a broad range of disease states through its ability to degrade heme and to produce carbon monoxide (CO, biliverdin/bilirubin, and the release of free iron with subsequent ferritin induction. With the present review we would like to highlight the important antioxidant role of the heme oxygenase system and especially discuss the contribution of the biliverdin, bilirubin and biliverdin reductase to these beneficial effects. The bilierdin reductase was reported to confer an antioxidant redox amplification cycle by which low, physiological bilirubin concentrations confer potent antioxidant protection via recycling of biliverdin from oxidized bilirubin by the biliverdin reductase, linking this sink for oxidants to the NADPH pool. To date the existence and role of this antioxidant redox cycle is still under debate and we present and discuss the pros and cons as well as our own findings on this topic.

  8. Differential labeling of free and disulfide-bound thiol functions in proteins.

    Science.gov (United States)

    Seiwert, Bettina; Hayen, Heiko; Karst, Uwe

    2008-01-01

    A method for the simultaneous determination of the number of free cysteine groups and disulfide-bound cysteine groups in proteins has been developed based on the sequential labeling of free and bound thiol functionalities with two ferrocene-based maleimide reagents. Liquid chromatography/electrochemistry/mass spectrometry was used to assign the N-(2-ferroceneethyl)maleimide (FEM) labeled free cysteine functionalities in a tryptic digest mixture, whereas a precursor ion scan enables the detection of peptides with ferrocenecarboxylic acid-(2-maleimidoyl)ethylamide (FMEA) labeled disulfide-bound cysteine groups after reduction. Fragment spectra of the labeled peptides yield an excellent coverage of b-type and y-type ions. The ferrocene labeled cysteines were fragmented as 412 Da (FEM) and 455 Da (FMEA). These fragment masses are significantly higher than unlabeled amino acids or dipeptides and are easily detected. The position of free and disulfide-bound cysteine may therefore be assigned in an amino acid sequence. PMID:17977013

  9. Dimerization of 1-Phenyl-1H-Tetrazole-5-Thiol over Metalloporphyrin Catalysts

    Directory of Open Access Journals (Sweden)

    Lin Yu

    2012-07-01

    Full Text Available In an alkaline methanol solution, dimerization of 1-phenyl-1H-tetrazole-5-thiol (HL was carried out over metalloporphyrin catalysts under mild conditions. The dimer product, 1,2-bis (1-phenyl-1H-tetrazol-5-yl disulfane (L-L, was characterized by determinations of infrared (IR, HPLC, NMR and elementary analysis respectively. In situ UV-Vis spectroscopic analysis and cyclic voltammetric (CV determinations suggested that the active intermediate for L-L formation is an axially ligated complex, RS-Mn?THPP, which decomposes into a Mn?THPP molecule and a stable radical (SR for coupling to form the disulfane. Meanwhile MnIITHPP molecule can be oxidized easily to form Mn?THPP species again by oxygen from the air for using in next catalytic circle.

  10. Separation and Purification of Palladium from Aqueous Solution by Thiol Derivatives

    International Nuclear Information System (INIS)

    The extraction of palladium from aqueous nitrate solution was studied with solvents containing 3,4 dithio toluene (H2TDT), bis(2,4,4-trimethylpentyl) phosphinic acid-mono thio (CYANEX 302) and dithio (CYNAEX 301) analogues. The effect of different parameters affecting the extraction of palladium was investigated. Comparative data are presented to illustrate the effect of composition (sulphur substitution) on the extraction and stripping characteristics of the three reagents. The selectivity range of the studied extractants was found to be in the order H2TDT > CYNAEX 301 > CYNAEX 302. The extraction of the Pd(II) complexes with respect to metal-thiol complexes has also been studied. Experimental data relating to palladium have been analysed numerically to determine the stoichiometry of the extracted species and its equilibrium constant. It was found that Pd(II) was extracted into the organic phase by the formation of the species RS-Pd(II).

  11. Single molecular switch based on thiol tethered iron(II)clathrochelate on gold

    International Nuclear Information System (INIS)

    Molecular electronics has been associated with high density nano-electronic devices. Developments of molecular electronic devices were based on reversible switching of molecules between the two conductive states. In this paper, self-assembled monolayers of dodecanethiol (DDT) and thiol tethered iron(II)clathrochelate (IC) have been prepared on gold film. The electrochemical and electronic properties of IC molecules inserted into the dodecanethiol monolayer (IC-DDT SAM) were investigated using voltammetric, electrochemical impedance spectroscopy (EIS), scanning tunneling microscopy (STM) and cross-wire tunneling measurements. The voltage triggered switching behaviour of IC molecules on mixed SAM was demonstrated. Deposition of polyaniline on the redox sites of IC-DDT SAM using electrochemical polymerization of aniline was performed in order to confirm that this monolayer acts as nano-patterned semiconducting electrode surface.

  12. An Improved Isotope Coded Affinity Tag Technology for Thiol Redox Proteomics

    Directory of Open Access Journals (Sweden)

    Ning Zhu

    2012-01-01

    Full Text Available Isotope Coded Affinity Tag (ICAT is a gel-free technology for quantitative proteomics. In ICAT procedure, strong cation exchange chromatography (SCX using increased potassium chloride gradient is recommended for peptide fractionation. Here we report optimization of hydrophilic interaction chromatography (HILIC as an alternative strategy for peptide fractionation of ICAT samples. HILIC exhibits high separation efficiency and does not require any downstream desalting steps. Compared to SCX based ICAT, integration of HILIC into the ICAT technology has resulted in high rates of protein identification, cysteine mapping, and quantification of cysteine-containing peptides. The improved technology has shown utility in thiol redox proteomics. Interestingly, results from HILIC ICAT and SCX ICAT are complementary. Implementation of both provides high coverage analysis of a complex proteome.

  13. An excellent ozone-resistant polymethylvinylsiloxane coating on natural rubber by thiol-ene click chemistry

    Directory of Open Access Journals (Sweden)

    N. Y. Ning

    2015-06-01

    Full Text Available Natural rubber (NR as renewable resource is a kind of cheap and versatile elastomer. A disadvantage of NR is that the ozone resistance is not good, which needs to be improved for its wider application. In this study, polymethylvinylsiloxane (PMVS coating on natural rubber (NR was realized for the first time by using thiol-ene click reaction under UV irradiation, simultaniously realizing the fast crosslinking of PMVS layer and the covalent crosslinking between PMVS layer and NR layer. As a result, a good interphase adhesion between PMVS coating and NR was obtained. The coating of the crosslinked PMVS layer on NR resulted in an obvious increase in the ozone resistance of NR. Our study provides a new and high efficient strategy to prepare elastomer materials with good ozone resistance.

  14. Differential regulation of tissue thiol-disulfide redox status in a murine model of peritonitis

    Directory of Open Access Journals (Sweden)

    Benton Shana M

    2012-10-01

    Full Text Available Abstract Background Glutathione (GSH/glutathione disulfide (GSSG and cysteine (Cys/cystine (CySS are major redox pools with important roles in cytoprotection. We determined the impact of septic peritonitis on thiol-disulfide redox status in mice. Methods FVB/N mice (612 week old; 8/group underwent laparotomy with cecal ligation and puncture (CLP or laparotomy alone (control. Sections of ileum, colon, lung and liver were obtained and GSH, GSSG, Cys and CySS concentrations determined by HPLC 24 h after laparotomy. Redox potential [Eh in millivolts (mV] of the GSH/GSSG and Cys/CySS pools was calculated using the Nernst equation. Data were analyzed by ANOVA (mean SE. Results GSH/GSSG Eh in ileum, colon, and liver was significantly oxidized in septic mice versus control mice (ileum: septic ?2024 versus control ?2282 mV; colon: -1958 versus ?2141 mV; and liver: -1943 vs. -2101 mV, all Ph was unchanged with CLP, while liver and lung Cys/CySS Eh became significantly more reducing (liver: septic = ?1033 versus control ?902 mV; lung: -1015 versus ?811 mV, each P Conclusions Septic peritonitis induced by CLP oxidizes ileal and colonic GSH/GSSG redox but Cys/CySS Eh remains unchanged in these intestinal tissues. In liver, CLP oxidizes the GSH/GSSG redox pool and CyS/CySS Eh becomes more reducing; in lung, CLP does not alter GSH/GSSG Eh, and Cys/CySS Eh is less oxidized. CLP-induced infection/inflammation differentially regulates major thiol-disulfide redox pools in this murine model.

  15. Thiol click modification of cyclic disulfide containing biodegradable polyurethane urea elastomers.

    Science.gov (United States)

    Fang, Jun; Ye, Sang-Ho; Wang, Jing; Zhao, Ting; Mo, Xiumei; Wagner, William R

    2015-05-11

    Although the thiol click reaction is an attractive tool for postpolymerization modification of thiolmers, thiol groups are easily oxidized, limiting the potential for covalent immobilization of bioactive molecules. In this study, a series of biodegradable polyurethane elastomers incorporating stable cyclic disulfide groups was developed and characterized. These poly(ester urethane)urea (PEUU-SS) polymers were based on polycaprolactone diol (PCL), oxidized dl-dithiothreitol (O-DTT), lysine diisocyanate (LDI), or butyl diisocyanate (BDI), with chain extension by putrescine. The ratio of O-DTT:PCL was altered to investigate different levels of potential functionalization. PEG acrylate was employed to study the mechanism and availability of both bulk and surface click modification of PEUU-SS polymers. All synthesized PEUU-SS polymers were elastic with breaking strengths of 38-45 MPa, while the PEUU-SS(LDI) polymers were more amorphous, possessing lower moduli and relatively small permanent deformations versus PEUU-SS(BDI) polymers. Variable bulk click modification of PEUU-SS(LDI) polymers was achieved by controlling the amount of reduction reagent, and rapid reaction rates occurred using a one-pot, two-step process. Likewise, surface click reaction could be carried out quickly under mild, aqueous conditions. Furthermore, a maleimide-modified affinity peptide (TPS) was successfully clicked on the surface of an electrospun PEUU-SS(BDI) fibrous sheet, which improved endothelial progenitor cell adhesion versus corresponding unmodified films. The cyclic disulfide containing biodegradable polyurethanes described provide an option for cardiovascular and other soft tissue regenerative medicine applications where a temporary, elastic scaffold with designed biofunctionality from a relatively simple click chemistry approach is desired. PMID:25891476

  16. Visible light initiated thiol-ene based reflection H-PDLCs

    Science.gov (United States)

    Natarajan, Lalgudi V.; Brown, Dean P.; Wofford, Jeremy M.; Tondiglia, Vincent P.; Sutherland, Richard L.; Lloyd, Pam; Jakubiak, Rachel; Vaia, Richard; Bunning, Timothy J.

    2005-08-01

    Multifunctional acrylate formulations containing nematic liquid crystals have been shown to form holographic polymer dispersed liquid crystal gratings (H-PDLCs) easily using ultra-violet AND/OR visible photoinitiators. Laser wavelengths of 364, 476, 488, 514, 532 and 647 nm have been used for the fabrication of the gratings. Recently, the use of a thiol-ene based monomer system has been shown to overcome some of the adverse effects like post polymerization, voltage creep, and non-uniform shrinkage incurred when using highly functional acrylate monomers. However, Bragg reflection gratings have only been demonstrated utilizing ultra-violet (UV) (363.8 nm Argon ion) photopolymerization. Using UV irradiation and single prism geometry limits the upper end of the reflection notch wavelength. In this work, we report on new visible photoinitiator systems developed for the formation of reflective H-PDLCs using thiol-ene monomers. Using these new photoinitiator systems, reflection notches have been routinely written from the visible to the near infrared (IR) regions. The visible photoinitiator systems included the photoinitiator and radical generator titanocene organo-metallic complex (commercially known as Irgacure 784 (Ciba-Geigy), Rhodamine 6G, Pyrromethene, and a radical generating organic peroxide as coinitiator. Reflection gratings were written using laser wavelengths 442, 488, and 532 nm with diffraction efficiencies (DEs) above 70%. Angle tuning allowed for gratings with reflection notches in the near IR (900-1500 nm) to be written using these initiator systems. Rhodamine 6G was found to be more efficient than the other two initiators. We discuss here this new chemistry, the morphology, and electro-optical properties of the reflection gratings.

  17. Synergistic Effects of Bismuth Thiols and Various Antibiotics Against Pseudomonas aeruginosa Biofilm

    Directory of Open Access Journals (Sweden)

    Maryam Varposhti

    2014-03-01

    Full Text Available Background: Pseudomonas aeruginosa is an opportunistic pathogen that takes advantages of some weaknesses in the immune system to initiate an infection. Biofilms of P. aeruginosa can cause chronic opportunistic infections in immunocompromised and elderly patients. This bacterium is considered as a model organism to study antibiotic resistance as well as biofilm formation. In the biofilm structures, bacteria are protected from many harmful environmental factors such as fluctuations in the level of oxygen and nutrients, and the alterations of pH as well as sensitivity to antibiotics. Decreased permeability of biofilms is one of the important reasons of antimicrobial resistance in bacteria. Objectives: In this study the anti-biofilm activity of bismuth thiols in combination with ciprofloxacin, imipenem and ceftazidime against the P. aeruginosa biofilm was investigated. Materials and Methods: Checkerboard method was used to test the susceptibility of biofilms against various antimicrobial combinations. The biofilm formation was measured by 2,3-bis (2-methoxy-4-nitro-5-sulfo-phenyl-2H-tetrazolium-5-carboxanilide (XTT colorimetric assay. The fractional bio-film inhibitory concentration was reported for each agent. Results: The combination of bismuth ethanedithiol with ciprofloxacin showed synergistic inhibitory effect on the P. aeruginosa biofilm formation. The combination of bismuth ethanedithiol ciprofloxacin, ceftazidime and imipenem showed synergistic inhibitory effects on the biofilm formation. Furthermore, the combination of bismuth ethanedithiol, imipenem and ceftazidime did not show any synergistic inhibitory effect on biofilm formation. Conclusions: Our studies show that using appropriate concentrations of bismuth thiols in combination with various antibiotics can act synergistically against P. aeruginosa biofilm formation. Keywords: Biofilms; Pseudomonas aeruginosa; Antibacterial Agents

  18. Vibrio harveyi Nitroreductase Is Also a Chromate Reductase

    OpenAIRE

    Kwak, Young Hak; Lee, Dong Seok; Kim, Han Bok

    2003-01-01

    The chromate reductase purified from Pseudomonas ambigua was found to be homologous with several nitroreductases. Escherichia coli DH5? and Vibrio harveyi KCTC 2720 nitroreductases were chosen for the present study, and their chromate-reducing activities were determined. A fusion between glutathione S-transferase (GST) and E. coli DH5? NfsA (GST-EcNfsA), a fusion between GST and E. coli DH5? NfsB (GST-EcNfsB), and a fusion between GST and V. harveyi KCTC 2720 NfsA (GST-VhNfsA) were prepare...

  19. Biofilm Modifies Expression of Ribonucleotide Reductase Genes in Escherichia coli

    OpenAIRE

    Cendra, Maria del Mar; Jurez, Antonio; Torrents, Eduard

    2012-01-01

    Ribonucleotide reductase (RNR) is an essential enzyme for all living organisms since is the responsible for the last step in the synthesis of the four deoxyribonucleotides (dNTPs) necessary for DNA replication and repair. In this work, we have investigated the expression of the three-RNR classes (Ia, Ib and III) during Escherichia coli biofilm formation. We show the temporal and spatial importance of class Ib and III RNRs during this process in two different E. coli wild-type strains, the com...

  20. Methylenetetrahydrofolate reductase (MTHFR) deficiency presenting as a rash.

    LENUS (Irish Health Repository)

    Crushell, Ellen

    2012-09-01

    We report on the case of a 2-year-old girl recently diagnosed with Methylenetetrahydrofolate reductase (MTHFR) deficiency who originally presented in the neonatal period with a distinctive rash. At 11 weeks of age she developed seizures, she had acquired microcephaly and developmental delay. The rash deteriorated dramatically following commencement of phenobarbitone; both rash and seizures abated following empiric introduction of pyridoxine and folinic acid as treatment of possible vitamin responsive seizures. We postulate that phenobarbitone in combination with MTHFR deficiency may have caused her rash to deteriorate and subsequent folinic acid was helpful in treating the rash and preventing further acute neurological decline as commonly associated with this condition.

  1. Inhibition of aldose reductase prevents colon cancer metastasis

    OpenAIRE

    Tammali, Ravinder; Reddy, Aramati B. M.; Saxena, Ashish; Rychahou, Piotr G; Evers, B.Mark; Qiu, Suimin; Awasthi, Sanjay; Ramana, Kota V.; Srivastava, Satish K

    2011-01-01

    Colon cancer is the third most common cause of cancer and is the second leading cause of cancer deaths in the USA. Although inhibition of aldose reductase (AR) is known to prevent human colon cancer cell growth in nude mice xenografts, the role of AR in the regulation of cancer metastasis is not known. We now demonstrate the mechanisms by which AR regulates colon cancer metastasis in vitro and in vivo. Inhibition of AR prevented the epidermal growth factor (EGF) or fibroblast growth factor (F...

  2. Foodomics platform for the assay of thiols in wines with fluorescence derivatization and ultra performance liquid chromatography mass spectrometry using multivariate statistical analysis.

    Science.gov (United States)

    Inoue, Koichi; Nishimura, Maiko; Tsutsui, Haruhito; Min, Jun Zhe; Todoroki, Kenichiro; Kauffmann, Jean-Michel; Toyo'oka, Toshimasa

    2013-02-13

    The presence of specific volatile and aminothiols in wine is associated with quality, worth, price, and taste. The identification of specific thiol-containing compounds in various wines has been reported in many valuable and interesting works. In this study, a novel foodomics assay of thiol-containing compounds, such as free aminothiols and related conjugates, was developed using ultra performance liquid chromatography (UPLC) with fluorescence (FL) and electrospray (ESI) time-of-flight mass spectrometric (TOF/MS) detections. FL specific derivatization was applied along with multivariate statistical analysis. First, the optimal experimental conditions were studied using representative thiols, such as l-cysteine, N-acetyl-l-cysteine, cysteamine, and l-glutathione, and then the UPLC-FL derivatization and separation steps were fixed for the subsequent screening of unknown thiol-containing compounds. The screening assay consisted of monitoring the UPLC-TOF/MS peaks of unknown thiols, which decreased due to the derivatization as compared to the nonderivatized thiols. The principal component analysis of the UPLC-TOF/MS data could be well-differentiated and categorized into two groups. The orthogonal signal correction partial least-squares discriminant analysis, the so-called S-plot, showed that the quality differentiation is directly related to the decrease of native thiols and increase of derivatized thiols. With this strategy, the mass difference from the derivatization reagent (+m/z 198) could be utilized for the identification of these thiols using the FL peaks retention time and metabolomics-databases. The presence of l-glutathione in rice wine was for the first time reported on the basis of the available metabolomics-databases and standard matching. This novel concept based on foodomics could be applied in food analysis for the ready screening of specific functional compounds by exploiting the various derivatization modes available. PMID:23339461

  3. Aldose reductase catalysis and crystallography. Insights from recent advances in enzyme structure and function.

    Science.gov (United States)

    Petrash, J M; Tarle, I; Wilson, D K; Quiocho, F A

    1994-08-01

    Enhanced metabolism of glucose via the polyol pathway may play an important role in the pathogenesis of diabetic retinopathy, neuropathy, and nephropathy. Aldose reductase catalyzes the NADPH-dependent conversion of glucose to sorbitol, the first step in the polyol pathway. Interruption of the polyol pathway by inhibition of aldose reductase holds considerable promise as a therapeutic measure to prevent or delay the onset and severity of these late complications of diabetes. Dramatic advances in our understanding of the molecular biology, enzymology, and three-dimensional structure of aldose reductase have occurred in recent years, providing new and challenging insights into the enzyme's catalytic mechanism. Recent developments in structure determination of aldose reductase and the implications for evaluation and development of aldose reductase inhibitors are summarized. PMID:8039602

  4. Trypanothione Reductase: A Viable Chemotherapeutic Target for Antitrypanosomal and Antileishmanial Drug Design

    Directory of Open Access Journals (Sweden)

    M. Omar F. Khan

    2007-01-01

    Full Text Available Trypanosomiasis and leishmaniasis are two debilitating disease groups caused by parasites of Trypanosoma and Leishmania spp. and affecting millions of people worldwide. A brief outline of the potential targets for rational drug design against these diseases are presented, with an emphasis placed on the enzyme trypanothione reductase. Trypanothione reductase was identified as unique to parasites and proposed to be an effective target against trypanosomiasis and leishmaniasis. The biochemical basis of selecting this enzyme as a target, with reference to the simile and contrast to human analogous enzyme glutathione reductase, and the structural aspects of its active site are presented. The process of designing selective inhibitors for the enzyme trypanothione reductase has been discussed. An overview of the different chemical classes of inhibitors of trypanothione reductase with their inhibitory activities against the parasites and their prospects as future chemotherapeutic agents are briefl y revealed.

  5. Aminoadipate reductase gene: a new fungal-specific gene for comparative evolutionary analyses

    Directory of Open Access Journals (Sweden)

    Miura Yoshiharu

    2002-04-01

    Full Text Available Abstract Background In fungi, aminoadipate reductase converts 2-aminoadipate to 2-aminoadipate 6-semialdehyde. However, other organisms have no homologue to the aminoadipate reductase gene and this pathway appears to be restricted to fungi. In this study, we designed degenerate primers for polymerase chain reaction (PCR amplification of a large fragment of the aminoadipate reductase gene for divergent fungi. Results Using these primers, we amplified DNA fragments from the archiascomycetous yeast Saitoella complicata and the black-koji mold Aspergillus awamori. Based on an alignment of the deduced amino acid sequences, we constructed phylogenetic trees. These trees are consistent with current ascomycete systematics and demonstrate the potential utility of the aminoadipete reductase gene for phylogenetic analyses of fungi. Conclusions We believe that the comparison of aminoadipate reductase among species will be useful for molecular ecological and evolutionary studies of fungi, because this enzyme-encoding gene is a fungal-specific gene and generally appears to be single copy.

  6. 20(S)-Ginsenoside Rh2 as aldose reductase inhibitor from Panax ginseng.

    Science.gov (United States)

    Fatmawati, Sri; Ersam, Taslim; Yu, Hongshan; Zhang, Chunzhi; Jin, Fengxie; Shimizu, Kuniyoshi

    2014-09-15

    The root of Panax ginseng C. A. Meyer (Araliaceae) is a well-known herbal medicine in East Asia. The major bioactive metabolites in this root are commonly identified as ginsenosides. A series of ginsenosides were determined for in vitro human recombinant aldose reductase. This Letter aims to clarify the structural requirement for aldose reductase inhibition. We discovered that only ginsenoside 20(S)-Rh2 showed potent against aldose reductase, with an IC50 of 147.3 ?M. These results implied that the stereochemistry of the hydroxyl group at C-20 may play an important role in aldose reductase inhibition. An understanding of these requirements is considered necessary in order to develop a new type of aldose reductase inhibitor. Furthermore, P. ginseng might be an important herbal medicine in preventing diabetic complications. PMID:25152999

  7. The effect of ionic and non-ionic surfactants on the growth, nitrate reductase and nitrite reductase activities of Spirodela polyrrhiza (L. Schleiden

    Directory of Open Access Journals (Sweden)

    Jzef Buczek

    2014-02-01

    Full Text Available Inclusion into the medium of 5 mgdm-3 of non-ionic (ENF or ionic (DBST surfactant caused 50-60% inhibition of nitrite reductase MR activity in S. polyrrhiza. At the same time, increased accumulation of NO2- in the plant tissues and lowering of the total and soluble protein contents were found. DBST also lowered the nitrate reductase (NR activity and the dry mass of the plants.

  8. Complex formation of copper (II) with 1,2,4-three-azole-thiol-5 at ionic strength 0.01 mol/l

    International Nuclear Information System (INIS)

    The process of complex formation of copper (II) with 1,2,4-three-azole-thiol-5 at ionic strength 0.01 mol/l NaNo3 was studied by means of potentiometric method. The values of stepped constants of formation of 1,2,4-three-azole-thiol complexes of copper (II) in temperature intervals 273-338 K were defined. The rates of thermodynamical functions of complex formation process in the system CuCl2-1,2,4-three-azole-thiol-5-H2O were evaluated. (author)

  9. Thioredoxin-thioredoxin reductase system of Streptomyces clavuligerus: sequences, expression, and organization of the genes.

    Science.gov (United States)

    Cohen, G; Yanko, M; Mislovati, M; Argaman, A; Schreiber, R; Av-Gay, Y; Aharonowitz, Y

    1993-08-01

    The genes that encode thioredoxin and thioredoxin reductase of Streptomyces clavuligerus were cloned, and their DNA sequences were determined. Previously, we showed that S. clavuligerus possesses a disulfide reductase with broad substrate specificity that biochemically resembles the thioredoxin oxidoreductase system and may play a role in the biosynthesis of beta-lactam antibiotics. It consists consists of two components, a 70-kDa NADPH-dependent flavoprotein disulfide reductase with two identical subunits and a 12-kDa heat-stable protein general disulfide reductant. In this study, we found, by comparative analysis of their predicted amino acid sequences, that the 35-kDa protein is in fact thioredoxin reductase; it shares 48.7% amino acid sequence identity with Escherichia coli thioredoxin reductase, the 12-kDa protein is thioredoxin, and it shares 28 to 56% amino acid sequence identity with other thioredoxins. The streptomycete thioredoxin reductase has the identical cysteine redox-active region--Cys-Ala-Thr-Cys--and essentially the same flavin adenine dinucleotide- and NADPH dinucleotide-binding sites as E. coli thioredoxin reductase and is partially able to accept E. coli thioredoxin as a substrate. The streptomycete thioredoxin has the same cysteine redox-active segment--Trp-Cys-Gly-Pro-Cys--that is present in virtually all eucaryotic and procaryotic thioredoxins. However, in vivo it is unable to donate electrons to E. coli methionine sulfoxide reductase and does not serve as a substrate in vitro for E. coli thioredoxin reductase. The S. clavuligerus thioredoxin (trxA) and thioredoxin reductase (trxB) genes are organized in a cluster. They are transcribed in the same direction and separated by 33 nucleotides. In contrast, the trxA and trxB genes of E. coli, the only other organism in which both genes have been characterized, are physically widely separated. PMID:8349555

  10. Functions of Flavin Reductase and Quinone Reductase in 2,4,6-Trichlorophenol Degradation by Cupriavidus necator JMP134?

    OpenAIRE

    Belchik, Sara Mae; XUN, Luying

    2007-01-01

    The tcpRXABCYD operon of Cupriavidus necator JMP134 is involved in the degradation of 2,4,6-trichlorophenol (2,4,6-TCP), a toxic pollutant. TcpA is a reduced flavin adenine dinucleotide (FADH2)-dependent monooxygenase that converts 2,4,6-TCP to 6-chlorohydroxyquinone. It has been implied via genetic analysis that TcpX acts as an FAD reductase to supply TcpA with FADH2, whereas the function of TcpB in 2,4,6-TCP degradation is still unclear. In order to provide direct biochemical evidence for t...

  11. [Structure and function of peroxisomal tetrameric carbonyl reductase].

    Science.gov (United States)

    Imamura, Yorishige

    2008-11-01

    In this paper, the structure and function of a new tetrameric carbonyl reductase (TCR) is reviewed. TCRs were purified from rabbit and pig heart, using 4-benzoylpyridine as a substrate. Partial peptide sequencing and cDNA cloning of rabbit and pig TCRs revealed that both enzymes belonged to the short-chain dehydrogenase/reductase family and that their subunits consisted of 260 amino acid residues. Rabbit and pig TCRs catalyzed the reduction of alkyl phenyl ketones, alpha-dicarbonyl compounds, quinones and retinals. Both enzymes were potently inhibited by flavonoids and fatty acids. 9,10-Phenanthrenequinone, which is efficiently reduced by rabbit and pig TCRs, mediated the formation of superoxide radical through its redox cycling in pig heart. The C-terminal sequences of rabbit and pig TCRs comprised a type 1 peroxisomal targeting signal (PTS1) Ser-Arg-Leu, suggesting that the enzymes are localized in the peroxisome. In fact, pig TCR was targeted into the peroxisomal matrix, in the case of transfection of HeLa cells with vectors expressing the enzyme. However, when the recombinant pig TCR was directly introduced into HeLa cells, the enzyme was not targeted into the peroxisomal matrix. The crystal structure of recombinant pig TCR demonstrated that the C-terminal PTS1 of each subunit of the enzyme was buried in the interior of the tetrameric molecule. These findings indicate that pig TCR is imported into the peroxisome as a monomer and then forms an active tetramer within this organelle. PMID:18981702

  12. The effect of 5?-reductase-2 deficiency on human fertility.

    Science.gov (United States)

    Kang, Hey-Joo; Imperato-McGinley, Julianne; Zhu, Yuan-Shan; Rosenwaks, Zev

    2014-02-01

    A most interesting and intriguing male disorder of sexual differentiation is due to 5?-reductase-2 isoenzyme deficiency. These male infants are born with ambiguous external genitalia due to a deficiency in their ability to catalyze the conversion of T to dihydrotestosterone. Dihydrotestosterone is a potent androgen responsible for differentiation of the urogenital sinus and genital tubercle into the external genitalia, urethra, and prostate. Affected males are born with a clitoral-like phallus, bifid scrotum, hypospadias, blind shallow vaginal pouch from incomplete closure of the urogenital sinus, and a rudimentary prostate. At puberty, the surge in mainly T production prompts virilization, causing most boys to choose gender reassignment to male. Fertility is a challenge for affected men for several reasons. Uncorrected cryptorchidism is associated with low sperm production, and there is evidence of defective transformation of spermatogonia into spermatocytes. The underdeveloped prostate and consequent low semen volumes affect sperm transport. In addition, semen may not liquefy due to a lack of prostate-specific antigen. In the present review, we discuss the 5?-reductase-2 deficiency syndrome and its impact on human fertility. PMID:24412121

  13. Spectroscopic, thermodynamic and kinetic properties of Candida nitratophila nitrate reductase.

    Science.gov (United States)

    Kay, C J; Barber, M J; Solomonson, L P; Kau, D; Cannons, A C; Hipkin, C R

    1990-01-01

    Visible spectra of oxidized and reduced Candida nitratophila assimilatory NAD(P)H:nitrate reductase yielded absorbance maxima of 413 nm and 423 nm, and 525 nm and 555 nm respectively, characteristic of a b5-type cytochrome. E.p.r. spectra of the partially reduced enzyme revealed a single Mo(V) species (g1 = 1.9957, g2 = 1.9664 and g3 = 1.9658) exhibiting superhyperfine coupling to a single proton [A(1H)av. = 1.4 mT]. Oxidation-reduction midpoint potentials (E'0) (25 degrees C, pH 7) for the haem and Mo-pterin prosthetic groups were determined by visible and e.p.r. potentiometric titrations and yielded values of E'0 = -174 mV (n = 1) for the haem and E'0 = -3 mV and E'0 = -27 mV for the Mo(VI)/Mo(V) and Mo(V)/Mo(IV) couples respectively. Comparison of initial rates of the NADH-oxidizing and nitrate-reducing partial activities at various ionic strengths indicated electron transfer from reduced haem to Mo was rate-limiting during turnover. These results suggest a close similarity between Candida nitratophila and Chlorella vulgaris nitrate reductases. PMID:2268283

  14. A mutant of barley lacking NADH-hydroxypyruvate reductase

    International Nuclear Information System (INIS)

    A mutant of barley, LaPr 88/29, deficient in peroxisomal NADH-hydroxypyruvate reductase (HPR) activity has been identified. Compared to the wild type the activities of NADH-HPR and NADPH-HPR were severely reduced but the mutant was still capable of fixing CO2 at rates equivalent to 75% of that of the wild type in air. Although lacking an enzyme in the main photorespiratory pathway, there appeared to be little disruption to photorespiratory metabolism as ammonia release, CO2 efflux and 14CO2 release from L-[U-14C] serine were similar in both mutant and wild type. LaPr 88/29 has been used to show that NADH-glyoxylate reductase (GR) and NADH-HPR are probably not catalyzed by the same enzyme in barley and that over 80% of the NADPH-HPR activity is due to the NADH-HPR enzyme. Immunological studies, using antibodies raised against spinach HPR, have shown that the NADH-dependent enzyme protein is absent in LaPr 88/29 but there appears to be enhanced synthesis of the NADPH-dependent enzyme protein

  15. The mixed-valent copper thiolate complex hexakis{?3-2-[(1,3-dimethylimidazolidene)amino]benzenethiolato}dicopper(II)tetracopper(I) bis(hexafluoridophosphate) acetonitrile disolvate dichloromethane disolvate

    OpenAIRE

    Neuba, Adam; Flo?rke, Ulrich; Henkel, Gerald

    2012-01-01

    The molecular structure of the title compound, [Cu4 ICu2 II(C11H14N3S)6](PF6)22CH3CN2CH2Cl2, shows a mixed-valent copper(I/II) thiolate complex with a distorted tetrahedral coordination of the CuI and CuII cations by one guanidine N atom and three S atoms each. Characteristic features of the Cu6S6 skeleton are a total of six chemically identical ?3-thiolate bridges and almost planar Cu2S2 units with a maximum deviation of 0.110?(1)? from the best plane. Each Cu2S2 uni...

  16. Functional complementation of a nitrate reductase defective mutant of a green alga Dunaliella viridis by introducing the nitrate reductase gene.

    Science.gov (United States)

    Sun, Yu; Gao, Xiaoshu; Li, Qiyun; Zhang, Qingqi; Xu, Zhengkai

    2006-08-01

    Nitrate reductase (NR) catalyzes NAD (P) H dependent reduction of nitrate to nitrite. Transformation systems have been established in several species of green algae by nitrate reductase gene functional complementation. In this report, an endogenous NR cDNA (3.4 kb) and a genomic fragment (14.6 kb) containing the NR gene (DvNIA1) were isolated from the D. viridis cDNA and genomic libraries respectively. Southern blot and Northern blot analyses showed that this gene exists as a single copy in D. viridis and is induced by nitrate. To obtain a NR defective mutant as a recipient strain, D. viridis cells were treated with a chemical mutagen and then cultured on a chlorate-containing plate to enrich chlorate tolerant mutants. Southern analysis showed that one isolate, B14, had a deletion in the DvNIA1 gene region. Using electroporation conditions determined in this laboratory, plasmid pDVNR containing the intact DvNIA1 gene has been electroporated into the defective mutant B14. Strains retaining a nitrate assimilation phenotype were obtained from nitrate plates after spreading the electroporated cells. In some individual strains, transcription of the introduced gene was detected. NR activity in these strains was slightly higher than that in the defective B14 cell, but excretion of nitrite into culture media was almost as high as that of the wild-type cell. Possible episomal presence of the introduced DNA in D. viridis is discussed. PMID:16797881

  17. Assaying phenothiazine derivatives as trypanothione reductase and glutathione reductase inhibitors by theoretical docking and molecular dynamics studies.

    Science.gov (United States)

    Iribarne, F; Paulino, M; Aguilera, S; Tapia, O

    2009-11-01

    A theoretical docking study, conducted on a sample of previously reported phenothiazine derivatives, at the binding sites of Trypanosoma cruzi trypanothione reductase (TR) and human erythrocyte glutathione reductase (GR), examines interaction energies (affinities) towards the parasite enzyme to check for selectivity with respect to the human counterpart. Phenothiazine compounds were previously shown to be TR inhibitors. The analysis of data collected from the docking procedure was undertaken both from the numeric and graphical standpoints, including the comparison of force field, energies, molecular contacts and spatial location of the different orientations that ligands acquired at the binding sites. Molecular Dynamics simulations were also carried out for derivatives with known quantitative inhibition kinetics (K(i)). The results indicate that (positively) charged phenothiazines attain larger interaction energies at TR active site, in line with previous experimental information. Suitable molecular size and shape is also needed to complement the electrostatic effect, as clearly evidenced by graphical analysis of output docked conformations. Docking energies values are reasonably well correlated with those obtained by Molecular Dynamics as well as with the experimental K(i) values, confirming once again the validity of this type of scoring methods to rapidly assess ligand-receptor affinities. Alongside newly discovered classes of TR inhibitors, the promazine (N-alkylaminopropylphenothiazine) nucleus should still be considered when good candidates are sought as leaders for selective TR inhibition. PMID:19801198

  18. Curcumin is a tight-binding inhibitor of the most efficient human daunorubicin reductase - Carbonyl reductase 1.

    Science.gov (United States)

    Hintzpeter, Jan; Hornung, Jan; Ebert, Bettina; Martin, Hans-Jrg; Maser, Edmund

    2015-06-01

    Curcumin is a major component of the plant Curcuma longa L. It is traditionally used as a spice and coloring in foods and is an important ingredient in curry. Curcuminoids have anti-oxidant and anti-inflammatory properties and gained increasing attention as potential neuroprotective and cancer preventive compounds. In the present study, we report that curcumin is a potent tight-binding inhibitor of human carbonyl reductase 1 (CBR1, Ki=223nM). Curcumin acts as a non-competitive inhibitor with respect to the substrate 2,3-hexandione as revealed by plotting IC50-values against various substrate concentrations and most likely as a competitive inhibitor with respect to NADPH. Molecular modeling supports the finding that curcumin occupies the cofactor binding site of CBR1. Interestingly, CBR1 is one of the most effective human reductases in converting the anthracycline anti-tumor drug daunorubicin to daunorubicinol. The secondary alcohol metabolite daunorubicinol has significantly reduced anti-tumor activity and shows increased cardiotoxicity, thereby limiting the clinical use of daunorubicin. Thus, inhibition of CBR1 may increase the efficacy of daunorubicin in cancer tissue and simultaneously decrease its cardiotoxicity. Western-blots demonstrated basal expression of CBR1 in several cell lines. Significantly less daunorubicin reduction was detected after incubating A549 cell lysates with increasing concentrations of curcumin (up to 60% less with 50?M curcumin), suggesting a beneficial effect in the co-treatment of anthracycline anti-tumor drugs together with curcumin. PMID:25541467

  19. MK386: a potent, selective inhibitor of the human type 1 5alpha-reductase.

    Science.gov (United States)

    Ellsworth, K; Azzolina, B; Baginsky, W; Bull, H; Chang, B; Cimis, G; Mitra, S; Toney, J; Bakshi, R K; Rasmusson, G R; Tolman, R L; Harris, G S

    1996-07-01

    Steroid 5alpha-reductase is required for the conversion of testosterone to dihydrotestosterone. Localization of type 1 5alpha-reductase in the sebaceous gland of skin offers the possibility for selective inhibition of this isozyme as a treatment for acne. The goals of these studies are to demonstrate the mechanism of inhibition of MK386 and its selectivity for type 1 5alpha-reductase. The apparent potency of MK386 differed depending on the source of the enzyme (i.e. recombinant vs. native), yet selectivity for type 1 5alpha-reductase was unchanged. Our results indicate that the apparent potency of MK386 is modulated by the membrane concentration of the assay. These results suggest that MK386 has a high affinity for the lipid-rich membrane environment of 5alpha-reductase. MK386 was also found to be a slow binding inhibitor of type 1 5alpha-reductase. However, the cause of this time-dependent inhibition is unrelated to partitioning of the inhibitor into the membrane because similar studies with type 2 5alpha-reductase indicate that MK386 is a reversible, competitive inhibitor. A number of counterscreens were developed to demonstrate that MK386 is a poor inhibitor of other steroid metabolizing enzymes. PMID:8903421

  20. Strategies for creating antifouling surfaces using selfassembled poly(ethylene glycol) thiol molecules

    DEFF Research Database (Denmark)

    Lokanathan, Arcot R.

    2011-01-01

    Microorganisms are one of the most important parts of our ecosystem influencing the sustenance of human society. The beneficial microbes are of high relevance to food industry, development of antibiotics and processing of many raw materials. Mankind has indeed benefitted a lot from large number of microbial species, but then the environment is also teeming with pathogenic microbes that pose serious threat to human health. Hence the success of human survival not only depends on exploiting the useful microbes but also on our ability to defend ourselves against the pathogenic ones. Microbes such as bacteria can exist either in free floating planktonic form or as biofilm which can defined as adherent microbial colonies embedded in slime or extracellular polymeric substance. Microbial biofilms are responsible for a large number of problems posing a serious safety threat within our society. Biofilms have substantial impact on human health, as many bacterial infections are caused by or involve biofilms. Biofilm infections are for example often associated with medical implants, as artificial surfaces in the human body provide a safe haven where biofilms can form. The food industry daily combats biofilms forming on the surfaces of equipment to avoid contamination of their products. In addition to posing a health problem, biofilms also pose technical problems, such as corrosion and reduced water flow in technical water systems. Removal of a biofilm is very tedious and sometimes even impossible because bacteria in biofilms are resilient towards antibiotic and biocides. It is therefore desirable to be able to prevent biofilm formation, rather than attempting to eliminate biofilms after they form. The formation of a microbial biofilm on a surface can be prevented by creating unfavourable conditions for the reversible, initial attachment of microbial cells. This effect can be obtained by grafting hydrophilic polymeric chains onto surfaces and thereby provide a steric barrier between the substrate surface and the microbial cell. Poly (ethylene glycol) (PEG) is one of the most widely used polymers for making non-adhesive coatings. The work presented in this thesis involves grafting PEG chains onto surfaces using different modifications of the grafting to technique. The main aim of studies presented in this thesis was to develop surfaces which would prevent bacteria from forming biofilm. The work focuses on novel strategies to self assemble PEG thiol monolayers with high graft density. One of the strategies investigated involved backfilling a self assembled layer of 2000 Da PEG thiol with shorter oligo (ethylene glycol) (OEG) thiol molecules to form a mixed monolayer. Detailed quantitative characterization of the backfilling process was carried out using x-ray photoelectron spectroscopy (XPS), contact angle measurements, and atomic force microscopy (AFM). These studies helped in understanding the arrangement of backfilled molecules and the extent of desorption of PEG molecules during the backfilling process. The PEG SAM backfilled with OEG molecules was tested for resistance towards serum adsorption and bacterial attachment, and was found to better resist fouling compared to PEG SAM that had not been backfilled. The second strategy involved PEG grafting using supercritical carbon dioxide (SFC), which is known to possess robust solvation properties. The idea was to demonstrate cloud point type grafting during self assembly using SFC, and produce PEG SAM of high graft density. The SFC based PEG grafting reported here is the first of its kind, and the results demonstrated that SFC is a good solvent for polymer grafting. The fouling properties of such layers were ascertained by quantitative protein adsorption studies and bacterial attachment studies. The detailed surface characterization of grafted polymeric layer and antifouling studies helped in development of novel ways to create antifouling surfaces which have the potential to counter problems caused by biofilms.

  1. Complex formation of vanadium (V) with 4-methyl-1,2,4-three-azole-thiol in 1 mol/l H2SO4

    International Nuclear Information System (INIS)

    The process of complex formation of vanadium (V) with 4-methyl-1,2,4-three-azole-thiol in 1 mol/l H2SO4 at 288-318 K was studied by means of potentiometric titration method. It was found that five complex forms were formed at interaction of vanadium with 4-methyl-1,2,4-three-azole-thiol-5. Stability constants of these complexes and their thermodynamical functions were evaluated. (author)

  2. Dietary sources of aldose reductase inhibitors: prospects for alleviating diabetic complications.

    Science.gov (United States)

    Saraswat, Megha; Muthenna, P; Suryanarayana, P; Petrash, J Mark; Reddy, G Bhanuprakash

    2008-01-01

    Activation of polyol pathway due to increased aldose reductase activity is one of the several mechanisms that have been implicated in the development of various secondary complications of diabetes. Though numerous synthetic aldose reductase inhibitors have been tested, these have not been very successful clinically. Therefore, a number of common plant/ natural products used in Indian culinary have been evaluated for their aldose reductase inhibitory potential in the present study. The aqueous extracts of 22 plant-derived materials were prepared and evaluated for the inhibitory property against rat lens and human recombinant aldose reductase. Specificity of these extracts towards aldose reductase was established by testing their ability to inhibit a closely related enzyme viz, aldehyde reductase. The ex vivo incubation of erythrocytes in high glucose containing medium was used to underscore the significance in terms of prevention of intracellular sorbitol accumulation. Among the 22 dietary sources tested, 10 showed considerable inhibitory potential against both rat lens and human recombinant aldose reductase. Prominent inhibitory property was found in spinach, cumin, fennel, lemon, basil and black pepper with an approximate IC50 of 0.2 mg/mL with an excellent selectivity towards aldose reductase. As against this, 10 to 20 times higher concentrations were required for 50% inhibition of aldehyde reductase. Reduction in the accumulation of intracellular sorbitol by the dietary extracts further substantiated their in vivo efficacy. The findings reported here indicate the scope of adapting life-style modifications in the form of inclusion of certain common sources in the diet for the management of diabetic complications. PMID:19114390

  3. Pyrroline-5-Carboxylate Reductase in Chlorella autotrophica and Chlorella saccharophila in Relation to Osmoregulation.

    Science.gov (United States)

    Lalibert, G; Hellebust, J A

    1989-11-01

    Pyrroline-5-carboxylate (P5C) reductase (EC 1.5.1.2), which catalyzes the reduction of P5C to proline, was partially purified from two Chlorella species; Chlorella autotrophica, a euryhaline marine alga that responds to increases in salinity by accumulating proline and ions, and Chlorella saccharophila, which does not accumulate proline for osmoregulation. From the elution profile of this enzyme from an anion exchange column in Tris-HCl buffer (pH 7.6), containing sorbitol and glycine betaine, it was shown that P5C reductase from C. autotrophica was a neutral protein whereas the enzyme from C. saccharophila was negatively charged. The kinetic mechanisms of the reductase was characteristic of a ping-pong mechanism with double competitive substrate inhibition. Both enzymes showed high specificity for NADH as cofactor. The affinities of the reductases for their substrates did not change when the cells were grown at different salinities. In both algae, the apparent K(m) values of the reductase for P5C and NADH were 0.17 and 0.10 millimolar, respectively. A fourfold increase in maximal velocity of the reductase was observed when C. autotrophica was transferred from 50 to 150% artificial sea water. Even though the reductase was inhibited by NaCl, KCl, and proline, it still showed appreciable activity in the presence of these compounds at molar concentrations. A possible role for the regulation of proline synthesis at the step catalyzed by P5C reductase is discussed in relation to the specificity of P5C reductase for NADH and its responses to salt treatments. PMID:16667157

  4. Pyrroline-5-Carboxylate Reductase in Chlorella autotrophica and Chlorella saccharophila in Relation to Osmoregulation 1

    Science.gov (United States)

    Lalibert, Gilles; Hellebust, Johan A.

    1989-01-01

    Pyrroline-5-carboxylate (P5C) reductase (EC 1.5.1.2), which catalyzes the reduction of P5C to proline, was partially purified from two Chlorella species; Chlorella autotrophica, a euryhaline marine alga that responds to increases in salinity by accumulating proline and ions, and Chlorella saccharophila, which does not accumulate proline for osmoregulation. From the elution profile of this enzyme from an anion exchange column in Tris-HCl buffer (pH 7.6), containing sorbitol and glycine betaine, it was shown that P5C reductase from C. autotrophica was a neutral protein whereas the enzyme from C. saccharophila was negatively charged. The kinetic mechanisms of the reductase was characteristic of a ping-pong mechanism with double competitive substrate inhibition. Both enzymes showed high specificity for NADH as cofactor. The affinities of the reductases for their substrates did not change when the cells were grown at different salinities. In both algae, the apparent Km values of the reductase for P5C and NADH were 0.17 and 0.10 millimolar, respectively. A fourfold increase in maximal velocity of the reductase was observed when C. autotrophica was transferred from 50 to 150% artificial sea water. Even though the reductase was inhibited by NaCl, KCl, and proline, it still showed appreciable activity in the presence of these compounds at molar concentrations. A possible role for the regulation of proline synthesis at the step catalyzed by P5C reductase is discussed in relation to the specificity of P5C reductase for NADH and its responses to salt treatments. PMID:16667157

  5. Phosphorylation state of HMG CoA reductase affects its catalytic activity and degradation.

    Science.gov (United States)

    Parker, R A; Miller, S J; Gibson, D M

    1986-01-01

    The expressed catalytic activity of liver microsomal HMG CoA reductase, the limiting enzyme in cholesterol synthesis, is reversibly diminished by phosphorylation in vitro. In intact hepatocytes the expressed activity of HMG CoA reductase is enhanced by incubation of cells with insulin, and diminished by treatment with glucagon or with mevalonate. In the latter situations the level of total reductase activity falls following initial inactivation (phosphorylation) of the enzyme. This observation suggested that the phosphorylated form of HMG CoA reductase is more sensitive to proteolysis. HMG CoA reductase is a 97,000 dalton (97 K) integral protein of the endoplasmic reticulum with a cytosolic domain that includes the catalytic site and serine residues that may be reversibly phosphorylated. In vitro the Ca2+-activated proteolytic enzyme, calpain, generates two catalytically-active fragments: a membrane bound 62 K and a soluble 53 K form of the enzyme which are quantified by specific immunoblot procedures. Cleavage of the native 97 K HMG CoA reductase is enhanced by pretreatment (inactivation) of microsomes with ATP (Mg2+) and liver reductase kinase compared to microsomes pretreated with protein phosphatase. This is reflected in a loss of 97 K reductase and an increase in the soluble 53 K form of the enzyme. Degradation of HMG CoA reductase in hepatocytes is partially blocked by lysosomotropic agents and insulin. A steady state model for the turnover of proteins subject to reversible phosphorylation has been developed which recognizes fractional degradative rate constants for the phosphorylated and dephosphorylated species. PMID:3028050

  6. Inhibition of glutathione synthesis eliminates the adaptive response of ascitic hepatoma 22 cells to nedaplatin that targets thioredoxin reductase

    International Nuclear Information System (INIS)

    Thioredoxin reductase (TrxR) is a target for cancer therapy and the anticancer mechanism of cisplatin involves TrxR inhibition. We hypothesize that the anticancer drug nedaplatin (NDP), an analogue of cisplatin and a second-generation platinum complex, also targets TrxR. Furthermore, we investigate whether the therapeutic efficacy of NDP can be enhanced by simultaneous modulation of 1) TrxR, via NDP, and 2) glutathione (GSH), via the GSH synthesis inhibitor buthionine sulfoximine (BSO). Mice bearing ascitic hepatoma 22 (H22) cells were treated with NDP alone or NDP plus BSO. TrxR activity of H22 cells was inhibited by NDP in a dose-dependent manner. A high correlation between the inhibition of TrxR activity at 6 h and the inhibition of ascitic fluid volume at 72 h was established (r = 0.978, p < 0.01). As an adaptive response, the viable ascitic cancer cells after NDP treatment displayed an enlarged cell phenotype, assembled with several-fold more antioxidant enzymes and GSH-predominant non-protein free thiols. This adaptive response was largely eliminated when BSO was co-administered with NDP, leading to the decimation of the H22 cell population without enhancing renal toxicity, since at this dose, NDP did not inhibit renal TrxR activity. In conclusion, the pharmacological effect of NDP involves TrxR inhibition, and the adaptive response of NDP-treated ascitic H22 cells can be efficiently counteracted by BSO. Simultaneous modulation of TrxR and GSH on ascitic H22 cells using NDP plus BSO greatly enhances therapeutic efficacy as compared with the single modulation of TrxR using NDP alone. -- Highlights: ? Nedaplatin at a pharmacological dose inhibits TrxR in cancer cells but not in kidney. ? The nedaplatin-treated cancer cells exhibit adaptive response. ? Buthionine sulfoximine inhibits glutathione in both cancer cells and kidney. ? Buthionine sulfoximine counteracts the adaptive response to the nedaplatin treatment. ? Buthionine sulfoximine does not aggravate renal toxicity of the nedaplatin treatment.

  7. Inhibition of glutathione synthesis eliminates the adaptive response of ascitic hepatoma 22 cells to nedaplatin that targets thioredoxin reductase

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yijun [School of Tea and Food Science, Anhui Agricultural University, Hefei 230036, Anhui (China); Lu, Hongjuan [Productivity Center of Jiangsu Province, Nanjing 210042, Jiangsu (China); Wang, Dongxu; Li, Shengrong; Sun, Kang; Wan, Xiaochun [School of Tea and Food Science, Anhui Agricultural University, Hefei 230036, Anhui (China); Taylor, Ethan Will [Department of Nanoscience, Joint School of Nanoscience and Nanoengineering, University of North Carolina at Greensboro, Greensboro, NC 27402 (United States); Zhang, Jinsong, E-mail: zjs@ahau.edu.cn [School of Tea and Food Science, Anhui Agricultural University, Hefei 230036, Anhui (China)

    2012-12-15

    Thioredoxin reductase (TrxR) is a target for cancer therapy and the anticancer mechanism of cisplatin involves TrxR inhibition. We hypothesize that the anticancer drug nedaplatin (NDP), an analogue of cisplatin and a second-generation platinum complex, also targets TrxR. Furthermore, we investigate whether the therapeutic efficacy of NDP can be enhanced by simultaneous modulation of 1) TrxR, via NDP, and 2) glutathione (GSH), via the GSH synthesis inhibitor buthionine sulfoximine (BSO). Mice bearing ascitic hepatoma 22 (H22) cells were treated with NDP alone or NDP plus BSO. TrxR activity of H22 cells was inhibited by NDP in a dose-dependent manner. A high correlation between the inhibition of TrxR activity at 6 h and the inhibition of ascitic fluid volume at 72 h was established (r = 0.978, p < 0.01). As an adaptive response, the viable ascitic cancer cells after NDP treatment displayed an enlarged cell phenotype, assembled with several-fold more antioxidant enzymes and GSH-predominant non-protein free thiols. This adaptive response was largely eliminated when BSO was co-administered with NDP, leading to the decimation of the H22 cell population without enhancing renal toxicity, since at this dose, NDP did not inhibit renal TrxR activity. In conclusion, the pharmacological effect of NDP involves TrxR inhibition, and the adaptive response of NDP-treated ascitic H22 cells can be efficiently counteracted by BSO. Simultaneous modulation of TrxR and GSH on ascitic H22 cells using NDP plus BSO greatly enhances therapeutic efficacy as compared with the single modulation of TrxR using NDP alone. -- Highlights: ? Nedaplatin at a pharmacological dose inhibits TrxR in cancer cells but not in kidney. ? The nedaplatin-treated cancer cells exhibit adaptive response. ? Buthionine sulfoximine inhibits glutathione in both cancer cells and kidney. ? Buthionine sulfoximine counteracts the adaptive response to the nedaplatin treatment. ? Buthionine sulfoximine does not aggravate renal toxicity of the nedaplatin treatment.

  8. Effect of alpha-tocopherol and alpha-tocopheryl quinone on the radiosensitivity of thiol-depleted mammalian cells

    International Nuclear Information System (INIS)

    The effect of hypoxic cell radiosensitizers is increased when mammalian cells are depleted of endogenous glutathione by buthionine sulphoximine pre-treatment in vitro; a similar gain has not been observed in tumors in vivo despite evidence of glutathione depletion in vivo following buthionine sulphoximine treatment. However, concentrations of biological reducing agents other than glutathione were not measured in the in vivo experiments. Other reducing agents found in tumors include alpha-tocopherol, which reduces the sensitizing efficiency of nitro-aromatic sensitizers in thiol-depleted mammalian cells. These data suggest that the failure to observe large gains in misonidazole sensitizing efficiency in thiol-depleted tumors in vivo may be due, in part, to the presence of biological reducing agents such as alpha-tocopherol

  9. Selective and Reversible Binding of Thiol-Functionalized Biomolecules on Polymers Prepared via Chemical Vapor Deposition Polymerization.

    Science.gov (United States)

    Ross, Aftin; Durmaz, Hakan; Cheng, Kenneth; Deng, Xiaopei; Liu, Yuwei; Oh, Jonathan; Chen, Zhan; Lahann, Joerg

    2015-05-12

    We use chemical vapor deposition polymerization to prepare a novel dibromomaleimide-functionalized polymer for selective and reversible binding of thiol-containing biomolecules on a broad range of substrates. We report the synthesis and CVD polymerization of 4-(3,4-dibromomaleimide)[2.2]paracyclophane to yield nanometer thick polymer coatings. Fourier transformed infrared spectroscopy and X-ray photoelectron spectroscopy confirmed the chemical composition of the polymer coating. The reactivity of the polymer coating toward thiol-functionalized molecules was confirmed using fluorescent ligands. As a proof of concept, the binding and subsequent release of cysteine-modified peptides from the polymer coating were also demonstrated via sum frequency generation spectroscopy. This reactive polymer coating provides a flexible surface modification approach to selectively and reversibly bind biomolecules on a broad range of materials, which could open up new opportunities in many biomedical sensing and diagnostic applications where specific binding and release of target analytes are desired. PMID:25869214

  10. X-ray magnetic circular dichroism and small angle neutron scattering studies of thiol capped gold nanoparticles.

    Energy Technology Data Exchange (ETDEWEB)

    de la Venta, J.; Bouzas, V.; Pucci, A.; Laguna-Marco, M. A.; Haskel, D.; te Velthuis, S. G. E; Hoffmann, A.; Lal, J.; Bleuel, M.; Ruggeri, G.; de Julian Fernandez, C.; Garcia, M. A.; Univ.Complutense de Madrid; Inst. de Magnetismo Aplicado; Univ. of Pisa; Lab. di Magnetismo Molecolare

    2009-01-01

    X-ray magnetic circular dichroism (XMCD) and Small Angle Neutron Scattering (SANS) measurements were performed on thiol capped Au nanoparticles (NPs) embedded into polyethylene. An XMCD signal of 0.8 {center_dot} 10{sup -4} was found at the Au L{sub 3} edge of thiol capped Au NPs embedded in a polyethylene matrix for which Superconducting Quantum Interference Device (SQUID) magnetometry yielded a saturation magnetization, M{sub s}, of 0.06 emu/g{sub Au}. SANS measurements showed that the 3.2 nm average-diameter nanoparticles are 28% polydispersed, but no detectable SANS magnetic signal was found with the resolution and sensitivity accessible with the neutron experiment. A comparison with previous experiments carried out on Au NPs and multilayers, yield to different values between XMCD signals and magnetization measured by SQUID magnetometer. We discuss the origin of those differences.

  11. X-ray magnetic circular dichroism and small angle neutron scattering study of thiol capped gold nanoparticles.

    Energy Technology Data Exchange (ETDEWEB)

    de la Venta, J.; Bouzas, V.; Pucci, A.; Laguna-Marco, M. A.; Haskel, D.; Pinel, E. F.; te Velthuis, S. G. E.; Hoffmann, A.; Lal, J.; Bleuel, M.; Ruggeri, G.; de Julian, C.; Garcia, M. A.; Univ. Complutense de Madrid; Inst. de Magnetismo Aplicado UCM; Univ. Pisa; Univ. di Padova

    2009-11-01

    X-ray magnetic circular dichroism (XMCD) and Small Angle Neutron Scattering (SANS) measurements were performed on thiol capped Au nanoparticles (NPs) embedded into polyethylene. An XMCD signal of 0.8 {center_dot} 10{sup -4} was found at the Au L{sub 3} edge of thiol capped Au NPs embedded in a polyethylene matrix for which Superconducting Quantum Interference Device (SQUID) magnetometry yielded a saturation magnetization, M{sub s}, of 0.06 emu/g{sub Au}. SANS measurements showed that the 3.2 nm average-diameter nanoparticles are 28% polydispersed, but no detectable SANS magnetic signal was found with the resolution and sensitivity accessible with the neutron experiment. A comparison with previous experiments carried out on Au NPs and multilayers, yield to different values between XMCD signals and magnetization measured by SQUID magnetometer. We discuss the origin of those differences.

  12. The S(2p) Core Level Binding Energies for Alternative Adsorption Sites and the Example of Thiol Self Assembly

    Science.gov (United States)

    Jia, Juanjuan; Esaulov, Vladimir; Kara, Abdelkader

    2015-03-01

    Results of an investigation of the characteristics of thiol SAMs obtained by vacuum evaporative adsorption, useful for reactive substrates, are presented along with core level binding energy (BE) calculations. Thiol ended SAMs of 1,4-benzenedimethanethiol (BDMT) are obtained by evaporation on Au. They display an unconventional BE structure at about 161 eV, which is close to a known BE of an S atom on Au. S(2p) core level BE calculations for molecules chemisorbed on hollow, bridge and atop sites are reported and suggest that the 161 eV peak is indeed due to an alternative adsorption site, which can be associated to an atop configuration. This must therefore not be confused with atomic sulfur and dissociation processes with S-C bond scission. Work partially supported by the U.S. Department of Energy Basic Energy Science under Contract No DE-FG02-11ER16243.

  13. SYNTHESIS, STRUCTURAL CHARACTERIZATION AND ENZYME INHIBITION STUDIES ON 5-(2-NITROSTYRYL)-1,3,4-OXADIAZOLE-2-THIOL DERIVATIVES

    Scientific Electronic Library Online (English)

    MUHAMMAD ATHAR, ABBASI; ADNAN, AKHTAR; , AZIZ-UR-REHMAN; KHADIJA, NAFEESA; SABAHAT ZAHRA, SIDDIQUI; KHALID MOHAMMED, KHAN; MUHAMMAD, ASHRAF; SYEDA ABIDA, EJAZ.

    2186-21-01

    Full Text Available In the present work, S-substituted derivatives of 5-(2-nitrostyryl)-1,3,4-oxadiazole-2-thiol (4) were synthesized by successive conversions of 3-(2-nitrophenyl) acrylic acid (1) into its respective ester, hydrazide and 1,3,4-oxadiazole. Finally the target compounds were obtained by reaction of 5-(2- [...] nitrostyryl)-1,3,4-oxadiazole-2-thiol (4) with a series of various electrophiles, (5a-I), in NN-dimethyl formamide (DMF) in the presence of sodium hydride (NaH). The structural characterization of these newly synthesized compounds was done by IR, H-NMR, HR-MS and EI-MS spectral data. All these compounds were evaluated for their enzyme inhibitory potentials and found to exhibit broad range spectrum against acetylcholinesterase, butyrylcholinestrase and lipoxygenase enzymes.

  14. Thiol-mediated oxidation of nonphenolic lignin model compounds by manganese peroxidase of Phanerochaete chrysosporium.

    Science.gov (United States)

    Wariishi, H; Valli, K; Renganathan, V; Gold, M H

    1989-08-25

    In the presence of MnII, H2O2, and glutathione (GSH), manganese peroxidase oxidized veratryl alcohol (I) to veratraldehyde (IV). Anisyl alcohol (II) and benzyl alcohol (III) were also oxidized by this system to their corresponding aldehydes (V and VI). In the presence of GSH, chemically prepared MnIII or gamma-irradiation also catalyzed the oxidation of I, II, and III to IV, V, and VI, respectively. GSH and dithiothreitol rapidly reduced MnIII to MnII in the absence of aromatic substrates and the dithiothreitol was oxidized to its disulfide (4,5-dihydroxyl-1,2-dithiane). These results indicate that the thiol is oxidized by enzyme-generated MnIII to a thiyl radical. The latter abstracts a hydrogen from the substrate, forming a benzylic radical which reacts with another thiyl radical to yield an intermediate which decomposes to the benzaldehyde product. In the presence of MnII, GSH, and H2O2, manganese peroxidase also oxidized 1-(4-ethoxy-3-methoxy-phenyl)-2-(4'-hydroxymethyl-2'-methoxyphenoxy)- 1,3-dihydroxypropane (XII) to yield vanillyl alcohol (VII), vanillin (VIII), 1-(4-ethoxy-3-methoxyphenyl)-1,3-dihydroxypropane (XVI), 1-(4-ethoxy-3-methoxyphenyl)-1-oxo-3-hydroxypropane (XIX), and several C alpha oxidized dimeric products. Abstraction of the C alpha (A ring) hydrogen of the dimer (XII) yields a benzylic radical, leading to C beta oxygen ether cleavage. The resultant intermediates yield the ketone (XIX) and vanillyl alcohol (VII) or vanillin (VIII). Alternatively, benzylic radical formation at the C' alpha position (B ring) leads to radical cleavage, yielding a quinone methide and a C beta radical, which yield vanillin and the 1,3-diol (XVI), respectively. In these reactions, MnIII oxidizes a thiol to a thiyl radical which subsequently abstracts a hydrogen from the substrate to form a benzylic radical. The latter undergoes nonenzymatic reactions to yield the final products. PMID:2760063

  15. Electronic Transport through Self Assembled Thiol Molecules: Effect of Monolayer Order, Dynamics and Temperature

    Science.gov (United States)

    Dholakia, Geetha; Fan, Wendy; Meyyappan, M.

    2005-01-01

    We present the charge transport and tunneling conductance of self assembled organic thiol molecules and discuss the influence of order and dynamics in the monolayer on the transport behavior and the effect of temperature. Conjugated thiol molecular wires and organometals such as terpyridine metal complexes provide a new platform for molecular electronic devices and we study their self assembly on Au(111) substrates by the scanning tunneling microscope. Determining the organization of the molecule and the ability to control the nature of its interface with the substrate is important for reliable performance of the molecular electronic devices. By concurrent scanning tunneling microscopy and spectroscopy studies on SAMs formed from oligo (phenelyne ethynelyne) monolayers with and without molecular order, we show that packing and order determine the response of a self assembled monolayer (SAM) to competing interactions. Molecular resolution STM imaging in vacuum shows that the OPES adopt an imcommensurate SAM structure on Au(111) with a rectangular unit cell. Tunneling spectroscopic measurements were performed on the SAM as a function of junction resistance. STS results show that the I-Vs are non linear and asymmetric due to the inherent asymmetry in the molecular structure, with larger currents at negative sample biases. The asymmetry increases with increasing junction resistance due to the asymmetry in the coupling to the leads. This is brought out clearly in the differential conductance, which also shows a gap at the Fermi level. We also studied the effect of order and dynamics in the monolayer on the charge transport and found that competing forces between the electric field, intermolecular interactions, tip-molecule physisorption and substrate-molecule chemisorption impact the transport measurements and its reliability and that the presence of molecular order is very important for reproducible transport measurements. Thus while developing new electronic platforms based on molecules, it is important to have a good control of the molecule-substrate interface, for the devices to perform reliably. While such a control would minimize fluctuations and dynamics in the ensemble, the real challenge is to develop device architectures that are tolerant to fluctuations, since they cannot be totally eliminated in these low dimensional soft systems. Results of temperature dependent STS measurements will also be discussed.

  16. Decaborane thiols as building blocks for self-assembled monolayers on metal surfaces.

    Science.gov (United States)

    Bould, Jonathan; Mach?ek, Jan; Londesborough, Michael G S; Macas, Ramn; Kennedy, John D; Bastl, Zden?k; Rupper, Patrick; Bae, Tom

    2012-02-01

    Three nido-decaborane thiol cluster compounds, [1-(HS)-nido-B(10)H(13)] 1, [2-(HS)-nido-B(10)H(13)] 2, and [1,2-(HS)(2)-nido-B(10)H(12)] 3 have been characterized using NMR spectroscopy, single-crystal X-ray diffraction analysis, and quantum-chemical calculations. In the solid state, 1, 2, and 3 feature weak intermolecular hydrogen bonding between the sulfur atom and the relatively positive bridging hydrogen atoms on the open face of an adjacent cluster. Density functional theory (DFT) calculations show that the value of the interaction energy is approximately proportional to the number of hydrogen atoms involved in the interaction and that these values are consistent with a related bridging-hydrogen atom interaction calculated for a B(18)H(22)C(6)H(6) solvate. Self-assembled monolayers (SAMs) of 1, 2, and 3 on gold and silver surfaces have been prepared and characterized using X-ray photoelectron spectroscopy. The variations in the measured sulfur binding energies, as thiolates on the surface, correlate with the (CC2) calculated atomic charge for the relevant boron vertices and for the associated sulfur substituents for the parent B(10)H(13)(SH) compounds. The calculated charges also correlate with the measured and DFT-calculated thiol (1)H chemical shifts. Wetting-angle measurements indicate that the hydrophilic open face of the cluster is directed upward from the substrate surface, allowing the bridging hydrogen atoms to exhibit a similar reactivity to that of the bulk compound. Thus, [PtMe(2)(PMe(2)Ph)(2)] reacts with the exposed and acidic B-H-B bridging hydrogen atoms of a SAM of 1 on a gold substrate, affording the addition of the metal moiety to the cluster. The XPS-derived stoichiometry is very similar to that for a SAM produced directly from the adsorption of [1-(HS)-7,7-(PMe(2)Ph)(2)-nido-7-PtB(10)H(11)] 4. The use of reactive boron hydride SAMs as templates on which further chemistry may be carried out is unprecedented, and the principle may be extended to other binary boron hydride clusters. PMID:22229807

  17. Feedback regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase in Saccharomyces cerevisiae.

    OpenAIRE

    Dimster-denk, D.; Thorsness, M. K.; Rine, J.

    1994-01-01

    In eukaryotic cells all isoprenoids are synthesized from a common precursor, mevalonate. The formation of mevalonate from 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) is catalyzed by HMG-CoA reductase and is the first committed step in isoprenoid biosynthesis. In mammalian cells, synthesis of HMG-CoA reductase is subject to feedback regulation at multiple molecular levels. We examined the state of feedback regulation of the synthesis of the HMG-CoA reductase isozyme encoded by the yeast ge...

  18. Purification, Characterization, and Overexpression of Flavin Reductase Involved in Dibenzothiophene Desulfurization by Rhodococcus erythropolis D-1

    OpenAIRE

    Matsubara, Toshiyuki; Ohshiro, Takashi; Nishina, Yoshihiro; Izumi, Yoshikazu

    2001-01-01

    The dibenzothiophene (DBT)-desulfurizing bacterium, Rhodococcus erythropolis D-1, removes sulfur from DBT to form 2-hydroxybiphenyl using four enzymes, DszC, DszA, DszB, and flavin reductase. In this study, we purified and characterized the flavin reductase from R. erythropolis D-1 grown in a medium containing DBT as the sole source of sulfur. It is conceivable that the enzyme is essential for two monooxygenase (DszC and DszA) reactions in vivo. The purified flavin reductase contains no chrom...

  19. Thiol-functionalization of metal-organic framework by a facile coordination-based postsynthetic strategy and enhanced removal of Hg2+ from water

    International Nuclear Information System (INIS)

    Highlights: ? A novel type of functionalized MOF for heavy metal removal. ? Functionalization of MOF by a facile coordination-based postsynthetic strategy. ? Thiol-functionalization of MOF has been realized for the first time. ? Enhanced removal of Hg2+ by thiol-functionalized MOFs. - Abstract: The presence of coordinatively unsaturated metal centers in metal-organic frameworks (MOFs) provides an accessible way to selectively functionalize MOFs through coordination bonds. In this work, we describe thiol-functionalization of MOFs by choosing a well known three-dimensional (3D) Cu-based MOF, i.e. [Cu3(BTC)2(H2O)3]n (HKUST-1, BTC = benzene-1,3,5-tricarboxylate), by a facile coordination-based postsynthetic strategy, and demonstrate their application for removal of heavy metal ion from water. A series of [Cu3(BTC)2]n samples stoichiometrically decorated with thiol groups has been prepared through coordination bonding of coordinatively unsaturated metal centers in HKUST-1 with SH group in dithioglycol. The obtained thiol-functionalized samples were characterized by powder X-ray diffraction, scanning electron microscope, energy dispersive X-ray spectroscopy, infrared spectroscopy, and N2 sorptiondesorption isothermal. Significantly, the thiol-functionalized [Cu3(BTC)2]n exhibited remarkably high adsorption affinity (Kd = 4.73 105 mL g?1) and high adsorption capacity (714.29 mg g?1) for Hg2+ adsorption from water, while the unfunctionalized HKUST-1 showed no adsorption of Hg2+ under the same condition.

  20. Speciation and determination of thiols in biological samples using high performance liquid chromatography-inductively coupled plasma-mass spectrometry and high performance liquid chromatography-Orbitrap MS.

    Science.gov (United States)

    Bakirdere, Sezgin; Bramanti, Emilia; D'ulivo, Alessandro; Ataman, O Yavuz; Mester, Zoltan

    2010-11-01

    An analytical method was developed for the determination of thiols in biological samples. Reverse phase chromatography coupled to ICP quadrupole MS or Orbitrap MS was employed for the separation and detection of thiols. For the determination of total thiols, oxidized thiols were reduced using dithiothreitol (DTT). Reduction efficiencies for species of interest were found to be close to 100%. Reduced thiols were derivatized by p-hydroxymercuribenzoate (PHMB) and then separated on a C8 column. Optimization of the extraction, separation and detection steps of the HPLC-ICP-MS and HPLC-Orbitrap MS methods was carried out. Detection limits for cysteine, homocysteine, selenocysteine, glutathione, selenomethionine and cysteinyl-glycine were found to be 18, 34, 39, 12, 128 and 103 fmol, respectively, using HPLC-Orbitrap MS and 730, 1110, 440, 1110 and 580 fmol for cysteine, homocysteine, selenocysteine, glutathione, and cysteinyl-glycine using HPLC-ICP-MS. Contrary to expectation, the LODs and RSDs are higher for the HPLC-ICP-MS instrument, therefore HPLC-Orbitrap MS was used for the determination of thiols in yeast samples. Three different brands of baker's yeast and a selenized yeast were analyzed. The GSH and cysteine levels found in these samples ranged from 4.45 to 17.87 ?mol g(-1) and 0.61 to 1.32 ?mol g(-1), respectively. PMID:20969989

  1. Fabrication of Thiol-Ene "Clickable" Copolymer-Brush Nanostructures on Polymeric Substrates via Extreme Ultraviolet Interference Lithography.

    Science.gov (United States)

    Dbner, Matthias; Gevrek, Tugce N; Sanyal, Amitav; Spencer, Nicholas D; Padeste, Celestino

    2015-06-01

    We demonstrate a new approach to grafting thiol-reactive nanopatterned copolymer-brush structures on polymeric substrates by means of extreme ultraviolet (EUV) interference lithography. The copolymer brushes were designed to contain maleimide functional groups as thiol-reactive centers. Fluoropolymer films were exposed to EUV radiation at the X-ray interference lithography beamline (XIL-II) at the Swiss Light Source, in order to create radical patterns on their surfaces. The radicals served as initiators for the copolymerization of thiol-ene "clickable" brushes, composed of a furan-protected maleimide monomer (FuMaMA) and different methacrylates, namely, methyl methacrylate (MMA), ethylene glycol methyl ether methacrylate (EGMA), or poly(ethylene glycol) methyl ether methacrylate (PEGMA). Copolymerization with ethylene-glycol-containing monomers provides antibiofouling properties to these surfaces. The number of reactive centers on the grafted brush structures can be tailored by varying the monomer ratios in the feed. Grafted copolymers were characterized by using attenuated total reflection infrared (ATR-IR) spectroscopy. The reactive maleimide methacrylate (MaMA) units were utilized to conjugate thiol-containing moieties using the nucleophilic Michael-addition reaction, which proceeds at room temperature without the need for any metal-based catalyst. Using this approach, a variety of functionalities was introduced to yield polyelectrolytes, as well as fluorescent and light-responsive polymer-brush structures. Functionalization of the brush structures was demonstrated via ATR-IR and UV-vis spectroscopy and fluorescence microscopy, and was also indicated by a color switch. Furthermore, grafted surfaces were generated via plasma activation, showing a strongly increased wettability for polyelectrolytes and a reversible switch in static water contact angle (CA) of up to 18 for P(EGMA-co-MaMA-SP) brushes, upon exposure to alternating visible and UV-light irradiation. PMID:25978723

  2. Identification of the Atomic Scale Structures of the Gold-Thiol Interfaces of Molecular Nanowires by Inelastic Tunneling Spectroscopy

    OpenAIRE

    Demir, Firuz; Kirczenow, George

    2012-01-01

    We examine theoretically the effects of the bonding geometries at the gold-thiol interfaces on the inelastic tunneling spectra of propanedithiolate (PDT) molecules bridging gold electrodes and show that inelastic tunneling spectroscopy combined with theory can be used to determine these bonding geometries experimentally. With the help of density functional theory, we calculate the relaxed geometries and vibrational modes of extended molecules each consisting of one or two PD...

  3. Adsorption Kinetics of 4-Amino-5-Phenyl-4H-1, 2, 4-Triazole-3-Thiol on Mild Steel Surface

    OpenAIRE

    Khadom, Anees A.; Musa, Ahmed Y.; Kadhum, Abdul Amir H.; Abu Bakar Mohamad; Mohd Sobri Takriff

    2010-01-01

    The adsorption of 4-amino-5-phenyl-4H-1, 2, 4-triazole-3-thiol (APTT) as a corrosion inhibitor on mild steel surface in hydrochloric acid (HCl) solution was studied using the weight loss technique. The surface coverage with the adsorbed APTT was used to calculate the free energy of adsorption, ?Gads, of APTT using Bockris-Swinkels isotherm. The dependence of free energy of adsorption, ?Gads, on the surface coverage, ?, is ascribed to the surface heterogeneity of the adsorbe...

  4. The preparation and physical properties of polysulfide-based elastomers through one-pot thiol-ene click reaction

    OpenAIRE

    Quan, Y. W.; Zhang, G. Z.; Fan, Z. K.; Chen, Q. M.

    2013-01-01

    In this paper, polysulfide-based elastomers were successfully prepared through a simple one-pot thiol-ene click reaction of the liquid polysulfide oligomer with bisphenol-A diacrylate resin. Real-time Fourier transform infrared spectroscopy (FTIR) analysis showed that the molecular weight of the liquid polysulfide oligomer had no effect on mercaptan functional group conversion. The obtained elastomers continued to keep low temperature flexibility of polysulfide except Elastomer-LP3, which was...

  5. Characterization of Arabidopsis thaliana cDNAs that render yeasts tolerant toward the thiol-oxidizing drug diamide.

    OpenAIRE

    Kushnir, S.; Babiychuk, E.; Kampfenkel, K.; Belles-boix, E.; Montagu, M.; Inze?, D.

    1995-01-01

    Diamide oxidizes cellular thiols and induces oxidative stress. To isolate plant genes which may, when overexpressed, increase tolerance of plants toward oxidative damage, an in vivo diamide tolerance screening in yeasts was used. An Arabidopsis cDNA library in a yeast expression vector was used to transform a yeast strain with intact antioxidant defense. Cells from approximately 10(5) primary transformants were selected for resistance to diamide. Three Arabidopsis cDNAs which confer diamide t...

  6. Missing links in understanding redox signaling via thiol/disulfide modulation: how is glutathione oxidized in plants?

    OpenAIRE

    Rahantaniaina, Marie-sylviane; Tuzet, Andre?e; Mhamdi, Amna; Noctor, Graham

    2013-01-01

    Glutathione is a small redox-active molecule existing in two main stable forms: the thiol (GSH) and the disulphide (GSSG). In plants growing in optimal conditions, the GSH:GSSG ratio is high in most cell compartments. Challenging environmental conditions are known to alter this ratio, notably by inducing the accumulation of GSSG, an effect that may be influential in the perception or transduction of stress signals. Despite the potential importance of glutathione status in redox signaling, the...

  7. Quantifying changes in the cellular thiol-disulfide status during differentiation of B cells into antibody-secreting plasma cells

    DEFF Research Database (Denmark)

    Hansen, Rosa Rebecca Erritze; Otsu, Mieko

    2013-01-01

    Plasma cells produce and secrete massive amounts of disulfide-containing antibodies. To accommodate this load on the secretory machinery, the differentiation of resting B cells into antibody-secreting plasma cells is accompanied by a preferential expansion of the secretory compartments of the cells and by an up-regulation of enzymes involved in redox regulation and protein folding. We have quantified the absolute levels of protein thiols, protein disulfides, and glutathionylated proteins in whole cells. The results show that while the global thiol-disulfide state is affected to some extent by the differentiation, steady-state levels of glutathionylated protein thiols are less than 0.3% of the total protein cysteines, even in fully differentiated cells, and the overall protein redox state is not affected until late in differentiation, when large-scale IgM production is ongoing. A general expansion of the ER does not affect global protein redox status until an extensive production of cargo proteins has started.

  8. Thiol redox chemistry: role of protein cysteine oxidation and altered redox homeostasis in allergic inflammation and asthma.

    Science.gov (United States)

    Hoffman, Sidra; Nolin, James; McMillan, David; Wouters, Emiel; Janssen-Heininger, Yvonne; Reynaert, Niki

    2015-06-01

    Asthma is a pulmonary disorder, with an estimated 300 million people affected worldwide. While it is thought that endogenous reactive oxygen species (ROS) and reactive nitrogen species (RNS) such as hydrogen peroxide and nitric oxide, are important mediators of natural physiological processes, inflammatory cells recruited to the asthmatic airways have an exceptional capacity for producing a variety of highly reactive ROS and RNS believed to contribute to tissue damage and chronic airways inflammation. Antioxidant defense systems form a tightly regulated network that maintains the redox environment of the intra- as well as extracellular environment. Evidence for an oxidant-antioxidant imbalance in asthmatic airways is demonstrated in a number of studies, revealing decreased total antioxidant capacity as well as lower levels of individual antioxidants. Thiols in the form of GSH and sulfhydryl groups of proteins are among the most susceptible oxidant-sensitive targets, and hence, studies investigating protein thiol redox modifications in biology and disease have emerged. This perspective offers an overview of the combined efforts aimed at the elucidation of mechanisms whereby cysteine oxidations contribute to chronic inflammation and asthma, as well as insights into potential cysteine thiol-based therapeutic strategies. PMID:25565397

  9. Thiol-ene click chemistry derived cationic cyclodextrin chiral stationary phase and its enhanced separation performance in liquid chromatography.

    Science.gov (United States)

    Yao, Xiaobin; Tan, Timothy Thatt Yang; Wang, Yong

    2014-01-24

    This work is the first demonstration of a simple thiol-ene click chemistry to anchor vinyl imidazolium ?-CD onto thiol silica to form a novel cationic native cyclodextrin (CD) chiral stationary phase (CSP). The CSP afforded high enantioseparation ability towards dansyl (Dns) amino acids, carboxylic aryl compounds and flavonoids in chiral HPLC. The current CSP demonstrates the highest resolving ability (selectivity >1.1, resolution >1.5) towards Dns amino acids in a mobile phase buffered at pH=6.5, with the resolution of Dns-dl-leucine as high as 6.97. 2,4-dichloride propionic acid (2,4-ClPOPA) was well resolved with the selectivity and resolution of 1.37 and 4.88, respectively. Compared to a previously reported native CD-CSP based on a triazole linkage, the current cationic CD-CSP shows a stronger retention and higher resolution towards acidic chiral compounds, ascribed to the propitious strong electrostatic attraction. Stability evaluation results indicated that thiol-ene reaction can provide a facile and robust approach for the preparation of positively charged CD CSPs. PMID:24411138

  10. Electrogenerated chemiluminescence from thiol-capped CdTe quantum dots and its sensing application in aqueous solution

    International Nuclear Information System (INIS)

    In this paper, the electrogenerated chemiluminescence (ECL) from thiol-capped CdTe quantum dots (QDs) was reported. The ECL emission was occurred at -1.1 V and reached a maximum value at -2.4 V when the potential was cycled between 0.0 and -2.5 V. The reduced species of CdTe QDs could react with the coreactants to produce the ECL emission. The CdTe QD concentration (6.64 x 10-7 mol L-1) of ECL is lower than that (1.0 x 10-3 mol L-1) of chemiluminescence (CL). Based on the enhancement of light emission from thiol-capped CdTe QDs by H2O2 in the negative electrode potential, a novel method for the determination of H2O2 was developed. The light intensity was linearly proportional to the concentration of H2O2 between 2.0 x 10-7 and 1.0 x 10-5 mol L-1 with a detection limit of 6.0 x 10-8 mol L-1. Compared with most of previous reports, the proposed method has higher sensitivity for the determination of H2O2. In addition, the ECL spectrum of thiol-capped CdTe QDs exhibited a peak at around 620 nm, which was substantially red shifted from the photoluminescence (PL) spectrum, suggesting the surface states play an important role in this ECL process

  11. Aptamer-based organic-silica hybrid affinity monolith prepared via "thiol-ene" click reaction for extraction of thrombin.

    Science.gov (United States)

    Wang, Zheng; Zhao, Jin-Cheng; Lian, Hong-Zhen; Chen, Hong-Yuan

    2015-06-01

    A novel strategy for preparing aptamer-based organic-silica hybrid monolithic column was developed via "thiol-ene" click chemistry. Due to the large specific surface area of the hybrid matrix and the simplicity, rapidness and high efficiency of "thiol-ene" click reaction, the average coverage density of aptamer on the organic-silica hybrid monolith reached 420pmol?L(-1). Human ?-thrombin can be captured on the prepared affinity monolithic column with high specificity and eluted by NaClO4 solution. N-p-tosyl-Gly-Pro-Arg p-nitroanilide acetate was used as the sensitive chromogenic substrate of thrombin. The thrombin enriched by this affinity column was detected with a detection of limit of 0.01?M by spectrophotometry. Furthermore, the extraction recovery of thrombin at 0.15?M in human serum was 91.8% with a relative standard deviation of 4.0%. These results indicated that "thiol-ene" click chemistry provided a promising technique to immobilize aptamer on organic-inorganic hybrid monolith and the easily-assembled affinity monolithic material could be used to realize highly selective recognition of trace proteins. PMID:25863371

  12. Eriocalyxin B-induced apoptosis in pancreatic adenocarcinoma cells through thiol-containing antioxidant systems and downstream signalling pathways.

    Science.gov (United States)

    Li, L; Yue, G G L; Pu, J X; Sun, H D; Fung, K P; Leung, P C; Han, Q B; Lau, C B S; Leung, P S

    2014-01-01

    Thiol-containing antioxidant systems play an important role in regulating cellular redox homeostasis. Several anti-cancer agents act by targeting these systems by inducing the production of reactive oxygen species (ROS). Our earlier studies have shown that Eriocalyxin B (EriB), a diterpenoid isolated from Isodon eriocalyx, possesses anti-pancreatic tumour activities in vitro and in vivo. The present study further demonstrated that only thiol-containing antioxidants, N-acetylcysteine (NAC) or dithiothreitol (DTT), inhibited EriB-induced cytotoxicity and apoptosis. EriB suppressed the glutathione and thioredoxin antioxidant systems, thus increasing the intracellular ROS levels and regulating the MAPK, NF?B pathways. Treatment with EriB depleted the intracellular thiol-containing proteins in CAPAN-2 cells. In vivo studies also showed that EriB treatment (2.5 mg/kg) reduced the pancreatic tumour weights significantly in nude mice with increased superoxide levels. Taken together, our results shed important new light on the molecular mechanisms of EriB acting as an apoptogenic agent and its therapeutic potential for pancreatic cancer. PMID:24894173

  13. Influence of indium-tin oxide surface structure on the ordering and coverage of carboxylic acid and thiol monolayers

    International Nuclear Information System (INIS)

    This paper analyses the variability of self-assembled monolayers (SAMs) formation on ITO depending on the substrate surface features. In particular, we report on the formation of carboxylic acid- and thiol-based SAMs on two lots of commercially prepared indium-tin oxide (ITO) thin films. Contact angle measurements, electrochemical experiments, and near-edge x-ray absorption fine structure (NEXAFS) spectroscopy showed that the quality of monolayers formed differed substantially between the two ITO batches. Only one of the two ITO substrates was capable of forming well-organized thiol- and carboxylic acid-based SAMs. In order to rationalize these observations, atomic force microscopy and x-ray diffraction analyses were carried out, and SAMs were prepared on ITO substrates fabricated by sputtering in our laboratories. An attempt was made to influence the film microstructure and surface morphology by varying substrate temperatures during ITO deposition. Good-quality thiol and carboxylic acid SAMs were obtained on one of the ITO substrates prepared in-house. While our characterization could not single out conclusively one specific parameter in ITO surface structure that could be responsible for good SAMs formation, we could point out homogeneous surface morphology as a relevant factor for the quality of the SAMs. Evidence was also found for ITO crystallographic orientation to be a parameter influencing SAMs organization

  14. Synthesis of antibacterial amphiphilic elastomer based on polystyrene-block-polyisoprene-block-polystyrene via thiol-ene addition

    International Nuclear Information System (INIS)

    A new type of amphiphilic antibacterial elastomer has been described. Thermoplastic elastomer, polystyreneblock-polyisopreneblock-polystyrene (PSb-PIb-PS) triblock copolymer was functionalized in toluene solution by free radical mercaptan addition in order to obtain an amphiphilic antibacterial elastomer. Thiol terminated PEG was grafted through the double bonds of PSb-PIb-PS via free radical thiol-ene coupling reaction. The antibacterial properties of the amphiphilic graft copolymers were observed. The original and the modified polymers were used to create microfibers in an electro-spinning process. Topology of the electrospun micro/nanofibers were studied by using scanning electron microscopy (SEM). The chemical structures of the amphiphilic comb type graft copolymers were elucidated by the combination of elemental analysis, 1H NMR, 13C NMR, GPC and FTIR. - Graphical abstract: Double bonds of polyisoprene units in polystyreneblock-polyisopreneblock-polystyrene triblock copolymer were partially capped with PEG containing mercapto end group via thiol-ene addition in order to obtain antibacterial amphiphilic elastomer. Nano fibers from amphiphilic graft polymers solution were produced by electrospinning. The PEG grafted copolymer inhibits very effectively bacterial growth. Highlights: ? A commercial synthetic elastomer was grafted with PEG to obtain amphiphilic elastomer. ? Amphiphilic elastomer shows antibacterial properties. ? Electrospun micro fibers of the amphiphilic elastomer tend to globular formation

  15. Pulse radiolysis studies on superoxide reductase from Treponema pallidum

    CERN Document Server

    Nivire, V; Fontecave, M; Houe-Levin, C

    2015-01-01

    Superoxide reductases (SORs) are small metalloenzymes, which catalyze reduction of O2*- to H2O2. The reaction of the enzyme from Treponema pallidum with superoxide was studied by pulse radiolysis methods. The first step is an extremely fast bi-molecular reaction of the ferrous center with O2, with a rate constant of 6 x 10 (8) M(-1) s(-1). A first intermediate is formed which is converted to a second one with a slower rate constant of 4800 s(-1). This latter value is 10 times higher than the corresponding one previously reported in the case of SOR from Desulfoarculus baarsii. The reconstituted spectra for the two intermediates are consistent with formation of transient iron-peroxide species.

  16. Methylenetetrahydrofolate reductase polymorphisms in myeloid leukemia patients from Northeastern Brazil

    Scientific Electronic Library Online (English)

    Cynara Gomes, Barbosa; Claudio Lima, Souza; Jos Pereira de, Moura Neto; Maria da Glria Bomfim, Arruda; Jos Henrique, Barreto; Mitermayer Galvo, Reis; Marilda Souza, Goncalves.

    Full Text Available Methylenetetrahydrofolate reductase (MTHFR: EC 1.5.1.20) polymorphisms are associated to acute lymphoid leukemia in different populations. We used the polymerase chain reaction and the restriction fragment length polymorphism method (PCR-RFLP) to investigate MTHFR C677T and A1298C polymorphism frequ [...] encies in 67 patients with chronic myeloid leukemia (CML), 27 with acute myeloid leukemia FAB subtype M3 (AML-M3) and 100 apparently healthy controls. The MTHFR mutant allele frequencies were as follows: CML = 17.2% for C677T, 21.6% for A1298C; AML-M3 = 22.2% for C677T, 24.1% for A1298C; and controls = 20.5% for C677T, 21% for A1298C. Taken together, our results provide evidence that MTHFR polymorphisms have no influence on the development of CML or AML-M3.

  17. Biofilm Modifies Expression of Ribonucleotide Reductase Genes in Escherichia coli

    Science.gov (United States)

    Cendra, Maria del Mar; Jurez, Antonio; Torrents, Eduard

    2012-01-01

    Ribonucleotide reductase (RNR) is an essential enzyme for all living organisms since is the responsible for the last step in the synthesis of the four deoxyribonucleotides (dNTPs) necessary for DNA replication and repair. In this work, we have investigated the expression of the three-RNR classes (Ia, Ib and III) during Escherichia coli biofilm formation. We show the temporal and spatial importance of class Ib and III RNRs during this process in two different E. coli wild-type strains, the commensal MG1655 and the enteropathogenic and virulent E2348/69, the prototype for the enteropathogenic E. coli (EPEC). We have established that class Ib RNR, so far considered cryptic, play and important role during biofilm formation. The implication of this RNR class under the specific growth conditions of biofilm formation is discussed. PMID:23050019

  18. Arylfurans as potential Trypanosoma cruzi trypanothione reductase inhibitors

    Directory of Open Access Journals (Sweden)

    Renata B de Oliveira

    2006-03-01

    Full Text Available The natural lignans veraguensin and grandisin have been reported to be active against Trypanosoma cruzi bloodstream forms. Aiming at the total synthesis of these and related compounds, we prepared three 2-arylfurans and eight 2,5-diarylfurans. They were evaluated for their potential as T. cruzi trypanothione reductase (TR inhibitors as well against the parasite's intracellular (amastigote and bloodstream (trypomastigote forms. Compound 12 was the most effective against TR with an IC50 of 48.5 M while 7 and 14 were active against amastigotes, inhibiting the parasite development by 60% at 20 g/ml (59 and 90 M, respectively. On the other hand, none of the compounds was significantly active against the parasite bloodstream forms even at 250 g/ml (0.6-1.5 mM.

  19. Arylfurans as potential Trypanosoma cruzi trypanothione reductase inhibitors

    Scientific Electronic Library Online (English)

    Renata B de, Oliveira; Aline BM, Vaz; Rosana O, Alves; Daniel B, Liarte; Claudio L, Donnici; Alvaro J, Romanha; Carlos L, Zani.

    2006-03-01

    Full Text Available The natural lignans veraguensin and grandisin have been reported to be active against Trypanosoma cruzi bloodstream forms. Aiming at the total synthesis of these and related compounds, we prepared three 2-arylfurans and eight 2,5-diarylfurans. They were evaluated for their potential as T. cruzi tryp [...] anothione reductase (TR) inhibitors as well against the parasite's intracellular (amastigote) and bloodstream (trypomastigote) forms. Compound 12 was the most effective against TR with an IC50 of 48.5 M while 7 and 14 were active against amastigotes, inhibiting the parasite development by 60% at 20 g/ml (59 and 90 M, respectively). On the other hand, none of the compounds was significantly active against the parasite bloodstream forms even at 250 g/ml (0.6-1.5 mM).

  20. Mechanism of inhibition of ribonucleotide reductase with motexafin gadolinium (MGd)

    International Nuclear Information System (INIS)

    Motexafin gadolinium (MGd) is an expanded porphyrin anticancer agent which selectively targets tumor cells and works as a radiation enhancer, with promising results in clinical trials. Its mechanism of action is oxidation of intracellular reducing molecules and acting as a direct inhibitor of mammalian ribonucleotide reductase (RNR). This paper focuses on the mechanism of inhibition of RNR by MGd. Our experimental data present at least two pathways for inhibition of RNR; one precluding subunits oligomerization and the other direct inhibition of the large catalytic subunit of the enzyme. Co-localization of MGd and RNR in the cytoplasm particularly in the S-phase may account for its inhibitory properties. These data can elucidate an important effect of MGd on the cancer cells with overproduction of RNR and its efficacy as an anticancer agent and not only as a general radiosensitizer.

  1. Inhibition of rat lens aldose reductase by quercetagetin and patuletin.

    Science.gov (United States)

    Li, S; Mao, W; Cao, X; Liang, S; Ding, Z; Li, N

    1991-03-01

    In this paper the results of inhibition of the Aldose reductase (AR) activity on Wistar rat lens by Quercetagetin extracted from Tagetes erects Linn and by Patuletin extracted from Tagetes patula Linn are reported. Quercetagetin inhibited AR of the rat lens by 93.9% at 10(-4) M, 76.0% at 10(-5) M and 13.3% at 10(-6) M. Patuletin inhibited AR of the rat lens by 100% at 10(-1) M, 80% at 10(-5) M and 22.7% at 10(-6) M respectively. The results show that these two flavones are lens AR Inhibitors, but further study should be done for the possibility of clinical application. PMID:1843126

  2. Regulation of ribonucleotide reductase by Spd1 involves multiple mechanisms

    DEFF Research Database (Denmark)

    Nestoras, Konstantinos; Mohammed, Asma Hadi

    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 by small inhibitory proteins that associate with the R1 catalytic subunit. In addition, the subcellular localization of the R2 subunit is regulated through the cell cycle and in response to DNA damage. We show that the fission yeast small RNR inhibitor Spd1 is intrinsically disordered and regulates R2 nuclear import, as predicted by its relationship to Saccharomyces cerevisiae Dif1. We demonstrate that Spd1 can interact with both R1 and R2, and show that the major restraint of RNR in vivo by Spd1 is unrelated to R2 subcellular localization. Finally, we identify a new behavior for RNR complexes that potentially provides yet another mechanism to regulate dNTP synthesis via modulation of RNR complex architecture.

  3. Conversion of Pipecolic Acid into Lysine in Penicillium chrysogenum Requires Pipecolate Oxidase and Saccharopine Reductase: Characterization of the lys7 Gene Encoding Saccharopine Reductase

    Science.gov (United States)

    Naranjo, Leopoldo; de Valmaseda, Eva Martin; Bauelos, Oscar; Lopez, Pilar; Riao, Jorge; Casqueiro, Javier; Martin, Juan F.

    2001-01-01

    Pipecolic acid is a component of several secondary metabolites in plants and fungi. This compound is useful as a precursor of nonribosomal peptides with novel pharmacological activities. In Penicillium chrysogenum pipecolic acid is converted into lysine and complements the lysine requirement of three different lysine auxotrophs with mutations in the lys1, lys2, or lys3 genes allowing a slow growth of these auxotrophs. We have isolated two P. chrysogenum mutants, named 7.2 and 10.25, that are unable to convert pipecolic acid into lysine. These mutants lacked, respectively, the pipecolate oxidase that converts pipecolic acid into piperideine-6-carboxylic acid and the saccharopine reductase that catalyzes the transformation of piperideine-6-carboxylic acid into saccharopine. The 10.25 mutant was unable to grow in Czapek medium supplemented with ?-aminoadipic acid. A DNA fragment complementing the 10.25 mutation has been cloned; sequence analysis of the cloned gene (named lys7) revealed that it encoded a protein with high similarity to the saccharopine reductase from Neurospora crassa, Magnaporthe grisea, Saccharomyces cerevisiae, and Schizosaccharomyces pombe. Complementation of the 10.25 mutant with the cloned gene restored saccharopine reductase activity, confirming that lys7 encodes a functional saccharopine reductase. Our data suggest that in P. chrysogenum the conversion of pipecolic acid into lysine proceeds through the transformation of pipecolic acid into piperideine-6-carboxylic acid, saccharopine, and lysine by the consecutive action of pipecolate oxidase, saccharopine reductase, and saccharopine dehydrogenase. PMID:11717275

  4. Molecular modeling and site-directed mutagenesis reveal the benzylisoquinoline binding site of the short-chain dehydrogenase/reductase salutaridine reductase.

    Science.gov (United States)

    Geissler, Ren; Brandt, Wolfgang; Ziegler, Jrg

    2007-04-01

    Recently, the NADPH-dependent short-chain dehydrogenase/reductase (SDR) salutaridine reductase (E.C. 1.1.1.248) implicated in morphine biosynthesis was cloned from Papaver somniferum. In this report, a homology model of the Papaver bracteatum homolog was created based on the x-ray structure of human carbonyl reductase 1. The model shows the typical alpha/beta-folding pattern of SDRs, including the four additional helices alphaF'-1 to alphaF'-4 assumed to prevent the dimerization of the monomeric short-chain dehydrogenases/reductases. Site-directed mutagenesis of asparagine-152, serine-180, tyrosine-236, and lysine-240 resulted in enzyme variants with strongly reduced performance or inactive enzymes, showing the involvement of these residues in the proton transfer system for the reduction of salutaridine. The strong preference for NADPH over NADH could be abolished by replacement of arginine residues 44 and 48 by glutamic acid, confirming the interaction between the arginines and the 2'-phosphate group. Docking of salutaridine into the active site revealed nine amino acids presumably responsible for the high substrate specificity of salutaridine reductase. Some of these residues are arranged in the right position by an additional alphaE' helix, which is not present in SDRs analyzed so far. Enzyme kinetic data from mutagenic replacement emphasize the critical role of these residues in salutaridine binding and provide the first data on the molecular interaction of benzylisoquinoline alkaloids with enzymes. PMID:17337529

  5. Lactococcus lactis Thioredoxin Reductase Is Sensitive to Light Inactivation

    DEFF Research Database (Denmark)

    Bjrnberg, Olof; Viennet, Thibault

    2015-01-01

    Thioredoxin, involved in numerous redox pathways, is maintained in the dithiol state by the nicotinamide adenine dinucleotide phosphate-dependent flavoprotein thioredoxin reductase (TrxR). Here, TrxR from Lactococcus lactis is compared with the well-characterized TrxR from Escherichia coli. The two enzymes belong to the same class of low-molecular weight thioredoxin reductases and display similar kcat values (?25 s-1) with their cognate thioredoxin. Remarkably, however, the L. lactis enzyme is inactivated by visible light and furthermore reduces molecular oxygen 10 times faster than E. coli TrxR. The rate of light inactivation under standardized conditions (?max = 460 nm and 4 C) was reduced at lowered oxygen concentrations and in the presence of iodide. Inactivation was accompanied by a distinct spectral shift of the flavin adenine dinucleotide (FAD) that remained firmly bound. High-resolution mass spectrometric analysis of heat-extracted FAD from light-damaged TrxR revealed a mass increment of 13.979 Da, relative to that of unmodified FAD, corresponding to the addition of one oxygen atom and the loss of two hydrogen atoms. Tandem mass spectrometry confined the increase in mass of the isoalloxazine ring, and the extracted modified cofactor reacted with dinitrophenyl hydrazine, indicating the presence of an aldehyde. We hypothesize that a methyl group of FAD is oxidized to a formyl group. The significance of this not previously reported oxidation and the exceptionally high rate of oxygen reduction are discussed in relation to other flavin modifications and the possible occurrence of enzymes with similar properties.

  6. Evidence for a hexaheteromeric methylenetetrahydrofolate reductase in Moorella thermoacetica.

    Science.gov (United States)

    Mock, Johanna; Wang, Shuning; Huang, Haiyan; Kahnt, Jrg; Thauer, Rudolf K

    2014-09-01

    Moorella thermoacetica can grow with H? and CO?, forming acetic acid from 2 CO? via the Wood-Ljungdahl pathway. All enzymes involved in this pathway have been characterized to date, except for methylenetetrahydrofolate reductase (MetF). We report here that the M. thermoacetica gene that putatively encodes this enzyme, metF, is part of a transcription unit also containing the genes hdrCBA, mvhD, and metV. MetF copurified with the other five proteins encoded in the unit in a hexaheteromeric complex with an apparent molecular mass in the 320-kDa range. The 40-fold-enriched preparation contained per mg protein 3.1 nmol flavin adenine dinucleotide (FAD), 3.4 nmol flavin mononucleotide (FMN), and 110 nmol iron, almost as predicted from the primary structure of the six subunits. It catalyzed the reduction of methylenetetrahydrofolate with reduced benzyl viologen but not with NAD(P)H in either the absence or presence of oxidized ferredoxin. It also catalyzed the reversible reduction of benzyl viologen with NADH (diaphorase activity). Heterologous expression of the metF gene in Escherichia coli revealed that the subunit MetF contains one FMN rather than FAD. MetF exhibited 70-fold-higher methylenetetrahydrofolate reductase activity with benzyl viologen when produced together with MetV, which in part shows sequence similarity to MetF. Heterologously produced HdrA contained 2 FADs and had NAD-specific diaphorase activity. Our results suggested that the physiological electron donor for methylenetetrahydrofolate reduction in M. thermoacetica is NADH and that the exergonic reduction of methylenetetrahydrofolate with NADH is coupled via flavin-based electron bifurcation with the endergonic reduction of an electron acceptor, whose identity remains unknown. PMID:25002540

  7. Fatty acyl-CoA reductases of birds

    Directory of Open Access Journals (Sweden)

    Hellenbrand Janine

    2011-12-01

    Full Text Available Abstract Background Birds clean and lubricate their feathers with waxes that are produced in the uropygial gland, a holocrine gland located on their back above the tail. The type and the composition of the secreted wax esters are dependent on the bird species, for instance the wax ester secretion of goose contains branched-chain fatty acids and unbranched fatty alcohols, whereas that of barn owl contains fatty acids and alcohols both of which are branched. Alcohol-forming fatty acyl-CoA reductases (FAR catalyze the reduction of activated acyl groups to fatty alcohols that can be esterified with acyl-CoA thioesters forming wax esters. Results cDNA sequences encoding fatty acyl-CoA reductases were cloned from the uropygial glands of barn owl (Tyto alba, domestic chicken (Gallus gallus domesticus and domestic goose (Anser anser domesticus. Heterologous expression in Saccharomyces cerevisiae showed that they encode membrane associated enzymes which catalyze a NADPH dependent reduction of acyl-CoA thioesters to fatty alcohols. By feeding studies of transgenic yeast cultures and in vitro enzyme assays with membrane fractions of transgenic yeast cells two groups of isozymes with different properties were identified, termed FAR1 and FAR2. The FAR1 group mainly synthesized 1-hexadecanol and accepted substrates in the range between 14 and 18 carbon atoms, whereas the FAR2 group preferred stearoyl-CoA and accepted substrates between 16 and 20 carbon atoms. Expression studies with tissues of domestic chicken indicated that FAR transcripts were not restricted to the uropygial gland. Conclusion The data of our study suggest that the identified and characterized avian FAR isozymes, FAR1 and FAR2, can be involved in wax ester biosynthesis and in other pathways like ether lipid synthesis.

  8. Structural modifications of gold thin films produced by thiol-derivatized single-stranded DNA immobilization

    International Nuclear Information System (INIS)

    Recent experiments have reported an opposite sign of the differential surface stress produced on gold-coated cantilevers by a thiol-derivatized single-stranded DNA (SH-DNA) immobilization process. The sign of the surface stress depends on the method used to evaporate the gold thin film, being compressive (negative) or tensile (positive) for e-beam or resistively deposited gold, respectively. This study investigates the origin of this effect by means of a combination of x-ray diffraction and x-ray photoelectron spectroscopy. Both e-beam and resistively grown gold thin films are characterized to find the subtle differences responsible for this intriguing stress behaviour. Somewhat remarkably, these studies show a tight relation between the surface structure of the gold overlayer and the SH-DNA immobilization efficiency. The average grain size variation seems to correlate well with the differential surface stress triggered by the SH-DNA immobilization previously reported. These results suggest that the relation of the probe molecules with the surface structure must be considered to understand surface stress changes. (paper)

  9. Photolithographic fabrication of solidliquid core waveguides by thiol-ene chemistry

    DEFF Research Database (Denmark)

    Sagar, Kaushal Shashikant; Gopalakrishnan, Nimi

    2011-01-01

    In this work we demonstrate an efficient and cleanroom compatible method for the fabrication of solidliquid core waveguides based on nanoporous polymers. We have used thiol-ene photo-grafting to tune and pattern the hydrophilicity of an originally hydrophobic nanoporous 1, 2-polybutadiene. The generated refractive index contrast between the patterned water-filled volume and the surrounding empty hydrophobic porous polymer allows for light confinement within the water-filled volumethe solidliquid core. The presented fabrication process is simple and fast. It allows a high degree of flexibility on the type and grade of surface chemistry imparted to the large nanoporous area depending upon the application. The fabrication does not need demanding chemical reaction conditions. Thus, it can be readily used on a standard silicon lithography bench. The propagation loss values reported in this work are comparable with literature values for state-of-the-art liquid-core waveguide devices. The demonstrated waveguide function added to the nanoporous polymer with a very high internal surface area makes the system interesting for many applications in different areas, such as diagnostics and bio-chemical sensing.

  10. Effect of cysteamine on redox-sensitive thiol-containing proteins in the duodenal mucosa.

    Science.gov (United States)

    Khomenko, Tetyana; Deng, Xiaoming; Jadus, Martin R; Szabo, Sandor

    2003-10-01

    Recent studies from our laboratory demonstrated that Egr-1 is upregulated in the rat duodenal mucosa during cysteamine-induced duodenal ulceration and that antisense egr-1 oligonucleotide aggravates the duodenal ulcers. This study was aimed to determine the effects of cysteamine on redox-sensitive Egr-1 transcriptional activity and on other thiol-containing proteins such as redox factor-1 (Ref-1) and thioredoxin (Trx). Here we demonstrate for the first time that cysteamine increases the expression and nuclear translocation of Egr-1, Ref-1, and Trx, and activates binding of Egr-1 to DNA. Moreover, we also show that Egr-1 forms a complex with other redox-sensitive transcription factors (e.g., AP-1, AP-2, NFATc, Sp1, PAX-5, MTF-1, c-Myb, and CREB) in rat duodenal mucosa and that cysteamine enhances the formation of these complexes. The antioxidant ebselen markedly elevated the nuclear Ref-1 expression and Egr-1/DNA binding, and decreased the ulcerogenic effect of cysteamine as did catalase. Thus, redox-sensitive signaling systems seem to play an important role in cysteamine-induced duodenal ulceration. PMID:13679060

  11. Purification and partial characterization of a thiol proteinase activating prokallikrein from the rat kidney cortex.

    Science.gov (United States)

    Nishii, M; Takaoka, M; Nakamura, M; Takenobu, Y; Morimoto, S

    1989-02-24

    The rat kidney cortex contains at least three kinds of prokallikrein-activating proteinase, and among these the one with the highest molecular weight was purified by a procedure including chromatography on CM-cellulose, concanavalin A-Sepharose, organomercurial-Sepharose 4B and Sephadex G-100. The resulting preparation was apparently homogeneous, as assessed by SDS-polyacrylamide gel electrophoresis. The molecular weight was estimated to be 57,000. The optimal pH for the activation of prokallikrein by the preparation, termed activator I, was around 4.5. Activator I was inhibited by E-64, iodoacetate and leupeptin, but not by PMSF and phosphoramidon. In immunodiffusion analysis, the antiserum to activator I formed an immunoprecipitin arc with the extract from the kidney cortex or submandibular gland, but not with that from the pancreas. These results indicate that activator I is a thiol proteinase with a molecular weight of 57,000. A proteinase immunologically identical with activator I appears to be present in the submandibular gland. PMID:2492829

  12. Biochemical Characterization of a Thiol-Activated, Oxidation Stable Keratinase from Bacillus pumilus KS12.

    Science.gov (United States)

    Rajput, Rinky; Sharma, Richa; Gupta, Rani

    2010-01-01

    An extracellular keratinase from Bacillus pumilus KS12 was purified by DEAE ion exchange chromatography. It was a 45?kDa monomer as determined by SDS PAGE analysis. It was found to be an alkaline, serine protease with pH and temperature optima of 10 and 60C, respectively. It was thiol activated with two- and eight-fold enhancement in presence of 10 mM DTT and ?-mercaptoethanol, respectively. In addition, its activity was stimulated in the presence of various surfactants, detergents, and oxidizing agents where a nearly 2- to 3-fold enhancement was observed in presence of H(2)O(2) and NaHClO(3). It hydrolyzed broad range of complex substrates including feather keratin, haemoglobin, fibrin, casein,and ?-keratin. Analysis of amidolytic activity revealed that it efficiently cleaved phenylalanine ? leucine ? alanine- p-nitroanilides. It also cleaved insulin B chain between Val(2)- Asn(3), Leu(6)-Cys(7) and His(10)-Leu(11) residues. PMID:21048858

  13. Biochemical Characterization of a Thiol-Activated, Oxidation Stable Keratinase from Bacillus pumilus KS12

    Directory of Open Access Journals (Sweden)

    Rinky Rajput

    2010-01-01

    Full Text Available An extracellular keratinase from Bacillus pumilus KS12 was purified by DEAE ion exchange chromatography. It was a 45?kDa monomer as determined by SDS PAGE analysis. It was found to be an alkaline, serine protease with pH and temperature optima of 10 and 60C?, respectively. It was thiol activated with two- and eight-fold enhancement in presence of 10 mM DTT and ?-mercaptoethanol, respectively. In addition, its activity was stimulated in the presence of various surfactants, detergents, and oxidizing agents where a nearly 2- to 3-fold enhancement was observed in presence of H2O2 and NaHClO3. It hydrolyzed broad range of complex substrates including feather keratin, haemoglobin, fibrin, casein,and ?-keratin. Analysis of amidolytic activity revealed that it efficiently cleaved phenylalanine ? leucine ? alanine- p-nitroanilides. It also cleaved insulin B chain between Val2- Asn3, Leu6-Cys7 and His10-Leu11 residues.

  14. The performance of thiol-terminated PEG-paclitaxel-conjugated gold nanoparticles.

    Science.gov (United States)

    Ding, Ya; Zhou, Ying-Ying; Chen, Huan; Geng, Dong-Dong; Wu, Dong-Yan; Hong, Jin; Shen, Wen-Bin; Hang, Tai-Jun; Zhang, Can

    2013-12-01

    A series of thiol-terminated polyethylene glycol (PEG)-paclitaxel (PTX) derivatives are designed and synthesized to fabricate PTX-conjugated gold nanoparticles (PTX@GNPs) and improve their overall performance. By extending the molecular weight of PEG from 400 to 1000 Da, the optimized water solubility of the conjugate reaches 184 mg/mL, equal to 4.6 10(5) times that of PTX alone (0.4 ?g/mL). High drug loading is obtained by eliminating the steric hindrance between PTX molecules on the surface of GNPs. The gold conjugate shows double simultaneous stimulation-induced drug release behavior in the presence of both esterase and high concentrations of glutathione. The synergic release characteristics of this conjugate results in significant performance improvements, including prolonged circulation due to high stability in vivo, targeted release of PTX inside tumor cells, and increased tumor cell killing efficiency. Improving the in vitro properties of the conjugate not only significantly enhances its therapeutic efficacy in a murine liver cancer model, but also allows drug-conjugated gold nanoparticles to be used as a promising nanoprodrug system in the cancer therapeutics. PMID:24055524

  15. Radio- and thermosensitivity of E. coli K1060 after thiol depletion by diethylmaleate

    International Nuclear Information System (INIS)

    The Escherichia coil auxotroph K 1060 has been grown in a medium supplemented with either oleic acid (18:1) or linolenic acid (18:3) and its radiosensitivity and thermosensitivity established using bacterial cell survival as the assay system. No difference in radiosensitivity was observed when oleic and linolenic grown cells were exposed to ?-radiation at room temperature. When heated at 490C linolenic grown cells were more sensitive than oleic grown cells. To investigate whether soluble -SH compounds, e.g., glutathione (GSH), were critical in protecting cells against radiation or heat, studies were performed using cell depleted of -SH by incubation with diethylmaleate (DEM). After reduction of water-soluble non-protein thiol compounds to 25% (10 mM DEM treatment) of control value, no major changes in radiosensitivity under oxic conditions were found. Radioresistance increased slightly when irradiation was performed under hypoxic conditions. Thermoresistance was clearly stimulated after DEM treatments between 1 and 10 mM DEM. The main conclusion of these experiments is that lowering the cellular level of reduced glutathione may not generally be correlated with a higher radio- and thermosensitivity. (orig.)

  16. Modification of VTMS hybrid monolith via thiol-ene click chemistry for capillary electrochromatography.

    Science.gov (United States)

    Wang, Keyi; Chen, Yingzhuang; Yang, Huihui; Li, Yi; Nie, Lihua; Yao, Shouzhuo

    2012-03-15

    An n-octadecanethiol (C(18))/3-mercapto-1-propane-sulfonate (MPS) modified organic-inorganic hybrid silica monolithic column possessing vinyl ligands through thiol-ene click chemistry for capillary electrochromatography (CEC) is described. The proposed column is prepared via the sol-gel process by in situ co-condensation using vinyltrimethoxysilane (VTMS) and tetra-methoxysilane (TMOS) as precursors. Examination by SEM shows that the capillary has homogenous macroporous morphology and is well attached to the inner wall of the capillary. The obtained C(18)-MPS-VTMS silica hybrid monolithic column demonstrated an enhanced hydrophilic property and could be applied as a reversed-phase stationary phase in CEC directly. Compared with unmodified VTMS silica hybrid monolithic column, stronger EOF was observed using this monolithic column. VTMS/TOMS ratios in the reaction mixture were varied and 1:3 was found to be optimum. Good separations of benzenes, aromatic amines, acids and peptides were achieved, the lowest plate height of ? 3?m was obtained, the peak symmetry range from 0.98 to 1.29. The resulting C(18)-MPS-VTMS silica hybrid monolithic column can be used in different separation modes, including reversed phase mode and ion exchange mode. PMID:22365679

  17. Simple thiol-ene click chemistry modification of SBA-15 silica pores with carboxylic acids.

    Science.gov (United States)

    Bordoni, Andrea V; Lombardo, M Vernica; Regazzoni, Alberto E; Soler-Illia, Galo J A A; Wolosiuk, Alejandro

    2015-07-15

    A straightforward approach for anchoring tailored carboxylic groups in mesoporous SiO2 colloidal materials is presented. The thiol-ene photochemical reaction between vinyltrimethoxysilane precursors and various thiocarboxylic acids which has, click chemistry features (i.e. high conversion yields, insensitivity to oxygen, mild reaction conditions), results in carboxylated silane precursors that can be readily used as surface modifiers. The carboxylic groups of acetic, undecanoic and succinic acid were immobilized on the silica mesopore walls of SBA-15 powders employing the synthesized silane precursors. Post-grafting has been confirmed through infrared spectrometry (FTIR), energy dispersive X-ray spectroscopy (EDS), elemental analysis (EA) and zeta potential measurements. Detailed field-emission gun scanning electron microscopy (FESEM) images and small angle X-ray scattering (SAXS) data revealed parallel mesopores and ordered mesostructures. It is shown that the immobilized COOH groups are chemically accessible for acid-base reactions as well as copper adsorption. Immobilization of easily synthesized tailored carboxylic modified alkoxide precursors within mesoporous systems provides a unique chemical nanoenvironment within these ordered frameworks. PMID:25845883

  18. Characterization of self-assembled thiols monolayers on gold surface by electrochemical impedance spectroscopy

    Scientific Electronic Library Online (English)

    Renata K., Mendes; Renato S., Freire; Carla P., Fonseca; Silmara, Neves; Lauro T., Kubota.

    2004-12-01

    Full Text Available Tiis de cadeias carbnicas com tamanhos diferentes e contendo grupo terminal COOH foram usados para formar monocamadas auto-organizadas sobre a superfcie de eletrodos de ouro. A transferncia de eltrons do par Fe(CN)6(3-/4-) para o eletrodo foi estudada em diferentes pH usando-se as tcnicas de v [...] oltametria cclica (CV) e de espectroscopia de impedncia eletroqumica (EIS). Mudanas no pH da soluo resultaram em variaes na carga do grupo terminal das monocamadas auto-organizadas e, conseqentemente, em alteraes na interao eletrosttica da SAM com as espcies eletroativas em soluo. Abstract in english Thiols with different alkyl chain length and containing COOH terminal group were self-assembled on gold electrodes. The electron transfer of Fe(CN)6(3-/4-) couple to the electrode was studied at different pH by means cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). Changes i [...] n solution pH resulted in the charge variation of the self-assembled monolayer (SAM) terminal group and, consequently, the electrostatic interaction of SAM with the electroactive species in the solution.

  19. Investigating thiol-modification on hyaluronan via carbodiimide chemistry using response surface methodology.

    Science.gov (United States)

    Santhanam, Sruthi; Liang, Jue; Baid, Rinku; Ravi, Nathan

    2015-07-01

    Hyaluronan (HA) is a naturally occurring glycosaminoglycan widely researched for its use as a biomaterial in tissue engineering, drug delivery, angiogenesis, and ophthalmic surgeries. The mechanical properties of this biomaterial can be altered to a required extent by chemically modifying the pendant reactive groups. However, derivatizing these polymers to a predetermined extent has been the Achilles heel for this process. In this study, we have investigated the factors controlling the derivatization of the carboxyl moieties of HA with amine containing thiol, cystamine dihydrochloride (Cys), via carbodiimide crosslinking chemistry. We used fractional factorial design to screen and identify the significant factor(s) affecting the reaction, and response surface methodology (RSM) to develop a model equation for predicting the degree of thiolation of HA. Also, we analyzed the reaction mechanism for potential side reactions. We observed that N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC) (mole ratio with repeat unit of HA) is the significant factor controlling the degree of amidation. The quadratic equations developed from RSM predict the formulation for a desired degree of amidation of HA and percentage of potential side product. Hence, derivatizing HA to a predetermined extent with minimal side product can be achieved using the statistical design of experiments. 2014 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 103: 2300-2308, 2015. PMID:25369214

  20. Crystal Structure of Mammalian Cysteine dioxygenase: A Novel Mononuclear Iron Center for Cysteine Thiol Oxidation

    Energy Technology Data Exchange (ETDEWEB)

    Simmons,C.; Liu, Q.; Huang, Q.; Hao, Q.; Begley, T.; Karplus, P.; Stipanuk, M.

    2006-01-01

    Cysteine dioxygenase is a mononuclear iron-dependent enzyme responsible for the oxidation of cysteine with molecular oxygen to form cysteinesulfinate. This reaction commits cysteine to either catabolism to sulfate and pyruvate or to the taurine biosynthetic pathway. Cysteine dioxygenase is a member of the cupin superfamily of proteins. The crystal structure of recombinant rat cysteine dioxygenase has been determined to 1.5 Angstroms resolution, and these results confirm the canonical cupin {beta}-sandwich fold and the rare cysteinyl-tyrosine intramolecular crosslink (between Cys93 and Tyr157) seen in the recently reported murine cysteine dioxygenase structure. In contrast to the catalytically inactive mononuclear Ni(II) metallocenter present in the murine structure, crystallization of a catalytically competent preparation of rat cysteine dioxygenase revealed a novel tetrahedrally coordinated mononuclear iron center involving three histidines (His86, His88, and His140) and a water molecule. Attempts to acquire a structure with bound ligand using either co-crystallization or soaks with cysteine revealed the formation of a mixed disulfide involving Cys164 near the active site, which may explain previously observed substrate inhibition. This work provides a framework for understanding the molecular mechanisms involved in thiol dioxygenation and sets the stage for exploring the chemistry of both the novel mononuclear iron center and the catalytic role of the cysteinyl-tyrosine linkage.

  1. Self-consistent GW calculations of electronic transport in thiol- and amine-linked molecular junctions

    DEFF Research Database (Denmark)

    Strange, M.; Rostgaard, Carsten

    2011-01-01

    The electronic conductance of a benzene molecule connected to gold electrodes via thiol, thiolate, or amino anchoring groups is calculated using nonequilibrium Green functions in combination with the fully self-consistent GW approximation for exchange and correlation. The calculated conductance of benzenedithiol and benzenediamine is one-fifth that predicted by standard density functional theory (DFT), in very good agreement with experiments. In contrast, the widely studied benzenedithiolate structure is found to have a significantly higher conductance due to the unsaturated sulfur bonds. These findings suggest that more complex gold-thiolate structures where the thiolate anchors are chemically passivated by Au adatoms are responsible for the measured conductance. Analysis of the energy level alignment obtained with DFT, Hartree-Fock, and GW reveals the importance of self-interaction corrections (exchange) on the molecule and dynamical screening at the metal-molecule interface. The main effect of the GW self-energy is to renormalize the level positions; however, its influence on the shape of molecular resonances also affects the conductance. Non-self-consistent G(0)W(0) calculations, starting from either DFT or Hartree-Fock, yield conductance values within 50% of the self-consistent GW results.

  2. PLACE OF 5?-REDUCTASE IN THE THERAPY FOR BENIGN PROSTATIC HYPERPLASIA

    Directory of Open Access Journals (Sweden)

    C. Novac

    2007-10-01

    Full Text Available Twelve years ago, finasteride, the first 5?-reductase inhibitor, was introduced as drug therapy for benign prostatic hyperplasia, and more recently dutasteride has emerged as an alternative. The efficacy, safety and ability of these 5?-reductase inhibitors to reverse the natural progression of benign prostatic hyperplasia have been convincingly demonstrated and both drugs are now welt established in the medical armamentarium against the disease. Given the multifactorial etiology of benign hyperplasia, the usefulness of 5?-reductase inhibitors in combination with a adrenergic blockers has also been investigated and justified in select patients. Wider applications of 5?-reductase inhibitors are also emerging, though their perhaps most important new role as chemopreventive agents remains unclear.

  3. Metabolism of the synthetic progestogen norethynodrel by human ketosteroid reductases of the aldo-keto reductase superfamily.

    Science.gov (United States)

    Jin, Yi; Duan, Ling; Chen, Mo; Penning, Trevor M; Kloosterboer, Helenius J

    2012-04-01

    Human ketosteroid reductases of the aldo-keto reductase (AKR) superfamily, i.e. AKR1C1-4, are implicated in the biotransformation of synthetic steroid hormones. Norethynodrel (NOR, 17?-ethynyl-17?-hydroxy-estra-5(10)-en-3-one), the progestin component of the first marketed oral contraceptive, is known to undergo rapid and extensive metabolism to 3?- and 3?-hydroxymetabolites. The ability of the four human AKR1C enzymes to catalyze the metabolism of NOR has now been characterized. AKR1C1 and AKR1C2 almost exclusively converted NOR to 3?-hydroxy NOR, while AKR1C3 gave 3?-hydroxy NOR as the main product and AKR1C4 predominantly formed 3?-hydroxy NOR. Individual AKR1C enzymes also displayed distinct kinetic properties in the reaction of NOR. In contrast, norethindrone (NET), the ?(4)-isomer of NOR and the most commonly used synthetic progestogen, was not a substrate for the AKR1C enzymes. NOR is also structurally identical to the hormone replacement therapeutic tibolone (TIB), except TIB has a methyl group at the 7?-position. Product profiles and kinetic parameters for the reduction of NOR catalyzed by each individual AKR1C isoform were identical to those for the reduction of TIB catalyzed by the respective isoform. These data suggest that the presence of the 7?-methyl group has a minimal effect on the stereochemical outcome of the reaction and kinetic behavior of each enzyme. Results indicate a role of AKR1C in the hepatic and peripheral metabolism of NOR to 3?- and 3?-hydroxy NOR and provide insights into the differential pharmacological properties of NOR, NET and TIB. PMID:22210085

  4. Escherichia coli delta 1-pyrroline-5-carboxylate reductase: gene sequence, protein overproduction and purification.

    OpenAIRE

    Deutch, A. H.; Smith, C. J.; Rushlow, K. E.; Kretschmer, P. J.

    1982-01-01

    The sequence of the Escherichia coli proC gene which encodes for delta 1-pyrroline-5-carboxylate (PCA) reductase was determined. Overproduction of the proC gene product via an expression plasmid carrying the bacteriophage lambda PL promoter allowed the purification to homogeneity of PCA reductase by affinity adsorption chromatography. NH2 and COOH-terminal analysis and amino acid composition of the purified proC protein is consistent with the gene sequence reported. The molecular weight of th...

  5. DT-diaphorase and cytochrome B5 reductase in human lung and breast tumours.

    OpenAIRE

    Mara?n, A.; La?pez Cerain, A.; Hamilton, E.; Lewis, A. D.; Martinez-pea?uela, J. M.; Idoate, M. A.; Bello, J.

    1997-01-01

    The level of expression of enzymes that can activate or detoxify bioreductive agents within tumours has emerged as an important feature in the development of these anti-tumour compounds. The levels of two such reductase enzymes have been determined in 19 human non-small-cell lung tumours and 20 human breast tumours, together with the corresponding normal tissue. DT-diaphorase (DTD) enzyme levels (both expression and activity) were determined in these samples. Cytochrome b5 reductase (Cytb5R) ...

  6. X-Ray crystal structure of GarRtartronate semialdehyde reductase from Salmonella typhimurium

    OpenAIRE

    Osipiuk, J.; Zhou, M; Moy, S.; Collart, F.; Joachimiak, A.

    2009-01-01

    Tartronate semialdehyde reductases (TSRs), also known as 2-hydroxy-3-oxopropionate reductases, catalyze the reduction of tartronate semialdehyde using NAD as cofactor in the final stage of D-glycerate biosynthesis. These enzymes belong to family of structurally and mechanically related ?-hydroxyacid dehydrogenases which differ in substrate specificity and catalyze reactions in specific metabolic pathways. Here, we present the crystal structure of GarR a TSR from Salmonella typhimurium determi...

  7. Aldose reductases influence prostaglandin f2? levels and adipocyte differentiation in male mouse and human species.

    Science.gov (United States)

    Pastel, Emilie; Pointud, Jean-Christophe; Loubeau, Galle; Dani, Christian; Slim, Karem; Martin, Gwenalle; Volat, Fanny; Sahut-Barnola, Isabelle; Val, Pierre; Martinez, Antoine; Lefranois-Martinez, Anne-Marie

    2015-05-01

    Aldose reductases (AKR1B) are widely expressed oxidoreductases whose physiological function remains elusive. Some isoforms are genuine prostaglandin F2? (PGF2?) synthases, suggesting they might influence adipose homeostasis because PGF2? inhibits adipogenesis. This was shown by Akr1b7 gene ablation in the mouse, which resulted in increased adiposity related to a lower PGF2? content in fat. Yet humans have no ortholog gene for Akr1b7, so the role of aldose reductases in human adipose homeostasis remains to be explored. We analyzed expression of genes encoding human and mouse aldose reductase isoforms in adipose tissues and differentiating adipocytes to assess conserved mechanisms regulating PGF2? synthesis and adipogenesis. The Akr1b3 gene encoded the most abundant isoform in mouse adipose tissue, whereas Akr1b7 encoded the only isoform enriched in the stromal vascular fraction. Most mouse aldose reductase gene expression peaked in early adipogenesis of 3T3-L1 cells and diminished with differentiation. In contrast with its mouse ortholog Akr1b3, AKR1B1 expression increased throughout differentiation of human multipotent adipose-derived stem cells, paralleling PGF2? release, whereas PGF2? receptor (FP) levels collapsed in early differentiation. Pharmacological inhibition of aldose reductase using Statil altered PGF2? production and enhanced human multipotent adipose-derived stem adipocyte differentiation. As expected, the adipogenic effects of Statil were counteracted by an FP agonist (cloprostenol). Thus, in both species aldose reductase-dependent PGF2? production could be important in early differentiation to restrict adipogenesis. PGF2? antiadipogenic signaling could then be toned down through the FP receptor or aldose reductases down-regulation in human and mouse cells, respectively. Our data suggest that aldose reductase inhibitors could have obesogenic potential. PMID:25730106

  8. Sepiapterin reductase expression is increased in Parkinsons disease brain tissue.

    OpenAIRE

    Tobin, Jennifer E.; CUI, Jing; Jemma B. Wilk; Latourelle, Jeanne C.; Laramie, Jason M; McKee, Ann C; Guttman, Mark; Karamohamed, Samer; DeStefano, Anita L.; Myers, Richard H.

    2007-01-01

    The PARK3 locus on chromosome 2p13 has shown linkage to both the development and age of onset of Parkinsons disease (PD). One candidate gene at this locus is sepiapterin reductase (SPR). Sepiapterin reductase catalyzes the final step in the biosynthetic pathway of tetrahydrobiopterin (BH4), an essential cofactor for aromatic amino acid hydrolases including tyrosine hydroxylase, the rate-limiting enzyme in dopamine synthesis. The expression of SPR was assayed using semiquantitative real-tim...

  9. New Insights into Biliverdin Reductase Functions: Linking Heme Metabolism to Cell Signaling

    Science.gov (United States)

    PhD Mahin D. Maines (University of Rochester Medical Center Department of Biochemistry and Biophysics)

    2005-12-01

    Biliverdin reductase (BVR) functions in cell signaling through three distinct tracks: a dual-specificity kinase that functions in the insulin receptor/MAPK pathways; a bzip-type transcription factor for ATF-2/CREB and HO-1 regulation; and a reductase that catalyzes the conversion of biliverdin to bilirubin. These, together with the protein?s primary and secondary features, intimately link BVR to the entire spectrum of cell-signaling cascades.

  10. Current status of 5?-reductase inhibitors in the treatment of benign hyperplasia of prostate

    OpenAIRE

    Kumar Vijay; Wahane Vishal

    2008-01-01

    Benign prostatic hyperplasia (BPH) is a common problem in aging men, which is associated with lower urinary tract symptoms. This condition is dependent on the presence of androgens for its progression, and medical therapy is the first-line treatment for BPH patients with moderate-to-severe symptoms and includes the use of either alpha 1-adrenergic blockers or 5?-reductase inhibitors. Adrenergic blocking drugs reduce the dynamic component while the 5?-reductase inhibitors reduce the ...

  11. Normal bone density in male pseudohermaphroditism due to 5a- reductase 2 deficiency

    OpenAIRE

    Costa Elaine Maria Frade; Arnhold Ivo Jorge Prado; Inacio Marlene; Mendonca Berenice Bilharinho

    2001-01-01

    Bone is an androgen-dependent tissue, but it is not clear whether the androgen action in bone depends on testosterone or on dihydrotestosterone. Patients with 5alpha-reductase 2 deficiency present normal levels of testosterone and low levels of dihydrotestosterone, providing an in vivo human model for the analysis of the effect of testosterone on bone. OBJECTIVE: To analyze bone mineral density in 4 adult patients with male pseudohermaphroditism due to 5alpha-reductase 2 deficiency. RESULTS: ...

  12. Aldose reductase deficiency in mice prevents azoxymethane-induced colonic preneoplastic aberrant crypt foci formation

    OpenAIRE

    Tammali, Ravinder; Reddy, Aramati B. M.; Kota V. Ramana; PETRASH, J. MARK; Srivastava, Satish K.

    2008-01-01

    Aldose reductase (AR; EC 1.1.1.21), an nicotinamide adenine dinucleotide phosphate-dependent aldoketo reductase, has been shown to be involved in oxidative stress signaling initiated by inflammatory cytokines, chemokines and growth factors. Recently, we have shown that inhibition of this enzyme prevents the growth of colon cancer cells in vitro as well as in nude mice xenografts. Herein, we investigated the mediation of AR in the formation of colonic preneoplastic aberrant crypt foci (ACF) u...

  13. Genetic Diversity of Benzoyl Coenzyme A Reductase Genes Detected in Denitrifying Isolates and Estuarine Sediment Communities

    OpenAIRE

    Song, Bongkeun; Ward, Bess B.

    2005-01-01

    Benzoyl coenzyme A (benzoyl-CoA) reductase is a central enzyme in the anaerobic degradation of organic carbon, which utilizes a common intermediate (benzoyl-CoA) in the metabolism of many aromatic compounds. The diversity of benzoyl-CoA reductase genes in denitrifying bacterial isolates capable of degrading aromatic compounds and in river and estuarine sediment samples from the Arthur Kill in New Jersey and the Chesapeake Bay in Maryland was investigated. Degenerate primers were developed fro...

  14. Structural and functional characterization of HMG-COA reductase from Artemisia annua

    OpenAIRE

    Kiran, Usha; Ram, Mauji; Khan, Mather Ali; Khan, Salim; Jha, Prabhakar; Alam, Afshar; Abdin, Malik Zainul

    2010-01-01

    Plants synthesize a great variety of isoprenoid products that are required not only for normal growth and development but also for their adaptive responses to environmental challenges. However, despite the remarkable diversity in the structure and function of plant isoprenoids, they all originate from a single metabolic precursor, mevalonic acid. The synthesis of mevalonic acid is catalysed by the enzyme, 3?hydroxy?3?methylglutaryl coenzyme A reductase (HMG? CoA reductase). T...

  15. Acceleration of an aldo-keto reductase by minimal loop engineering

    OpenAIRE

    Krump, C.; Vogl, M.; Brecker, L; Nidetzky, B; Kratzer, R.

    2014-01-01

    Aldo-keto reductases tighten coenzyme binding by forming a hydrogen bond across the pyrophosphate group of NAD(P)(H). Mutation of the hydrogen bonding anchor Lys24 in Candida tenuis xylose reductase prevents fastening of the safety belt around NAD(H). The loosened NAD(H) binding leads to increased turnover numbers (kcat) for reductions of bulky-bulky ketones at constant substrate and coenzyme affinities (i.e. Km Ketone, Km NADH).

  16. Comparison of the Stereospecificity and Immunoreactivity of NADH-Ferricyanide Reductases in Plant Membranes

    OpenAIRE

    Fredlund, Kenneth M.; Struglics, Andr; Widell, Susanne; Askerlund, Per; Kader, Jean-Claude; Mller, Ian M

    1994-01-01

    The substrate stereospecificity of NADH-ferricyanide reductase activities in the inner mitochondrial membrane and peroxisomal membrane of potato (Solanum tuberosum L.) tubers, spinach (Spinacea oleracea L.) leaf plasma membrane, and red beetroot (Beta vulgaris L.) tonoplast were all specific for the [beta]-hydrogen of NADH, whereas the reductases in wheat root (Triticum aestivum L.) endoplasmic reticulum and potato tuber outer mitochondrial membrane were both [alpha]-hydrogen specific. In all...

  17. Essential Protein-Protein Interactions between Plasmodium falciparum Thymidylate Synthase and Dihydrofolate Reductase Domains*

    OpenAIRE

    Shallom, Shamira; Zhang, Kai; JIANG, Lei; Rathod, Pradipsinh K

    1999-01-01

    In Plasmodium falciparum, dihydrofolate reductase and thymidylate synthase activities are conferred by a single 70-kDa bifunctional polypeptide (DHFR-TS, dihydrofolate reductase-thymidylate synthase) which assembles into a functional 140-kDa homodimer. In mammals, the two enzymes are smaller distinct molecules encoded on different genes. A 27-kDa amino domain of malarial DHFR-TS is sufficient to provide DHFR activity, but the structural requirements for TS function have not been established. ...

  18. Discovery of potent pteridine reductase inhibitors to guide antiparasite drug development

    OpenAIRE

    Cavazzuti, Antonio; Paglietti, Giuseppe; Hunter, William N.; Gamarro, Francisco; Piras, Sandra; Loriga, Mario; Alleca, Sergio; Corona, Paola; McLuskey, Karen; Tulloch, Lindsay; Gibellini, Federica; Ferrari, Stefania; Costi, Maria Paola

    2008-01-01

    Pteridine reductase (PTR1) is essential for salvage of pterins by parasitic trypanosomatids and is a target for the development of improved therapies. To identify inhibitors of Leishmania major and Trypanosoma cruzi PTR1, we combined a rapid-screening strategy using a folate-based library with structure-based design. Assays were carried out against folate-dependent enzymes including PTR1, dihydrofolate reductase (DHFR), and thymidylate synthase. Affinity profiling determi...

  19. X-ray structure of trypanothione reductase from Crithidia fasciculata at 2. 4- angstrom resolution

    Energy Technology Data Exchange (ETDEWEB)

    Kuriyan, J.; Xiangpeng Kong; Krishna, T.S.R.; Murgolo, N.J.; Field, H.; Cerami, A.; Henderson, G.B. (Rockefeller Univ., New York, NY (United States)); Sweet, R.M. (Brookhaven National Lab., Upton, NY (United States))

    1991-10-01

    Trypanosomes and related protozoan parasites lack glutathione reductase and possess instead a closely related enzyme that serves as the reductant of a bis(glutathione)-spermidien conjugate, trypanothione. The human and parasite enzymes have mutually exclusive substrate specificities, providing a route for the design of therapeutic agents by specific inhibition of the parasite enzyme. The authors report here the three-dimensional structure of trypanothione reductase from Crithidia fasciculata and show that it closely resembles the structure of human glutathione reductase. In particular, the core structure surrounding the catalytic machinery is almost identical in the two enzymes. However, significant differences are found at the substrate binding sites. A cluster of basic residues in glutathione reductase is replaced by neutral, hydrophobic, or acidic residues in trypanothione reductase, consistent with the nature of the spermidine linkage and the change in overall charge of the substrate from {minus}2 to +1, respectively. The binding site is more open in trypanothione reductase due to rotations of about 4{degree} in the domains that form in site, with relative shifts of as much as 2-3 {angstrom} in residues that can interact with potential inhibitors and complement previous modeling and mutagenesis studies on the two enzymes.

  20. A soluble 3-hydroxy-3-methylglutaryl-CoA reductase in the protozoan Trypanosoma cruzi

    DEFF Research Database (Denmark)

    Pena Diaz, Javier; Montalvetti, A

    1997-01-01

    We report the isolation and characterization of a genomic clone containing the open reading frame sequence for 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase from Trypanosoma cruzi, the causative agent of Chagas' disease. The protozoan gene encoded for a smaller polypeptide than the rest of the genes described from eukaryotic organisms and the deduced amino acid sequence could be aligned with the C-terminal half of animal and plant reductases exhibiting pronounced similarity to other eukaryotic counterparts. Further examination of the 5' flanking region by cDNA analysis and establishment of the splice acceptor sites clearly indicated that the corresponding mRNA apparently lacks sequences encoding a membrane N-terminal domain. The reductase gene is a single copy and is located on a chromosome of 1.36 Mb as determined by contour-clamped homogeneous electric field electrophoresis. The overall cellular distribution of enzymic activity was investigated after differential centrifugation of Trypanosoma cell extracts. Reductase activity was primarily associated with the cellular soluble fraction because 95% of the total cellular activity was recovered in the supernatant and was particularly sensitive to proteolytic inactivation. Furthermore the enzyme can be efficiently overexpressed in a highly active form by using the expression vector pET-11c. Thus Trypanosoma cruzi HMG-CoA reductase is unique in the sense that it totally lacks the membrane-spanning sequences present in all eukaryotic HMG-CoA reductases so far characterized.

  1. The catalytic mechanism of NADH-dependent reduction of 9,10-phenanthrenequinone by Candida tenuis xylose reductase reveals plasticity in an aldo-keto reductase active site

    OpenAIRE

    Pival, Simone L.; Klimacek, Mario; Nidetzky, Bernd

    2009-01-01

    Abstract Despite their widely varying physiological functions in carbonyl metabolism, Candida tenuis xylose reductase (AKR2B5) and many related enzymes of the aldo-keto reductase protein superfamily utilise 9,10 phenanthrenequinone (PQ) as a common in vitro substrate for NAD(P)H-dependent reduction. The catalytic roles of the conserved active-site residues (Tyr51, Lys80, His113) of AKR2B5 in the conversion of the reactive ?-dicarbonyl moiety of PQ are not well understood. Using wi...

  2. Stereoselective synthesis of (R)-3-quinuclidinol through asymmetric reduction of 3-quinuclidinone with 3-quinuclidinone reductase of Rhodotorula rubra.

    Science.gov (United States)

    Uzura, Atsuko; Nomoto, Fumiki; Sakoda, Akiko; Nishimoto, Yukifumi; Kataoka, Michihiko; Shimizu, Sakayu

    2009-06-01

    A novel nicotinamide adenine dinucleotide phosphate-dependent carbonyl reductase, 3-quinuclidinone reductase, was isolated from Rhodotorula rubra JCM3782. The enzyme catalyzes the asymmetric reduction of 3-quinuclidinone to (R)-3-quinuclidinol. The gene encoding the enzyme was also cloned and sequenced. A 819-bp nucleotide fragment was confirmed to be the gene encoding the 3-quinuclidinone reductase by agreement of the internal amino acid sequences of the purified enzyme. The gene encodes a total of 272 amino acid residues, and the deduced amino acid sequence shows similarity to those of several short-chain dehydrogenase/reductase family proteins. An expression vector, pWKLQ, which contains the full length 3-quinuclidinone reductase gene was constructed. Using Escherichia coli cells coexpressing the 3-quinuclidinone reductase and glucose dehydrogenase (cofactor regeneration enzyme) genes, 618 mM 3-quinuclidinone was almost stiochiometrically converted to (R)-3-quinuclidinol with an >99.9% enantiomeric excess within 21 h of reaction. PMID:19234697

  3. Discovery and characterization of a second mammalian thiol dioxygenase, cysteamine dioxygenase.

    Science.gov (United States)

    Dominy, John E; Simmons, Chad R; Hirschberger, Lawrence L; Hwang, Jesse; Coloso, Relicardo M; Stipanuk, Martha H

    2007-08-31

    There are only two known thiol dioxygenase activities in mammals, and they are ascribed to the enzymes cysteine dioxygenase (CDO) and cysteamine (2-aminoethanethiol) dioxygenase (ADO). Although many studies have been dedicated to CDO, resulting in the identification of its gene and even characterization of the tertiary structure of the protein, relatively little is known about cysteamine dioxygenase. The failure to identify the gene for this protein has significantly hampered our understanding of the metabolism of cysteamine, a product of the constitutive degradation of coenzyme A, and the synthesis of taurine, the final product of cysteamine oxidation and the second most abundant amino acid in mammalian tissues. In this study we identified a hypothetical murine protein homolog of CDO (hereafter called ADO) that is encoded by the gene Gm237 and belongs to the DUF1637 protein family. When expressed as a recombinant protein, ADO exhibited significant cysteamine dioxygenase activity in vitro. The reaction was highly specific for cysteamine; cysteine was not oxidized by the enzyme, and structurally related compounds were not competitive inhibitors of the reaction. When overexpressed in HepG2/C3A cells, ADO increased the production of hypotaurine from cysteamine. Similarly, when endogenous expression of the human ADO ortholog C10orf22 in HepG2/C3A cells was reduced by RNA-mediated interference, hypotaurine production decreased. Western blots of murine tissues with an antibody developed against ADO showed that the protein is ubiquitously expressed with the highest levels in brain, heart, and skeletal muscle. Overall, these data suggest that ADO is responsible for endogenous cysteamine dioxygenase activity. PMID:17581819

  4. Cadmium sensitivity, uptake, subcellular distribution and thiol induction in a marine diatom: Recovery from cadmium exposure

    International Nuclear Information System (INIS)

    Studies in the recovery from metal stress and the tolerance development to metal exposure of aquatic organisms are important for the understanding of epidemic pollution. In this study, the responses of a marine diatom, Thalassiosira nordenskioeldii, following recovery from environmental cadmium (Cd) stress were investigated. The diatoms were exposed to different concentrations of Cd for 7 days, and were then allowed different periods of time to recover. The Cd sensitivity increased after recovery from Cd stress, followed by a gradual restoration. The extent of restoration depended on both the recovery time and the environmental Cd stress during the exposure period. A complete restoration of Cd tolerance proved to be impossible for cells pre-exposed to High-Cd. The Cd cellular burden and subcellular Cd concentration decreased to the control level within the first day of recovery, indicating that the elevated sensitivity may have been due to the accumulation of functional damage caused by Cd exposure instead of a result of physical Cd accumulation. The rapid change in phytochelatins (PC) to both the increase in and the withdrawal of environmental Cd stress made it a good quantitative bioindicator of environmental Cd contamination. However, the relationships between Cd distribution in the metal sensitive fraction (MSF-Cd) or intracellular Cd to thiol ratio (intra-Cd/PC-SH) and the relative change in the median inhibition [Cd2+] ([Cd2+]-based-IC2+] ([Cd2+]-based-IC50, i.e., Cd sensitivity) differed for the various exposure and recovery periods tested. Our study suggests that more attention should be given to the recovery of aquatic organisms from episodic metal exposure.

  5. Cadmium sensitivity, uptake, subcellular distribution and thiol induction in a marine diatom: Recovery from cadmium exposure

    Energy Technology Data Exchange (ETDEWEB)

    Wang Mengjiao [State Key Laboratory in Marine Pollution, Section of Marine Ecology and Biotechnology, Division of Life Science, Hong Kong University of Science and Technology (HKUST), Clear Water Bay, Kowloon (Hong Kong); Wang Wenxiong, E-mail: wwang@ust.hk [State Key Laboratory in Marine Pollution, Section of Marine Ecology and Biotechnology, Division of Life Science, Hong Kong University of Science and Technology (HKUST), Clear Water Bay, Kowloon (Hong Kong)

    2011-01-25

    Studies in the recovery from metal stress and the tolerance development to metal exposure of aquatic organisms are important for the understanding of epidemic pollution. In this study, the responses of a marine diatom, Thalassiosira nordenskioeldii, following recovery from environmental cadmium (Cd) stress were investigated. The diatoms were exposed to different concentrations of Cd for 7 days, and were then allowed different periods of time to recover. The Cd sensitivity increased after recovery from Cd stress, followed by a gradual restoration. The extent of restoration depended on both the recovery time and the environmental Cd stress during the exposure period. A complete restoration of Cd tolerance proved to be impossible for cells pre-exposed to High-Cd. The Cd cellular burden and subcellular Cd concentration decreased to the control level within the first day of recovery, indicating that the elevated sensitivity may have been due to the accumulation of functional damage caused by Cd exposure instead of a result of physical Cd accumulation. The rapid change in phytochelatins (PC) to both the increase in and the withdrawal of environmental Cd stress made it a good quantitative bioindicator of environmental Cd contamination. However, the relationships between Cd distribution in the metal sensitive fraction (MSF-Cd) or intracellular Cd to thiol ratio (intra-Cd/PC-SH) and the relative change in the median inhibition [Cd{sup 2+}] ([Cd{sup 2+}]-based-IC{sub 50}, i.e., Cd sensitivity) differed for the various exposure and recovery periods tested. Our study suggests that more attention should be given to the recovery of aquatic organisms from episodic metal exposure.

  6. Antioxidant defences of Spironucleus vortens: Glutathione is the major non-protein thiol.

    Science.gov (United States)

    Williams, C F; Yarlett, N; Aon, M A; Lloyd, D

    2014-08-01

    The aerotolerant hydrogenosome-containing piscine diplomonad, Spironucleus vortens, is able to withstand high fluctuations in O? tensions during its life cycle. In the current study, we further investigated the O? scavenging and antioxidant defence mechanisms which facilitate the survival of S. vortens under such oxidizing conditions. Closed O? electrode measurements revealed that the S. vortens ATCC 50386 strain was more O? tolerant than a freshly isolated S. vortens intestinal strain (Sv1). In contrast to the related human diplomonad, Giardia intestinalis, RP-HPLC revealed the major non-protein thiols of S. vortens to be glutathione (GSH, 776 nmol/10? cells) with cysteine and H2S as minor peaks. Furthermore, antioxidant proteins of S. vortens were assayed enzymatically and revealed that S. vortens possesses superoxide dismutase and NADH oxidase (883 and 37.5nmol/min/mg protein, respectively), but like G. intestinalis, lacks catalase and peroxidase activities. Autofluorescence of NAD(P)H and FAD alongside the fluorescence of the GSH-adduct in monochlorobimane-treated live organisms allowed the monitoring of redox balances before and after treatment with inhibitors, metronidazole and auranofin. H?O? was emitted into the exterior of S. vortens at a rate of 2.85 pmol/min/10? cells. Metronidazole and auranofin led to depletion of S. vortens intracellular NAD(P)H pools and an increase in H?O? release with concomitant oxidation of GSH, respectively. Garlic-derived compounds completely inhibited O? consumption by S. vortens (ajoene oil), or significantly depleted the intracellular GSH pool of the organism (allyl alcohol and DADS). Hence, antioxidant defence mechanisms of S. vortens may provide novel targets for parasite chemotherapy. PMID:25101875

  7. Surface plasmon resonance and magnetism of thiol-capped gold nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Guerrero, E; Munoz-Marquez, M A; Fernandez, A [Instituto de Ciencia de Materiales de Sevilla, CSIC-US, Avenida Americo Vespucio 49, E-41092 Sevilla (Spain); Garcia, M A; Crespo, P; Fernandez-Pinel, E; Hernando, A [Instituto de Magnetismo Aplicado (UCM-ADIF-CSIC), PO Box 155, E-28230 Las Rozas, Madrid (Spain)], E-mail: miguel.angel@icmse.csic.es

    2008-04-30

    Surface plasmon resonance measurements and magnetic characterization studies have been carried out for two types of thiol-capped gold nanoparticles (NPs) with similar diameters between 2.0 and 2.5 nm and different organic molecules linked to the sulfur atom: dodecanethiol and tiopronin. In addition, Au NPs capped with tetraoctyl ammonium bromide have also been included in the investigation since such capping molecules weakly interact with the gold surface atoms and, therefore, this system can be used as a model for naked gold NPs; such particles presented a bimodal size distribution with diameters around 1.5 and 5 nm. The plasmon resonance is non-existent for tiopronin-capped NPs, whereas a trace of such a feature is observed for NPs covered with dodecanethiol molecules and a bulk-like feature is measured for NPs capped with tetralkyl ammonium salts. These differences would indicate that the modification of the surface electronic structure of the Au NPs depends on the geometry and self-assembling capabilities of the capping molecules and on the electric charge transferred between Au and S atoms. Regarding the magnetization, dodecanethiol-capped NPs have a ferromagnetic-like behaviour, while the NPs capped with tiopronin exhibit a paramagnetic behaviour and tetralkyl ammonium-protected NPs are diamagnetic across the studied temperature range; straight chains with a well-defined symmetry axis can induce orbital momentum on surface electrons close to the binding atoms. The orbital momentum not only contributes to the magnetization but also to the local anisotropy, giving rise to permanent magnetism. Due to the domain structure of the adsorbed molecules, orbital momentum is not induced for tiopronin-capped NPs and the charge transfer only induces a paramagnetic spin component.

  8. Characterization of thiol-specific antioxidant 1 (TSA1) of Candida albicans.

    Science.gov (United States)

    Shin, Duck Hyang; Jung, Saem; Park, So Jeong; Kim, Yeo Jung; Ahn, Jung Min; Kim, Wankee; Choi, Wonja

    2005-08-01

    We previously identified several proteins that are differentially expressed in pathogenic hyphae by comparing protein profiles of yeast and hyphae of Candida albicans. One of these, thiol-specific antioxidant 1 (TSA1), attracted our attention because it may play some roles in surviving an unfavourable oxidative environment created by host cells. Two alleles of the C. albicans TSA1 (CaTSA1) gene are located in opposite orientation on the same chromosome. Using PCR-directed disruption cassettes and URA-Blaster, a series of deletion mutants that lack one to four copies were constructed to examine the functions of CaTSA1. Northern and Western analyses showed that both the transcript and protein products of CaTSA1 decreased proportionally to the disrupted copy number and were completely absent in the null mutant, indicating that all four TSA1 copies are equally functional at the transcriptional level. Intracellular H2O2 increased by an order of magnitude in deletion mutants lacking three to four copies, suggesting that CaTsa1p is not a redundant H2O2 scavenger. CaTsa1p was indispensable for yeast-to-hyphal transition when C. albicans was cultured under oxidative stress. The level of its oxidized form increased approximately five-fold in hyphal cells, whereas that of the reduced form increased two-fold compared to yeast cells. The ratio of oxidized to reduced form was increased three-fold in hyphal cells. This overall increase was found to be controlled at the post-transcriptional level. Interestingly, CaTsa1p is translocated to the nucleus of hyphal cells. These findings may be of biological significance for differentiation and pathogenicity. PMID:16134099

  9. Physical Incorporation of NADPH-cytochrome P450 Reductase and Cytochrome P450 into Phospholipid Vesicles using Glycocholate and Biobeads*

    OpenAIRE

    Reed, James R.; Brignac-huber, Lauren M.; Backes, Wayne L.

    2007-01-01

    In a previous study from our lab (Drug Metab. Disp. (2006) 34, 660666), we found several limitations with published methods (cholate gel filtration and cholate dialysis) for the incorporation of cytochromes P450 and P450 reductase into phospholipid vesicles. We found that a significant proportion of reductase was not incorporated in the vesicles when the amount of reductase was equal to or greater than that of CYP2B4 in the systems reconstituted with phosphatidylcholine. Furthermore, imple...

  10. Development of Transformation System of Verticillium lecanii (Lecanicillium spp.) (Deuteromycotina: Hyphomycetes) Based on Nitrate Reductase Gene of Aspergillus nidulans

    OpenAIRE

    Hasan, Saba; Singh, Rana Inder; Singh, Sandhu Sardul

    2011-01-01

    A heterologous transformation system was developed for V. lecanii based on the complementation of a nitrate reductase mutant. Nitrate reductase mutants were obtained by resistance to chlorate in a rate of 23.24% when compared to other mutations that lead to the chlorate resistance. Mutant no. 01 and 04 was chosen for the transformation experiments. Plasmid pBT was used as transformation vector containing the Aspergillus nidulans nitrate reductase gene. A frequency of approximately 3 transform...

  11. In vivo regulation of human mononuclear leukocyte 3-hydroxy-3-methylglutaryl coenzyme A reductase. Studies in normal subjects.

    OpenAIRE

    Harwood, H. J.; Bridge, D. M.; Stacpoole, P. W.

    1987-01-01

    In vivo regulation of microsomal HMG CoA reductase activity was investigated in freshly isolated mononuclear leukocytes from 26 healthy adult males. Reductase activity exhibited a diurnal rhythm and decreased during fasting. Enzyme activity was also modulated in vivo by alterations in dietary and plasma cholesterol, suggesting the existence of an operative cholesterol feedback regulatory system. A single, high cholesterol meal decreased reductase activity within 2 h. In addition, rapid deplet...

  12. Feedback Regulation of Cholesterol Synthesis: Sterol-Accelerated Ubiquitination and Degradation of HMG CoA Reductase

    OpenAIRE

    Debose-boyd, Russell A.

    2008-01-01

    HMG CoA reductase produces mevalonate, an important intermediate in the synthesis of cholesterol and essential nonsterol isoprenoids. The reductase is subject to an exorbitant amount of feedback control through multiple mechanisms that are mediated by sterol and nonsterol end-products of mevalonate metabolism. Here, I will discuss recent advances that shed light on one mechanism for control of reductase, which involves rapid degradation of the enzyme. Accumulation of certain sterols triggers ...

  13. 3-Hydroxy-3-methylglutaryl-coenzyme A reductase from Haloferax volcanii: purification, characterization, and expression in Escherichia coli.

    OpenAIRE

    Bischoff, K. M.; Rodwell, V. W.

    1996-01-01

    Prior work from this laboratory characterized eukaryotic (hamster) and eubacterial (Pseudomonas mevalonii) 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductases. We report here the characterization of an HMG-CoA reductase from the third domain, the archaea. HMG-CoA reductase of the halobacterium Haloferax volcanii was initially partially purified from extracts of H. volcanii. Subsequently, a portion of the H. volcanii lovastatin (formerly called mevinolin) resistance marker mev was subcl...

  14. Structural and kinetic determinants of aldehyde reduction by aldose reductase.

    Science.gov (United States)

    Srivastava, S; Watowich, S J; Petrash, J M; Srivastava, S K; Bhatnagar, A

    1999-01-01

    Aldose reductase (AR) is a member of the aldo-keto reductase superfamily. Due to its ability to catalyze the formation of sorbitol from glucose during hyperglycemic and hypertonic stress, the aldose-reducing property of AR has been accepted as its main physiological and pathological function. Nonetheless, AR is a poor catalyst for glucose reduction and displays active-site properties unexpected of a carbohydrate-binding protein. We, therefore, examined the catalytic properties of AR with a series of naturally occurring aldehydes, compatible in their hydrophobicity to the large apolar active site of the enzyme. Our results show that recombinant human AR is an efficient catalyst for the reduction of medium- to long-chain unbranched saturated and unsaturated aldehydes. The enzyme displayed selective preference for saturated aldehydes, such as hexanal, and unsaturated aldehydes, such as trans-2-octenal and nonenal as well as their 4-hydroxy derivatives. Short-chain aldehydes such as propanal and acrolein were reduced less efficiently. Branched derivatives of acrolein or its glutathione conjugate (GS-propanal) were, however, reduced with high efficiency. In the absence of NADPH, the alpha, beta unsaturated aldehydes caused covalent modification of the enzyme. On the basis of electrospray mass spectrometric analysis of the wild-type and site-directed mutants of AR (in which the solvent exposed cysteines were individually replaced with serine), the site of modification was identified to be the active-site residue, Cys 298. The unsaturated aldehydes, however, did not modify the enzyme bound to NADPH and did not inactivate the enzyme during catalysis. Modeling studies indicate that the large hydrophobic active site of AR can accommodate a large number of aldehydes without changes in the structure of the binding site or movement of side chains. High hydrophobicity due to long alkyl chains or apolar substituents appears to stabilize the interaction of the aldehyde substrates with the enzyme. Apparently, such hydrophobic interactions provide substrate selectivity and catalytic efficiency of the order achievable by hydrogen bonding. Since several of the aldehydes reduced by AR are either environmental and pharmacological pollutants or products of lipid peroxidation, the present studies provide the basis of future investigations on the role of AR in regulating aldehyde metabolism particularly under pathological states associated with oxidative stress and/or aldehyde toxicity. PMID:9890881

  15. Isobutyraldehyde production from Escherichia coli by removing aldehyde reductase activity

    Directory of Open Access Journals (Sweden)

    Rodriguez Gabriel M

    2012-06-01

    Full Text Available Abstract Background Increasing global demand and reliance on petroleum-derived chemicals will necessitate alternative sources for chemical feedstocks. Currently, 99% of chemical feedstocks are derived from petroleum and natural gas. Renewable methods for producing important chemical feedstocks largely remain unaddressed. Synthetic biology enables the renewable production of various chemicals from microorganisms by constructing unique metabolic pathways. Here, we engineer Escherichia coli for the production of isobutyraldehyde, which can be readily converted to various hydrocarbons currently derived from petroleum such as isobutyric acid, acetal, oxime and imine using existing chemical catalysis. Isobutyraldehyde can be readily stripped from cultures during production, which reduces toxic effects of isobutyraldehyde. Results We adopted the isobutanol pathway previously constructed in E. coli, neglecting the last step in the pathway where isobutyraldehyde is converted to isobutanol. However, this strain still overwhelmingly produced isobutanol (1.5?g/L/OD600 (isobutanol vs 0.14?g/L/OD600 (isobutyraldehyde. Next, we deleted yqhD which encodes a broad-substrate range aldehyde reductase known to be active toward isobutyraldehyde. This strain produced isobutanol and isobutyraldehyde at a near 1:1 ratio, indicating further native isobutyraldehyde reductase (IBR activity in E. coli. To further eliminate isobutanol formation, we set out to identify and remove the remaining IBRs from the E. coli genome. We identified 7 annotated genes coding for IBRs that could be active toward isobutyraldehyde: adhP, eutG, yiaY, yjgB, betA, fucO, eutE. Individual deletions of the genes yielded only marginal improvements. Therefore, we sequentially deleted all seven of the genes and assessed production. The combined deletions greatly increased isobutyraldehyde production (1.5?g/L/OD600 and decreased isobutanol production (0.4?g/L/OD600. By assessing production by overexpression of each candidate IBR, we reveal that AdhP, EutG, YjgB, and FucO are active toward isobutyraldehyde. Finally, we assessed long-term isobutyraldehyde production of our best strain containing a total of 15 gene deletions using a gas stripping system with in situ product removal, resulting in a final titer of 35?g/L after 5?days. Conclusions In this work, we optimized E. coli for the production of the important chemical feedstock isobutyraldehyde by the removal of IBRs. Long-term production yielded industrially relevant titers of isobutyraldehyde with in situ product removal. The mutational load imparted on E. coli in this work demonstrates the versatility of metabolic engineering for strain improvements.

  16. Structural basis for the high all-trans-retinaldehyde reductase activity of the tumor marker AKR1B10

    OpenAIRE

    Gallego, Oriol; Ruiz, F. Xavier; Arde?vol, Albert; Domi?nguez, Marta; Alvarez, Rosana; Lera, Angel R.; Rovira, Carme; Farre?s, Jaume; Fita, Ignacio; Pare?s, Xavier

    2007-01-01

    AKR1B10 is a human aldo-keto reductase (AKR) found to be elevated in several cancer types and in precancerous lesions. In vitro, AKR1B10 exhibits a much higher retinaldehyde reductase activity than any other human AKR, including AKR1B1 (aldose reductase). We here demonstrate that AKR1B10 also acts as a retinaldehyde reductase in vivo. This activity may be relevant in controlling the first step of retinoic acid synthesis. Up-regulation of AKR1B10, resulting in retinoic acid depletion, may lead...

  17. Simple quantitative determination of potent thiols at ultratrace levels in wine by derivatization and high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) analysis.

    Science.gov (United States)

    Capone, Dimitra L; Ristic, Renata; Pardon, Kevin H; Jeffery, David W

    2015-01-20

    Volatile sulfur compounds contribute characteristic aromas to foods and beverages and are widely studied, because of their impact on sensory properties. Certain thiols are particularly important to the aromas of roasted coffee, cooked meat, passion fruit, grapefruit, and guava. These same thiols enhance the aroma profiles of different wine styles, imparting pleasant aromas reminiscent of citrus and tropical fruits (due to 3-mercaptohexan-1-ol, 3-mercaptohexyl acetate, 4-mercapto-4-methylpentan-2-one), roasted coffee (2-furfurylthiol), and struck flint (benzyl mercaptan), at nanogram-per-liter levels. In contrast to the usual gas chromatography (GC) approaches, a simple and unique high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed for routine analysis of five wine thiols, using 4,4'-dithiodipyridine (DTDP) as a derivatizing agent and polydeuterated internal standards for maximum accuracy and precision. DTDP reacted rapidly with thiols at wine pH and provided stable derivatives, which were enriched by solid-phase extraction (SPE) prior to analysis by HPLC-MS/MS. All steps were optimized and the method was validated in different wine matrices, with method performance being comparable to a well-optimized but more cumbersome gas chromatography-mass spectrometry (GC-MS) method. A range of commercial wines was analyzed with the new method, revealing the distribution of the five thiols in white, red, ros, and sparkling wine styles. PMID:25562625

  18. Characterization of thiol-conjugated metabolites of ginger components shogaols in mouse and human rrine and modulation of the glutathione levels in cancer cells by [6]-shogaol

    Science.gov (United States)

    Chen, Huadong; Soroka, Dominique N.; Hu, Yuhui; Chen, Xiaoxin; Sang, Shengmin

    2013-01-01

    Scope Shogaols, a series of major constituents in dried ginger with the most abundant being [6]-, [8]-, and [10]-shogaols, show much higher anti-cancer potencies than gingerols. Previously, we reported the mercapturic acid pathway as a major metabolic route for [6]-shogaol in mice. However, it is still unclear how the side chain length affects the metabolism of shogaols and how shogaols are metabolized in humans. Methods and results We first investigate the metabolism of [10]-shogaol in mouse urine, and then investigate the biotransformation of shogaols in human urine. Our results show that eight major thiol-conjugated metabolites of [10]-shogaol were detected in mouse urine, while six major thiol-conjugated metabolites of [6]-shogaol, two thiol-conjugated metabolites of [8]-shogaol, and two thiol-conjugated metabolites of [10]-shogaol were detected in urine collected from human after drinking ginger tea, using liquid chromatography/electrospray ionization tandem mass spectrometry. Our results clearly indicate the mercapturic acid pathway is a major metabolic route for [10]-shogaol in mice and for shogaols in human. Furthermore, we also investigated the regulation of glutathione (GSH) by [6]-shogaol in human colon cancer cells HCT-116. Our results show [6]-shogaol, after initially depleting glutathione levels, can subsequently restore and increase GSH levels over time. Conclusion Shogaols are metabolized extensively in mouse and human to form thiol-conjugated metabolites and GSH might play an important role in the cancer preventative activity of ginger. PMID:23322393

  19. Arabidopsis CDS blastp result: AK071633 [KOME

    Lifescience Database Archive (English)

    Full Text Available AK071633 J023102J23 At1g07080.1 gamma interferon ... responsive lysosomal thiol reductase family pro ... n / GILT family protein similar to SP|P13284 Gamma-interferon ... inducible lysosomal thiol reductase precursor {Hom ... o sapiens}; contains Pfam profile PF03227: Gamma interferon ... inducible lysosomal thiol reductase (GILT) 6e-66 ...

  20. Arabidopsis CDS blastp result: AK106050 [KOME

    Lifescience Database Archive (English)

    Full Text Available AK106050 001-206-F01 At1g07080.1 gamma interferon ... responsive lysosomal thiol reductase family pr ... n / GILT family protein similar to SP|P13284 Gamma-interferon ... inducible lysosomal thiol reductase precursor {Hom ... o sapiens}; contains Pfam profile PF03227: Gamma interferon ... inducible lysosomal thiol reductase (GILT) 6e-63 ...

  1. Studies on aldose reductase inhibitors from natural products. IV. Constituents and aldose reductase inhibitory effect of Chrysanthemum morifolium, Bixa orellana and Ipomoea batatas.

    Science.gov (United States)

    Terashima, S; Shimizu, M; Horie, S; Morita, N

    1991-12-01

    The hot water extracts of Chrysanthemum morifolium, Bixa orellana and Ipomoea batatas, were found to have potent inhibitory activity towards lens aldose reductase (AR). Ellagic acid (4) was isolated from C. morifolium and I. batatas, isoscutellarein (7) from B. orellana and 3,5-dicaffeoylquinic acid (10) from I. batatas, respectively, as potent inhibitors. PMID:1814628

  2. Expression, purification, crystallization and preliminary X-ray analysis of perakine reductase, a new member of the aldo-keto reductase enzyme superfamily from higher plants

    International Nuclear Information System (INIS)

    Perakine reductase, a novel member of the aldo-keto reductase enzyme superfamily of higher plants, is involved in the biosynthesis of monoterpenoid indole alkaloids in the Indian medicinal plant Rauvolfia serpentina. The enzyme has been crystallized in C-centered orthorhombic space group and diffracts to 2.0 resolution. Perakine reductase (PR) is a novel member of the aldo-keto reductase enzyme superfamily from higher plants. PR from the plant Rauvolfia serpentina is involved in the biosynthesis of monoterpenoid indole alkaloids by performing NADPH-dependent reduction of perakine, yielding raucaffrinoline. However, PR can also reduce cinnamic aldehyde and some of its derivatives. After heterologous expression of a triple mutant of PR in Escherichia coli, crystals of the purified and methylated enzyme were obtained by the hanging-drop vapour-diffusion technique at 293 K with 100 mM sodium citrate pH 5.6 and 27% PEG 4000 as precipitant. Crystals belong to space group C2221 and diffract to 2.0 , with unit-cell parameters a = 58.9, b = 93.0, c = 143.4

  3. Genetic diversity of benzoyl coenzyme A reductase genes detected in denitrifying isolates and estuarine sediment communities.

    Science.gov (United States)

    Song, Bongkeun; Ward, Bess B

    2005-04-01

    Benzoyl coenzyme A (benzoyl-CoA) reductase is a central enzyme in the anaerobic degradation of organic carbon, which utilizes a common intermediate (benzoyl-CoA) in the metabolism of many aromatic compounds. The diversity of benzoyl-CoA reductase genes in denitrifying bacterial isolates capable of degrading aromatic compounds and in river and estuarine sediment samples from the Arthur Kill in New Jersey and the Chesapeake Bay in Maryland was investigated. Degenerate primers were developed from the known benzoyl-CoA reductase genes from Thauera aromatica, Rhodopseudomonas palustris, and Azoarcus evansii. PCR amplification detected benzoyl-CoA reductase genes in the denitrifying isolates belonging to alpha-, beta-, or gamma-Proteobacteria as well as in the sediment samples. Phylogenetic analysis, sequence similarity comparison, and conserved indel determination grouped the new sequences into either the bcr type (found in T. aromatica and R. palustris) or the bzd type (found in A. evansii). All the Thauera strains and the isolates from the genera Acidovorax, Bradyrhizobium, Paracoccus, Ensifer, and Pseudomonas had bcr-type benzoyl-CoA reductases with amino acid sequence similarities of more than 97%. The genes detected from Azarocus strains were assigned to the bzd type. A total of 50 environmental clones were detected from denitrifying consortium and sediment samples, and 28 clones were assigned to either the bcr or the bzd type of benzoyl-CoA reductase genes. Thus, we could determine the genetic capabilities for anaerobic degradation of aromatic compounds in sediment communities of the Chesapeake Bay and the Arthur Kill on the basis of the detection of two types of benzoyl-CoA reductase genes. The detected genes have future applications as genetic markers to monitor aromatic compound degradation in natural and engineered ecosystems. PMID:15812036

  4. Adsorption structure of trithiophene-thiols self-assembled on Au(111) studied by NEXAFS and STM

    International Nuclear Information System (INIS)

    Full text: The structure of aromatic thiols adsorbed on Au(111) is derived from a delicate balance involving ?-interaction of aromatic with metal d band and a ?- ? interaction among aromatic rings. In the present study, we combine C K-edge NEXAFS and in-air STM to investigate the orientation and adsorption structure of two trithiophene derivatives, 2,2':5',2'-terthiophene- 5-thiol (5tTP) and 2,2':5',2'-terthiophene-5-methanethiol (5mTP), adsorbed respectively on Au(111) at sub-monolayer and full monolayer coverages. The 5mTP molecule has one methylene unit inserted between the trithiophene and thiol functional group, which provides flexibility in packing adsorbate molecules as well as strengthens metal-thiolate bond. XPS results indicate that both 5tTP and 5mTP adsorb on Au(111) via a cleavage of the SH bond to form thiolate. The surface coverage of the 5tTP is judged to be 30% less than that of the 5mTP based on XPS and XAS data. The orientation of the aromatic plane of the thiophene with respect to Au surface is obtained by analyzing polarization-dependent C K-edge NEXAFS data. Result suggests that the 5mTP adsorbs in a more upright configuration than the 5tTP does. The tilt angle for aromatic plane of the 5tTP is estimated to be 30 deg away from Au surface normal, whereas aromatic plane for the 5mTP is tilted even closer to the surface normal. The sequential development of adsorption structures will be presented and discussed in conjunction with atomically-resoled in conjunction with atomically-resolved STM data

  5. Silver activates mast cells through reactive oxygen species production and a thiol-sensitive store-independent Ca2+ influx.

    Science.gov (United States)

    Yoshimaru, Tetsuro; Suzuki, Yoshihiro; Inoue, Toshio; Niide, Osamu; Ra, Chisei

    2006-06-01

    In genetically susceptible human and/or experimental animals, heavy metals such as mercury, gold, and silver have been shown to highly induce adverse immunological reactions such as allergy and autoimmunity, in which mast cell degranulation is implicated as playing a role. We previously reported that silver activates mast cells and induces Ca2+ influx without stimulating intracellular signaling events required for activation of store-operated Ca2+ channels (SOCs). The purpose of the present study was to elucidate the possible involvement of reactive oxygen species (ROS) in the biological effects of silver. Analysis using oxidant-sensitive fluorescent probes such as dichlorodihydrofluorescein and scopoletin, as well as MCLA-amplified chemiluminescence, showed that silver induced intracellular production and/or extracellular release of ROS. Silver induced mast cell degranulation in a Ca2+ -dependent manner. Unlike IgE antigen, silver-induced Ca2+ influx was not affected by depletion of internal Ca2+ stores. Instead, the metal-induced Ca2+ influx was abolished and reversed by the cell-impermeant thiol-reducing agent dithiothreitol, indicating the regulation by oxidation of vicinal thiols on the cell surface. Consistent with this view, Ca2+ influx was blocked by the glutathione peroxidase mimetic ebselen (2-phenyl-1,2-benzisoselenazol-3(2H)-one) and the superoxide dismutase mimetic manganese(III) tetrakis 4-(benzoic acid)porphyrin, but not by exogenously added catalase or superoxide dismutase. These findings indicate that silver evokes the release of ROS and oxidation of thiols critical for the activation of a Ca2+ channel other than SOC. Such a novel ROS-dependent pathway might play a role in mast cell degranulation in metal-induced allergic and autoimmune reactions. PMID:16716896

  6. A second target of benzamide riboside: dihydrofolate reductase.

    Science.gov (United States)

    Roussel, Breton; Johnson-Farley, Nadine; Kerrigan, John E; Scotto, Kathleen W; Banerjee, Debabrata; Felczak, Krzysztof; Pankiewicz, Krzysztof W; Gounder, Murugesan; Lin, HongXia; Abali, Emine Ercikan; Bertino, Joseph R

    2012-11-01

    Dihydrofolate reductase (DHFR) is an essential enzyme involved in de novo purine and thymidine biosynthesis. For several decades, selective inhibition of DHFR has proven to be a potent therapeutic approach in the treatment of various cancers including acute lymphoblastic leukemia, non-Hodgkin's lymphoma, osteogenic sarcoma, carcinoma of the breast, and head and neck cancer. Therapeutic success with DHFR inhibitor methotrexate (MTX) has been compromised in the clinic, which limits the success of MTX treatment by both acquired and intrinsic resistance mechanisms. We report that benzamide riboside (BR), via anabolism to benzamide adenine dinucleotide (BAD) known to potently inhibit inosine monophosphate dehydrogenase (IMPDH), also inhibits cell growth through a mechanism involving downregulation of DHFR protein. Evidence to support this second site of action of BR includes the finding that CCRF-CEM/R human T-cell lymphoblasic leukemia cells, resistant to MTX as a consequence of gene amplification and overexpression of DHFR, are more resistant to BR than are parental cells. Studies of the mechanism by which BR lowers DHFR showed that BR, through its metabolite BAD, reduced NADP and NADPH cellular levels by inhibiting nicotinamide adenine dinucleotide kinase (NADK). As consequence of the lack of NADPH, DHFR was shown to be destabilized. We suggest that, inhibition of NADK is a new approach to downregulate DHFR and to inhibit cell growth. PMID:22954684

  7. Kinetic mechanism of sheep liver NADPH-dependent aldehyde reductase.

    Science.gov (United States)

    De Jongh, K S; Schofield, P J; Edwards, M R

    1987-01-01

    The kinetic mechanism of the major sheep liver aldehyde reductase (ALR1) was studied with three aldehyde substrates: p-nitrobenzaldehyde, pyridine-3-aldehyde and D-glucuronate. In each case the enzyme mechanism was sequential and product-inhibition studies were consistent with an ordered Bi Bi mechanism, with the coenzymes binding to the free enzyme. Binding studies were used to investigate the interactions of substrates, products and inhibitors with the free enzyme. These provided evidence for the binding of D-glucuronate, L-gulonate and valproate, as well as NADP+ and NADPH. The enzyme was inhibited by high concentrations of D-glucuronate in a non-competitive manner, indicating that this substrate was able to bind to the free enzyme and to the E X NADP+ complex at elevated concentrations. Although the enzyme was inhibited by high pyridine-3-aldehyde concentrations, there was no evidence for the binding of this substrate to the free enzyme. Sheep liver ALR1 was inhibited by the ionized forms of alrestatin, sorbinil, valproate, 2-ethylhexanoate and phenobarbitone, indicating the presence of an anion-binding site similar to that described for the pig liver enzyme, which interacts with inhibitors and substrates containing a carboxy group. Sorbinil, valproate and 2-ethylhexanoate inhibited the enzyme uncompetitively at low concentrations and non-competitively at high concentrations, whereas phenobarbitone and alrestatin were non-competitive and uncompetitive inhibitors respectively. The significance of these results with respect to inhibitor and substrate binding is discussed. PMID:3593233

  8. Mechanistic studies of ribonucleoside triphosphate reductase from Lactobacillus leichmannii

    International Nuclear Information System (INIS)

    The mechanism of action of the adenosylcobalamin (AdoCbl)-dependent ribonucleoside triphosphate reductase (RTPR) was investigated using isotope effect and substrate specificity studies. These experiments were conducted on RTPR purified by a new method from Lactobacillus leichmannii. Isotope effect studies using [3'-3H]UTP and [3'-3H]ATP demonstrated that the 3' C-H bond of the nucleotide is cleaved in order to cleave the 2' C-OH bond. AdoCbl does not act as a direct H abstractor from the 3' position of the substrate, but instead is thought to act as a radical chain initiator to generate an amino acid radical on the enzyme. Further support for this enzyme mediated cleavage of the 3' C-H bond of the nucleotide and the novel role of AdoCbl came from studies using [3'3H]2'-chloro-2'-deoxyuridine 5'-triphosphate ([3'-3H]CIUTP). Evidence is presented that during the course of this reaction, the 3H abstracted from the 3' position of [3'-3H]CIUTP was either exchanged with the solvent or returned to the ? face of the 2' position to produce [2'3H]-2'-deoxy-3'-ketoUTP. This result demonstrates that RTPR is capable of catalyzing a rearrangement reaction. The significance of the RTPR-catalyzed rearrangement with respect to the AdoCbl-dependent enzymes which catalyze rearrangements is discussed

  9. Structural basis of stereospecific reduction by quinuclidinone reductase.

    Science.gov (United States)

    Takeshita, Daijiro; Kataoka, Michihiko; Miyakawa, Takuya; Miyazono, Ken-Ichi; Kumashiro, Shoko; Nagai, Takahiro; Urano, Nobuyuki; Uzura, Atsuko; Nagata, Koji; Shimizu, Sakayu; Tanokura, Masaru

    2014-01-01

    Chiral molecule (R)-3-quinuclidinol, a valuable compound for the production of various pharmaceuticals, is efficiently synthesized from 3-quinuclidinone by using NADPH-dependent 3-quinuclidinone reductase (RrQR) from Rhodotorula rubra. Here, we report the crystal structure of RrQR and the structure-based mutational analysis. The enzyme forms a tetramer, in which the core of each protomer exhibits the ?/? Rossmann fold and contains one molecule of NADPH, whereas the characteristic substructures of a small lobe and a variable loop are localized around the substrate-binding site. Modeling and mutation analyses of the catalytic site indicated that the hydrophobicity of two residues, I167 and F212, determines the substrate-binding orientation as well as the substrate-binding affinity. Our results revealed that the characteristic substrate-binding pocket composed of hydrophobic amino acid residues ensures substrate docking for the stereospecific reaction of RrQR in spite of its loose interaction with the substrate. PMID:24507746

  10. Inhibition of aldose reductase by Gentiana lutea extracts.

    Science.gov (United States)

    Akileshwari, Chandrasekhar; Muthenna, Puppala; Nastasijevi?, Branislav; Joksi?, Gordana; Petrash, J Mark; Reddy, Geereddy Bhanuprakash

    2012-01-01

    Accumulation of intracellular sorbitol due to increased aldose reductase (ALR2) activity has been implicated in the development of various secondary complications of diabetes. Thus, ALR2 inhibition could be an effective strategy in the prevention or delay of certain diabetic complications. Gentiana lutea grows naturally in the central and southern areas of Europe. Its roots are commonly consumed as a beverage in some European countries and are also known to have medicinal properties. The water, ethanol, methanol, and ether extracts of the roots of G. lutea were subjected to in vitro bioassay to evaluate their inhibitory activity on the ALR2. While the ether and methanol extracts showed greater inhibitory activities against both rat lens and human ALR2, the water and ethanol extracts showed moderate inhibitory activities. Moreover, the ether and methanol extracts of G. lutea roots significantly and dose-dependently inhibited sorbitol accumulation in human erythrocytes under high glucose conditions. Molecular docking studies with the constituents commonly present in the roots of G. lutea indicate that a secoiridoid glycoside, amarogentin, may be a potential inhibitor of ALR2. This is the first paper that shows G. lutea extracts exhibit inhibitory activity towards ALR2 and these results suggest that Gentiana or its constituents might be useful to prevent or treat diabetic complications. PMID:22844269

  11. B-factor Analysis and Conformational Rearrangement of Aldose Reductase.

    Science.gov (United States)

    Balendiran, Ganesaratnam K; Pandian, J Rajendran; Drake, Evin; Vinayak, Anubhav; Verma, Malkhey; Cascio, Duilio

    2014-01-01

    The NADPH-dependent reduction of glucose reaction that is catalyzed by Aldose Reductase (AR) follows a sequential ordered kinetic mechanism in which the co-factor NADPH binds to the enzyme prior to the aldehyde substrate. The kinetic/structural experiments have found a conformational change involving a hinge-like movement of a surface loop (residues 213-224) which is anticipated to take place upon the binding of the diphosphate moiety of NADPH. The reorientation of this loop, expected to permit the release of NADP(+), represents the rate-limiting step of the catalytic mechanism. This study reveals: 1) The Translation/Libration/Screw (TLS) analysis of absolute B-factors of apo AR crystal structures indicates that the 212-224 loop might move as a rigid group. 2) Residues that make the flexible loop slide in the AR binary and ternary complexes. 3) The normalized B-factors separate this segment into three different clusters with fewer residues. PMID:25364319

  12. Determination of Nitrate Reductase Assay Depending on the Microbial Growth

    International Nuclear Information System (INIS)

    A rapid micro-dilution assay for determination of the antimicrobial susceptibility of different bacterial isolates was developed. This assay is based on the ability of the most of viable organisms to reduce nitrates. The MIC or MBC could be determined by nitrate reductase (NR) only after 30 to 90 min of incubation depending on the behaviour of microbial growth. Bacterial viability is detected by a positive nitrite reduction rather than visible turbidity. The nitrate reduction assay was compared with standard micro-assay using 250 isolates of different taxa against 10 antibiotics belonging to different classes. An excellent agreement of 82.5 % was found between the two methods and only 17.5 % of 1794 trials showed difference in the determined MIC by tow-dilution interval above or below the MIC determined by the turbidimetric method under the same test conditions. However, the nitrate reduction assay was more rapid and sensitive in detecting viable bacteria and so, established an accurate estimate of the minimal inhibitory concentration (MIC) or the minimal bacterial concentration (MBC). The nitrate reduction assay offers the additional advantage that it could be used to determine the MBC without having to subculture the broth. 232 cases of resistance were detected by NR and 4 different media were tested for susceptibility test. The bacterial isolates were exposed to ultra violet (UV) light for different period

  13. ADP-ribosylation of dinitrogenase reductase in Rhodobacter capsulatus

    International Nuclear Information System (INIS)

    In the photosynthetic bacterium Rhodobacter capsulatus, nitrogenase is regulated by a reversible covalent modification of Fe protein or dinitrogenase reductase (Rc2). The linkage of the modifying group to inactive Rc2 was found to be sensitive to alkali and to neutral hydroxylamine. Complete release of the modifying group was achieved by incubation of inactive Rc2 in 0.4 or 1 M hydroxylamine. After hydroxylamine treatment of the Rc2 preparation, the modifying group could be isolated and purified by affinity chromatography and ion-exchange HPLC. The modifying group comigrated with ADP-ribose on both ion-exchange HPLC and thin-layer chromatography. Analyses by 31P NMR spectroscopy and mass spectrometry provided further evidence that the modifying group was ADP-ribose. The NMR spectrum of inactive Rc2 exhibited signals characteristic of ADP-ribose; integration of these signals allowed calculation of a molar ration ADP-ribose/Rc2 of 0.63. A hexapeptide carrying the ADP-ribose moiety was purified from a subtilisin digest of inactive Rc2. The structure of this peptide, determined by amino acid analysis and sequencing, is Gly-Arg(ADP-ribose)-Gly-Val-Ile-Thr. This structure allows identification of the binding site for ADP-ribose as Arg 101 of the polypeptide chain of Rc2. It is concluded that nitrogenase activity in R. capsulatus is regulated by reversible ADP-ribosylation of a specific arginyl residue of dinitrogenase reductaseuctase

  14. Binding of Natural and Synthetic Polyphenols to Human Dihydrofolate Reductase

    Directory of Open Access Journals (Sweden)

    Jos Neptuno Rodrguez-Lpez

    2009-12-01

    Full Text Available Dihydrofolate reductase (DHFR is the subject of intensive investigation since it appears to be the primary target enzyme for antifolate drugs. Fluorescence quenching experiments show that the ester bond-containing tea polyphenols (--epigallocatechin gallate (EGCG and (--epicatechin gallate (ECG are potent inhibitors of DHFR with dissociation constants (KD of 0.9 and 1.8 ?M, respectively, while polyphenols lacking the ester bound gallate moiety [e.g., (--epigallocatechin (EGC and (--epicatechin (EC] did not bind to this enzyme. To avoid stability and bioavailability problems associated with tea catechins we synthesized a methylated derivative of ECG (3-O-(3,4,5-trimethoxybenzoyl-(--epicatechin; TMECG, which effectively binds to DHFR (KD = 2.1 ?M. In alkaline solution, TMECG generates a stable quinone methide product that strongly binds to the enzyme with a KD of 8.2 nM. Quercetin glucuronides also bind to DHFR but its effective binding was highly dependent of the sugar residue, with quercetin-3-xyloside being the stronger inhibitor of the enzyme with a KD of 0.6 ?M. The finding that natural polyphenols are good inhibitors of human DHFR could explain the epidemiological data on their prophylactic effects for certain forms of cancer and open a possibility for the use of natural and synthetic polyphenols in cancer chemotherapy.

  15. Aldo-keto reductases and bioactivation/detoxication.

    Science.gov (United States)

    Jin, Yi; Penning, Trevor M

    2007-01-01

    Aldo-keto reductases (AKRs) are soluble NAD(P)(H) oxidoreductases that primarily reduce aldehydes and ketones to primary and secondary alcohols, respectively. The ten known human AKR enzymes can turnover a vast range of substrates, including drugs, carcinogens, and reactive aldehydes. They play central roles in the metabolism of these agents, and this can lead to either their bioactivation or detoxication. AKRs are Phase I drug metabolizing enzymes for a variety of carbonyl-containing drugs and are implicated in cancer chemotherapeutic drug resistance. They are involved in tobacco-carcinogenesis because they activate polycyclic aromatic trans-dihydrodiols to yield reactive and redox active o-quinones, but they also catalyze the detoxication of nicotine derived nitrosamino ketones. They also detoxify reactive aldehydes formed from exogenous toxicants, e.g., aflatoxin, endogenous toxicants, and those formed from the breakdown of lipid peroxides. AKRs are stress-regulated genes and play a central role in the cellular response to osmotic, electrophilic, and oxidative stress. PMID:16970545

  16. Benzamidomethylation with (Benzamidomethyl)triethylammonium Chloride. 2. A Simple Method for Benzamidomethylation of Thiols, Amines and Carboxylic Acids

    OpenAIRE

    Emil Popovski; Drazen Vikic-Topic; Ljiljana Klisarova

    2000-01-01

    Thiols and amines were benzamidomethylated in water solution at room temperature with (benzamidomethyl)triethylammonium chloride (1) in the presence of a small quantity of triethylamine (pH>9). Benzamidomethyl thioethers (3a-d) and (benzamidomethyl) amines or di(benzamidomethyl)amines (5) were obtained in high yields (>90%) as well as S(CH2NHBz)2 in a reaction of 1 with Na2S. Benzamidomethyl esters RCOOCH2NHBz were obtained (60-75%) in reactions of carboxylic acids with 1 in chloroform or dio...

  17. Adsorption Kinetics of 4-Amino-5-Phenyl-4H-1, 2, 4-Triazole-3-Thiol on Mild Steel Surface

    Directory of Open Access Journals (Sweden)

    Anees A. Khadom

    2010-01-01

    Full Text Available The adsorption of 4-amino-5-phenyl-4H-1, 2, 4-triazole-3-thiol (APTT as a corrosion inhibitor on mild steel surface in hydrochloric acid (HCl solution was studied using the weight loss technique. The surface coverage with the adsorbed APTT was used to calculate the free energy of adsorption, ?Gads, of APTT using Bockris-Swinkels isotherm. The dependence of free energy of adsorption, ?Gads, on the surface coverage, ?, is ascribed to the surface heterogeneity of the adsorbent. The effect of APTT was discussed from the adsorption model viewpoint. The adsorption of APTT molecules on the surface occurs without modifying the kinetic of corrosion process.

  18. Adsorption Kinetics of 4-Amino-5-Phenyl-4H-1, 2, 4-Triazole-3-Thiol on Mild Steel Surface

    Scientific Electronic Library Online (English)

    Anees A., Khadom; Ahmed Y., Musa; Abdul Amir H., Kadhum; Abu Bakar, Mohamad; Mohd Sobri, Takriff.

    Full Text Available The adsorption of 4-amino-5-phenyl-4H-1, 2, 4-triazole-3-thiol (APTT) as a corrosion inhibitor on mild steel surface in hydrochloric acid (HCl) solution was studied using the weight loss technique. The surface coverage with the adsorbed APTT was used to calculate the free energy of adsorption, ?Gad [...] s, of APTT using Bockris-Swinkels isotherm. The dependence of free energy of adsorption, ?Gads, on the surface coverage, ?, is ascribed to the surface heterogeneity of the adsorbent. The effect of APTT was discussed from the adsorption model viewpoint. The adsorption of APTT molecules on the surface occurs without modifying the kinetic of corrosion process.

  19. In situ preparation of fluorescent CdTe quantum dots with small thiols and hyperbranched polymers as co-stabilizers

    OpenAIRE

    Shi, Yunfeng; Ma, Zhimin; Cui, Ningning; Liu, Yanli; Hou, Xiaoyu; Du, Weimin; Liu, Lin; Gangsheng, Tong

    2014-01-01

    A new strategy for in situ preparation of highly fluorescent CdTe quantum dots (QDs) with 3-mercaptopropionic acid (MPA) and hyperbranched poly(amidoamine)s (HPAMAM) as co-stabilizers was proposed in this paper. MPA and HPAMAM were added in turn to coordinate Cd2+. After adding NaHTe and further microwave irradiation, fluorescent CdTe QDs stabilized by MPA and HPAMAM were obtained. Such a strategy avoids the aftertreatment of thiol-stabilized QDs in their bioapplication and provides an opport...

  20. Electronic structure, magnetic properties, and microstructural analysis of thiol-functionalized Au nanoparticles: role of chemical and structural parameters in the ferromagnetic behaviour

    Energy Technology Data Exchange (ETDEWEB)

    Guerrero, Estefania; Munoz-Marquez, Miguel A., E-mail: miguel.angel@icmse.csic.e [Instituto de Ciencia de Materiales de Sevilla (CSIC-US) (Spain); Fernandez-Pinel, Enrique; Crespo, Patricia; Hernando, Antonio [Instituto de Magnetismo Aplicado (UCM-ADIF-CSIC) (Spain); Fernandez, Asuncion [Instituto de Ciencia de Materiales de Sevilla (CSIC-US) (Spain)

    2008-12-15

    Gold nanoparticles (NPs) have been stabilized with a variety of thiol-containing molecules in order to change their chemical and physical properties; among the possible capping systems, alkane chains with different lengths, a carboxylic acid and a thiol-containing biomolecule (tiopronin) have been selected as protecting shells for the synthesized NPs; the NPs solubility in water or organic solvents is determined by the protecting molecule. A full microstructural characterization of these NPs is presented in the current research work. It has been shown that NPs capped with alkanethiol chains have a marked ferromagnetic behaviour which might also be dependent on the chain length. The simultaneous presence of Au-Au and Au-S bonds together with a reduced particle diameter, and the formation of an ordered monolayer protective shell, have proved to be key parameters for the ferromagnetic-like behaviour exhibited by thiol-functionalized gold NPs.