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Sample records for arabinoxylan feruloyl transferase

  1. Quantitative Profiling of Feruloylated Arabinoxylan Side-Chains from Graminaceous Cell Walls

    Science.gov (United States)

    Schendel, Rachel R.; Meyer, Marleen R.; Bunzel, Mirko

    2016-01-01

    Graminaceous arabinoxylans are distinguished by decoration with feruloylated monosaccharidic and oligosaccharidic side-chains. Although it is hypothesized that structural complexity and abundance of these feruloylated arabinoxylan side-chains may contribute, among other factors, to resistance of plant cell walls to enzymatic degradation, quantitative profiling approaches for these structural units in plant cell wall materials have not been described yet. Here we report the development and application of a rapid and robust method enabling the quantitative comparison of feruloylated side-chain profiles in cell wall materials following mildly acidic hydrolysis, C18-solid phase extraction (SPE), reduction under aprotic conditions, and liquid chromatography with diode-array detection/mass spectrometry (LC-DAD/MS) separation and detection. The method was applied to the insoluble fiber/cell wall materials isolated from 12 whole grains: wild rice (Zizania aquatica L.), long-grain brown rice (Oryza sativa L.), rye (Secale cereale L.), kamut (Triticum turanicum Jakubz.), wheat (Triticum aestivum L.), spelt (Triticum spelta L.), intermediate wheatgrass (Thinopyrum intermedium), maize (Zea mays L.), popcorn (Zea mays L. var. everta), oat (Avena sativa L.) (dehulled), barley (Hordeum vulgare L.) (dehulled), and proso millet (Panicum miliaceum L.). Between 51 and 96% of the total esterified monomeric ferulates were represented in the quantified compounds captured in the feruloylated side-chain profiles, which confirms the significance of these structures to the global arabinoxylan structure in terms of quantity. The method provided new structural insights into cereal grain arabinoxylans, in particular, that the structural moiety α-l-galactopyranosyl-(1→2)-β-d-xylopyranosyl-(1→2)-5-O-trans-feruloyl-l-arabinofuranose (FAXG), which had previously only been described in maize, is ubiquitous to cereal grains. PMID:26834763

  2. Inhibition of Intestinal α-Glucosidase and Glucose Absorption by Feruloylated Arabinoxylan Mono- and Oligosaccharides from Corn Bran and Wheat Aleurone

    Science.gov (United States)

    Malunga, Lovemore Nkhata; Eck, Peter; Beta, Trust

    2016-01-01

    The effect of feruloylated arabinoxylan mono- and oligosaccharides (FAXmo) on mammalian α-glucosidase and glucose transporters was investigated using human Caco-2 cells, rat intestinal acetone powder, and Xenopus laevis oocytes. The isolated FAXmo from wheat aleurone and corn bran were identified to have degree of polymerization (DP) of 4 and 1, respectively, by HPLC-MS. Both FAXmo extracts were effective inhibitors of sucrase and maltase functions of the α-glucosidase. The IC50 for FAXmo extracts on Caco-2 cells and rat intestinal α-glucosidase was 1.03–1.65 mg/mL and 2.6–6.5 mg/mL, respectively. Similarly, glucose uptake in Caco-2 cells was inhibited up to 40%. The inhibitory effect of FAXmo was dependent on their ferulic acid (FA) content (R = 0.95). Sodium independent glucose transporter 2 (GLUT2) activity was completely inhibited by FAXmo in oocytes injected to express GLUT2. Our results suggest that ferulic acid and feruloylated arabinoxylan mono-/oligosaccharides have potential for use in diabetes management. PMID:27073693

  3. RNA interference suppression of genes in glycosyl transferase families 43 and 47 in wheat starchy endosperm causes large decreases in arabinoxylan content

    OpenAIRE

    Lovegrove, Alison; Wilkinson, Mark D; Freeman, Jackie; Pellny, Till K.; Tosi, Paola; Saulnier, Luc; Shewry, Peter R.; Mitchell, Rowan A. C.

    2013-01-01

    The cell walls of wheat (Triticum aestivum) starchy endosperm are dominated by arabinoxylan (AX), accounting for 65% to 70% of the polysaccharide content. Genes within two glycosyl transferase (GT) families, GT43 (IRREGULAR XYLEM9 [IRX9] and IRX14) and GT47 (IRX10), have previously been shown to be involved in the synthesis of the xylan backbone in Arabidopsis, and close homologs of these have been implicated in the synthesis of xylan in other species. Here, homologs of IRX10 TaGT47_2 and IRX...

  4. Enzymatic hydrolysis of corn bran arabinoxylan

    DEFF Research Database (Denmark)

    Agger, Jane

    This thesis concerns enzymatic hydrolysis of corn bran arabinoxylan. The work has focused on understanding the composition and structure of corn bran with specific interest in arabinoxylan with the main purpose of targeting enzymatic hydrolysis for increased yields. Corn bran has been used as a...... model substrate because it represents a readily available agroindustrial side product with upgrading potentials. Corn bran originates from the wet-milling process in corn starch processing, is the outmost layers of the corn kernel and is particularly rich in pentose monosaccharides comprising the major...... components of arabinoxylan. Corn bran is one of the most recalcitrant cereal byproducts with arabinoxylans of particular heterogeneous nature. It is also rich in feruloyl derived substitutions, which are responsible for extensive cross-linking between arabinoxylan molecules and thereby participate in a...

  5. The potato suberin feruloyl transferase FHT which accumulates in the phellogen is induced by wounding and regulated by abscisic and salicylic acids.

    Science.gov (United States)

    Boher, Pau; Serra, Olga; Soler, Marçal; Molinas, Marisa; Figueras, Mercè

    2013-08-01

    The present study provides new insights on the role of the potato (Solanum tuberosum) suberin feruloyl transferase FHT in native and wound tissues, leading to conclusions about hitherto unknown properties of the phellogen. In agreement with the enzymatic role of FHT, it is shown that its transcriptional activation and protein accumulation are specific to tissues that undergo suberization such as the root boundary layers of the exodermis and the endodermis, along with the tuber periderm. Remarkably, FHT expression and protein accumulation within the periderm is restricted to the phellogen derivative cells with phellem identity. FHT levels in the periderm are at their peak near harvest during periderm maturation, with the phellogen becoming meristematically inactive and declining thereafter. However, periderm FHT levels remain high for several months after harvest, suggesting that the inactive phellogen retains the capacity to synthesize ferulate esters. Tissue wounding induces FHT expression and the protein accumulates from the first stages of the healing process onwards. FHT is up-regulated by abscisic acid and down-regulated by salicylic acid, emphasizing the complex regulation of suberin synthesis and wound healing. These findings open up new prospects important for the clarification of the suberization process and yield important information with regard to the skin quality of potatoes. PMID:23918964

  6. Dietary fibers - purification, structure and their health benefits with particular reference to feruloyl arabinoxylans

    DEFF Research Database (Denmark)

    Rao, Shyama Prasad; Muralikrishna, G

    2011-01-01

    -digestible carbohydrates have been collectively referred to as ‘dietary fibre’ (Hipsley, 1953). Some of these carbohydrates are of particular interest to the food industry for the purpose of developing ‘functional foods’, i.e., foods that are able to exert positive health effects. Non-digestible oligo/polysaccharides are......The nature of carbohydrates present in the food is growing field of interest within the food industry due to the potential of some of them to help prevent diseases of lifestyle. Non-glycemic carbohydrates, i.e., those carbohydrates (or their components) that are not absorbed in the small intestine...... and, therefore, transit down to become fermented in the colon, have drawn lot of attention. In fact, food carbohydrates can be broadly classified on the basis of their in vivo digestibility into digestible and non-digestible carbohydrates (Table1) (Asp, 1996; Englyst et al., 1992). Non...

  7. Structural Characterisation by ESI-MS of Feruloylated Arabino-oligosaccharides Synthesised by Chemoenzymatic Esterification

    Directory of Open Access Journals (Sweden)

    Paul Christakopoulos

    2007-07-01

    Full Text Available The chemoenzymatic synthesis of feruloylated arabino-oligosaccharides has been achieved, using a feruloyl esterase type C from Sporotrichum thermophile (StFaeC.The structure of the feruloylated products was confirmed by ESI-MSn.

  8. Arabinoxylans, gut microbiota and immunity.

    Science.gov (United States)

    Mendis, Mihiri; Leclerc, Estelle; Simsek, Senay

    2016-03-30

    Arabinoxylan (AX) is a non-starch polysaccharide found in many cereal grains and is considered as a dietary fiber. Despite their general structure, there is structural heterogeneity among AX originating from different botanical sources. Furthermore, the extraction procedure and hydrolysis by xylolytic enzymes can further render differences to theses AX. The aim of this review was to address the effects of AX on the gut bacteria and their immunomodulatory properties. Given the complex structure of AX, we also aimed to discuss how the structural heterogeneity of AX affects its role in bacterial growth and immunomodulation. The existing literature indicates the role of fine structural details of AX on its potential as polysaccharides that can impact the gut associated microbial growth and immune system. PMID:26794959

  9. Maize Arabinoxylan Gels as Protein Delivery Matrices

    Directory of Open Access Journals (Sweden)

    Ana Luisa Martínez-López

    2009-04-01

    Full Text Available The laccase induced gelation of maize bran arabinoxylans at 2.5% (w/v in the presence of insulin or β-lactoglobulin at 0.1% (w/v was investigated. Insulin and β-lacto-globulin did not modify either the gel elasticity (9 Pa or the cross-links content (0.03 and 0.015 mg di- and triferulic acids/mg arabinoxylan, respectively. The protein release capability of the gel was also investigated. The rate of protein release from gels was dependent on the protein molecular weight. The apparent diffusion coefficient was 0.99 × 10-7 and 0.79 × 10-7 cm2/s for insulin (5 kDa and β-lactoglobulin (18 kDa, respectively. The results suggest that maize bran arabinoxylan gels can be potential candidates for the controlled release of proteins.

  10. Pilot-scale bioreactor production and long term stability of feruloyl soy glycerides

    Science.gov (United States)

    Soybean oil was transesterified with ethyl ferulate at 60 °C using the immobilized lipase Candida antartica lipase B (Novozym 435) to produce a mixture of feruloylated monoacylglycerols and feruloylated diacylglycerols homologues, collectively referred to as feruloylated soy glycerides (FSG). A 1-to...

  11. Concentration and shear rate dependence of solution viscosity for arabinoxylans from different sources

    Science.gov (United States)

    Arabinoxylans are cell wall polysaccharides abundant in plants. Alkaline extraction is commonly used to isolate arabinoxylans from cell wall rich materials, such as cereal brans, crop residues etc. While arabinoxylans from certain sources such as wheat endosperm, corn bran and rye bran have been wid...

  12. Arabinoxylan Microspheres: Structural and Textural Characteristics

    Directory of Open Access Journals (Sweden)

    Yolanda López-Franco

    2013-04-01

    Full Text Available The aim of this research was to study the structural and textural characteristics of maize bran arabinoxylan (MBAX microspheres. The laccase-induced cross-linking process was monitored by storage (G' and loss (G'' moduli changes in a 4% (w/v MBAX solution. The G' and G'' values at the plateau region were 215 and 4 Pa, respectively. After gelation, the content of ferulic acid dimers decreased from 0.135 to 0.03 µg/mg MBAX, suggesting the formation of ferulated structures unreleased by mild alkaline hydrolysis. MBAX microspheres presented an average diameter of 531 µm and a swelling ratio value (q of 18 g water/g MBAX. The structural parameters of MBAX microspheres were calculated from equilibrium swelling experiments, presenting an average mesh size of 52 nm. Microstructure and textural properties of dried MBAX microspheres were studied by scanning electron microscopy and nitrogen adsorption/desorption isotherms, respectively, showing a heterogeneous mesoporous and macroporous structure throughout the network.

  13. Both wheat (Triticum aestivum) bran arabinoxylans and gut flora-mediated fermentation products protect human colon cells from genotoxic activities of 4-hydroxynonenal and hydrogen peroxide.

    Science.gov (United States)

    Glei, Michael; Hofmann, Thomas; Küster, Katrin; Hollmann, Jürgen; Lindhauer, Meinolf G; Pool-Zobel, Beatrice L

    2006-03-22

    Dietary fibers are fermented by the gut flora to yield short chain fatty acids (SCFAs), which inhibit the growth of tumor cells, induce glutathione S-transferases (GSTs), and protect cells from the genotoxic activity of 4-hydroxynonenal (HNE). Here, we investigated effects of wheat bran-derived arabinoxylans and fermentation products on these parameters of chemoprevention. Newly isolated water extractable (WeAx) and alkali extractable arabinoxylans (AeAx) were fermented under anaerobic conditions with human feces. Resulting fermentation supernatants (FSs) were analyzed for SCFAs and used to treat HT29 colon cancer cells. Cell growth, cytotoxicity, antigenotoxicity against hydrogen peroxide (H2O2) or HNE, and GST activity were determined. Nonfermented WeAx decreased H2O2-induced DNA damage by 64%, thus demonstrating chemoprotective properties by this nonfermented wheat bran fiber. The fermentation of WeAx and AeAx resulted in 3-fold increases of SCFA, but all FSs (including the control without arabinoxylans) inhibited the growth of the HT29 cells, reduced the genotoxicity of HNE, and enhanced the activity of GSTs (FS WeAx, 2-fold; FS AeAx, 1.7-fold; and control FS, 1.4-fold), which detoxify HNE. Thus, increases in SCFAs were not reflected by enhanced functional effects. The conclusion is that fermentation mixtures contain modulatory compounds that arise from the feces and might add to the effectiveness of SCFAs. PMID:16536580

  14. Arabinoxylan content and characterisation throughout the bread-baking process

    Science.gov (United States)

    End-use quality of wheat (Triticum aestivum L.) is influenced in a variety of ways by non-starch polysaccharides, especially arabinoxylans (AX). The assessment of AX content and structural properties is often performed on flour and extrapolated to predict the role that AX may play in baked products....

  15. In vitro degradation of covalently cross-linked arabinoxylan hydrogels by bifidobacteria.

    Science.gov (United States)

    Martínez-López, Ana L; Carvajal-Millan, Elizabeth; Micard, Valérie; Rascón-Chu, Agustín; Brown-Bojorquez, Francisco; Sotelo-Cruz, Norberto; López-Franco, Yolanda L; Lizardi-Mendoza, Jaime

    2016-06-25

    Arabinoxylan gels with different cross-linking densities, swelling ratios, and rheological properties were obtained by increasing the concentration of arabinoxylan from 4 to 6% (w/v) during oxidative gelation by laccase. The degradation of these covalently cross-linked gels by a mixture of two Bifidobacterium strains (Bifidobacterium longum and Bifidobacterium adolescentis) was investigated. The kinetics of the evolution of structural morphology of the arabinoxylan gel, the carbohydrate utilization profiles and the bacterial production of short-acid fatty acid (SCFA) were measured. Scanning electron microscopy analysis of the degraded gels showed multiple cavity structures resulting from the bacterial action. The total SCFA decreased when the degree of cross-linking increased in the gels. A slower fermentation of arabinoxylan chains was obtained for arabinoxylan gels with more dense network structures. These results suggest that the differences in the structural features and properties studied in this work affect the degradation time of the arabinoxylan gels. PMID:27083795

  16. POD promoted oxidative gelation of water-extractable arabinoxylan through ferulic acid dimers. Evidence for its negative effect on malt filterability.

    Science.gov (United States)

    Wu, Dianhui; Zhou, Ting; Li, Xiaomin; Cai, Guolin; Lu, Jian

    2016-04-15

    As a major component of non-starch polysaccharide in barley, arabinoxylan (AX) plays an important role in quality traits of malt and the final beer product. The Chinese barley malt has encountered filterability problems for a long time. The main reason caused by barley cultivar has been accepted in the malting and brewing industries. In our previous proteomic study, the peroxidase (POD) BP1 was found to be in quite high abundant in the filterability defect Chinese barley malt. Therefore, the present study tried to verify its negative effect on filterability, by surveying its activity in different malt samples and detecting effects of POD on AX gelation and filterability. The results showed that the activity of POD, as well as the content of AX bounded ferulic acid, were both negatively correlated with filterability, while the feruloyl esterase activity was positively correlated with it. In addition, AX gelation catalyzed by POD caused worse filterability, and the natural inhibitor of POD, vitamin C, could blocked the cross linking catalyzed by POD and thus improve the filterability. These results all suggested the great negative effect of POD on malt filterability. PMID:26616970

  17. Common and Distant Structural Characteristics of Feruloyl Esterase Families from Aspergillus oryzae

    DEFF Research Database (Denmark)

    Udatha, D. B. R. K. Gupta; Mapelli, Valeria; Panagiotou, Gianni;

    2012-01-01

    Background: Feruloyl esterases (FAEs) are important biomass degrading accessory enzymes due to their capability of cleaving the ester links between hemicellulose and pectin to aromatic compounds of lignin, thus enhancing the accessibility of plant tissues to cellulolytic and hemicellulolytic enzy...

  18. Transferases in Polymer Chemistry

    NARCIS (Netherlands)

    van der Vlist, Jeroen; Loos, Katja; Palmans, ARA; Heise, A

    2010-01-01

    Transferases are enzymes that catalyze reactions in which a group is transferred from one compound to another. This makes these enzymes ideal catalysts for polymerization reactions. In nature, transferases are responsible for the synthesis of many important natural macromolecules. In synthetic polym

  19. Purification and Characterization of a Feruloyl Esterase from the Intestinal Bacterium Lactobacillus acidophilus

    OpenAIRE

    Wang, Xiaokun; Geng, Xin; Egashira, Yukari; Sanada, Hiroo

    2004-01-01

    Dietary ferulic acid (FA), a significant antioxidant substance, is currently the subject of extensive research. FA in cereals exists mainly as feruloylated sugar ester. To release FA from food matrices, it is necessary to cleave ester cross-linking by feruloyl esterase (FAE) (hydroxycinnamoyl esterase; EC 3.1.1.73). In the present study, the FAE from a human typical intestinal bacterium, Lactobacillus acidophilus, was isolated, purified, and characterized for the first time. The enzyme was pu...

  20. Physicochemical characterization and evaluation of suspending properties of arabinoxylan from Ispaghula (Plantago ovata) husk.

    Science.gov (United States)

    Bashir, Sajid; Erum, Alia; Saghir, Shazia; Tulain, Umme Ruqia; Rashid, Ayesha

    2014-11-01

    The purpose of this study is to evaluate the use of arabinoxylan as potential suspending agent, an effective alternative to commercially used excipients for the preparation of pharmaceutical suspensions. Alkali extraction was done to separate arabinoxylan from ispaghula (Plantago ovata) seed husk by alkali extraction its physicochemical characterization was done and the suspending properties of arabinoxylan isolated were evaluated comparatively with those of bentonite at different concentration ranges of 0.125,0.25,0.5 and 1% in Zinc oxide suspension. The parameters employed for evaluation were sedimentation volume, degree of flocculation, flow rate, density, pH, redispersibility, microbiological evaluation and particle size analysis. Physicochemical characterization of arabinoxylan indicates its suitability as excipient as it has fair flow properties, low moisture content and almost neutral pH. Arabinoxylan at low conc. 0.125% showed sedimentation volume comparable to commercially used suspending agents such as bentonite 1% while suspensions containing higher concentrations such as 0.25% (sedimentation volume 92%), 0.5% (sedimentation volume 94%) and 1% conc. (sedimentation volume 98%) of arabinoxylan remained almost completely suspended during study period of 7 days. Formulations containing 0.125% and 0.25% arabinoxylan as suspending agents are easily redispersible as compared to bentonite containing formulation while formulation containing 0.5% arabinoxylan are moderately redispersible while formulation containing 1% suspending agent gel upon storage and was not redispersible. Furthermore arabinoxylan produces stable, highly flocculated suspension, which fulfilled microbiological, and particle size specifications, however the formulations containing higher arabinoxylan 1% concentration gel upon storage. So it is concluded that arabinoxylan could be used as effective suspending agent at low concentrations in Zinc oxide suspension. PMID:25362588

  1. Arabinoxylan activates Dectin-1 and modulates particulate beta-glucan-induced Dectin-1 activation

    NARCIS (Netherlands)

    Sahasrabudhe, Neha M.; Schols, Henk A.; Faas, Marijke M.; de Vos, Paul

    2016-01-01

    ScopeArabinoxylan is one of the most commonly consumed dietary fiber. Immunomodulation by arabinoxylan is documented but the mechanisms by which these immune-effects are accomplished are unknown. Methods and resultsBy applying reporter cell lines for Toll-like receptors (TLRs) and Dectin-1, we demon

  2. Impact of water extractable arabinoxylan from rye bran on the frozen steamed bread dough quality.

    Science.gov (United States)

    Wang, Pei; Tao, Han; Jin, Zhengyu; Xu, Xueming

    2016-06-01

    Impact of water extractable arabinoxylan from rye bran on frozen steamed bread dough quality was investigated in terms of the bread characteristics, ice crystallization, yeast activity as well as the gluten molecular weight distribution and glutenin macropolymer content in the present study. Results showed that water extractable arabinoxylan significantly improved bread characteristics during the 60-day frozen storage. Less water was crystallized in the water extractable arabinoxylan dough during storage, which could explain the alleviated yeast activity loss. For all the frozen dough samples, more soluble high molecular weight (Mw ≈ 91,000-688,000) and low molecular weight (Mw ≈ 91,000-16,000) proteins were derived from glutenin macropolymer depolymerization. Nevertheless, water extractable arabinoxylan dough developed higher glutenin macropolymer content with lowered level of soluble low molecular weight proteins throughout the storage. This study suggested water extractable arabinoxylan from rye bran had great potential to be served as an effective frozen steamed bread dough improver. PMID:26830568

  3. Enzymatic Glycosylation by Transferases

    DEFF Research Database (Denmark)

    Blixt, Klas Ola; Razi, Nahid

    Glycosyltransferases are important biological catalysts in cellular systems generating complex cell surface glycans involved in adhesion and signaling processes. Recent advances in glycoscience have increased the demands to access significant amount of glycans representing the glycome. Glycosyltr...... representing terminal sequences of glycoproteins and glycolipids using recombinant transferases. Transferases are also being explored in the context of solid-phase synthesis, immobilized on resins and over expression in vivo by engineered bacteria....

  4. Water Extractable Arabinoxylan Aerogels Prepared by Supercritical CO2 Drying

    Directory of Open Access Journals (Sweden)

    Agustín Rascón-Chu

    2013-05-01

    Full Text Available Water extractable arabinoxylan (WEAX aerogels were prepared by extracting the solvent from the alcogels (WEAX hydrogels with an alcohol as the solvent with carbon dioxide under supercritical conditions. WEAX aerogels were characterized using scanning electron microscopy and adsorption and desorption nitrogen isotherms. The micrographs indicate a heterogeneous porous network structure in WEAX aerogel. Adsorption/desorption nitrogen isotherms of this material were type IV, which confirm that this material possess a mesoporous structure. WEAX aerogels rehydration capability was evaluated and the water absorption mechanism was determined. The WEAX aerogels water absorption mechanism was non-Fickian (n = 0.54.

  5. Application of cross-linked and hydrolyzed arabinoxylans in baking of model rye bread.

    Science.gov (United States)

    Buksa, Krzysztof; Nowotna, Anna; Ziobro, Rafał

    2016-02-01

    The role of water extractable arabinoxylan with varying molar mass and structure (cross-linked vs. hydrolyzed) in the structure formation of rye bread was examined using a model bread. Instead of the normal flour, the dough contained starch, arabinoxylan and protein, which were isolated from rye wholemeal. It was observed that the applied mixes of these constituents result in a product closely resembling typical rye bread, even if arabinoxylan was modified (by cross-linking or hydrolysis). The levels of arabinoxylan required for bread preparation depended on its modification and mix composition. At 3% protein, the maximum applicable level of poorly soluble cross-linked arabinoxylan was 3%, as higher amounts of this preparation resulted in an extensively viscous dough and diminished bread volume. On the other hand highly soluble, hydrolyzed arabinoxylan could be used at a higher level (6%) together with larger amounts of rye protein (3% or 6%). Further addition of arabinoxylan leads to excessive water absorption, resulting in a decreased viscosity of the dough during baking and insufficient gas retention. PMID:26304439

  6. Genotypic and Environmental Variations of Arabinoxylan Content and Endoxylanase Activity in Barley Grains

    Institute of Scientific and Technical Information of China (English)

    ZHANG Xiao-qin; XUE Da-wei; WU Fei-bo; ZHANG Guo-ping

    2013-01-01

    Arabinoxylan (AX) content in barley grains is an important quality determinant when barley is used as raw material of malt or beer production. The cultivar and environmental variations of total arabinoxylan (TAX), water extractable arabinoxylan (WEAX) and endoxylanase activity (EA) were investigated using eight barley cultivars growing at seven locations with diverse environmental conditions. The results showed that both barley cultivar and location significantly affected the TAX, WEAX and EA levels, but the variations of TAX content and EA were mainly attributed to cultivar, while the impact of location on WEAX content was greater than that of cultivar. Correlation analysis indicated that TAX was significantly correlated to WUAX.

  7. Molecular Features of Wheat Endosperm Arabinoxylan Inclusion in Functional Bread

    Directory of Open Access Journals (Sweden)

    Weili Li

    2013-06-01

    Full Text Available Arabinoxylan (AX is a major dietary fibre component found in a variety of cereals. Numerous health benefits of arabinoxylans have been reported to be associated with their solubility and molecular features. The current study reports the development of a functional bread using a combination of AX-enriched material (AEM and optimal commercial endoxylanase. The total AX content of bread was increased to 8.2 g per 100 g available carbohydrates. The extractability of AX in breads with and without endoxylanase was determined. The results demonstrate that water-extractable AX (WE-AX increased progressively through the bread making process. The application of endoxylanase also increased WE-AX content. The presence of 360 ppm of endoxylanase had positive effects on the bread characteristics in terms of bread volume and firmness by converting the water unextractable (WU-AX to WE-AX. In addition, the molecular weight (Mw distribution of the WE-AX of bread with and without endoxylanase was characterized by size-exclusion chromatography. The results show that as the portion of WE-AX increased, the amount of high Mw WE-AX (higher than 100 kDa decreased, whereas the amount of low Mw WE-AX (lower than 100 kDa increased from 33.2% to 44.2% through the baking process. The low Mw WE-AX further increased to 75.5% with the application of the optimal endoxylanase (360 ppm.

  8. Expression of feruloyl esterase A from Aspergillus terreus and its application in biomass degradation.

    Science.gov (United States)

    Zhang, Shuai-Bing; Wang, Le; Liu, Yan; Zhai, Huan-Chen; Cai, Jing-Ping; Hu, Yuan-Sen

    2015-11-01

    Feruloyl esterases (FAEs) are key enzymes involved in the complete biodegradation of lignocelluloses, which could hydrolyze the ester bonds between hemicellulose and lignin. The coding sequence of a feruloyl esterase A (AtFaeA) was cloned from Aspergillus terreus and the recombinant AtFaeA was constitutively expressed in Pichia pastoris. The SDS-PAGE analysis of purified AtFaeA showed two protein bands owing to the different extent of glycosylation, and the recombinant AtFaeA had an optimum temperature of 50°C and an optimum pH of 5.0. The substrate utilization and primary sequence identity of AtFaeA demonstrated that it is a type-A feruloyl esterase. The hydrolysis of corn stalk and corncob by xylanase from Aspergillus niger could be significantly improved in concert with recombinant AfFaeA. PMID:26282562

  9. Kinetics of enzyme-catalyzed cross-linking of feruloylated arabinan from sugar beet

    DEFF Research Database (Denmark)

    Abang Zaidel, Dayang Norulfairuz; Arnous, Anis; Holck, Jesper;

    2011-01-01

    the kinetics of HRP catalyzed cross-linking of FA esterified to α-(1,5)-linked arabinans are affected by the length of the arabinan chains carrying the feruloyl substitutions. The kinetics of the HRP-catalyzed cross-linking of four sets of arabinan samples from sugar beet pulp, having different...... molecular weights and hence different degrees of polymerization, were monitored by the disappearance of FA absorbance at 316 nm. MALDI-TOF/TOF-MS analysis confirmed that the sugar beet arabinans were feruloyl-substituted, and HPLC analysis verified that the amounts of diFAs increased when FA levels...

  10. Debranching of soluble wheat arabinoxylan dramatically enhances recalcitrant binding to cellulose

    DEFF Research Database (Denmark)

    Selig, Michael J.; Thygesen, Lisbeth G.; Felby, Claus; Master, Emma R.

    2015-01-01

    The presence of xylan is a detriment to the enzymatic saccharification of cellulose in lignocelluloses. The inhibition of the processive cellobiohydrolase Cel7A by soluble wheat arabinoxylan is shown here to increase by 50 % following enzymatic treatment with a commercially-purified α-l-arabinofu......The presence of xylan is a detriment to the enzymatic saccharification of cellulose in lignocelluloses. The inhibition of the processive cellobiohydrolase Cel7A by soluble wheat arabinoxylan is shown here to increase by 50 % following enzymatic treatment with a commercially-purified α...... considerably increased the rate and rigidity of arabinoxylan mass association with cellulose. These data also suggest significant xylan–xylan adlayer formation occurs following initial binding of debranched arabinoxylan. From this, we speculate the inhibitory effects of xylan to cellulases may result from...... reduced enzymatic access via the dense association of xylan with cellulose....

  11. Effects of Wheat Bran Extract Containing Arabinoxylan Oligosaccharides on Gastrointestinal Parameters in Healthy Preadolescent Children

    NARCIS (Netherlands)

    Francois, Isabelle E. J. A.; Lescroart, Olivier; Veraverbeke, Wim S.; Marzorati, Massimo; Possemiers, Sam; Hamer, Henrike; Windey, Karen; Welling, Gjalt W.; Delcour, Jan A.; Courtin, Christophe M.; Verbeke, Kristin; Broekaert, Willem F.

    2014-01-01

    Objectives: We assessed whether wheat bran extract (WBE) containing arabinoxylan-oligosaccharides (AXOS) elicited a prebiotic effect and modulated gastrointestinal (GI) parameters in healthy preadolescent children upon consumption in a beverage. Methods: This double-blind randomized placebo-controll

  12. Structural Variation and Content of Arabinoxylans in Endosperm and Bran of Durum Wheat (Triticum turgidum L.).

    Science.gov (United States)

    Marcotuli, Ilaria; Hsieh, Yves S-Y; Lahnstein, Jelle; Yap, Kuok; Burton, Rachel Anita; Blanco, Antonio; Fincher, Geoffrey Bruce; Gadaleta, Agata

    2016-04-13

    Arabinoxylans are one group of dietary fiber components in cereal grains, and specific health benefits have been linked with their molecular fine structures and hence with physicochemical properties such as solubility in aqueous media. To characterize the fiber quality for functional foods, starchy endosperm and bran fractions from 11 durum wheat lines were analyzed for total and water-soluble arabinoxylans, (1,3;1,4)-β-glucan, and bound ferulic acid. The arabinoxylan contents ranged from 11 to 16.4% (w/w) in bran and from 1.5 to 1.8% in the starchy endosperm. Of the starchy endosperm arabinoxylans, 37% was soluble in water. No correlation was found between arabinoxylan content and bound ferulic acid in bran, although a relatively high level of this antioxidant was found in endosperm (38.3 μg/g endosperm flour). Enzymatic fingerprinting was performed to define the major fine structural features of arabinoxylans from both regions of the grain. Five major oligosaccharides released by xylanase hydrolysis were identified and characterized in the 11 durum lines. In addition, DP5, DP6, and DP7 oligosaccharides containing five, six, and seven pentosyl residues, respectively, were purified. PMID:27018210

  13. A monoclonal antibody to feruloylated (1→4)-β-D-galactan

    DEFF Research Database (Denmark)

    Clausen, Mads Hartvig; Ralet, Marie-Christine; Willats, William G. T.;

    2004-01-01

    -(trans-feruloyl)-beta-D-galactopyransoyl]-(1-->4)-D-galactopyranose (Gal(2)F). LM9 is therefore a useful antibody probe for the analysis of phenolic substitution of cell wall pectic polymers and of cell wall structure in the Amaranthaceae including sugar beet (Beta vulgaris L.) and spinach (Spinacia oleracea L.)....

  14. Overproduction of the Aspergillus niger feruloyl esterase for pulp bleaching application

    NARCIS (Netherlands)

    Record, E.; Asther, M.; Sigoillot, C.; Pagès, S.; Punt, P.J.; Delattre, M.; Haon, M.; Hondel, C.A.M.J.J. van den; Sigoillot, J.C.; Lesage-Meessen, L.; Asther, M.

    2003-01-01

    A well-known industrial fungus for enzyme production, Aspergillus niger, was selected to produce the feruloyl esterase FAEA by homologous overexpression for pulp bleaching application. The gpd gene promoter was used to drive FAEA expression. Changing the nature and concentration of the carbon source

  15. Covalently Cross-Linked Arabinoxylans Films for Debaryomyces hansenii Entrapment.

    Science.gov (United States)

    González-Estrada, Ramsés; Calderón-Santoyo, Montserrat; Carvajal-Millan, Elizabeth; Ascencio Valle, Felipe de Jesús; Ragazzo-Sánchez, Juan Arturo; Brown-Bojorquez, Francisco; Rascón-Chu, Agustín

    2015-01-01

    In the present study, wheat water extractable arabinoxylans (WEAX) were isolated and characterized, and their capability to form covalently cross-linked films in presence of Debaryomyces hansenii was evaluated. WEAX presented an arabinose to xylose ratio of 0.60, a ferulic acid and diferulic acid content of 2.1 and 0.04 µg∙mg(-1) WEAX, respectively and a Fourier Transform Infra-Red (FT-IR) spectrum typical of WEAX. The intrinsic viscosity and viscosimetric molecular weight values for WEAX were 3.6 dL∙g(-1) and 440 kDa, respectively. The gelation of WEAX (1% w/v) with and without D. hansenii (1 × 10(7) CFU∙cm(-2)) was rheologically investigated by small amplitude oscillatory shear. The entrapment of D. hansenii decreased gel elasticity from 1.4 to 0.3 Pa, probably by affecting the physical interactions between WEAX chains. Covalently cross-linked WEAX films containing D. hansenii were prepared by casting. Scanning electron microscopy images show that WEAX films containing D. hansenii were porous and consisted of granular-like and fibre microstructures. Average tensile strength, elongation at break and Young's modulus values dropped when D. hansenii was present in the film. Covalently cross-lined WEAX containing D. hansenii could be a suitable as a functional entrapping film. PMID:26102070

  16. Covalently Cross-Linked Arabinoxylans Films for Debaryomyces hansenii Entrapment

    Directory of Open Access Journals (Sweden)

    Ramsés González-Estrada

    2015-06-01

    Full Text Available In the present study, wheat water extractable arabinoxylans (WEAX were isolated and characterized, and their capability to form covalently cross-linked films in presence of Debaryomyces hansenii was evaluated. WEAX presented an arabinose to xylose ratio of 0.60, a ferulic acid and diferulic acid content of 2.1 and 0.04 µg∙mg−1 WEAX, respectively and a Fourier Transform Infra-Red (FT-IR spectrum typical of WEAX. The intrinsic viscosity and viscosimetric molecular weight values for WEAX were 3.6 dL∙g−1 and 440 kDa, respectively. The gelation of WEAX (1% w/v with and without D. hansenii (1 × 107 CFU∙cm−2 was rheologically investigated by small amplitude oscillatory shear. The entrapment of D. hansenii decreased gel elasticity from 1.4 to 0.3 Pa, probably by affecting the physical interactions between WEAX chains. Covalently cross-linked WEAX films containing D. hansenii were prepared by casting. Scanning electron microscopy images show that WEAX films containing D. hansenii were porous and consisted of granular-like and fibre microstructures. Average tensile strength, elongation at break and Young’s modulus values dropped when D. hansenii was present in the film. Covalently cross-lined WEAX containing D. hansenii could be a suitable as a functional entrapping film.

  17. Revisiting the structural features of arabinoxylans from brewers' spent grain.

    Science.gov (United States)

    Coelho, Elisabete; Rocha, M Angélica M; Moreira, Ana S P; Domingues, M Rosário M; Coimbra, Manuel A

    2016-03-30

    The brewers' spent grain (BSG) arabinoxylans (AX) have been described to be composed by a backbone of (β1→4)-linked xylose residues containing only single units of arabinose as side chains. However, this is not in accordance with the structural features of AX from other cereal sources. Aiming to disclose the possibility of additional structural details, fractions enriched in AX were obtained by sequential extraction from BSG. The AX richest fraction was hydrolysed with xylanase, fractioned by size-exclusion chromatography, and analysed by electrospray tandem mass spectrometry (ESI-MS(n)). Methylation analysis showed that the amount of terminally linked arabinose residues was not in accordance with the number of xylose branching points. This was due to the presence of O-acetyl, hexose, hexuronic acid, and methylated uronic acid residues. AXs presenting these structural features can be a potential source of a large screening prebiotic, providing, in the same molecule, areas of fast and slow probiotic fermentation rates. PMID:26794960

  18. Composite Films of Arabinoxylan and Fibrous Sepiolite: Morphological, Mechanical, and Barrier Properties

    DEFF Research Database (Denmark)

    Sárossy, Zsuzsa; Blomfeldt, J.O.; Hedenqvist, Mikael S.;

    2012-01-01

    enhance the properties of rye flour arabinoxylan. Composite films cast from arabinoxylan solutions and sepiolite suspensions in water were transparent or semitransparent at additive loadings in the 2.5−10 wt % range. Scanning electron microscopy showed that the sepiolite was well dispersed in the......Hemicelluloses represent a largely unutilized resource for future bioderived films in packaging and other applications. However, improvement of film properties is needed in order to transfer this potential into reality. In this context, sepiolite, a fibrous clay, was investigated as an additive to......(ethylene glycol) methyl ether (mPEG) plasticizer addition. Incorporation of sepiolite did not significantly influence the thermal degradation or the gas barrier properties of arabinoxylan films, which is likely a consequence of sepiolite fiber morphology. In summary, sepiolite was shown to have potential as an...

  19. Review on technological and scientific aspects of feruloyl esterases: A versatile enzyme for biorefining of biomass.

    Science.gov (United States)

    Gopalan, Nishant; Rodríguez-Duran, L V; Saucedo-Castaneda, G; Nampoothiri, K Madhavan

    2015-10-01

    With increasing focus on sustainable energy, bio-refining from lignocellulosic biomass has become a thrust area of research. With most of the works being focused on biofuels, significant efforts are also being directed towards other value added products. Feruloyl esterases (EC. 3.1.1.73) can be used as a tool for bio-refining of lignocellulosic material for the recovery and purification of ferulic acid and related hydroxycinnamic acids ubiquitously found in the plant cell wall. More and more genes coding for feruloyl esterases have been mined out from various sources to allow efficient enzymatic release of ferulic acid and allied hydroxycinnamic acids (HCAs) from plant-based biomass. A sum up on enzymatic extraction of HCAs and its recovery from less explored agro residual by-products is still a missing link and this review brushes up the achieved landmarks so far in this direction and also covers a detailed patent search on this biomass refining enzyme. PMID:26159377

  20. A new feruloyl amide derivative from the fruits of Tribulus terrestris.

    Science.gov (United States)

    Zhang, Xiaopo; Wei, Na; Huang, Jian; Tan, Yinfeng; Jin, Dejun

    2012-01-01

    A new feruloyl amide derivative, named tribulusamide C, was isolated from the fruits of Tribulus terrestris. Its structure was determined on the basis of spectroscopic analysis including IR, 1-D-, 2-D-NMR and HR-ESI-MS. The structure of tribulusamide C was characterised by a unit of pyrrolidine-2,5-dione, which distinguished it from other lignanamides previously isolated from the fruits of T. terrestris. PMID:22149942

  1. A novel feruloyl esterase from rumen microbial metagenome: Gene cloning and enzyme characterization in the release of mono- and diferulic acids

    Science.gov (United States)

    A feruloyl esterase (FAE) gene was isolated from a rumen microbial metagenome, cloned into E. coli, and expressed in active form. The enzyme (RuFae4) was classified as a Type D feruloyl esterase based on its action on synthetic substrates and ability to release diferulates. The RuFae4 alone releas...

  2. Films from spruce galactoglucomannan blended with poly(vinyl alcohol), corn arabinoxylan and konjac glucomannan

    Science.gov (United States)

    The improvement of mechanical properties of spruce galactoglucomannan (GGM)-based films was sought by blending GGM with each of poly(vinyl alcohol) (PVOH), corn arabinoxylan (cAX), and konjac glucomannan (KGM). The blend ratios were 3:1, 1:1, and 1:3(w/w), and in addition films were made from each o...

  3. Hydroxycinnamic acid bound arabinoxylans from millet brans-structural features and antioxidant activity.

    Science.gov (United States)

    Bijalwan, Vandana; Ali, Usman; Kesarwani, Atul Kumar; Yadav, Kamalendra; Mazumder, Koushik

    2016-07-01

    Hydroxycinnamic acid bound arabinoxylans (HCA-AXs) were extracted from brans of five Indian millet varieties and response surface methodology was used to optimize the extraction conditions. The optimal condition to obtain highest yield of millet HCA-AXs was determined as follows: time 61min, temperature 66°C, ratio of solvent to sample 12ml/g. Linkage analysis indicated that hydroxycinnamic acid bound arabinoxylan from kodo millet (KM-HCA-AX) contained comparatively low branched arabinoxylan consisting of 14.6% mono-substituted, 1.2% di-substituted and 41.2% un-substituted Xylp residues. The HPLC analysis of millet HCA-AXs showed significant variation in the content of three major bound hydroxycinnamic acids (caffeic, p-coumaric and ferulic acid). The antioxidant activity of millet HCA-AXs were evaluated using three in vitro assay methods (DPPH, FRAP and β-carotene linoleate emulsion assays) which suggested both phenolic acid composition and structural characteristics of arabinoxylans could be correlated to their antioxidant potential, the detailed structural analysis revealed that low substituted KM-HCA-AX exhibited relatively higher antioxidant activity compared to other medium and highly substituted HCA-AXs from finger (FM), proso (PM), barnyard (BM) and foxtail (FOXM) millet. PMID:27050114

  4. Effect of suppression of arabinoxylan synthetic genes in wheat endosperm on chain length of arabinoxylan and extract viscosity.

    Science.gov (United States)

    Freeman, Jackie; Lovegrove, Alison; Wilkinson, Mark David; Saulnier, Luc; Shewry, Peter Robert; Mitchell, Rowan Andrew Craig

    2016-01-01

    Arabinoxylan (AX) is the dominant component within wheat (Triticum aestivum L.) endosperm cell walls, accounting for 70% of the polysaccharide. The viscosity of aqueous extracts from wheat grain is a key trait influencing the processing for various end uses, and this is largely determined by the properties of endosperm AX. We have previously shown dramatic effects on endosperm AX in transgenic wheat by down-regulating either TaGT43_2 or TaGT47_2 genes (orthologues to IRX9 and IRX10 in Arabidopsis, respectively) implicated in AX chain extension and the TaXAT1 gene responsible for monosubstitution by 3-linked arabinose. Here, we use these transgenic lines to investigate the relationship between amounts of AX in soluble and insoluble fractions, the chain-length distribution of these measured by intrinsic viscosity and the overall effect on extract viscosity. In transgenic lines expressing either the TaGT43_2 or TaGT47_2 RNAi transgenes, the intrinsic viscosities of water-extractable (WE-AX) and of a water-insoluble alkaline-extracted fraction (AE-AX) were decreased by between 10% and 50% compared to control lines. In TaXAT1 RNAi lines, there was a 15% decrease in intrinsic viscosity of WE-AX but no consistent effect on that of AE-AX. All transgenic lines showed decreases in extract viscosity with larger effects in TaGT43_2 and TaGT47_2 RNAi lines (by up to sixfold) than in TaXAT1 RNAi lines (by twofold). These effects were explained by the decreases in amount and chain length of WE-AX, with decreases in amount having the greater influence. Extract viscosity from wheat grain can therefore be greatly decreased by suppression of single gene targets. PMID:25819752

  5. Contribution of Disulfide Bridges to the Thermostability of a Type A Feruloyl Esterase from Aspergillus usamii

    OpenAIRE

    Xin Yin; Die Hu; Jian-Fang Li; Yao He; Tian-Di Zhu; Min-Chen Wu

    2015-01-01

    The contribution of disulfide bridges to the thermostability of a type A feruloyl esterase (AuFaeA) from Aspergillus usamii E001 was studied by introducing an extra disulfide bridge or eliminating a native one from the enzyme. MODIP and DbD, two computational tools that can predict the possible disulfide bridges in proteins for thermostability improvement, and molecular dynamics (MD) simulations were used to design the extra disulfide bridge. One residue pair A126-N152 was chosen, and the res...

  6. Rice Bran Feruloylated Oligosaccharides Activate Dendritic Cells via Toll-Like Receptor 2 and 4 Signaling

    OpenAIRE

    Chi Chen Lin; Hua Han Chen; Yu Kuo Chen; Hung Chia Chang; Ping Yi Lin; I-Hong Pan; Der-Yuan Chen; Chuan Mu Chen; Su Yi Lin

    2014-01-01

    This work presents the effects of feruloylated oligosaccharides (FOs) of rice bran on murine bone marrow-derived dendritic cells (BMDCs) and the potential pathway through which the effects are mediated. We found that FOs induced phenotypic maturation of DCs, as shown by the increased expression of CD40, CD80/CD86 and MHC-I/II molecules. FOs efficiently induced maturation of DCs generated from C3H/HeN or C57BL/6 mice with normal toll-like receptor 4 (TLR-4) or TLR-2 but not DCs from mice with ...

  7. Decreased Polysaccharide Feruloylation Compromises Plant Cell Wall Integrity and Increases Susceptibility to Necrotrophic Fungal Pathogens

    Science.gov (United States)

    Reem, Nathan T.; Pogorelko, Gennady; Lionetti, Vincenzo; Chambers, Lauran; Held, Michael A.; Bellincampi, Daniela; Zabotina, Olga A.

    2016-01-01

    The complexity of cell wall composition and structure determines the strength, flexibility, and function of the primary cell wall in plants. However, the contribution of the various components to cell wall integrity (CWI) and function remains unclear. Modifications of cell wall composition can induce plant responses known as CWI control. In this study, we used transgenic expression of the fungal feruloyl esterase AnFAE to examine the effect of post-synthetic modification of Arabidopsis and Brachypodium cell walls. Transgenic Arabidopsis plants expressing AnFAE showed a significant reduction of monomeric ferulic acid, decreased amounts of wall-associated extensins, and increased susceptibility to Botrytis cinerea, compared with wild type. Transgenic Brachypodium showed reductions in monomeric and dimeric ferulic acids and increased susceptibility to Bipolaris sorokiniana. Upon infection, transgenic Arabidopsis and Brachypodium plants also showed increased expression of several defense-related genes compared with wild type. These results demonstrate a role, in both monocot and dicot plants, of polysaccharide feruloylation in plant CWI, which contributes to plant resistance to necrotrophic pathogens. PMID:27242834

  8. Decreased Polysaccharide Feruloylation Compromises Plant Cell Wall Integrity and Increases Susceptibility to Necrotrophic Fungal Pathogens

    Directory of Open Access Journals (Sweden)

    Nathan T Reem

    2016-05-01

    Full Text Available The complexity of cell wall composition and structure determines the strength, flexibility, and function of the primary cell wall in plants. However, the contribution of the various components to cell wall integrity and function remains unclear. Modifications of cell wall composition can induce plant responses known as Cell Wall Integrity control. In this study, we used transgenic expression of the fungal feruloyl esterase AnFAE to examine the effect of post-synthetic modification of Arabidopsis and Brachypodium cell walls. Transgenic Arabidopsis plants expressing AnFAE showed a significant reduction of monomeric ferulic acid, increased amounts of wall-associated extensins, and increased susceptibility to Botrytis cinerea, compared with wild type. Transgenic Brachypodium showed reductions in monomeric and dimeric ferulic acids and increased susceptibility to Bipolaris sorokiniana. Upon infection, transgenic Arabidopsis and Brachypodium plants also showed increased expression of several defense-related genes compared with wild type. These results demonstrate a role, in both monocot and dicot plants, of polysaccharide feruloylation in plant cell wall integrity, which contributes to plant resistance to necrotrophic pathogens.

  9. Decreased Polysaccharide Feruloylation Compromises Plant Cell Wall Integrity and Increases Susceptibility to Necrotrophic Fungal Pathogens.

    Science.gov (United States)

    Reem, Nathan T; Pogorelko, Gennady; Lionetti, Vincenzo; Chambers, Lauran; Held, Michael A; Bellincampi, Daniela; Zabotina, Olga A

    2016-01-01

    The complexity of cell wall composition and structure determines the strength, flexibility, and function of the primary cell wall in plants. However, the contribution of the various components to cell wall integrity (CWI) and function remains unclear. Modifications of cell wall composition can induce plant responses known as CWI control. In this study, we used transgenic expression of the fungal feruloyl esterase AnFAE to examine the effect of post-synthetic modification of Arabidopsis and Brachypodium cell walls. Transgenic Arabidopsis plants expressing AnFAE showed a significant reduction of monomeric ferulic acid, decreased amounts of wall-associated extensins, and increased susceptibility to Botrytis cinerea, compared with wild type. Transgenic Brachypodium showed reductions in monomeric and dimeric ferulic acids and increased susceptibility to Bipolaris sorokiniana. Upon infection, transgenic Arabidopsis and Brachypodium plants also showed increased expression of several defense-related genes compared with wild type. These results demonstrate a role, in both monocot and dicot plants, of polysaccharide feruloylation in plant CWI, which contributes to plant resistance to necrotrophic pathogens. PMID:27242834

  10. Novel feruloyl esterase from Lactobacillus fermentum NRRL B-1932 and analysis of the recombinant enzyme produced in Escherichia coli.

    Science.gov (United States)

    Using agar plates containing ethyl ferulate as the sole carbon source, 33 Lactobacillus strains were screened for feruloyl esterase (FE) activity. Among a dozen species showing a clearing zone on the opaque plate containing ethyl ferulate, Lactobacillus fermentum NRRL B-1932 demonstrated the stronge...

  11. Feruloyl esterases as a tool for the release of phenolic compounds from agro-industrial by-products

    NARCIS (Netherlands)

    Benoit, Isabelle; Navarro, David; Marnet, Nathalie; Rakotomanomana, Nnjara; Lesage-Meessen, Laurence; Sigoillot, Jean-Claude; Asther, Marcel; Asther, Michèle

    2006-01-01

    Agro-industrial by-products are a potential source of added-value phenolic acids with promising applications in the food and pharmaceutical industries. Here two purified feruloyl esterases from Aspergillus niger, FAEA and FAEB were tested for their ability to release phenolic acids such as caffeic a

  12. Enzymatic Cross-Linking of Alkali Extracted Arabinoxylans: Gel Rheological and Structural Characteristics

    Directory of Open Access Journals (Sweden)

    Agustin Rascón-Chu

    2011-09-01

    Full Text Available Ferulated arabinoxylans were alkali-extracted from wheat bran at different incubation times (0.0, 0.5, 1.0, 1.5 and 2.0 h. Wheat bran ferulated arabinoxylans (WBAX arabinose-to-xylose ratio, ferulic acid content, intrinsic viscosity and viscosimetric molecular weight values decreased as the incubation time of extraction increased. WBAX enzymatic cross-linking capability was affected by incubation time while an increase in WBAX concentration from 5 to 6% (w/v favored gelation. The WBAX gels formed presented a macroporous structure with mesh size ranging from 40 to 119 nm and hardness values varying from 1.7 to 5 N.

  13. Resistant starch and arabinoxylan augment SCFA absorption, but affect postprandial glucose and insulin responses differently - CORRIGENDUM

    DEFF Research Database (Denmark)

    Ingerslev, Anne Krog; Theil, Peter Kappel; Hedemann, Mette Skou;

    2015-01-01

    glucose was observed in RSD fed pigs (203 mmol/h, P = 0.02). The NPF of total SCFA, acetate, propionate, and butyrate were high, intermediate, and low (P < 0.01) in AXD, RSD, and WSD fed pigs, respectively, with the largest relative increase for butyrate in response to arabinoxylan supplementation. In......The effects of increased colonic fermentation of dietary fibres (DF) on net portal flux (NPF) of carbohydrate-derived metabolites (glucose, SCFA and especially butyrate), hormones (insulin, C-peptide, GLP-1, GIP) and NEFA were studied in a healthy catheterised pig model. Six 59 ± 3.8 kg pigs were...... conclusion, RSD and AXD had different effects on net portal insulin and glucose flux, suggesting different impact of arabinoxylan and resistant starch on human health....

  14. Arabinoxylan Oligosaccharide Hydrolysis by Family 43 and 51 Glycosidases from Lactobacillus brevis DSM 20054

    OpenAIRE

    Michlmayr, Herbert; Hell, Johannes; Lorenz, Cindy; Böhmdorfer, Stefan; Rosenau, Thomas; Kneifel, Wolfgang

    2013-01-01

    Due to their potential prebiotic properties, arabinoxylan-derived oligosaccharides [(A)XOS] are of great interest as functional food and feed ingredients. While the (A)XOS metabolism of Bifidobacteriaceae has been extensively studied, information regarding lactic acid bacteria (LAB) is still limited in this context. The aim of the present study was to fill this important gap by characterizing candidate (A)XOS hydrolyzing glycoside hydrolases (GHs) identified in the genome of Lactobacillus bre...

  15. Interactions of arabinoxylan and (1,3)(1,4)-β-glucan with cellulose networks.

    Science.gov (United States)

    Mikkelsen, Deirdre; Flanagan, Bernadine M; Wilson, Sarah M; Bacic, Antony; Gidley, Michael J

    2015-04-13

    To identify interactions of relevance to the structure and properties of the primary cell walls of cereals and grasses, we used arabinoxylan and (1,3)(1,4)-β-glucan, major polymers in cereal/grass primary cell walls, to construct composites with cellulose produced by Gluconacetobacter xylinus. Both polymers associated prolifically with cellulose without becoming rigid or altering the nature or extent of cellulose crystallinity. Mechanical properties were modestly affected compared with xyloglucan or pectin (characteristic components of nongrass primary cell walls) composites with cellulose. In situ depletion of arabinoxylan arabinose side chains within preformed cellulose composites resulted in phase separation, with only limited enhancement of xylan-cellulose interactions. These results suggest that arabinoxylan and (1 → 3)(1 → 4)-β-d-glucan are not functional homologues for either xyloglucan or pectin in the way they interact with cellulose networks. Association of cell-wall polymers with cellulose driven by entropic amelioration of high energy cellulose/water interfaces should be considered as a third type of interaction within cellulose-based cell walls, in addition to molecular binding (enthalpic driving force) exhibited by, for example, xyloglucans or mannans, and interpenetrating networks based on, for example, pectins. PMID:25756836

  16. Extraction and chemical characterization of rye arabinoxylan and the effect of β-glucan on the mechanical and barrier properties of cast arabinoxylan films

    DEFF Research Database (Denmark)

    Sárossy, Zsuzsa; Tenkanen, Maija; Pitkänen, Leena;

    2013-01-01

    properties (between 0.9 and 1.0 cm3 mm/m2 d kPa). However, the water vapor permeability increased with addition of increasing amounts of BG to WE-AX. To our knowledge, this is the first study on the effect of β-glucans on the material and permeability properties of arabinoxylan-based films. © 2012 Elsevier......Water-extractable hemicellulose (WEH) fractions, containing approximately 65% arabinoxylans (WE-AX) and 20% mixed-linkage b-glucans were isolated from rye bran. In addition, water-extractable mixedlinkage β-glucans (BG) were isolated from oat bran as a reference material. The β-glucan content of....../mol. The material properties of films prepared from the rye hemicellulose isolate and WE-AX as such, or with varying amounts of added BG (20:80; 50:50; 80:20 ratios) were studied. Prior removal of β-glucan from the isolate decreased the tensile strength of the films significantly as well as the elongation...

  17. MIF proteins are not glutathione transferase homologs.

    OpenAIRE

    Pearson, W R

    1994-01-01

    Although macrophage migration inhibitory factor (MIF) proteins conjugate glutathione, sequence analysis does not support their homology to other glutathione transferases. Glutathione transferases are not detected with MIF proteins in searches of protein sequence databases, and MIF proteins do not share significant sequence similarity with glutathione transferases. Homology cannot be demonstrated by multiple sequence alignment or evolutionary tree construction; such methods assume that the pro...

  18. Feruloylated and Nonferuloylated Arabino-oligosaccharides from Sugar Beet Pectin Selectively Stimulate the Growth of Bifidobacterium spp. in Human Fecal in Vitro Fermentations

    DEFF Research Database (Denmark)

    Holck, Jesper; Lorentzen, Andrea; Vigsnæs, Louise Kristine; Licht, Tine Rask; Mikkelsen, Jørn Dalgaard; Meyer, Anne S.

    2011-01-01

    The side chains of the rhamnogalacturonan I fraction in sugar beet pectin are particularly rich in arabinan moieties, which may be substituted with feruloyl groups. In this work the arabinan-rich fraction resulting from sugar beet pulp based pectin production was separated by Amberlite XAD...... bioactive feruloylated arabino-oligosaccharides from sugar beet pulp and an initial indication of the potentially larger bifidogenic effect of relatively long-chain arabino-oligosaccharides as opposed to short-chain arabino-oligosaccharides....

  19. Effects of a Bioavailable Arabinoxylan-enriched White Bread Flour on Postprandial Glucose Response in Normoglycemic Subjects.

    Science.gov (United States)

    Giulia Falchi, Anna; Grecchi, Ilaria; Muggia, Chiara; Palladini, Giuseppina; Perlini, Stefano

    2016-11-01

    The beneficial effects of soluble fibers on carbohydrate metabolism are well documented. In this regard, we tested an arabinoxylan-enriched white bread flour, obtained by a patented process by which the bran extracted from the milling process is enzymatically hydrolyzed in order to separate the soluble fraction fiber from the insoluble fiber. We recruited 24 healthy normoglycemic volunteers [Age 34-61 ± 12.5 y; Body Mass Index (BMI) 22.1 ± 2.5 kg/m(2); Waist circumference (WC) 84.43 ± 8.0 cm; Fat Mass (FM) 22.7 ± 8.0%] attending the Dietetics Outpatient Clinic of the Internal Medicine Department at IRCCS Policlinico S. Matteo Foundation, University of Pavia, Pavia, Italy. Subjects acutely consumed arabinoxylan-enriched white bread (weight: 100 g) or isoenergetic control breads, in a double-blind crossover study design. Plasma glucose levels were measured just before bread administration and 30 minutes afterwards. The 30-minute peak postprandial glucose concentrations after arabinoxylan-enriched meals were significantly lower than after the control meal (107±4.6 mg/dL vs. 121 ± 5.2 mg/dL; p < 0.05). The here-reported results show how postprandial glucose responses were improved by ingestion of the arabinoxylan-enriched meal. Further studies are needed to clarify whether daily consumption of arabinoxylan-enriched bread will benefit patients with type 2 diabetes mellitus. PMID:27049812

  20. Enzyme catalyzed oxidative cross-linking of feruloylated pectic polysaccharides from sugar beet

    DEFF Research Database (Denmark)

    Abang Zaidel, Dayang Norulfairuz

    has been our intention to study the rheological properties of cross-linked feruloylated arabinanoligosaccharide, however the attempt has not been fully achieved. It might be due to small molecular weight of the arabinan (?1.3 kDa) which prevented the measurement of the rheological properties since the......Sugar beet pulp is a byproduct from sugar production consisting mainly of cellulose and pectic polysaccharide. Its utilization has been mostly as feedstock due to its high content of energy and fiber. This study emphasizes on the utilization of the pectin and arabinan fractions extracted from sugar...... beet pulp as a potential starting material for production of pectin derived products which could help maintain the competitiveness of the sugar beet based industry. The overall objective of this study has been focusing on understanding the kinetics of enzyme catalyzed oxidative crosslinking of...

  1. Production of Feruloyl Esterase from Aspergillus niger by Solid-State Fermentation on Different Carbon Sources

    Directory of Open Access Journals (Sweden)

    Shiyi Ou

    2011-01-01

    Full Text Available A mixture of wheat bran with maize bran as a carbon source and addition of (NH4SO4 as nitrogen source was found to significantly increase production of feruloyl esterase (FAE enzyme compared with wheat bran as a sole carbon and nitrogen source. The optimal conditions in conical flasks were carbon source (30 g to water 1 : 1, maize bran to wheat bran 1 : 2, (NH4SO4 1.2 g and MgSO4 70 mg. Under these conditions, FAE activity was 7.68 mU/g. The FAE activity on the mixed carbon sources showed, high activity against the plant cell walls contained in the cultures.

  2. How well do the substrates KISS the enzyme? Molecular docking program selection for feruloyl esterases

    DEFF Research Database (Denmark)

    Udatha, D. B. R. K. Gupta; Sugaya, Nobuyoshi; Olsson, Lisbeth; Panagiotou, Gianni

    2012-01-01

    Molecular docking is the most commonly used technique in the modern drug discovery process where computational approaches involving docking algorithms are used to dock small molecules into macromolecular target structures. Over the recent years several evaluation studies have been reported by...... independent scientists comparing the performance of the docking programs by using default 'black box' protocols supplied by the software companies. Such studies have to be considered carefully as the docking programs can be tweaked towards optimum performance by selecting the parameters suitable for the...... target of interest. In this study we address the problem of selecting an appropriate docking and scoring function combination (88 docking algorithm-scoring functions) for substrate specificity predictions for feruloyl esterases, an industrially relevant enzyme family. We also propose the 'Key Interaction...

  3. Optimization of reaction conditions for enzymatic viscosity reduction and hydrolysis of wheat arabinoxylan in an industrial ethanol fermentation residue

    DEFF Research Database (Denmark)

    Sørensen, H.R.; Pedersen, S.; Meyer, Anne Boye Strunge

    2006-01-01

    arabinoxylan in the different vinasse fractions irrespective of the state of solubility of the substrate material. The levels of liberated arabinose and xylose increased with increased dry matter concentration during enzymatic hydrolysis in the vinasse and the vinasse supernatant, but at the same time......, increased substrate dry matter concentrations gave corresponding linear decreases in the hydrolytic efficiency as evaluated from levels of monosaccharide release per weight unit dry matter. The study thus documents that enzymatic arabinoxylan hydrolysis of the vinasse significantly decreases the vinasse......This study examined enzyme-catalyzed viscosity reduction and evaluated the effects of substrate dry matter concentration on enzymatic degradation of arabinoxylan in a fermentation residue, "vinasse", resulting from industrial ethanol manufacture on wheat. Enzymatic catalysis was accomplished with a...

  4. Postprandial effects of test meals including concentrated arabinoxylan and whole grain rye in subjects with the metabolic syndrome

    DEFF Research Database (Denmark)

    Hartvigsen, M L; Lærke, H N; Bjørnshave, Ann;

    2014-01-01

    grain rye kernels on postprandial glucose, insulin, free fatty acids (FFA), gut hormones, SCFA and appetite in subjects with the metabolic syndrome (MetS). SUBJECTS/METHODS: Fifteen subjects with MetS participated in this acute, randomised, cross-over study. The test meals each providing 50 g of...... digestible carbohydrate were as follows: semolina porridge added concentrated arabinoxylan (AX), rye kernels (RK) or concentrated arabinoxylan combined with rye kernels (AXRK) and semolina porridge as control (SE). A standard lunch was served 4 h after the test meals. Blood samples were drawn during a 6-h...... meal glucose response. It remains to be tested in a long-term study if a beneficial effect on the glucose response of the isolated arabinoxylan will be related to the SCFA production....

  5. In vitro fermentation of arabinoxylan oligosaccharides and low molecular mass arabinoxylans with different structural properties from wheat (Triticum aestivum L.) bran and psyllium (Plantago ovata Forsk) seed husk.

    Science.gov (United States)

    Pollet, Annick; Van Craeyveld, Valerie; Van de Wiele, Tom; Verstraete, Willy; Delcour, Jan A; Courtin, Christophe M

    2012-02-01

    Ball milling was used for producing complex arabinoxylan oligosaccharides (AXOS) and low molecular mass arabinoxylans (AX) from wheat bran, pericarp-enriched wheat bran, and psyllium seed husk. The arabinose to xylose ratio of the samples produced varied between 0.14 and 0.92, and their average degree of polymerization (avDP) ranged between 42 and 300. Their fermentation for 48 h in an in vitro system using human colon suspensions was compared to enzymatically produced wheat bran AXOS with an arabinose to xylose ratio of 0.22 and 0.34 and an avDP of 4 and 40, respectively. Degrees of AXOS fermentation ranged from 28% to 50% and were lower for the higher arabinose to xylose ratio and/or higher avDP materials. Arabinose to xylose ratios of the unfermented fractions exceeded those of their fermented counterparts, indicating that molecules less substituted with arabinose were preferably fermented. Xylanase, arabinofuranosidase, and xylosidase activities increased with incubation time. Enzyme activities in the samples containing psyllium seed husk AX or psyllium seed husk AXOS were generally higher than those in the wheat bran AXOS preparations. Fermentation gave rise to unbranched short-chain fatty acids. Concentrations of acetic, propionic, and butyric acids increased to 1.9-2.6, 1.9-2.8, and 1.3-2.0 times their initial values, respectively, after 24 h incubation. Results show that the human intestinal microbiota can at least partially use complex AXOS and low molecular mass AX. The tested materials are thus interesting physiologically active carbohydrates. PMID:22224418

  6. Crystallization and preliminary X-ray analysis of a novel halotolerant feruloyl esterase identified from a soil metagenomic library

    International Nuclear Information System (INIS)

    A novel feruloyl esterase (EstF27) identified from a soil metagenomic library has been crystallized and a complete data set was collected from a single cooled crystal using an in-house X-ray source. Feruloyl esterase cleaves the ester linkage formed between ferulic acid and polysaccharides in plant cell walls and thus has wide potential industrial applications. A novel feruloyl esterase (EstF27) identified from a soil metagenomic library was crystallized and a complete data set was collected from a single cooled crystal using an in-house X-ray source. The crystal diffracted to 2.9 Å resolution and belonged to space group P212121, with unit-cell parameters a = 94.35, b = 106.19, c = 188.51 Å, α = β = γ = 90.00°. A Matthews coefficient of 2.55 Å3 Da−1, with a corresponding solvent content of 51.84%, suggested the presence of ten protein subunits in the asymmetric unit

  7. FILMS FROM SPRUCE GALACTOGLUCOMANNAN BLENDED WITH POLY(VINYL ALCOHOL), CORN ARABINOXYLAN, AND KONJAC GLUCOMANNAN

    OpenAIRE

    Kirsi S. Mikkonen; Yadav, Madhav P.; Peter Cooke; Stefan Willför; Hicks, Kevin B; Maija Tenkanen

    2008-01-01

    The improvement of mechanical properties of spruce galactoglucomannan (GGM)-based films was sought by blending GGM with each of poly(vinyl alcohol) (PVOH), corn arabinoxylan (cAX), and konjac glucomannan (KGM). The blend ratios were 3:1, 1:1, and 1:3 (w/w), and in addition films were made from each of the polymers alone. Glycerol was used as plasticizer. Adding other polymers increased the elongation at break of GGM blend films. The tensile strength of films increased with increasing amount o...

  8. Characterization of Water Extractable Arabinoxylans from a Spring Wheat Flour: Rheological Properties and Microstructure

    Directory of Open Access Journals (Sweden)

    Alma Campa-Mada

    2013-07-01

    Full Text Available In the present study water extractable arabinoxylans (WEAX from a Mexican spring wheat flour (cv. Tacupeto F2001 were isolated, characterized and gelled and the gel rheological properties and microstructure were investigated. These WEAX presented an arabinose to xylose ratio of 0.66, a ferulic acid and diferulic acid content of 0.526 and 0.036 µg/mg WEAX, respectively and a Fourier Transform Infra-Red (FT-IR spectrum typical of arabinoxylans. The intrinsic viscosity and viscosimetric molecular weight values for WEAX were 3.5 dL/g and 504 kDa, respectively. WEAX solution at 2% (w/v formed gels induced by a laccase as cross-linking agent. Cured WEAX gels registered storage (G’ and loss (G’’ modulus values of 31 and 5 Pa, respectively and a diferulic acid content of 0.12 µg/mg WEAX, only traces of triferulic acid were detected. Scanning electron microscopy analysis of the lyophilized WEAX gels showed that this material resembles that of an imperfect honeycomb.

  9. Comparison of different twin-screw extraction conditions for the production of arabinoxylans.

    Science.gov (United States)

    Jacquemin, Leslie; Mogni, Assad; Zeitoun, Rawan; Guinot, Cécile; Sablayrolles, Caroline; Saulnier, Luc; Pontalier, Pierre-Yves

    2015-02-13

    The aim of this article is to compare two different sets of optimal conditions for twin-screw extraction of xylans and define their influence on the purification steps, combining ultrafiltration and industrial chromatography. Two xylan extracts were obtained by twin-screw extrusion of straw and bran. Condition 1 used a high straw/bran ratio (equal to 6) and high sodium hydroxide content, and condition 2 used a lower straw/bran ratio (equal to 2) and low sodium hydroxide content. Arabinoxylan extraction yields are slightly higher for conditions with low straw content (5.1% versus 4.4%). Nevertheless, these recovery yields remain between 9% and 10%. Ultrafiltration is as efficient as evaporation for polysaccharide concentration, with lower energy consumption, but also demineralizes the solution. The combination of ultrafiltration and chromatography gives partial purification of the extract with a final arabinoxylan purity ranging from 16% to 26%. This is slightly higher than by direct precipitation, but limited because all the large molecules such as proteins and lignins were retained by ultrafiltration. PMID:25458276

  10. Production and partial characterization of arabinoxylan-degrading enzymes by Penicillium brasilianum under solid-state fermentation

    DEFF Research Database (Denmark)

    Panagiotou, Gianni; Granouillet, P.; Olsson, Lisbeth

    2006-01-01

    The production of a battery of arabinoxylan-degrading enzymes by the fungus Penicillium brasilianum grown on brewer's spent grain (BSG) under solid-state fermentation was investigated. Initial moisture content, initial pH, temperature, and nitrogen source content were optimized to achieve maximum...

  11. Effects of arabinoxylan-oligosaccharides (AXOS) on juvenile Siberian sturgeon (Acipenser baerii) performance, immune responses and gastrointestinal microbial community

    NARCIS (Netherlands)

    Geraylou, Z.; Souffreau, C.; Rurangwa, E.; Hondt, D' S.; Callewaert, L.; Courtin, C.M.; Delcour, J.A.; Buyse, J.; Ollevier, F.

    2012-01-01

    Arabinoxylan-oligosaccharides (AXOS) are a newly discovered class of candidate prebiotics that exert different properties depending on their structure. In this study the effects of two different structures of AXOS, namely AXOS-32-0.30 (average degree of polymerization: 32, average degree of substitu

  12. Production and partial characterization of alkaline feruloyl esterases by Fusarium oxysporum during submerged batch cultivation

    DEFF Research Database (Denmark)

    Topakas, E.; Christakopoulos, Paul

    2004-01-01

    Production of feruloyl esterases (FAEs) by Fusarium oxysporum was enhanced by optimization of initial pH of the culture medium, the type and concentration of nitrogen and carbon source. Submerged batch cultivation in a laboratory bioreactor (17 1) produced activity at 82 nkat g(-1) dry substrate....... Production of FAE does not therefore, require FA, however, production is diminished by the removal of esterified FA from the growth substrate. Optimal FAE activity was observed at pH 7 and 50 degreesC with 68 and 55% activity at pH 8 and pH 9, respectively. The esterase was fully stable at pH 5-8 and up to...... 40 degreesC and retained 72 and 40% of its activity after 6 h at pH 9 and pH 10, respectively. After separation by isoelectric focusing electrophoresis, a zymogram indicated one major FAE activity exhibiting pI value of 10.5....

  13. Rice Bran Feruloylated Oligosaccharides Activate Dendritic Cells via Toll-Like Receptor 2 and 4 Signaling

    Directory of Open Access Journals (Sweden)

    Chi Chen Lin

    2014-04-01

    Full Text Available This work presents the effects of feruloylated oligosaccharides (FOs of rice bran on murine bone marrow-derived dendritic cells (BMDCs and the potential pathway through which the effects are mediated. We found that FOs induced phenotypic maturation of DCs, as shown by the increased expression of CD40, CD80/CD86 and MHC-I/II molecules. FOs efficiently induced maturation of DCs generated from C3H/HeN or C57BL/6 mice with normal toll-like receptor 4 (TLR-4 or TLR-2 but not DCs from mice with mutated TLR4 or TLR2. The mechanism of action of FOs may be mediated by increased phosphorylation of ERK, p38 and JNK mitogen-activated protein kinase (MAPKs and increased NF-kB activity, which are important signaling molecules downstream of TLR-4 and TLR-2. These data suggest that FOs induce DCs maturation through TLR-4 and/or TLR-2 and that FOs might have potential efficacy against tumor or virus infection or represent a candidate-adjuvant approach for application in immunotherapy and vaccination.

  14. Feruloylated Oligosaccharides from Maize Bran Modulated the Gut Microbiota in Rats.

    Science.gov (United States)

    Ou, Juan-Ying; Huang, Jun-Qing; Song, Yuan; Yao, Sheng-Wen; Peng, Xi-Chun; Wang, Ming-Fu; Ou, Shi-Yi

    2016-06-01

    Corn bran is a byproduct produced from corn milling; it is rich in ferulic acid and hemicellulose. In this research, the effects of feruloylated oligosaccharides (FOs) from maize bran on the microbial diversity and profiles in rat feces were investigated through 16S rRNA sequencing. FOs significantly increased bacterial richness and diversity compared with the control and xylooligosaccharides (XOS) alone. In comparison with the control group and the group administrated with XOS, FOs orally administered at 300 mg/kg increased OTU in feces by 57.0 and 24.8 %, and Chao value by 93.4 and 37.6 %, respectively. FOs also influenced obesity- and diabetes-associated bacteria. Oral administration of FOs at 300 mg/kg decreased the ratio of Firmicutes to Bacteroidetes from 477.7:1 to 55.1:1; greatly increased the reads of bacteria that were previously found resistant against diabetes in rats, such as Actinobacteria, Bacteroides, and Lactobacillus; whereas decreased diabetes-prone bacteria, such as Clostridium and Firmicutes. PMID:27165128

  15. Rice bran feruloylated oligosaccharides activate dendritic cells via Toll-like receptor 2 and 4 signaling.

    Science.gov (United States)

    Lin, Chi Chen; Chen, Hua Han; Chen, Yu Kuo; Chang, Hung Chia; Lin, Ping Yi; Pan, I-Hong; Chen, Der-Yuan; Chen, Chuan Mu; Lin, Su Yi

    2014-01-01

    This work presents the effects of feruloylated oligosaccharides (FOs) of rice bran on murine bone marrow-derived dendritic cells (BMDCs) and the potential pathway through which the effects are mediated. We found that FOs induced phenotypic maturation of DCs, as shown by the increased expression of CD40, CD80/CD86 and MHC-I/II molecules. FOs efficiently induced maturation of DCs generated from C3H/HeN or C57BL/6 mice with normal toll-like receptor 4 (TLR-4) or TLR-2 but not DCs from mice with mutated TLR4 or TLR2. The mechanism of action of FOs may be mediated by increased phosphorylation of ERK, p38 and JNK mitogen-activated protein kinase (MAPKs) and increased NF-kB activity, which are important signaling molecules downstream of TLR-4 and TLR-2. These data suggest that FOs induce DCs maturation through TLR-4 and/or TLR-2 and that FOs might have potential efficacy against tumor or virus infection or represent a candidate-adjuvant approach for application in immunotherapy and vaccination. PMID:24762969

  16. Enzymatic hydrolysis of water-soluble wheat arabinoxylan. 1. Synergy between alpha-L-arabinofuranosidases, endo-1,4-beta-xylanases, and beta-xylosidase activities

    DEFF Research Database (Denmark)

    Sørensen, H.R.; Meyer, Anne Boye Strunge; Pedersen, S.

    2003-01-01

    Hydrolysis of arabinoxylan is an important prerequisite for improved utilization of wheat hemicellulose in the ethanol fermentation industry. This study investigates the individual and combined efficiencies of three commercial, cellulytic and hemicellulytic enzyme preparations, Celluclast 1.5 L, ...

  17. Ingestion of a novel galactoglucomannan oligosaccharide-arabinoxylan (GGMO-AX) complex affected growth performance and fermentative and immunological characteristics of broiler chicks challenged with Salmonella typhimurium

    Science.gov (United States)

    Fermentable carbohydrates may enhance the ability of the gastrointestinal tract to defend against a pathogenic infection. We hypothesized that a galactoglucomannan oligosaccharide-arabinoxylan (GGMO-AX) complex would positively impact immune status and prevent colonization and shedding in Salmonell...

  18. Application of model bread baking in the examination of arabinoxylan-protein complexes in rye bread.

    Science.gov (United States)

    Buksa, Krzysztof

    2016-09-01

    The changes in molecular mass of arabinoxylan (AX) and protein caused by bread baking process were examined using a model rye bread. Instead of the normal flour, the dough contained starch, water-extractable AX and protein which were isolated from rye wholemeal. From the crumb of selected model breads, starch was removed releasing AX-protein complexes, which were further examined by size exclusion chromatography. On the basis of the research, it was concluded that optimum model mix can be composed of 3-6% AX and 3-6% rye protein isolate at 94-88% of rye starch meaning with the most similar properties to low extraction rye flour. Application of model rye bread allowed to examine the interactions between AX and proteins. Bread baked with a share of AX, rye protein and starch, from which the complexes of the highest molar mass were isolated, was characterized by the strongest structure of the bread crumb. PMID:27185141

  19. Impact of wheat flour-associated endoxylanases on arabinoxylan in dough after mixing and resting.

    Science.gov (United States)

    Dornez, Emmie; Gebruers, Kurt; Cuyvers, Sven; Delcour, Jan A; Courtin, Christophe M

    2007-08-22

    The impact of varying levels of endoxylanase activity in wheat flour on arabinoxylan (AX) in mixed and rested dough was studied using eight industrially milled wheat flour fractions with varying endoxylanase activity levels. Analysis of the levels of reducing end xylose (RX) and solubilized AX (S-AX) formed during mixing and resting and their correlation with the endoxylanase activity in the flour milling fractions showed that solubilization of AX during the mixing phase is mainly due to mechanical forces, while solubilization of AX during resting is caused by endoxylanase activity. Moreover, solubilization of AX during the dough resting phase is more outspoken than that during the mixing phase. Besides endoxylanase activity, there were significant xylosidase and arabinofuranosidase activities during the dough resting phase. The results indicate that wheat flour-associated endoxylanases can alter part of the AX in dough, thereby changing their functionality in bread making and potentially affecting dough and end product properties. PMID:17661495

  20. Arabinoxylan-mediated synthesis of gold and silver nanoparticles having exceptional high stability.

    Science.gov (United States)

    Amin, Muhammad; Iram, Fozia; Iqbal, Mohammad S; Saeed, Muhammad Z; Raza, Mohsin; Alam, Shehzad

    2013-02-15

    A green synthesis of highly stable gold and silver nanoparticles (NPs) using arabinoxylan (AX) from ispaghula (Plantago ovata) seed husk is being reported. The NPs were synthesized by stirring a mixture of AX and HAuCl(4)·H(2)O or AgNO(3), separately, below 100 °C for less than an hour, where AX worked as the reducing and the stabilizing agent. The synthesized NPs were characterized by surface plasmon resonance (SPR) spectroscopy, transmission electron microscopy (TEM), atomic force microscopy (AFM), and X-ray diffraction (XRD). The particle size was (silver: 5-20 nm and gold: 8-30 nm) found to be dependent on pH, temperature, reaction time and concentrations of AX and the metal salts used. The NPs were poly-dispersed with a narrow range. They were stable for more than two years time. PMID:23399234

  1. FILMS FROM SPRUCE GALACTOGLUCOMANNAN BLENDED WITH POLY(VINYL ALCOHOL, CORN ARABINOXYLAN, AND KONJAC GLUCOMANNAN

    Directory of Open Access Journals (Sweden)

    Kirsi S. Mikkonen

    2008-02-01

    Full Text Available The improvement of mechanical properties of spruce galactoglucomannan (GGM-based films was sought by blending GGM with each of poly(vinyl alcohol (PVOH, corn arabinoxylan (cAX, and konjac glucomannan (KGM. The blend ratios were 3:1, 1:1, and 1:3 (w/w, and in addition films were made from each of the polymers alone. Glycerol was used as plasticizer. Adding other polymers increased the elongation at break of GGM blend films. The tensile strength of films increased with increasing amount of PVOH and KGM, but the effect of cAX was the opposite. Dynamic mechanical analysis showed two separate loss modulus peaks for blends of GGM and PVOH, but a single peak for all other films. Optical and scanning electron microscopy confirmed good miscibility of GGM with cAX and KGM. In contrast, films blended from GGM and PVOH showed phase separation when examined by microscopy.

  2. Rye Arabinoxylans: Molecular Structure, Physicochemical Properties and Physiological Effects in the Gastrointestinal Tract

    DEFF Research Database (Denmark)

    Bach Knudsen, Knud Erik; Lærke, Helle Nygaard

    2010-01-01

    Arabinoxylans (AX) are the main dietary fiber (DF) polysaccharides in rye where they represent ≈55% of the total polysaccharides. Rye AX consist of a backbone of (1→4)-β-d-xylopyranosyl residues (X) mainly substituted with α-l-arabinofuranosyl residues (A) to varying degrees at the O-2 position...... and 12 g of AX-rich wheat fiber to a breakfast meal has significantly lowered postprandial glucose and insulin response. Studies with hypercholesterolemic pigs fed rye buns rich in AX have resulted in dramatic reductions in plasma total and LDL cholesterol, whereas a gender difference was seen in...... studies on the effect of AX on plasma lipids in humans. Only certain species of bacteria from the human gut produce the enzymes needed for the degradation of AX. Nevertheless, wheat AX stimulate prebiotic bacteria presumably brought about by cross feeding of lactobacilli and bifidobacteria with...

  3. Family 42 carbohydrate-binding modules display multiple arabinoxylan-binding interfaces presenting different ligand affinities.

    Science.gov (United States)

    Ribeiro, Teresa; Santos-Silva, Teresa; Alves, Victor D; Dias, Fernando M V; Luís, Ana S; Prates, José A M; Ferreira, Luís M A; Romão, Maria J; Fontes, Carlos M G A

    2010-10-01

    Enzymes that degrade plant cell wall polysaccharides display a modular architecture comprising a catalytic domain bound to one or more non-catalytic carbohydrate-binding modules (CBMs). CBMs display considerable variation in primary structure and are grouped into 59 sequence-based families organized in the Carbohydrate-Active enZYme (CAZy) database. Here we report the crystal structure of CtCBM42A together with the biochemical characterization of two other members of family 42 CBMs from Clostridium thermocellum. CtCBM42A, CtCBM42B and CtCBM42C bind specifically to the arabinose side-chains of arabinoxylans and arabinan, suggesting that various cellulosomal components are targeted to these regions of the plant cell wall. The structure of CtCBM42A displays a beta-trefoil fold, which comprises 3 sub-domains designated as alpha, beta and gamma. Each one of the three sub-domains presents a putative carbohydrate-binding pocket where an aspartate residue located in a central position dominates ligand recognition. Intriguingly, the gamma sub-domain of CtCBM42A is pivotal for arabinoxylan binding, while the concerted action of beta and gamma sub-domains of CtCBM42B and CtCBM42C is apparently required for ligand sequestration. Thus, this work reveals that the binding mechanism of CBM42 members is in contrast with that of homologous CBM13s where recognition of complex polysaccharides results from the cooperative action of three protein sub-domains presenting similar affinities. PMID:20637315

  4. GGT (Gamma-Glutamyl Transferase) Test

    Science.gov (United States)

    ... be limited. Home Visit Global Sites Search Help? GGT Share this page: Was this page helpful? Also ... How is it used? The gamma-glutamyl transferase (GGT) test may be used to determine the cause ...

  5. Optimization of reaction conditions for enzymatic viscosity reduction and hydrolysis of wheat arabinoxylan in an industrial ethanol fermentation residue

    DEFF Research Database (Denmark)

    Sørensen, H.R.; Pedersen, S.; Meyer, Anne Boye Strunge

    2006-01-01

    This study examined enzyme-catalyzed viscosity reduction and evaluated the effects of substrate dry matter concentration on enzymatic degradation of arabinoxylan in a fermentation residue, "vinasse", resulting from industrial ethanol manufacture on wheat. Enzymatic catalysis was accomplished with...... viscosity and that a compromise in the dry matter must be found if enzymatic efficiency must be balanced with monosaccharide yields.......This study examined enzyme-catalyzed viscosity reduction and evaluated the effects of substrate dry matter concentration on enzymatic degradation of arabinoxylan in a fermentation residue, "vinasse", resulting from industrial ethanol manufacture on wheat. Enzymatic catalysis was accomplished with a...... increased enzyme dosage and treatment time at pH 5, 50 degrees C, 5 wt % vinasse dry matter. After 24 It of enzymatic treatment, 76-84%, 75-80%, and 43-47%, respectively, of the theoretically maximal arabinose, xylose, and glucose releases were achieved, indicating that the viscosity decrease was a result...

  6. Dependency of the hydrogen bonding capacity of the solvent anion on the thermal stability of feruloyl esterases in ionic liquid systems

    DEFF Research Database (Denmark)

    Zeuner, Birgitte; Ståhlberg, Tim; Nguyen van Buu, Olivier;

    2011-01-01

    Three feruloyl esterases, EC 3.1.1.73, (FAEs), namely FAE A from Aspergillus niger (AnFaeA), FAE C from Aspergillus nidulans (AndFaeC), and the FAE activity in a commercial b-glucanase mixture from Humicola insolens (Ultraflo L) were tested for their ability to catalyse esterification of sinapic ...

  7. The interplay of descriptor-based computational analysis with pharmacophore modeling builds the basis for a novel classification scheme for feruloyl esterases

    DEFF Research Database (Denmark)

    Udatha, D.B.R.K. Gupta; Kouskoumvekaki, Irene; Olsson, Lisbeth;

    2011-01-01

    One of the most intriguing groups of enzymes, the feruloyl esterases (FAEs), is ubiquitous in both simple and complex organisms. FAEs have gained importance in biofuel, medicine and food industries due to their capability of acting on a large range of substrates for cleaving ester bonds and...

  8. Characterization of water and alkali extractable arabinoxylan from wheat and rye under standardized conditions.

    Science.gov (United States)

    Buksa, Krzysztof; Praznik, Werner; Loeppert, Renate; Nowotna, Anna

    2016-03-01

    Arabinoxylans (AXs) are an important component of wheat and rye dough. They bind water, contribute to the formation of viscous dough and improve the quality of bread. For the application of AX fractions in bread making process, it is useful to record a quality profile of wheat fractions compared to the quality profile of rye fractions under standardized conditions. In this work water and alkali extractable AX containing fractions, from wheat- and rye wholemeal, were extracted under standardized conditions and characterized. For analysis of composition, structural features, and molecular dimension a combination of chemical, physicochemical, enzymatic and chromatographic techniques was applied. The molar mass distributions obtained by means of an innovative colorimetric pentose detection in the eluted SEC fractions were comparable for all under standardized conditions extracted AXs. The determined molar masses of AXs extracted both from wheat- and from rye grain were close to 2.0 × 10(5) g/mol for water extractable AXs and 3.0 × 10(5) g/mol for alkali extractable AXs. Different susceptibility to endoxylanase treatment, having been observed as differences in the SEC profiles, may be evidence of structural differences between AXs depending on their origin. The viscosities of AX solutions were strongly influenced by their molar mass and structure; samples being less susceptible to endoxylanase provided solutions of higher viscosity. PMID:27570263

  9. Entrapment of Probiotics in Water Extractable Arabinoxylan Gels: Rheological and Microstructural Characterization

    Directory of Open Access Journals (Sweden)

    Adriana Morales-Ortega

    2014-03-01

    Full Text Available Due to their porous structure, aqueous environment and dietary fiber nature arabinoxylan (AX gels could have potential applications for colon-specific therapeutic molecule delivery. In addition, prebiotic and health related effects of AX have been previously demonstrated. It has been also reported that cross-linked AX can be degraded by bacteria from the intestinal microbiota. However, AX gels have not been abundantly studied as carrier systems and there is no information available concerning their capability to entrap cells. In this regard, probiotic bacteria such as Bifidobacterium longum have been the focus of intense research activity lately. The objective of this research was to investigate the entrapment of probiotic B. longum in AX gels. AX solution at 2% (w/v containing B. longum (1 × 107 CFU/cm formed gels induced by laccase as cross-linking agent. The entrapment of B. longum decreased gel elasticity from 31 to 23 Pa, probably by affecting the physical interactions taking place between WEAX chains. Images of AX gels containing B. longum viewed under a scanning electron microscope show the gel network with the bacterial cells entrapped inside. The microstructure of these gels resembles that of an imperfect honeycomb. The results suggest that AX gels can be potential candidates for the entrapment of probiotics.

  10. Evaluation of the prebiotic potential of arabinoxylans from brewer's spent grain.

    Science.gov (United States)

    Reis, Sofia F; Gullón, Beatriz; Gullón, Patricia; Ferreira, Susana; Maia, Cláudio J; Alonso, José L; Domingues, Fernanda C; Abu-Ghannam, Nissreen

    2014-11-01

    Arabinoxylans (AX) consumption has been related to the treatment and prevention of cardiovascular diseases, type II diabetes, colorectal cancer and obesity. The beneficial health effects are conferred through gut microbiota modulation, and therefore, they have been proposed as potential slowly fermentable prebiotic candidates. As the mechanisms are not yet well understood, the prebiotic potential of AX from brewer's spent grain (BSG) has been investigated. Two types of AX from BSG (AX1 and AX2) of different length and branching averages were fermented with human faecal inocula and compared to fermented cultures containing a commercial prebiotic (fructooligosaccharide (FOS)) and cultures with no added carbohydrate (control). Results demonstrated that the AX were extensively metabolised after 48 h of fermentation. The pH decreased along fermentation and the lowest value was achieved in AX1 cultures. The production of short chain fatty acids (SCFA) was higher in AX cultures than in cultures containing FOS and controls, with AX1 presenting the highest concentrations. The stimulatory effect of beneficial bacteria was higher in AX cultures, and AX2 presented the highest positive effect. Prebiotic potential of AX from BSG was confirmed by the production of SCFA and the modulation of gut microbiota, especially by the high increase in bifidobacteria populations. PMID:25117549

  11. Purification and characterisation of arabinoxylan arabinofuranohydrolase I responsible for the filterability of barley malt.

    Science.gov (United States)

    Li, Xiaomin; Gao, Fei; Cai, Guolin; Jin, Zhao; Lu, Jian; Dong, Jianjun; Yin, Hua; Yu, Junhong; Yang, Mei

    2015-05-01

    Dan'er is a widely grown malt barley cultivar in China, but its filterability defects have severely impeded its application in beer brewing. Previous investigations have suggested that we should identify the malt filterability correlated proteins, one of which was postulated to be arabinoxylan arabinofuranohydrolase I (AXAH-I). To verify this hypothesis, we purified AXAH-I from Dan'er malt, characterised its enzyme performance, and investigated its influence on filterability by adding different amounts of purified enzyme to the mash. With 6 mU g(-1) malt AXAH-I supplemented, the wort separation rate increased by 31.8%, viscosity decreased by 3.6%, and the endosperm reserve contents declined concomitantly. Unexpectedly, the wort turbidity increased with increasing AXAH-I. We also tried to optimise the use of currently available commercial enzyme products for filterability improvement in beer brewing, by supplementing them with purified AXAH-I and β-amylase. AXAH-I could be a functional component for novel commercial enzyme products in the beer industry. PMID:25529682

  12. Enzymatic Xylose Release from Pretreated Corn Bran Arabinoxylan: Differential Effects of Deacetylation and Deferuloylation on Insoluble and Soluble Substrate Fractions

    DEFF Research Database (Denmark)

    Agger, Jane; Viksø-Nielsen, Ander; Meyer, Anne S.

    2010-01-01

    In the present work enzymatic hydrolysis of arabinoxylan from pretreated corn bran (190 °C, 10 min) was evaluated by measuring the release of xylose and arabinose after treatment with a designed minimal mixture of monocomponent enzymes consisting of α-l-arabinofuranosidases, an endoxylanase, and a...... β-xylosidase. The pretreatment divided the corn bran material 50:50 into soluble and insoluble fractions having A:X ratios of 0.66 and 0.40, respectively. Addition of acetyl xylan esterase to the monocomponent enzyme mixture almost doubled the xylose release from the insoluble substrate fraction and...

  13. Ball milling improves extractability and affects molecular properties of psyllium (Plantago ovata Forsk) seed husk arabinoxylan.

    Science.gov (United States)

    Van Craeyveld, Valerie; Delcour, Jan A; Courtin, Christophe M

    2008-12-10

    Psyllium (Plantago ovata Forsk) seed husk (PSH) is very rich in arabinoxylan (AX). However, its high gelling capacity and the complex nature of the AX make it difficult to process. In this study, ball milling was investigated as a tool for enhancing PSH AX water extractability and molecular mass (MM). A 48 h laboratory-scale ball mill treatment under standardized optimal conditions reduced the PSH average particle size from 161 microm for the untreated sample to 6 microm. Concurrently, it increased the water-extractable AX (WE-AX) level from 13 (untreated PSH) to 90% of the total PSH AX. While the WE-AX of the untreated PSH had a peak MM of 216 kDa and an arabinose to xylose (A/X) ratio of 0.20, WE-AX fragments from ball mill-pretreated PSH had a peak MM of 22 kDa and an A/X ratio of 0.31. Ball milling further drastically reduced the intrinsic viscosity of PSH extracts and their water-holding capacity. Prolonged treatment brought almost all AX (98%) in solution and yielded WE-AX fragments with an even higher A/X ratio (0.42) and a lower peak MM (11 kDa). While impact and jet milling of PSH equally led to significant reductions in particle size, these technologies only marginally affected the water extractability of PSH AX. This implies that ball milling affects PSH particles and their constituent molecules differently than impact and jet milling. PMID:19007123

  14. Enzymatic Hydrolysis of Wheat Arabinoxylan by a Recombinant "Minimal" Enzyme Cocktail Containing beta-Xylosidase and Novel endo-1,4-beta-Xylanase and alpha-L-Arabinofuranosidase Activities

    DEFF Research Database (Denmark)

    Sørensen, Hanne R.; Pedersen, Sven; Jørgensen, Christel T.; Meyer, Anne Boye Strunge

    2007-01-01

    -xylanase from H. insolens (Xyl III), and a GH3 beta-xylosidase from Trichoderma reesei (beta-xyl) released 322 mg of arabinose and 512 mg of xylose per gram of water-soluble wheat arabinoxylan dry matter and 150 mg of arabinose and 266 mg of xylose per gram of water-insoluble wheat arabinoxylan dry matter after...... wheat arabinoxylan substrates were hydrolyzed with a combination of Ultraflo L and Celluclast 1.5 L, two commercially available enzyme preparations produced by H. insolens and T. reesei....

  15. Glutathione Transferase (GST)-Activated Prodrugs

    OpenAIRE

    Andrea Calderan; Paolo Ruzza

    2013-01-01

    Glutathione transferase (formerly GST) catalyzes the inactivation of various electrophile-producing anticancer agents via conjugation to the tripeptide glutathione. Moreover, several data link the overexpression of some GSTs, in particular GSTP1-1, to both natural and acquired resistance to various structurally unrelated anticancer drugs. Tumor overexpression of these proteins has provided a rationale for the search of GST inhibitors and GST activated cytotoxic prodrugs. In the present review...

  16. Promiscuity and Selectivity in Phosphoryl Transferases

    OpenAIRE

    Barrozo, Alexandre

    2016-01-01

    Phosphoryl transfers are essential chemical reactions in key life processes, including energy production, signal transduction and protein synthesis. They are known for having extremely low reaction rates in aqueous solution, reaching the scale of millions of years. In order to make life possible, enzymes that catalyse phosphoryl transfer, phosphoryl transferases, have evolved to be tremendously proficient catalysts, increasing reaction rates to the millisecond timescale. Due to the nature of ...

  17. CATECHOL-O-METHYL TRANSFERASE AND SCHIZOPHRENIA

    OpenAIRE

    Šagud, Marina; Műck-Šeler, Dorotea; Mihaljević-Peleš, Alma; Vuksan-Ćusa, Bjanka; Živković, Maja; Jakovljević, Miro; Pivac, Nela

    2010-01-01

    Catechol-O-methyl transferase (COMT) is an enzyme involved in the degradation of dopamine. The most commonly examined polymorphism within the COMT gene is Val108/158Met polymorphism, which results in three to fourfold difference in COMT enzyme activity. It is particularely important in prefrontal cortex, since COMT activity is the most important regulator of the prefrontal dopamine function. Given the association between schizophrenia and decreased dopamine activity in the prefrontal corte...

  18. Diets high in resistent starch and arabinoxylan modulate digestion processes and SCFA pool size in the large intestine and faecal microbial composition in pigs

    DEFF Research Database (Denmark)

    Nielsen, Tina Skau; Lærke, Helle Nygaard; Theil, Peter Kappel;

    2014-01-01

    The effects of a high level of dietary fibre (DF) either as arabinoxylan (AX) or resistant starch (RS) on digestion processes, SCFA concentration and pool size in various intestinal segments and on the microbial composition in the faeces were studied in a model experiment with pigs. A total of...

  19. Influence of jet-cooking and pH on extraction and molecular weight of ß-glucan and arabinoxylan from barley (Hordeum vulgare Prowashonupana)

    Science.gov (United States)

    Food processing conditions may affect the solubility and molecular weight of beta-glucans and arabinoxylans in cereal products. This can dramatically affect the functional and physiological properties of the final products. Therefore, the purpose of the research was to explore the effects of jet-c...

  20. Concentrated Arabinoxylan but Not Concentrated Beta-Glucan in Wheat Bread Has Similar Effects on Postprandial Insulin as Whole-Grain Rye in Porto-arterial Catheterized Pigs

    DEFF Research Database (Denmark)

    Christensen, Kirstine Lykke; Hedemann, Mette Skou; Lærke, Helle Nygaard;

    2013-01-01

    The acute glycemic effects of concentrated dietary fibers (DF) versus whole-grain rye were studied in portoarterial catheterized pigs. Two white wheat breads with wheat arabinoxylan (AX) or oat beta-glucan (BG), two rye breads with intact rye kernels (RK) or milled rye (GR), and a low DF white...

  1. SIKLODEKSTRIN GLIKOSIL TRANSFERASE DAN PEMANFAATANNYA DALAM INDUSTRI [Cyclodextrin Glycosyl Transferase and its application in industries

    Directory of Open Access Journals (Sweden)

    Budiasih Wahyuntari

    2005-12-01

    Full Text Available Cyclodextrin glycosyl transferase (CGT-ase is mainly produced by Bacilli. Systematical name of the enzyme is E.C. 2.4.1.19 a-1,4 glucan-4-glycosyl transferase. The enzyme catalyzes hydrolysis of starch intramolecular, and intermolecular transglycosylation of a-1,4, glucan chains. Cyclodextrins are a-1,4 linked cyclic oligosaccharides resulting from enzymatic degradation of starch by cyclodextrin glycosyl transferase through untramolecular transglycosylation. The major cyclodextrins are made up of 6, 7 and 8 glucopyranose units which are known as a-, b-, and y-cyclodextrin. All CGT-ase catalyze three kinds of cyclodextrins, the proportion of the cyclodextrins depends on the enzyme source and reaction conditions. The intermolecular transglycosylation ability of the enzyme has been applied in transfering glycosyl residues into suitable acceptor. Transglycosylation by the enzymes have been tested to improve solubility of some flavonoids and to favor precipitation ci some glycosides.

  2. Ferulic acid release and 4-vinylguaiacol formation during brewing and fermentation: indications for feruloyl esterase activity in Saccharomyces cerevisiae.

    Science.gov (United States)

    Coghe, Stefan; Benoot, Koen; Delvaux, Filip; Vanderhaegen, Bart; Delvaux, Freddy R

    2004-02-11

    The release of ferulic acid and the subsequent thermal or enzymatic decarboxylation to 4-vinylguaiacol are inherent to the beer production process. Phenolic, medicinal, or clove-like flavors originating from 4-vinylguaiacol frequently occur in beer made with wheat or wheat malt. To evaluate the release of ferulic acid and the transformation to 4-vinylguaiacol, beer was brewed with different proportions of barley malt, wheat, and wheat malt. Ferulic acid as well as 4-vinylguaiacol levels were determined by HPLC at several stages of the beer production process. During brewing, ferulic acid was released at the initial mashing phase, whereas moderate levels of 4-vinylguaiacol were formed by wort boiling. Higher levels of the phenolic flavor compound were produced during fermentations with brewery yeast strains of the Pof(+) phenotype. In beer made with barley malt, ferulic acid was mainly released during the brewing process. Conversely, 60-90% of ferulic acid in wheat or wheat malt beer was hydrolyzed during fermentation, causing higher 4-vinylguaiacol levels in these beers. As cereal enzymes are most likely inactivated during wort boiling, the additional release of ferulic acid during fermentation suggests the activity of feruloyl esterases produced by brewer's yeast. PMID:14759156

  3. Hypoglycemic and Hypotensive Activity of a Root Extract of Smilax aristolochiifolia, Standardized on N-trans-Feruloyl-Tyramine

    Directory of Open Access Journals (Sweden)

    Carol Arely Botello Amaro

    2014-07-01

    Full Text Available The metabolic syndrome (MS is a condition consisting of various metabolic abnormalities that are risk factors for developing kidney failure, cardiovascular, vascular and cerebrovascular diseases, among others. The prevalence of this syndrome shows a marked increase. The aim of this study was to investigate the pharmacological effect of Smilax aristolochiifolia root on some components of MS and obtain some of the active principle using chromatographic techniques. The compound isolated was N-trans-feruloyl tyramine NTF (1, and its structure was determined by spectroscopic and spectrometric analyses. The whole extract and the standardized fractions were able to control the weight gain around 30%; the fraction rich in NTF was able to decrease the hypertriglyceridemia by 60%. The insulin resistance decreased by approximately 40%; the same happened with blood pressure, since the values of systolic and diastolic pressure fell on average 31% and 37% respectively, to levels comparable to normal value. The treatment also had an immunomodulatory effect on the low-grade inflammation associated with obesity, since it significantly decreased the relative production of pro-inflammatory cytokines regarding anti-inflammatory cytokines, both kidney and adipose tissue. Therefore it can be concluded that the extract and fractions of Smilax aristolochiifolia root with NTF are useful to counteract some symptoms of MS in animal models.

  4. The Wood Rot Ascomycete Xylaria polymorpha Produces a Novel GH78 Glycoside Hydrolase That Exhibits α-l-Rhamnosidase and Feruloyl Esterase Activities and Releases Hydroxycinnamic Acids from Lignocelluloses

    OpenAIRE

    Nghi, Do Huu; Bittner, Britta; Kellner, Harald; Jehmlich, Nico; Ullrich, René; Pecyna, Marek J.; Nousiainen, Paula; Sipilä, Jussi; Huong, Le Mai; Hofrichter, Martin; Liers, Christiane

    2012-01-01

    Soft rot (type II) fungi belonging to the family Xylariaceae are known to substantially degrade hardwood by means of their poorly understood lignocellulolytic system, which comprises various hydrolases, including feruloyl esterases and laccase. In the present study, several members of the Xylariaceae were found to exhibit high feruloyl esterase activity during growth on lignocellulosic materials such as wheat straw (up to 1,675 mU g−1) or beech wood (up to 80 mU g−1). Following the ester-clea...

  5. Variations in Content and Extractability of Durum Wheat (Triticum turgidum L. var durum Arabinoxylans Associated with Genetic and Environmental Factors

    Directory of Open Access Journals (Sweden)

    Daniela Sgrulletta

    2011-07-01

    Full Text Available Arabinoxylans (AX represent the most abundant components of non-starch polysaccharides in wheat, constituting about 70% of cell wall polysaccharides. An important property of AX is their ability to form highly viscous water solutions; this peculiarity has a significant impact on the technological characteristics of wheat and determines the physiologically positive influence in consumption. Durum wheat (Triticum turgidum L. var durum, the raw material for pasta production, is one of the most important crops in Italy. As part of a large project aimed at improving durum wheat quality, the characterization of the nutritional and technological aspects of whole grains was considered. Particular attention was addressed to identify the best suited genotypes for the production of innovative types of pasta with enhanced functional and organoleptic properties. The objective of the present study was to investigate the genetic variability of AX by examining a group of durum wheat genotypes collected at two localities in Italy for two consecutive years. The environmental influence on AX content and extractability was also evaluated. Variability in the AX fraction contents was observed; the results indicated that AX fractions of durum wheat grain can be affected by the genotype and environment characteristics and the different contribution of genotype and environment to total variation was evidenced. The genotype × environment (G × E interaction was significant for all examined traits, the variations due to G × E being lower than that of genotype or environment. The data and the statistical analysis allowed identification of the Italian durum wheat varieties that were consistently higher in total arabinoxilans; in addition, principal component analysis biplots illustrated that for arabinoxylan fractions some varieties responded differently in various environment climatic conditions.

  6. Variations in content and extractability of durum wheat (Triticum turgidum L. var durum) Arabinoxylans associated with genetic and environmental factors.

    Science.gov (United States)

    Ciccoritti, Roberto; Scalfati, Giulia; Cammerata, Alessandro; Sgrulletta, Daniela

    2011-01-01

    Arabinoxylans (AX) represent the most abundant components of non-starch polysaccharides in wheat, constituting about 70% of cell wall polysaccharides. An important property of AX is their ability to form highly viscous water solutions; this peculiarity has a significant impact on the technological characteristics of wheat and determines the physiologically positive influence in consumption. Durum wheat (Triticum turgidum L. var durum), the raw material for pasta production, is one of the most important crops in Italy. As part of a large project aimed at improving durum wheat quality, the characterization of the nutritional and technological aspects of whole grains was considered. Particular attention was addressed to identify the best suited genotypes for the production of innovative types of pasta with enhanced functional and organoleptic properties. The objective of the present study was to investigate the genetic variability of AX by examining a group of durum wheat genotypes collected at two localities in Italy for two consecutive years. The environmental influence on AX content and extractability was also evaluated. Variability in the AX fraction contents was observed; the results indicated that AX fractions of durum wheat grain can be affected by the genotype and environment characteristics and the different contribution of genotype and environment to total variation was evidenced. The genotype × environment (G × E) interaction was significant for all examined traits, the variations due to G × E being lower than that of genotype or environment. The data and the statistical analysis allowed identification of the Italian durum wheat varieties that were consistently higher in total arabinoxilans; in addition, principal component analysis biplots illustrated that for arabinoxylan fractions some varieties responded differently in various environment climatic conditions. PMID:21845095

  7. Glutathione S-transferases as risk factors in prostate cancer

    DEFF Research Database (Denmark)

    Autrup, Judith; Thomassen, L.H.; Olsen, J.H.;

    1999-01-01

    Glutathione S-transferases are enzymes involved in the metabolism of carcinogens and in the defence against reactive oxygen species. Genetic polymorphisms have been detected in glutathione S-transferases M1, T1 and P1, and some of these polymorphisms have been associated with an increased risk of...

  8. Effects of concentrated arabinoxylan and β-glucan compared with refined wheat and whole grain rye on glucose and appetite in subjects with the metabolic syndrome

    DEFF Research Database (Denmark)

    Hartvigsen, M L; Gregersen, S; Lærke, H N; Holst, J J; Knudsen, Knud Erik Bach; Hermansen, K

    2014-01-01

    grain on glucose, hormone responses and appetite in subjects with the metabolic syndrome (MetS). SUBJECTS/METHODS: Fifteen subjects with MetS participated in this acute, randomised, cross-over intervention study. The test breads provided 50 g of digestible carbohydrate: wheat bread with concentrated...... arabinoxylan (AX) or β-glucan (BG), rye bread with kernels (RK) and wheat bread (WB) as control. Blood samples were drawn for 270 min to determine glucose, insulin, glucagon-like peptide-1, glucose-dependent insulinotropic peptide (GIP) and ghrelin. Appetite score was addressed every 30 min. Ad libitum energy......BACKGROUND/OBJECTIVES: Several studies emphasise that arabinoxylan and β-glucan have more beneficial effects on glucose metabolism than low-dietary fibre (DF) meals. Less attention has been paid to the effects of concentrated DF compared with whole grain. We compared the effects of DF and whole...

  9. Inter-conversion of catalytic abilities in a bifunctional carboxyl/feruloyl-esterase from earthworm gut metagenome.

    Science.gov (United States)

    Vieites, José María; Ghazi, Azam; Beloqui, Ana; Polaina, Julio; Andreu, José M; Golyshina, Olga V; Nechitaylo, Taras Y; Waliczek, Agnes; Yakimov, Michail M; Golyshin, Peter N; Ferrer, Manuel

    2010-01-01

    Carboxyl esterases (CE) exhibit various reaction specificities despite of their overall structural similarity. In present study we have exploited functional metagenomics, saturation mutagenesis and experimental protein evolution to explore residues that have a significant role in substrate discrimination. We used an enzyme, designated 3A6, derived from the earthworm gut metagenome that exhibits CE and feruloyl esterase (FAE) activities with p-nitrophenyl and cinnamate esters, respectively, with a [(k(cat)/K(m))](CE)/[(k(cat)/K(m))](FAE) factor of 17. Modelling-guided saturation mutagenesis at specific hotspots (Lys(281), Asp(282), Asn(316) and Lys(317)) situated close to the catalytic core (Ser(143)/Asp(273)/His(305)) and a deletion of a 34-AA-long peptide fragment yielded mutants with the highest CE activity, while cinnamate ester bond hydrolysis was effectively abolished. Although, single to triple mutants with both improved activities (up to 180-fold in k(cat)/K(m) values) and enzymes with inverted specificity ((k(cat)/K(m))(CE)/(k(cat)/K(m))(FAE) ratio of ∼0.4) were identified, no CE inactive variant was found. Screening of a large error-prone PCR-generated library yielded by far less mutants for substrate discrimination. We also found that no significant changes in CE activation energy occurs after any mutation (7.3 to -5.6 J mol(-1)), whereas a direct correlation between loss/gain of FAE function and activation energies (from 33.05 to -13.7 J mol(-1)) was found. Results suggest that the FAE activity in 3A6 may have evolved via introduction of a limited number of 'hot spot' mutations in a common CE ancestor, which may retain the original hydrolytic activity due to lower restrictive energy barriers but conveys a dynamic energetically favourable switch of a second hydrolytic reaction. PMID:21255305

  10. Nomenclature for mammalian soluble glutathione transferases.

    Science.gov (United States)

    Mannervik, Bengt; Board, Philip G; Hayes, John D; Listowsky, Irving; Pearson, William R

    2005-01-01

    The nomenclature for human soluble glutathione transferases (GSTs) is extended to include new members of the GST superfamily that have been discovered, sequenced, and shown to be expressed. The GST nomenclature is based on primary structure similarities and the division of GSTs into classes of more closely related sequences. The classes are designated by the names of the Greek letters: Alpha, Mu, Pi, etc., abbreviated in Roman capitals: A, M, P, and so on. (The Greek characters should not be used.) Class members are distinguished by Arabic numerals and the native dimeric protein structures are named according to their subunit composition (e.g., GST A1-2 is the enzyme composed of subunits 1 and 2 in the Alpha class). Soluble GSTs from other mammalian species can be classified in the same manner as the human enzymes, and this chapter presents the application of the nomenclature to the rat and mouse GSTs. PMID:16399376

  11. Production and functional characterisation of arabinoxylan-oligosaccharides from wheat (Triticum aestivum L.) bran and psyllium (Plantago ovata Forsk) seed husk

    OpenAIRE

    Van Craeyveld, Valerie

    2009-01-01

    Prebiotica zijn voedingscomponenten die niet geabsorbeerd of gehydrolyse erd kunnen worden door enzymen van het gastrointestinaal stelsel, maar d ie in de dikke darm selectief gefermenteerd worden door bepaalde types b acteriën die een gezondheids-bevorderend effect uitoefenen op hun gasthe er. Naast inuline en fructo-oligosachariden, de meest bestudeerde prebio tica, vormen arabinoxylan-oligosachariden (AXOS) mogelijk een nieuwe kla sse van prebiotische componenten. AXOS kunnen uit tarwezeme...

  12. Biochemical genetics of glutathione-S-transferase in man.

    OpenAIRE

    Board, P G

    1981-01-01

    Glutathione-S-transferases from liver and erythrocytes have been separated by starch gel electrophoresis and localized by a specific staining procedure. The data suggest that the most active glutathione-S-transferases in liver are the products of two autosomal loci, GST1 and GST2. Both these loci are polymorphic, and there is evidence that a common null allele exists at the GST1 locus. The glutathione-S-transferase expressed in erythrocytes is the product of a third locus, GST3, and is not po...

  13. The Genetic Architecture of Murine Glutathione Transferases.

    Directory of Open Access Journals (Sweden)

    Lu Lu

    Full Text Available Glutathione S-transferase (GST genes play a protective role against oxidative stress and may influence disease risk and drug pharmacokinetics. In this study, massive multiscalar trait profiling across a large population of mice derived from a cross between C57BL/6J (B6 and DBA2/J (D2--the BXD family--was combined with linkage and bioinformatic analyses to characterize mechanisms controlling GST expression and to identify downstream consequences of this variation. Similar to humans, mice show a wide range in expression of GST family members. Variation in the expression of Gsta4, Gstt2, Gstz1, Gsto1, and Mgst3 is modulated by local expression QTLs (eQTLs in several tissues. Higher expression of Gsto1 in brain and liver of BXD strains is strongly associated (P < 0.01 with inheritance of the B6 parental allele whereas higher expression of Gsta4 and Mgst3 in brain and liver, and Gstt2 and Gstz1 in brain is strongly associated with inheritance of the D2 parental allele. Allele-specific assays confirmed that expression of Gsto1, Gsta4, and Mgst3 are modulated by sequence variants within or near each gene locus. We exploited this endogenous variation to identify coexpression networks and downstream targets in mouse and human. Through a combined systems genetics approach, we provide new insight into the biological role of naturally occurring variants in GST genes.

  14. Gelation of wheat arabinoxylans in the presence of Cu(+2) and in aqueous mixtures with cereal β-glucans.

    Science.gov (United States)

    Skendi, Adriana; Biliaderis, Costas G

    2016-07-15

    The effect of copper ions (0.63-0.16mM) on the rheological behavior of arabinoxylan (AX) aqueous solutions was investigated. Moreover, the influence of β-glucan addition (BG, 0.5-3% w/v) on the gelation of mixed AX/BG solutions with and without addition of the peroxidase/H2O2 was examined. Generally, gels formed with inclusion of transition metal-ions differed from those obtained by adding peroxidase/H2O2. Copper ions induced viscosity increase of the AX-solutions and form stronger thermoreversible gels with increasing ion-concentration; optimal gelation was at 15°C. For added β-glucan at levels >1%, the lower the concentration and the higher the molecular weight of β-glucan, the weaker the gelling ability of the mixed AX/BG system treated with peroxidase/H2O2. The polysaccharide-ratio affected both the gelling rate and the network melting temperature, with the β-glucan itself giving the strongest network. Calorimetry provided evidence for existence of β-glucan ordered domains in the mixed gel structures of AX/BG1, indicative of phase separation events. PMID:26948614

  15. Classification of wheat varieties based on structural features of arabinoxylans as revealed by endoxylanase treatment of flour and grain.

    Science.gov (United States)

    Ordaz-Ortiz, José Juan; Devaux, Marie-Françoise; Saulnier, Luc

    2005-10-19

    Arabinoxylans (AX) are cell wall polysaccharides of complex structure involved in many aspects of wheat flour end uses. The study of the variations of AX structure can lead to the identification of genes involved in their biosynthesis, and thus in the control of the various aspects of grain quality related to their presence. A method is proposed to identify AX variations directly in whole grain by enzymatic degradation. An endoxylanase from Trichoderma viride was used to extract AX from a collection of 20 wheat cultivars (Triticum aestivum L.). Enzymatic degradation products were analyzed by HPAEC and multivariate analysis techniques (principal component analysis, canonical correlation analysis, and cluster analysis) were applied to analyze chromatographic data. The method evidenced variations in the proportion of mono- and disubstitution of the xylan backbone by arabinose side chains, allowing classification of the different varieties according to the structural features of AX. A similar classification was obtained starting from flour or whole grain, indicating that the method was specific of AX from endosperm tissues. In conclusion, the method combining endoxylanase treatment of wheat grain and the analysis of degradation products, e.g., enzymatic fingerprinting, can be applied to collections of wheat cultivars, and possibly other cereals in order to establish quantitative trait loci related to the biosynthesis of AX. PMID:16218687

  16. Effect of heat and drought stress on the structure and composition of arabinoxylan and β-glucan in wheat grain.

    Science.gov (United States)

    Rakszegi, Marianna; Lovegrove, Alison; Balla, Krisztina; Láng, László; Bedő, Zoltán; Veisz, Ottó; Shewry, Peter R

    2014-02-15

    The effects of heat (H), drought (D) and H+D (from 12th day after heading for 15 days) on the dietary fiber content and composition (arabinoxylan (AX) and β-glucan) of three winter wheat varieties (Plainsman V, Mv Magma and Fatima 2) were determined. Results showed that H and D stress decreased the TKW, the β-glucan contents of the seeds and the quantity of the DP3+DP4 units, while the protein and AX contents increased. The highest amounts of AX and proteins were in the H+D stressed samples with heat stress also increasing the water extractability (WE) of the AX. However, while the content of AX content was generally increased by all stresses, drought stress had negative effect on the AX content of the drought tolerant Plainsman V. Fatima 2 behaved similarly to Plainsman V as regards to its drought tolerance, but was very sensitive to heat stress, while Mv Magma was the most resistant to heat stress. PMID:24507319

  17. Enzymatic Hydrolysis of Wheat Arabinoxylan by a Recombinant "Minimal" Enzyme Cocktail Containing beta-Xylosidase and Novel endo-1,4-beta-Xylanase and alpha-L-Arabinofuranosidase Activities

    DEFF Research Database (Denmark)

    Sørensen, Hanne R.; Pedersen, Sven; Jørgensen, Christel T.;

    2007-01-01

    This study describes the identification of the key enzyme activities required in a "minimal" enzyme cocktail able to catalyze hydrolysis of water-soluble and water-insoluble wheat arabinoxylan and whole vinasse, a fermentation effluent resulting from industrial ethanol manufacture from wheat. The...

  18. Steroid sulfatase and sulfuryl transferase activities in human brain tumors

    Czech Academy of Sciences Publication Activity Database

    Kříž, L.; Bičíková, M.; Mohapl, M.; Hill, M.; Černý, Ivan; Hampl, R.

    2008-01-01

    Roč. 109, č. 1 (2008), s. 31-39. ISSN 0960-0760 Institutional research plan: CEZ:AV0Z40550506 Keywords : dehydroepiandrosterone * steroid sulfatase * steroid sulfuryl transferase * brain Subject RIV: CC - Organic Chemistry Impact factor: 2.827, year: 2008

  19. Glutathione S-Transferase Isoenzymes from Streptomyces griseus

    OpenAIRE

    Dhar, Kajari; Dhar, Alok; Rosazza, John P. N.

    2003-01-01

    An inducible, cytosolic glutathione S-transferase (GST) was purified from Streptomyces griseus. GST isoenzymes with pI values of 6.8 and 7.9 used standard GST substrates including 1-chloro-2,4-dinitrobenzene. GST had subunit and native Mrs of 24 and 48, respectively, and the N-terminal sequence SMILXYWDIIRGLPAH.

  20. METAL-INDUCED INHIBITION OF GLUTATHIONE S-TRANSFERASES

    Science.gov (United States)

    The glutathione S-transferases comprise a group of multi-functional enzymes involved in the biotransformation/detoxication of a broad spectrum of hydrophobic compounds bearing an electrophilic center. The enzymes facilitate the nucleophilic attack of the -SH group of reduced glut...

  1. Phosphorylation and inhibition of. gamma. -glutamyl transferase activity by cAMP-dependent protein kinase

    Energy Technology Data Exchange (ETDEWEB)

    Kolesnichenko, L.S.; Chernov, N.N.

    1986-10-20

    It was shown that preparations of bovine kidney ..gamma..-glutamyl transferase of differing degrees of purity are phosphorylated by cAMP-dependent protein kinase. This is accompanied by a decrease in both the transferase and hydrolase activities of the enzyme. Consequently, ..gamma..-glutamyl transferase may serve as the substrate and target of the regulation of cAMP-dependent protein kinase.

  2. Lactobacillus fermentum CRL1446 Ameliorates Oxidative and Metabolic Parameters by Increasing Intestinal Feruloyl Esterase Activity and Modulating Microbiota in Caloric-Restricted Mice.

    Science.gov (United States)

    Russo, Matias; Fabersani, Emanuel; Abeijón-Mukdsi, María C; Ross, Romina; Fontana, Cecilia; Benítez-Páez, Alfonso; Gauffin-Cano, Paola; Medina, Roxana B

    2016-01-01

    The purpose of this study was to determine whether the administration of the feruloyl esterase (FE)-producing strain Lactobacillus fermentum CRL1446 enhances metabolic and oxidative parameters in caloric-restricted (CR) mice. Balb/c male mice were divided into ad libitum fed Group (ALF Group), CR diet Group (CR Group) and CR diet plus L. fermentum Group (CR-Lf Group). CR diet was administered during 45 days and CRL1446 strain was given in the dose of 10⁸ cells/mL/day/mouse. FE activity was determined in intestinal mucosa and content at Day 1, 20 and 45. Triglyceride, total cholesterol, glucose, thiobarbituric acid reactive substances (TBARS) levels and glutathione reductase activity were determined in plasma. Gut microbiota was evaluated by high-throughput sequencing of 16S rRNA gene amplicons. At Day 45, total intestinal FE activity in CR-Lf Group was higher (p = 0.020) than in CR and ALF groups and an improvement in both metabolic (reductions in triglyceride (p = 0.0025), total cholesterol (p = 0.005) and glucose (p < 0.0001) levels) and oxidative (decrease of TBARS levels and increase of plasmatic glutathione reductase activity (p = 0.006)) parameters was observed, compared to ALF Group. CR diet increased abundance of Bacteroidetes and CRL1446 administration increased abundance of Bifidobacterium and Lactobacillus genus. L. fermentun CRL1446 exerted a bifidogenic effect under CR conditions. PMID:27399766

  3. Effects of Resistant Starch and Arabinoxylan on Parameters Related to Large Intestinal and Metabolic Health in Pigs Fed Fat-Rich Diets

    DEFF Research Database (Denmark)

    Nielsen, Tina Skau; Theil, Peter Kappel; Purup, Stig;

    2015-01-01

    This study compared the effects of a resistant starch (RS)-rich, arabinoxylan (AX)-rich, or low-DF Western-style control diet (all high-fat) on large intestinal gene expression, adiposity, and glycemic response parameters in pigs. Animals were slaughtered after 3 weeks of treatment. Plasma butyrate...... concentration was higher following the high-DF diets, whereas plasma glucose, insulin, and insulin resistance increased after 3 weeks irrespective of diet. The mRNA abundance in the large intestine of genes involved in nutrient transport, immune response, and intestinal permeability was affected by segment...... (cecum, proximal, mid or distal colon) and some genes also by diet. In contrast, there was no diet-induced effect on adipose mRNA abundance or adipocyte size. Overall, a high level of RS or AX did not demonstrate strong beneficial effects on large intestinal gene expression as indicators of colonic...

  4. Molecular weight changes of arabinoxylans of wheat and rye incurred by the digestion processes in the upper gastrointestinal tract of pigs

    DEFF Research Database (Denmark)

    Legall, Maud; Eybye, Karin; Knudsen, Knud Erik Bach

    difference was found between the water extract and the ileal soluble phase when the wheat based diets were fed. The viscosity of the ileal phase is the highest after feeding the RAF diet. In conclusion, the MWw of AX from wheat diets is not changed during passage of the upper intestinal tract of pigs......Twenty cannulated-pigs were fed wheat flour (WFL), wheat whole grain (WWG), wheat aleurone flour (WAF) and rye aleurone flour (RAF) differing by their arabinoxylans (AX) proportions. After ileal collection of digesta (0800–1800), the soluble phase was extracted. The weight average molecular weight...... (MWw) of the ileal soluble phase and of the dietary water extracts were determined by high-performance size exclusion chromatography. The MWw value of the ileal soluble phase of digesta after feeding the RAF diet was reduced by approximately 25% compared with the dietary water extract whereas no...

  5. Monoclonal antibodies against human placental glutathione transferase (class pi).

    Science.gov (United States)

    Massoud, R; Lo Bello, M; De Stefano, E; Molino, A; Zelaschi, D; Federici, G

    1991-02-01

    Five monoclonal antibodies (MAbs) were produced in a mouse hybridoma system against human placental glutathione transferase (GST pi). Four of these monoclonal antibodies, named 461 to 464, were of immunoglobulin G class, whereas the monoclonal antibody 465 was of IgA class. All these MAbs specifically recognized the glutathione transferase from human placenta (class pi) showing no cross reactivity against the basic and the neutral forms of GST from human liver. When each MAb was incubated with the GST pi, no inhibition of enzymatic activity towards 1-chloro-2,4-dinitrobenzene was observed except for MAb 465 which showed a slight inhibition to a serial dilution of 1:128. PMID:1709614

  6. A randomised, double-blind, placebo controlled cross-over study to determine the gastrointestinal effects of consumption of arabinoxylan-oligosaccharides enriched bread in healthy volunteers

    Directory of Open Access Journals (Sweden)

    Walton Gemma E

    2012-06-01

    Full Text Available Abstract Background Prebiotics are food ingredients, usually non-digestible oligosaccharides, that are selectively fermented by populations of beneficial gut bacteria. Endoxylanases, altering the naturally present cereal arabinoxylans, are commonly used in the bread industry to improve dough and bread characteristics. Recently, an in situ method has been developed to produce arabinoxylan-oligosaccharides (AXOS at high levels in breads through the use of a thermophilic endoxylanase. AXOS have demonstrated potentially prebiotic properties in that they have been observed to lead to beneficial shifts in the microbiota in vitro and in murine, poultry and human studies. Methods A double-blind, placebo controlled human intervention study was undertaken with 40 healthy adult volunteers to assess the impact of consumption of breads with in situ produced AXOS (containing 2.2 g AXOS compared to non-endoxylanase treated breads. Volatile fatty acid concentrations in faeces were assessed and fluorescence in situ hybridisation was used to assess changes in gut microbial groups. Secretory immunoglobulin A (sIgA levels in saliva were also measured. Results Consumption of AXOS-enriched breads led to increased faecal butyrate and a trend for reduced iso-valerate and fatty acids associated with protein fermentation. Faecal levels of bifidobacteria increased following initial control breads and remained elevated throughout the study. Lactobacilli levels were elevated following both placebo and AXOS-breads. No changes in salivary secretory IgA levels were observed during the study. Furthermore, no adverse effects on gastrointestinal symptoms were reported during AXOS-bread intake. Conclusions AXOS-breads led to a potentially beneficial shift in fermentation end products and are well tolerated.

  7. Glutathione S-transferases in human liver cancer.

    OpenAIRE

    Hayes, P C; May, L.; Hayes, J. D.; Harrison, D J

    1991-01-01

    An immunohistochemical study of glutathione S-transferase (GST) expression in hepatocellular carcinoma and cholangiocarcinoma is described. Unlike most animal models of hepatic malignancy pi class GST was not consistently overexpressed in hepatocellular carcinoma. This tumour type either predominantly expressed alpha class GST or failed to express GST. By contrast, cholangiocarcinoma always expressed pi class GST, presumably reflecting the tissue of origin, since in human biliary epithelium p...

  8. Analysis of the glutathione S-transferase (GST) gene family

    OpenAIRE

    Nebert Daniel W; Vasiliou Vasilis

    2004-01-01

    Abstract The glutathione S-transferase (GST) gene family encodes genes that are critical for certain life processes, as well as for detoxication and toxification mechanisms, via conjugation of reduced glutathione (GSH) with numerous substrates such as pharmaceuticals and environmental pollutants. The GST genes are upregulated in response to oxidative stress and are inexplicably overexpressed in many tumours, leading to problems during cancer chemotherapy. An analysis of the GST gene family in...

  9. Purification and properties of glutathione transferase from Issatchenkia orientalis.

    OpenAIRE

    Tamaki, H.; Kumagai, H.; Tochikura, T

    1989-01-01

    Glutathione transferase (GST) (EC 2.5.1.18) was purified from a cell extract of Issatchenkia orientalis, and two GST isoenzymes were isolated. They had molecular weights of 37,500 and 40,000 and were designated GST Y-1 and GST Y-2, respectively. GST Y-1 and GST Y-2 gave single bands with molecular weights of 22,000 and 23,500, respectively, on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. GST Y-1 and GST Y-2 were immunologically distinguished from each other. GST Y-1 showed speci...

  10. 21 CFR 862.1315 - Galactose-1-phosphate uridyl transferase test system.

    Science.gov (United States)

    2010-04-01

    ... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1315 Galactose-1-phosphate uridyl transferase test system. (a)...

  11. A glutathione s-transferase confers herbicide tolerance in rice

    Directory of Open Access Journals (Sweden)

    Tingzhang Hu

    2014-07-01

    Full Text Available Plant glutathione S-transferases (GSTs have been a focus of attention due to their role in herbicide detoxification. OsGSTL2 is a glutathione S-transferase, lambda class gene from rice (Oryza sativa L.. Transgenic rice plants over-expressing OsGSTL2 were generated from rice calli by the use of an Agrobacterium transformation system, and were screened by a combination of hygromycin resistance, PCR and Southern blot analysis. In the vegetative tissues of transgenic rice plants, the over-expression of OsGSTL2 not only increased levels of OsGSTL2 transcripts, but also GST and GPX expression, while reduced superoxide. Transgenic rice plants also showed higher tolerance to glyphosate and chlorsulfuron, which often contaminate agricultural fields. The findings demonstrate the detoxification role of OsGSTL2 in the growth and development of rice plants. It should be possible to apply the present results to crops for developing herbicide tolerance and for limiting herbicide contamination in the food chain.

  12. Spontaneous and 5-azacytidine-induced reexpression of ornithine carbamoyl transferase in hepatoma cells.

    OpenAIRE

    Delers, A; Szpirer, J; Szpirer, C; Saggioro, D.

    1984-01-01

    Rat hepatoma cells that do not synthesize the hepatic enzyme ornithine carbamoyl transferase spontaneously give rise to producing cells at a low frequency. Reexpression of this differentiation trait is strongly increased by 5-azacytidine treatment, suggesting that hypermethylation plays a critical role in the impaired expression of the ornithine carbamoyl transferase gene in hepatoma cells.

  13. Arabinoxylan rice bran (MGN-3/Biobran) provides protection against whole-body γ-irradiation in mice via restoration of hematopoietic tissues

    International Nuclear Information System (INIS)

    The aim of the current study is to examine the protective effect of MGN-3 on overall maintenance of hematopoietic tissue after γ-irradiation. MGN-3 is an arabinoxylan from rice bran that has been shown to be a powerful antioxidant and immune modulator. Swiss albino mice were treated with MGN-3 prior to irradiation and continued to receive MGN-3 for 1 or 4 weeks. Results were compared with mice that received radiation (5 Gy γ rays) only, MGN-3 (40 mg/kg) only and control mice (receiving neither radiation nor MGN-3). At 1 and 4 weeks post-irradiation, different hematological, histopathological and biochemical parameters were examined. Mice exposed to irradiation alone showed significant depression in their complete blood count (CBC) except for neutrophilia. Additionally, histopathological studies showed hypocellularity of their bone marrow, as well as a remarkable decrease in splenic weight/relative size and in number of megakaryocytes. In contrast, pre-treatment with MGN-3 resulted in protection against irradiation-induced damage to the CBC parameters associated with complete bone marrow cellularity, as well as protection of the aforementioned splenic changes. Furthermore, MGN-3 exerted antioxidative activity in whole-body irradiated mice, and provided protection from irradiation-induced loss of body and organ weight. In conclusion, MGN-3 has the potential to protect progenitor cells in the bone marrow, which suggests the possible use of MGN-3/Biobran as an adjuvant treatment to counteract the severe adverse side effects associated with radiation therapy

  14. Recombinant baculovirus vectors expressing glutathione-S-transferase fusion proteins.

    Science.gov (United States)

    Davies, A H; Jowett, J B; Jones, I M

    1993-08-01

    Recombinant baculoviruses are a popular means of producing heterologous protein in eukaryotic cells. Purification of recombinant proteins away from the insect cell background can, however, remain an obstacle for many developments. Recently, prokaryotic fusion protein expression systems have been developed allowing single-step purification of the heterologous protein and specific proteolytic cleavage of the affinity tag moiety from the desired antigen. Here we report the introduction of these attributes to the baculovirus system. "Baculo-GEX" vectors enable baculovirus production of fusion proteins with the above advantages, but in a eukaryotic post-translational processing environment. Glutathione-S-transferase (GST) fusions are stable cytoplasmic proteins in insect cells and may therefore be released by sonication alone, avoiding the solubility problems and detergent requirements of bacterial systems. Thus large amounts of authentic antigen may be purified in a single, non-denaturing step. PMID:7763917

  15. From glutathione transferase to pore in a CLIC

    CERN Document Server

    Cromer, B A; Morton, C J; Parker, M W; 10.1007/s00249-002-0219-1

    2002-01-01

    Many plasma membrane chloride channels have been cloned and characterized in great detail. In contrast, very little is known about intracellular chloride channels. Members of a novel class of such channels, called the CLICs (chloride intracellular channels), have been identified over the last few years. A striking feature of the CLIC family of ion channels is that they can exist in a water- soluble state as well as a membrane-bound state. A major step forward in understanding the functioning of these channels has been the recent crystal structure determination of one family member, CLIC1. The structure confirms that CLICs are members of the glutathione S- transferase superfamily and provides clues as to how CLICs can insert into membranes to form chloride channels. (69 refs).

  16. Ghrelin O-Acyl Transferase: Bridging Ghrelin and Energy Homeostasis

    Directory of Open Access Journals (Sweden)

    Andrew Shlimun

    2011-01-01

    Full Text Available Ghrelin O-acyl transferase (GOAT is a recently identified enzyme responsible for the unique n-acyl modification of ghrelin, a multifunctional metabolic hormone. GOAT structure and activity appears to be conserved from fish to man. Since the acyl modification is critical for most of the biological actions of ghrelin, especially metabolic functions, GOAT emerged as a very important molecule of interest. The research on GOAT is on the rise, and several important results reiterating its significance have been reported. Notable among these discoveries are the identification of GOAT tissue expression patterns, effects on insulin secretion, blood glucose levels, feeding, body weight, and metabolism. Several attempts have been made to design and test synthetic compounds that can modulate endogenous GOAT, which could turn beneficial in favorably regulating whole body energy homeostasis. This paper will focus to provide an update on recent advances in GOAT research and its broader implications in the regulation of energy balance.

  17. Functional analysis and localisation of a delta-class glutathione S-transferase from Sarcoptes scabiei.

    Science.gov (United States)

    Pettersson, Eva U; Ljunggren, Erland L; Morrison, David A; Mattsson, Jens G

    2005-01-01

    The mite Sarcoptes scabiei causes sarcoptic mange, or scabies, a disease that affects both animals and humans worldwide. Our interest in S. scabiei led us to further characterise a glutathione S-transferase. This multifunctional enzyme is a target for vaccine and drug development in several parasitic diseases. The S. scabiei glutathione S-transferase open reading frame reported here is 684 nucleotides long and yields a protein with a predicted molecular mass of 26 kDa. Through phylogenetic analysis the enzyme was classified as a delta-class glutathione S-transferase, and our paper is the first to report that delta-class glutathione S-transferases occur in organisms other than insects. The recombinant S. scabiei glutathione S-transferase was expressed in Escherichia coli via three different constructs and purified for biochemical analysis. The S. scabiei glutathione S-transferase was active towards the substrate 1-chloro-2,4-dinitrobenzene, though the positioning of fusion partners influenced the kinetic activity of the enzyme. Polyclonal antibodies raised against S. scabiei glutathione S-transferase specifically localised the enzyme to the integument of the epidermis and cavities surrounding internal organs in adult parasites. However, some minor staining of parasite intestines was observed. No staining was seen in host tissues, nor could we detect any antibody response against S. scabiei glutathione S-transferase in sera from naturally S. scabiei infected dogs or pigs. Additionally, the polyclonal sera raised against recombinant S. scabiei glutathione S-transferase readily detected a protein from mites, corresponding to the predicted size of native glutathione S-transferase. PMID:15619514

  18. A search for synbiotics: effects of enzymatically modified arabinoxylan and Butyrivibrio fibrisolvens on short-chain fatty acids in the cecum content and plasma of rats

    DEFF Research Database (Denmark)

    Nielsen, Tina Skau; Jensen, Bent Borg; Purup, Stig;

    2016-01-01

    ) versus a Western-style control diet (WSD) low in dietary fiber with or without orally administrated Butyrivibrio fibrisolvens, a butyrate producer, on the SCFA pool in the cecal content and feces and the SCFA concentration in the blood of rats. The pool of acetate, butyrate and total SCFA was more than......Identification of dietary strategies to increase large intestinal production and absorption of short-chain fatty acids (SCFAs), especially butyrate, is of great interest due to the possible health promoting effects. We explored the effect of an enzymatically modified arabinoxylan-rich diet (EAXD...

  19. Glutathione Transferase GSTπ In Breast Tumors Evaluated By Three Techniques

    Directory of Open Access Journals (Sweden)

    Rafael Molina

    1993-01-01

    Full Text Available The glutathione transferases are involved in intracellular detoxification reactions. One of these, GSTπ, is elevated in some breast cancer cells, particularly cells selected for resistance to anticancer agents. We evaluated GSTπ expression in 60 human breast tumors by three techniques, immunohistochemistry, Northern hybridization, and Western blot analysis. There was a significant positive correlation between the three methods, with complete concordance seen in 64% of the tumors. There was strong, inverse relationship between GSTπ expression and steroid receptor status with all of the techniques utili zed. [n addition, there was a trend toward higher GSTπ expression in poorly differentiated tumors, but no correlation was found between tumor GSTπ content and DNA ploidy or %S-phase. GSTπ expression was also detected in adjacent benign breast tissue as well as infiltrating lymphocytes; this expression may contribute to GSTπ measurements using either Northern hybridization or Western blot analysis. These re sults suggest that immunohistochemistry is the method of choice for measuring GSTπ in breast tumors.

  20. Modulation of Rab GTPase function by a protein phosphocholine transferase.

    Science.gov (United States)

    Mukherjee, Shaeri; Liu, Xiaoyun; Arasaki, Kohei; McDonough, Justin; Galán, Jorge E; Roy, Craig R

    2011-09-01

    The intracellular pathogen Legionella pneumophila modulates the activity of host GTPases to direct the transport and assembly of the membrane-bound compartment in which it resides. In vitro studies have indicated that the Legionella protein DrrA post-translationally modifies the GTPase Rab1 by a process called AMPylation. Here we used mass spectrometry to investigate post-translational modifications to Rab1 that occur during infection of host cells by Legionella. Consistent with in vitro studies, DrrA-mediated AMPylation of a conserved tyrosine residue in the switch II region of Rab1 was detected during infection. In addition, a modification to an adjacent serine residue in Rab1 was discovered, which was independent of DrrA. The Legionella effector protein AnkX was required for this modification. Biochemical studies determined that AnkX directly mediates the covalent attachment of a phosphocholine moiety to Rab1. This phosphocholine transferase activity used CDP-choline as a substrate and required a conserved histidine residue located in the FIC domain of the AnkX protein. During infection, AnkX modified both Rab1 and Rab35, which explains how this protein modulates membrane transport through both the endocytic and exocytic pathways of the host cell. Thus, phosphocholination of Rab GTPases represents a mechanism by which bacterial FIC-domain-containing proteins can alter host-cell functions. PMID:21822290

  1. Glutathione S-transferase activity and glutathione S-transferase mu expression in subjects with risk for colorectal cancer.

    Science.gov (United States)

    Szarka, C E; Pfeiffer, G R; Hum, S T; Everley, L C; Balshem, A M; Moore, D F; Litwin, S; Goosenberg, E B; Frucht, H; Engstrom, P F

    1995-07-01

    The glutathione S-transferases (alpha, mu, and pi), a family of Phase II detoxication enzymes, play a critical role in protecting the colon mucosa by catalyzing the conjugation of dietary carcinogens with glutathione. We investigated the efficacy of using the glutathione S-transferase (GST) activity of blood lymphocytes and GST-mu expression as biomarkers of risk for colorectal cancer. GST activity was measured in the blood lymphocytes of control individuals (n = 67) and in the blood lymphocytes (n = 60) and colon tissue (n = 34) of individuals at increased risk for colon cancer. Total GST activity was determined spectrophotometrically with the use of 1-chloro-2,4-dinitrobenzene as a substrate. The ability to express the um subclass of GST was determined with the use of an ELISA. Although interindividual variability in the GST activity of blood lymphocytes was greater than 8-fold (range, 16.7-146.8 nmol/min/mg), the GST activity of blood lymphocytes and colon tissue within an individual was constant over time and was unrelated to sex, age, or race. The GST activity of blood lymphocytes from high-risk individuals was significantly lower than that of blood lymphocytes from control individuals (P GST-mu phenotype and risk for colorectal cancer. Blood lymphocytes from high-risk individuals unable to express GST-mu had lower levels of GST activity than did those from control subjects with the GST-mu null phenotype; however, this difference was significant in male subjects only (P GST activity of the two tissue types (Spearman's rank correlation, r = 0.87; P GST activity of blood lymphocytes may be used to identify high-risk individuals with decreased protection from this Phase II detoxication enzyme who may benefit from clinical trials evaluating GST modulators as chemopreventive agents for colorectal cancer. The GST activity of blood lymphocytes may also be used in colorectal cancer chemoprevention trials to monitor the responsiveness of colon tissue to regimens that

  2. Serum fucosyl transferase activity and serum fucose levels as diagnostic tools in malignancy.

    Directory of Open Access Journals (Sweden)

    Sen,Umi

    1983-12-01

    Full Text Available Glycoproteins play a significant role in neoplastic transformations. Both the levels of fucose and the activity of fucosyl transferase, which mediates the assembly of the oligosaccharide moieties of the glycoprotein chains, have been found to be elevated in neoplastic conditions. Since these elevations are common features of a variety of neoplastic cells, these two have been designated as non-specific markers of malignancy. In the present study, the fucose level and fucosyl transferase activity were determined in the sera of cancer patients and an attempt was made to establish a relationship between the two. It was found that both the fucose levels and fucosyl transferase activities showed considerable elevation in the five cancer groups studied, establishing them as useful diagnostic parameters. However, it was also observed that the rate of increased fucosyl transferase activity was not fully reflected in the resulting serum fucose levels in a few cases.

  3. 结构预测及分子对接方法研究几种水解产物对新阿魏酸酯酶的抑制作用%Structure Prediction and Molecular Docking Studies on the Inhibitory Effect of Several Hydrolyzate on a Novel Feruloyl Esterase

    Institute of Scientific and Technical Information of China (English)

    程凡升; 张茂秋; 程凡杰; 生吉萍; 陈婧雨; 郑鄢燕; 申琳

    2012-01-01

    Feruloyl esterases, one of the key enzymes in biomass degradation, show great prospects in food, textile, forge and pulp industries. In this study, a feruloyl esterase with low sequence identity was studied, which was originated from China Holstein cow rumen metagenomic library. Possible catalytic mechanisms were discussed based on conserved domains and tertiary structure modelling. Molecular docking studies show that the predicted affinity energy exhibits liner relations to experimental Vmax values and hydrogen bond may play a key role on catalytic efficiency. In addition, molecular docking study indicates that L-arabinobiose, D-xylose, D-glucose and trans-ferulic acid may have inhibitory effect on feruloyl esterase activity and the inhibition studies verify this hypothesis. This study may pave a way on rational design of feruloyl esterase and provide guidance on ferulic acid production from lignocellulose materials.%应用基于生物信息学的蛋白质结构预测方法,探讨了来源于荷斯坦奶牛瘤胃宏基因组文库的一个新型阿魏酸酯酶( FAE-SH1)的结构及可能的催化机制.同时通过对该酶的预测结构与4种模式底物的对接研究发现,实验所测得酶促反应动力参数Vmax与对接的亲和能存在线性关系,底物与酶形成的氢键可能是影响催化效率的关键因素.同时,本研究发现D-木糖、L-阿拉伯糖、D-葡萄糖及阿魏酸能够对FAE-SH1的水解反应产生抑制作用,并对其进行了验证.

  4. Glutathione S-transferase isoenzymes in relation to their role in detoxification of xenobiotics.

    OpenAIRE

    Vos, R.M.E.

    1989-01-01

    The glutathione S-transferases (GST) are a family of isoenzymes serving a major part in the biotransformation of many reactive compounds. The isoenzymes from rat, man and mouse are divided into three classes, alpha, mu and pi, on the basis of similar structural and enzymatic properties.The main function of the glutathione S-transferases isthe catalysisof the conjugation of electrophilic, hydrophobic compounds with the tripeptide glutathione (GSH). In addition, some of the isoenzymes are capab...

  5. Serum fucosyl transferase activity and serum fucose levels as diagnostic tools in malignancy.

    OpenAIRE

    Sen,Umi; Guha,Subhas; Chowdhury, J Roy

    1983-01-01

    Glycoproteins play a significant role in neoplastic transformations. Both the levels of fucose and the activity of fucosyl transferase, which mediates the assembly of the oligosaccharide moieties of the glycoprotein chains, have been found to be elevated in neoplastic conditions. Since these elevations are common features of a variety of neoplastic cells, these two have been designated as non-specific markers of malignancy. In the present study, the fucose level and fucosyl transferase activi...

  6. Biological role of sialosyl transferase activity in rat brain

    International Nuclear Information System (INIS)

    The purpose of this dissertation is to obtain new evidence that will support or refute the existence of an ecto sialosyltransferse activity (STase) that has been described in the synaptic plasma membrane (SPM). This STase has been proposed to transfer sialic acid (NANA) to endogenous SPM gangliosides. Preparations of rat brain synaptosomes were assayed for STase by incubation with CMP-(14C)NANA, and measuring radioactivity transferred to the endogenous gangliosides. The activity was found to be 0.84 pmoles NANA transferred per mg protein per hour. The product specificity for STase was determined by the incorporation of label into individual ganglioside species. Subfractions were produced from rat brain that were enriched in Golgi membranes, synaptosomes, and SPM as judged by EM morphology and marker enzymes. The Golgi fraction had over 3 fold greater STase activity than synaptosomes, while SPM were enriched 2.5 fold over the synaptosomes from which they came. The labeling pattern of endogenous gangliosides was quite different by the Golgi STase. An unknown compound in the ganglioside extracts was specifically labeled, but gangliosides were not labeled with specificity by the Golgi transferase. The synaptosomal and SPM labeling patterns were identical and were characterized by GD3 specificity. Therefore the STase of SPM is not due to Golgi contamination. Intact neurons were assayed for STase by the use of brain cortical slices. Slices incubated that labeled CMP-NANA (available for cell surface reactions) produced the GD3-specific labeling pattern. These results suggest that the GD3-specific sialosyltransferase is a cell surface ecto-enzyme

  7. Analysis of Arabidopsis glutathione-transferases in yeast.

    Science.gov (United States)

    Krajewski, Matthias P; Kanawati, Basem; Fekete, Agnes; Kowalski, Natalie; Schmitt-Kopplin, Philippe; Grill, Erwin

    2013-07-01

    The genome of Arabidopsis thaliana encodes 54 functional glutathione transferases (GSTs), classified in seven clades. Although plant GSTs have been implicated in the detoxification of xenobiotics, such as herbicides, extensive redundancy within this large gene family impedes a functional analysis in planta. In this study, a GST-deficient yeast strain was established as a system for analyzing plant GSTs that allows screening for GST substrates and identifying substrate preferences within the plant GST family. To this end, five yeast genes encoding GSTs and GST-related proteins were simultaneously disrupted. The resulting yeast quintuple mutant showed a strongly reduced conjugation of the GST substrates 1-chloro-2,4-dinitrobenzene (CDNB) and 4-chloro-7-nitro-2,1,3-benzoxadiazole (NBD-Cl). Consistently, the quintuple mutant was hypersensitive to CDNB, and this phenotype was complemented by the inducible expression of Arabidopsis GSTs. The conjugating activity of the plant GSTs was assessed by in vitro enzymatic assays and via analysis of exposed yeast cells. The formation of glutathione adducts with dinitrobenzene was unequivocally verified by stable isotope labeling and subsequent accurate ultrahigh-resolution mass spectrometry (ICR-FTMS). Analysis of Arabidopsis GSTs encompassing six clades and 42 members demonstrated functional expression in yeast by using CDNB and NBD-Cl as model substrates. Subsequently, the established yeast system was explored for its potential to screen the Arabidopsis GST family for conjugation of the fungicide anilazine. Thirty Arabidopsis GSTs were identified that conferred increased levels of glutathionylated anilazine. Efficient anilazine conjugation was observed in the presence of the phi, tau, and theta clade GSTs including AtGSTF2, AtGSTF4, AtGSTF6, AtGSTF8, AtGSTF10, and AtGSTT2, none of which had previously been known to contribute to fungicide detoxification. ICR-FTMS analysis of yeast extracts allowed the simultaneous detection and

  8. An efficient arabinoxylan-debranching α-L-arabinofuranosidase of family GH62 from Aspergillus nidulans contains a secondary carbohydrate binding site.

    Science.gov (United States)

    Wilkens, Casper; Andersen, Susan; Petersen, Bent O; Li, An; Busse-Wicher, Marta; Birch, Johnny; Cockburn, Darrell; Nakai, Hiroyuki; Christensen, Hans E M; Kragelund, Birthe B; Dupree, Paul; McCleary, Barry; Hindsgaul, Ole; Hachem, Maher Abou; Svensson, Birte

    2016-07-01

    An α-L-arabinofuranosidase of GH62 from Aspergillus nidulans FGSC A4 (AnAbf62A-m2,3) has an unusually high activity towards wheat arabinoxylan (WAX) (67 U/mg; k cat = 178/s, K m = 4.90 mg/ml) and arabinoxylooligosaccharides (AXOS) with degrees of polymerisation (DP) 3-5 (37-80 U/mg), but about 50 times lower activity for sugar beet arabinan and 4-nitrophenyl-α-L-arabinofuranoside. α-1,2- and α-1,3-linked arabinofuranoses are released from monosubstituted, but not from disubstituted, xylose in WAX and different AXOS as demonstrated by NMR and polysaccharide analysis by carbohydrate gel electrophoresis (PACE). Mutants of the predicted general acid (Glu(188)) and base (Asp(28)) catalysts, and the general acid pK a modulator (Asp(136)) lost 1700-, 165- and 130-fold activities for WAX. WAX, oat spelt xylan, birchwood xylan and barley β-glucan retarded migration of AnAbf62A-m2,3 in affinity electrophoresis (AE) although the latter two are neither substrates nor inhibitors. Trp(23) and Tyr(44), situated about 30 Å from the catalytic site as seen in an AnAbf62A-m2,3 homology model generated using Streptomyces thermoviolaceus SthAbf62A as template, participate in carbohydrate binding. Compared to wild-type, W23A and W23A/Y44A mutants are less retarded in AE, maintain about 70 % activity towards WAX with K i of WAX substrate inhibition increasing 4-7-folds, but lost 77-96 % activity for the AXOS. The Y44A single mutant had less effect, suggesting Trp(23) is a key determinant. AnAbf62A-m2,3 seems to apply different polysaccharide-dependent binding modes, and Trp(23) and Tyr(44) belong to a putative surface binding site which is situated at a distance of the active site and has to be occupied to achieve full activity. PMID:26946172

  9. The Influence of Prebiotic Arabinoxylan Oligosaccharides on Microbiota Derived Uremic Retention Solutes in Patients with Chronic Kidney Disease: A Randomized Controlled Trial

    Science.gov (United States)

    Evenepoel, Pieter; de Loor, Henriette; Delcour, Jan A.; Courtin, Christophe M.; Kuypers, Dirk; Augustijns, Patrick; Verbeke, Kristin; Meijers, Björn

    2016-01-01

    The colonic microbial metabolism is a key contributor to uremic retention solutes accumulating in patients with CKD, relating to adverse outcomes and insulin resistance. Whether prebiotics can reduce intestinal generation of these microbial metabolites and improve insulin resistance in CKD patients not yet on dialysis remains unknown. We performed a randomized, placebo-controlled, double-blind, cross-over study in 40 patients with eGFR between 15 and 45 ml/min/1.73 m2. Patients were randomized to sequential treatment with prebiotic arabinoxylan oligosaccharides (AXOS) (10 g twice daily) and maltodextrin for 4 weeks, or vice versa, with a 4-week wash-out period between both intervention periods. Serum levels and 24h urinary excretion of p-cresyl sulfate, p-cresyl glucuronide, indoxyl sulfate, trimethylamine N-oxide and phenylacetylglutamine were determined at each time point using liquid chromatography—tandem mass spectrometry. In addition, insulin resistance was estimated by the homeostatic model assessment (HOMA-IR). A total of 39 patients completed the study. We observed no significant effect of AXOS on serum p-cresyl sulfate (P 0.42), p-cresyl glucuronide (P 0.59), indoxyl sulfate (P 0.70) and phenylacetylglutamine (P 0.41) and a small, albeit significant decreasing effect on serum trimethylamine N-oxide (P 0.04). There were neither effect of AXOS on 24h urinary excretion of p-cresyl sulfate (P 0.31), p-cresyl glucuronide (P 0.23), indoxyl sulfate (P 0.87) and phenylacetylglutamine (P 0.43), nor on 24h urinary excretion of trimethylamine N-oxide (P 0.97). In addition, we observed no significant change in HOMA-IR (P 0.93). In conclusion, we could not demonstrate an influence of prebiotic AXOS on microbiota derived uremic retention solutes and insulin resistance in patients with CKD not yet on dialysis. Further study is necessary to elucidate whether prebiotic therapy with other characteristics, higher cumulative exposure or in different patient populations may

  10. Glucomannan synthesis in pea epicotyls: the mannose and glucose transferases.

    Science.gov (United States)

    Piro, G; Zuppa, A; Dalessandro, G; Northcote, D H

    1993-01-01

    Membrane fractions and digitonin-solubilized enzymes prepared from stem segments isolated from the third internode of etiolated pea seedlings (Pisum sativum L. cv. Alaska) catalyzed the synthesis of a beta-1,4-[14C]mannan from GDP-D-[U-14C]-mannose, a mixed beta-1,3- and beta-1,4-[14C]glucan from GDP-D-[U-14C]-glucose and a beta-1,4-[14C]-glucomannan from both GDP-D-[U-14C]mannose and GDP-D-[U-14C]glucose. The kinetics of the membrane-bound and soluble mannan and glucan synthases were determined. The effects of ions, chelators, inhibitors of lipid-linked saccharides, polyamines, polyols, nucleotides, nucleoside-diphosphate sugars, acetyl-CoA, group-specific chemical probes, phospholipases and detergents on the membrane-bound mannan and glucan synthases were investigated. The beta-glucan synthase had different properties from other preparations which bring about the synthesis of beta-1,3-glucans (callose) and mixed beta-1,3- and beta-1,4- glucans and which use UDP-D-glucose as substrate. It also differed from xyloglucan synthase because in the presence of several concentrations of UDP-D-xylose in addition to GDP-D-glucose no xyloglucan was formed. Using either the membrane-bound or the soluble mannan synthase, GDP-D-glucose acted competitively in the presence of GDP-D-mannose to inhibit the incorporation of mannose into the polymer. This was not due to an inhibition of the transferase activity but was a result of the incorporation of glucose residues from GDP-D-glucose into a glucomannan. The kinetics and the composition of the synthesized glucomannan depended on the ratio of the concentrations of GDP-D-glucose and GDP-D-mannose that were available. Our data indicated that a single enzyme has an active centre that can use both GDP-D-mannose and GDP-D-glucose to bring about the synthesis of the heteropolysaccharide. PMID:7685647

  11. IDENTIFYING GENES CONTROLLING FERULATE CROSS-LINKING FORMATION IN GRASS CELL WALLS

    Energy Technology Data Exchange (ETDEWEB)

    de O Buanafina, Marcia Maria

    2013-10-16

    formation or genes encoding transcription factors that control feruloylation. So it will require further investigations to confirm if we have a mutation on the ferulloyltransferase gene(s). We have also identified severe phenotypes which showed a significant change in the level of cell wall ferulates and sugars and have not survived. As this genotype did not reach flowering stage there was no seed production and so further analysis could not be done. 3. Candidate Gene Approach: Because of the likely long time expected to generate and identify candidate with mutation(s) on the feruloyltransferase gene, from our screening, we have in addition taken a bioinformatics approach in order to try to identify candidates gene(s) involved in feruloylation. Homologues of the rice feruloyl transferase genes belonging to Pfam PF02458 family were identified in Brachypodium distachyon by blasting EST sequences of putative rice arabinoxylan feruloyl transferase genes against Brachypodium and homologous sequences identified were tested for their expression level in Brachypodium. Sequences of the two Brachypodium genes, which showed highest expression and similarity to rice sequences, were used to design primers for construction of RNAi and over-expression vectors. These were transformed into Brachypodium using Agrobacterium transformation and plants generated have been analyzed for levels of cell wall ferulates and diferulates over generations T0 to T2 or T3. Our data shows a significant reduction if ferulates monomers and dimers from plants generated from RNAi::BdAT2 over 2-3 generations indicating that this gene might be a positive candidate for feruloylation in Brachypodium. However when BdAT2 was up regulated there was not much increase in the level of ferulates as would be expected. This lack of effect on the level of cell wall ferulates could be due to the CaMV::35S promoter used to drive the expression of the putative BdAT2 gene. We have shown previously that Aspergillus FAEA

  12. Alteration of glutathione S-transferase properties during the development of Micromelalopha troglodyta larvae (Lepidoptera: Notodontidae)

    Institute of Scientific and Technical Information of China (English)

    TANG Fang; ZHANG Xiu-bo; LIU Yu-sheng; GAO Xi-wu

    2011-01-01

    Micromelalopha troglodyta (Graeser) is an important pest ofpoplar in China. Glutathione S-transferases (GSTs) are known to beresponsible for adaptation mechanisms of M. Troglodyta. The activitiesand kinetic constants of glutathione S-transferases in M. Troglodyta werestudied. Significant differences in glutathione S-transferase activity andkinetic characteristics were observed among five instars of M. Troglodytalarvae. Furthermore, the inhibition of glutathione S-transferase activity infive instars by 24 inhibitors was conducted. The results show the inhibi-tion of GST activity of different instars by 24 inhibitors was different.For GST activity in the 1st instar chlorpyrifos, lambda-cyhalothrin,endosulfan, abamectin, fipronil and pyridaben were the best inhibitorstested, and for GST activity in the 2nd instar, tannic acid and quercetinwere the most potent inhibitors tested, and for GST activity in the 3rdinstar, the inhibitory effects of quercetin, chlorpyrifos andlambda-cyhalothrin were the highest, and for GST activity in the 4thinstar, quercetin and lambda-cyhalothrin were the best inhibitors, and theinhibitory effect of pboxim was the highest for GST activity in the 5thinstar. Our results show that glutathione S-transferases in different iustarsare qualitatively different in isozyme composition and thus different insensitivity to inhibitors.

  13. Disparate Metabolic Responses in Mice Fed a High-Fat Diet Supplemented with Maize-Derived Non-Digestible Feruloylated Oligo- and Polysaccharides Are Linked to Changes in the Gut Microbiota.

    Directory of Open Access Journals (Sweden)

    Junyi Yang

    Full Text Available Studies have suggested links between colonic fermentation of dietary fibers and improved metabolic health. The objectives of this study were to determine if non-digestible feruloylated oligo- and polysaccharides (FOPS, a maize-derived dietary fiber, could counteract the deleterious effects of high-fat (HF feeding in mice and explore if metabolic benefits were linked to the gut microbiota. C57BL/6J mice (n = 8/group were fed a low-fat (LF; 10 kcal% fat, HF (62 kcal% fat, or HF diet supplemented with FOPS (5%, w/w. Pronounced differences in FOPS responsiveness were observed: four mice experienced cecal enlargement and enhanced short chain fatty acid production, indicating increased cecal fermentation (F-FOPS. Only these mice displayed improvements in glucose metabolism compared with HF-fed mice. Blooms in the gut microbial genera Blautia and Akkermansia were observed in three of the F-FOPS mice; these shifts were associated with reductions in body and adipose tissue weights compared with the HF-fed control mice. No improvements in metabolic markers or weights were detected in the four mice whose gut microbiota did not respond to FOPS. These findings demonstrate that FOPS-induced improvements in weight gain and metabolic health in mice depended on the ability of an individual's microbiota to ferment FOPS.

  14. The psychrophilic bacterium Pseudoalteromonas halosplanktis TAC125 possesses a gene coding for a cold-adapted feruloyl esterase activity that shares homology with esterase enzymes from gamma-proteobacteria and yeast.

    Science.gov (United States)

    Aurilia, Vincenzo; Parracino, Antonietta; Saviano, Michele; Rossi, Mose'; D'Auria, Sabato

    2007-08-01

    The complete genome of the psychrophilic bacteria Pseudoalteromonas haloplanktis TAC 125, recently published, owns a gene coding for a putative esterase activity corresponding to the ORF PSHAa1385, also classified in the Carbohydrate Active Enzymes database (CAZY) belonging to family 1 of carbohydrate esterase proteins. This ORF is 843 bp in length and codes for a protein of 280 amino acid residues. In this study we characterized and cloned the PSHAa1385 gene in Escherichia coli. We also characterized the recombinant protein by biochemical and biophysical methodologies. The PSHAa1385 gene sequence showed a significant homology with several carboxyl-esterase and acetyl-esterase genes from gamma-proteobacteria genera and yeast. The recombinant protein exhibited a significant activity towards pNP-acetate, alpha-and beta-naphthyl acetate as generic substrates, and 4-methylumbelliferyl p-trimethylammonio cinnamate chloride (MUTMAC) as a specific substrate, indicating that the protein exhibits a feruloyl esterase activity that it is displayed by similar enzymes present in other organisms. Finally, a three-dimensional model of the protein was built and the amino acid residues involved in the catalytic function of the protein were identified. PMID:17543477

  15. Nuclear translocation of glutathione transferase omega is a progression marker in Barrett's esophagus

    DEFF Research Database (Denmark)

    Piaggi, Simona; Marchi, Santino; Ciancia, Eugenio;

    2009-01-01

    fraction of BE patients. This study was aimed to investigate the possible role of glutathione-S-transferase-omega 1 (GSTO1), a recently discovered member of the glutathione-S-transferase family, as a progression marker in the Barrett's disease in order to improve the diagnosis of NiN in BE and to......N were equally divided between nuclear, cytoplasmic and diffuse staining (2 each, respectively). Experiments in vitro showed that in human HeLa cancer cells, GSTO1 translocates into the nucleus as a consequence of heath shock. These findings suggested that the nuclear translocation of glutathione...

  16. Origin and evolution of the Peptidyl Transferase Center from proto-tRNAs

    Directory of Open Access Journals (Sweden)

    Sávio T. Farias

    2014-01-01

    Full Text Available We tested the hypothesis of Tamura (2011 [3] that molecules of tRNA gave origin to ribosomes, particularly to the Peptidyl Transferase Center (PTC of the 23S ribosomal RNA. We reconstructed the ancestral sequences from all types of tRNA and compared them in their sequences with the current PTC of 23S ribosomal RNA from different organisms. We built an ancestral sequence of proto-tRNAs that showed a remarkable overall identity of 50.53% with the catalytic site of PTC. We conclude that the Peptidyl Transferase Center was indeed originated by the fusion of ancestral sequences of proto-tRNA.

  17. The role of human demographic history in determining the distribution and frequency of transferase-deficient galactosaemia mutations

    NARCIS (Netherlands)

    J.M. Flanagan; G. McMahon; S.H. Brendan Shia; P. Fitzpatrick; O. Tighe; C. O'Neill; P. Briones; L. Gort; L. Kozak; A. Magee; E. Naughten; B. Radomyska; M. Schwartz; J.S. Shin; W.M. Strobl; L.A. Tyfield; H.R. Waterham; H. Russell; G. Bertorelle; J.K.V. Reichardt; P.D. Mayne; D.T. Croke

    2010-01-01

    Classical or transferase-deficient galactosaemia is an inherited metabolic disorder caused by mutation in the human Galactose-1-phosphate uridyl transferase (GALT) gene. Of some 170 causative mutations reported, fewer than 10% are observed in more than one geographic region or ethnic group. To bette

  18. Purification of human hepatic glutathione S-transferases and the development of a radioimmunoassay for their measurement in plasma

    International Nuclear Information System (INIS)

    A purification scheme is described for six human hepatic glutathione S-transferases from a single liver. Five of the transferases comprised Ya monomers and had a molecular mass of 44000. The remaining enzyme comprised Yb monomers and had a molecular mass of 47000. Data are presented demonstrating that there are at least two distinct Ya monomers. A radioimmunoassay has been developed that has sufficient precision and sensitivity to allow direct measurement of glutathione S-transferase concentrations in unextracted plasma. A comparison of aminotransferase and glutathione S-transferase levels, in three patients who had taken a paracetamol overdose, indicated that glutathione S-transferase measurements provided a far more sensitive index of hepatocellular integrity than the more conventional aminotransferase measurements. (Auth.)

  19. Purification of human hepatic glutathione S-transferases and the development of a radioimmunoassay for their measurement in plasma

    Energy Technology Data Exchange (ETDEWEB)

    Hayes, J.D.; Gilligan, D.; Beckett, G.J. (Edinburgh Univ. (UK). Dept. of Clinical Chemistry); Chapman, B.J. (Royal Infirmary, Edinburgh (UK))

    1983-10-31

    A purification scheme is described for six human hepatic glutathione S-transferases from a single liver. Five of the transferases comprised Ya monomers and had a molecular mass of 44000. The remaining enzyme comprised Yb monomers and had a molecular mass of 47000. Data are presented demonstrating that there are at least two distinct Ya monomers. A radioimmunoassay has been developed that has sufficient precision and sensitivity to allow direct measurement of glutathione S-transferase concentrations in unextracted plasma. A comparison of aminotransferase and glutathione S-transferase levels, in three patients who had taken a paracetamol overdose, indicated that glutathione S-transferase measurements provided a far more sensitive index of hepatocellular integrity than the more conventional aminotransferase measurements.

  20. Meat consumption, N-acetyl transferase 1 and 2 polymorphism and risk of breast cancer, in Danish postmenopausal women

    DEFF Research Database (Denmark)

    Egeberg, Rikke; Olsen, Anja; Autrup, Herman;

    2008-01-01

    total meat intake and red meat intake and breast cancer risk were confined to intermediate/fast N-acetyl transferase 2 acetylators (P-interaction=0.03 and 0.04). Our findings support an association between meat consumption and breast cancer risk and that N-acetyl transferase 2 polymorphism has a......The aim of this study was to investigate whether polymorphisms in N-acetyl transferase 1 and 2 modify the association between meat consumption and risk of breast cancer. A nested case-control study was conducted among 24697 postmenopausal women included in the 'Diet, Cancer and Health' cohort study...... increment in intake. Compared with slow acetylators, the IRR (95% confidence interval) among fast N-acetyl transferase 1 acetylators was 1.43 (1.03-1.99) and 1.13 (0.83-1.54) among intermediate/fast N-acetyl transferase 2 acetylators. Interaction analyses revealed that the positive associations between...

  1. In vitro fermentation patterns of rice bran components by human gut microbiota

    Science.gov (United States)

    Rice bran is a rich source of bioactive components that can promote gastrointestinal health. However, bran is removed during polishing. Among those, feruloylated arabinoxylan oligosaccharides (FAXO) and rice bran polyphenolics (RBPP) are hypothesized to have positive impacts on human gut microbiota ...

  2. EFSA Panel on Dietetic Products, Nutrition and Allergies (NDA); Scientific Opinion on the substantiation of health claims related to arabinoxylan produced from wheat endosperm and reduction of post-prandial glycaemic responses (ID 830) pursuant to Article 13(1) of Regulation (EC) No 1924/2006

    DEFF Research Database (Denmark)

    Tetens, Inge

    Member States or directly from stakeholders. The food constituent that is the subject of the health claim is “wheat grain fibre”. From the references and information provided, the Panel assumes that the food constituent that is responsible for the claimed effect is arabinoxylan from wheat endosperm. The...... claim in relation to arabinoxylan produced from wheat endosperm and reduction of post-prandial glycaemic responses. The scientific substantiation is based on the information provided by the Member States in the consolidated list of Article 13 health claims and references that EFSA has received from...... responses. The Panel considers that the reduction of post-prandial glycaemic responses (as long as post-prandial insulinaemic responses are not disproportionally increased) may be a beneficial physiological effect. In weighing the evidence, the Panel took into account that one well-designed intervention...

  3. The role of glutathione S-transferase and claudin-1 gene polymorphisms in contact sensitization

    DEFF Research Database (Denmark)

    Ross-Hansen, K; Linneberg, A; Johansen, J D;

    2013-01-01

    BACKGROUND: Contact sensitization is frequent in the general population and arises from excessive or repeated skin exposure to chemicals and metals. However, little is known about its genetic susceptibility. OBJECTIVES: To determine the role of polymorphisms of glutathione S-transferase (GST) genes...

  4. Effect of glutathione S-transferases on the survival of patients with acute myeloid leukaemia

    DEFF Research Database (Denmark)

    Autrup, Judith; Hokland, Peter; Pedersen, Lars;

    2002-01-01

    The objective of the study was to investigate the effect of genetic polymorphisms in glutathione S-transferases (GST) on the survival of acute myeloid leukaemia patients receiving adriamycin induction therapy. A total of 89 patients were included in the study. Patients who carried at least one GSTM...

  5. 21 CFR 573.130 - Aminoglycoside 3′-phospho- transferase II.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Aminoglycoside 3â²-phospho- transferase II. 573.130 Section 573.130 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN... genetically modified cotton, oilseed rape, and tomatoes in accordance with the following prescribed...

  6. Development of isoform-specific sensors of polypeptide GalNAc-transferase activity

    DEFF Research Database (Denmark)

    Song, Lina; Bachert, Collin; Schjoldager, Katrine T; Clausen, Henrik; Linstedt, Adam D

    2014-01-01

    Humans express up to 20 isoforms of GalNAc-transferase (herein T1-T20) that localize to the Golgi apparatus and initiate O-glycosylation. Regulation of this enzyme family affects a vast array of proteins transiting the secretory pathway and diseases arise upon misregulation of specific isoforms...

  7. Plasmodium spp. membrane glutathione S-transferases: detoxification units and drug targets

    Directory of Open Access Journals (Sweden)

    Andreas Martin Lisewski

    2014-10-01

    Full Text Available Membrane glutathione S-transferases from the class of membrane-associated proteins in eicosanoid and glutathione metabolism (MAPEG form a superfamily of detoxification enzymes that catalyze the conjugation of reduced glutathione (GSH to a broad spectrum of xenobiotics and hydrophobic electrophiles. Evolutionarily unrelated to the cytosolic glutathione S-transferases, they are found across bacterial and eukaryotic domains, for example in mammals, plants, fungi and bacteria in which significant levels of glutathione are maintained. Species of genus Plasmodium, the unicellular protozoa that are commonly known as malaria parasites, do actively support glutathione homeostasis and maintain its metabolism throughout their complex parasitic life cycle. In humans and in other mammals, the asexual intraerythrocytic stage of malaria, when the parasite feeds on hemoglobin, grows and eventually asexually replicates inside infected red blood cells (RBCs, is directly associated with host disease symptoms and during this critical stage GSH protects the host RBC and the parasite against oxidative stress from parasite-induced hemoglobin catabolism. In line with these observations, several GSH-dependent Plasmodium enzymes have been characterized including glutathione reductases, thioredoxins, glyoxalases, glutaredoxins and glutathione S-transferases (GSTs; furthermore, GSH itself have been found to associate spontaneously and to degrade free heme and its hydroxide, hematin, which are the main cytotoxic byproducts of hemoglobin catabolism. However, despite the apparent importance of glutathione metabolism for the parasite, no membrane associated glutathione S-transferases of genus Plasmodium have been previously described. We recently reported the first examples of MAPEG members among Plasmodium spp.

  8. Glutathione-S-transferase genotype and p53 mutations in adenocarcinoma of the small intestine

    DEFF Research Database (Denmark)

    Pedersen, Lisbeth Nørum; Kærlev, Linda; Teglbjærg, Peter Stubbe;

    2003-01-01

    Adenocarcinoma of the small intestine (ASI) is a rare disease of unknown aetiology. The glutathione S-transferase M1 (GSTM1) enzyme catalyses the detoxification of compounds involved in carcinogenesis of adenocarcinoma of the stomach, colon and lung, including constituents of tobacco smoke. We...... differences. Thus p53 does not seem to be the target of carcinogens acting in the small intestine....

  9. Habitual consumption of fruits and vegetables: associations with human rectal glutathione S-transferase

    NARCIS (Netherlands)

    Wark, P.A.; Grubben, M.J.A.L.; Peters, W.H.M.; Nagengast, F.M.; Kampman, E.; Kok, F.J.; Veer, van 't P.

    2004-01-01

    The glutathione (GSH)/glutathione S-transferase (GST) system is an important detoxification system in the gastrointestinal tract. A high activity of this system may benefit cancer prevention. The aim of the study was to assess whether habitual consumption of fruits and vegetables, especially citrus

  10. Role of genetic polymorphism of glutathione-s-transferase T1 and microsomal epoxide hydrolase in aflatoxin-associated hepatocellular carcinoma

    NARCIS (Netherlands)

    Tiemersma, E.W.; Omer, R.E.; Bunschoten, A.; Veer, van't P.; Kok, F.J.; Idrsi, M.O.; Kampman, E.

    2001-01-01

    Exposure to aflatoxins is a risk factor for hepatocellular carcinoma (HCC). Aflatoxins occur in peanut butter and are metabolized by genetically polymorphic enzymes such as glutathione-S-transferases encoded by glutathione-S-transferase mu 1 gene (GSTM1) and glutathione-S-transferase theta 1 gene (G

  11. Functional dissection of the bipartite active site of the class I coenzyme A (CoA)-transferase succinyl-CoA:acetate CoA-transferase

    Science.gov (United States)

    Murphy, Jesse; Mullins, Elwood; Kappock, T.

    2016-05-01

    Coenzyme A (CoA)-transferases catalyze the reversible transfer of CoA from acyl-CoA thioesters to free carboxylates. Class I CoA-transferases produce acylglutamyl anhydride intermediates that undergo attack by CoA thiolate on either the internal or external carbonyl carbon atoms, forming distinct tetrahedral intermediates less than 3 Å apart. In this study, crystal structures of succinyl-CoA:acetate CoA-transferase (AarC) from Acetobacter aceti are used to examine how the Asn347 carboxamide stabilizes the internal oxyanion intermediate. A structure of the active mutant AarC-N347A bound to CoA revealed both solvent replacement of the missing contact and displacement of the adjacent Glu294, indicating that Asn347 both polarizes and orients the essential glutamate. AarC was crystallized with the nonhydrolyzable acetyl-CoA (AcCoA) analogue dethiaacetyl-CoA (1a) in an attempt to trap a closed enzyme complex containing a stable analogue of the external oxyanion intermediate. One active site contained an acetylglutamyl anhydride adduct and truncated 1a, an unexpected result hinting at an unprecedented cleavage of the ketone moiety in 1a. Solution studies confirmed that 1a decomposition is accompanied by production of near-stoichiometric acetate, in a process that seems to depend on microbial contamination but not AarC. A crystal structure of AarC bound to the postulated 1a truncation product (2a) showed complete closure of one active site per dimer but no acetylglutamyl anhydride, even with acetate added. These findings suggest that an activated acetyl donor forms during 1a decomposition; a working hypothesis involving ketone oxidation is offered. The ability of 2a to induce full active site closure furthermore suggests that it subverts a system used to impede inappropriate active site closure on unacylated CoA.

  12. Immunolabeling of Gamma-glutamyl transferase 5 in Normal Human Tissues Reveals Expression and Localization Differs from Gamma-glutamyl transferase 1

    OpenAIRE

    Hanigan, Marie H.; Gillies, Elizabeth M.; Wickham, Stephanie; Wakeham, Nancy; Wirsig-Wiechmann, Celeste R

    2014-01-01

    Gamma-glutamyl transferase (GGT5) was discovered due to its ability to convert leukotriene C4 (LTC4, a glutathione S-conjugate) to LTD4 and may have an important role in the immune system. However, it was not known which cells express the enzyme in humans. We have developed a sensitive and specific antibody that can be used to detect human GGT5 on western blots and in fixed tissue sections. We localized GGT5 expression in normal human tissues. We observed GGT5 expressed by macrophages present...

  13. A novel plant glutathione S-transferase/peroxidase suppresses Bax lethality in yeast

    DEFF Research Database (Denmark)

    Kampranis, S C; Damianova, R; Atallah, M;

    2000-01-01

    for the identification of plant genes, which inhibit either directly or indirectly the lethal phenotype of Bax. Using this method a number of cDNA clones were isolated, the more potent of which encodes a protein homologous to the class theta glutathione S-transferases. This Bax-inhibiting (BI) protein...... was expressed in Escherichia coli and found to possess glutathione S-transferase (GST) and weak glutathione peroxidase (GPX) activity. Expression of Bax in yeast decreases the intracellular levels of total glutathione, causes a substantial reduction of total cellular phospholipids, diminishes the...... mitochondrial membrane potential, and alters the intracellular redox potential. Co-expression of the BI-GST/GPX protein brought the total glutathione levels back to normal and re-established the mitochondrial membrane potential but had no effect on the phospholipid alterations. Moreover, expression of BI...

  14. Regiospecificity of placental metabolism by cytochromes P450 and glutathione S-transferase.

    Science.gov (United States)

    McRobie, D J; Glover, D D; Tracy, T S

    1996-01-01

    The placenta possesses the ability to metabolize numerous xenobiotics and endogenous steroids. However, it is unknown whether regional differences in these enzymatic reactions exist in the human placenta. To this end, we undertook a study of four regions of the placenta, the chorionic plate, maternal surface, placental margin and whole tissue, to assess the activities of cytochrome P450 1A1 and 19A1 (aromatase) and glutathione S-stransferase in these fractions. No differences in either P450 1A1 or glutathione S-transferase activities were noted among any of the placental fractions. However, with respect to P450 19A1 activity, the placental margin differed significantly from all other fractions (p < 0.05). This study demonstrates that whole tissue samples of the human placenta are adequate for placental cytochrome P450 and glutathione S-transferase metabolism studies. PMID:8938464

  15. Glutathione and gamma-glutamyl transferases are involved in the formation of cadmium-glutathione complex.

    Science.gov (United States)

    Adamis, Paula Daniela Braga; Mannarino, Sérgio Cantú; Eleutherio, Elis Cristina Araújo

    2009-05-01

    In a wild-type strain of Saccharomyces cerevisiae, cadmium induces the activities of both gamma-glutamyl transferase (gamma-GT) and glutathione transferase 2 (Gtt2). However, Gtt2 activity did not increase under gamma-GT or Ycf1 deficiencies, suggesting that the accumulation of glutathione-cadmium in the cytosol inhibits Gtt2. On the other hand, the balance between the cytoplasmic and vacuolar level of glutathione seems to regulate gamma-GT activity, since this enzyme was not activated in a gtt2 strain. Taken together, these results suggest that gamma-GT and Gtt2 work together to remove cadmium from the cytoplasm, a crucial mechanism for metal detoxification that is dependent on glutathione. PMID:19345220

  16. Serum gamma-glutamyl transferase: A novel biomarker for coronary artery disease

    OpenAIRE

    Mao, Yu; Qi, Xiaolong; Xu, Wenjun; Song, Haoming; Xu, Mingxin; Ma, Wanrong; Zhou, Lin

    2014-01-01

    Background Atherosclerosis is a chronic inflammatory process, in which oxidative stress is the key event. Gamma-glutamyl transferase (GGT) is a cellular production of oxidants. We aimed to elucidate the relationship of serum GGT levels and coronary artery disease (CAD) in a Chinese population. Material/Methods A total of 513 adult subjects who had undergone coronary angiography were enrolled in the study. Clinical characteristics, coronary angiography, and serum samples were collected from al...

  17. γ-Glutamyl transferase 7 is a novel regulator of glioblastoma growth

    OpenAIRE

    Bui, Timothy T; Nitta, Ryan T.; Kahn, Suzana A.; Razavi, Seyed-Mostafa; Agarwal, Maya; Aujla, Parvir; Gholamin, Sharareh; Recht, Lawrence; Li, Gordon

    2015-01-01

    Background Glioblastoma (GBM) is the most malignant primary brain tumor in adults, with a median survival time of one and a half years. Traditional treatments, including radiation, chemotherapy, and surgery, are not curative, making it imperative to find more effective treatments for this lethal disease. γ-Glutamyl transferase (GGT) is a family of enzymes that was shown to control crucial redox-sensitive functions and to regulate the balance between proliferation and apoptosis. GGT7 is a nove...

  18. Glutathione S-Transferase Polymorphisms, Passive Smoking, Obesity, and Heart Rate Variability in Nonsmokers

    OpenAIRE

    Probst-Hensch, Nicole M.; Imboden, Medea; Dietrich, Denise Felber; Barthélemy, Jean-Claude; Ackermann-Liebrich, Ursula; Berger, Wolfgang; Gaspoz, Jean-Michel; Schwartz, Joel David

    2008-01-01

    Background: Disturbances of heart rate variability (HRV) may represent one pathway by which second-hand smoke (SHS) and air pollutants affect cardiovascular morbidity and mortality. The mechanisms are poorly understood. Objectives: We investigated the hypothesis that oxidative stress alters cardiac autonomic control. We studied the association of polymorphisms in oxidant-scavenging glutathione S-transferase (GST) genes and their interactions with SHS and obesity with HRV. Methods: A total of ...

  19. Characterisation of Dermanyssus gallinae glutathione S-transferases and their potential as acaricide detoxification proteins

    OpenAIRE

    Bartley, Kathryn; Wright, Harry W.; Bull, Robert S.; Huntley, John F; Nisbet, Alasdair J

    2015-01-01

    Background Glutathione S-transferases (GSTs) facilitate detoxification of drugs by catalysing the conjugation of the reduced glutathione (GSH) to electrophilic xenobiotic substrates and therefore have a function in multi-drug resistance. As a result, knowledge of GSTs can inform both drug resistance in, and novel interventions for, the control of endo- and ectoparasite species. Acaricide resistance and the need for novel control methods are both pressing needs for Dermanyssus gallinae, a high...

  20. Gamma-glutamyl transferase activity in fetal serum, maternal serum, and amniotic fluid during gestation.

    OpenAIRE

    Moniz, C; Nicolaides, K H; Keys, D.; Rodeck, C H

    1984-01-01

    Gamma-glutamyl transferase activity was measured in fetal serum, maternal serum, and amniotic fluid in 173 pregnancies from 15 to 40 weeks' gestation. Fetal serum was obtained in the second trimester by fetoscopy and in the third trimester by umbilical cord puncture at caesarian section or vaginal delivery. Enzyme activities in maternal blood (10 IU/1, SD 2) and fetal blood (88 IU/1, SD 20) remained relatively constant throughout gestation, whereas in the amniotic fluid there was a significan...

  1. Functional and physical interaction between the histone methyl transferase Suv39H1 and histone deacetylases

    OpenAIRE

    Vaute, Olivier; Nicolas, Estelle; Vandel, Laurence; Trouche, Didier

    2002-01-01

    The histone methyl transferase Suv39H1 is involved in silencing by pericentric heterochromatin. It specifically methylates K9 of histone H3, thereby creating a high affinity binding site for HP1 proteins. We and others have shown recently that it is also involved in transcriptional repression by the retinoblastoma protein Rb. Strikingly, both HP1 localisation and repression by Rb also require, at least in part, histone deacetylases. We found here that repression of a heterologous promoter by ...

  2. Methionine sulfoxide reductase regulates brain catechol-O-methyl transferase activity

    OpenAIRE

    Moskovitz, Jackob; Walss-Bass, Consuelo; Cruz, Dianne A.; Thompson, Peter M.; Bortolato, Marco

    2014-01-01

    Catechol-O-methyl transferase (COMT) plays a key role in the degradation of brain dopamine (DA). Specifically, low COMT activity results in higher DA levels in the prefrontal cortex (PFC), thereby reducing the vulnerability for attentional and cognitive deficits in both psychotic and healthy individuals. COMT activity is markedly reduced by a non-synonymous SNP that generates a valine-to-methionine substitution on the residue 108/158, by means of as-yet incompletely understood posttranslation...

  3. Glutathione Transferase from Trichoderma virens Enhances Cadmium Tolerance without Enhancing Its Accumulation in Transgenic Nicotiana tabacum

    OpenAIRE

    Dixit, Prachy; Mukherjee, Prasun K.; Ramachandran, V.; Eapen, Susan

    2011-01-01

    Background Cadmium (Cd) is a major heavy metal pollutant which is highly toxic to plants and animals. Vast agricultural areas worldwide are contaminated with Cd. Plants take up Cd and through the food chain it reaches humans and causes toxicity. It is ideal to develop plants tolerant to Cd, without enhanced accumulation in the edible parts for human consumption. Glutathione transferases (GST) are a family of multifunctional enzymes known to have important roles in combating oxidative stresses...

  4. Studies on Human and Drosophila melanogaster Glutathione Transferases of Biomedical and Biotechnological Interest

    OpenAIRE

    Mazari, Aslam M.A.

    2016-01-01

    Glutathione transferases (GSTs, EC.2.5.1.18) are multifunctional enzymes that are universally distributed in all cellular life forms. They play important roles in metabolism and detoxication of endogenously produced toxic compounds and xenobiotics. GSTs have gained considerable interest over the years for biomedical and biotechnological applications due to their involvement in the conjugation of glutathione (GSH) to a vast array of chemical species. Additionally, the emergence of non-detoxify...

  5. Genetic polymorphism for glutathione-S-transferase mu in asbestos cement workers.

    OpenAIRE

    Jakobsson, K; Rannug, A.; Alexandrie, A K; Rylander, L; Albin, M; Hagmar, L

    1994-01-01

    OBJECTIVE--To investigate whether a lack of glutathione-S-transferase mu (GSTM1) activity was related to an increased risk for adverse outcome after asbestos exposure. METHODS--A study was made of 78 male former asbestos cement workers, with retrospective cohort data on exposure, radiographical findings, and lung function. Venous blood samples were obtained for the analysis of GSTM1 polymorphism by the polymerase chain reaction technique. Chest x ray films were classified according to the Int...

  6. Frequency of Galactose-1-phosphate Uridyl Transferase Gene Mutations in Healthy Population of Croatia

    OpenAIRE

    Barišić, Karmela; Rumora, Lada; Grdić, Marija; JURETIĆ, DUBRAVKA

    2008-01-01

    Galactosemia is a human disease caused by deficient activity of each one of the three enzymes involved in galactose metabolism, galactokinase (GALK), galactose-1-phosphate uridyl transferase (GALT) and UDP-galactose-4-epimerase (GALE). Absence or deficiency of GALT activity results in classical galactosemia. This disorder exhibits allelic heterogeneity in different populations and ethnic groups. The aim of this study was to search for galactosemia mutations Q188R, N314D, and K285N in healthy ...

  7. Expression Profiling of Selected Glutathione Transferase Genes in Zea mays (L.) Seedlings Infested with Cereal Aphids

    OpenAIRE

    Hubert Sytykiewicz; Grzegorz Chrzanowski; Paweł Czerniewicz; Iwona Sprawka; Iwona Łukasik; Sylwia Goławska; Cezary Sempruch

    2014-01-01

    The purpose of this report was to evaluate the expression patterns of selected glutathione transferase genes (gst1, gst18, gst23 and gst24) in the tissues of two maize (Zea mays L.) varieties (relatively resistant Ambrozja and susceptible Tasty Sweet) that were colonized with oligophagous bird cherry-oat aphid (Rhopalosiphum padi L.) or monophagous grain aphid (Sitobion avenae L.). Simultaneously, insect-triggered generation of superoxide anion radicals (O2 •−) in infested Z. mays plants was ...

  8. Glutathione-S-transferase P protects against endothelial dysfunction induced by exposure to tobacco smoke

    OpenAIRE

    Conklin, Daniel J.; Haberzettl, Petra; Prough, Russell A.; Bhatnagar, Aruni

    2009-01-01

    Exposure to tobacco smoke impairs endothelium-dependent arterial dilation. Reactive constituents of cigarette smoke are metabolized and detoxified by glutathione-S-transferases (GSTs). Although polymorphisms in GST genes are associated with the risk of cancer in smokers, the role of these enzymes in regulating the cardiovascular effects of smoking has not been studied. The P isoform of GST (GSTP), which catalyzes the conjugation of electrophilic molecules in cigarette smoke such as acrolein, ...

  9. Isolation and Characterization of a Theta Glutathione S-transferase Gene from Panax ginseng Meyer

    OpenAIRE

    Kim, Yu-Jin; Lee, Ok Ran; Lee, Sungyoung; Kim, Kyung-Tack; Yang, Deok-Chun

    2012-01-01

    Plants have versatile detoxification systems to encounter the phytotoxicity of the wide range of natural and synthetic compounds present in the environment. Glutathione S-transferase (GST) is an enzyme that detoxifies natural and exogenous toxic compounds by conjugation with glutathione (GSH). Recently, several roles of GST giving stress tolerance in plants have demonstrated, but little is known about the role of ginseng GSTs. Therefore, this work aimed to provide further information on the G...

  10. Expression of glutathione S-transferases in normal and malignant pancreas: an immunohistochemical study.

    OpenAIRE

    Collier, J D; Bennett, M K; Hall, A.; Cattan, A R; Lendrum, R.; Bassendine, M F

    1994-01-01

    The glutathione S-transferases (GSTs) are a family of detoxification and metabolising enzymes, which have been linked with the susceptibility of tissues to environmental carcinogens and resistance of tumours to chemotherapy. Environmental carcinogens have been implicated in the pathogenesis of pancreatic carcinoma, which is also a tumour characterised by marked chemotherapeutic drug resistance. In this study 26 pancreatic adenocarcinoma and 12 normal pancreatic samples were examined immunohis...

  11. Predicted binding of certain antifilarial compounds with glutathione-S-transferase of human Filariids

    OpenAIRE

    Saeed, Mohd; Baig, Mohd. Hassan; Bajpai, Preeti; Srivastava, Ashwini Kumar; Ahmad, Khurshid; Mustafa, Huma

    2013-01-01

    Glutathione-S-transferase is a major phase-II detoxification enzyme in parasitic helminthes. Previous research highlights the importance of GSTs in the establishment of chronic infections in cytotoxic microenvironments. Filarial nematodes depend on these detoxification enzymes for their survival in the host. GST plays an important role in filariasis and other diseases. GST from W.bancrofti and B.malayi are very much different from human GST. This structural difference makes GST potential chem...

  12. Dietary Patterns and Serum Gamma-Glutamyl Transferase in Japanese Men and Women

    OpenAIRE

    . .

    2015-01-01

    Background Although specific foods and nutrients have been examined as potential determinants of serum gamma-glutamyl transferase (GGT) concentrations, the relationship between dietary patterns and GGT remains unknown. The present cross-sectional study aimed to determine relationships between dietary patterns and GGT concentrations, and the effects of lifestyle factors on GGT. Methods Relationships between dietary patterns and GGT were analyzed in 9803 Japanese individuals (3723 men and 6080 ...

  13. Study on N-acetylgucosaminyl transferase and the uptake of the correlated imaging agent

    International Nuclear Information System (INIS)

    N-acetylglucosaminyl transferase(GnT) is related to the development of tumor and the cancer patients' prognosis by effecting the change of glucose's chain. Study on the transform of glycosyltransferase is benefit to the comprehension of the mechanism of biological behavior. The noninvasive diagnostic and treating methods of tumor will be provided along with the development of new imaging agent of tumor glucose analogue and its mechanism defined clearly. (authors)

  14. Modeling analysis of GST (glutathione-S-transferases) from Wuchereria bancrofti and Brugia malayi

    OpenAIRE

    Bhargavi, Rayavarapu; Vishwakarma, Siddharth; Murty, Upadhyayula Suryanarayana

    2005-01-01

    GST (glutathione S-transferases) are a family of detoxification enzymes that catalyze the conjugation of reduced GSH (glutathione) to xenobiotic (endogenous electrophilic) compounds. GST from Wb (Wuchereria bancrofti) and Bm (Brugia malayi) are significantly different from human GST in sequence and structure. Thus, Wb-GST and Bm-GST are potential chemotherapeutic targets for anti-filarial treatment. Comparison of modeled Wb and Bm GST with human GST show structural difference between them. An...

  15. Exploiting the Substrate Promiscuity of Hydroxycinnamoyl-CoA:Shikimate Hydroxycinnamoyl Transferase to Reduce Lignin

    OpenAIRE

    Eudes, Aymerick; Pereira, Jose H.; Yogiswara, Sasha; Wang, George; Teixeira Benites, Veronica; Baidoo, Edward E.K.; Lee, Taek Soon; Adams, Paul D; Keasling, Jay D.; Loqué, Dominique

    2016-01-01

    Lignin poses a major challenge in the processing of plant biomass for agro-industrial applications. For bioengineering purposes, there is a pressing interest in identifying and characterizing the enzymes responsible for the biosynthesis of lignin. Hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyl transferase (HCT; EC 2.3.1.133) is a key metabolic entry point for the synthesis of the most important lignin monomers: coniferyl and sinapyl alcohols. In this study, we investigated the substrate prom...

  16. Wild-type HTT modulates the enzymatic activity of the neuronal palmitoyl transferase HIP14

    OpenAIRE

    Huang, Kun; Shaun S Sanders; Kang, Rujun; Carroll, Jeffrey B; Sutton, Liza; Wan, Junmei; Singaraja, Roshni; Young, Fiona B.; Liu, Lili; El-Husseini, Alaa; Davis, Nicholas G.; Hayden, Michael R.

    2011-01-01

    Huntington disease (HD) is caused by polyglutamine expansion in the huntingtin (HTT) protein. Huntingtin-interacting protein 14 (HIP14), one of 23 DHHC domain-containing palmitoyl acyl transferases (PATs), binds to HTT and robustly palmitoylates HTT at cysteine 214. Mutant HTT exhibits reduced palmitoylation and interaction with HIP14, contributing to the neuronal dysfunction associated with HD. In this study, we confirmed that, among 23 DHHC PATs, HIP14 and its homolog DHHC-13 (HIP14L) are t...

  17. Glutathione S-transferases of Aulacorthum solani and Acyrthosiphon pisum: partial purification and characterization.

    OpenAIRE

    Francis, Frédéric; Haubruge, Eric; Gaspar, Charles; Dierickx, P. J.

    2001-01-01

    Glutathione S-transferases (GST) play an important role in the detoxification of many substances including allelochemicals from plants. Brassicaceae plants contain glucosinolates and emit volatile isothiocyanates which affect the GST system. A comparison of the GST of two aphid species, the generalist Aulacorthum solani found on Brassicaceae and the Fabaceae specialist Acyrthosiphon pisum, was made to try to explain their respective feeding behaviour. Differences of GST were determined among ...

  18. Homology between O-linked GlcNAc transferases and proteins of the glycogen phosphorylase superfamily.

    Science.gov (United States)

    Wrabl, J O; Grishin, N V

    2001-11-30

    The O-linked GlcNAc transferases (OGTs) are a recently characterized group of largely eukaryotic enzymes that add a single beta-N-acetylglucosamine moiety to specific serine or threonine hydroxyls. In humans, this process may be part of a sugar regulation mechanism or cellular signaling pathway that is involved in many important diseases, such as diabetes, cancer, and neurodegeneration. However, no structural information about the human OGT exists, except for the identification of tetratricopeptide repeats (TPR) at the N terminus. The locations of substrate binding sites are unknown and the structural basis for this enzyme's function is not clear. Here, remote homology is reported between the OGTs and a large group of diverse sugar processing enzymes, including proteins with known structure such as glycogen phosphorylase, UDP-GlcNAc 2-epimerase, and the glycosyl transferase MurG. This relationship, in conjunction with amino acid similarity spanning the entire length of the sequence, implies that the fold of the human OGT consists of two Rossmann-like domains C-terminal to the TPR region. A conserved motif in the second Rossmann domain points to the UDP-GlcNAc donor binding site. This conclusion is supported by a combination of statistically significant PSI-BLAST hits, consensus secondary structure predictions, and a fold recognition hit to MurG. Additionally, iterative PSI-BLAST database searches reveal that proteins homologous to the OGTs form a large and diverse superfamily that is termed GPGTF (glycogen phosphorylase/glycosyl transferase). Up to one-third of the 51 functional families in the CAZY database, a glycosyl transferase classification scheme based on catalytic residue and sequence homology considerations, can be unified through this common predicted fold. GPGTF homologs constitute a substantial fraction of known proteins: 0.4% of all non-redundant sequences and about 1% of proteins in the Escherichia coli genome are found to belong to the GPGTF

  19. Functional Identification of Proteus mirabilis eptC gene encoding a Core Lipopolysaccharide Phosphoethanolamine Transferase

    OpenAIRE

    Eleonora Aquilini; Susana Merino; Knirel, Yuriy A.; Miguel Regué; Tomás, Juan M.

    2014-01-01

    By comparison of the Proteus mirabilis HI4320 genome with known lipopolysaccharide (LPS) phosphoethanolamine transferases, three putative candidates (PMI3040, PMI3576, and PMI3104) were identified. One of them, eptC (PMI3104) was able to modify the LPS of two defined non-polar core LPS mutants of Klebsiella pneumoniae that we use as surrogate substrates. Mass spectrometry and nuclear magnetic resonance showed that eptC directs the incorporation of phosphoethanolamine to the O-6 of l-glycer...

  20. Pharmacogenetics of azathioprine in inflammatory bowel disease: A role for glutathione-S-transferase?

    OpenAIRE

    Stocco, Gabriele; Pelin, Marco; Franca, Raffaella; De Iudicibus, Sara; Cuzzoni, Eva; Favretto, Diego; Martelossi, Stefano; Ventura, Alessandro; Decorti, Giuliana

    2014-01-01

    Azathioprine is a purine antimetabolite drug commonly used to treat inflammatory bowel disease (IBD). In vivo it is active after reaction with reduced glutathione (GSH) and conversion to mercaptopurine. Although this reaction may occur spontaneously, the presence of isoforms M and A of the enzyme glutathione-S-transferase (GST) may increase its speed. Indeed, in pediatric patients with IBD, deletion of GST-M1, which determines reduced enzymatic activity, was recently associated with reduced s...

  1. The Quest for Functional Quasi-Species in Glutathione Transferase Libraries

    OpenAIRE

    Rúnarsdóttir, Arna

    2010-01-01

    Glutathione transferases (GSTs) are good candidates for investigations of enzyme evolution, due to their broad substrate specificities and structural homology. The primary role of GSTs is to act as phase II detoxifying enzymes protecting the cell from toxic compounds of both endo- and exogenous origins. The detoxification is conducted via conjugation with glutathione (GSH), which facilitates their removal from the body. The work presented in this thesis has supported a theory for enzyme evolu...

  2. The Stereochemical Course of 4-Hydroxy-2-nonenal Metabolism by Glutathione S-Transferases*S⃞

    OpenAIRE

    Balogh, Larissa M.; Roberts, Arthur G.; Shireman, Laura M.; Greene, Robert J.; Atkins, William M.

    2008-01-01

    4-Hydroxy-2-nonenal (HNE) is a toxic aldehyde generated during lipid peroxidation and has been implicated in a variety of pathological states associated with oxidative stress. Glutathione S-transferase (GST) A4-4 is recognized as one of the predominant enzymes responsible for the metabolism of HNE. However, substrate and product stereoselectivity remain to be fully explored. The results from a product formation assay indicate that hGSTA4-4 exhibits a modest preference ...

  3. Glutathione transferase A4-4 resists adduction by 4-hydroxynonenal☆

    OpenAIRE

    Shireman, Laura M.; Kripps, Kimberly A.; Balogh, Larissa M.; Conner, Kip P.; Whittington, Dale; Atkins, William M.

    2010-01-01

    4-Hydroxy-2-trans-nonenal (HNE) is a lipid peroxidation product that contributes to the pathophysiology of several diseases with components of oxidative stress. The electrophilic nature of HNE results in covalent adduct formation with proteins, fatty acids and DNA. However, it remains unclear whether enzymes that metabolize HNE avoid inactivation by it. Glutathione transferase A4-4 (GST A4-4) plays a significant role in the elimination of HNE by conjugating it with glutathione (GSH), with cat...

  4. Glutathione S Transferases Polymorphisms Are Independent Prognostic Factors in Lupus Nephritis Treated with Cyclophosphamide

    OpenAIRE

    Audemard-Verger, Alexandra; Martin Silva, Nicolas; Verstuyft, Céline; Costedoat-Chalumeau, Nathalie; Hummel, Aurélie; Le Guern, Véronique; Sacré, Karim; Meyer, Olivier; Daugas, Eric; Goujard, Cécile; Sultan, Audrey; Lobbedez, Thierry; Galicier, Lionel; Pourrat, Jacques; Le Hello, Claire

    2016-01-01

    Objective To investigate association between genetic polymorphisms of GST, CYP and renal outcome or occurrence of adverse drug reactions (ADRs) in lupus nephritis (LN) treated with cyclophosphamide (CYC). CYC, as a pro-drug, requires bioactivation through multiple hepatic cytochrome P450s and glutathione S transferases (GST). Methods We carried out a multicentric retrospective study including 70 patients with proliferative LN treated with CYC. Patients were genotyped for polymorphisms of the ...

  5. Glutathione S-transferase polymorphisms, passive smoking, obesity, and heart rate variability in nonsmokers

    OpenAIRE

    Probst-Hensch, N.M.; Imboden, M.; Felber Dietrich, D; Barthélémy, Jean Claude; Ackermann-Liebrich, U; Berger, W.; Gaspoz, Jean-Michel; Schwartz, J.

    2008-01-01

    BACKGROUND: Disturbances of heart rate variability (HRV) may represent one pathway by which second-hand smoke (SHS) and air pollutants affect cardiovascular morbidity and mortality. The mechanisms are poorly understood. OBJECTIVES: We investigated the hypothesis that oxidative stress alters cardiac autonomic control. We studied the association of polymorphisms in oxidant-scavenging glutathione S-transferase (GST) genes and their interactions with SHS and obesity with HRV. METHODS: A total of ...

  6. Alpha-class glutathione transferases as steroid isomerases and scaffolds for protein redesign

    OpenAIRE

    Pettersson, Pär L.

    2002-01-01

    The present work focuses on the glutathione transferase (GST) Alpha-class enzymes, their characteristics as steroid isomerases and structural plasticity as malleable scaffolds for protein design. The GSTs are a family of detoxication enzymes that appears to have a wider variety of additional functions. Kinetic steady-state parameters for human GST A1-1 with the steroid isomerase substrate Δ5-androstene-3,17-dione (AD), an intermediate in steroid hormone biosynthesis, were determined. It was e...

  7. Glutathione-S-Transferases As Determinants of Cell Survival and Death

    OpenAIRE

    Tew, Kenneth D.; Townsend, Danyelle M.

    2012-01-01

    Significance: The family of glutathione S-transferases (GSTs) is part of a cellular Phase II detoxification program composed of multiple isozymes with functional human polymorphisms that have the capacity to influence individual response to drugs and environmental stresses. Catalytic activity is expressed through GST dimer-mediated thioether conjugate formation with resultant detoxification of a variety of small molecule electrophiles. Recent Advances: More recent work indicates that in addit...

  8. Proteomic and Immunochemical Characterization of Glutathione Transferase as a New Allergen of the Nematode Ascaris lumbricoides

    OpenAIRE

    Nathalie Acevedo; Jens Mohr; Josefina Zakzuk; Martin Samonig; Peter Briza; Anja Erler; Anna Pomés; Huber, Christian G.; Fatima Ferreira; Luis Caraballo

    2013-01-01

    Helminth infections and allergy have evolutionary and clinical links. Infection with the nematode Ascaris lumbricoides induces IgE against several molecules including invertebrate pan-allergens. These antibodies influence the pathogenesis and diagnosis of allergy; therefore, studying parasitic and non-parasitic allergens is essential to understand both helminth immunity and allergy. Glutathione transferases (GSTs) from cockroach and house dust mites are clinically relevant allergens and compa...

  9. Erythromycin binding is reduced in ribosomes with conformational alterations in the 23 S rRNA peptidyl transferase loop

    DEFF Research Database (Denmark)

    Douthwaite, S; Aagaard, C

    1993-01-01

    induced by mutations in the peptidyl transferase loop, and to determine how these changes affect drug interaction. Mutations at positions 2057 (G-->A) and 2058 (A-->G, or -->U), all of which confer drug resistance, induce a more open conformation in the peptidyl transferase loop. Erythromycin still...... protects against chemical modification in the mutant peptidyl transferase loops, but the affinity of the drug interaction is reduced 20-fold in the 2057A mutant, 10(3)-fold in the 2058U mutant and 10(4)-fold in the 2058G mutant. Single mutations at position 2032 in the adjacent hairpin loop, which have...... previously been shown to alter drug tolerances, gave no detectable effects on the structure of the peptidyl transferase loop or on erythromycin binding. Dual mutations at positions 2032 and 2058, however, induce a marked change in the rRNA conformation with opening of the phylogenetically conserved base...

  10. Mapping important nucleotides in the peptidyl transferase centre of 23 S rRNA using a random mutagenesis approach

    DEFF Research Database (Denmark)

    Porse, B T; Garrett, R A

    1995-01-01

    Random mutations were generated in the lower half of the peptidyl transferase loop in domain V of 23 S rRNA from Escherichia coli using a polymerase chain reaction (PCR) approach, a rapid procedure for identifying mutants and a plasmid-based expression system. The effects of 21 single......-site mutations, at 18 different positions, on cell growth, mutant rRNA incorporation into ribosomes and peptidyl transferase activity of the mutant ribosomes were analysed. The general importance of the whole region for the peptidyl transferase centre was emphasized by the finding that 14 of the mutants were...... sick, or very sick, when ribosomes containing chromosomal-encoded 23 S rRNA were inhibited by erythromycin, and all except one of these exhibited low levels of peptidyl transferase activity in their mutated ribosomes. Two mutations, psi 2580-->C and U2584-->G that both yielded inactive ribosomes were...

  11. Interactions of photoactive DNAs with terminal deoxynucleotidyl transferase: Identification of peptides in the DNA binding domain

    International Nuclear Information System (INIS)

    Terminal deoxynucleotidyl transferase (terminal transferase) was specifically modified in the DNA binding site by a photoactive DNA substrate (hetero-40-mer duplex containing eight 5-azido-dUMP residues at one 3' end). Under optimal photolabeling conditions, 27-40% of the DNA was covalently cross-linked to terminal transferase. The specificity of the DNA and protein interaction was demonstrated by protection of photolabeling at the DNA binding domain with natural DNA substrates. In order to recover high yields of modified peptides from limited amounts of starting material, protein modified with 32P-labeled photoactive DNA and digested with trypsin was extracted 4 times with phenol followed by gel filtration chromatography. All peptides not cross-linked to DNA were extracted into the phenol phase while the photolyzed DNA and the covalently cross-linked peptides remained in the aqueous phase. The 32P-containing peptide-DNA fraction was subjected to amino acid sequence analysis. Two sequences, Asp221-Lys231 (peptide B8) and Cys234-Lys249 (peptide B10), present in similar yield, were identified. Structure predictions placed the two peptides in an α-helical array of 39 angstrom which would accommodate a DNA helix span of 11 nucleotides. These peptides share sequence similarity with a region in DNA polymerase β that has been implicated in the binding of DNA template

  12. Expression of polypeptide GalNAc-transferases in stratified epithelia and squamous cell carcinomas

    DEFF Research Database (Denmark)

    Mandel, U; Hassan, H; Therkildsen, M H;

    1999-01-01

    human GalNAc-T1, -T2, and -T3. Application of this panel of novel antibodies revealed that GalNAc- transferases are differentially expressed in different cell lines, in spermatozoa, and in oral mucosa and carcinomas. For example, GalNAc-T1 and -T2 but not -T3 were highly expressed in WI38 cells, and Gal......NAc-T3 but not GalNAc-T1 or -T2 was expressed in spermatozoa. The expression patterns in normal oral mucosa were found to vary with cell differentiation, and for GalNAc-T2 and -T3 this was reflected in oral squamous cell carcinomas. The expression pattern of GalNAc-T1 was on the other hand changed in......Mucin-type O-glycosylation is initiated by a large family of UDP-GalNAc: polypeptide N -acetyl-galactosaminyltransferases (GalNAc-transferases). Individual GalNAc-transferases appear to have different functions and Northern analysis indicates that they are differently expressed in different organs...

  13. Characterization of Affinity-Purified Isoforms of Acinetobacter calcoaceticus Y1 Glutathione Transferases

    Directory of Open Access Journals (Sweden)

    Chin-Soon Chee

    2014-01-01

    Full Text Available Glutathione transferases (GST were purified from locally isolated bacteria, Acinetobacter calcoaceticus Y1, by glutathione-affinity chromatography and anion exchange, and their substrate specificities were investigated. SDS-polyacrylamide gel electrophoresis revealed that the purified GST resolved into a single band with a molecular weight (MW of 23 kDa. 2-dimensional (2-D gel electrophoresis showed the presence of two isoforms, GST1 (pI 4.5 and GST2 (pI 6.2 with identical MW. GST1 was reactive towards ethacrynic acid, hydrogen peroxide, 1-chloro-2,4-dinitrobenzene, and trans,trans-hepta-2,4-dienal while GST2 was active towards all substrates except hydrogen peroxide. This demonstrated that GST1 possessed peroxidase activity which was absent in GST2. This study also showed that only GST2 was able to conjugate GSH to isoproturon, a herbicide. GST1 and GST2 were suggested to be similar to F0KLY9 (putative glutathione S-transferase and F0KKB0 (glutathione S-transferase III of Acinetobacter calcoaceticus strain PHEA-2, respectively.

  14. A study of the prognostic role of serum fucose and fucosyl transferase in cancer of the uterine cervix.

    OpenAIRE

    Sen, Urmi; Guha,Subhas; Chowdhury, J Roy

    1985-01-01

    Serum fucose levels and fucosyl transferase activities have been designated as nonspecific markers of malignancy, and play an important role in the diagnosis of different types of malignancies. In the present study, attempts were made to determine the prognostic significance of these markers in patients with cancer of the uterine cervix after therapy. It was found that both serum fucose and fucosyl transferase, which were elevated in untreated patients declined significantly in patients respo...

  15. The degradation of arabinoxylan-rich cell walls in digesta obtained from piglets fed wheat-based diets varies depending on digesta collection site, type of cereal, and source of exogenous xylanase.

    Science.gov (United States)

    Pedersen, N R; Azem, E; Broz, J; Guggenbuhl, P; Le, D M; Fojan, P; Pettersson, D

    2012-12-01

    The objective of the present study was to compare the ability of experimental and commercial xylanases to degrade, in vitro, the arabinoxylan (AX) fraction in digesta from 28-d-old piglets fed a wheat (Triticum aestivum)-based diet (49% wheat). Pigs were euthanized at 1, 2, 3, or 4 h after feeding; stomach and ileum contents were isolated and frozen and later used for the in vitro studies. Xylan solubilization provided information regarding the ability of the enzymes to degrade AX during the harsh in vivo conditions prevailing in the gastrointestinal tract. The hydrolytic capacity of a commercial xylanase was compared with that of an experimental xylanase using stomach digesta (pH 1.8) obtained at 4 h after feeding. Relative to the control, both enzymes increased (P < 0.001) xylan solubilization 3-fold. In the ileal digesta (1 h), xylan solubilization was increased by 36% (P < 0.001). Inclusion of arabinofuranosidases (Ara f) with xylanases increased xylan solubilization in stomach samples (P = 0. 007 and P = 0. 030) but not in ileal samples (P = 0.873 and P = 0.997). Our results illustrate clearly the importance of using different conditions and substrates when enzyme performance is studied in vitro as a prescreening tool for setting up in vivo trials. PMID:23365312

  16. Preparation of feruloyl esterase and application in feed of broiler%阿魏酸酯酶的制备及在肉鸡饲料中的应用

    Institute of Scientific and Technical Information of China (English)

    杨道秀; 许文江; 王林林; 蔡婀娜; 李夏兰

    2013-01-01

    为了降低饲料中非淀粉多糖质量抗营养因子,提高肉鸡对饲料养分的利用,本文研究了固态发酵阿魏酸酯酶的工艺及其对饲料养分利用率的影响。结果表明:在麦麸与麦糟质量比为2∶8、蛋白胨与酵母粉质量比为2∶4、含水量为40%、接种量为10%、培养时间为6天、培养温度为33℃条件下,阿魏酸酯酶的酶活达到了29.49 U/g,较未优化前提高了5.11倍。将浓缩后的酶液与溢多酶AF831添加入肉鸡饲料中,发现添加溢多酶及阿魏酸酯酶可以提高干物质、粗蛋白、粗灰分、酸性洗涤纤维等利用率,但作用不明显,对中性洗涤纤维改善效果极显著(P<0.01)。%To reduce non-starch polysaccharides anti-nutritional factor in feed and promote intake of nutrient,the solid-state fermentation technology of feruloyl esterase (FAE) and the influence on nutrient utilization rate were studied. Under optimized conditions of 2∶8 of the ratio for wheat bran and brewers’ spent grain (BSG),2∶4 of the ratio for peptone and yeast,40%of water content,10%of inoculum,6 days of incubation and 33 ℃ of incubation temperature,the activity of FAE was improved to five times compared to unoptimized conditions and the maximum activity was 29.49 U/g. The combination of FAE and Yiduoli enzyme was mixed in feed for broiler. FAE and Yiduoli enzyme could improve the utilization of dry matter,crude protein,crude ash,acidic detergent fiber,but the improvement was limited. The utilization of neutral detergent fiber increased markedly (P<0.01).

  17. Xenotransplantation of galactosyl-transferase knockout, CD55, CD59, CD39, and fucosyl-transferase transgenic pig kidneys into baboons.

    Science.gov (United States)

    Le Bas-Bernardet, S; Tillou, X; Poirier, N; Dilek, N; Chatelais, M; Devallière, J; Charreau, B; Minault, D; Hervouet, J; Renaudin, K; Crossan, C; Scobie, L; Cowan, P J; d'Apice, A J F; Galli, C; Cozzi, E; Soulillou, J P; Vanhove, B; Blancho, G

    2011-11-01

    Galactosyl-transferase knockout (GT-KO) pigs represent the latest major progress to reduce immune reactions in xenotransplantation. However, their organs are still subject to rapid humoral rejection involving complement activation requiring the ongoing development of further genetic modifications in the pig. In a pig-to-baboon renal transplantation setting, we have used donor pigs that are not only GT-KO, but also transgenic for human CD55 (hCD55), hCD59, hCD39, and fucosyl-transferase (hHT). We studied kidney xenograft survival, physiological and immunologic parameters, xenogeneic rejection characteristics, as well as viral transmission aspects among two groups of baboons: control animals (n = 2), versus those (n = 4) treated with a cocktail of cyclophosphamide, tacrolimus, mycophenolate mofetil, steroids, and a recombinant human C1 inhibitor. Whereas control animals showed clear acute humoral rejection at around day 4, the treated animals showed moderately improved graft survival with rejection at around 2 weeks posttransplantation. Biopsies showed signs of acute vascular rejection (interstitial hemorrhage, glomerular thrombi, and acute tubular necrosis) as well as immunoglobulin (Ig)M and complement deposition in the glomerular and peritubular capillaries. The low level of preformed non-Gal-α1.3Gal IgM detected prior to transplantation increased at 6 days posttransplantation, whereas induced IgG appeared after day 6. No porcine endogenous retrovirus (PERV) transmission was detected in any transplanted baboon. Thus, surprisingly, organs from the GT-KO, hCD55, hCD59, hCD39, and hHT transgenic donors did not appear to convey significant protection against baboon anti-pig antibodies and complement activation, which obviously continue to be significant factors under a suboptimal immunosuppression regimen. The association, timing, and doses of immunosuppressive drugs remain critical. They will have to be optimized to achieve longer graft survivals. PMID:22099813

  18. Compensatory expression and substrate inducibility of γ-glutamyl transferase GGT2 isoform in Arabidopsis thaliana

    OpenAIRE

    Destro, Tiziana; Prasad, Dinesh; Martignago, Damiano; Lliso Bernet, Ignacio; Trentin, Anna Rita; Renu, Indu Kumari; Ferretti, Massimo; Masi, Antonio

    2010-01-01

    γ-Glutamyl transferases (GGT; EC 2.3.2.2) are glutathione-degrading enzymes that are represented in Arabidopsis thaliana by a small gene family of four members. Two isoforms, GGT1 and GGT2, are apoplastic, sharing broad similarities in their amino acid sequences, but they are differently expressed in the tissues: GGT1 is expressed in roots, leaves, and siliques, while GGT2 was thought to be expressed only in siliques. It is demonstrated here that GGT2 is also expressed in wild-type roots, alb...

  19. Can we use serum gamma-glutamyl transferase levels to predict early mortality in stroke?

    OpenAIRE

    Akinci, Emine; Doğan, Nurettin Özgür; GÜMÜŞ, Haluk; Akilli, Nazire Belgin

    2014-01-01

    Objective: Serum gamma-glutamyl transferase (GGT) is a marker for alcohol consumption and hepatobiliary diseases. There are reports on the prognostic role of GGT in coronary artery diseases and stroke. The aim of our study was to identify the potential differences in GGT levels in different types of stroke, and to evaluate the correlation between GGT and 30-day mortality. Method: Patients diagnosed with stroke in emergency department between 01.01.2010 and 30.12.2012 was included in the study...

  20. Is serum gamma-glutamyl transferase a good marker of alcohol intake in stroke patients?

    OpenAIRE

    Peck, K.; Shinton, R; Beevers, G.

    1990-01-01

    Serial serum gamma-glutamyl transferase (GGT) levels were estimated in 23 consecutive patients admitted to hospital with a diagnosis of acute stroke. The proportion of patients with elevated GGT levels in the initial, 36-hour and 72-hour samples was 13%, 30% and 24% respectively, suggesting a transient rise following a stroke. Patients with a history of diabetes mellitus had an initial serum GGT level 21 IU/l (95% confidence interval 6 to 37) higher than non-diabetics. We conclude that GGT le...

  1. Glutathione S-transferase and Catalase gene polymorphisms with Type 2 diabetes mellitus.

    OpenAIRE

    Pushpank Vats; Honey Chandra; Monisha Banerjee

    2013-01-01

    Background and Aim: Antioxidant enzymes such as glutathione S-transferases (GSTs) and catalase (CAT) play important roles in cellular defense by detoxifying various toxic substrates and can be used as important biomarkers for T2DM. The aim of the present work was to study the association of GST and CAT gene polymorphism with T2DM cases and controls in north Indian population. Materials and Methods: Polymorphic GST gene isoforms, GSTM1, T1 and P1 were investigated in 201 healthy control su...

  2. Generation of Active Bovine Terminal Deoxynucleotidyl Transferase (TdT in E.coli

    Directory of Open Access Journals (Sweden)

    Wee Liang Kuan

    2010-08-01

    Full Text Available A synthetic gene encoding bovine terminal deoxynucleotidyl transferase (TdT was generated, cloned into an expression vector and expressed in E.coli. The effects of altering culture and induction conditions on the nature of recombinant protein production were investigated. This led to the expression of active recombinant bovine TdT in E.coli. After purification and characterisation, the activity of the enzyme was assessed in a biological assay for apoptosis. The process described in this report enables the economical production of TdT for high throughput applications.

  3. Cefadroxil potency as cancer co-therapy candidate by glutathione s-transferase mechanism

    OpenAIRE

    Tri Yuliani; Sudibyo Martono; Sansan Sukamdani Tjipto; Muhammad Yusuf Putroutomo; Irwan Desyanto Raharjo Indartono

    2013-01-01

    Background: Glutathione S-transferases (GSTs) havean important role in the detoxification of electrophiles,such as some anticancer drugs. Compounds with phenolicand/or α,b-unsaturated carbonyl group have been knownas GSTs inhibitor in vitro. Cefadroxil in vitro decreasedGST-Pi activity but not GSTs in rat kidney cytosol.GST inhibitor in a specific organ and of a specific classis needed for safety in cancer chemotherapy. The studyaims to observe the effect of cefadroxil on GSTs in vivoin rat k...

  4. Characterization of Affinity-Purified Isoforms of Acinetobacter calcoaceticus Y1 Glutathione Transferases

    OpenAIRE

    Chin-Soon Chee; Irene Kit-Ping Tan; Zazali Alias

    2014-01-01

    Glutathione transferases (GST) were purified from locally isolated bacteria, Acinetobacter calcoaceticus Y1, by glutathione-affinity chromatography and anion exchange, and their substrate specificities were investigated. SDS-polyacrylamide gel electrophoresis revealed that the purified GST resolved into a single band with a molecular weight (MW) of 23 kDa. 2-dimensional (2-D) gel electrophoresis showed the presence of two isoforms, GST1 (pI 4.5) and GST2 (pI 6.2) with identical MW. GST1 was r...

  5. Glutathione S-transferase pi localizes in mitochondria and protects against oxidative stress.

    OpenAIRE

    Goto, Shinji; Kawakatsu, Miho; Izumi, Shin-ichi; Urata, Yoshishige; Kageyama, Kan; Ihara, Yoshito; Koji, Takehiko; Kondo, Takahito

    2009-01-01

    Glutathione S-transferases (GSTs) are multifunctional enzymes involved in the protection of cellular components against anti-cancer drugs or peroxidative stress. Previously we found that GST pi, an isoform of the GSTs, is transported into the nucleus. In the present study, we found that GST pi is present in mitochondria as well as in the cytosol and nucleus in mammalian cell lines. A construct comprising the 84 amino acid residues in the amino-terminal region of GST pi and green fluorescent p...

  6. Cloning, expression and analysis of the olfactory glutathione S-transferases in coho salmon

    OpenAIRE

    Espinoza, Herbert M.; Shireman, Laura M.; McClain, Valerie; Atkins, William; Gallagher, Evan P.

    2012-01-01

    The glutathione S-transferases (GSTs) provide cellular protection by detoxifying xenobiotics, maintaining redox status, and modulating secondary messengers, all of which are critical to maintaining olfaction in salmonids. Here, we characterized the major coho salmon olfactory GSTs (OlfGSTs), namely omega, pi, and rho subclasses. OlfGST omega contained an open reading frame of 720 bp and encoded a protein of 239 amino acids. OlfGST pi and OlfGST rho contained open reading frames of 727 and 681...

  7. Physicochemical consequences of the perdeuteriation of glutathione S-transferase from S. japonicum

    OpenAIRE

    Brockwell, David; Yu, Lu; Cooper, Serena; Mccleland, Steven; Cooper, Alan; Attwood, David; Gaskell, Simon J.; Barber, Jill

    2001-01-01

    Glutathione S-transferase (GST) from Schistosoma japonicum has been prepared in both normal protiated (pGST) and fully deuteriated (dGST) form by recombinant DNA technology. Electrospray mass spectrometry showed that the level of deuteriation in dGST was 96% and was homogeneous across the sample. This result is attributed to the use of a deuterium-tolerant host Escherichia coli strain in the preparation of the protein. 10 heteroatom-bound deuteriums (in addition to the carbon-bound deuteriums...

  8. Mechanism of activation of mouse liver microsomal glutations S—transferase by paracetamol treatment

    Institute of Scientific and Technical Information of China (English)

    ZhenY; LouYJ

    2002-01-01

    Microsomal glutathion S-transferase(mGST) is one of the important detoxifcation enzymes in vivo,its modifying activation by drugs has been paid more attention to in pertinent field recently.This study was to explore the influence of paracetamol(Par) on mGST and its possible mechanism in vivo,and to further reveal the biological significance.Par is metabolized to N-acetyl-p-benzoquinone imine(NAPQI) by CYP2E1 and mGST is activated by sulfhydryl modification.

  9. Fucosylation of xyloglucan: localization of the transferase in dictyosomes of pea stem cells

    International Nuclear Information System (INIS)

    Microsomal membranes from elongating regions of etiolated Pisum sativum stems were separated by rate-zonal centrifugation on Renografin gradients. The transfer of labeled fucose and xylose from GDP-[14C] fucose and UDP-[14C]xylose to xyloglucan occurred mainly in dictyosome-enriched fractions. No transferase activity was detected in secretory vesicle fractions. Pulse-chase experiments using pea stem slices incubated with [3H]fucose suggest that xyloglucan chains are fucosylated and their structure completed within the dictyosomes, before being transported to the cell wall by secretory vesicles

  10. Micronuclei rate and hypoxanthine phosphoribosyl transferase mutation in radon-exposed rats

    Institute of Scientific and Technical Information of China (English)

    Fengmei Cui; Saijun Fan; Mingjiang Hu; Jihua Nie; Hongmei Li; Jian Tong

    2008-01-01

    The genetic changes in rats with radon exposure were studied by the micronucleus technology and detection of hypoxanthine phosphoribosyl transferase (hprt) mutations.The rate of the micronuclei in peripheral blood lymphocytes and tracheal-bronchial epithelial cells in the radon-inhaled rats was higher than that of the controls (P < 0.05).A similar result was obtained from the hprt assay,which showed a higher mutation frequency in radon-exposed rats.Our results suggested that micronuclei rate and hprt deficiency could be used as biomarkers for the genetic changes with radon exposure.

  11. Association of catechol-o-methyl transferase gene polymorphism with prostate cancer and benign prostatic hyperplasia

    OpenAIRE

    Omrani, Mir Davood; Bazargani, Soroush; Bagheri, Morteza; Yazdan-nejad, Hamed

    2009-01-01

    BACKGROUND: A single nucleotide variation within catechol-o-methyl transferase (COMT) gene may alter the COMT enzyme activity level. Polymorphism of Val158Met in the COMT gene has been related to malignancy. In this regard, a study was carried out to find a possible association between the COMT gene polymorphism in patients with sporadic prostate cancer (PCa) and benign prostatic hyperplasia (BPH). METHODS: All types of COMT158 Val/Met polymorphism were carried out using ASO-PCR method in 41 ...

  12. Expression of glutathione transferases in corneal cell lines, corneal tissues and a human cornea construct.

    Science.gov (United States)

    Kölln, Christian; Reichl, Stephan

    2016-06-15

    Glutathione transferase (GST) expression and activity were examined in a three-dimensional human cornea construct and were compared to those of excised animal corneas. The objective of this study was to characterize phase II enzyme expression in the cornea construct with respect to its utility as an alternative to animal cornea models. The expression of the GSTO1-1 and GSTP1-1 enzymes was investigated using immunofluorescence staining and western blotting. The level of total glutathione transferase activity was determined using 1-chloro-2,4- dinitrobenzene as the substrate. Furthermore, the levels of GSTO1-1 and GSTP1-1 activity were examined using S-(4-nitrophenacyl)glutathione and ethacrynic acid, respectively, as the specific substrates. The expression and activity levels of these enzymes were examined in the epithelium, stroma and endothelium, the three main cellular layers of the cornea. In summary, the investigated enzymes were detected at both the protein and functional levels in the cornea construct and the excised animal corneas. However, the enzymatic activity levels of the human cornea construct were lower than those of the animal corneas. PMID:27113863

  13. Structural and thermodynamic properties of kappa class glutathione transferase from Camelus dromedarius.

    Science.gov (United States)

    Malik, Ajamaluddin; Fouad, Dalia; Labrou, Nikolaos E; Al-Senaidy, Abdulrahman M; Ismael, Mohamed A; Saeed, Hesham M; Ataya, Farid S

    2016-07-01

    The Arabian camel, Camelus dromedarius is naturally adapted to extreme desert climate and has evolved protective mechanisms to limit oxidative stress. The mitochondrial kappa class glutathione transferase enzyme is a member of GST supergene family that represents an important enzyme group in cellular Phase II detoxification machinery and is involved in the protection against oxidative stress and xenobiotics. In the present study, C. dromedarius kappa class glutathione transferase (CdGSTK1-1) was cloned, expressed in E. coli BL21, purified and its structural, thermodynamic and unfolding pathway was investigated. The results showed that CdGSTK1-1 has unique trimeric structure, exhibits low thermostability and a complex equilibrium unfolding profile. It unfolds through three folding states with formation of thinly populated intermediate species. The melting points (Tm) of the first unfolding transition was 40.3±0.2°C and Tm of the second unfolding transition was 49.1±0.1°C. The van't Hoff enthalpy of the first and second transition were 298.7±13.2 and 616.5±2.4kJ/mol, respectively. Moreover, intrinsic fluorescence and near-UV CD studies indicates that substrate binding does not leads to major conformational changes in CdGSTK1-1. PMID:27044344

  14. Characterisation of the Candida albicans Phosphopantetheinyl Transferase Ppt2 as a Potential Antifungal Drug Target.

    Directory of Open Access Journals (Sweden)

    Katharine S Dobb

    Full Text Available Antifungal drugs acting via new mechanisms of action are urgently needed to combat the increasing numbers of severe fungal infections caused by pathogens such as Candida albicans. The phosphopantetheinyl transferase of Aspergillus fumigatus, encoded by the essential gene pptB, has previously been identified as a potential antifungal target. This study investigated the function of its orthologue in C. albicans, PPT2/C1_09480W by placing one allele under the control of the regulatable MET3 promoter, and deleting the remaining allele. The phenotypes of this conditional null mutant showed that, as in A. fumigatus, the gene PPT2 is essential for growth in C. albicans, thus fulfilling one aspect of an efficient antifungal target. The catalytic activity of Ppt2 as a phosphopantetheinyl transferase and the acyl carrier protein Acp1 as a substrate were demonstrated in a fluorescence transfer assay, using recombinant Ppt2 and Acp1 produced and purified from E.coli. A fluorescence polarisation assay amenable to high-throughput screening was also developed. Therefore we have identified Ppt2 as a broad-spectrum novel antifungal target and developed tools to identify inhibitors as potentially new antifungal compounds.

  15. Association of ORCA/LRWD1 with repressive histone methyl transferases mediates heterochromatin organization.

    Science.gov (United States)

    Giri, Sumanprava; Prasanth, Supriya G

    2015-11-01

    Heterochromatin mostly constitutes tightly packaged DNA, decorated with repressive histone marks, including histone H3 methylated at lysine 9, histone H4 methylated at lysine 20 and histone H3 methylated at lysine 27. Each of these marks is incorporated by specific histone lysine methyl transferases. While constitutive heterochromatin enriched with H3K9me3 and H4K20me3 occur within repetitive elements, including centromeres and telomeres, the facultative heterochromatin resides on the inactive X-chromosome and contains H3K27me3 mark. Origin recognition complex-associated (ORCA/LRWD1) protein is required for the initiation of DNA replication and also plays crucial roles in heterochromatin organization. ORCA associates with constitutive and facultative heterochromatin in human cells and binds to repressive histone marks. We demonstrate that ORCA binds to multiple repressive histone methyl transferases including G9a, GLP, Suv39h1 (H3K9me2/3), Suv420h1/h2 (H4K20me2/3) and EZH2 (H3K27me3). Removal of ORCA from human cells causes aberrations in the chromatin architecture. We propose that ORCA acts as a scaffold protein that enables the formation of multiple histone lysine methyltransferase complexes at heterochromatic sites thereby facilitating chromatin organization. PMID:26765314

  16. Glucose-induced expression of MIP-1 genes requires O-GlcNAc transferase in monocytes

    Energy Technology Data Exchange (ETDEWEB)

    Chikanishi, Toshihiro [Institute of Molecular and Cellular Biosciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032 (Japan); ERATO, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi-shi, Saitama 332-0012 (Japan); Fujiki, Ryoji; Hashiba, Waka; Sekine, Hiroki; Yokoyama, Atsushi [ERATO, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi-shi, Saitama 332-0012 (Japan); Kato, Shigeaki, E-mail: uskato@mail.ecc.u-tokyo.ac.jp [Institute of Molecular and Cellular Biosciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032 (Japan); ERATO, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi-shi, Saitama 332-0012 (Japan)

    2010-04-16

    O-glycosylation has emerged as an important modification of nuclear proteins, and it appears to be involved in gene regulation. Recently, we have shown that one of the histone methyl transferases (MLL5) is activated through O-glycosylation by O-GlcNAc transferase (OGT). Addition of this monosaccharide is essential for forming a functional complex. However, in spite of the abundance of OGT in the nucleus, the impact of nuclear O-glycosylation by OGT remains largely unclear. To address this issue, the present study was undertaken to test the impact of nuclear O-glycosylation in a monocytic cell line, THP-1. Using a cytokine array, MIP-1{alpha} and -1{beta} genes were found to be regulated by nuclear O-glycosylation. Biochemical purification of the OGT interactants from THP-1 revealed that OGT is an associating partner for distinct co-regulatory complexes. OGT recruitment and protein O-glycosylation were observed at the MIP-1{alpha} gene promoter; however, the known OGT partner (HCF-1) was absent when the MIP-1{alpha} gene promoter was not activated. From these findings, we suggest that OGT could be a co-regulatory subunit shared by functionally distinct complexes supporting epigenetic regulation.

  17. A novel lysophosphatidylcholine acyl transferase activity is expressed by peroxiredoxin 6.

    Science.gov (United States)

    Fisher, Aron B; Dodia, Chandra; Sorokina, Elena M; Li, Haitao; Zhou, Suiping; Raabe, Tobias; Feinstein, Sheldon I

    2016-04-01

    The phospholipase A2(PLA2) activity of peroxiredoxin (Prdx)6 has important physiological roles in the synthesis of lung surfactant and in the repair of peroxidized cell membranes. These functions require the activity of a lysophospholipid acyl transferase as a critical component of the phospholipid remodeling pathway. We now describe a lysophosphatidylcholine acyl transferase (LPCAT) activity for Prdx6 that showed a strong preference for lysophosphatidylcholine (LPC) as the head group and for palmitoyl CoA in the acylation reaction. The calculated kinetic constants for acylation wereKm18 μM andVmax30 nmol/min/mg protein; theVmaxwas increased 25-fold by phosphorylation of the protein whileKmwas unchanged. Study of recombinant protein in vitro and in mouse pulmonary microvascular endothelial cells infected with a lentiviral vector construct indicated that amino acid D31 is crucial for LPCAT activity. A linear incorporation of labeled fatty acyl CoA into dipalmitoyl phosphatidylcholine (PC) indicated that LPC generated by Prdx6 PLA2activity remained bound to the enzyme for the reacylation reaction. Prdx6 is the first LPCAT enzyme with demonstrated cytoplasmic localization. Thus, Prdx6 is a complete enzyme comprising both PLA2and LPCAT activities for the remodeling pathway of PC synthesis or for repair of membrane lipid peroxidation. PMID:26830860

  18. Glutathione Transferases Superfamily: Cold-Inducible Expression of Distinct GST Genes in Brassica oleracea.

    Science.gov (United States)

    Vijayakumar, Harshavardhanan; Thamilarasan, Senthil Kumar; Shanmugam, Ashokraj; Natarajan, Sathishkumar; Jung, Hee-Jeong; Park, Jong-In; Kim, HyeRan; Chung, Mi-Young; Nou, Ill-Sup

    2016-01-01

    Plants, as sessile organisms, can suffer serious growth and developmental consequences under cold stress conditions. Glutathione transferases (GSTs, EC 2.5.1.18) are ubiquitous and multifunctional conjugating proteins, which play a major role in stress responses by preventing oxidative damage by reactive oxygen species (ROS). Currently, understanding of their function(s) during different biochemical and signaling pathways under cold stress condition remain unclear. In this study, using combined computational strategy, we identified 65 Brassica oleracea glutathione transferases (BoGST) and characterized them based on evolutionary analysis into 11 classes. Inter-species and intra-species duplication was evident between BoGSTs and Arabidopsis GSTs. Based on localization analyses, we propose possible pathways in which GST genes are involved during cold stress. Further, expression analysis of the predicted putative functions for GST genes were investigated in two cold contrasting genotypes (cold tolerance and susceptible) under cold condition, most of these genes were highly expressed at 6 h and 1 h in the cold tolerant (CT) and cold susceptible (CS) lines, respectively. Overall, BoGSTU19, BoGSTU24, BoGSTF10 are candidate genes highly expressed in B. oleracea. Further investigation of GST superfamily in B. oleracea will aid in understanding complex mechanism underlying cold tolerance in plants. PMID:27472324

  19. Structure of Human O-GlcNAc Transferase and its Complex with a Peptide Substrate

    Energy Technology Data Exchange (ETDEWEB)

    M Lazarus; Y Nam; J Jiang; P Sliz; S Walker

    2011-12-31

    The essential mammalian enzyme O-linked {beta}-N-acetylglucosamine transferase (O-GlcNAc transferase, here OGT) couples metabolic status to the regulation of a wide variety of cellular signalling pathways by acting as a nutrient sensor. OGT catalyses the transfer of N-acetylglucosamine from UDP-N-acetylglucosamine (UDP-GlcNAc) to serines and threonines of cytoplasmic, nuclear and mitochondrial proteins, including numerous transcription factors, tumour suppressors, kinases, phosphatases and histone-modifying proteins. Aberrant glycosylation by OGT has been linked to insulin resistance, diabetic complications, cancer and neurodegenerative diseases including Alzheimer's. Despite the importance of OGT, the details of how it recognizes and glycosylates its protein substrates are largely unknown. We report here two crystal structures of human OGT, as a binary complex with UDP (2.8 {angstrom} resolution) and as a ternary complex with UDP and a peptide substrate (1.95 {angstrom}). The structures provide clues to the enzyme mechanism, show how OGT recognizes target peptide sequences, and reveal the fold of the unique domain between the two halves of the catalytic region. This information will accelerate the rational design of biological experiments to investigate OGT's functions; it will also help the design of inhibitors for use as cellular probes and help to assess its potential as a therapeutic target.

  20. Crystallization and preliminary X-ray analysis of glutathione transferases from cyanobacteria

    International Nuclear Information System (INIS)

    Glutathione S-transferases (GSTs) are a group of detoxifying enzymes that are found in animals, plants and microorganisms. Here, the crystallizations of two cyanobacterial GSTs are reported with the aim of determining their atomic structures. Glutathione S-transferases (GSTs) are a group of multifunctional enzymes that are found in animals, plants and microorganisms. Their primary function is to remove toxins derived from exogenous sources or the products of metabolism from the cell. Mammalian GSTs have been extensively studied, in contrast to bacterial GSTs which have received relatively scant attention. A new class of GSTs called Chi has recently been identified in cyanobacteria. Chi GSTs exhibit a high glutathionylation activity towards isothiocyanates, compounds that are normally found in plants. Here, the crystallization of two GSTs are presented: TeGST produced by Thermosynechococcus elongates BP-1 and SeGST from Synechococcus elongates PCC 6301. Both enzymes formed crystals that diffracted to high resolution and appeared to be suitable for further X-ray diffraction studies. The structures of these GSTs may shed further light on the evolution of GST catalytic activity and in particular why these enzymes possess catalytic activity towards plant antimicrobial compounds

  1. The pleuromutilin drugs tiamulin and valnemulin bind to the RNA at the peptidyl transferase centre on the ribosome

    DEFF Research Database (Denmark)

    Poulsen, S M; Karlsson, M; Johansson, L B;

    2001-01-01

    The pleuromutilin antibiotic derivatives, tiamulin and valnemulin, inhibit protein synthesis by binding to the 50S ribosomal subunit of bacteria. The action and binding site of tiamulin and valnemulin was further characterized on Escherichia coli ribosomes. It was revealed that these drugs are...... strong inhibitors of peptidyl transferase and interact with domain V of 23S RNA, giving clear chemical footprints at nucleotides A2058-9, U2506 and U2584-5. Most of these nucleotides are highly conserved phylogenetically and functionally important, and all of them are at or near the peptidyl transferase...... centre and have been associated with binding of several antibiotics. Competitive footprinting shows that tiamulin and valnemulin can bind concurrently with the macrolide erythromycin but compete with the macrolide carbomycin, which is a peptidyl transferase inhibitor. We infer from these and previous...

  2. Mice Deficient in Glutathione Transferase Zeta/Maleylacetoacetate Isomerase Exhibit a Range of Pathological Changes and Elevated Expression of Alpha, Mu, and Pi Class Glutathione Transferases

    Science.gov (United States)

    Lim, Cindy E.L.; Matthaei, Klaus I.; Blackburn, Anneke C.; Davis, Richard P.; Dahlstrom, Jane E.; Koina, Mark E.; Anders, M.W.; Board, Philip G.

    2004-01-01

    Glutathione transferase zeta (GSTZ1-1) is the major enzyme that catalyzes the metabolism of α-halo acids such as dichloroacetic acid, a carcinogenic contaminant of chlorinated water. GSTZ1-1 is identical with maleylacetoacetate isomerase, which catalyzes the penultimate step in the catabolic pathways for phenylalanine and tyrosine. In this study we have deleted the Gstz1 gene in BALB/c mice and characterized their phenotype. Gstz1−/− mice do not have demonstrable activity with maleylacetone and α-halo acid substrates, and other GSTs do not compensate for the loss of this enzyme. When fed a standard diet, the GSTZ1-1-deficient mice showed enlarged liver and kidneys as well as splenic atrophy. Light and electron microscopic examination revealed multifocal hepatitis and ultrastructural changes in the kidney. The addition of 3% (w/v) phenylalanine to the drinking water was lethal for young mice (<28 days old) and caused liver necrosis, macrovesicular steatosis, splenic atrophy, and a significant loss of circulating leukocytes in older surviving mice. GSTZ1-1-deficient mice showed constitutive induction of alpha, mu, and pi class GSTs as well as NAD(P)H:quinone oxidoreductase 1. The overall response is consistent with the chronic accumulation of a toxic metabolite(s). We detected the accumulation of succinylacetone in the serum of deficient mice but cannot exclude the possibility that maleylacetoacetate and maleylacetone may also accumulate. PMID:15277241

  3. A study of the prognostic role of serum fucose and fucosyl transferase in cancer of the uterine cervix.

    Directory of Open Access Journals (Sweden)

    Sen,Urmi

    1985-04-01

    Full Text Available Serum fucose levels and fucosyl transferase activities have been designated as nonspecific markers of malignancy, and play an important role in the diagnosis of different types of malignancies. In the present study, attempts were made to determine the prognostic significance of these markers in patients with cancer of the uterine cervix after therapy. It was found that both serum fucose and fucosyl transferase, which were elevated in untreated patients declined significantly in patients responsive to therapy at different follow-up intervals, but not in patients unresponsive to therapy.

  4. Development of radioimmunoassay for gamma-glutamyl transferase using pancreatic enzyme

    International Nuclear Information System (INIS)

    A radioimmunoassay (RIA) for the determination of gamma-glutamyl transferase (GGT) was developed using human pancreatic enzyme as antigen. The assay allows the determination of GGT in concentrations as low as 80 ng/ml, and it is reproducible and specific. A good parallel relation was demonstrated between the standard curve and dilution curves for serum, urine, bile, and partially purified kidney GGT. In normal individuals, the mean serum concentration of GGT determined by RIA was found to be 3.43 μg/ml (SD+-1.20). Enzyme activity calculated from the GGT concentration measured by the radioimmunoassay using a regression equation was approximately twice as great as that determined by conventional enzyme assay. (author)

  5. Relation of Gamma-Glutamyl Transferase to Cardiovascular Events in Patients With Acute Coronary Syndromes.

    Science.gov (United States)

    Ndrepepa, Gjin; Braun, Siegmund; Cassese, Salvatore; Fusaro, Massimiliano; Laugwitz, Karl-Ludwig; Schunkert, Heribert; Kastrati, Adnan

    2016-05-01

    The prognostic value of gamma-glutamyl transferase (GGT) in patients with acute coronary syndromes (ACS) has been incompletely investigated. We investigated this clinically relevant question in 2,534 consecutive patients with ACS who underwent percutaneous coronary intervention (PCI). GGT activity was measured before PCI procedure in all patients. Statin therapy at hospital discharge was prescribed in 94% of the patients. The primary outcome was 3-year mortality. Patients were divided into 3 groups: the group with GGT in the first tertile (GGT estimates were calculated per SD increase in the logarithmic scale of GGT activity). In conclusion, in contemporary patients with ACS treated with PCI and on statin therapy, elevated GGT activity was associated with the increased risk of all-cause and noncardiac mortality but not with the risk of cardiac mortality. PMID:26956636

  6. Glutathione S-transferases variants as risk factors in Alzheimer's disease.

    Science.gov (United States)

    Wang, Tengfei

    2015-10-01

    Glutathione S-transferase (GST) was suggested as an important contributor to Alzheimer's disease (AD). The GSTs polymorphisms have been investigated as candidate genetic risk factors for AD, yet results remained uncertain. Therefore, we performed a meta-analysis to clarify the relationship of GSTs polymorphisms with the occurrence of AD. PubMed, Embase, Cochrane library and Alzgene databases were searched and potential literatures were selected. Pooled analyses and subgroup analyses were conducted, and also publication bias tests and cumulative meta-analysis. This meta-analysis suggested null associations between polymorphisms of GSTM1, GSTT1, GSTM3, GSTP1, GSTO1 and AD risk. GSTs variants may not have an impact on the morbidity of Alzheimer's disease. Further well designed researches are required to confirm these findings of the current study. PMID:25981226

  7. Differential roles of tau class glutathione S-transferases in oxidative stress

    DEFF Research Database (Denmark)

    Kilili, Kimiti G; Atanassova, Neli; Vardanyan, Alla;

    2004-01-01

    The plant glutathione S-transferase BI-GST has been identified as a potent inhibitor of Bax lethality in yeast, a phenotype associated with oxidative stress and disruption of mitochondrial functions. Screening of a tomato two-hybrid library for BI-GST interacting proteins identified five homologous...... Tau class GSTs, which readily form heterodimers between them and BI-GST. All six LeGSTUs were found to be able to protect yeast cells from prooxidant-induced cell death. The efficiency of each LeGSTU was prooxidant-specific, indicating a different role for each LeGSTU in the oxidative stress......-response mechanism. The prooxidant protective effect of all six proteins was suppressed in the absence of YAP1, a transcription factor that regulates hydroperoxide homeostasis in Saccharomyces cerevisiae, suggesting a role for the LeGSTUs in the context of the YAP1-dependent stress-responsive machinery. The...

  8. Biochemical properties of an omega-class glutathione S-transferase of the silkmoth, Bombyx mori.

    Science.gov (United States)

    Yamamoto, Kohji; Nagaoka, Sumiharu; Banno, Yutaka; Aso, Yoichi

    2009-05-01

    A cDNA encoding an omega-class glutathione S-transferase of the silkmoth, Bombyx mori (bmGSTO), was cloned by reverse transcriptase-polymerase chain reaction. The resulting clone was sequenced and deduced for amino acid sequence, which revealed 40, 40, and 39% identities to omega-class GSTs from human, pig, and mouse, respectively. A recombinant protein (rbmGSTO) was functionally overexpressed in Escherichia coli cells in a soluble form and purified to homogeneity. rbmGSTO was able to catalyze the biotranslation of glutathione with 1-chloro-2,4-dinitrobenzene, a model substrate for GST, as well as with 4-hydroxynonenal, a product of lipid peroxidation. This enzyme was shown to have high affinity for organophosphorus insecticide and was present abundantly in silkmoth strain exhibiting fenitrothion resistance. These results indicate that bmGSTO could be involved in the increase in level of insecticide resistance for lepidopteran insects. PMID:19022397

  9. Design of a monomeric human glutathione transferase GSTP1, a structurally stable but catalytically inactive protein.

    Science.gov (United States)

    Abdalla, Abdel-Monem; Bruns, Christopher M; Tainer, John A; Mannervik, Bengt; Stenberg, Gun

    2002-10-01

    By the introduction of 10 site-specific mutations in the dimer interface of human glutathione transferase P1-1 (GSTP1-1), a stable monomeric protein variant, GSTP1, was obtained. The monomer had lost the catalytic activity but retained the affinity for a number of electrophilic compounds normally serving as substrates for GSTP1-1. Fluorescence and circular dichroism spectra of the monomer and wild-type proteins were similar, indicating that there are no large structural differences between the subunits of the respective proteins. The GSTs have potential as targets for in vitro evolution and redesign with the aim of developing proteins with novel properties. To this end, a monomeric GST variant may have distinct advantages. PMID:12468717

  10. Response of Glutathione and Glutathione S-transferase in Rice Seedlings Exposed to Cadmium Stress

    Institute of Scientific and Technical Information of China (English)

    ZHANG Chun-hua; GE Ying

    2008-01-01

    A hydroponic culture experiment was done to investigate the effect of Cd stress on glutathione content(GSH)and glutathione S-transferase(GST,EC 2.5.1.18)activity in rice seedlings.The rice growth was severely inhibited when Cd level in the solution was higher than 10 mg/L.In rice shoots,GSH content and GST activity increased with the increasing Cd level,while in roots,GST was obviously inhibited by Cd treatments.Compared with shoots,the rice roots had higher GSH content and GST activity,indicating the ability of Cd detoxification was much higher in roots than in shoots.There was a significant correlation between Cd level and GSH content or GST activity,suggesting that both parameters may be used as biomarkers of Cd stress in rice.

  11. Structural basis for the interaction of antibiotics with peptidyl transferase center in eubacteria

    Energy Technology Data Exchange (ETDEWEB)

    Schlunzen, Frank; Zarivach, Raz; Harms, Jörg; Bashan, Anat; Tocilj, Ante; Albrecht, Renate; Yonath, Ada; Franceschi, Francois (WIS-I); (Max Planck Germany)

    2009-10-07

    Ribosomes, the site of protein synthesis, are a major target for natural and synthetic antibiotics. Detailed knowledge of antibiotic binding sites is central to understanding the mechanisms of drug action. Conversely, drugs are excellent tools for studying the ribosome function. To elucidate the structural basis of ribosome-antibiotic interactions, we determined the high-resolution X-ray structures of the 50S ribosomal subunit of the eubacterium Deinococcus radiodurans, complexed with the clinically relevant antibiotics chloramphenicol, clindamycin and the three macrolides erythromycin, clarithromycin and roxithromycin. We found that antibiotic binding sites are composed exclusively of segments of 23S ribosomal RNA at the peptidyl transferase cavity and do not involve any interaction of the drugs with ribosomal proteins. Here we report the details of antibiotic interactions with the components of their binding sites. Our results also show the importance of putative Mg{sup +2} ions for the binding of some drugs. This structural analysis should facilitate rational drug design.

  12. Serum glutathione transferase does not respond to indole-3-carbinol: A pilot study

    Directory of Open Access Journals (Sweden)

    Daniel R McGrath

    2010-05-01

    Full Text Available Daniel R McGrath1, Hamid Frydoonfar2, Joshua J Hunt3, Chris J Dunkley3, Allan D Spigelman41Ipswich Hospital, Ipswich, UK; 2Hunter Pathology Service, New South Wales; 3Royal Newcastle Centre, Newcastle; 4St Vincent’s Clinical School, Sydney, AustraliaBackground: Despite the well recognized protective effect of cruciferous vegetables against various cancers, including human colorectal cancers, little is known about how this effect is conferred. It is thought that some phytochemicals found only in these vegetables confer the protection. These compounds include the glucosinolates, of which indole-3-carbinol is one. They are known to induce carcinogen-metabolizing (phase II enzymes, including the glutathione S-transferase (GST family. Other effects in humans are not well documented. We wished to assess the effect of indole-3-carbinol on GST enzymes.Methods: We carried out a placebo-controlled human volunteer study. All patients were given 400 mg daily of indole-3-carbinol for three months, followed by placebo. Serum samples were tested for the GSTM1 genotype by polymerase chain reaction. Serum GST levels were assessed using enzyme-linked immunosorbent assay and Western Blot methodologies.Results: Forty-nine volunteers completed the study. GSTM1 genotypes were obtained for all but two volunteers. A slightly greater proportion of volunteers were GSTM1-positive, in keeping with the general population. GST was detected in all patients. Total GST level was not affected by indole-3-carbinol dosing compared with placebo. Although not statistically significant, the GSTM1 genotype affected the serum GST level response to indole-3-carbinol.Conclusion: Indole-3-carbinol does not alter total serum GST levels during prolonged dosing.Keywords: pilot study, colorectal cancer, glutathione transferase, human, indole-3-carbinol

  13. A glutathione transferase from Agrobacterium tumefaciens reveals a novel class of bacterial GST superfamily.

    Directory of Open Access Journals (Sweden)

    Katholiki Skopelitou

    Full Text Available In the present work, we report a novel class of glutathione transferases (GSTs originated from the pathogenic soil bacterium Agrobacterium tumefaciens C58, with structural and catalytic properties not observed previously in prokaryotic and eukaryotic GST isoenzymes. A GST-like sequence from A. tumefaciens C58 (Atu3701 with low similarity to other characterized GST family of enzymes was identified. Phylogenetic analysis showed that it belongs to a distinct GST class not previously described and restricted only in soil bacteria, called the Eta class (H. This enzyme (designated as AtuGSTH1-1 was cloned and expressed in E. coli and its structural and catalytic properties were investigated. Functional analysis showed that AtuGSTH1-1 exhibits significant transferase activity against the common substrates aryl halides, as well as very high peroxidase activity towards organic hydroperoxides. The crystal structure of AtuGSTH1-1 was determined at 1.4 Å resolution in complex with S-(p-nitrobenzyl-glutathione (Nb-GSH. Although AtuGSTH1-1 adopts the canonical GST fold, sequence and structural characteristics distinct from previously characterized GSTs were identified. The absence of the classic catalytic essential residues (Tyr, Ser, Cys distinguishes AtuGSTH1-1 from all other cytosolic GSTs of known structure and function. Site-directed mutagenesis showed that instead of the classic catalytic residues, an Arg residue (Arg34, an electron-sharing network, and a bridge of a network of water molecules may form the basis of the catalytic mechanism. Comparative sequence analysis, structural information, and site-directed mutagenesis in combination with kinetic analysis showed that Phe22, Ser25, and Arg187 are additional important residues for the enzyme's catalytic efficiency and specificity.

  14. Functional promiscuity correlates with conformational heterogeneity in A-class glutathione S-transferases.

    Science.gov (United States)

    Hou, Liming; Honaker, Matthew T; Shireman, Laura M; Balogh, Larissa M; Roberts, Arthur G; Ng, Kei-Cheuk; Nath, Abhinav; Atkins, William M

    2007-08-10

    The structurally related glutathione S-transferase isoforms GSTA1-1 and GSTA4-4 differ greatly in their relative catalytic promiscuity. GSTA1-1 is a highly promiscuous detoxification enzyme. In contrast, GSTA4-4 exhibits selectivity for congeners of the lipid peroxidation product 4-hydroxynonenal. The contribution of protein dynamics to promiscuity has not been studied. Therefore, hydrogen/deuterium exchange mass spectrometry (H/DX) and fluorescence lifetime distribution analysis were performed with glutathione S-transferases A1-1 and A4-4. Differences in local dynamics of the C-terminal helix were evident as expected on the basis of previous studies. However, H/DX demonstrated significantly greater solvent accessibility throughout most of the GSTA1-1 sequence compared with GSTA4-4. A Phe-111/Tyr-217 aromatic-aromatic interaction in A4-4, which is not present in A1-1, was hypothesized to increase core packing. "Swap" mutants that eliminate this interaction from A4-4 or incorporate it into A1-1 yield H/DX behavior that is intermediate between the wild type templates. In addition, the single Trp-21 residue of each isoform was exploited to probe the conformational heterogeneity at the intrasubunit domain-domain interface. Excited state fluorescence lifetime distribution analysis indicates that this core residue is more conformationally heterogeneous in GSTA1-1 than in GSTA4-4, and this correlates with greater stability toward urea denaturation for GSTA4-4. The fluorescence distribution and urea sensitivity of the mutant proteins were intermediate between the wild type templates. The results suggest that the differences in protein dynamics of these homologs are global. The results suggest also the possible importance of extensive conformational plasticity to achieve high levels of functional promiscuity, possibly at the cost of stability. PMID:17561509

  15. Polymerase θ is a robust terminal transferase that oscillates between three different mechanisms during end-joining

    Science.gov (United States)

    Kent, Tatiana; Mateos-Gomez, Pedro A; Sfeir, Agnel; Pomerantz, Richard T

    2016-01-01

    DNA polymerase θ (Polθ) promotes insertion mutations during alternative end-joining (alt-EJ) by an unknown mechanism. Here, we discover that mammalian Polθ transfers nucleotides to the 3’ terminus of DNA during alt-EJ in vitro and in vivo by oscillating between three different modes of terminal transferase activity: non-templated extension, templated extension in cis, and templated extension in trans. This switching mechanism requires manganese as a co-factor for Polθ template-independent activity and allows for random combinations of templated and non-templated nucleotide insertions. We further find that Polθ terminal transferase activity is most efficient on DNA containing 3’ overhangs, is facilitated by an insertion loop and conserved residues that hold the 3’ primer terminus, and is surprisingly more proficient than terminal deoxynucleotidyl transferase. In summary, this report identifies an unprecedented switching mechanism used by Polθ to generate genetic diversity during alt-EJ and characterizes Polθ as among the most proficient terminal transferases known. DOI: http://dx.doi.org/10.7554/eLife.13740.001 PMID:27311885

  16. LIGNIFICATION IN TRANSGENICS DEFICIENT IN P-COUMARATE 3-HYDROXYLASE (C3H) AND THE ASSOCIATED HYDROXYCINNAMOYL TRANSFERASE (HCT)

    Science.gov (United States)

    The effects on lignification of downregulating most of the genes for enzymes on the monolignol biosynthetic pathway have been reasonably well studied in angiosperms. The exception to this is the crucial hydroxylase, cinnamate 3-hydroxylase (C3H), and its associated hydroxycinnamyl transferase (HCT),...

  17. Tet Proteins Connect the O-Linked N-acetylglucosamine Transferase Ogt to Chromatin in Embryonic Stem Cells

    DEFF Research Database (Denmark)

    Vella, Pietro; Scelfo, Andrea; Jammula, Sriganesh;

    2013-01-01

    O-linked N-acetylglucosamine (O-GlcNAc) transferase (Ogt) activity is essential for embryonic stem cell (ESC) viability and mouse development. Ogt is present both in the cytoplasm and the nucleus of different cell types and catalyzes serine and threonine glycosylation. We have characterized the...

  18. Systemic catechol-O-methyl transferase inhibition enables the D1 agonist radiotracer R-[11C]SKF 82957

    DEFF Research Database (Denmark)

    Palner, Mikael; McCormick, Patrick; Parkes, Jun; Knudsen, Gitte M; Wilson, Alan A

    2010-01-01

    R-[(11)C]-SKF 82957 is a high-affinity and potent dopamine D(1) receptor agonist radioligand, which gives rise to a brain-penetrant lipophilic metabolite. In this study, we demonstrate that systemic administration of catechol-O-methyl transferase (COMT) inhibitors blocks this metabolic pathway...

  19. Crystal structure of a murine α-class glutathione S-transferase involved in cellular defense against oxidative stress

    NARCIS (Netherlands)

    Krengel, Ute; Schröter, Klaus-Hasso; Hoier, Helga; Arkema, Anita; Kalk, Kor H.; Zimniak, Piotr; Dijkstra, Bauke W.

    1998-01-01

    Glutathione S-transferases (GSTs) are ubiquitous multifunctional enzymes which play a key role in cellular detoxification. The enzymes protect the cells against toxicants by conjugating them to glutathione. Recently, a novel subgroup of α-class GSTs has been identified with altered substrate specifi

  20. Immunohistochemical localization of glutathione-S-transferase and glutathione peroxidase in adult Syrian hamster tissues and during kidney development.

    OpenAIRE

    Oberley, T. D.; Oberley, L. W.; Slattery, A. F.; Elwell, J. H.

    1991-01-01

    Tissues from adult Syrian hamsters were studied with immunoperoxidase techniques using polyclonal antibodies to glutathione-S-transferase (rat liver and human placental enzymes) and human erythrocyte glutathione peroxidase. Most tissues immunostained similarly with these antibodies. Most notable was the cytoplasmic staining of mesenchyme tissues, especially smooth muscle, by all three antibodies. Epithelial cells stained distinctively, but usually less intensely than mesenchyme. Epithelial ce...

  1. Purification of β-mannosyl transferase that synthesizes Man-β-GlcNAc-GlcNAc-PP-dolichol

    International Nuclear Information System (INIS)

    The β-mannosyl transferase that catalyzes the transfer of mannose from GDP-mannose to GlcNAc-GlcNAc-PP-dolichol (GlcNAc2-lipid) to form Man-β-GlcNAc2-lipid was solubilized from the microsomal fraction of pig aorta by treatment with 0.5% NP-40. The enzyme was purified about 115-fold using DEAE-cellulose, hydroxylapatite, and epoxy-activated Sepharose. The purified enzyme was free of α 1.3 and α1,6-mannosyl transferases since the only product seen when enzyme was incubated with GDP-[14C]-mannose and GlcNAc2-lipid was Man-β-GlcNAc2-lipid. The oligosaccharide portion of this lipid was released by mild acid hydrolysis and characterized as Man-β-GlcNAc-GlcNAc by gel filtration, as well as susceptibility to β-mannosidase and resistance to a α-mannosidase. This partially purified enzyme was stabilized by adding 20% glycerol and 0.5 mM dithiothreitol to the storage buffer. Thus, the transferase was stable for 5 or 6 days at 00 and could be kept for a month at -200. The activity was greatly stimulated by detergent with optimum activity being seen at 0.1% NP-40. However, phospholipids had no effect. The transferase had a pH optimum of 7.0, and showed an almost absolute requirement for Mg++, with maximum activity at 5 mM. The K/sub m/ for GDP-mannose was about 5 x 10-7 M, and for GlcNAc2-lipid about 1 x 10-6 M. The transferase was competitively inhibited by a variety of guanosine nucleotides

  2. Structural snapshots along the reaction pathway of Yersinia pestis RipA, a putative butyryl-CoA transferase

    Energy Technology Data Exchange (ETDEWEB)

    Torres, Rodrigo; Lan, Benson; Latif, Yama; Chim, Nicholas [UC Irvine, 2212 Natural Sciences I, Irvine, CA 92697 (United States); Goulding, Celia W., E-mail: celia.goulding@uci.edu [UC Irvine, 2212 Natural Sciences I, Irvine, CA 92697 (United States); UC Irvine, 2302 Natural Sciences I, Irvine, CA 92697 (United States)

    2014-04-01

    The crystal structures of Y. pestis RipA mutants were determined to provide insights into the CoA transferase reaction pathway. Yersinia pestis, the causative agent of bubonic plague, is able to survive in both extracellular and intracellular environments within the human host, although its intracellular survival within macrophages is poorly understood. A novel Y. pestis three-gene rip (required for intracellular proliferation) operon, and in particular ripA, has been shown to be essential for survival and replication in interferon γ-induced macrophages. RipA was previously characterized as a putative butyryl-CoA transferase proposed to yield butyrate, a known anti-inflammatory shown to lower macrophage-produced NO levels. RipA belongs to the family I CoA transferases, which share structural homology, a conserved catalytic glutamate which forms a covalent CoA-thioester intermediate and a flexible loop adjacent to the active site known as the G(V/I)G loop. Here, functional and structural analyses of several RipA mutants are presented in an effort to dissect the CoA transferase mechanism of RipA. In particular, E61V, M31G and F60M RipA mutants show increased butyryl-CoA transferase activities when compared with wild-type RipA. Furthermore, the X-ray crystal structures of E61V, M31G and F60M RipA mutants, when compared with the wild-type RipA structure, reveal important conformational changes orchestrated by a conserved acyl-group binding-pocket phenylalanine, Phe85, and the G(V/I)G loop. Binary structures of M31G RipA and F60M RipA with two distinct CoA substrate conformations are also presented. Taken together, these data provide CoA transferase reaction snapshots of an open apo RipA, a closed glutamyl-anhydride intermediate and an open CoA-thioester intermediate. Furthermore, biochemical analyses support essential roles for both the catalytic glutamate and the flexible G(V/I)G loop along the reaction pathway, although further research is required to fully

  3. Structural snapshots along the reaction pathway of Yersinia pestis RipA, a putative butyryl-CoA transferase

    International Nuclear Information System (INIS)

    The crystal structures of Y. pestis RipA mutants were determined to provide insights into the CoA transferase reaction pathway. Yersinia pestis, the causative agent of bubonic plague, is able to survive in both extracellular and intracellular environments within the human host, although its intracellular survival within macrophages is poorly understood. A novel Y. pestis three-gene rip (required for intracellular proliferation) operon, and in particular ripA, has been shown to be essential for survival and replication in interferon γ-induced macrophages. RipA was previously characterized as a putative butyryl-CoA transferase proposed to yield butyrate, a known anti-inflammatory shown to lower macrophage-produced NO levels. RipA belongs to the family I CoA transferases, which share structural homology, a conserved catalytic glutamate which forms a covalent CoA-thioester intermediate and a flexible loop adjacent to the active site known as the G(V/I)G loop. Here, functional and structural analyses of several RipA mutants are presented in an effort to dissect the CoA transferase mechanism of RipA. In particular, E61V, M31G and F60M RipA mutants show increased butyryl-CoA transferase activities when compared with wild-type RipA. Furthermore, the X-ray crystal structures of E61V, M31G and F60M RipA mutants, when compared with the wild-type RipA structure, reveal important conformational changes orchestrated by a conserved acyl-group binding-pocket phenylalanine, Phe85, and the G(V/I)G loop. Binary structures of M31G RipA and F60M RipA with two distinct CoA substrate conformations are also presented. Taken together, these data provide CoA transferase reaction snapshots of an open apo RipA, a closed glutamyl-anhydride intermediate and an open CoA-thioester intermediate. Furthermore, biochemical analyses support essential roles for both the catalytic glutamate and the flexible G(V/I)G loop along the reaction pathway, although further research is required to fully

  4. Two Pear Glutathione S-Transferases Genes Are Regulated during Fruit Development and Involved in Response to Salicylic Acid, Auxin, and Glucose Signaling

    OpenAIRE

    Hai-Yan Shi; Zheng-Hong Li; Yu-Xing Zhang; Liang Chen; Di-Ying Xiang; Yu-Feng Zhang

    2014-01-01

    Two genes encoding putative glutathione S-transferase proteins were isolated from pear (Pyrus pyrifolia) and designated PpGST1 and PpGST2. The deduced PpGST1 and PpGST2 proteins contain conserved Glutathione S-transferase N-terminal domain (GST_N) and Glutathione S-transferase, C-terminal domain (GST_C). Using PCR amplification technique, the genomic clones corresponding to PpGST1 and PpGST2 were isolated and shown to contain two introns and a singal intron respectively with typical GT/AG bou...

  5. Evaluation of gamma gluthamyl transferase and uric acid levels in arsenic exposed subject

    Directory of Open Access Journals (Sweden)

    Ceylan Bal

    2015-06-01

    Full Text Available Objective: Arsenic is a metal with a widespread industrial usage and causing oxidative stress. Studies shows serum uric acid and gamma gluthamyl transferase (GGT levels are increasing in oxidative stress. The aim of this study is to evaluate the effect of arsenic exposure on serum uric acid and GGT levels. Methods: 500 patients who refer to Ankara Occupational Disease Hospital between 2010 to 2014 for periodic examination and urinary arsenic, serum uric acid and serum GGT levels assessed are included in this study. 268 patients with urinary arsenic levels over 35μg/L are defined as exposed and below 35μg/L are controls. Results: Data of 500 patients were analysed. 268 of them had high urine arsenic levels and 232 had normal urine arsenic levels. In the high urine arsenic level group the median serum uric acid level was 5.4 (2.60-7.20 and median serum GGT level was 27 (10-51 in the other group with normal urine arsenic levels the median serum uric acid level was 4.9 (2.5-7 and median serum GGT level was 22 (10-52. The difference between two groups was statistically significant (p value: 0.002 and <0.001 respectively Conclusion: Arsenic exposure may be associated with hyperuricemia and high levels of GGT and with prospective studies the causal relationship between arsenic exposure and hyperuricemia and GGT can be revealed.

  6. Genomic heterogeneity and instability in colorectal cancer: spectral karyotyping, glutathione transferase-Ml and ras.

    Science.gov (United States)

    Bartos, Jeremy D; Stoler, Daniel L; Matsui, Sei-ichi; Swede, Helen; Willmott, Lyndsay J; Sait, Sheila N; Petrelli, Nicholas J; Anderson, Garth R

    2004-12-21

    Genomic instability in cancer is frequently described as being either chromosomal instability or microsatellite instability, although when events within chromosomes are monitored, extensive intrachromosomal instability is also found. Spectral karyotyping was used to visualize how extensively genomic instability gives rise to intratumor genomic heterogeneity in sporadic colorectal carcinomas. Two factors were then examined which might relate to intrachromosomal instability in colorectal cancers: the presence of the glutathione transferase-Ml gene to detoxify potential carcinogens, and the presence of activated ras which has been associated with chromosomal instability when first expressed. Intrachromosomal genomic instability was previously determined by inter-(simple sequence repeat) PCR (inter-SSR PCR) and by fractional allelic loss rate for 348 markers. GSTM1 status was determined for each of 49 tumors through use of specific PCR, and 28 of the tumors showed the GSTM1 null genotype. A significant association was found between GSTMl-null status and elevated inter-(simple sequence repeat) PCR instability. In contrast, no association was found with fractional allelic loss rate. The first exons of the K-ras and H-ras oncogenes were sequenced in 72 colorectal cancers; 19 of the tumors had a mutation in codon 12 of the K-ras gene (24.5%), but no H-ras mutations were found. A weak correlation (p=0.10) was observed between mutant K-ras and inter-(simple sequence repeat) PCR genomic instability, and no association existed with fractional allelic loss rate. PMID:15542115

  7. Development of pyrethroid-like fluorescent substrates for glutathione S-transferase

    Science.gov (United States)

    Huang, Huazhang; Yao, Hongwei; Liu, Jun-Yan; Samra, Aman I.; Kamita, Shizuo G.; Cornel, Anthony J.; Hammock, Bruce D.

    2012-01-01

    The availability of highly sensitive substrates is critical for the development of precise and rapid assays for detecting changes in glutathione S-transferase (GST) activity that are associated with GST-mediated metabolism of insecticides. In this study, six pyrethroid-like compounds were synthesized and characterized as substrates for insect and mammalian GSTs. All of the substrates were esters composed of the same alcohol moiety, 7-hydroxy-4-methylcoumarin, and acid moieties that structurally mimic some commonly used pyrethroid insecticides including cypermethrin and cyhalothrin. CpGSTD1, a recombinant Delta class GST from the mosquito Culex pipiens, metabolized our pyrethroid-like substrates with both chemical and geometric (i.e., the cis-isomers were metabolized at 2- to 5-fold higher rates than the corresponding trans-isomers) preference. A GST preparation from mouse liver also metabolized most of our pyrethroid-like substrates with both chemical and geometric preference but at 10- to 170-fold lower rates. CpGSTD1 and mouse GSTs metabolized CDNB, a general GST substrate, at more than 200-fold higher rates than our novel pyrethroid-like substrates. There was a 10-fold difference in the specificity constant (kcat/KM ratio) of CpGSTD1 for CDNB and those of CpGSTD1 for cis-DCVC and cis-TFMCVC suggesting that cis-DCVC and cis-TFMCVC may be useful for the detection of GST-based metabolism of pyrethroids in mosquitoes. PMID:23000005

  8. Glutathione S-transferase K1 genotype and overweight status in schizophrenia patients: A pilot study.

    Science.gov (United States)

    Oniki, Kentaro; Kamihashi, Ryoko; Tomita, Tetsu; Ishioka, Masamichi; Yoshimori, Yuki; Osaki, Natsumi; Tsuchimine, Shoko; Sugawara, Norio; Kajiwara, Ayami; Morita, Kazunori; Miyata, Keishi; Otake, Koji; Nakagawa, Kazuko; Ogata, Yasuhiro; Saruwatari, Junji; Yasui-Furukori, Norio

    2016-05-30

    Elevated oxidative stress in mitochondria and mitochondrial dysfunction are associated with weight gain in schizophrenia (SCZ) patients. Glutathione S-transferase kappa 1 (GSTK1) protects cells against exogenous and endogenous oxidative stress in the mitochondria. This exploratory study investigated the possible effects of a common GSTK1 polymorphism (rs1917760, G-1308T) on the risk for overweight status among 329 SCZ patients and 305 age- and gender-matched controls and on the GSTK1 mRNA level in peripheral blood mononuclear cells among 14 SCZ patients. The GSTK1 T/T genotype was associated with having a higher BMI value among SCZ male patients, whereas this genotype tended to be associated with a lower BMI value among female patients. Conversely, these associations were not observed among the controls. The GSTK1 T/T genotype was associated with decreased GSTK1 mRNA level among SCZ patients. The GSTK1 T/T genotype may be a novel risk factor for the prediction of overweight status in SCZ male patients, although the results of this pilot study should be verified by a larger study. PMID:27010189

  9. Purification and characterization of a glutathione S-transferase from Mucor mucedo.

    Science.gov (United States)

    Hamed, Ragaa R; Abu-Shady, Mohamed R; El-Beih, Fawkia M; Abdalla, Abdel-Monem A; Afifi, Ola M

    2005-01-01

    An intracellular glutathione transferase was purified to homogenity from the fungus, Mucor mucedo, using DEAE-cellulose ion-exchange and glutathione affinity chromatography. Gel filtration chromatography and SDS-PAGE revealed that the purified GST is a homodimer with approximate native and subunit molecular mass of 53 kDa and 23.4 kDa, respectively. The enzyme has a pI value of 4.8, a pH optimum at pH 8.0 and apparent activation energy (Ea) of 1.42 kcal mol(-1). The purified GST acts readily on CDNB with almost negligible peroxidase activity and the activity was inhibited by Cibacron Blue (IC50 0.252 microM) and hematin (IC50 3.55 microM). M. mucedo GST displayed a non-Michaelian behavior. At low (0.1-0.3 mM) and high (0.3-2 mM) substrate concentration, Km (GSH) was calculated to be 0.179 and 0.65 mM, whereas Km(CDNB) was 0.531 and 11 mM and k(cat) was 39.8 and 552 s(-1), respectively. The enzyme showed apparent pKa values of 6-6.5 and 8.0. PMID:16209109

  10. Glutathione S-Transferase Ω 1 variation does not influence age at onset of Huntington's disease

    Directory of Open Access Journals (Sweden)

    Saft Carsten

    2004-03-01

    Full Text Available Abstract Background Huntington's disease (HD is a fully penetrant, autosomal dominantly inherited disorder associated with abnormal expansions of a stretch of perfect CAG repeats in the 5' part of the IT15 gene. The number of repeat units is highly predictive for the age at onset (AO of the disorder. But AO is only modestly correlated with repeat length when intermediate HD expansions are considered. Circumstantial evidence suggests that additional features of the HD course are based on genetic traits. Therefore, it may be possible to investigate the genetic background of HD, i.e. to map the loci underlying the development and progression of the disease. Recently an association of Glutathione S-Transferase Ω 1 (GSTO1 and possibly of GSTO2 with AO was demonstrated for, both, Alzheimer's (AD and Parkinson's disease (PD. Methods We have genotyped the polymorphisms rs4925 GSTO1 and rs2297235 GSTO2 in 232 patients with HD and 228 controls. Results After genotyping GSTO1 and GSTO2 polymorphisms, firstly there was no statistically significant difference in AO for HD patients, as well as secondly for HD patients vs. controls concerning, both, genotype and allele frequencies, respectively. Conclusion The GSTO1 and GSTO2 genes flanked by the investigated polymorphisms are not comprised in a primary candidate region influencing AO in HD.

  11. Septins guide microtubule protrusions induced by actin-depolymerizing toxins like Clostridium difficile transferase (CDT).

    Science.gov (United States)

    Nölke, Thilo; Schwan, Carsten; Lehmann, Friederike; Østevold, Kristine; Pertz, Olivier; Aktories, Klaus

    2016-07-12

    Hypervirulent Clostridium difficile strains, which are associated with increased morbidity and mortality, produce the actin-ADP ribosylating toxin Clostridium difficile transferase (CDT). CDT depolymerizes actin, causes formation of microtubule-based protrusions, and increases pathogen adherence. Here, we show that septins (SEPT) are essential for CDT-induced protrusion formation. SEPT2, -6, -7, and -9 accumulate at predetermined protrusion sites and form collar-like structures at the base of protrusions. The septin inhibitor forchlorfenuron or knockdown of septins inhibits protrusion formation. At protrusion sites, septins colocalize with the GTPase Cdc42 (cell division control protein 42) and its effector Borg (binder of Rho GTPases), which act as up-stream regulators of septin polymerization. Precipitation and surface plasmon resonance studies revealed high-affinity binding of septins to the microtubule plus-end tracking protein EB1, thereby guiding incoming microtubules. The data suggest that CDT usurps conserved regulatory principles involved in microtubule-membrane interaction, depending on septins, Cdc42, Borgs, and restructuring of the actin cytoskeleton. PMID:27339141

  12. Effects of gestational and overt diabetes on placental cytochromes P450 and glutathione S-transferase.

    Science.gov (United States)

    Glover; McRobie; Tracy

    1998-07-01

    Objective: Animal and in vivo human studies have observed that diabetes alters the expression of hepatic metabolizing cytochrome P450 (CYP) and glutathione S-transferase (GST) enzymes. The placenta has the ability to metabolize a number of xenobiotic and endogenous compounds by processes similar to those seen in the liver. Our objective was to compare placental xenobiotic metabolizing activity in diabetics to matched non-diabetic controls to determine if the presence of diabetes alters placental xenobiotic metabolizing activity.Methods: The catalytic activities of 7-ethoxyresorufin-O-deethylation [EROD] (CYP1A1), chlorzoxazone 6-hydroxylation (CYP2E1), dextromethorphan N-demethylation (CYP3A4), dextromethorphan O-demethylation (CYP2D6), and 1-chloro-2,4-dinitrobenzene (CDNB) conjugation with glutathione (GST) from placentas of diet controlled (class A1) and insulin-dependent (class A2) gestational diabetics and overt diabetics were compared to matched controls.Results: No differences in EROD activity were observed among overt or gestational diabetics and their respectively matched controls. CYP2E1, 2D6, and 3A4 enzyme activity were not detected in human placentas. In contrast, GST activity was significantly reduced by 30% (P <.05) in overt diabetics as compared to their matched controls and gestational diabetics.Conclusion: Pregnant women with overt diabetes have reduced GST activity in the placenta, which could potentially result in exposure of the fetus to harmful reactive electrophilic metabolites. PMID:10838356

  13. Prevalence of glutathione S-transferase gene deletions and their effect on sickle cell patients

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    Pandey Sanjay

    2012-01-01

    Full Text Available BACKGROUND: Glutathione S-transferase gene deletions are known detoxification agents and cause oxidative damage. Due to the different pathophysiology of anemia in thalassemia and sickle cell disease, there are significant differences in the pathophysiology of iron overload and iron-related complications in these disorders. OBJECTIVE: The aim of this study was to estimate the frequency of the GSTM1 and GSTT1 genotypes in sickle cell disease patients and their effect on iron status. METHODS: Forty sickle cell anemia and sixty sickle ß-thalassemia patients and 100 controls were evaluated to determine the frequency of GST gene deletions. Complete blood counts were performed by an automated cell analyzer. Hemoglobin F, hemoglobin A, hemoglobin A2 and hemoglobin S were measured and diagnosis of patients was achieved by high performance liquid chromatography with DNA extraction by the phenol-chloroform method. The GST null genotype was determined using multiplex polymerase chain reaction and serum ferritin was measured using an ELISA kit. Statistical analysis was by EpiInfo and GraphPad statistics software. RESULTS: An increased frequency of the GSTT1 null genotype (p-value = 0.05 was seen in the patients. The mean serum ferritin level was higher in patients with the GST genotypes than in controls; this was statistically significant for all genotypes except GSTM1, however the higher levels of serum ferritin were due to blood transfusions in patients. CONCLUSION: GST deletions do not play a direct role in iron overload of sickle cell patients.

  14. PLLA-PCys co-electrospun fibers for capture and elution of glutathione S-transferase

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    The copolymer poly(L-lactic acid)-b-poly(L-cysteine) (PLA-b-PCys) was co-electrospun with PLGA into ultrafine fibers. The reduced glutathione (GSH) was conjugated to the fiber surfaces via disulfide bonds. The glutathione S-transferase (GST) was captured onto the GSH fibers via specific substrate-enzyme interaction between the bound GSH and GST. The captured GST was eluted with free GSH aqueous solution and lyophilized to get pure GST powders. The results show that the GSH moieties on the fiber surface retain the bioactivity of the free GSH and thus they can bind specifically with GST and the GST in solution is captured onto the fiber surface. In addition, the bound GSH is not as active as free GSH so that the captured GST can be eluted off from the fiber by free GSH aqueous solution. Based on this principle, GST itself or its fused proteins can be separated and purified very easily. The preliminary purification efficiency is 6.5 mg·(gPCys)-1. Further improvements are undertaken.

  15. Prolactin confers resistance against cisplatin in breast cancer cells by activating glutathione-S-transferase.

    Science.gov (United States)

    LaPensee, Elizabeth W; Schwemberger, Sandy J; LaPensee, Christopher R; Bahassi, El Mustapha; Afton, Scott E; Ben-Jonathan, Nira

    2009-08-01

    Resistance to chemotherapy is a major obstacle for successful treatment of breast cancer patients. Given that prolactin (PRL) acts as an anti-apoptotic/survival factor in the breast, we postulated that it antagonizes cytotoxicity by chemotherapeutic drugs. Treatment of breast cancer cells with PRL caused variable resistance to taxol, vinblastine, doxorubicin and cisplatin. PRL prevented cisplatin-induced G(2)/M cell cycle arrest and apoptosis. In the presence of PRL, significantly less cisplatin was bound to DNA, as determined by mass spectroscopy, and little DNA damage was seen by gamma-H2AX staining. PRL dramatically increased the activity of glutathione-S-transferase (GST), which sequesters cisplatin in the cytoplasm; this increase was abrogated by Jak and mitogen-activated protein kinase inhibitors. PRL upregulated the expression of the GSTmu, but not the pi, isozyme. A GST inhibitor abrogated antagonism of cisplatin cytotoxicity by PRL. In conclusion, PRL confers resistance against cisplatin by activating a detoxification enzyme, thereby reducing drug entry into the nucleus. These data provide a rational explanation for the ineffectiveness of cisplatin in breast cancer, which is characterized by high expression of both PRL and its receptor. Suppression of PRL production or blockade of its actions should benefit patients undergoing chemotherapy by allowing for lower drug doses and expanded drug options. PMID:19443905

  16. The role of glutathione-S-transferase polymorphisms in ovarian cancer survival.

    Science.gov (United States)

    Nagle, Christina M; Chenevix-Trench, Georgia; Spurdle, Amanda B; Webb, Penelope M

    2007-01-01

    Resistance to chemotherapy represents one of the most important causes of treatment failure in patients with ovarian cancer. Common polymorphisms in the glutathione-S-transferase (GSTM1, GSTP1 and GSTT1) family have been implicated in chemoresistence and ovarian cancer survival. In this study, we have analysed Australian women diagnosed with primary invasive epithelial ovarian cancer between 1985 and 1997, using DNA extracted from peripheral blood and archival uninvolved (normal) tissues. GSTP1 genotypes were determined using ABI Prism 7700 Sequence Detection System methodology (n=448) and GSTT1 and GSTM1 genotypes using PCR-agarose methodology (n=239). We observed a significant survival advantage among carriers of GSTP1 Ile105Val GG/GA genotype (HR 0.77, 95% confidence interval (CI) 0.61-0.99,p=0.04) and a non-significant survival advantage among women who were homozygous for the GSTM1 and GSTT1 deletion variants. There was also evidence of an additive effect, with a stronger survival benefit in women carrying three low function GST genotypes (GSTM1 null, GSTT1 null and GSTP1 GA/GG) (HR 0.47, 95% CI 0.22-1.02). The results of this study, the largest to date, are consistent with a number of previous smaller studies which have also observed that reduced GST function was associated with better survival outcomes in patients with ovarian cancer. PMID:17084623

  17. Immunoprophylactic potential of filarial glutathione-s-transferase in lymphatic filariaisis

    Institute of Scientific and Technical Information of China (English)

    BalM; MandalN; AcharyKG; DasMK; KarSK

    2011-01-01

    Objective:To elucidates the immunoprophylactic potential of glutathion-s-transferase (GST) from cattle filarial parasite Setaria digitata (S. digitata) against lymphatic filariasis. Methods:GST was purified through affinity chromatography (SdGST) and chacterized by SDS-PAGE and Nano-LC MS/MS analysis. Antibody isotypes to SdGST were measured by ELISA. Antibody dependant cellular cytotoxicity (ADCC) was performed in vitro using sera from immunized animals and immune individuals. T-cell proliferation and cytokine response to SdGST in different groups of filariasis were measured. Immunoprophylactic potential of SdGST was evaluate in animal model. Results: SdGST exhibited 30-fold enhancement of enzyme activity over crude parasitic extract. It was found to be 26 kDa by SDS-PAGE. Nano LC-MS/MS analysis followed by blast search showed 100%homology with Dirofilaria immitis (D. immitis) and only 43%with Homo sapiens (H. sapiens). Immunoblotting analysis showed putatively immune individuals carry significant level of antibodies to SdGST as compared with microfilaraemics. Immunized sera and sera endemic normal could neutralize the enzymatic activity of SdGST and inducing in vitro cytotoxicity of microfilariae. Peripheral blood mononuclear cells (PBMC) from endemic normals upon stimulation with SdGST showed a mixed type of Th1/Th2 response. SdGST immunization clear microfilariae from circulation in S. digitata implanted mastomys. Conclusions:The heterologous GST could be potentially developed as a vaccine candidate against lymphatic filarial parasite.

  18. Recognition and Detoxification of the Insecticide DDT by Drosophila melanogaster Glutathione S-Transferase D1

    Energy Technology Data Exchange (ETDEWEB)

    Low, Wai Yee; Feil, Susanne C.; Ng, Hooi Ling; Gorman, Michael A.; Morton, Craig J.; Pyke, James; McConville, Malcolm J.; Bieri, Michael; Mok, Yee-Foong; Robin, Charles; Gooley, Paul R.; Parker, Michael W.; Batterham, Philip (SVIMR-A); (Melbourne)

    2010-06-14

    GSTD1 is one of several insect glutathione S-transferases capable of metabolizing the insecticide DDT. Here we use crystallography and NMR to elucidate the binding of DDT and glutathione to GSTD1. The crystal structure of Drosophila melanogaster GSTD1 has been determined to 1.1 {angstrom} resolution, which reveals that the enzyme adopts the canonical GST fold but with a partially occluded active site caused by the packing of a C-terminal helix against one wall of the binding site for substrates. This helix would need to unwind or be displaced to enable catalysis. When the C-terminal helix is removed from the model of the crystal structure, DDT can be computationally docked into the active site in an orientation favoring catalysis. Two-dimensional {sup 1}H,{sup 15}N heteronuclear single-quantum coherence NMR experiments of GSTD1 indicate that conformational changes occur upon glutathione and DDT binding and the residues that broaden upon DDT binding support the predicted binding site. We also show that the ancestral GSTD1 is likely to have possessed DDT dehydrochlorinase activity because both GSTD1 from D. melanogaster and its sibling species, Drosophila simulans, have this activity.

  19. A glutathione S-transferase gene associated with antioxidant properties isolated from Apis cerana cerana

    Science.gov (United States)

    Liu, Shuchang; Liu, Feng; Jia, Haihong; Yan, Yan; Wang, Hongfang; Guo, Xingqi; Xu, Baohua

    2016-06-01

    Glutathione S-transferases (GSTs) are an important family of multifunctional enzymes in aerobic organisms. They play a crucial role in the detoxification of exogenous compounds, especially insecticides, and protection against oxidative stress. Most previous studies of GSTs in insects have largely focused on their role in insecticide resistance. Here, we isolated a theta class GST gene designated AccGSTT1 from Apis cerana cerana and aimed to explore its antioxidant and antibacterial attributes. Analyses of homology and phylogenetic relationships suggested that the predicted amino acid sequence of AccGSTT1 shares a high level of identity with the other hymenopteran GSTs and that it was conserved during evolution. Quantitative real-time PCR showed that AccGSTT1 is most highly expressed in adult stages and that the expression profile of this gene is significantly altered in response to various abiotic stresses. These results were confirmed using western blot analysis. Additionally, a disc diffusion assay showed that a recombinant AccGSTT1 protein may be roughly capable of inhibiting bacterial growth and that it reduces the resistance of Escherichia coli cells to multiple adverse stresses. Taken together, these data indicate that AccGSTT1 may play an important role in antioxidant processes under adverse stress conditions.

  20. Expression profiling of selected glutathione transferase genes in Zea mays (L. seedlings infested with cereal aphids.

    Directory of Open Access Journals (Sweden)

    Hubert Sytykiewicz

    Full Text Available The purpose of this report was to evaluate the expression patterns of selected glutathione transferase genes (gst1, gst18, gst23 and gst24 in the tissues of two maize (Zea mays L. varieties (relatively resistant Ambrozja and susceptible Tasty Sweet that were colonized with oligophagous bird cherry-oat aphid (Rhopalosiphum padi L. or monophagous grain aphid (Sitobion avenae L.. Simultaneously, insect-triggered generation of superoxide anion radicals (O2•- in infested Z. mays plants was monitored. Quantified parameters were measured at 1, 2, 4, 8, 24, 48 and 72 h post-initial aphid infestation (hpi in relation to the non-infested control seedlings. Significant increases in gst transcript amounts were recorded in aphid-stressed plants in comparison to the control seedlings. Maximal enhancement in the expression of the gst genes in aphid-attacked maize plants was found at 8 hpi (gst23 or 24 hpi (gst1, gst18 and gst24 compared to the control. Investigated Z. mays cultivars formed excessive superoxide anion radicals in response to insect treatments, and the highest overproduction of O2•- was noted 4 or 8 h after infestation, depending on the aphid treatment and maize genotype. Importantly, the Ambrozja variety could be characterized as having more profound increments in the levels of gst transcript abundance and O2•- generation in comparison with the Tasty Sweet genotype.

  1. Expression profiling of selected glutathione transferase genes in Zea mays (L.) seedlings infested with cereal aphids.

    Science.gov (United States)

    Sytykiewicz, Hubert; Chrzanowski, Grzegorz; Czerniewicz, Paweł; Sprawka, Iwona; Łukasik, Iwona; Goławska, Sylwia; Sempruch, Cezary

    2014-01-01

    The purpose of this report was to evaluate the expression patterns of selected glutathione transferase genes (gst1, gst18, gst23 and gst24) in the tissues of two maize (Zea mays L.) varieties (relatively resistant Ambrozja and susceptible Tasty Sweet) that were colonized with oligophagous bird cherry-oat aphid (Rhopalosiphum padi L.) or monophagous grain aphid (Sitobion avenae L.). Simultaneously, insect-triggered generation of superoxide anion radicals (O2•-) in infested Z. mays plants was monitored. Quantified parameters were measured at 1, 2, 4, 8, 24, 48 and 72 h post-initial aphid infestation (hpi) in relation to the non-infested control seedlings. Significant increases in gst transcript amounts were recorded in aphid-stressed plants in comparison to the control seedlings. Maximal enhancement in the expression of the gst genes in aphid-attacked maize plants was found at 8 hpi (gst23) or 24 hpi (gst1, gst18 and gst24) compared to the control. Investigated Z. mays cultivars formed excessive superoxide anion radicals in response to insect treatments, and the highest overproduction of O2•- was noted 4 or 8 h after infestation, depending on the aphid treatment and maize genotype. Importantly, the Ambrozja variety could be characterized as having more profound increments in the levels of gst transcript abundance and O2•- generation in comparison with the Tasty Sweet genotype. PMID:25365518

  2. Role of glutathione S-transferases in the spinocerebellar ataxia type 2 clinical phenotype.

    Science.gov (United States)

    Almaguer-Gotay, D; Almaguer-Mederos, L E; Aguilera-Rodríguez, R; Estupiñán-Rodríguez, A; González-Zaldivar, Y; Cuello-Almarales, D; Laffita-Mesa, J M; Vázquez-Mojena, Y

    2014-06-15

    Spinocerebellar ataxia type 2 (SCA2) is a neurodegenerative and incurable hereditary disorder caused by a CAG repeat expansion mutation on ATXN2 gene. The identification of reliable biochemical markers of disease severity is of paramount significance for the development and assessment of clinical trials. In order to evaluate the potential use of glutathione-S-transferase (GST) activity as a biomarker for SCA2, a case-control study in 38 affected, presymptomatic individuals or healthy controls was conducted. An enlarged sample of 121 affected individuals was set to assess the impact of GST activity on SCA2 clinical expression. There was a significant increase in GST activity in affected individuals relative to controls, although sensibility and specificity were not high. GST activity was not significantly influenced by sex, age, disease duration or CAG repeat size and did not significantly influence disease severity markers. These findings show a disruption of in vivo GST activity in SCA2, suggesting a role for oxidative stress in the neurodegenerative process. PMID:24780439

  3. Immunohistochemical localization of glutathione S-transferase-pi in human colorectal polyps

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    AIM: To investigate the distribution of the placental form of glutathione-S-transferase (GST) in colon polyps in order to evaluate the role of GST-pi in these tissues. METHODS: Sixteen polyp tissues removed at colon- oscopy were examined. Tissues were investigated his- tologicaUy and ultrastructurally. GST-pi expression was also analysed immunohistochemically, using peroxidase anti-peroxidase (PAP) method and immunogold label- ling method, for light and electron microscope respec- tively. RESULTS: All polyp tissues examined were adenoma of low, mild and high- grade dysplasia as shown in the histopathological reports. Nevertheless, the examina- tion of the above specimens with electron microscope revealed that 3 of 9 adenoma of mild dysplasia had ultrastuctural features similar to high-grade dysplasia adenoma. GST-pi was variably expressed in adenoma, with the lowest relative levels occurring in low-grade adenoma and the highest levels found in high-grade adenoma. GST-pi was located mainly in undifferentiat- ed epithelial cells. GST-pi positive particles were found in the cytoplasm and especially in the nucleus adjacent to the nuclear membrane of these cells. CONCLUSION: The overexpression of GST-pi in mild- grade adenomas with significant subcellular changes and in the majority of high-grade dysplasia adenoma suggests that this might be related to the carcinogenetic proceeding. Immunohistochemical localization of GST-pi in combination with ultrastructural changes indicate that GST-pi might be a sensitive agent for the detection of preneoplastic transformations in adenoma.

  4. Highly ordered protein nanorings designed by accurate control of glutathione S-transferase self-assembly.

    Science.gov (United States)

    Bai, Yushi; Luo, Quan; Zhang, Wei; Miao, Lu; Xu, Jiayun; Li, Hongbin; Liu, Junqiu

    2013-07-31

    Protein self-assembly into exquisite, complex, yet highly ordered architectures represents the supreme wisdom of nature. However, precise manipulation of protein self-assembly behavior in vitro is a great challenge. Here we report that by taking advantage of the cooperation of metal-ion-chelating interactions and nonspecific protein-protein interactions, we achieved accurate control of the orientation of proteins and their self-assembly into protein nanorings. As a building block, we utilized the C2-symmetric protein sjGST-2His, a variant of glutathione S-transferase from Schistosoma japonicum having two properly oriented His metal-chelating sites on the surface. Through synergic metal-coordination and non-covalent interactions, sjGST-2His self-assembled in a fixed bending manner to form highly ordered protein nanorings. The diameters of the nanorings can be regulated by tuning the strength of the non-covalent interaction network between sjGST-2His interfaces through variation of the ionic strength of the solution. This work provides a de novo design strategy that can be applied in the construction of novel protein superstructures. PMID:23865524

  5. Increased transcription of Glutathione S-transferases in acaricide exposed scabies mites

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    Currie Bart J

    2010-05-01

    Full Text Available Abstract Background Recent evidence suggests that Sarcoptes scabiei var. hominis mites collected from scabies endemic communities in northern Australia show increasing tolerance to 5% permethrin and oral ivermectin. Previous findings have implicated detoxification pathways in developing resistance to these acaricides. We investigated the contribution of Glutathione S-transferase (GST enzymes to permethrin and ivermectin tolerance in scabies mites using biochemical and molecular approaches. Results Increased in vitro survival following permethrin exposure was observed in S. scabiei var. hominis compared to acaricide naïve mites (p in vitro permethrin susceptibility, confirming GST involvement in permethrin detoxification. Assay of GST enzymatic activity in mites demonstrated that S. scabiei var. hominis mites showed a two-fold increase in activity compared to naïve mites (p S. scabiei var. canis- mu 1 (p S. scabiei var. hominis mites collected from a recurrent crusted scabies patient over the course of ivermectin treatment. Conclusions These findings provide further support for the hypothesis that increased drug metabolism and efflux mediate permethrin and ivermectin resistance in scabies mites and highlight the threat of emerging acaricide resistance to the treatment of scabies worldwide. This is one of the first attempts to define specific genes involved in GST mediated acaricide resistance at the transcriptional level, and the first application of such studies to S. scabiei, a historically challenging ectoparasite.

  6. Transcriptional Responses of Glutathione Transferase Genes in Ruditapes philippinarum Exposed to Microcystin-LR

    Directory of Open Access Journals (Sweden)

    Bruno Reis

    2015-04-01

    Full Text Available Glutathione Transferases (GSTs are phase II detoxification enzymes known to be involved in the molecular response against microcystins (MCs induced toxicity. However, the individual role of the several GST isoforms in the MC detoxification process is still unknown. In this study, the time-dependent changes on gene expression of several GST isoforms (pi, mu, sigma 1, sigma 2 in parallel with enzymatic activity of total GST were investigated in gills and hepatopancreas of the bivalve Ruditapes philippinarum exposed to pure MC-LR (10 and 100 µg/L. No significant changes in GST enzyme activities were found on both organs. In contrast, MC-LR affected the transcriptional activities of these detoxification enzymes both in gills and hepatopancreas. GST transcriptional changes in gills promoted by MC-LR were characterized by an early (12 h induction of mu and sigma 1 transcripts. On the other hand, the GST transcriptional changes in hepatopancreas were characterized by a later induction (48 h of mu transcript, but also by an early inhibition (6 h of the four transcripts. The different transcription patterns obtained for the tested GST isoforms in this study highlight the potential divergent physiological roles played by these isoenzymes during the detoxification of MC-LR.

  7. Measurement of mouse liver glutathione S-transferase activity by the integrated method

    Institute of Scientific and Technical Information of China (English)

    廖飞; 李甲初; 康格非; 曾昭淳; 左渝萍

    2003-01-01

    Objective: The integrated method was investigated to measure Vm/Km of mouse liver glutathione S-transferase (GST) activity on GSH and 7-Cl-4-nitrobenzofurazozan. Methods: Presetting concentration of one substrate twenty-fold above the others and taking maximum product absorbance Am as parameter while Km as constant, Vm/Km was obtained by nonlinear fitting of GST reaction curve to the integrated Michaelis-Menten equation ln [Am/(Am-Ai)]+Ai/(ε×Km)=(Vm/Km)×ti (1). Results: Vm/Km for GST showed slight dependence on initial substrate concentration and data range, but it was resistant to background absorbance, error in reaction origin and small deviation in presetting Km. Vm/Km was proportional to the amount of GST with upper limit higher than that by initial rate. There was close correlation between Vm/Km and initial rate of the same GST. Consistent results were obtained by this integrated method and classical initial rate method for the measurement of mouse liver GST. Conclusion: With the concentration of one substrate twenty-fold above the others, this integrated method was reliable to measure the activity of enzyme on two substrates, and substrate concentration of the lower one close to its apparent Km was able to be used.

  8. Antibiotic inhibition of the movement of tRNA substrates through a peptidyl transferase cavity

    DEFF Research Database (Denmark)

    Porse, B T; Rodriguez-Fonseca, C; Leviev, I;

    1996-01-01

    The present review attempts to deal with movement of tRNA substrates through the peptidyl transferase centre on the large ribosomal subunit and to explain how this movement is interrupted by antibiotics. It builds on the concept of hybrid tRNA states forming on ribosomes and on the observed...... movement of the 5' end of P-site-bound tRNA relative to the ribosome that occurs on peptide bond formation. The 3' ends of the tRNAs enter, and move through, a catalytic cavity where antibiotics are considered to act by at least three primary mechanisms: (i) they interfere with the entry of the aminoacyl...... moiety into the catalytic cavity before peptide bond formation; (ii) they inhibit movement of the nascent peptide along the peptide channel, a process that may generally involve destabilization of the peptidyl tRNA, and (iii) they prevent movement of the newly deacylated tRNA between the P/P and hybrid P...

  9. Identification of Small-Molecule Frequent Hitters of Glutathione S-Transferase-Glutathione Interaction.

    Science.gov (United States)

    Brenke, Jara K; Salmina, Elena S; Ringelstetter, Larissa; Dornauer, Scarlett; Kuzikov, Maria; Rothenaigner, Ina; Schorpp, Kenji; Giehler, Fabian; Gopalakrishnan, Jay; Kieser, Arnd; Gul, Sheraz; Tetko, Igor V; Hadian, Kamyar

    2016-07-01

    In high-throughput screening (HTS) campaigns, the binding of glutathione S-transferase (GST) to glutathione (GSH) is used for detection of GST-tagged proteins in protein-protein interactions or enzyme assays. However, many false-positives, so-called frequent hitters (FH), arise that either prevent GST/GSH interaction or interfere with assay signal generation or detection. To identify GST-FH compounds, we analyzed the data of five independent AlphaScreen-based screening campaigns to classify compounds that inhibit the GST/GSH interaction. We identified 53 compounds affecting GST/GSH binding but not influencing His-tag/Ni(2+)-NTA interaction and general AlphaScreen signals. The structures of these 53 experimentally identified GST-FHs were analyzed in chemoinformatic studies to categorize substructural features that promote interference with GST/GSH binding. Here, we confirmed several existing chemoinformatic filters and more importantly extended them as well as added novel filters that specify compounds with anti-GST/GSH activity. Selected compounds were also tested using different antibody-based GST detection technologies and exhibited no interference clearly demonstrating specificity toward their GST/GSH interaction. Thus, these newly described GST-FH will further contribute to the identification of FH compounds containing promiscuous substructures. The developed filters were uploaded to the OCHEM website (http://ochem.eu) and are publicly accessible for analysis of future HTS results. PMID:27044684

  10. Glutathione S-transferase P influences redox and migration pathways in bone marrow.

    Directory of Open Access Journals (Sweden)

    Jie Zhang

    Full Text Available To interrogate why redox homeostasis and glutathione S-transferase P (GSTP are important in regulating bone marrow cell proliferation and migration, we isolated crude bone marrow, lineage negative and bone marrow derived-dendritic cells (BMDDCs from both wild type (WT and knockout (Gstp1/p2(-/- mice. Comparison of the two strains showed distinct thiol expression patterns. WT had higher baseline and reactive oxygen species-induced levels of S-glutathionylated proteins, some of which (sarco-endoplasmic reticulum Ca2(+-ATPase regulate Ca(2+ fluxes and subsequently influence proliferation and migration. Redox status is also a crucial determinant in the regulation of the chemokine system. CXCL12 chemotactic response was stronger in WT cells, with commensurate alterations in plasma membrane polarization/permeability and intracellular calcium fluxes; activities of the downstream kinases, ERK and Akt were also higher in WT. In addition, expression levels of the chemokine receptor CXCR4 and its associated phosphatase, SHP-2, were higher in WT. Inhibition of CXCR4 or SHP2 decreased the extent of CXCL12-induced migration in WT BMDDCs. The differential surface densities of CXCR4, SHP-2 and inositol trisphosphate receptor in WT and Gstp1/p2(-/- cells correlated with the differential CXCR4 functional activities, as measured by the extent of chemokine-induced directional migration and differences in intracellular signaling. These observed differences contribute to our understanding of how genetic ablation of GSTP causes different levels of myeloproliferation and migration [corrected

  11. Gamma-Glutamyl Transferase Levels in Patients with Acute Ischemic Stroke

    Directory of Open Access Journals (Sweden)

    Nurbanu Gurbuzer

    2014-01-01

    Full Text Available Objective. The aim of this study was to investigate the relationship between gamma-glutamyl transferase (GGT levels, cerebrovascular risk factors, and distribution of cerebral infarct areas in patients with acute ischemic stroke (AIS. Patients and Methods. Sixty patients with AIS and 44 controls who had not cerebrovascular disease were included in the study. The patients were divided into four groups according to the location of the infarct area and evaluated as for GGT levels and the presence of diabetes mellitus (DM, hypertension (HT, and hyperlipidemia (HL. Results. The frequency of DM, HT, and HL and gender distributions were similar. The mean GGT levels were significantly higher in patients with AIS and those with relatively larger areas of infarction (P<0.05. Increased mean GGT levels were found in the subgroup with hypertension, higher LDL-cholesterol, and triglyceride levels among cases with AIS (P<0.05. Conclusion. Higher GGT levels in AIS patients reinforce the relationship of GGT with inflammation and oxidative stress. The observation of higher GGT levels in patients with relatively larger areas of infarction is indicative of a positive correlation between increases in infarct areas and elevated GGT levels.

  12. Glutathione S-transferases of Aulacorthum solani and Acyrthosiphon pisum: partial purification and characterization.

    Science.gov (United States)

    Francis, F; Haubruge, E; Gaspar, C; Dierickx, P J

    2001-05-01

    Glutathione S-transferases (GST) play an important role in the detoxification of many substances including allelochemicals from plants. Brassicaceae plants contain glucosinolates and emit volatile isothiocyanates which affect the GST system. A comparison of the GST of two aphid species, the generalist Aulacorthum solani found on Brassicaceae and the Fabaceae specialist Acyrthosiphon pisum, was made to try to explain their respective feeding behaviour. Differences of GST were determined among the two aphid species based on purification by affinity chromatography, SDS-PAGE and on kinetic studies. Purification yields using an epoxy-activated Sepharose 6B column were highly different for the two aphid species (18% and 34% for A. solani and A. pisum, respectively). These variations were confirmed by SDS-PAGE. While only a 27-kDa band was observed for A. pisum, two bands of approximately 25-kDa were visualized for the generalist aphid, A. solani. Considering the kinetic results, differences of Km and Vmax were observed following the aphid species when a range of substrates (CDNB and DCNB) and GSH concentrations were tested. Studies on the detoxification enzymes of generalist and specialist herbivores would be undertaken to determine accurately the effect of the host plant on the organisms eating them, particularly in terms of biochemical and ecological advantages. PMID:11337260

  13. Use of heterologously-expressed cytochrome P450 and glutathione transferase enzymes in toxicity assays.

    Science.gov (United States)

    Guengerich, F Peter; Wheeler, James B; Chun, Young-Jin; Kim, Donghak; Shimada, Tsutomu; Aryal, Pramod; Oda, Yoshimitsu; Gillam, Elizabeth M J

    2002-12-27

    Our groups have had a long-term interest in utilizing bacterial systems in the characterization of bioactivation and detoxication reactions catalyzed by cytochrome P450 (P450) and glutathione transferase (GST) enzymes. Bacterial systems remain the first choice for initial screens with new chemicals and have advantages, including high-throughput capability. Most human P450s of interest in toxicology have been readily expressed in Escherichia coli with only minor sequence modification. These enzymes can be readily purified and used in assays of activation of chemicals. Bicistronic systems have been developed in order to provide the auxiliary NADPH-P450 reductase. Alternative systems involve these enzymes expressed together within bacteria. In one approach, a lac selection system is used with E. coli and has been applied to the characterization of inhibitors of P450s 1A2 and 1B1, as well as in basic studies involving random mutagenesis. Another approach utilizes induction of the SOS (umu) response in Salmonella typhimurium, and systems have now been developed with human P450s 1A1, 1A2, 1B1, 2C9, 2D6, 2E1, and 3A4, which have been used to report responses from heterocyclic amines. S. typhimurium his reporter systems have also been used with GSTs, first to demonstrate the role of rat GST 5-5 in the activation of dihalomethanes. These systems have been used to compare these GSTs with regard to activation of dihaloalkanes and potential toxicity. PMID:12505322

  14. Functional Identification of Proteus mirabilis eptC Gene Encoding a Core Lipopolysaccharide Phosphoethanolamine Transferase

    Directory of Open Access Journals (Sweden)

    Eleonora Aquilini

    2014-04-01

    Full Text Available By comparison of the Proteus mirabilis HI4320 genome with known lipopolysaccharide (LPS phosphoethanolamine transferases, three putative candidates (PMI3040, PMI3576, and PMI3104 were identified. One of them, eptC (PMI3104 was able to modify the LPS of two defined non-polar core LPS mutants of Klebsiella pneumoniae that we use as surrogate substrates. Mass spectrometry and nuclear magnetic resonance showed that eptC directs the incorporation of phosphoethanolamine to the O-6 of l-glycero-d-mano-heptose II. The eptC gene is found in all the P. mirabilis strains analyzed in this study. Putative eptC homologues were found for only two additional genera of the Enterobacteriaceae family, Photobacterium and Providencia. The data obtained in this work supports the role of the eptC (PMI3104 product in the transfer of PEtN to the O-6 of l,d-HepII in P. mirabilis strains.

  15. Cefadroxil potency as cancer co-therapy candidate by glutathione s-transferase mechanism

    Directory of Open Access Journals (Sweden)

    Tri Yuliani

    2013-03-01

    Full Text Available Background: Glutathione S-transferases (GSTs havean important role in the detoxification of electrophiles,such as some anticancer drugs. Compounds with phenolicand/or α,b-unsaturated carbonyl group have been knownas GSTs inhibitor in vitro. Cefadroxil in vitro decreasedGST-Pi activity but not GSTs in rat kidney cytosol.GST inhibitor in a specific organ and of a specific classis needed for safety in cancer chemotherapy. The studyaims to observe the effect of cefadroxil on GSTs in vivoin rat kidney cytosol and then compare it to those seenfor liver, lung, and spleen in vivo.Methods: Cefadroxil was given twice a day byforcefeeding for five days. Rat kidney cytosol was thenprepared and its protein concentration was determined.Cytosolic total GST, GST-Mu and GST-Pi activitieswere monitored by a continuous spectrophotometricmethod using the following substrates: 1-chloro,2,4-dinitrobenzene (CDNB (non-specific substrate,1,2-dichloro-4-nitrobenzene (DCNB for GST-Mu, andethacrynic acid (EA for GST-Pi.Results: The data showed that cefadroxil significantlyincreased the activity of GSTs, GST-Mu, and GSTPiin rat kidney cytosol (8.75%, 47.81%, and 6.67%respectively.Conclusion: Cefadroxil did not inhibit GSTs, GST-Mu,and GST-Pi in rat kidney in vivo indicating that it doesnot inhibit chemotherapy detoxification by GSTs, GSTMu,and GST-Pi in normal kidney cells.

  16. Molecular characterization of zeta class glutathione S-transferases from Pinus brutia Ten.

    Indian Academy of Sciences (India)

    E. Oztetik; F. Kockar; M. Alper; M. Iscan

    2015-09-01

    Glutathione transferases (GSTs; EC 2.5.1.18) play important roles in stress tolerance and metabolic detoxification in plants. In higher plants, studies on GSTs have focussed largely on agricultural plants. There is restricted information about molecular characterization of GSTs in gymnosperms. To date, only tau class GST enzymes have been characterized from some pinus species. For the first time, the present study reports cloning and molecular characterization of two zeta class GST genes, namely PbGSTZ1 and PbGSTZ2 from Pinus brutia Ten., which is an economically important pine native to the eastern Mediterranean region and have to cope with several environmental stress conditions. The PbGSTZ1 gene was isolated from cDNA, whereas PbGSTZ2 was isolated from genomic DNA. Sequence analysis of PbGSTZ1 and PbGSTZ2 revealed the presence of an open reading frame of 226 amino acids with typical consensus sequences of the zeta class plant GSTs. Protein and secondary structure prediction analysis of two zeta class PbGSTZs have shared common features of other plant zeta class GSTs. Genomic clone, PbGSTZ2 gene, is unexpectedly intronless. Extensive sequence analysis of PbGSTZ2, with cDNA clone, PbGSTZ1, revealed 87% identity at nucleotide and 81% identity at amino acid levels with 41 amino acids differences suggesting that genomic PbGSTZ2 gene might be an allelic or a paralogue version of PbGSTZ1.

  17. Methionine sulfoxide reductase regulates brain catechol-O-methyl transferase activity.

    Science.gov (United States)

    Moskovitz, Jackob; Walss-Bass, Consuelo; Cruz, Dianne A; Thompson, Peter M; Bortolato, Marco

    2014-10-01

    Catechol-O-methyl transferase (COMT) plays a key role in the degradation of brain dopamine (DA). Specifically, low COMT activity results in higher DA levels in the prefrontal cortex (PFC), thereby reducing the vulnerability for attentional and cognitive deficits in both psychotic and healthy individuals. COMT activity is markedly reduced by a non-synonymous single-nucleotide polymorphism (SNP) that generates a valine-to-methionine substitution on the residue 108/158, by means of as-yet incompletely understood post-translational mechanisms. One post-translational modification is methionine sulfoxide, which can be reduced by the methionine sulfoxide reductase (Msr) A and B enzymes. We used recombinant COMT proteins (Val/Met108) and mice (wild-type (WT) and MsrA knockout) to determine the effect of methionine oxidation on COMT activity and COMT interaction with Msr, through a combination of enzymatic activity and Western blot assays. Recombinant COMT activity is positively regulated by MsrA, especially under oxidative conditions, whereas brains of MsrA knockout mice exhibited lower COMT activity (as compared with their WT counterparts). These results suggest that COMT activity may be reduced by methionine oxidation, and point to Msr as a key molecular determinant for the modulation of COMT activity in the brain. The role of Msr in modulating cognitive functions in healthy individuals and schizophrenia patients is yet to be determined. PMID:24735585

  18. Trichinella spiralis: low vaccine potential of glutathione S-transferase against infections in mice.

    Science.gov (United States)

    Li, Ling Ge; Wang, Zhong Quan; Liu, Ruo Dan; Yang, Xuan; Liu, Li Na; Sun, Ge Ge; Jiang, Peng; Zhang, Xi; Zhang, Gong Yuan; Cui, Jing

    2015-06-01

    We have previously reported that Trichinella spiralis glutathione-S-transferase (TsGST) gene is an up-regulated gene in intestinal infective larvae (IIL) compared to muscle larvae (ML). In this study, the TsGST gene was cloned, and recombinant TsGST (rTsGST) was produced. Anti-rTsGST serum recognized the native TsGST by Western blotting in crude antigens of ML, adult worm (AW) and newborn larvae (NBL) of T. spiralis, but not in ML excretory-secretory (ES) antigens. Expression of TsGST was observed in all different developmental stages (IIL, AW, NBL and ML). An immunolocalization analysis identified TsGST in the cuticle, stichosome and genital primordium of the parasite. The rTsGST had GST enzymatic activity. After a challenge infection with T. spiralis larvae, mice immunized with rTsGST displayed a 35.71% reduction in adult worms and a 38.55% reduction in muscle larvae. The vaccination of mice with rTsGST induced the Th1/Th2-mixed type of immune response with Th2 predominant (high levels of IgG1) and partial protective immunity against T. spiralis infection. PMID:25757368

  19. Glutathione S-transferase in aquatic macro-invertebrates and its interaction with different organic micropollutants

    Energy Technology Data Exchange (ETDEWEB)

    Dierickx, P.J.

    1984-12-01

    In higher organisms, glutathione S-transferase (GST) plays a key role in the detoxification of a large number of xenobiotics. In the present work the presence of GST in aquatic macro-invertebrates and its possible significance as a detoxification mechanism of organic micropollutants in the aquatic environment is investigated. So far, GST has been found in 20 macro-invertebrates (in adults as well as in larvae) and in insects as well as in other animal groups. The GST activities were relatively high, ranging from 10 to 600% of the activity found in rat liver. The interaction of quinones, o-chloranil and chlorophenoxyalkyl acids with the GST activity, in extracts from three different macro-invertebrates, revealed an inhibition which was quite similar to that previously found for rat liver GST. In Tubifex tubifex extracts at least three different GST isoenzymes could be demonstrated. These partially purified isoenzymes were used for the kinetic analysis of GST inhibition by 2,4-dichlorophenoxyalkyl acid and 1,4-benzoquinone, using Lineweaver--Burk plots. The same kinetic patterns were observed as for rat liver GST. The results demonstrate that the interactions of the compounds investigated with aquatic macro-invertebrate and with rat liver GST are in very good agreement. It is concluded that macro-invertebrate GST can play a key role in the detoxification of organic micropollutants in the aquatic environment.

  20. Glutathione transferase A4-4 resists adduction by 4-hydroxynonenal.

    Science.gov (United States)

    Shireman, Laura M; Kripps, Kimberly A; Balogh, Larissa M; Conner, Kip P; Whittington, Dale; Atkins, William M

    2010-12-15

    4-Hydroxy-2-trans-nonenal (HNE) is a lipid peroxidation product that contributes to the pathophysiology of several diseases with components of oxidative stress. The electrophilic nature of HNE results in covalent adduct formation with proteins, fatty acids and DNA. However, it remains unclear whether enzymes that metabolize HNE avoid inactivation by it. Glutathione transferase A4-4 (GST A4-4) plays a significant role in the elimination of HNE by conjugating it with glutathione (GSH), with catalytic activity toward HNE that is dramatically higher than the homologous GST A1-1 or distantly related GSTs. To determine whether enzymes that metabolize HNE resist its covalent adduction, the rates of adduction of these GST isoforms were compared and the functional effects of adduction on catalytic properties were determined. Although GST A4-4 and GST A1-1 have striking structural similarity, GST A4-4 was insensitive to adduction by HNE under conditions that yield modest adduction of GST A1-1 and extensive adduction of GST P1-1. Furthermore, adduction of GST P1-1 by HNE eliminated its activity toward the substrates 1-chloro-2,4-dinitrobenzene (CDNB) and toward HNE itself. HNE effects on GST A4-4 and A1-1 were less significant. The results indicate that enzymes that metabolize HNE may have evolved structurally to resist covalent adduction by it. PMID:20836986

  1. Glutathione transferase A4-4 resists adduction by 4-hydroxynonenal☆

    Science.gov (United States)

    Shireman, Laura M.; Kripps, Kimberly A.; Balogh, Larissa M.; Conner, Kip P.; Whittington, Dale; Atkins, William M.

    2010-01-01

    4-Hydroxy-2-trans-nonenal (HNE) is a lipid peroxidation product that contributes to the pathophysiology of several diseases with components of oxidative stress. The electrophilic nature of HNE results in covalent adduct formation with proteins, fatty acids and DNA. However, it remains unclear whether enzymes that metabolize HNE avoid inactivation by it. Glutathione transferase A4-4 (GST A4-4) plays a significant role in the elimination of HNE by conjugating it with glutathione (GSH), with catalytic activity toward HNE that is dramatically higher than the homologous GST A1-1 or distantly related GSTs. To determine whether enzymes that metabolize HNE resist its covalent adduction, the rates of adduction of these GST isoforms were compared and the functional effects of adduction on catalytic properties were determined. Although GST A4-4 and GST A1-1 have striking structural similarity, GST A4-4 was insensitive to adduction by HNE under conditions that yield modest adduction of GST A1-1 and extensive adduction of GST P1-1. Furthermore, adduction of GST P1-1 by HNE eliminated its activity toward the substrates 1-chloro- 2,4-dinitrobenzene (CDNB) and toward HNE itself. HNE effects on GST A4-4 and A1-1 were less significant. The results indicate that enzymes that metabolize HNE may have evolved structurally to resist covalent adduction by it. PMID:20836986

  2. Inhibition of insect glutathione S-transferase (GST) by conifer extracts.

    Science.gov (United States)

    Wang, Zhiling; Zhao, Zhong; Abou-Zaid, Mamdouh M; Arnason, John T; Liu, Rui; Walshe-Roussel, Brendan; Waye, Andrew; Liu, Suqi; Saleem, Ammar; Cáceres, Luis A; Wei, Qin; Scott, Ian M

    2014-12-01

    Insecticide synergists biochemically inhibit insect metabolic enzyme activity and are used both to increase the effectiveness of insecticides and as a diagnostic tool for resistance mechanisms. Considerable attention has been focused on identifying new synergists from phytochemicals with recognized biological activities, specifically enzyme inhibition. Jack pine (Pinus banksiana Lamb.), black spruce (Picea mariana (Mill.) BSP.), balsam fir (Abies balsamea (L.) Mill.), and tamarack larch (Larix laricina (Du Roi) Koch) have been used by native Canadians as traditional medicine, specifically for the anti-inflammatory and antioxidant properties based on enzyme inhibitory activity. To identify the potential allelochemicals with synergistic activity, ethanol crude extracts and methanol/water fractions were separated by Sephadex LH-20 chromatographic column and tested for in vitro glutathione S-transferase (GST) inhibition activity using insecticide-resistant Colorado potato beetle, Leptinotarsa decemlineata (Say) midgut and fat-body homogenate. The fractions showing similar activity were combined and analyzed by ultra pressure liquid chromatography-mass spectrometry. A lignan, (+)-lariciresinol 9'-p-coumarate, was identified from P. mariana cone extracts, and L. laricina and A. balsamea bark extracts. A flavonoid, taxifolin, was identified from P. mariana and P. banksiana cone extracts and L. laricina bark extracts. Both compounds inhibit GST activity with taxifolin showing greater activity compared to (+)-lariciresinol 9'-p-coumarate and the standard GST inhibitor, diethyl maleate. The results suggested that these compounds can be considered as potential new insecticide synergists. PMID:25270601

  3. Rat liver glutathione S-transferase activity stimulation following acute cadmium or manganese intoxication

    International Nuclear Information System (INIS)

    The effect of cadmium or manganese administration on rat liver glutathione S-transferase (GST) has been investigated. The activity of this enzyme in liver cytosol, where almost all the cellular activity is present, had increased by more than 36% 24 h after a single i.p. injection of CdCl2 (2.5 mg kg-1 b.w.) or MnCl2 (2.0 mg kg-1 b.w.). After shorter and longer time intervals, a lower enzyme activity stimulation was observed in both cases. When liver cytosol was incubated for 10 min with 75 μM CdCl2 or 40 μM MnCl2, no effect was observed on enzyme activity. The increase in GST following cadmium or manganese administration was blocked by prior administration of actinomycin D, indicative of a possible transcription-dependent response. The liver soluble GST from both control and metal-treated rats was not at all affected by Vitamin E, in the range of 20-300 μM. By contrast, hematin was seen to be a competitive inhibitor of this liver enzyme from both types of rats by using CDNB as substrate and the Ki value was equal to 0.22 μM. The possibility that under the conditions used class alpha GST isoenzymes are affected by cadmium or manganese is discussed

  4. A phosphopantetheinyl transferase that is essential for mitochondrial fatty acid biosynthesis.

    Science.gov (United States)

    Guan, Xin; Chen, Hui; Abramson, Alex; Man, Huimin; Wu, Jinxia; Yu, Oliver; Nikolau, Basil J

    2015-11-01

    In this study we report the molecular genetic characterization of the Arabidopsis mitochondrial phosphopantetheinyl transferase (mtPPT), which catalyzes the phosphopantetheinylation and thus activation of mitochondrial acyl carrier protein (mtACP) of mitochondrial fatty acid synthase (mtFAS). This catalytic capability of the purified mtPPT protein (encoded by AT3G11470) was directly demonstrated in an in vitro assay that phosphopantetheinylated mature Arabidopsis apo-mtACP isoforms. The mitochondrial localization of the AT3G11470-encoded proteins was validated by the ability of their N-terminal 80-residue leader sequence to guide a chimeric GFP protein to this organelle. A T-DNA-tagged null mutant mtppt-1 allele shows an embryo-lethal phenotype, illustrating a crucial role of mtPPT for embryogenesis. Arabidopsis RNAi transgenic lines with reduced mtPPT expression display typical phenotypes associated with a deficiency in the mtFAS system, namely miniaturized plant morphology, slow growth, reduced lipoylation of mitochondrial proteins, and the hyperaccumulation of photorespiratory intermediates, glycine and glycolate. These morphological and metabolic alterations are reversed when these plants are grown in a non-photorespiratory condition (i.e. 1% CO2 atmosphere), demonstrating that they are a consequence of a deficiency in photorespiration due to the reduced lipoylation of the photorespiratory glycine decarboxylase. PMID:26402847

  5. A glutathione S-transferase gene associated with antioxidant properties isolated from Apis cerana cerana.

    Science.gov (United States)

    Liu, Shuchang; Liu, Feng; Jia, Haihong; Yan, Yan; Wang, Hongfang; Guo, Xingqi; Xu, Baohua

    2016-06-01

    Glutathione S-transferases (GSTs) are an important family of multifunctional enzymes in aerobic organisms. They play a crucial role in the detoxification of exogenous compounds, especially insecticides, and protection against oxidative stress. Most previous studies of GSTs in insects have largely focused on their role in insecticide resistance. Here, we isolated a theta class GST gene designated AccGSTT1 from Apis cerana cerana and aimed to explore its antioxidant and antibacterial attributes. Analyses of homology and phylogenetic relationships suggested that the predicted amino acid sequence of AccGSTT1 shares a high level of identity with the other hymenopteran GSTs and that it was conserved during evolution. Quantitative real-time PCR showed that AccGSTT1 is most highly expressed in adult stages and that the expression profile of this gene is significantly altered in response to various abiotic stresses. These results were confirmed using western blot analysis. Additionally, a disc diffusion assay showed that a recombinant AccGSTT1 protein may be roughly capable of inhibiting bacterial growth and that it reduces the resistance of Escherichia coli cells to multiple adverse stresses. Taken together, these data indicate that AccGSTT1 may play an important role in antioxidant processes under adverse stress conditions. PMID:27126403

  6. Glutathione-S-transferase in Nereis succinea (Polychaeta) and its induction by xeno-estrogen.

    Science.gov (United States)

    Ayoola, James A O; García-Alonso, Javier; Hardege, Jörg D

    2011-10-01

    The need to replace or at least to reduce the use of vertebrates in toxicity tests is a timely major concern in research and industry but to date, efforts made to minimize their use are still far from complete. Increasing demands for toxicity tests put considerable pressures upon the development of future fast and efficient test methods using invertebrates. In fact, to date, few studies provide links between biochemical and cellular effects of xeno-estrogens in aquatic invertebrates. Glutathione-S-transferase (GST) activity, as a biomarker of stress exposure, was measured in the population of clamworms (Nereis succinea) from Cardiff Bay. In addition, we examined the effect of single exposure to nonylphenol (NP) on this enzymatic activity. Field study results showed a relationship between the worm's size, reproductive status, and GST activity from the field population. In addition, we show a significant increase in the GST activity at 100 μg/L NP with sex-specific responses. The xeno-estrogens, which could affect reproduction of nereid by interfering in normal endocrinological pathways, are eliminated through GST by conjugation with glutathione. This work shows for the first time that GST activity depends on sex and stage of the clamworms and also that the xeno-estrogen NP induces its activity. This study supports the use of this species as a bioindicator of aquatic pollution and lays the foundation to causally link toxic exposure with reproductive output. PMID:20549611

  7. Structure, function and disease relevance of Omega-class glutathione transferases.

    Science.gov (United States)

    Board, Philip G; Menon, Deepthi

    2016-05-01

    The Omega-class cytosolic glutathione transferases (GSTs) have distinct structural and functional attributes that allow them to perform novel roles unrelated to the functions of other GSTs. Mammalian GSTO1-1 has been found to play a previously unappreciated role in the glutathionylation cycle that is emerging as significant mechanism regulating protein function. GSTO1-1-catalyzed glutathionylation or deglutathionylation of a key signaling protein may explain the requirement for catalytically active GSTO1-1 in LPS-stimulated pro-inflammatory signaling through the TLR4 receptor. The observation that ML175 a specific GSTO1-1 inhibitor can block LPS-stimulated inflammatory signaling has opened a new avenue for the development of novel anti-inflammatory drugs that could be useful in the treatment of toxic shock and other inflammatory disorders. The role of GSTO2-2 remains unclear. As a dehydroascorbate reductase, it could contribute to the maintenance of cellular redox balance and it is interesting to note that the GSTO2 N142D polymorphism has been associated with multiple diseases including Alzheimer's disease, Parkinson's disease, familial amyotrophic lateral sclerosis, chronic obstructive pulmonary disease, age-related cataract and breast cancer. PMID:26993125

  8. Glutathione S-Transferase of Brown Planthoppers (Nilaparvata lugens) Is Essential for Their Adaptation to Gramine-Containing Host Plants

    OpenAIRE

    Sun, Xiao-Qin; Zhang, Mao-Xin; Yu, Jing-Ya; Jin, Yu; Ling, Bing; Du, Jin-Ping; Li, Gui-Hua; Qin, Qing-Ming; Cai, Qing-Nian

    2013-01-01

    Plants have evolved complex processes to ward off attacks by insects. In parallel, insects have evolved mechanisms to thwart these plant defenses. To gain insight into mechanisms that mediate this arms race between plants and herbivorous insects, we investigated the interactions between gramine, a toxin synthesized by plants of the family Gramineae, and glutathione S transferase (GST), an enzyme found in insects that is known to detoxify xenobiotics. Here, we demonstrate that rice (Oryza sati...

  9. Mimicking Insect Communication: Release and Detection of Pheromone, Biosynthesized by an Alcohol Acetyl Transferase Immobilized in a Microreactor

    OpenAIRE

    Lourdes Muñoz; Nikolay Dimov; Gerard Carot-Sans; Bula, Wojciech P.; Angel Guerrero; Gardeniers, Han J. G. E.

    2012-01-01

    Infochemical production, release and detection of (Z,E)-9,11-tetradecadienyl acetate, the major component of the pheromone of the moth Spodoptera littoralis is achieved in a novel microfluidic system, designed to mimic the final step of the pheromone biosynthesis by immobilized recombinant alcohol acetyl transferase. The microfluidic system is part of an "artificial gland", i.e. a chemoemitter that comprises a microreactor connected to a microevaporator and is able to produce a...

  10. Determination of glutathione-S-transferase traces in preparations of p53 C-terminal domain (aa320-393)

    Czech Academy of Sciences Publication Activity Database

    Brázdová, Marie; Kizek, René; Havran, Luděk; Paleček, Emil

    2002-01-01

    Roč. 55, 1/2 (2002), s. 115-118. ISSN 1567-5394 R&D Projects: GA AV ČR IAA4004110; GA ČR GV204/97/K084; GA ČR GA204/00/D049; GA MZd NC5343 Institutional research plan: CEZ:AV0Z5004920 Keywords : p53 * glutathione-S-transferase determination * constant current chronopotentiometry Subject RIV: BO - Biophysics Impact factor: 1.463, year: 2002

  11. Identification and Characterization of Seven Glutathione S-Transferase Genes from Citrus Red Mite, Panonychus citri (McGregor)

    OpenAIRE

    Liao, Chong-Yu; Zhang, Kun; Niu, Jin-Zhi; Ding, Tian-Bo; Zhong, Rui; Xia, Wen-Kai; Dou, Wei; Wang, Jin-Jun

    2013-01-01

    The citrus red mite, Panonychus citri (McGregor), is a global citrus pest, and has developed severe resistance to several types of acaricides. However, the molecular mechanisms of resistance in this mite remain unknown. In this study, seven full-length cDNAs encoding glutathione S-transferases (GSTs) genes were identified and characterized in P. citri. The effects of pyridaben and fenpropathrin exposure on the expression of these genes were also investigated. Phylogenetic analysis revealed th...

  12. ANALISIS GEN PENYANDI Schistosoma japonicum Gluthation s Transferase (SJ26GST) DI DATARAN TINGGI LINDU, SULAWESI TENGAH INDONESIA

    OpenAIRE

    Anis Nurwidayati; Triwibowo A. Garjito; Phetisya Pamela Frederika Sumolang; Risti Risti

    2015-01-01

    AbstractSchistosomiasis is only found at Napu and Lindu highland, Central Sulawesi in Indonesia. Schistosomiasis still as a public health problem, with its prevalence increase every year. The large scale by mass drug treatment using praziquantel has done to reduce the prevalence since 1980. To look for the possibility evidence of the development of resistance in S. japonicumto praziquantel in endemic areas by analysis of Schistosoma japonicumGluthation S Transferase (Sj26gst) Coding Gene. Mol...

  13. 23S rRNA nucleotides in the peptidyl transferase center are essential for tryptophanase operon induction.

    Science.gov (United States)

    Yang, Rui; Cruz-Vera, Luis R; Yanofsky, Charles

    2009-06-01

    Distinct features of the ribosomal peptide exit tunnel are known to be essential for recognition of specific amino acids of a nascent peptidyl-tRNA. Thus, a tryptophan residue at position 12 of the peptidyl-tRNA TnaC-tRNA(Pro) leads to the creation of a free tryptophan binding site within the ribosome at which bound tryptophan inhibits normal ribosome functions. The ribosomal processes that are inhibited are hydrolysis of TnaC-tRNA(Pro) by release factor 2 and peptidyl transfer of TnaC of TnaC-tRNA(Pro) to puromycin. These events are normally performed in the ribosomal peptidyl transferase center. In the present study, changes of 23S rRNA nucleotides in the 2585 region of the peptidyl transferase center, G2583A and U2584C, were observed to reduce maximum induction of tna operon expression by tryptophan in vivo without affecting the concentration of tryptophan necessary to obtain 50% induction. The growth rate of strains with ribosomes with either of these changes was not altered appreciably. In vitro analyses with mutant ribosomes with these changes showed that tryptophan was not as efficient in protecting TnaC-tRNA(Pro) from puromycin action as wild-type ribosomes. However, added tryptophan did prevent sparsomycin action as it normally does with wild-type ribosomes. These findings suggest that these two mutational changes act by reducing the ability of ribosome-bound tryptophan to inhibit peptidyl transferase activity rather than by reducing the ability of the ribosome to bind tryptophan. Thus, the present study identifies specific nucleotides within the ribosomal peptidyl transferase center that appear to be essential for effective tryptophan induction of tna operon expression. PMID:19329641

  14. Selective Reversible Inhibition of Liver Carnitine Palmitoyl-Transferase 1 by Teglicar Reduces Gluconeogenesis and Improves Glucose Homeostasis

    OpenAIRE

    Conti, Roberto; Mannucci, Edoardo; Pessotto, Pompeo; Tassoni, Emanuela; Carminati, Paolo; Giannessi, Fabio; Arduini, Arduino

    2011-01-01

    OBJECTIVE We have developed a new antihyperglycemic agent (teglicar) through the selective and reversible inhibition of the liver isoform of carnitine palmitoyl-transferase 1 (L-CPT1). RESEARCH DESIGN AND METHODS Glucose production was investigated in isolated hepatocytes and during pancreatic clamps in healthy rats. Chronic treatments on C57BL/6J, db/db, high-fat fed mice, and rats were performed to understand glucose metabolism and insulin sensitivity. RESULTS In isolated hepatocytes, tegli...

  15. Molecular characterization of two galactosemia mutations: correlation of mutations with highly conserved domains in galactose-1-phosphate uridyl transferase.

    OpenAIRE

    Reichardt, J K; Packman, S; Woo, S L

    1991-01-01

    Galactosemia is an autosomal recessive disorder of human galactose metabolism caused by deficiency of the enzyme galactose-1-phosphate uridyl transferase (GALT). The molecular basis of this disorder is at present not well understood. We report here two missense mutations which result in low or undetectable enzymatic activity. First, we identified at nucleotide 591 a transition which substitutes glutamine 188 by arginine. The mutated glutamine is not only highly conserved in evolution (conserv...

  16. Increased Sensitivity of Glutathione S-Transferase P-Null Mice to Cyclophosphamide-Induced Urinary Bladder Toxicity

    OpenAIRE

    Conklin, Daniel J.; Haberzettl, Petra; Lesgards, Jean-Francois; Prough, Russell A.; Srivastava, Sanjay; Bhatnagar, Aruni

    2009-01-01

    Hemorrhagic cystitis and diffuse inflammation of the bladder, common side effects of cyclophosphamide (CY) treatment, have been linked to the generation of acrolein derived from CY metabolism. Metabolic removal of acrolein involves multiple pathways, which include reduction, oxidation, and conjugation with glutathione. Herein, we tested the hypothesis that glutathione S-transferase P (GSTP), the GST isoform that displays high catalytic efficiency with acrolein, protects against CY-induced uro...

  17. Distribution of glutathione S-transferase isoenzymes in human kidney: basis for possible markers of renal injury.

    OpenAIRE

    Harrison, D J; Kharbanda, R; Cunningham, D S; McLellan, L I; Hayes, J. D.

    1989-01-01

    To determine whether the tissue distribution of glutathione S-transferase (GST) isoenzymes could define the precise nature of renal injury, 13 adult kidneys were studied, using specific antibodies raised against purified isoenzymes. Basic GST stained strongly proximal convoluted tubules and some medullary tubules; acidic GST stained strongly distal convoluted tubules and medullary tubules; neutral GST stained similarly to acidic GST, but weaker, and microsomal GST stained glomerular and inter...

  18. Effect of cadmium on glutathione S-transferase and metallothionein gene expression in coho salmon liver, gill and olfactory tissues

    OpenAIRE

    Espinoza, Herbert M.; Williams, Chase R.; Gallagher, Evan P.

    2011-01-01

    The glutathione S-transferases (GSTs) are a multifunctional family of phase II enzymes that detoxify a variety of environmental chemicals, reactive intermediates, and secondary products of oxidative damage. GST mRNA expression and catalytic activity have been used as biomarkers of exposure to environmental chemicals. However, factors such as species differences in induction, partial analyses of multiple GST isoforms, and lack of understanding of fish GST gene regulation, have confounded the u...

  19. Proteomic Profiling of Cytosolic Glutathione Transferases from Three Bivalve Species: Corbicula fluminea, Mytilus galloprovincialis and Anodonta cygnea

    OpenAIRE

    José Carlos Martins; Alexandre Campos; Hugo Osório; Rute da Fonseca; Vítor Vasconcelos

    2014-01-01

    Suspension-feeding bivalves are considered efficient toxin vectors with a relative insensitivity to toxicants compared to other aquatic organisms. This fact highlights the potential role of detoxification enzymes, such as glutathione transferases (GSTs), in this bivalve resistance. Nevertheless, the GST system has not been extensively described in these organisms. In the present study, cytosolic GSTs isoforms (cGST) were surveyed in three bivalves with different habitats and life strategies: ...

  20. Glutathione S-transferase (GST) gene polymorphisms, cigarette smoking and colorectal cancer risk among Chinese in Singapore

    OpenAIRE

    Koh, Woon-Puay; Nelson, Heather H.; Yuan, Jian-Min; Van den Berg, David; Jin, Aizhen; Wang, Renwei; Yu, Mimi C.

    2011-01-01

    Cigarette smoking is a risk factor for colorectal cancer. Putative colorectal procarcinogens in tobacco smoke include polycyclic aromatic hydrocarbons and heterocyclic aromatic amines that are known substrates of glutathione S-transferases (GSTs). This study examined the influence of functional GST gene polymorphisms on the smoking–colorectal cancer association in a population known to be minimally exposed to dietary sources of these procarcinogens. Incident cases of colorectal cancer (n = 48...

  1. Distinct and cooperative activities of HESO1 and URT1 nucleotidyl transferases in microRNA turnover in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Bin Tu

    2015-04-01

    Full Text Available 3' uridylation is increasingly recognized as a conserved RNA modification process associated with RNA turnover in eukaryotes. 2'-O-methylation on the 3' terminal ribose protects micro(miRNAs from 3' truncation and 3' uridylation in Arabidopsis. Previously, we identified HESO1 as the nucleotidyl transferase that uridylates most unmethylated miRNAs in vivo, but substantial 3' tailing of miRNAs still remains in heso1 loss-of-function mutants. In this study, we found that among nine other potential nucleotidyl transferases, UTP:RNA uridylyltransferase 1 (URT1 is the single most predominant nucleotidyl transferase that tails miRNAs. URT1 and HESO1 prefer substrates with different 3' end nucleotides in vitro and act cooperatively to tail different forms of the same miRNAs in vivo. Moreover, both HESO1 and URT1 exhibit nucleotidyl transferase activity on AGO1-bound miRNAs. Although these enzymes are able to add long tails to AGO1-bound miRNAs, the tailed miRNAs remain associated with AGO1. Moreover, tailing of AGO1-bound miRNA165/6 drastically reduces the slicing activity of AGO1-miR165/6, suggesting that tailing reduces miRNA activity. However, monouridylation of miR171a by URT1 endows the miRNA the ability to trigger the biogenesis of secondary siRNAs. Therefore, 3' tailing could affect the activities of miRNAs in addition to leading to miRNA degradation.

  2. Association of Glutathione S-Transferase P1 (GSTP1) Polymorphism with Tourette Syndrome in Taiwanese Patients

    OpenAIRE

    Shen, Che-Piao; Chou, I-Ching; Liu, Hsin-Ping; Lee, Cheng-Chun; Tsai, Yuhsin; Wu, Bor-Tsang; Hsu, Ban-Dar; Lin, Wei-Yong; Tsai, Fuu-Jen

    2014-01-01

    The etiology of Tourette syndrome (TS) is multifactorial. TS vulnerability may be associated with genetic and environmental factors. From the genetic point of view, TS is heterogeneous. Previous studies showed that some single-nucleotide polymorphisms (SNPs) of the glutathione-S-transferase P1 (GSTP1) gene can affect cellular proliferation and apoptotic activity and TS is a neurodevelopmental disorder. We guessed that there was a relationship between TS and genetic variants of the GSTP1 gene....

  3. Substrate Specificity Combined with Stereopromiscuity in Glutathione Transferase A4-4-dependent Metabolism of 4-Hydroxynonenal

    OpenAIRE

    Balogh, Larissa M.; Le Trong, Isolde; Kripps, Kimberly A.; Shireman, Laura M.; Stenkamp, Ronald E.; Zhang, Wei; Mannervik, Bengt; Atkins, William M.

    2010-01-01

    Conjugation to glutathione (GSH) by glutathione transferase A4-4 (GSTA4-4) is a major route of elimination for the lipid peroxidation product 4-hydroxynonenal (HNE), a toxic compound that contributes to numerous diseases. Both enantiomers of HNE are presumed to be toxic, and GSTA4-4 has negligible stereoselectivity towards them, despite its high catalytic chemospecificity for alkenals. In contrast to the highly flexible, and substrate promiscuous, GSTA1-1 isoform that has poor catalytic effic...

  4. Expression Patterns of Glutathione Transferase Gene (GstI) in Maize Seedlings Under Juglone-Induced Oxidative Stress

    OpenAIRE

    Hubert Sytykiewicz

    2011-01-01

    Juglone (5-hydroxy-1,4-naphthoquinone) has been identified in organs of many plant species within Juglandaceae family. This secondary metabolite is considered as a highly bioactive substance that functions as direct oxidant stimulating the production of reactive oxygen species (ROS) in acceptor plants. Glutathione transferases (GSTs, E.C.2.5.1.18) represent an important group of cytoprotective enzymes participating in detoxification of xenobiotics and limiting oxidative damages of cellular ma...

  5. Glutathione S transferase (GSTP 1, GSTM 1, and GSTT 1) gene polymorphisms in Egyptian patients with acute myeloid leukemia

    OpenAIRE

    Nasr, Aml S.; Rania M Sami; Noha Y Ibrahim; Dalia O Darwish

    2015-01-01

    BACKGROUND: The super family of glutathione S-transferases (GSTs) is composed of multiple isoenzymes with significant evidence of functional polymorphic variation. GSTs detoxify potentially mutagenic and toxic DNA-reactive electrophiles, including metabolites of several chemotherapeutic agents, some of which are suspected human carcinogens. Polymorphisms within the phase II metabolizer enzymes GST T1, GST M1, and GST P1 affect the body's ability to detoxify a range of potential leukemogens en...

  6. Molecular characterization of glutathione S-transferase, endothelial nitric oxide synthase and Vitamin D receptor genes in breast cancer cases

    OpenAIRE

    Rizk El-Baz(1); Azza Ismail(2) ; Maher Amer(2); Mai Elshahat(3); Amira Kazamel(2); Ahmad Settin

    2012-01-01

    Background: Enzymes of the Glutathione S-transferase system (GST) modulate the effects of exposure to several cytotoxic and genotoxic agents. Nitric oxide (NO) is constitutively synthesized in the endothelium by endothelial nitric oxide synthase (eNOS) and acts as a pleiotropic regulator involved in carcinogenesis. Vitamin D levels may influence breast cancer development. The vitamin D receptor (VDR) is a crucial mediator for the cellular effects of vitamin D and additionally interacts with o...

  7. Urine α-Glutathione S-Transferase, systemic inflammation and arterial function in juvenile type 1 diabetes.

    OpenAIRE

    Holmquist, Peter; Liuba, Petru

    2012-01-01

    BACKGROUND: Despite marked improvement in therapy and monitoring of patients with insulin-dependent (type 1) diabetes, diabetic nephropathy remains a serious complication, with subsequent end-stage renal disease in about 20% of cases. OBJECTIVE: To investigate in young patients with type 1 diabetes whether urine α-Glutathione S-transferase to creatinine ratio (α-GST:crea) relates to markers of systemic inflammation and subclinical vasculopathy. DESIGN: Children and adolescents ...

  8. Dual Localization of Glutathione S-Transferase in the Cytosol and Mitochondria: Implications in Oxidative Stress, Toxicity and Disease

    OpenAIRE

    Raza, Haider

    2011-01-01

    Glutathione (GSH) conjugating enzymes, glutathione S-transferases (GSTs) are present in different subcellular compartments including cytosol, mitochondria, endoplasmic reticulum, nucleus and plasma membrane. The regulation and function of GSTs have implications in cell growth, oxidative stress, as well as in disease progression and prevention. Of the several mitochondria localized forms, GSTK (GST kappa) is mitochondria-specific since it contains N-terminal canonical and cleavable mitochondri...

  9. Dual protective role for Glutathione S-transferase class pi against VCD-induced ovotoxicity in the rat ovary1

    OpenAIRE

    Keating, Aileen F.; Sen, Nivedita; Sipes, I. Glenn; Hoyer, Patricia B.

    2010-01-01

    The occupational chemical 4-vinylcyclohexene diepoxide (VCD) selectively destroys ovarian small pre-antral follicles in rats and mice via apoptosis. Detoxification of VCD can occur through glutathione conjugation, catalyzed by glutathione S-transferase (GST) enzymes. Further, GST class pi (GSTp) can negatively regulate JNK activity through protein:protein interactions in extra-ovarian tissues. Dissociation of this protein complex in the face of chemical exposure releases the inhibition of pro...

  10. QUANTITATIVE IMAGE CYTOMETRY OF HEPATOCYTES EXPRESSING GAMMA-GLUTAMYL TRANSPEPTIDASE AND GLUTATHIONE S-TRANSFERASE IN DIETHYLNITROSAMINE-INITIATED RATS TREATED WITH PHENOBARBITAL AND/OR PHTHALATE ESTERS

    Science.gov (United States)

    Image cytometry was used to quantify the volume of liver tissue expressing two widely accepted biochemical markers of neoplasia, gammaglutamyl transpeptidase (GGT) and the placental isozyme of glutathione s-transferase (GST-P). ats were treated with hepatocarcinogen, diethylnitro...

  11. Neuroantibodies (NAB) in African-American Children: Associations with Gender, Glutathione-S-Transferase (GST)Pi Polymorphisms (SNP) and Heavy Metals

    Science.gov (United States)

    CONTACT (NAME ONLY): Hassan El-Fawal Abstract Details PRESENTATION TYPE: Platform or Poster CURRENT CATEGORY: Neurodegenerative Disease | Biomarkers | Neurotoxicity, Metals KEYWORDS: Autoantibodies, Glutathione-S-Transferase, DATE/TIME LAST MODIFIED: DATE/TIME SUBMITTED: Abs...

  12. Effect of trans-acting factor on rat glutathione S-transferase P1 gene transcription regulation in tumor cells

    Institute of Scientific and Technical Information of China (English)

    刘东远; 廖名湘; 左瑾; 方福德

    2002-01-01

    Objective To investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells. Methods The binding of trans-acting factor(s) to two enhancers of the rGSTP1 gene, glutathione S-transferase P enhancer Ⅰ (GPEI) and glutathione S-transferase P enhancer Ⅱ-1 (GPEⅡ-1), was identified by an electrophoretic mobility shift assay (EMSA). The molecular weight of trans-acting factor was measured in a UV cross-linking experiment. Results Trans-acting factor interacting with the core sequence of GPEI (cGPEI) were found in human cervical adenocarcinoma cell line (HeLa) and rat hepatoma cell line (CBRH7919). These proteins were not expressed in normal rat liver. Although specific binding proteins that bound to GPEⅡ-1 were detected in all three cell types, a 64 kDa binding protein that exists in HeLa and CBRH7919 cells was absent in normal rat liver. Conclusion cGPEI, GPEII specific binding proteins expressed in HeLa and CBRH7919 cells may play an important role in the high transcriptional level of the rGSTP1 gene in tumor cells.

  13. Structures of a putative ζ-class glutathione S-transferase from the pathogenic fungus Coccidioides immitis

    International Nuclear Information System (INIS)

    The pathogenic fungus C. immitis causes coccidioidomycosis, a potentially fatal disease. Here, apo and glutathione-bound crystal structures of a previously uncharacterized protein from C. immitis that appears to be a ζ-class glutathione S-transferase are presented. Coccidioides immitis is a pathogenic fungus populating the southwestern United States and is a causative agent of coccidioidomycosis, sometimes referred to as Valley Fever. Although the genome of this fungus has been sequenced, many operons are not properly annotated. Crystal structures are presented for a putative uncharacterized protein that shares sequence similarity with ζ-class glutathione S-transferases (GSTs) in both apo and glutathione-bound forms. The apo structure reveals a nonsymmetric homodimer with each protomer comprising two subdomains: a C-terminal helical domain and an N-terminal thioredoxin-like domain that is common to all GSTs. Half-site binding is observed in the glutathione-bound form. Considerable movement of some components of the active site relative to the glutathione-free form was observed, indicating an induced-fit mechanism for cofactor binding. The sequence homology, structure and half-site occupancy imply that the protein is a ζ-class glutathione S-transferase, a maleylacetoacetate isomerase (MAAI)

  14. Assignment of Biochemical Functions to Glycosyl Transferase Genes Which Are Essential for Biosynthesis of Exopolysaccharides in Sphingomonas Strain S88 and Rhizobium leguminosarum

    OpenAIRE

    Pollock, Thomas J.; van Workum, Wilbert A. T.; Thorne, Linda; Mikolajczak, Marcia J.; Yamazaki, Motohide; Kijne, Jan W.; Armentrout, Richard W.

    1998-01-01

    Glycosyl transferases which recognize identical substrates (nucleotide-sugars and lipid-linked carbohydrates) can substitute for one another in bacterial polysaccharide biosynthesis, even if the enzymes originate in different genera of bacteria. This substitution can be used to identify the substrate specificities of uncharacterized transferase genes. The spsK gene of Sphingomonas strain S88 and the pssDE genes of Rhizobium leguminosarum were identified as encoding glucuronosyl-(β1→4)-glucosy...

  15. In Vitro and in Vivo Effects of Three Different Mitragyna speciosa Korth Leaf Extracts on Phase II Drug Metabolizing Enzymes—Glutathione Transferases (GSTs)

    OpenAIRE

    Sharif Mahsufi Mansor; Mohd Ikram Mohd Said; Surash Ramanathan; Mohd Nizam Mordi; Sabariah Ismail; Juzaili Azizi

    2010-01-01

    In the present study, we investigate the effects of three different Mitragyna speciosa extracts, namely methanolic, aqueous and total alkaloid extracts, on glutathione transferase-specific activity in male Sprague Dawley rat liver cytosol in vitro and in vivo. In the in vitro study, the effect of Mitragyna speciosa extracts (0.01 to 750 µg/mL) against the specific activity of glutathione transferases was examined in rat liver cytosolic fraction from untreated rats. Our data show concentration...

  16. Arylamine N-acetyl Transferase (NAT) in the blue secretion of Telescopium telescopium: xenobiotic metabolizing enzyme as a biomarker for detection of environmental pollution

    OpenAIRE

    Gorain, Bapi; Chakraborty, Sumon; Pal, Murari Mohan; Sarkar, Ratul; Samanta, Samir Kumar; Karmakar, Sanmoy; Sen, Tuhinadri

    2014-01-01

    Telescopium telescopium, a marine mollusc collected from Sundarban mangrove, belongs to the largest mollusca phylum in the world and exudes a blue secretion when stimulated mechanically. The blue secretion was found to metabolize (preferentially) para-amino benzoic acid, a substrate for N-acetyl transferase (NAT), thereby indicating acetyl transferase like activity of the secretion. Attempts were also made to characterise bioactive fraction of the blue secretion and to further use this as a b...

  17. Glutathione S-transferase P protects against cyclophosphamide-induced cardiotoxicity in mice

    Energy Technology Data Exchange (ETDEWEB)

    Conklin, Daniel J., E-mail: dj.conklin@louisville.edu [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Institute of Molecular Cardiology, University of Louisville, Louisville, KY 40292 (United States); Haberzettl, Petra; Jagatheesan, Ganapathy; Baba, Shahid [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Institute of Molecular Cardiology, University of Louisville, Louisville, KY 40292 (United States); Merchant, Michael L. [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Division of Nephrology, Department of Medicine, University of Louisville, Louisville, KY 40292 (United States); Prough, Russell A. [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Department of Biochemistry and Molecular Biology, University of Louisville, Louisville, KY 40292 (United States); Williams, Jessica D. [University of Cincinnati College of Medicine, Internal Medicine, Cincinnati, OH 45267 (United States); Prabhu, Sumanth D. [Division of Cardiovascular Disease, University of Alabama-Birmingham, Birmingham, AL 35294 (United States); Bhatnagar, Aruni [Diabetes and Obesity Center, University of Louisville, Louisville, KY 40292 (United States); Institute of Molecular Cardiology, University of Louisville, Louisville, KY 40292 (United States); Department of Biochemistry and Molecular Biology, University of Louisville, Louisville, KY 40292 (United States)

    2015-06-01

    High-dose chemotherapy regimens using cyclophosphamide (CY) are frequently associated with cardiotoxicity that could lead to myocyte damage and congestive heart failure. However, the mechanisms regulating the cardiotoxic effects of CY remain unclear. Because CY is converted to an unsaturated aldehyde acrolein, a toxic, reactive CY metabolite that induces extensive protein modification and myocardial injury, we examined the role of glutathione S-transferase P (GSTP), an acrolein-metabolizing enzyme, in CY cardiotoxicity in wild-type (WT) and GSTP-null mice. Treatment with CY (100–300 mg/kg) increased plasma levels of creatine kinase-MB isoform (CK·MB) and heart-to-body weight ratio to a significantly greater extent in GSTP-null than WT mice. In addition to modest yet significant echocardiographic changes following acute CY-treatment, GSTP insufficiency was associated with greater phosphorylation of c-Jun and p38 as well as greater accumulation of albumin and protein–acrolein adducts in the heart. Mass spectrometric analysis revealed likely prominent modification of albumin, kallikrein-1-related peptidase, myoglobin and transgelin-2 by acrolein in the hearts of CY-treated mice. Treatment with acrolein (low dose, 1–5 mg/kg) also led to increased heart-to-body weight ratio and myocardial contractility changes. Acrolein induced similar hypotension in GSTP-null and WT mice. GSTP-null mice also were more susceptible than WT mice to mortality associated with high-dose acrolein (10–20 mg/kg). Collectively, these results suggest that CY cardiotoxicity is regulated, in part, by GSTP, which prevents CY toxicity by detoxifying acrolein. Thus, humans with low cardiac GSTP levels or polymorphic forms of GSTP with low acrolein-metabolizing capacity may be more sensitive to CY toxicity. - Graphical abstract: Cyclophosphamide (CY) treatment results in P450-mediated metabolic formation of phosphoramide mustard and acrolein (3-propenal). Acrolein is either metabolized and

  18. Glutathione S-transferase P protects against cyclophosphamide-induced cardiotoxicity in mice

    International Nuclear Information System (INIS)

    High-dose chemotherapy regimens using cyclophosphamide (CY) are frequently associated with cardiotoxicity that could lead to myocyte damage and congestive heart failure. However, the mechanisms regulating the cardiotoxic effects of CY remain unclear. Because CY is converted to an unsaturated aldehyde acrolein, a toxic, reactive CY metabolite that induces extensive protein modification and myocardial injury, we examined the role of glutathione S-transferase P (GSTP), an acrolein-metabolizing enzyme, in CY cardiotoxicity in wild-type (WT) and GSTP-null mice. Treatment with CY (100–300 mg/kg) increased plasma levels of creatine kinase-MB isoform (CK·MB) and heart-to-body weight ratio to a significantly greater extent in GSTP-null than WT mice. In addition to modest yet significant echocardiographic changes following acute CY-treatment, GSTP insufficiency was associated with greater phosphorylation of c-Jun and p38 as well as greater accumulation of albumin and protein–acrolein adducts in the heart. Mass spectrometric analysis revealed likely prominent modification of albumin, kallikrein-1-related peptidase, myoglobin and transgelin-2 by acrolein in the hearts of CY-treated mice. Treatment with acrolein (low dose, 1–5 mg/kg) also led to increased heart-to-body weight ratio and myocardial contractility changes. Acrolein induced similar hypotension in GSTP-null and WT mice. GSTP-null mice also were more susceptible than WT mice to mortality associated with high-dose acrolein (10–20 mg/kg). Collectively, these results suggest that CY cardiotoxicity is regulated, in part, by GSTP, which prevents CY toxicity by detoxifying acrolein. Thus, humans with low cardiac GSTP levels or polymorphic forms of GSTP with low acrolein-metabolizing capacity may be more sensitive to CY toxicity. - Graphical abstract: Cyclophosphamide (CY) treatment results in P450-mediated metabolic formation of phosphoramide mustard and acrolein (3-propenal). Acrolein is either metabolized and

  19. Association of catechol-o-methyl transferase gene polymorphism with prostate cancer and benign prostatic hyperplasia

    Directory of Open Access Journals (Sweden)

    mir davood omrani

    2009-08-01

    Full Text Available

    • BACKGROUND: A single nucleotide variation within  atechol-o-methyl transferase (COMT gene may alter the COMT enzyme activity level. Polymorphism of Val158Met in the COMT gene has been related to malignancy. In this regard, a study was carried out to find a possible association between the COMT gene polymorphism in patients with sporadic prostate cancer (PCa and benign prostatic hyperplasia (BPH.
    • METHODS: All types of COMT158 Val/Met polymorphism were carried out using ASO-PCR method in 41 patients with prostate cancer, 193 patients with benign prostatic hyperplasia and 107 healthy male individuals.
    • RESULTS: The results of this study showed that the frequency of low producer allele A at codon 158 of the  OMT gene is significantly different in BPH group compared to normal male control group (OR, 95% CI, p value 1.95: 1.46, 2.44, 0.021, respectively. However no significant difference was noticed when the comparison was made between prostate cancer group and normal male control group and also between BPH and PCa groups.
    • CONCLUSIONS: Decreased level of catechol-o-methyl transferase gene

    • Exploiting the Substrate Promiscuity of Hydroxycinnamoyl-CoA:Shikimate Hydroxycinnamoyl Transferase to Reduce Lignin

      Science.gov (United States)

      Eudes, Aymerick; Pereira, Jose H.; Yogiswara, Sasha; Wang, George; Teixeira Benites, Veronica; Baidoo, Edward E.K.; Lee, Taek Soon; Adams, Paul D.; Keasling, Jay D.; Loqué, Dominique

      2016-01-01

      Lignin poses a major challenge in the processing of plant biomass for agro-industrial applications. For bioengineering purposes, there is a pressing interest in identifying and characterizing the enzymes responsible for the biosynthesis of lignin. Hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyl transferase (HCT; EC 2.3.1.133) is a key metabolic entry point for the synthesis of the most important lignin monomers: coniferyl and sinapyl alcohols. In this study, we investigated the substrate promiscuity of HCT from a bryophyte (Physcomitrella) and from five representatives of vascular plants (Arabidopsis, poplar, switchgrass, pine and Selaginella) using a yeast expression system. We demonstrate for these HCTs a conserved capacity to acylate with p-coumaroyl-CoA several phenolic compounds in addition to the canonical acceptor shikimate normally used during lignin biosynthesis. Using either recombinant HCT from switchgrass (PvHCT2a) or an Arabidopsis stem protein extract, we show evidence of the inhibitory effect of these phenolics on the synthesis of p-coumaroyl shikimate in vitro, which presumably occurs via a mechanism of competitive inhibition. A structural study of PvHCT2a confirmed the binding of a non-canonical acceptor in a similar manner to shikimate in the active site of the enzyme. Finally, we exploited in Arabidopsis the substrate flexibility of HCT to reduce lignin content and improve biomass saccharification by engineering transgenic lines that overproduce one of the HCT non-canonical acceptors. Our results demonstrate conservation of HCT substrate promiscuity and provide support for a new strategy for lignin reduction in the effort to improve the quality of plant biomass for forage and cellulosic biofuels. PMID:26858288

  1. Cloning and characterization of two glutathione S-transferases from pyrethroid resistant Culex pipiens

    Science.gov (United States)

    Samra, Aman I; Kamita, Shizuo G; Yao, Hong-Wei; Cornel, Anthony J; Hammock, Bruce D

    2013-01-01

    BACKGROUND The Marin strain of Culex pipiens Say is a pyrethroid-resistant population that was collected in Marin County, California, in 2001 and subsequently maintained in the laboratory under regular permethrin exposure. RESULTS In this study, two genes, CpGSTd1 and CpGSTd2, encoding glutathione S-transferase (GST) were cloned from Cx. pipiens Marin. Phylogenetic analysis of the deduced amino acid sequences, CpGSTD1 and CpGSTD2, of these genes indicated that they belong to the Delta class of insect GSTs. The nucleotide and deduced amino acid sequences of CpGSTd1 and CpGSTd2 were 59% and 48% identical, respectively. CpGSTD1 and CpGSTD2 were expressed in Escherichia coli and purified by affinity chromatography. The recombinant GSTs exhibited unique selectivity towards the general GST substrates CDNB and DCNB, and also differed in their sensitivity to known inhibitors of GSTs. CpGSTD1 exhibited peroxidase activity with cumene hydroperoxide, while CpGSTD2 appeared to lack this activity. CpGSTD1 was able to metabolize DDT, while DDT metabolism by CpGSTD2 was not detectable. CpGSTD1 and CpGSTD2 showed no detectable metabolism of permethrin. Gene expression of CpGSTd1 and CpGSTd2 in Marin mosquitoes was elevated by about 2-fold in comparison to that found in a pyrethroid-sensitive mosquito strain. CONCLUSION Our results indicated that CpGSTD1 and CpGSTD2 have unique biochemical characteristics but they did not appear to play major roles in permethrin resistance in Marin mosquitoes. PMID:22290868

  2. Characterization of Ser73 in Arabidopsis thaliana Glutathione S-transferase zeta class

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Glutathione S-transferases (GSTs) are ubiquitous detoxifying superfamily enzymes. The zeta class GST from Arabidopsis thaliana (AtGSTZ) can efficiently degrade dichloroacetic acid (DCA), which is a common carcinogenic contaminant in drinking water. Ser73 in AtGSTZ is a conserved residue at Glutathione binding site (G-site). Compared with the equivalent residues in other GSTs, the catalytic and structural properties of Ser73 were poorly investigated. In this article, site-saturation mutagenesis was performed to characterize the detailed role of Ser73. The DCA de.chlorinating (DCA-DC) activity showed that most of the mutants had less than 3% of the wild-type activity, except S73T and $73A showing 43.48% and 21.62% of the wild-type activity, respectively, indicating that position 73 in AtGSTZ showed low mutational substitutability. Kinetic experiments revealed that mutants S73T, $73A, and S73G showed low binding affinity and catalytic efficiency toward DCA, 1.8-, 3.1-, and 10.7- fold increases in KmDcA values and 4.0-, 9.6-, and 34.1- fold decreases in KcatDCA/KmDCA values, respectively, compared to the wild type. Thermostability and refolding experiments showed that the wild type maintalned more thermostability and recovered activity. These results demonstrated the important role of Set73 in catalytic activity and structural stability of the enzyme. Such properties of Set73 could be particularly crucial to the molecular evolution of AtGSTZ and might,therefore, help explain why Ser73 is conserved in all GSTs. This conclusion might provide insights into the directed evolution of the DCA-DC activity of AtGSTZ.

  3. Genetic polymorphism in three glutathione s-transferase genes and breast cancer risk; TOPICAL

    International Nuclear Information System (INIS)

    The role of the glutathione S-transferase (GST) enzyme family is to detoxify environmental toxins and carcinogens and to protect organisms from their adverse effects, including cancer. The genes GSTM1, GSTP1, and GSTT1 code for three GSTs involved in the detoxification of carcinogens, such as polycyclic aromatic hydrocarbons (PAHs) and benzene. In humans, GSTM1 is deleted in about 50% of the population, GSTT1 is absent in about 20%, whereas the GSTP1 gene has a single base polymorphism resulting in an enzyme with reduced activity. Epidemiological studies indicate that GST polymorphisms increase the level of carcinogen-induced DNA damage and several studies have found a correlation of polymorphisms in one of the GST genes and an increased risk for certain cancers. We examined the role of polymorphisms in genes coding for these three GST enzymes in breast cancer. A breast tissue collection consisting of specimens of breast cancer patients and non-cancer controls was analyzed by polymerase chain reaction (PCR) for the presence or absence of the GSTM1 and GSTT1 genes and for GSTP1 single base polymorphism by PCR/RFLP. We found that GSTM1 and GSTT1 deletions occurred more frequently in cases than in controls, and GSTP1 polymorphism was more frequent in controls. The effective detoxifier (putative low-risk) genotype (defined as presence of both GSTM1 and GSTT1 genes and GSTP1 wild type) was less frequent in cases than controls (16% vs. 23%, respectively). The poor detoxifier (putative high-risk) genotype was more frequent in cases than controls. However, the sample size of this study was too small to provide conclusive results

  4. Structural analysis of an epsilon-class glutathione transferase from housefly, Musca domestica.

    Science.gov (United States)

    Nakamura, Chihiro; Yajima, Shunsuke; Miyamoto, Toru; Sue, Masayuki

    2013-01-25

    Glutathione transferases (GSTs) play an important role in the detoxification of insecticides, and as such, they are a key contributor to enhanced resistance to insecticides. In the housefly (Musca domestica), two epsilon-class GSTs (MdGST6A and MdGST6B) that share high sequence homology have been identified, which are believed to be involved in resistance against insecticides. The structural determinants controlling the substrate specificity and enzyme activity of MdGST6s are unknown. The aim of this study was to crystallize and perform structural analysis of the GST isozyme, MdGST6B. The crystal structure of MdGST6B complexed with reduced glutathione (GSH) was determined at a resolution of 1.8 Å. MdGST6B was found to have a typical GST folding comprised of N-terminal and C-terminal domains. Arg113 and Phe121 on helix 4 were shown to protrude into the substrate binding pocket, and as a result, the entrance of the substrate binding pocket was narrower compared to delta- and epsilon-class GSTs from Africa malaria vector Anopheles gambiae, agGSTd1-6 and agGSTe2, respectively. This substrate pocket narrowing is partly due to the presence of a π-helix in the middle of helix 4. Among the six residues that donate hydrogen bonds to GSH, only Arg113 was located in the C-terminal domain. Ala substitution of Arg113 did not have a significant effect on enzyme activity, suggesting that the Arg113 hydrogen bond does not play a crucial role in catalysis. On the other hand, mutation at Phe108, located just below Arg113 in the binding pocket, reduced the affinity and catalytic activity to both GSH and the electrophilic co-substrate, 1-chloro-2,4-dinitrobenzene. PMID:23268341

  5. A critical perspective of the diverse roles of O-GlcNAc transferase in chromatin.

    Science.gov (United States)

    Gambetta, Maria Cristina; Müller, Jürg

    2015-12-01

    O-linked β-N-Acetylglucosamine (O-GlcNAc) is a posttranslational modification that is catalyzed by O-GlcNAc transferase (Ogt) and found on a plethora of nuclear and cytosolic proteins in animals and plants. Studies in different model organisms revealed that while O-GlcNAc is required for selected processes in Caenorhabditis elegans and Drosophila, it has evolved to become required for cell viability in mice, and this has challenged investigations to identify cellular functions that critically require this modification in mammals. Nevertheless, a principal cellular process that engages O-GlcNAcylation in all of these species is the regulation of gene transcription. Here, we revisit several of the primary experimental observations that led to current models of how O-GlcNAcylation affects gene expression. In particular, we discuss the role of the stable association of Ogt with the transcription factors Hcf1 and Tet, the two main Ogt-interacting proteins in nuclei of mammalian cells. We also critically evaluate the evidence that specific residues on core histones, including serine 112 of histone 2B (H2B-S112), are O-GlcNAcylated in vivo and discuss possible physiological effects of these modifications. Finally, we review our understanding of the role of O-GlcNAcylation in Drosophila, where recent studies suggest that the developmental defects in Ogt mutants are all caused by lack of O-GlcNAcylation of a single transcriptional regulator, the Polycomb repressor protein Polyhomeotic (Ph). Collectively, this reexamination of the experimental evidence suggests that a number of recently propagated models about the role of O-GlcNAcylation in transcriptional control should be treated cautiously. PMID:25894967

  6. Cloning of a glutathione S-transferase decreasing during differentiation of HL60 cell line

    International Nuclear Information System (INIS)

    By sequencing the Expressed Sequence Tags of human dermal papilla cDNA library, we identified a clone named K872 of which the expression decreased during differentiation of HL60 cell line. K872 plasmid DNA was isolated according to QIA plasmid extraction kit (Qiagen GmbH, Germany). The nucleotide sequencing was performed by Sanger's method with K872 plasmid DNA. The most updated GenBank EMBL necleic acid banks were searched through the internet by using BLAST (Basic Local Alignment Search Tools) program. Northern bots were performed using RNA isolated from various human tissues and cancer cell lines. The gene expression of the fusion protein was achieved by His-Patch Thiofusion expression system and the protein product was identified on SDS-PAGE. K872 clone is 1006 nucleotides long, and has a coding region of 675 nucleotides and a 3' non-coding region of 280 nucleotides. The presumed open reading frame starting at the 5' terminus of K872 encodes 226 amino acids, including the initiation methionine residue. The amino acid sequence deduced from the open reading frame of K872 shares 70% identity with that of rat glutathione S-transferase kappa 1 (rGSTK1). The transcripts were expressed inh a variety of human tissues and cancer cells. The levels of transcript were relatively high in those tissues such as heart, skeletal muscle, and peripheral blood leukocyte. It is noteworthy that K872 was found to be abundantly expressed in colorectal cancer and melanoma cell lines. Homology search result suggests that K872 clone is the human homolog of the rGSTK1 which is known to be involved in the resistance of cytotoxic therapy. We propose that meticulous functional analysis should be followed to confirm that

  7. Farnesyl transferase inhibitors induce extended remissions in transgenic mice with mature B cell lymphomas

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    Refaeli Yosef

    2008-05-01

    Full Text Available Abstract Background We have used a mouse model based on overexpression of c-Myc in B cells genetically engineered to be self-reactive to test the hypothesis that farnesyl transferase inhibitors (FTIs can effectively treat mature B cell lymphomas. FTIs are undergoing clinical trials to treat both lymphoid and non-lymphoid malignancies and we wished to obtain evidence to support the inclusion of B cell lymphomas in future trials. Results We report that two FTIs, L-744,832 and SCH66336, blocked the growth of mature B cell lymphoma cells in vitro and in vivo. The FTI treatment affected the proliferation and survival of the transformed B cells to a greater extent than naïve B cells stimulated with antigen. In syngeneic mice transplanted with the transgenic lymphoma cells, L-744,832 treatment prevented the growth of the tumor cells and the morbidity associated with the resulting lymphoma progression. Tumors that arose from transplantation of the lymphoma cells regressed with as little as three days of treatment with L-744,832 or SCH66336. Treatment of these established lymphomas with L-744,832 for seven days led to long-term remission of the disease in approximately 25% of animals. Conclusion FTI treatment can block the proliferation and survival of self-reactive transformed B cells that overexpress Myc. In mice transplanted with mature B cell lymphomas, we found that FTI treatment led to regression of disease. FTIs warrant further consideration as therapeutic agents for mature B cell lymphomas and other lymphoid tumors.

  8. Detection and adequacy evaluation of erythrocyte glutathione transferase on levels of circulating toxins in hemodialysis patients.

    Science.gov (United States)

    Yin, Rui; Qiu, Hui; Zuo, Huaiyun; Cui, Min; Zhai, Nailiang; Zheng, Hongguang; Zhang, Dewei; Huo, Ping; Hong, Min

    2016-08-01

    To explore detection and adequacy evaluation of erythrocyte glutathione S transferase (GST) on levels of circulating toxins in hemodialysis patients in Qinhuangdao region in China, this study divided 84 cases of long-term, end-stage hemodialysis patients into 2 groups: one group of 33 cases of adequate hemodialysis (spKt/V ≥ 1.3) and another group of 51 cases of inadequate hemodialysis (spKt/V GST, creatinine, high sensitivity C-reactive protein (hs-CRP), transferrin saturation (TSAT), parathyroid hormone (PTH), interleukin-2,6,8 (IL-2,6,8) and tumor necrosis factor-a (TNF-a) in the hemodialysis group were significantly higher than those in the control group (P GST, IL-2, 6, 8, and TNF-a levels in the inadequate hemodialysis group were significantly higher than in the adequate hemodialysis group (P GST and spKt/V, IL-2, IL-6, IL-8, and TNF-a have a positive correlation (P 0.05). There were 23 patients with levels of spKt/V ≥ 1.3 after adjusting the dialysis solution for 51 cases of inadequate hemodialysis patients, and the GST level after the adjustment was significantly lower than that before the adjustment, but still higher than that in the adequate dialysis group. This concludes that the maintenance of hemodialysis in patients has certain relevance on spKt/V and associated inflammatory factors. Through the study, it can be determined that GST can effectively respond to adequate hemodialysis, which has a guiding significance on adjusting the blood dialysis solution in clinical practice. PMID:27121915

  9. Glutathione S-Transferase Regulation in Calanus finmarchicus Feeding on the Toxic Dinoflagellate Alexandrium fundyense

    Science.gov (United States)

    Roncalli, Vittoria; Jungbluth, Michelle J.; Lenz, Petra H.

    2016-01-01

    The effect of the dinoflagellate, Alexandrium fundyense, on relative expression of glutathione S-transferase (GST) transcripts was examined in the copepod Calanus finmarchicus. Adult females were fed for 5-days on one of three experimental diets: control (100% Rhodomonas spp.), low dose of A. fundyense (25% by volume, 75% Rhodomonas spp.), and high dose (100% A. fundyense). Relative expression of three GST genes was measured using RT-qPCR on days 0.5, 1, 2 and 5 in two independent experiments. Differential regulation was found for the Delta and the Sigma GSTs between 0.5 to 2 days, but not on day 5 in both experiments. The third GST, a microsomal, was not differentially expressed in either treatment or day. RT-qPCR results from the two experiments were similar, even though experimental females were collected from the Gulf of Maine on different dates and their reproductive output differed. In the second experiment, expression of 39 GSTs was determined on days 2 and 5 using RNA-Seq. Global gene expression analyses agreed with the RT-qPCR results. Furthermore, the RNA-Seq measurements indicated that only four GSTs were differentially expressed under the experimental conditions, and the response was small in amplitude. In summary, the A. fundyense diet led to a rapid and transient response in C. finmarchicus in three cytosolic GSTs, while a fourth GST (Omega I) was significantly up-regulated on day 5. Although there was some regulation of GSTs in response the toxic dinoflagellate, the tolerance to A. fundyense by C. finmarchicus is not dependent on the long-term up-regulation of specific GSTs. PMID:27427938

  10. Genetic polymorphism in three glutathione s-transferase genes and breast cancer risk

    Energy Technology Data Exchange (ETDEWEB)

    Woldegiorgis, S.; Ahmed, R.C.; Zhen, Y.; Erdmann, C.A.; Russell, M.L.; Goth-Goldstein, R.

    2002-04-01

    The role of the glutathione S-transferase (GST) enzyme family is to detoxify environmental toxins and carcinogens and to protect organisms from their adverse effects, including cancer. The genes GSTM1, GSTP1, and GSTT1 code for three GSTs involved in the detoxification of carcinogens, such as polycyclic aromatic hydrocarbons (PAHs) and benzene. In humans, GSTM1 is deleted in about 50% of the population, GSTT1 is absent in about 20%, whereas the GSTP1 gene has a single base polymorphism resulting in an enzyme with reduced activity. Epidemiological studies indicate that GST polymorphisms increase the level of carcinogen-induced DNA damage and several studies have found a correlation of polymorphisms in one of the GST genes and an increased risk for certain cancers. We examined the role of polymorphisms in genes coding for these three GST enzymes in breast cancer. A breast tissue collection consisting of specimens of breast cancer patients and non-cancer controls was analyzed by polymerase chain reaction (PCR) for the presence or absence of the GSTM1 and GSTT1 genes and for GSTP1 single base polymorphism by PCR/RFLP. We found that GSTM1 and GSTT1 deletions occurred more frequently in cases than in controls, and GSTP1 polymorphism was more frequent in controls. The effective detoxifier (putative low-risk) genotype (defined as presence of both GSTM1 and GSTT1 genes and GSTP1 wild type) was less frequent in cases than controls (16% vs. 23%, respectively). The poor detoxifier (putative high-risk) genotype was more frequent in cases than controls. However, the sample size of this study was too small to provide conclusive results.

  11. Labeling embryonic stem cells with enhanced green fluorescent protein on the hypoxanthineguanine phosphoribosyl transferase locus

    Institute of Scientific and Technical Information of China (English)

    滕路; 孟国良; 刑阳; 尚克刚; 王小珂; 顾军

    2003-01-01

    Objective To label embryonic stem (ES) cells with enhanced green fluorescent protein (EGF P) on the hypoxanthineguanine phosphoribosyl transferase (HPRT) gene locus for t he first time to provide a convenient and efficient way for cell tracking and ma nipulation in the studies of transplantation and stem cell therapy.Methods Homologous fragments were obtained by polymerase chain reaction (PCR), from whic h the gene targeting vector pHPRT-EGFP was constructed. The linearized vector was introduced into ES cells by electroporation. The G418r6TGr cell clones were obtained after selection with G418 and 6TG media. The integration patterns of these resistant cell clones were identified with Southern blotting.Results EGFP expressing ES cells on the locus of HPRT were successfu lly generated. They have normal properties, such as karyotype, viability and di fferentiation ability. The green fluorescence of EGFP expressing cells was main tained in propagation of the ES cells for more than 30 passages and in different iated cells. Cultured in suspension, the "green" ES cells aggregated and forme d embryoid bodies, retaining the green fluorescence at varying developmental sta ges. The "green" embryoid bodies could expand and differentiate into various t ypes of cells, exhibiting ubiquitous green fluorescence. Conclusions This generation of "green" targeted ES cells is described in an efficient proto col for obtaining the homologous fragments by PCR. Introducing the marker gene in the genome of ES cells, we should be able to manipulate them in vitro and use them as vehicles in cell-replacement therapy as well as for other biomedical a nd research purposes.

  12. Ethnicity and glutathione S-transferase (GSTM1/GSTT1 polymorphisms in a Brazilian population

    Directory of Open Access Journals (Sweden)

    G.J.F. Gattás

    2004-04-01

    Full Text Available The distribution of polymorphisms related to glutathione S-transferases (GST has been described in different populations, mainly for white individuals. We evaluated the distribution of GST mu (GSTM1 and theta (GSTT1 genotypes in 594 individuals, by multiplex PCR-based methods, using amplification of the exon 7 of CYP1A1 gene as an internal control. In São Paulo, 233 whites, 87 mulattos, and 137 blacks, all healthy blood-donor volunteers, were tested. In Bahia, where black and mulatto populations are more numerous, 137 subjects were evaluated. The frequency of the GSTM1 null genotype was significantly higher among whites (55.4% than among mulattos (41.4%; P = 0.03 and blacks (32.8%; P < 0.0001 from São Paulo, or Bahian subjects in general (35.7%; P = 0.0003. There was no statistically different distribution among any non-white groups. The distribution of GSTT1 null genotype among groups did not differ significantly. The agreement between self-reported and interviewer classification of skin color in the Bahian group was low. The interviewer classification indicated a gradient of distribution of the GSTM1 null genotype from whites (55.6% to light mulattos (40.4%, dark mulattos (32.0% and blacks (28.6%. However, any information about race or ethnicity should be considered with caution regarding the bias introduced by different data collection techniques, specially in countries where racial admixture is intense, and ethnic definition boundaries are loose. Because homozygous deletions of GST gene might be associated with cancer risk, a better understanding of chemical metabolizing gene distribution can contribute to risk assessment of humans exposed to environmental carcinogens.

  13. Exploiting the Substrate Promiscuity of Hydroxycinnamoyl-CoA:Shikimate Hydroxycinnamoyl Transferase to Reduce Lignin.

    Science.gov (United States)

    Eudes, Aymerick; Pereira, Jose H; Yogiswara, Sasha; Wang, George; Teixeira Benites, Veronica; Baidoo, Edward E K; Lee, Taek Soon; Adams, Paul D; Keasling, Jay D; Loqué, Dominique

    2016-03-01

    Lignin poses a major challenge in the processing of plant biomass for agro-industrial applications. For bioengineering purposes, there is a pressing interest in identifying and characterizing the enzymes responsible for the biosynthesis of lignin. Hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyl transferase (HCT; EC 2.3.1.133) is a key metabolic entry point for the synthesis of the most important lignin monomers: coniferyl and sinapyl alcohols. In this study, we investigated the substrate promiscuity of HCT from a bryophyte (Physcomitrella) and from five representatives of vascular plants (Arabidopsis, poplar, switchgrass, pine and Selaginella) using a yeast expression system. We demonstrate for these HCTs a conserved capacity to acylate with p-coumaroyl-CoA several phenolic compounds in addition to the canonical acceptor shikimate normally used during lignin biosynthesis. Using either recombinant HCT from switchgrass (PvHCT2a) or an Arabidopsis stem protein extract, we show evidence of the inhibitory effect of these phenolics on the synthesis of p-coumaroyl shikimate in vitro, which presumably occurs via a mechanism of competitive inhibition. A structural study of PvHCT2a confirmed the binding of a non-canonical acceptor in a similar manner to shikimate in the active site of the enzyme. Finally, we exploited in Arabidopsis the substrate flexibility of HCT to reduce lignin content and improve biomass saccharification by engineering transgenic lines that overproduce one of the HCT non-canonical acceptors. Our results demonstrate conservation of HCT substrate promiscuity and provide support for a new strategy for lignin reduction in the effort to improve the quality of plant biomass for forage and cellulosic biofuels. PMID:26858288

  14. Deficiency of glutathione transferase zeta causes oxidative stress and activation of antioxidant response pathways.

    Science.gov (United States)

    Blackburn, Anneke C; Matthaei, Klaus I; Lim, Cindy; Taylor, Matthew C; Cappello, Jean Y; Hayes, John D; Anders, M W; Board, Philip G

    2006-02-01

    Glutathione S-transferase (GST) zeta (GSTZ1-1) plays a significant role in the catabolism of phenylalanine and tyrosine, and a deficiency of GSTZ1-1 results in the accumulation of maleylacetoacetate and its derivatives maleylacetone (MA) and succinylacetone. Induction of GST subunits was detected in the liver of Gstz1(-/-) mice by Western blotting with specific antisera and high-performance liquid chromatography analysis of glutathione affinity column-purified proteins. The greatest induction was observed in members of the mu class. Induction of NAD(P)H:quinone oxidoreductase 1 and the catalytic and modifier subunits of glutamate-cysteine ligase was also observed. Many of the enzymes that are induced in Gstz1(-/-) mice are regulated by antioxidant response elements that respond to oxidative stress via the Keap1/Nrf2 pathway. It is significant that diminished glutathione concentrations were also observed in the liver of Gstz1(-/-) mice, which supports the conclusion that under normal dietary conditions, the accumulation of electrophilic intermediates such as maleylacetoacetate and MA results in a high level of oxidative stress. Elevated GST activities in the livers of Gstz1(-/-) mice suggest that GSTZ1-1 deficiency may alter the metabolism of some drugs and xenobiotics. Gstz1(-/-) mice given acetaminophen demonstrated increased hepatotoxicity compared with wild-type mice. This toxicity may be attributed to the increased GST activity or the decreased hepatic concentrations of glutathione, or both. Patients with acquired deficiency of GSTZ1-1 caused by therapeutic exposure to dichloroacetic acid for the clinical treatment of lactic acidosis may be at increased risk of drug- and chemical-induced toxicity. PMID:16278372

  15. Glutathione S Transferases Polymorphisms Are Independent Prognostic Factors in Lupus Nephritis Treated with Cyclophosphamide.

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    Alexandra Audemard-Verger

    Full Text Available To investigate association between genetic polymorphisms of GST, CYP and renal outcome or occurrence of adverse drug reactions (ADRs in lupus nephritis (LN treated with cyclophosphamide (CYC. CYC, as a pro-drug, requires bioactivation through multiple hepatic cytochrome P450s and glutathione S transferases (GST.We carried out a multicentric retrospective study including 70 patients with proliferative LN treated with CYC. Patients were genotyped for polymorphisms of the CYP2B6, CYP2C19, GSTP1, GSTM1 and GSTT1 genes. Complete remission (CR was defined as proteinuria ≤0.33g/day and serum creatinine ≤124 µmol/l. Partial remission (PR was defined as proteinuria ≤1.5g/day with a 50% decrease of the baseline proteinuria value and serum creatinine no greater than 25% above baseline.Most patients were women (84% and 77% were Caucasian. The mean age at LN diagnosis was 41 ± 10 years. The frequency of patients carrying the GST null genotype GSTT1-, GSTM1-, and the Ile→105Val GSTP1 genotype were respectively 38%, 60% and 44%. In multivariate analysis, the Ile→105Val GSTP1 genotype was an independent factor of poor renal outcome (achievement of CR or PR (OR = 5.01 95% CI [1.02-24.51] and the sole factor that influenced occurrence of ADRs was the GSTM1 null genotype (OR = 3.34 95% CI [1.064-10.58]. No association between polymorphisms of cytochrome P450s gene and efficacy or ADRs was observed.This study suggests that GST polymorphisms highly impact renal outcome and occurrence of ADRs related to CYC in LN patients.

  16. The stereochemical course of 4-hydroxy-2-nonenal metabolism by glutathione S-transferases.

    Science.gov (United States)

    Balogh, Larissa M; Roberts, Arthur G; Shireman, Laura M; Greene, Robert J; Atkins, William M

    2008-06-13

    4-Hydroxy-2-nonenal (HNE) is a toxic aldehyde generated during lipid peroxidation and has been implicated in a variety of pathological states associated with oxidative stress. Glutathione S-transferase (GST) A4-4 is recognized as one of the predominant enzymes responsible for the metabolism of HNE. However, substrate and product stereoselectivity remain to be fully explored. The results from a product formation assay indicate that hGSTA4-4 exhibits a modest preference for the biotransformation of S-HNE in the presence of both enantiomers. Liquid chromatography mass spectrometry analyses using the racemic and enantioisomeric HNE substrates explicitly demonstrate that hGSTA4-4 conjugates glutathione to both HNE enantiomers in a completely stereoselective manner that is not maintained in the spontaneous reaction. Compared with other hGST isoforms, hGSTA4-4 shows the highest degree of stereoselectivity. NMR experiments in combination with simulated annealing structure determinations enabled the determination of stereochemical configurations for the GSHNE diastereomers and are consistent with an hGSTA4-4-catalyzed nucleophilic attack that produces only the S-configuration at the site of conjugation, regardless of substrate chirality. In total these results indicate that hGSTA4-4 exhibits an intriguing combination of low substrate stereoselectivity with strict product stereoselectivity. This behavior allows for the detoxification of both HNE enantiomers while generating only a select set of GSHNE diastereomers with potential stereochemical implications concerning their effects and fates in biological tissues. PMID:18424441

  17. Cloning, expression and analysis of the olfactory glutathione S-transferases in coho salmon.

    Science.gov (United States)

    Espinoza, Herbert M; Shireman, Laura M; McClain, Valerie; Atkins, William; Gallagher, Evan P

    2013-03-15

    The glutathione S-transferases (GSTs) provide cellular protection by detoxifying xenobiotics, maintaining redox status, and modulating secondary messengers, all of which are critical to maintaining olfaction in salmonids. Here, we characterized the major coho salmon olfactory GSTs (OlfGSTs), namely omega, pi, and rho subclasses. OlfGST omega contained an open reading frame of 720bp and encoded a protein of 239 amino acids. OlfGST pi and OlfGST rho contained open reading frames of 627 and 681nt, respectively, and encoded proteins of 208 and 226 amino acids. Whole-protein mass spectrometry yielded molecular weights of 29,950, 23,354, and 26,655Da, respectively, for the GST omega, pi, and rho subunits. Homology modeling using four protein-structure prediction algorithms suggest that the active sites in all three OlfGST isoforms resembled counterparts in other species. The olfactory GSTs conjugated prototypical GST substrates, but only OlfGST rho catalyzed the demethylation of the pesticide methyl parathion. OlfGST pi and rho exhibited thiol oxidoreductase activity toward 2-hydroxyethyl disulfide (2-HEDS) and conjugated 4-hydroxynonenal (HNE), a toxic aldehyde with neurodegenerative properties. The kinetic parameters for OlfGST pi conjugation of HNE were K(M)=0.16 ± 0.06mM and V(max)=0.5 ± 0.1μmolmin⁻¹mg⁻¹, whereas OlfGST rho was more efficient at catalyzing HNE conjugation (K(M)=0.022 ± 0.008 mM and V(max)=0.47 ± 0.05μmolmin⁻¹mg⁻¹). Our findings indicate that the peripheral olfactory system of coho expresses GST isoforms that detoxify certain electrophiles and pesticides and that help maintain redox status and signal transduction. PMID:23261526

  18. Glutathione S-transferase genotypes modify lung function decline in the general population: SAPALDIA cohort study

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    Ackermann-Liebrich Ursula

    2007-01-01

    Full Text Available Abstract Background Understanding the environmental and genetic risk factors of accelerated lung function decline in the general population is a first step in a prevention strategy against the worldwide increasing respiratory pathology of chronic obstructive pulmonary disease (COPD. Deficiency in antioxidative and detoxifying Glutathione S-transferase (GST gene has been associated with poorer lung function in children, smokers and patients with respiratory diseases. In the present study, we assessed whether low activity variants in GST genes are also associated with accelerated lung function decline in the general adult population. Methods We examined with multiple regression analysis the association of polymorphisms in GSTM1, GSTT1 and GSTP1 genes with annual decline in FEV1, FVC, and FEF25–75 during 11 years of follow-up in 4686 subjects of the prospective SAPALDIA cohort representative of the Swiss general population. Effect modification by smoking, gender, bronchial hyperresponisveness and age was studied. Results The associations of GST genotypes with FEV1, FVC, and FEF25–75 were comparable in direction, but most consistent for FEV1. GSTT1 homozygous gene deletion alone or in combination with GSTM1 homozygous gene deletion was associated with excess decline in FEV1 in men, but not women, irrespective of smoking status. The additional mean annual decline in FEV1 in men with GSTT1 and concurrent GSTM1 gene deletion was -8.3 ml/yr (95% confidence interval: -12.6 to -3.9 relative to men without these gene deletions. The GSTT1 effect on the FEV1 decline comparable to the observed difference in FEV1 decline between never and persistent smoking men. Effect modification by gender was statistically significant. Conclusion Our results suggest that genetic GSTT1 deficiency is a prevalent and strong determinant of accelerated lung function decline in the male general population.

  19. Enhanced tolerance and remediation of anthracene by transgenic tobacco plants expressing a fungal glutathione transferase gene

    International Nuclear Information System (INIS)

    Highlights: → Transgenic plants expressing a TvGST gene were tested for tolerance, uptake and degradation of anthracene. → Transgenic plants were more tolerant to anthracene and take up more anthracene from soil and solutions compared to control plants. → Using in vitro T1 seedlings, we showed that anthracene-a three fused benzene ring compound was phytodegraded to naphthalene derivatives, having two benzene rings. → This is the first time that a transgenic plant was shown to have the potential to phytodegrade anthracene. - Abstract: Plants can be used for remediation of polyaromatic hydrocarbons, which are known to be a major concern for human health. Metabolism of xenobiotic compounds in plants occurs in three phases and glutathione transferases (GST) mediate phase II of xenobiotic transformation. Plants, although have GSTs, they are not very efficient for degradation of exogenous recalcitrant xenobiotics including polyaromatic hydrocarbons. Hence, heterologous expression of efficient GSTs in plants may improve their remediation and degradation potential of xenobiotics. In the present study, we investigated the potential of transgenic tobacco plants expressing a Trichoderma virens GST for tolerance, remediation and degradation of anthracene-a recalcitrant polyaromatic hydrocarbon. Transgenic plants with fungal GST showed enhanced tolerance to anthracene compared to control plants. Remediation of 14C uniformly labeled anthracene from solutions and soil by transgenic tobacco plants was higher compared to wild-type plants. Transgenic plants (T0 and T1) degraded anthracene to naphthalene derivatives, while no such degradation was observed in wild-type plants. The present work has shown that in planta expression of a fungal GST in tobacco imparted enhanced tolerance as well as higher remediation potential of anthracene compared to wild-type plants.

  20. Extração, purificação e avaliação da atividade da glutationa S-Transferase de fígado bovino Extraction of glutathione s-transferase from bovine liver

    Directory of Open Access Journals (Sweden)

    Maria Célia Lopes Torres

    2006-04-01

    Full Text Available Considerando a ação detoxificante da enzima Glutationa S-Transferase (GST, importante contra o estresse oxidativo, câncer e outras doenças degenerativas, com este estudo, objetivou-se avaliar a atividade dessa enzima extraída de fígado bovino e avaliar a estabilidade em condições de refrigeração (5(0C. O fígado bovino foi selecionado por ser matéria prima disponível comercialmente e de baixo custo. A extração foi realizada em quatro etapas (homogeneização/centrifugação, passagem em coluna contendo dietilaminoetil-celulose (DEAE-celulose, precipitação com sulfato de amônia e passagem em coluna contendo Carboximetilcelulose (CMC. O extrato obtido apresentou atividade com o 1 cloro 2, 4 dinitrobenzeno, na presença de glutationa reduzida. O extrato final apresentou atividade específica 5 vezes maior que o extrato bruto centrifugado e estabilidade da atividade enzimática foi mantida nas condições de 5(0C, durante 70 dias.Considering the detoxication functions of Glutathione S-transferase (GST enzyme, that is important against oxidative stress, cancer and others degenerative diseases, this study aimed to evaluate the stability and activity of Glutathione S-transferase extracted from bovine liver, which is commercially available at low cost. The extraction was done in four steps (homogenization/centrifugation, passage through column containing diethylaminoethyl-cellulose (DEAE, precipitation with ammonium sulfate and passing through column of carboxy-methyl-cellulose (CMC. The extract thus obtained showed activity with 1 chloro 2, 4 dinitrobenzene, in the presence of reduced glutation. The specific activity of the final extract was 5 times greater than the crude centrifuged extract, and was stable for 70 days when stored at 5 ºC.

  1. Habitual consumption of fruits and vegetables: associations with human rectal glutathione S-transferase.

    Science.gov (United States)

    Wark, Petra A; Grubben, Marina J A L; Peters, Wilbert H M; Nagengast, Fokko M; Kampman, Ellen; Kok, Frans J; van 't Veer, Pieter

    2004-11-01

    The glutathione (GSH)/glutathione S-transferase (GST) system is an important detoxification system in the gastrointestinal tract. A high activity of this system may benefit cancer prevention. The aim of the study was to assess whether habitual consumption of fruits and vegetables, especially citrus fruits and brassica and allium vegetables, is positively associated with parameters reflecting the activity of the GSH/GST enzyme system in human rectal mucosa. GST enzyme activity, GST isoenzyme levels of GST-alpha (A1-1, A1-2 and A2-2), -mu (M1-1) and -pi (P1-1), and GSH levels were measured in rectal biopsies from 94 subjects. Diet, lifestyle, GSTM1 and GSTT1 null polymorphisms were assessed. Mean GST enzyme activity was 237 nmol/min/mg protein (SD = 79). Consumption of citrus fruits was positively associated with GST enzyme activity [difference between high and low consumption: 28.9 (95% confidence interval (CI) = 9.3-48.6) nmol/min/mg protein], but was not associated with the other parameters. A positive association with brassica vegetables was found among carriers of the GSTM1-plus genotype [difference between high and low consumption: 22.6 (95% CI = 0.2-45.0) nmol/min/mg protein], but not among GSTM1-null individuals (-25.8 nmol/min/mg protein, 95% CI = -63.3-11.8). This is in line with a positive association between consumption of brassica vegetables and GSTM isoenzyme level [difference between high and low consumption: 67.5%, 95% CI = (6.8-162.7)]. Consumption of allium vegetables was not associated with GST enzyme activity, but negatively with GSTP1-1 levels [difference between high and low consumption: -23.3%, 95% CI = (-35.5; -8.6)]. Associations were similar among those with the GSTT1-plus and GSTT1-null genotype. In conclusion, variations in habitual consumption of fruits, particularly citrus fruits, and of vegetables, in particular brassica vegetables, among those with the GSTM1-plus genotype, may contribute to variations in human rectal GST enzyme

  2. Inhibition characteristics of hypericin on rat small intestine glutathione-S-transferases.

    Science.gov (United States)

    Tuna, Gamze; Kulaksiz Erkmen, Gulnihal; Dalmizrak, Ozlem; Dogan, Arin; Ogus, I Hamdi; Ozer, Nazmi

    2010-10-01

    Glutathione-S-transferases constitute a family of enzymes involving in the detoxification of xenobiotics, signalling cascades and serving as ligandins or/and catalyzing the conjugation of various chemicals and drugs. The widely expressed cytosolic GST-pi is a marker protein in various cancers and its increased concentration is linked to drug resistance. GST-pi is autoregulated by S-glutathionylation and it catalyzes the S-glutathionylation of other proteins in response to oxidative or nitrosative stress. S-glutathionylation of GST-pi results in multimer formation and the breakage of ligand binding interactions with c-Jun NH(2)-terminal kinase (JNK). Another widely expressed GST enzyme, GST-alpha is assumed as a marker in hepatocellular damage, is implicated in cancer, asthma, cardiovascular disease and response to chemotherapy. Although, it was shown that hypericin binds and inhibits GST-alpha and GST-pi, the inhibition characteristics have not been investigated in detail. The aim of this study was to investigate the effects of hypericin on major GSTs; GST-alpha and GST-pi purified from rat small intestine. When GSH used as varied substrate the inhibition pattern with hypericin was uncompetitive for GST-alpha (K(i)=0.16 + or - 0.02 microM) and noncompetitive for GST-pi (K(i) = 2.46 + or - 0.43 microM). While using CDNB (1-chloro-2,4-dinitrobenzene) as the varied substrate, the inhibition patterns were noncompetitive for GST-alpha and competitive for GST-pi; K(i) values for GST-alpha and GST-pi were 1.91 + or - 0.21 and 0.55 + or - 0.07 microM, respectively. Since hypericin accumulated in cancer cells and important in photodynamic therapy (PDT), inhibition of GST-alpha and GST-pi by hypericin might increase the effectivity of the treatment. Considering that GST-pi is responsible for the drug resistance its inhibition might increase the benefit obtained from chemotherapy. PMID:20637187

  3. Glutathione S-transferase activity in follicular fluid from women undergoing ovarian stimulation: role in maturation.

    Science.gov (United States)

    Meijide, Susana; Hernández, M Luisa; Navarro, Rosaura; Larreategui, Zaloa; Ferrando, Marcos; Ruiz-Sanz, José Ignacio; Ruiz-Larrea, M Begoña

    2014-10-01

    Female infertility involves an emotional impact for the woman, often leading to a state of anxiety and low self-esteem. The assisted reproduction techniques (ART) are used to overcome the problem of infertility. In a first step of the in vitro fertilization therapy women are subjected to an ovarian stimulation protocol to obtain mature oocytes, which will result in competent oocytes necessary for fertilization to occur. Ovarian stimulation, however, subjects the women to a high physical and psychological stress, thus being essential to improve ART and to find biomarkers of dysfunction and fertility. GSH is an important antioxidant, and is also used in detoxification reactions, catalysed by glutathione S-transferases (GST). In the present work, we have investigated the involvement of GST in follicular maturation. Patients with fertility problems and oocyte donors were recruited for the study. From each woman follicles at two stages of maturation were extracted at the preovulatory stage. Follicular fluid was separated from the oocyte by centrifugation and used as the enzyme source. GST activity was determined based on its conjugation with 3,4-dichloronitrobenzene and the assay was adapted to a 96-well microplate reader. The absorbance was represented against the incubation time and the curves were adjusted to linearity (R(2)>0.990). Results showed that in both donors and patients GST activity was significantly lower in mature oocytes compared to small ones. These results suggest that GST may play a role in the follicle maturation by detoxifying xenobiotics, thus contributing to the normal development of the oocyte. Supported by FIS/FEDER (PI11/02559), Gobierno Vasco (Dep. Educación, Universiades e Investigación, IT687-13), and UPV/EHU (CLUMBER UFI11/20 and PES13/58). The work was approved by the Ethics Committee of the UPV/EHU (CEISH/96/2011/RUIZLARREA), and performed according to the UPV/EHU and IVI-Bilbao agreement (Ref. 2012/01). PMID:26461371

  4. Proteomic and immunochemical characterization of glutathione transferase as a new allergen of the nematode Ascaris lumbricoides.

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    Nathalie Acevedo

    Full Text Available Helminth infections and allergy have evolutionary and clinical links. Infection with the nematode Ascaris lumbricoides induces IgE against several molecules including invertebrate pan-allergens. These antibodies influence the pathogenesis and diagnosis of allergy; therefore, studying parasitic and non-parasitic allergens is essential to understand both helminth immunity and allergy. Glutathione transferases (GSTs from cockroach and house dust mites are clinically relevant allergens and comparative studies between them and the GST from A. lumbricoides (GSTA are necessary to evaluate their allergenicity. We sought to analyze the allergenic potential of GSTA in connection with the IgE response to non-parasitic GSTs. IgE to purified GSTs from Ascaris (nGSTA and rGSTA, house dust mites (rDer p 8, nBlo t 8 and rBlo t 8, and cockroach (rBla g 5 was measured by ELISA in subjects from Cartagena, Colombia. Also, multidimensional proteomic approaches were used to study the extract of A. lumbricoides and investigate the existence of GST isoforms. We found that among asthmatics, the strength of IgE levels to GSTA was significantly higher than to mite and cockroach GSTs, and there was a strong positive correlation between IgE levels to these molecules. Specific IgE to GSTA was found in 13.2% of controls and 19.5% of asthmatics. In addition nGSTA induced wheal and flare in skin of sensitized asthmatics indicating that it might be of clinical relevance for some patients. Frequency and IgE levels to GSTA were higher in childhood and declined with age. At least six GST isoforms in A. lumbricoides bind human IgE. Four isoforms were the most abundant and several amino acid substitutions were found, mainly on the N-terminal domain. In conclusion, a new allergenic component of Ascaris has been discovered; it could have clinical impact in allergic patients and influence the diagnosis of mite and cockroach allergy in tropical environments.

  5. Glutathione S-Transferase Gene Polymorphisms and Treatment Outcome in Cervical Cancer Patients under Concomitant Chemoradiation.

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    Mohammad Abbas

    Full Text Available Cisplatin based concomitant chemoradiation (CRT is the standard treatment for locally advanced cervical cancer (CC. Glutathione S-transferase (GST, a phase II antioxidant enzyme is induced by oxidative stress generated by drugs and reactive oxidants. The present study was undertaken to evaluate the association of GSTM1, T1 and P1 polymorphisms with the outcome of CRT treatment in CC patients.A total of 227 cervical cancer patients with stages IIB-IIIB treated with the same chemoradiotherapy regimen were enrolled and genotyped for GSTM1, T1 and P1 gene polymorphisms by multiplex polymerase chain reaction (mPCR and PCR-restriction fragment length polymorphism (PCR-RFLP. Overall survival was evaluated using Kaplan-Meier survival function and Cox proportional hazards model. All data were analyzed using SPSS (version 21.0.Stratified analysis showed that GSTM1 null (M1- genotype was associated with a significantly better survival among patients with stage IIB cervical cancer (log-rank P = 0.004 than cases with stage IIIA/IIIB. Death and recurrence were significantly higher in patients with GSTM1 present genotype (M1+ (P = 0.037 and P = 0.003 respectively and those with M1- showed reduced hazard of death with an adjusted hazard ratio 'HR' of 0.47 (95% CI, 0.269-0.802, P = 0.006. Women with M1- genotype as well as in combination with GSTT1 null (T1-, GSTP1 (AG+GG and GSTT1 null/GSTP1 (AG+GG showed better survival and also reduced risk of death (HR = 0.31, P = 0.016; HR = 0.45, P = 0.013; HR = 0.31, P = 0.02 respectively.To the best of our knowledge, this is the first study to correlate the association of GSTM1, T1 and P1 gene polymorphisms with treatment outcome of CRT treated CC patients. Our results suggested that individuals with GSTM1 null genotype and in combination with GSTT1 null and GSTP1 (AG+GG had a survival advantage. Such genetic studies may provide prognostic information in CRT treated CC patients.

  6. Prognostic significance of glutathione S-transferase-pi in invasive breast cancer.

    Science.gov (United States)

    Huang, Jingxiang; Tan, Puay-Hoon; Thiyagarajan, Jayabaskar; Bay, Boon-Huat

    2003-06-01

    Glutathione S-transferase pi (GST-pi), a Phase II detoxification enzyme, has recently been implicated in protection against apoptosis. Expression of GST-pi and Bcl-2 protein, an established apoptosis marker, was analyzed by immunohistochemistry in 116 cases of infiltrative ductal breast carcinomas in Singapore women. The markers were correlated with apoptosis detected by the TUNEL method and clinico-pathological parameters. There were 67 (58%) GST-pi-positive breast tumors and 43 (37%) Bcl-2-positive tumors. In a large proportion of GST-pi-positive/Bcl-2-positive tumors, there was a distinct accumulation of the GST-pi enzyme within the nucleus of cancer cells when examined by double immunofluorescence labeling under confocal microscopy. GST-pi immunoreactivity was not significantly correlated with any of the traditional histologic factors known to influence prognosis, whereas Bcl-2 overexpression was associated with reduced size of primary tumor (P =.021) and positive estrogen receptor status (P =.001). Univariate analysis revealed that GST-pi-positive, Bcl-2-positive, and lower histological grade tumors had decreased levels of apoptosis (P =.024, P =.011, and P =.029, respectively). However, multivariate analysis showed that histological grade and Bcl-2, but not GST-pi, immunoreactivity were correlated with apoptotic status. The Kaplan-Meier disease-free survival curves showed a significant difference between GST-pi-positive and GST-pi-negative breast cancer cases (P =.002). Disease-free survival in patients with GST-pi-positive tumors was also worse than that in patients with GST-pi-negative tumors in the group who had adjuvant chemotherapy (P =.04). In patients who were lymph node positive, GST-pi immunopositivity was found to influence disease-free survival. Recurrence of tumors was also significantly affected by GST-pi immunoreactivity (relative risk of 8.1). The findings indicate that GST-pi-positive tumors are more aggressive and have a poorer prognosis than

  7. Chemical Reactivity Window Determines Prodrug Efficiency toward Glutathione Transferase Overexpressing Cancer Cells.

    Science.gov (United States)

    van Gisbergen, Marike W; Cebula, Marcus; Zhang, Jie; Ottosson-Wadlund, Astrid; Dubois, Ludwig; Lambin, Philippe; Tew, Kenneth D; Townsend, Danyelle M; Haenen, Guido R M M; Drittij-Reijnders, Marie-José; Saneyoshi, Hisao; Araki, Mika; Shishido, Yuko; Ito, Yoshihiro; Arnér, Elias S J; Abe, Hiroshi; Morgenstern, Ralf; Johansson, Katarina

    2016-06-01

    Glutathione transferases (GSTs) are often overexpressed in tumors and frequently correlated to bad prognosis and resistance against a number of different anticancer drugs. To selectively target these cells and to overcome this resistance we previously have developed prodrugs that are derivatives of existing anticancer drugs (e.g., doxorubicin) incorporating a sulfonamide moiety. When cleaved by GSTs, the prodrug releases the cytostatic moiety predominantly in GST overexpressing cells, thus sparing normal cells with moderate enzyme levels. By modifying the sulfonamide it is possible to control the rate of drug release and specifically target different GSTs. Here we show that the newly synthesized compounds, 4-acetyl-2-nitro-benzenesulfonyl etoposide (ANS-etoposide) and 4-acetyl-2-nitro-benzenesulfonyl doxorubicin (ANS-DOX), function as prodrugs for GSTA1 and MGST1 overexpressing cell lines. ANS-DOX, in particular, showed a desirable cytotoxic profile by inducing toxicity and DNA damage in a GST-dependent manner compared to control cells. Its moderate conversion of 500 nmol/min/mg, as catalyzed by GSTA1, seems hereby essential since the more reactive 2,4-dinitrobenzenesulfonyl doxorubicin (DNS-DOX) (14000 nmol/min/mg) did not display a preference for GSTA1 overexpressing cells. DNS-DOX, however, effectively killed GSTP1 (20 nmol/min/mg) and MGST1 (450 nmol/min/mg) overexpressing cells as did the less reactive 4-mononitrobenzenesulfonyl doxorubicin (MNS-DOX) in a MGST1-dependent manner (1.5 nmol/min/mg) as shown previously. Furthermore, we show that the mechanism of these prodrugs involves a reduction in GSH levels as well as inhibition of the redox regulatory enzyme thioredoxin reductase 1 (TrxR1) by virtue of their electrophilic sulfonamide moiety. TrxR1 is upregulated in many tumors and associated with resistance to chemotherapy and poor patient prognosis. Additionally, the prodrugs potentially acted as a general shuttle system for DOX, by overcoming resistance

  8. Proteomic analysis of glutathione S-transferase isoforms in mouse liver mitochondria

    Institute of Scientific and Technical Information of China (English)

    Hai-Dan Sun; Ya-Wei Ru; Dong-Juan Zhang; Song-Yue Yin; Liang Yin; Ying-Ying Xie; You-Fei Guan; Si-Qi Liu

    2012-01-01

    AIM:To survey glutathione (GSH) S-transferase (GST)isoforms in mitochondria and to reveal the isoforms' biological significance in diabetic mice.METHODS:The presence of GSTs in mouse liver mitochondria was systematically screened by two proteomic approaches,namely,GSH affinity chromatography/two dimensional electrophoresis (2DE/MALDI TOF/TOFMS) and SDS-PAGE/LC ESI MS/MS.The proteomic results were further confirmed by Western blotting using monoclonal antibodies against GSTs.To evaluate the liver mitochondrial GSTs quantitatively,calibration curves were generated by the loading amounts of individual recombinant GST protein vs the relative intensities elicited from the Western blotting.An extensive comparison of the liver mitochondrial GSTs was conducted between normal and db/db diabetic mice.Student's t test was adopted for the estimation of regression and significant difference.RESULTS:Using GSH affinity/2DF/MALDI TOF/TOF MS,three GSTs,namely,alpha3,mu1 and pi1,were identified; whereas five GSTs,alpha3,mu1,pi1,kappa1 and zeta1,were detected in mouse liver mitochondria using SDS-PAGE/LC ESI MS/MS,of these GSTs,GST kappa1 was reported as a specific mitochondrial GST.The R2 values of regression ranged between values of about 0.86 and 0.98,which were acceptable for the quantification.Based on the measurement of the GST abundances in liver mitochondria of normal and diabetic mice,the four GSTs,alpha3,kappa1,mu1 and zeta1,were found to be almost comparable between the two sets of animals,whereas,lower GST pi1 was detected in the diabetic mice compared with normal ones,the signal of Western blotting in control and db/ db diabetic mice liver mitochondria is 134.61 ± 53.84vs 99.74 ± 46.2,with P < 0.05.CONCLUSION:Our results indicate that GSTs exist widely in mitochondria and its abundances of mitochondrial GSTs might be tissue-dependent and disease-related.

  9. In vivo induction of phase II detoxifying enzymes, glutathione transferase and quinone reductase by citrus triterpenoids

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    Ahmad Hassan

    2010-09-01

    Full Text Available Abstract Background Several cell culture and animal studies demonstrated that citrus bioactive compounds have protective effects against certain types of cancer. Among several classes of citrus bioactive compounds, limonoids were reported to prevent different types of cancer. Furthermore, the structures of citrus limonoids were reported to influence the activity of phase II detoxifying enzymes. The purpose of the study was to evaluate how variations in the structures of citrus limonoids (namely nomilin, deacetyl nomilin, and isoobacunoic acid and a mixture of limonoids would influence phase II enzyme activity in excised tissues from a mouse model. Methods In the current study, defatted sour orange seed powder was extracted with ethyl acetate and subjected to silica gel chromatography. The HPLC, NMR and mass spectra were used to elucidate the purity and structure of compounds. Female A/J mice were treated with three limonoids and a mixture in order to evaluate their effect on phase II enzymes in four different tissues. Assays for glutathione S-transferase and NAD(PH: quinone reductase (QR were used to evaluate induction of phase II enzymatic activity. Results The highest induction of GST against 1-chloro-2,4-dinitrobenzene (CDNB was observed in stomach (whole, 58% by nomilin, followed by 25% isoobacunoic acid and 19% deacetyl nomilin. Deacetyl nomilin in intestine (small as well as liver significantly reduced GST activity against CDNB. Additionally isoobacunoic acid and the limonoid mixture in liver demonstrated a significant reduction of GST activity against CDNB. Nomilin significantly induced GST activity against 4-nitroquinoline 1-oxide (4NQO, intestine (280% and stomach (75% while deacetyl nomilin showed significant induction only in intestine (73%. Induction of GST activity was also observed in intestine (93% and stomach (45% treated with the limonoid mixture. Finally, a significant induction of NAD(PH: quinone reductase (QR activity was

  10. Comparative study on glutathione transferases of rat brain and testis under the stress of phenobarbitol and β-methylcholanthrene

    Institute of Scientific and Technical Information of China (English)

    THYAGARAJU K.; HEMAVATHI B.; VASUNDHARA K.; RAO A.D.; DEVI K.N.

    2005-01-01

    A comparative study was made on the tissue specific expression of glutathione transferases (GST) in brain and testis after exposure of rat to phenobarbitol (PB) and 3-methylcholanthrene (MC). Glutathione transferases, a family of multifunctional proteins are involved in intracellular transport processes and in detoxication of electrophilic xenobiotics by catalyzing reactions such as conjugation, isomerization, reduction and thiolysis. On purification, the yield of GST proteins by affinity chromatography was 39% in testis and 32% in brain. The affinity purified testis GSTs were resolved by chromatofocusing into six anionic and four cationic isozymes, and in brain glutathione transferases were resolved into four anionic and three cationic isozymes, suggesting the presence of multiple isozymes with Yc, Yb, Y3 and Yδ in both of them. In testis and brain, these isozymes at identical pI values showed variable functions with a battery of substrates and the cationic isozymes of brain and testis showed identical properties in CHP (cumene hydroperoxide) at pH values of above 7.0. Substrate specificity studies and immunoblot analysis of testis and brain proteins revealed that they play a predominant role in the detoxication of phenobarbitol or 3-methylcholanthrene. Expression of the isozymes in testis and brain on exposure to PB and MC indicated elevated subunit variation. In both testis and brain, Yδ ofπclass was expressed on PB treatment and Yc of α class and Y3 of μ class was expressed in MC treated testis and only Yc was predominantly expressed in MC treated brain. Thus these subunits expression is considered as markers for carcinogenesis and specific to chemical toxicity under phenobarbitol and 13-methylcholanthrene stress.

  11. The Fusarium oxysporum gnt2, encoding a putative N-acetylglucosamine transferase, is involved in cell wall architecture and virulence.

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    Loida López-Fernández

    Full Text Available With the aim to decipher the molecular dialogue and cross talk between Fusarium oxysporum f.sp. lycopersci and its host during infection and to understand the molecular bases that govern fungal pathogenicity, we analysed genes presumably encoding N-acetylglucosaminyl transferases, involved in glycosylation of glycoproteins, glycolipids, proteoglycans or small molecule acceptors in other microorganisms. In silico analysis revealed the existence of seven putative N-glycosyl transferase encoding genes (named gnt in F. oxysporum f.sp. lycopersici genome. gnt2 deletion mutants showed a dramatic reduction in virulence on both plant and animal hosts. Δgnt2 mutants had αalterations in cell wall properties related to terminal αor β-linked N-acetyl glucosamine. Mutant conidia and germlings also showed differences in structure and physicochemical surface properties. Conidial and hyphal aggregation differed between the mutant and wild type strains, in a pH independent manner. Transmission electron micrographs of germlings showed strong cell-to-cell adherence and the presence of an extracellular chemical matrix. Δgnt2 cell walls presented a significant reduction in N-linked oligosaccharides, suggesting the involvement of Gnt2 in N-glycosylation of cell wall proteins. Gnt2 was localized in Golgi-like sub-cellular compartments as determined by fluorescence microscopy of GFP::Gnt2 fusion protein after treatment with the antibiotic brefeldin A or by staining with fluorescent sphingolipid BODIPY-TR ceramide. Furthermore, density gradient ultracentrifugation allowed co-localization of GFP::Gnt2 fusion protein and Vps10p in subcellular fractions enriched in Golgi specific enzymatic activities. Our results suggest that N-acetylglucosaminyl transferases are key components for cell wall structure and influence interactions of F. oxysporum with both plant and animal hosts during pathogenicity.

  12. Phosphoethanolamine Transferase LptA in Haemophilus ducreyi Modifies Lipid A and Contributes to Human Defensin Resistance In Vitro.

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    Michael P Trombley

    Full Text Available Haemophilus ducreyi resists the cytotoxic effects of human antimicrobial peptides (APs, including α-defensins, β-defensins, and the cathelicidin LL-37. Resistance to LL-37, mediated by the sensitive to antimicrobial peptide (Sap transporter, is required for H. ducreyi virulence in humans. Cationic APs are attracted to the negatively charged bacterial cell surface. In other gram-negative bacteria, modification of lipopolysaccharide or lipooligosaccharide (LOS by the addition of positively charged moieties, such as phosphoethanolamine (PEA, confers AP resistance by means of electrostatic repulsion. H. ducreyi LOS has PEA modifications at two sites, and we identified three genes (lptA, ptdA, and ptdB in H. ducreyi with homology to a family of bacterial PEA transferases. We generated non-polar, unmarked mutants with deletions in one, two, or all three putative PEA transferase genes. The triple mutant was significantly more susceptible to both α- and β-defensins; complementation of all three genes restored parental levels of AP resistance. Deletion of all three PEA transferase genes also resulted in a significant increase in the negativity of the mutant cell surface. Mass spectrometric analysis revealed that LptA was required for PEA modification of lipid A; PtdA and PtdB did not affect PEA modification of LOS. In human inoculation experiments, the triple mutant was as virulent as its parent strain. While this is the first identified mechanism of resistance to α-defensins in H. ducreyi, our in vivo data suggest that resistance to cathelicidin LL-37 may be more important than defensin resistance to H. ducreyi pathogenesis.

  13. An Entamoeba histolytica ADP-ribosyl transferase from the diphtheria toxin family modifies the bacterial elongation factor Tu.

    Science.gov (United States)

    Avila, Eva E; Rodriguez, Orlando I; Marquez, Jaqueline A; Berghuis, Albert M

    2016-06-01

    ADP-ribosyl transferases are enzymes involved in the post-translational modification of proteins; they participate in multiple physiological processes, pathogenesis and host-pathogen interactions. Several reports have characterized the functions of these enzymes in viruses, prokaryotes and higher eukaryotes, but few studies have reported ADP-ribosyl transferases in lower eukaryotes, such as parasites. The locus EHI_155600 from Entamoeba histolytica encodes a hypothetical protein that possesses a domain from the ADP-ribosylation superfamily; this protein belongs to the diphtheria toxin family according to a homology model using poly-ADP-ribosyl polymerase 12 (PARP12 or ARTD12) as a template. The recombinant protein expressed in Escherichia coli exhibited in vitro ADP-ribosylation activity that was dependent on the time and temperature. Unlabeled βNAD(+), but not ADP-ribose, competed in the enzymatic reaction using biotin-βNAD(+) as the ADP-ribose donor. The recombinant enzyme, denominated EhToxin-like, auto-ADP-ribosylated and modified an acceptor from E. coli that was identified by MS/MS as the elongation factor Tu (EF-Tu). To the best of our knowledge, this is the first report to identify an ADP-ribosyl transferase from the diphtheria toxin family in a protozoan parasite. The known toxins from this family (i.e., the diphtheria toxin, the Pseudomonas aeruginosa toxin Exo-A, and Cholix from Vibrio cholerae) modify eukaryotic elongation factor two (eEF-2), whereas the amoeba EhToxin-like modified EF-Tu, which is another elongation factor involved in protein synthesis in bacteria and mitochondria. PMID:27234208

  14. Benzene exposure assessed by metabolite excretion in Estonian oil shale mineworkers: influence of glutathione s-transferase polymorphisms

    DEFF Research Database (Denmark)

    Sørensen, Mette; Poole, Jason; Autrup, Herman;

    2004-01-01

    oil shale mine were compared with the excretion in workers engaged in various production assignments above ground. In addition, possible modifying effects of genetic polymorphisms in glutathione S-transferases T1 (GSTT1), M1 (GSTM1), and P1 (GSTP1) on the excretion of S-PMA and t,t-MA were......,t-MA and S-PMA excretion were significantly higher in smokers compared with nonsmokers. Subjects carrying the GSTT1 wild-type excreted higher concentrations of S-PMA than subjects carrying the null genotype, suggesting that it is a key enzyme in the glutathione conjugation that leads to S-PMA. The results...

  15. Characterization and heterospecific expression of cDNA clones of genes in the maize GSH S-transferase multigene family.

    OpenAIRE

    Grove, G; Zarlengo, R P; Timmerman, K P; Li, N Q; Tam, M F; Tu, C P

    1988-01-01

    We have isolated from a constructed lambda gt11 expression library two classes of cDNA clones encoding the entire sequence of the maize GSH S-transferases GST I and GST III. Expression of a full-length GST I cDNA in E. coli resulted in the synthesis of enzymatically active maize GST I that is immunologically indistinguishable from the native GST I. Another GST I cDNA with a truncated N-terminal sequence is also active in heterospecific expression. Our GST III cDNA sequence differs from the ve...

  16. Glutathione-S-Transferase: A Minor Allergen in Birch Pollen due to Limited Release from Hydrated Pollen

    OpenAIRE

    Stephan Deifl; Christian Zwicker; Eva Vejvar; Claudia Kitzmüller; Gabriele Gadermaier; Birgit Nagl; Susanne Vrtala; Peter Briza; Zlabinger, Gerhard J.; Beatrice Jahn-Schmid; Fatima Ferreira; Barbara Bohle

    2014-01-01

    Background Recently, a protein homologous to glutathione-S-transferases (GST) was detected in prominent amounts in birch pollen by proteomic profiling. As members of the GST family are relevant allergens in mites, cockroach and fungi we investigated the allergenic relevance of GST from birch (bGST). Methodology bGST was expressed in Escherichia coli, purified and characterized by mass spectrometry. Sera from 217 birch pollen-allergic patients were tested for IgE-reactivity to bGST by ELISA. T...

  17. Cloning and expression of a cDNA encoding a maize glutathione-S-transferase in E. coli.

    OpenAIRE

    Moore, R. E.; Davies, M S; O'Connell, K M; Harding, E I; Wiegand, R C; Tiemeier, D C

    1986-01-01

    The isolation and characterization of a family of maize glutathione-S-transferases (GST's) has been described previously. These enzymes are designated GSTs I, II and III based on size, substrate specificity and responsiveness to safeners. GST III has been shown to act on the herbicide alachlor as well as the commonly used substrate 1-chloro-2,4-dinitrobenzene (CDNB). Clones were isolated from a maize cDNA library in lambda gt10. Three clones contained the entire coding region for GST III. The...

  18. Identification of a highly reactive sulphydryl group in human placental glutathione transferase by a site-directed fluorescent reagent.

    Science.gov (United States)

    Lo Bello, M; Petruzzelli, R; De Stefano, E; Tenedini, C; Barra, D; Federici, G

    1990-04-24

    A fluorescent maleimide derivative, N-(4-anilino-1-naphthyl) maleimide (ANM), a specific probe for thiol groups, reacted with human placental glutathione transferase (GST, EC 2.5.1.18), causing a complete inactivation of the enzyme in a few minutes. The modified enzyme was denatured, alkylated and digested with (L-1-tosylamide-2-phenylethyl chloromethyl ketone)-trypsin. The tryptic digest was analysed by HPLC and a fluorescent peptide was obtained. The sequence of this peptide allowed us, by a comparison with a well known primary structure, to assign the position 47 to the most reactive cysteine of GST enzyme. PMID:2335245

  19. Effects of high-intensity intermittent training on carnitine palmitoyl transferase activity in the gastrocnemius muscle of rats

    OpenAIRE

    L.C. Carnevali Jr; Eder, R.; F.S. Lira; Lima, W. P.; Gonçalves, D. C.; N.E. Zanchi; H. Nicastro; Lavoie, J.M.; M.C.L. Seelaender

    2012-01-01

    We examined the capacity of high-intensity intermittent training (HI-IT) to facilitate the delivery of lipids to enzymes responsible for oxidation, a task performed by the carnitine palmitoyl transferase (CPT) system in the rat gastrocnemius muscle. Male adult Wistar rats (160-250 g) were randomly distributed into 3 groups: sedentary (Sed, N = 5), HI-IT (N = 10), and moderate-intensity continuous training (MI-CT, N = 10). The trained groups were exercised for 8 weeks with a 10% (HI-IT) and a ...

  20. Does occupational exposure to solvents and pesticides in association with glutathione S-transferase A1, M1, P1, and T1 polymorphisms increase the risk of bladder cancer? The Belgrade case-control study.

    Directory of Open Access Journals (Sweden)

    Marija G Matic

    Full Text Available OBJECTIVE: We investigated the role of the glutathione S-transferase A1, M1, P1 and T1 gene polymorphisms and potential effect modification by occupational exposure to different chemicals in Serbian bladder cancer male patients. PATIENTS AND METHODS: A hospital-based case-control study of bladder cancer in men comprised 143 histologically confirmed cases and 114 age-matched male controls. Deletion polymorphism of glutathione S-transferase M1 and T1 was identified by polymerase chain reaction method. Single nucleotide polymorphism of glutathione S-transferase A1 and P1 was identified by restriction fragment length polymorphism method. As a measure of effect size, odds ratio (OR with corresponding 95% confidence interval (95%CI was calculated. RESULTS: The glutathione S-transferase A1, T1 and P1 genotypes did not contribute independently toward the risk of bladder cancer, while the glutathione S-transferase M1-null genotype was overrepresented among cases (OR = 2.1, 95% CI = 1.1-4.2, p = 0.032. The most pronounced effect regarding occupational exposure to solvents and glutathione S-transferase genotype on bladder cancer risk was observed for the low activity glutathione S-transferase A1 genotype (OR = 9.2, 95% CI = 2.4-34.7, p = 0.001. The glutathione S-transferase M1-null genotype also enhanced the risk of bladder cancer among subjects exposed to solvents (OR = 6,5, 95% CI = 2.1-19.7, p = 0.001. The risk of bladder cancer development was 5.3-fold elevated among glutathione S-transferase T1-active patients exposed to solvents in comparison with glutathione S-transferase T1-active unexposed patients (95% CI = 1.9-15.1, p = 0.002. Moreover, men with glutathione S-transferase T1-active genotype exposed to pesticides exhibited 4.5 times higher risk in comparison with unexposed glutathione S-transferase T1-active subjects (95% CI = 0.9-22.5, p = 0.067. CONCLUSION: Null or low-activity genotypes of the

  1. Crystallization and preliminary X-ray diffraction analysis of a glutathione S-transferase from Xylella fastidiosa

    International Nuclear Information System (INIS)

    Glutathione S-transferase from X. fastidiosa (xfGST) has been overexpressed in E. coli, purified and crystallized. Diffraction data were collected to 2.23 Å. Glutathione S-transferases (GSTs) form a group of multifunctional isoenzymes that catalyze the glutathione-dependent conjugation and reduction reactions involved in the cellular detoxification of xenobiotic and endobiotic compounds. GST from Xylella fastidiosa (xfGST) was overexpressed in Escherichia coli and purified by conventional affinity chromatography. In this study, the crystallization and preliminary X-ray analysis of xfGST is described. The purified protein was crystallized by the vapour-diffusion method, producing crystals that belonged to the triclinic space group P1. The unit-cell parameters were a = 47.73, b = 87.73, c = 90.74 Å, α = 63.45, β = 80.66, γ = 94.55°. xfGST crystals diffracted to 2.23 Å resolution on a rotating-anode X-ray source

  2. A simple colorimetric assay for specific detection of glutathione-S transferase activity associated with DDT resistance in mosquitoes.

    Directory of Open Access Journals (Sweden)

    Evangelia Morou

    Full Text Available BACKGROUND: Insecticide-based methods represent the most effective means of blocking the transmission of vector borne diseases. However, insecticide resistance poses a serious threat and there is a need for tools, such as diagnostic tests for resistance detection, that will improve the sustainability of control interventions. The development of such tools for metabolism-based resistance in mosquito vectors lags behind those for target site resistance mutations. METHODOLOGY/PRINCIPAL FINDINGS: We have developed and validated a simple colorimetric assay for the detection of Epsilon class Glutathione transferases (GST-based DDT resistance in mosquito species, such as Aedes aegypti, the major vector of dengue and yellow fever worldwide. The colorimetric assay is based on the specific alkyl transferase activity of Epsilon GSTs for the haloalkene substrate iodoethane, which produces a dark blue colour highly correlated with AaGSTE2-2-overexpression in individual mosquitoes. The colour can be measured visually and spectrophotometrically. CONCLUSIONS/SIGNIFICANCE: The novel assay is substantially more sensitive compared to the gold standard CDNB assay and allows the discrimination of moderate resistance phenotypes. We anticipate that it will have direct application in routine vector monitoring as a resistance indicator and possibly an important impact on disease vector control.

  3. Acute cadmium intoxication induces alpha-class glutathione S-transferase protein synthesis and enzyme activity in rat liver

    International Nuclear Information System (INIS)

    Acute cadmium intoxication affects glutathione S-transferase (GST) in rat liver. It has been found that 24 h after i.p. cadmium administration to rats, at a dose of 2.5 mg CdCl2 kg-1 body weight, the activity of this enzyme in liver cytosol increased by 40%. A less stimulatory effect persisted till 48 h and thereafter the enzyme activity normalized. Since, GST isoenzymes belong to different classes in mammalian tissues, we used quantitative immunoassays to verify which family of GST isoenzymes is influenced by this intoxication. Only alpha-class glutathione S-transferase (α-GST) proteins were detected in rat liver cytosol and their level increased by about 25%, 24 h after cadmium treatment. No pi-GST isoforms were found in liver cytosol from either normal or cadmium-treated rats. Co-administration of actinomycin D with cadmium normalized both the protein level and the activity of α-GST, suggesting that some effect occurs on enzyme transcription of these isoenzymes by this metal. On the other hand, it seems unlikely that the stimulatory effect is due to the high level of peroxides caused by lipid peroxidation, since Vitamin E administration strongly reduced the TBARS level, but did not cause any GST activity decrease

  4. Crystallization and preliminary X-ray diffraction analysis of a glutathione S-transferase from Xylella fastidiosa

    Energy Technology Data Exchange (ETDEWEB)

    Garcia, Wanius, E-mail: wanius@if.sc.usp.br [Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Travensolo, Regiane F. [Grupo de Bioanalítica, Microfabricação e Separações, Instituto de Química de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Rodrigues, Nathalia C.; Muniz, João R. C. [Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Caruso, Célia S. [Grupo de Bioanalítica, Microfabricação e Separações, Instituto de Química de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Lemos, Eliana G. M. [Laboratório de Bioquímica de Microrganismos e de Plantas, Departamento de Tecnologia, UNESP, Jaboticabal (Brazil); Araujo, Ana Paula U. [Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Carrilho, Emanuel, E-mail: wanius@if.sc.usp.br [Grupo de Bioanalítica, Microfabricação e Separações, Instituto de Química de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil)

    2008-02-01

    Glutathione S-transferase from X. fastidiosa (xfGST) has been overexpressed in E. coli, purified and crystallized. Diffraction data were collected to 2.23 Å. Glutathione S-transferases (GSTs) form a group of multifunctional isoenzymes that catalyze the glutathione-dependent conjugation and reduction reactions involved in the cellular detoxification of xenobiotic and endobiotic compounds. GST from Xylella fastidiosa (xfGST) was overexpressed in Escherichia coli and purified by conventional affinity chromatography. In this study, the crystallization and preliminary X-ray analysis of xfGST is described. The purified protein was crystallized by the vapour-diffusion method, producing crystals that belonged to the triclinic space group P1. The unit-cell parameters were a = 47.73, b = 87.73, c = 90.74 Å, α = 63.45, β = 80.66, γ = 94.55°. xfGST crystals diffracted to 2.23 Å resolution on a rotating-anode X-ray source.

  5. The role of human demographic history in determining the distribution and frequency of transferase-deficient galactosaemia mutations.

    LENUS (Irish Health Repository)

    Flanagan, J M

    2010-02-01

    Classical or transferase-deficient galactosaemia is an inherited metabolic disorder caused by mutation in the human Galactose-1-phosphate uridyl transferase (GALT) gene. Of some 170 causative mutations reported, fewer than 10% are observed in more than one geographic region or ethnic group. To better understand the population history of the common GALT mutations, we have established a haplotyping system for the GALT locus incorporating eight single nucleotide polymorphisms and three short tandem repeat markers. We analysed haplotypes associated with the three most frequent GALT gene mutations, Q188R, K285N and Duarte-2 (D2), and estimated their age. Haplotype diversity, in conjunction with measures of genetic diversity and of linkage disequilibrium, indicated that Q188R and K285N are European mutations. The Q188R mutation arose in central Europe within the last 20 000 years, with its observed east-west cline of increasing relative allele frequency possibly being due to population expansion during the re-colonization of Europe by Homo sapiens in the Mesolithic age. K285N was found to be a younger mutation that originated in Eastern Europe and is probably more geographically restricted as it arose after all major European population expansions. The D2 variant was found to be an ancient mutation that originated before the expansion of Homo sapiens out of Africa.

  6. Effect of municipal waste water effluent upon the expression of Glutathione S-transferase isoenzymes of brine shrimp Artemia.

    Science.gov (United States)

    Grammou, Athina; Papadimitriou, Chrisa; Samaras, Peter; Vasara, Eleni; Papadopoulos, Athanasios I

    2011-06-01

    Multiple isoenzymes of the detoxification enzyme family Glutathione S-transferase are expressed in the brine shrimp Artemia. The number of the major ones detected in crude extract by means of chromatofocusing varied between three and four, depending on the age. Two isoenzymes, one alkaline and one neutral (with corresponding isoelectric points of 8.5 and 7.2) appear to be dominant in all three developmental stages studied, (24, 48, and 72 h after hatching). Culturing Artemia for 48 h after hatching, in artificial sea water prepared by municipal wastewater effluent resulted to significant alterations of the isoenzyme profile. In comparison to organisms cultured for the same period of time in artificial sea water prepared by filtered tap water, the expression of the alkaline isoenzyme decreased by 62% while that of the neutral isoenzyme increased by 58%. Furthermore, the enzyme activity of the major isoenzyme of the acidic area increased by more than two folds. It is worth mentioning that although the specific activity of the total enzyme in the whole body homogenate was elevated, no statistically significant alteration of the Km value was observed. These findings suggest that study of the isoenzyme profile of Glutathione S-transferase may offer high sensitivity in detecting environmental pollution and needs to be further investigated. PMID:21429555

  7. Glutathione transferase (GST) as a candidate molecular-based biomarker for soil toxin exposure in the earthworm Lumbricus rubellus

    International Nuclear Information System (INIS)

    The earthworm Lumbricus rubellus (Hoffmeister, 1843) is a terrestrial pollution sentinel. Enzyme activity and transcription of phase II detoxification superfamily glutathione transferases (GST) is known to respond in earthworms after soil toxin exposure, suggesting GST as a candidate molecular-based pollution biomarker. This study combined sub-proteomics, bioinformatics and biochemical assay to characterise the L. rubellus GST complement as pre-requisite to initialise assessment of the applicability of GST as a biomarker. L. rubellus possesses a range of GSTs related to known classes, with evidence of tissue-specific synthesis. Two affinity-purified GSTs dominating GST protein synthesis (Sigma and Pi class) were cloned, expressed and characterised for enzyme activity with various substrates. Electrospray ionisation mass spectrometry (ESI-MS) and tandem mass spectrometry (MS/MS) following SDS-PAGE were superior in retaining subunit stability relative to two-dimensional gel electrophoresis (2-DE). This study provides greater understanding of Phase II detoxification GST superfamily status of an important environmental pollution sentinel organism. - This study currently provides the most comprehensive view of the Phase II detoxification enzyme superfamily of glutathione transferases within the important environmental pollution sentinel earthworm Lumbricus rubellus.

  8. In-house preparation of hydrogels for batch affinity purification of glutathione S-transferase tagged recombinant proteins

    Directory of Open Access Journals (Sweden)

    Buhrman Jason S

    2012-09-01

    Full Text Available Abstract Background Many branches of biomedical research find use for pure recombinant proteins for direct application or to study other molecules and pathways. Glutathione affinity purification is commonly used to isolate and purify glutathione S-transferase (GST-tagged fusion proteins from total cellular proteins in lysates. Although GST affinity materials are commercially available as glutathione immobilized on beaded agarose resins, few simple options for in-house production of those systems exist. Herein, we describe a novel method for the purification of GST-tagged recombinant proteins. Results Glutathione was conjugated to low molecular weight poly(ethylene glycol diacrylate (PEGDA via thiol-ene “click” chemistry. With our in-house prepared PEGDA:glutathione (PEGDA:GSH homogenates, we were able to purify a glutathione S-transferase (GST green fluorescent protein (GFP fusion protein (GST-GFP from the soluble fraction of E. coli lysate. Further, microspheres were formed from the PEGDA:GSH hydrogels and improved protein binding to a level comparable to purchased GSH-agarose beads. Conclusions GSH containing polymers might find use as in-house methods of protein purification. They exhibited similar ability to purify GST tagged proteins as purchased GSH agarose beads.

  9. Cantharidin Impedes Activity of Glutathione S-Transferase in the Midgut of Helicoverpa armigera Hübner.

    Science.gov (United States)

    Khan, Rashid Ahmed; Liu, Ji Yuan; Rashid, Maryam; Wang, Dun; Zhang, Ya Lin

    2013-01-01

    Previous investigations have implicated glutathione S-transferases (GSTs) as one of the major reasons for insecticide resistance. Therefore, effectiveness of new candidate compounds depends on their ability to inhibit GSTs to prevent metabolic detoxification by insects. Cantharidin, a terpenoid compound of insect origin, has been developed as a bio-pesticide in China, and proves highly toxic to a wide range of insects, especially lepidopteran. In the present study, we test cantharidin as a model compound for its toxicity, effects on the mRNA transcription of a model Helicoverpa armigera glutathione S-transferase gene (HaGST) and also for its putative inhibitory effect on the catalytic activity of GSTs, both in vivo and in vitro in Helicoverpa armigera, employing molecular and biochemical methods. Bioassay results showed that cantharidin was highly toxic to H. armigera. Real-time qPCR showed down-regulation of the HaGST at the mRNA transcript ranging from 2.5 to 12.5 folds while biochemical assays showed in vivo inhibition of GSTs in midgut and in vitro inhibition of rHaGST. Binding of cantharidin to HaGST was rationalized by homology and molecular docking simulations using a model GST (1PN9) as a template structure. Molecular docking simulations also confirmed accurate docking of the cantharidin molecule to the active site of HaGST impeding its catalytic activity. PMID:23528854

  10. Cantharidin Impedes Activity of Glutathione S-Transferase in the Midgut of Helicoverpa armigera Hübner

    Directory of Open Access Journals (Sweden)

    Ya Lin Zhang

    2013-03-01

    Full Text Available Previous investigations have implicated glutathione S-transferases (GSTs as one of the major reasons for insecticide resistance. Therefore, effectiveness of new candidate compounds depends on their ability to inhibit GSTs to prevent metabolic detoxification by insects. Cantharidin, a terpenoid compound of insect origin, has been developed as a bio-pesticide in China, and proves highly toxic to a wide range of insects, especially lepidopteran. In the present study, we test cantharidin as a model compound for its toxicity, effects on the mRNA transcription of a model Helicoverpa armigera glutathione S-transferase gene (HaGST and also for its putative inhibitory effect on the catalytic activity of GSTs, both in vivo and in vitro in Helicoverpa armigera, employing molecular and biochemical methods. Bioassay results showed that cantharidin was highly toxic to H. armigera. Real-time qPCR showed down-regulation of the HaGST at the mRNA transcript ranging from 2.5 to 12.5 folds while biochemical assays showed in vivo inhibition of GSTs in midgut and in vitro inhibition of rHaGST. Binding of cantharidin to HaGST was rationalized by homology and molecular docking simulations using a model GST (1PN9 as a template structure. Molecular docking simulations also confirmed accurate docking of the cantharidin molecule to the active site of HaGST impeding its catalytic activity.

  11. The relationship of glutathione-S-transferases copy number variation and indoor air pollution to symptoms and markers of respiratory disease

    DEFF Research Database (Denmark)

    Hersoug, Lars-Georg; Brasch-Andersen, Charlotte; Husemoen, Lise Lotte Nystrup;

    2012-01-01

    Introduction: Exposure to particulate matter (PM) may induce inflammation and oxidative stress in the airways. Carriers of null polymorphisms of glutathione S-transferases (GSTs), which detoxify reactive oxygen species, may be particularly susceptible to the effects of PM. Objectives: To investig......Introduction: Exposure to particulate matter (PM) may induce inflammation and oxidative stress in the airways. Carriers of null polymorphisms of glutathione S-transferases (GSTs), which detoxify reactive oxygen species, may be particularly susceptible to the effects of PM. Objectives: To....... The relationship of glutathione-S-transferases copy number variation and indoor air pollution to symptoms and markers of respiratory disease. Clin Respir J 2011; DOI:10.1111/j.1752-699X.2011.00258.x....

  12. UV-induced modifications in the peptidyl transferase loop of 23S rRNA dependent on binding of the streptogramin B antibiotic, pristinamycin IA

    DEFF Research Database (Denmark)

    Porse, B T; Kirillov, S V; Awayez, M J;

    1999-01-01

    The naturally occurring streptogramin B antibiotic, pristinamycin IA, which inhibits peptide elongation, can produce two modifications in 23S rRNA when bound to the Escherichia coli 70S ribosome and irradiated at 365 nm. Both drug-induced effects map to highly conserved nucleotides within the...... functionally important peptidyl transferase loop of 23S rRNA at positions m2A2503/psi2504 and G2061/A2062. The modification yields are influenced strongly, and differentially, by P-site-bound tRNA and strongly by some of the peptidyl transferase antibiotics tested, with chloramphenicol producing a shift in the...... latter modification to A2062/C2063. Pristinamycin IA can also produce a modification on binding to deproteinized, mature 23S rRNA, at position U2500/C2501. The same modification occurs on an approximately 37-nt fragment, encompassing positions approximately 2496-2532 of the peptidyl transferase loop that...

  13. Extração, purificação e avaliação da atividade da glutationa S-Transferase de fígado bovino Extraction of glutathione s-transferase from bovine liver

    OpenAIRE

    Maria Célia Lopes Torres; Nilda de Fátima Ferreira Soares; Jos�� Antônio Marques Pereira

    2006-01-01

    Considerando a ação detoxificante da enzima Glutationa S-Transferase (GST), importante contra o estresse oxidativo, câncer e outras doenças degenerativas, com este estudo, objetivou-se avaliar a atividade dessa enzima extraída de fígado bovino e avaliar a estabilidade em condições de refrigeração (5(0)C). O fígado bovino foi selecionado por ser matéria prima disponível comercialmente e de baixo custo. A extração foi realizada em quatro etapas (homogeneização/centrifugação, passagem em coluna ...

  14. Glutathione S-transferase in the midgut tissue of gypsy moth (Lymantria dispar) caterpillars exposed to dietary cadmium.

    Science.gov (United States)

    Vlahović, Milena; Ilijin, Larisa; Mrdaković, Marija; Todorović, Dajana; Matić, Dragana; Lazarević, Jelica; Mataruga, Vesna Perić

    2016-06-01

    Activity of glutathione S-transferase (GST) in midgut of gypsy moth caterpillars exposed to 10 and 30μg Cd/g dry food was examined. Based on the enzyme reaction through conjugation with glutathione, overall activity remained unaltered after acute and chronic treatment. No-observed-effect-concentration (10μg Cd/g dry food) significantly increased activity only after 3-day recovery following cadmium administration. Almost all comparisons of the indices of phenotypic plasticity revealed statistically significant differences. Despite the facts that GST has important role in xenobiotic biotransformation, our results indicate that this enzyme in insect midgut does not represent the key factor in cadmium detoxification. PMID:27084993

  15. ANALISIS GEN PENYANDI Schistosoma japonicum Gluthation s Transferase (SJ26GST DI DATARAN TINGGI LINDU, SULAWESI TENGAH INDONESIA

    Directory of Open Access Journals (Sweden)

    Anis Nurwidayati

    2015-01-01

    Full Text Available AbstractSchistosomiasis is only found at Napu and Lindu highland, Central Sulawesi in Indonesia. Schistosomiasis still as a public health problem, with its prevalence increase every year. The large scale by mass drug treatment using praziquantel has done to reduce the prevalence since 1980. To look for the possibility evidence of the development of resistance in S. japonicumto praziquantel in endemic areas by analysis of Schistosoma japonicumGluthation S Transferase (Sj26gst Coding Gene. Moleculer laboratory study was conducted to analyse the sequences of S. japonicumgluthation s transferase gene (Sj26GST. DNA was extracted from adult S. japonicumusing isopropanol. Sj26GST gene was amplified used gradient PCR. The PCR result then run with electrophoresis and viewed using gel-doc. The Sj26GST band was cut and purified using Gene Aid Purification kitand amplified by PCR cycle sequencing, and the product was sequenced using Abi PRISM 310 Genetic analyser. The gene sequences of Sj26GST analysis showed that the homology was very high between isolate from Indonesia and several isolates from China that known still susceptible to praziquantel.. The results indicate that there was no evidence for reduced susceptibility of S. japonicum to praziquantel despite its extensive use in the endemic areas of Napu and Lindu for more than 20 years.Keywords : Drug Resistance, Praziquantel, Schistosoma Japonicum, SchistosomiasisAbstrak Schistosomiasis di Indonesia ditemukan di Dataran Tinggi Lindu, Napu, dan Bada Sulawesi Tengah. Schistosomiasis masih menjadi masalah kesehatan dengan angka kasus yang berfluktuasi setiap tahun. Obat praziquantel telah digunakan secara massal sejak tahun 1980an, sehingga perlu dilakukan analisis kerentanan cacing Schistosoma japonicumterhadap praziquantel. Penelitian ini  bertujuan  untuk  mengidentifikasi  kerentanan  cacing S. japonicum terhadap praziquantel di Dataran Tinggi Lindu, dengan analisis secara molekuler gen penyandi

  16. The role of glutathione S-transferase P in signaling pathways and S-glutathionylation in cancer.

    Science.gov (United States)

    Tew, Kenneth D; Manevich, Yefim; Grek, Christina; Xiong, Ying; Uys, Joachim; Townsend, Danyelle M

    2011-07-15

    Glutathione S-transferase P is abundantly expressed in some mammalian tissues, particularly those associated with malignancies. While the enzyme can catalyze thioether bond formation between some electrophilic chemicals and GSH, novel nondetoxification functions are now ascribed to it. This review summarizes recent material that implicates GSTP in mediating S-glutathionylation of specific clusters of target proteins and in reactions that define a negative regulatory role in some kinase pathways through ligand or protein:protein interactions. It is becoming apparent that GSTP participates in the maintenance of cellular redox homeostasis through a number of convergent and divergent mechanisms. Moreover, drug platforms that have GSTP as a target have produced some interesting preclinical and clinical candidates. PMID:21558000

  17. Movement of the 3'-end of tRNA through the peptidyl transferase centre and its inhibition by antibiotics

    DEFF Research Database (Denmark)

    Kirillov, Stanislav; Porse, Bo Torben; Vester, Birthe;

    1997-01-01

    Determining how antibiotics inhibit ribosomal activity requires a detailed understanding of the interactions and relative movement of tRNA, mRNA and the ribosome. Recent models for the formation of hybrid tRNA binding sites during the elongation cycle have provided a basis for re-evaluating earlier...... experimental data and, especially, those relevant to substrate movements through the peptidyl transferase centre. With the exception of deacylated tRNA, which binds at the E-site, ribosomal interactions of the 3'-ends of the tRNA substrates generate only a small part of the total free energy of tRNA......-ribosome binding. Nevertheless, these relatively weak interactions determine the unidirectional movement of tRNAs through the ribosome and, moreover, they appear to be particularly susceptible to perturbation by antibiotics. Here we summarise current ideas relating particularly to the movement of the 3'-ends of tRNA...

  18. Mimicking insect communication: release and detection of pheromone, biosynthesized by an alcohol acetyl transferase immobilized in a microreactor.

    Directory of Open Access Journals (Sweden)

    Lourdes Muñoz

    Full Text Available Infochemical production, release and detection of (Z,E-9,11-tetradecadienyl acetate, the major component of the pheromone of the moth Spodoptera littoralis, is achieved in a novel microfluidic system designed to mimic the final step of the pheromone biosynthesis by immobilized recombinant alcohol acetyl transferase. The microfluidic system is part of an "artificial gland", i.e., a chemoemitter that comprises a microreactor connected to a microevaporator and is able to produce and release a pre-defined amount of the major component of the pheromone from the corresponding (Z,E-9,11-tetradecadienol. Performance of the entire chemoemitter has been assessed in electrophysiological and behavioral experiments. Electroantennographic depolarizations of the pheromone produced by the chemoemitter were ca. 40% relative to that evoked by the synthetic pheromone. In a wind tunnel, the pheromone released from the evaporator elicited on males a similar attraction behavior as 3 virgin females in most of the parameters considered.

  19. Garlic organosulfur compounds upregulate the expression of the pi class of glutathione S-transferase in rat primary hepatocytes.

    Science.gov (United States)

    Tsai, Chia-Wen; Yang, Jaw-Ji; Chen, Haw-Wen; Sheen, Lee-Yan; Lii, Chong-Kuei

    2005-11-01

    The chemopreventive property of garlic is related in part to its induction of phase II detoxification enzymes. In the present study, we investigated the modulatory effect of 3 garlic organosulfur compounds, i.e., diallyl sulfide (DAS), diallyl disulfide (DADS), and diallyl trisulfide (DATS), which differ in their number of sulfur atoms, on the gene expression of the pi class of glutathione S-transferase (GSTP). Hepatocytes isolated from male Sprague-Dawley rats were cultured with 50-200 micromol/L of DAS, DADS, or DATS for 24 h. DADS and DATS increased GST activity toward ethacrynic acid by 40 and 66%, respectively (P effectiveness of 3 garlic allyl sulfides on GSTP expression was related to the number of sulfur atoms in the molecules, and GPE I was responsible for this upregulation. PMID:16251611

  20. Transcriptome-wide identification and expression analysis of glutathione S-transferase genes involved in flavonoids accumulation in Dracaena cambodiana.

    Science.gov (United States)

    Zhu, Jia-Hong; Li, Hui-Liang; Guo, Dong; Wang, Ying; Dai, Hao-Fu; Mei, Wen-Li; Peng, Shi-Qing

    2016-07-01

    Dragon's blood is a traditional medicine widely used in the world, and the main components of which are flavonoids. However, little is known about its formation mechanism. Previous studies indicate that plant glutathione S-transferase (GST) genes are involved in transportation of flavonoids from cytosolic synthesis to vacuolar accumulation. In this study, 20 Dracaena cambodiana GST genes (DcGSTs) were identified based on transcriptome database. Phylogenetic analysis revealed that 20 DcGSTs belonged to seven different classes. Tissue-specific expression analysis suggested that DcGSTs displayed differential expressions either in their transcript abundance or expression patterns under normal growth conditions. The transcript profiles of three DcGSTs in response to the inducer of dragon's blood were strongly correlated with flavonoids biosynthetic genes, consistent with dragon's blood accumulation. Our survey provides useful information for future studies on GST genes involved in dragon's blood formation in D. cambodiana. PMID:27208821

  1. Inductoin of Radioresistance by Overexpression of Glutathione S-Transferase K1 (hGSTK1) in MCF-7 Cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jae Chul [Kyungpook National University College of Medicine, Taegu (Korea, Republic of); Shin, Sei One [Yeungnam University College of Medicine, Taegu (Korea, Republic of)

    2001-12-15

    Purpose : This study was conducted to assess the effects of x-irradiation on the expression of the novel glutathione S-transferase K1 gene. Materials and methods : Human glutathione S-transferase K1 (hGSTK1) DNA was purified and ligated to a pcDNA3.1/Myc-His(+) vector for the overexpression of hGSTK1 gene. MCF-7 cells were transfected with or without the recombinant hGSTK1 gene, and irradiated with 6 MV x-ray. After incubation of 14 days, cell survival was measured and compared. The expression of hGSTK1 and the effect of x- irradiation on hGSTK1 expression were also estimated in MCF-7 cells transfected with or without the hGSTK1 gene by RT-PCR. Results : Following 2 to 12 Gy of x-irradiation, the cell survivals were higher in the MCF-7 cells transfected with the hGSTK1 gene than in those without transfection. Despite the higher cell survival in the hGSTK1-transfected cells, RT-PCR for hGSTK1 mRNA revealed no significant differences according to radiation dose, fractionation, and time after irradiation. Conclusion : The MCF-7 cells transfected with the hGSTK1 gene showed higher cell survival than those without transfection of the gene. The hGSTK1 gene might be associated with the radiosensitivity of MCF-7 cell line and further analysis should be needed.

  2. Effects of gestational and overt diabetes on human placental cytochromes P450 and glutathione S-transferase.

    Science.gov (United States)

    McRobie, D J; Glover, D D; Tracy, T S

    1998-04-01

    The placenta possesses the ability to metabolize a number of xenobiotics and endogenous compounds by processes similar to those seen in the liver. Animal and in vivo studies have observed that the presence of diabetes alters the expression of hepatic metabolizing enzymes (cytochrome P450 and glutathione S-transferase); however, it is unknown whether similar alterations occur in the human placenta. To evaluate whether diabetes has any effect of placental xenobiotic metabolizing activity, the catalytic activities of 7-ethoxyresorufin O-deethylation (EROD, CYP1A1), chlorzoxazone 6-hydroxylation (CYP2E1), dextromethorphan N-demethylation (CYP3A4), dextromethorphan O-demethylation (CYP2D6), and 1-chloro-2, 4-dinitrobenzene (CDNB) conjugation with glutathione (glutathione S-transferase, GST) from placentas of diet (class A1) and insulin-dependent (class A2) gestational diabetics and overt diabetics were compared with matched controls. EROD activity (CYP1A1) ranged from 0.29 to 2.67 pmol/min/mg protein. However, no differences were observed among overt or gestational diabetics and their respective matched controls. CDNB conjugation (GST) ranged from 0.275 to 1.65 units/min/mg protein. In contrast to that observed with CYP1A1, a small but statistically significant reduction in GST activity was noted in overt diabetics as compared with their matched controls and gestational diabetics. CYP2E1, 2D6, and 3A4 enzymatic activities were not detected in human placental tissue. GST protein was detectable in all tissues studied, but no CYP protein could be detected in any of the tissues. Thus, it seems that pregnant women with overt diabetes have reduced GST activity in the placenta, which could potentially result in the exposure of the fetus to harmful electrophiles. However, the full clinical significance of this finding remains to be elucidated. PMID:9531526

  3. Expression, purification, crystallization and structure determination of two glutathione S-transferase-like proteins from Shewanella oneidensis

    International Nuclear Information System (INIS)

    The production and purification of recombinant SoGST3 and SoGST6, two GST-like proteins from S. oneidensis, are reported and preliminary crystallographic studies of crystals of the recombinant enzymes are presented. Genome analysis of Shewanella oneidensis, a Gram-negative bacterium with an unusual repertoire of respiratory and redox capabilities, revealed the presence of six glutathione S-transferase-like genes (sogst1–sogst6). Glutathione S-transferases (GSTs; EC 2.5.1.18) are found in all kingdoms of life and are involved in phase II detoxification processes by catalyzing the nucleophilic attack of reduced glutathione on diverse electrophilic substrates, thereby decreasing their reactivity. Structure–function studies of prokaryotic GST-like proteins are surprisingly underrepresented in the scientific literature when compared with eukaryotic GSTs. Here, the production and purification of recombinant SoGST3 (SO-1576) and SoGST6 (SO-4697), two of the six GST-like proteins in S. oneidensis, are reported and preliminary crystallographic studies of crystals of the recombinant enzymes are presented. SoGST3 was crystallized in two different crystal forms in the presence of GSH and DTT that diffracted to high resolution: a primitive trigonal form in space group P31 that exhibited merohedral twinning with a high twin fraction and a primitive monoclinic form in space group P21. SoGST6 yielded primitive orthorhombic crystals in space group P212121 from which diffraction data could be collected to medium resolution after application of cryo-annealing protocols. Crystal structures of both SoGST3 and SoGST6 have been determined based on marginal search models by maximum-likelihood molecular replacement as implemented in the program Phaser

  4. Differential substrate specificity and kinetic behavior of Escherichia coli YfdW and Oxalobacter formigenes formyl coenzyme A transferase.

    Science.gov (United States)

    Toyota, Cory G; Berthold, Catrine L; Gruez, Arnaud; Jónsson, Stefán; Lindqvist, Ylva; Cambillau, Christian; Richards, Nigel G J

    2008-04-01

    The yfdXWUVE operon appears to encode proteins that enhance the ability of Escherichia coli MG1655 to survive under acidic conditions. Although the molecular mechanisms underlying this phenotypic behavior remain to be elucidated, findings from structural genomic studies have shown that the structure of YfdW, the protein encoded by the yfdW gene, is homologous to that of the enzyme that mediates oxalate catabolism in the obligate anaerobe Oxalobacter formigenes, O. formigenes formyl coenzyme A transferase (FRC). We now report the first detailed examination of the steady-state kinetic behavior and substrate specificity of recombinant, wild-type YfdW. Our studies confirm that YfdW is a formyl coenzyme A (formyl-CoA) transferase, and YfdW appears to be more stringent than the corresponding enzyme (FRC) in Oxalobacter in employing formyl-CoA and oxalate as substrates. We also report the effects of replacing Trp-48 in the FRC active site with the glutamine residue that occupies an equivalent position in the E. coli protein. The results of these experiments show that Trp-48 precludes oxalate binding to a site that mediates substrate inhibition for YfdW. In addition, the replacement of Trp-48 by Gln-48 yields an FRC variant for which oxalate-dependent substrate inhibition is modified to resemble that seen for YfdW. Our findings illustrate the utility of structural homology in assigning enzyme function and raise the question of whether oxalate catabolism takes place in E. coli upon the up-regulation of the yfdXWUVE operon under acidic conditions. PMID:18245280

  5. Synergistic and independent actions of multiple terminal nucleotidyl transferases in the 3' tailing of small RNAs in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Xiaoyan Wang

    2015-04-01

    Full Text Available All types of small RNAs in plants, piwi-interacting RNAs (piRNAs in animals and a subset of siRNAs in Drosophila and C. elegans are subject to HEN1 mediated 3' terminal 2'-O-methylation. This modification plays a pivotal role in protecting small RNAs from 3' uridylation, trimming and degradation. In Arabidopsis, HESO1 is a major enzyme that uridylates small RNAs to trigger their degradation. However, U-tail is still present in null hen1 heso1 mutants, suggesting the existence of (an enzymatic activities redundant with HESO1. Here, we report that UTP: RNA uridylyltransferase (URT1 is a functional paralog of HESO1. URT1 interacts with AGO1 and plays a predominant role in miRNA uridylation when HESO1 is absent. Uridylation of miRNA is globally abolished in a hen1 heso1 urt1 triple mutant, accompanied by an extensive increase of 3'-to-5' trimming. In contrast, disruption of URT1 appears not to affect the heterochromatic siRNA uridylation. This indicates the involvement of additional nucleotidyl transferases in the siRNA pathway. Analysis of miRNA tailings in the hen1 heso1 urt1 triple mutant also reveals the existence of previously unknown enzymatic activities that can add non-uridine nucleotides. Importantly, we show HESO1 may also act redundantly with URT1 in miRNA uridylation when HEN1 is fully competent. Taken together, our data not only reveal a synergistic action of HESO1 and URT1 in the 3' uridylation of miRNAs, but also independent activities of multiple terminal nucleotidyl transferases in the 3' tailing of small RNAs and an antagonistic relationship between uridylation and trimming. Our results may provide further insight into the mechanisms of small RNA 3' end modification and stability control.

  6. Novel functional association of rat testicular membrane-associated cytosolic glutathione S transferases and cyclooxygenase in vitro

    Institute of Scientific and Technical Information of China (English)

    S. Neeraja; B. Ramakrishna; A. S. Sreenath; G. V. Reddy; P. R. K. Reddy; P. Reddanna

    2005-01-01

    Aim: To analyze the role of cytosolic glutathione S-transferases (cGSTs) and membrane-associated cytosolic GSTs (macGSTs) in prostaglandin biosynthesis and to evaluate the possible interaction between glutathione S-transferases (GSTs) and cyclooxygenase (COX) in vitro. Methods: SDS-PAGE analysis was undertaken for characterization of GSTs, thin layer chromatography (TLC) to monitor the effect of GSTs on prostaglandin biosynthesis from arachidonic acid (AA) and spectrophotometric assays were done for measuring activity levels of COX and GSTs. Results:SDS-PAGE analysis indicates that macGSTs have molecular weights in the range of 25-28 kDa. In a coupled assay involving GSTs, arachidonic acid and cyclooxygenase-1, rat testicular macGSTs produced prostaglandin E2 and F2α,while the cGSTs caused the generation of prostaglandin D2, E2 and F2α. In vitro interaction studies on GSTs and COX at the protein level have shown dose-dependent inhibition of COX activity by macGSTs and vice versa. This effect,however, is not seen with cGSTs. The inhibitory effect of COX on macGST activity was relieved with increasing concentrations of reduced glutathione (GSH) but not with 1-chloro 2,4-dinitrobenzene (CDNB). The inhibition of COX by macGSTs, on the other hand, was potentiated by glutathione. Conclusion: We isolated and purified macGSTs and cGSTs from rat testis and analyzed their involvement in prostaglandin biosynthesis. These studies reveal a reversible functional interaction between macGSTs and COX in vitro, with possible interactions between them at the GSH binding site of macGSTs.

  7. Sites of interaction of streptogramin A and B antibiotics in the peptidyl transferase loop of 23 S rRNA and the synergism of their inhibitory mechanisms

    DEFF Research Database (Denmark)

    Porse, B T; Garrett, R A

    1999-01-01

    nucleotides in the peptidyl transferase loop of 23 S rRNA, including the two mutated nucleotides. An rRNA footprinting study, performed both in vivo and in vitro, on the A and B components complexed to Bacillus megaterium ribosomes, indicated that similar drug-induced effects occur on free ribosomes and...

  8. GAMMA-GLUTAMYL TRANSFERASE (GGT) ACTIVITY AND BIOCHEMICAL CHARACTERIZATION OF RAT VISCERAL YOLK-SAC DURING GESTATION WITH OR WITHOUT TRYPAN BLUE EXPOSURE

    Science.gov (United States)

    Yolk-sacs from untreated Sprague Dawley rat conceptuses were removed on days 9-18 of gestation and examined for gamma-glutamyl transferase (GGT), alkaline phosphatase (AP), lactate dehydrogenase (LDH) and glutamic-oxaloacetic transaminase (GOT) activities. All enzyme activities w...

  9. Conjugation of isoprene monoepoxides with glutathione, catalyzed by α, μ, π and θ-class glutathione S-transferases of rat and man

    NARCIS (Netherlands)

    Bogaards, J.J.P.; Venekamp, J.C.; Salmon, F.G.C.; Bladeren, P.J. van

    1999-01-01

    In the present study, the enzymatic conjugation of the isoprene monoepoxides 3,4 epoxy-3-methyl-1-butene (EPOX-I) and 3,4-epoxy-2-methyl-1-butene (EPOX-II) with glutathione was investigated, using purified glutathione S-transferases (GSTs) of the α, μ, π and θ-class of rat and man. HPLC analysis of

  10. Pooled analysis and meta-analysis of glutathione S-transferase M1 and bladder cancer: A HuGE review

    DEFF Research Database (Denmark)

    Engel, Lawrence S.; Taioli, Emanuela; Pfeiffer, Ruth;

    2002-01-01

    Smoking is a known risk factor for bladder cancer. The product of the GSTM1 gene, glutathione S-transferase M1 (GSTM1), is involved in the detoxification of polycyclic aromatic hydrocarbons found in tobacco smoke; a homozygous deletion of this gene in approximately 50% of Caucasians and Asians...

  11. Copy number variation in glutathione S-transferases M1 and T1 and ischemic vascular disease: four studies and meta-analyses

    DEFF Research Database (Denmark)

    Nørskov, Marianne S; Frikke-Schmidt, Ruth; Loft, Steffen;

    2011-01-01

    Glutathione S-transferases (GSTs) M1 and T1 detoxify products of oxidative stress and may protect against atherosclerosis and ischemic vascular disease (IVD). We tested the hypothesis that copy number variation (CNV) in GSTM1 and GSTT1 genes, known to be associated with stepwise decreases in...

  12. The role of the glutathione S-transferase genes GSTT1, GSTM1, and GSTP1 in acetaminophen-poisoned patients

    DEFF Research Database (Denmark)

    Buchard, Anders; Eefsen, Martin; Semb, Synne;

    2012-01-01

    The aim of this study was to assess if genetic variants in the glutathione-S-transferase genes GST-T1, M1, and P1 reflect risk factors in acetaminophen (APAP)-poisoned patients assessed by investigation of the relation to prothrombin time (PT), which is a sensitive marker of survival in these...

  13. Characterization of Alcohol Acyl Transferase and 1-Aminocyclopropane-1-Carboxylate Synthase Gene Expression and Volatile Compound Emission during Apple Fruit Development and Ripening

    Science.gov (United States)

    Alcohol acyl transferase (AAT) catalyzes the last step of volatile ester biosynthesis, and in this study, expression of four apple AAT genes was investigated in the peel of two apple cultivars with relatively high (‘Golden Delicious’) or low (‘Granny Smith’) volatile ester production. All four AAT ...

  14. Pleiotropic effects of polymorphism of the gene diacylglycerol-O-transferase 1 (DGAT1) in the mammary gland tissue of dairy cows

    NARCIS (Netherlands)

    Mach Casellas, N.; Blum, Y.; Bannink, A.; Causeur, D.; Houee-Bigot, M.; Lagarrigue, S.; Smits, M.A.

    2012-01-01

    Microarray analysis was used to identify genes whose expression in the mammary gland of Holstein-Friesian dairy cows was affected by the nonconservative Ala to Lys amino acid substitution at position 232 in exon VIII of the diacylglycerol-O-transferase 1 (DGAT1) gene. Mammary gland biopsies of 9 hom

  15. No elevation of glutathione S-transferase-a1-1 by amiodarone loading in intensive care unit patients with atrial fibrillation.

    NARCIS (Netherlands)

    Hilkens, M.; Pickkers, P.; Peters, W.H.M.; Hoeven, J.G. van der

    2009-01-01

    Hepatocellular toxicity is a putative side-effect of amiodarone. The hepatic detoxification enzyme glutathione S-transferase-A1-1 (GSTA1-1) is a sensitive indicator of hepatocellular damage. We investigated the occurrence of subclinical liver injury, as measured by plasma GSTA1-1 in intensive care u

  16. Glutathione S-transferase phenotypes in relation to genetic variation and fruit and vegetable consumption in an endoscopy-based population.

    NARCIS (Netherlands)

    Tijhuis, M.J.; Visker, M.H.P.W.; Aarts, J.; Peters, W.H.M.; Roelofs, H.M.J.; Camp, L.O. den; Rietjens, I.M.C.M.; Boerboom, A.M.A.; Nagengast, F.M.; Kok, F.J.; Kampman, E.

    2007-01-01

    High glutathione S-transferase (GST) activity may contribute to colorectal cancer prevention. Functional polymorphisms are known in the GSTM1, GSTT1, GSTA1 and GSTP1 genes. The influence of these GST polymorphisms and recent fruit and vegetable consumption on GST levels and activity has not been inv

  17. GLUTATHIONE-S-TRANSFERASE ACTIVITY AND ISOENZYME COMPOSITION IN BENIGN OVARIAN-TUMORS, UNTREATED MALIGNANT OVARIAN-TUMORS, AND MALIGNANT OVARIAN-TUMORS AFTER PLATINUM CYCLOPHOSPHAMIDE CHEMOTHERAPY

    NARCIS (Netherlands)

    VANDERZEE, AGJ; VANOMMEN, B; MEIJER, C; HOLLEMA, H; VANBLADEREN, PJ; DEVRIES, EGE

    1992-01-01

    Glutathione S-transferase (GST) isoenzyme composition, isoenzyme quantities and enzymatic activity were investigated in benign (n = 4) ovarian tumours and malignant ovarian tumours, before (n = 20) and after (n = 16) chemotherapy. Enzymatic activity of GST in cytosols was measured by determining 1-c

  18. Targeted cytosine deaminase-uracil phosphoribosyl transferase suicide gene therapy induces small cell lung cancer-specific cytotoxicity and tumor growth delay

    DEFF Research Database (Denmark)

    Christensen, Camilla L; Gjetting, Torben; Poulsen, Thomas Tuxen;

    2010-01-01

    deaminase (YCD) gene alone or fused with the yeast uracil phosphoribosyl transferase (YUPRT) gene followed by administration of 5-fluorocytosine (5-FC) prodrug. Experimental design: The YCD gene or the YCD-YUPRT gene was placed under regulation of the SCLC-specific promoter insulinoma-associated 1 (INSM1...

  19. Purification of a glutathione S-transferase and a glutathione conjugate-specific dehydrogenase involved in isoprene metabolism in Rhodococcus sp. strain AD45

    NARCIS (Netherlands)

    Hylckama Vlieg, Johan E.T. van; Kingma, Jaap; Kruizinga, Wim; Janssen, Dick B.

    1999-01-01

    A glutathione S transferase (GST) with activity toward 1,2-eposy-2-methyl-3-butene (isoprene monoxide) and cis-1,2-dichloroepoxyethane was purified from the isoprene-utilizing bacterium Rhodococcus sp. strain AD45, The homodimeric enzyme (two subunits of 27 kDa each) catalyzed the glutathione (GSH)-

  20. Correlation of Rutin Accumulation with 3-O-Glucosyl Transferase and Phenylalanine Ammonia-lyase Activities During the Ripening of Tomato Fruit

    NARCIS (Netherlands)

    Capanoglu, E.; Beekwilder, J.; Matros, A.; Boyacioglu, D.; Hall, R.D.; Mock, H.P.

    2012-01-01

    In tomato, the predominant flavonoid is quercetin-3-rutinoside (rutin). In this study, we aim to investigate the phenylalanine ammonia-lyase (PAL) and the quercetin-3-O-glucosyl transferase (3-GT) reactions in the formation of rutin during tomato fruit ripening. Tomatoes of the Moneymaker variety at

  1. High-throughput genotyping of copy number variation in glutathione S-transferases M1 and T1 using real-time PCR in 20,687 individuals

    DEFF Research Database (Denmark)

    Norskov, M.S.; Frikke-Schmidt, R.; Loft, S.;

    2009-01-01

    OBJECTIVES: Characteristic for the genes encoding glutathione S-transferase (GST) M1 and GSTT1 is a null allele, suggested to increase susceptibility to chronic diseases. We report an optimized method for the determination of copy number variation (CNV) in GST genes. DESIGN AND METHODS: Real-time...

  2. Succinyl-CoA:acetoacetate transferase deficiency : identification of a new patient with a neonatal onset and review of the literature

    NARCIS (Netherlands)

    Niezen-Koning, K E; Wanders, R J; Ruiter, J P; Ijlst, L; Visser, G; Reitsma-Bierens, W C; Heijmans, Hugo; Reijngoud, D J; Smit, G P

    1997-01-01

    UNLABELLED: We describe the clinical symptoms and biochemical findings of a patient with succinyl-CoA:acetoacetate transferase deficiency who presented in the neonatal period and review the current literature on this subject. Our patient was initially suspected to have distal renal tubular acidosis,

  3. Succinyl-CoA : acetoacetate transferase deficiency: identification of a new patient with a neonatal onset and review of the literature

    NARCIS (Netherlands)

    NiezenKoning, KE; Ijlst, L; Visser, G; ReitsmaBierens, WCC; Heymans, HSA; Reijngoud, DJ; Smit, GPA; Ruiter, Jos P. N.

    1997-01-01

    We describe the clinical symptoms and biochemical findings of a patient with succinyl-CoA:acetoacetate transferase deficiency who presented in the neonatal period and review the current literature on this subject. Our patient was initially suspected to have distal renal tubular acidosis, and subsequ

  4. Cold sensitivity in rice (Oryza sativa L.) is strongly correlated with a naturally occurring I99V mutation in the multifunctional glutathione transferase isoenzyme GSTZ2

    Science.gov (United States)

    GSTZs (zeta class glutathione transferases) belong to a highly conserved subfamily of soluble GSTs found in species ranging from fungi and plants to animals. GSTZ is identical to MAAI (maleylacetoacetate isomerase), which functions in tyrosine catabolism by catalyzing the isomerization of MAA (maley...

  5. Identification of a novel UDP-N-acetylglucosamine enolpyruvyl transferase (MurA) from Vibrio fischeri that confers high fosfomycin resistance in Escherichia coli

    Digital Repository Service at National Institute of Oceanography (India)

    Kumar, S.; Parvathi, A; Hernandez, R.L.; Cadle, K.M.; Varela, M.F.

    MurA [UDP-N-acetylglucosamine (UDP-NAG) enolpyruvyl transferase] is a key enzyme involved in bacterial cell wall peptidoglycan synthesis and a target for the antimicrobial agent fosfomycin, a structural analog of the MurA substrate phosphoenol...

  6. Addition of Rice Bran Arabinoxylan to Curcumin Therapy May Be of Benefit to Patients With Early-Stage B-Cell Lymphoid Malignancies (Monoclonal Gammopathy of Undetermined Significance, Smoldering Multiple Myeloma, or Stage 0/1 Chronic Lymphocytic Leukemia): A Preliminary Clinical Study.

    Science.gov (United States)

    Golombick, Terry; Diamond, Terrence H; Manoharan, Arumugam; Ramakrishna, Rajeev

    2016-06-01

    Hypothesis Prior studies on patients with early B-cell lymphoid malignancies suggest that early intervention with curcumin may lead to delay in progressive disease and prolonged survival. These patients are characterized by increased susceptibility to infections. Rice bran arabinoxylan (Ribraxx) has been shown to have immunostimulatory, anti-inflammatory, and proapoptotic effects. We postulated that addition of Ribraxx to curcumin therapy may be of benefit. Study design Monoclonal gammopathy of undetermined significance (MGUS)/smoldering multiple myeloma (SMM) or stage 0/1 chronic lymphocytic leukemia (CLL) patients who had been on oral curcumin therapy for a period of 6 months or more were administered both curcumin (as Curcuforte) and Ribraxx. Methods Ten MGUS/SMM patients and 10 patients with stage 0/1 CLL were administered 6 g of curcumin and 2 g Ribraxx daily. Blood samples were collected at baseline and at 2-month intervals for a period of 6 months, and various markers were monitored. MGUS/SMM patients included full blood count (FBC); paraprotein; free light chains/ratio; C-reactive protein (CRP)and erythrocyte sedimentation rate (ESR); B2 microglobulin and immunological markers. Markers monitored for stage 0/1 CLL were FBC, CRP and ESR, and immunological markers. Results Of 10 MGUS/SMM patients,5 (50%) were neutropenic at baseline, and the Curcuforte/Ribraxx combination therapy showed an increased neutrophil count, varying between 10% and 90% among 8 of the 10 (80%) MGUS/SMM patients. An additional benefit of the combination therapy was the potent effect in reducing the raised ESR in 4 (44%) of the MGUS/SMM patients. Conclusion Addition of Ribraxx to curcumin therapy may be of benefit to patients with early-stage B-cell lymphoid malignancies. PMID:27154182

  7. Orotate phosphoribosyl transferase MoPyr5 is involved in uridine 5'-phosphate synthesis and pathogenesis of Magnaporthe oryzae.

    Science.gov (United States)

    Qi, Zhongqiang; Liu, Muxing; Dong, Yanhan; Yang, Jie; Zhang, Haifeng; Zheng, Xiaobo; Zhang, Zhengguang

    2016-04-01

    Orotate phosphoribosyl transferase (OPRTase) plays an important role in de novo and salvage pathways of nucleotide synthesis and is widely used as a screening marker in genetic transformation. However, the function of OPRTase in plant pathogens remains unclear. In this study, we characterized an ortholog of Saccharomyces cerevisiae Ura5, the OPRTase MoPyr5, from the rice blast fungus Magnaporthe oryzae. Targeted gene disruption revealed that MoPyr5 is required for mycelial growth, appressorial turgor pressure and penetration into plant tissues, invasive hyphal growth, and pathogenicity. Interestingly, the ∆Mopyr5 mutant is also involved in mycelial surface hydrophobicity. Exogenous uridine 5'-phosphate (UMP) restored vegetative growth and rescued the defect in pathogenicity on detached barley and rice leaf sheath. Collectively, our results show that MoPyr5 is an OPRTase for UMP biosynthesis in M. oryzae and indicate that UTP biosynthesis is closely linked with vegetative growth, cell wall integrity, and pathogenicity of fungus. Our results also suggest that UMP biosynthesis would be a good target for the development of novel fungicides against M. oryzae. PMID:26810198

  8. Some novel features of glutathione transferase from juvenile catfish (Clarias gariepinus exposed to lindane-contaminated water

    Directory of Open Access Journals (Sweden)

    Yetunde Adedolapo Ojopagogo

    2015-03-01

    Full Text Available Catfish are hardy in nature and it is not known whether the presence of efficient detoxication enzymes is partly responsible for this trait. To investigate this, we have assessed induction of glutathione transferase (GST in 10-week-old juvenile catfish (Clarias gariepinus exposed to graded concentrations of lindane, an organochlorine insecticide, and characterised the purified enzyme from groups having the highest and statistically significant induction. Some of the unique properties observed for the purified enzyme are a high Km (1.72±0.21 mM for the electrophilic model substrate, 1-chloro-2,4-dinitrobenzene (CDNB and a very low catalytic rate (Vmax=0.130±0.010 units/mg protein. The kcat/Km being 55.4±0.2 M−1 s−1. The enzyme is present in high concentration in the organism, the main isoform accounts for about 5.6% of the total soluble protein, probably to compensate for the observed kinetic imperfection. Since these properties are generally not known for a detoxication enzyme, we suggest that they may form part of the organism׳s own adaptation to its polluted environment.

  9. Nucleocytoplasmic human O-GlcNAc transferase is sufficient for O-GlcNAcylation of mitochondrial proteins

    Science.gov (United States)

    Trapannone, Riccardo; Mariappa, Daniel; Ferenbach, Andrew T.; vanAalten, Daan M.F.

    2016-01-01

    O-linked N-acetylglucosamine modification (O-GlcNAcylation) is a nutrient-dependent protein post-translational modification (PTM), dynamically and reversibly driven by two enzymes: O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) that catalyse the addition and the removal of the O-GlcNAc moieties to/from serine and threonine residues of target proteins respectively. Increasing evidence suggests involvement of O-GlcNAcylation in many biological processes, including transcription, signalling, neuronal development and mitochondrial function. The presence of a mitochondrial O-GlcNAc proteome and a mitochondrial OGT (mOGT) isoform has been reported. We explored the presence of mOGT in human cell lines and mouse tissues. Surprisingly, analysis of genomic sequences indicates that this isoform cannot be expressed in most of the species analysed, except some primates. In addition, we were not able to detect endogenous mOGT in a range of human cell lines. Knockdown experiments and Western blot analysis of all the predicted OGT isoforms suggested the expression of only a single OGT isoform. In agreement with this, we demonstrate that overexpression of the nucleocytoplasmic OGT (ncOGT) isoform leads to increased O-GlcNAcylation of mitochondrial proteins, suggesting that ncOGT is necessary and sufficient for the generation of the O-GlcNAc mitochondrial proteome. PMID:27048592

  10. Glutathione S-transferase of brown planthoppers (Nilaparvata lugens is essential for their adaptation to gramine-containing host plants.

    Directory of Open Access Journals (Sweden)

    Xiao-Qin Sun

    Full Text Available Plants have evolved complex processes to ward off attacks by insects. In parallel, insects have evolved mechanisms to thwart these plant defenses. To gain insight into mechanisms that mediate this arms race between plants and herbivorous insects, we investigated the interactions between gramine, a toxin synthesized by plants of the family Gramineae, and glutathione S transferase (GST, an enzyme found in insects that is known to detoxify xenobiotics. Here, we demonstrate that rice (Oryza sativa, a hydrophytic plant, also produces gramine and that rice resistance to brown planthoppers (Nilaparvata lugens, BPHs is highly associated with in planta gramine content. We also show that gramine is a toxicant that causes BPH mortality in vivo and that knockdown of BPH GST gene nlgst1-1 results in increased sensitivity to diets containing gramine. These results suggest that the knockdown of key detoxification genes in sap-sucking insects may provide an avenue for increasing their sensitivity to natural plant-associated defense mechanisms.

  11. Volatile Gas Production by Methyl Halide Transferase: An In Situ Reporter Of Microbial Gene Expression In Soil.

    Science.gov (United States)

    Cheng, Hsiao-Ying; Masiello, Caroline A; Bennett, George N; Silberg, Jonathan J

    2016-08-16

    Traditional visual reporters of gene expression have only very limited use in soils because their outputs are challenging to detect through the soil matrix. This severely restricts our ability to study time-dependent microbial gene expression in one of the Earth's largest, most complex habitats. Here we describe an approach to report on dynamic gene expression within a microbial population in a soil under natural water levels (at and below water holding capacity) via production of methyl halides using a methyl halide transferase. As a proof-of-concept application, we couple the expression of this gas reporter to the conjugative transfer of a bacterial plasmid in a soil matrix and show that gas released from the matrix displays a strong correlation with the number of transconjugant bacteria that formed. Gas reporting of gene expression will make possible dynamic studies of natural and engineered microbes within many hard-to-image environmental matrices (soils, sediments, sludge, and biomass) at sample scales exceeding those used for traditional visual reporting. PMID:27415416

  12. Glutathione S-transferase SlGSTE1 in Spodoptera litura may be associated with feeding adaptation of host plants.

    Science.gov (United States)

    Zou, Xiaopeng; Xu, Zhibin; Zou, Haiwang; Liu, Jisheng; Chen, Shuna; Feng, Qili; Zheng, Sichun

    2016-03-01

    Spodoptera litura is polyphagous pest insect and feeds on plants of more than 90 families. In this study the role of glutathione S-transferase epilson 1 (slgste1) in S. litura in detoxification was examined. This gene was up-regulated in the midgut of S. litura at the transcriptional and protein levels when the insect fed on Brassica juncea or diet containing phytochemicals such as indole-3-carbinol and allyl-isothiocyanate that are metabolic products of sinigrin and glucobrassicin in B. juncea. The SlGSTE1 could catalyze the conjugation of reduced glutathione and indole-3-carbinol and allyl-isothiocyanate, as well as xanthotoxin, which is a furanocoumarin, under in vitro condition. When the expression of Slgste1 in the larvae was suppressed with RNAi, the larval growth and feeding rate were decreased. Furthermore, the up-regulated expression of the SlGSTE1 protein in the midgut of larvae that fed on different host plants was detected by 2-DE and ESI/MS analysis. The feeding adaptation from the most to the least of the larvae for the various host plants was Brassica alboglabra, Brassica linn. Pekinensis, Cucumis sativus, Ipomoea batatas, Arachis hypogaea and Capsicum frutescens. All the results together suggest that Slgste1 is a critical detoxifying enzyme that is induced by phytochmicals in the host plants and, inter alia, may be related to host plant adaptation of S. litura. PMID:26631599

  13. A novel glutathione-S transferase immunosensor based on horseradish peroxidase and double-layer gold nanoparticles.

    Science.gov (United States)

    Lu, Dingqiang; Lu, Fuping; Pang, Guangchang

    2016-06-01

    GSTs, a biotransformation enzyme group, can perform metabolism, drug transfer and detoxification functions. Rapid detection of the GSTs with more sensitive approaches is of great importance. In the current study, a novel double-layer gold nanoparticles-electrochemical immunosensor electrode (DGN-EIE) immobilized with Glutathione S-Transferase (GST) antibody derived from Balb/c mice was developed. To increase the fixed quantity of antibodies and electrochemical signal, an electrochemical biosensing signal amplification system was utilized with gold nanoparticles-thionine-chitosan absorbing horseradish peroxidase (HRP). In addition, transmission electron microscope (TEM) was used to characterize the nanogold solution. To evaluate the quality of DGN-EIE, the amperometric I-t curve method was applied to determine the GST in PBS. The results showed that the response current had a good linear correlation with the GST concentration ranged from 0.1-10(4) pg/mL. The lowest detection limit was found at 0.03 pg/mL(S/N = 3). The linear equation was deduced as △I/% = 7.386lgC + 22.36 (R(2) = 0.998). Moreover, it was validated with high sensitivity and reproducibility. Apparently, DGN-EIE may be a very useful tool for monitoring the GST. PMID:27220630

  14. Immunohistochemical detection of glutathione S-transferase (GST)-π in head and neck carcinoma and its change by radiotherapy

    International Nuclear Information System (INIS)

    It is well known that the 'placental form of glutathione S-transferase' (GST-π) is present in high concentrations in most human carcinomas. However, its expression in head and neck carcinomas have not yet been reported. The author investigated the expression of GST-π in the tissue of pharyngeal and laryngeal carcinomas by the immunohistochemical procedure. GST-π was positive in 80% of laryngeal carcinomas (35 cases) and 52.8% of pharyngeal carcinomas (36 cases). As a result, well differentiated squamous cell carcinomas showed stronger expression of GST-π than poorly differentiated squamous cell carcinomas. Although normal epithelia of the pharynx and larynx showed negative GST-π, it should be noticed that 54.6% of precancerous epithelia (11 cases) showed positive GST-π. Most patients treated with radiotherapy showed the diminution of GST-π expression after irradiation. However, correlation between the strength of initial GST-π expression and the effectiveness of radiotherapy was not observed (p<0.01). (author)

  15. Identification and Characterization of Seven Glutathione S-Transferase Genes from Citrus Red Mite, Panonychus citri (McGregor

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    Chong-Yu Liao

    2013-12-01

    Full Text Available The citrus red mite, Panonychus citri (McGregor, is a global citrus pest, and has developed severe resistance to several types of acaricides. However, the molecular mechanisms of resistance in this mite remain unknown. In this study, seven full-length cDNAs encoding glutathione S-transferases (GSTs genes were identified and characterized in P. citri. The effects of pyridaben and fenpropathrin exposure on the expression of these genes were also investigated. Phylogenetic analysis revealed that the seven GSTs genes in P. citri cloned in this study belong to three different cytosolic classes, including four in mu, two in delta and one in zeta. Among these seven GSTs genes, the relative expression level of PcGSTm1 was significantly higher in adult than in the other life stages (egg, larvae and nymph. Compared with the control, the mRNA levels of the seven GST genes did not change significantly following exposure to pyridaben at LC10. However, RT-qPCR results showed that, when exposed to LC10 of fenpropathrin, six GSTs gene (PcGSTm1, PcGSTm3, PcGSTm4, PcGSTd1, PcGSTd2 and PcGSTz1 transcripts increased in a time-dependent manner. This is the first insight into the molecular characteristics of GSTs gene cDNAs in P. citri. The elevated GSTs gene transcripts following exposure to fenpropathrin might be one of the mechanisms involved in detoxification of this acaricide.

  16. Identification and characterization of seven glutathione S-transferase genes from citrus red mite, Panonychus citri (McGregor).

    Science.gov (United States)

    Liao, Chong-Yu; Zhang, Kun; Niu, Jin-Zhi; Ding, Tian-Bo; Zhong, Rui; Xia, Wen-Kai; Dou, Wei; Wang, Jin-Jun

    2013-01-01

    The citrus red mite, Panonychus citri (McGregor), is a global citrus pest, and has developed severe resistance to several types of acaricides. However, the molecular mechanisms of resistance in this mite remain unknown. In this study, seven full-length cDNAs encoding glutathione S-transferases (GSTs) genes were identified and characterized in P. citri. The effects of pyridaben and fenpropathrin exposure on the expression of these genes were also investigated. Phylogenetic analysis revealed that the seven GSTs genes in P. citri cloned in this study belong to three different cytosolic classes, including four in mu, two in delta and one in zeta. Among these seven GSTs genes, the relative expression level of PcGSTm1 was significantly higher in adult than in the other life stages (egg, larvae and nymph). Compared with the control, the mRNA levels of the seven GST genes did not change significantly following exposure to pyridaben at LC10. However, RT-qPCR results showed that, when exposed to LC10 of fenpropathrin, six GSTs gene (PcGSTm1, PcGSTm3, PcGSTm4, PcGSTd1, PcGSTd2 and PcGSTz1) transcripts increased in a time-dependent manner. This is the first insight into the molecular characteristics of GSTs gene cDNAs in P. citri. The elevated GSTs gene transcripts following exposure to fenpropathrin might be one of the mechanisms involved in detoxification of this acaricide. PMID:24351815

  17. Thymidylate Synthase, Thymidine Phosphorylase and Orotate Phosphoribosyl Transferase Levels as Predictive Factors of Chemotherapy in Oral Squamous Cell Carcinoma

    International Nuclear Information System (INIS)

    We conducted a clinicopathologic study on protein and mRNA levels of thymidylate synthase (TS), thymidine phosphorylase (TP) and orotate phosphoribosyl transferase (OPRT) using biopsy tissue specimens before treatment. The mRNA levels have been measured in tumor cells microdissected from paraffin-embedded specimens (Danenberg Tumor Profile method: DTP method). We studied the mRNA and protein expression as effect predictive factors in chemotherapy. The subjects consisted of 20 cases of untreated oral squamous cell carcinoma who had undergone chemotherapy with TS-1 (16 males and 4 females, tongue in 8 cases, upper gingiva in 3 cases, lower gingiva in 3 cases, buccal mucosa in 5 cases and floor of the mouth in 1 case). TS gene expressions of the responders were lower than those for the nonresponders. Furthermore, regarding males who were less than 70 years of age, stage I and II, well differentiated type and tongue, TS mRNA expression of the responders were lower than that for the nonresponders. The mRNA expression of OPRT for the male responders was lower than that for the nonresponders. No remarkable difference was observed by immunohistochemistry. In this study, the measurement of the TS levels using the DTP method may potentially act as a predictive factor of antitumor effectiveness

  18. Different effects of nine clausenamide ennatiomers on liver glutathione biosynthesis and glutathione S-transferase activity in mice

    Institute of Scientific and Technical Information of China (English)

    Yu-qun WU; Li-de LIU; Hua-ling WEI; Geng-tao LIU

    2006-01-01

    Aim: To study the effects of nine synthetic clausenamide with different stereo structures on liver glutathione (GSH) biosynthesis and glutathione S-transferase (GST) activity in mice. Methods: The nine test compounds were racemic mixtures and their ennatiomers of clausenamide, neoclausenamide and epineoclausenamide. Mice were administered clausenamide 250 mg/kg once daily for 3 consecutive days, ig, and were killed 24 h after the last dosing. The mouse liver cytosol GSH and GST were determined with related biochemical methods. Results: Nine clausenamides exhibited different effects on liver GSH and GST. Of nine clausenamides, only (+) and (±)clausenamide markedly increased liver cytosol GSH content. The mechanism of increasing liver GSH content of (+)clausenamide is mainly due to stimulating the key limiting enzyme γ-glutamylcysteine synthetase (γ-GCS) activity for GSH biosynthesis. The other test clausenamides had no such effect on liver GSH. All of the nine clausenamides induced a significant increase of GST activity. Conclusion: The effects of clausenamide ennatiomers on liver GST and GSH varied with the alterations of their spatial structures. (+)Clausenamide stimulated liver GSH biosynthesis through enhancingγ-GCS activity.

  19. Problematic detoxification of estrogen quinones by NAD(P)H-dependent quinone oxidoreductase and glutathione-S-transferase.

    Science.gov (United States)

    Chandrasena, R Esala P; Edirisinghe, Praneeth D; Bolton, Judy L; Thatcher, Gregory R J

    2008-07-01

    Estrogen exposure through early menarche, late menopause, and hormone replacement therapy increases the risk factor for hormone-dependent cancers. Although the molecular mechanisms are not completely established, DNA damage by quinone electrophilic reactive intermediates, derived from estrogen oxidative metabolism, is strongly implicated. A current hypothesis has 4-hydroxyestrone-o-quinone (4-OQE) acting as the proximal estrogen carcinogen, forming depurinating DNA adducts via Michael addition. One aspect of this hypothesis posits a key role for NAD(P)H-dependent quinone oxidoreductase (NQO1) in the reduction of 4-OQE and protection against estrogen carcinogenesis, despite two reports that 4-OQE is not a substrate for NQO1. 4-OQE is rapidly and efficiently trapped by GSH, allowing measurement of NADPH-dependent reduction of 4-OQE in the presence and absence of NQO1. 4-OQE was observed to be a substrate for NQO1, but the acceleration of NADPH-dependent reduction by NQO1 over the nonenzymic reaction is less than 10-fold and at more relevant nanomolar concentrations of substrate is less than 2-fold. An alternative detoxifying enzyme, glutathione-S-transferase, was observed to be a target for 4-OQE, rapidly undergoing covalent modification. These results indicate that a key role for NQO1 and GST in direct detoxification of 4-hydroxy-estrogen quinones is problematic. PMID:18588320

  20. m5C RNA and m5C DNA methyl transferases use different cysteine residues as catalysts.

    Science.gov (United States)

    Liu, Y; Santi, D V

    2000-07-18

    A family of RNA m(5)C methyl transferases (MTases) containing over 55 members in eight subfamilies has been identified recently by an iterative search of the genomic sequence databases by using the known 16S rRNA m(5)C 967 MTase, Fmu, as an initial probe. The RNA m(5)C MTase family contained sequence motifs that were highly homologous to motifs in the DNA m(5)C MTases, including the ProCys sequence that contains the essential Cys catalyst of the functionally similar DNA-modifying enzymes; it was reasonable to assign the Cys nucleophile to be that in the conserved ProCys. The family also contained an additional conserved Cys residue that aligns with the nucleophilic catalyst in m(5)U54 tRNA MTase. Surprisingly, the mutant of the putative Cys catalyst in the ProCys sequence was active and formed a covalent complex with 5-fluorocytosine-containing RNA, whereas the mutant at the other conserved Cys was inactive and unable to form the complex. Thus, notwithstanding the highly homologous sequences and similar functions, the RNA m(5)C MTase uses a different Cys as a catalytic nucleophile than the DNA m(5)C MTases. The catalytic Cys seems to be determined, not by the target base that is modified, but by whether the substrate is DNA or RNA. The function of the conserved ProCys sequence in the RNA m(5)C MTases remains unknown. PMID:10899996

  1. Dynamic interplay between catalytic and lectin domains of GalNAc-transferases modulates protein O-glycosylation

    Science.gov (United States)

    Lira-Navarrete, Erandi; de Las Rivas, Matilde; Compañón, Ismael; Pallarés, María Carmen; Kong, Yun; Iglesias-Fernández, Javier; Bernardes, Gonçalo J. L.; Peregrina, Jesús M.; Rovira, Carme; Bernadó, Pau; Bruscolini, Pierpaolo; Clausen, Henrik; Lostao, Anabel; Corzana, Francisco; Hurtado-Guerrero, Ramon

    2015-05-01

    Protein O-glycosylation is controlled by polypeptide GalNAc-transferases (GalNAc-Ts) that uniquely feature both a catalytic and lectin domain. The underlying molecular basis of how the lectin domains of GalNAc-Ts contribute to glycopeptide specificity and catalysis remains unclear. Here we present the first crystal structures of complexes of GalNAc-T2 with glycopeptides that together with enhanced sampling molecular dynamics simulations demonstrate a cooperative mechanism by which the lectin domain enables free acceptor sites binding of glycopeptides into the catalytic domain. Atomic force microscopy and small-angle X-ray scattering experiments further reveal a dynamic conformational landscape of GalNAc-T2 and a prominent role of compact structures that are both required for efficient catalysis. Our model indicates that the activity profile of GalNAc-T2 is dictated by conformational heterogeneity and relies on a flexible linker located between the catalytic and the lectin domains. Our results also shed light on how GalNAc-Ts generate dense decoration of proteins with O-glycans.

  2. Null genotype of glutathione S-transferase M1 is associated with senile cataract susceptibility in non-smoker females

    International Nuclear Information System (INIS)

    In the present study, we investigated whether the polymorphisms of glutathione S-transferase M1 (GSTM1) and T1 (GSTT1) genes are risk factors of cataract among Iranian population in a molecular epidemiological way. Blood samples from 150 subjects with cataract (72 male; 78 female) and 150 age- and sex-matched healthy persons were collected. Both patient and control groups were unrelated Iranian Muslims. Using PCR-based method, the genotypes were determined. The null GSTM1 genotype was associated with a 2.38-fold increase in the risk of developing cataract (OR=2.38; 95% CI=1.46-3.89; P=0.0003). After stratification by sex of subjects, the association was apparent only among women (OR=3.20; 95% CI=1.58-6.52; P=0.0007). The GSTT1 null genotype was associated with a 1.10-fold increased risk of developing cataract, but this association was not statistically significant. After stratification by sex of subjects, same results were obtained. Female patients with null genotype for GSTM1 and no history of smoking had a 3.45-fold increased cataract risk (P<0.05), whereas females who were null for GSTM1 and having history of smoking were not at increased risk of cataract

  3. Effects of Inhibitors of [Delta]24(25)-Sterol Methyl Transferase on the Ultrastructure of Epimastigotes of Trypanosoma cruzi

    Science.gov (United States)

    Braga, Marina V.; Magaraci, Filippo; Orenes Lorente, Silvia; Gilbert, Ian; de Souza, Wanderley

    2005-12-01

    Trypanosoma cruzi is the ethiological agent of Chagas disease. New compounds are being developed based on the biosynthesis and function of sterols, because T. cruzi has a requirement for specific endogenous sterols for growth and survival. Sterol biosynthesis inhibitors (SBIs) are drugs commonly used against fungal diseases. These drugs act by depleting essential and specific membrane components and/or inducing the accumulation of toxic intermediary or lateral products of the biosynthetic pathway. In this work we present the effects of WSP488, WSP501, and WSP561, specific inhibitors of [Delta]24(25)-sterol methyl transferase, on the ultrastructure of T. cruzi epimastigotes. All three drugs inhibited parasite multiplication at low concentrations, with IC50 values of 0.48, 0.44, and 0.48 [mu]M, respectively, and induced marked morphological changes including (a) blockage of cell division; (b) swelling of the mitochondrion, with several projections and depressions; (c) swelling of the perinuclear space; (d) presence of autophagosomes and myelin-like figures; (e) enlargement of the flagellar pocket and of a cytoplasmic vacuole located in close association with the flagellar pocket; (f) detachment of the membrane of the cell body; and (g) formation of a vesicle at the surface of the parasite between the flagellar pocket and the cytostome. Our results show that these drugs are potent in vitro inhibitors of growth of T. cruzi.

  4. Cloning and sequencing of protein L-isoaspartyl O-methyl transferase of Salmonella Typhimurium isolated from poultry

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    S. K. Dixit

    2014-09-01

    Full Text Available Aim: To clone the Salmonella Typhimurium protein L-isoaspartyl O-methyl transferase (PIMT enzyme and to analyze the sequence with PIMT gene of other pathogenic serovars of Salmonella. Materials and Methods: Salmonella Typhimurium strain E-2375 was procured from the National Salmonella Center, IVRI. The genomic DNA was isolated from Salmonella Typhimurium. Polymerase chain reaction (PCR was carried out to amplify PIMT gene using the designed primers. The PCR product was cloned into pET28c plasmid vector and transformed into Escherichia coli DH5α cells. The plasmid was isolated from E. coli and was sequenced. The sequence was analyzed and submitted in Genbank. Results: The PCR product revealed a distinct amplicon of 627 bp. The clone was confirmed by PCR. Sequencing data revealed 100% homology between PIMT sequences from Salmonella Typhimurium strain E-2375 (used in the current study and PIMT sequences of standard reported strain (Salmonella Typhimurium str. LT2 in NCBI data base. This submitted sequence in Genbank having accession no. KJ575536. Conclusions: PIMT gene of Salmonella is highly conserved in most of the pathogenic Salmonella serovars. The PIMT clone can be used to isolate PIMT protein. This PIMT protein will be helpful to identify isoaspartate containing proteins thus can help in study Salmonella virulence.

  5. Plasma Hypoxanthine-Guanine Phosphoribosyl Transferase Activity in Bottlenose Dolphins Contributes to Avoiding Accumulation of Non-recyclable Purines

    Science.gov (United States)

    López-Cruz, Roberto I.; Crocker, Daniel E.; Gaxiola-Robles, Ramón; Bernal, Jaime A.; Real-Valle, Roberto A.; Lugo-Lugo, Orlando; Zenteno-Savín, Tania

    2016-01-01

    Marine mammals are exposed to ischemia/reperfusion and hypoxia/reoxygenation during diving. During oxygen deprivation, adenosine triphosphate (ATP) breakdown implies purine metabolite accumulation, which in humans is associated with pathological conditions. Purine recycling in seals increases in response to prolonged fasting and ischemia. Concentrations of metabolites and activities of key enzymes in purine metabolism were examined in plasma and red blood cells from bottlenose dolphins (Tursiops truncatus) and humans. Hypoxanthine and inosine monophosphate concentrations were higher in plasma from dolphins than humans. Plasma hypoxanthine-guanine phosphoribosyl transferase (HGPRT) activity in dolphins suggests an elevated purine recycling rate, and a mechanism for avoiding accumulation of non-recyclable purines (xanthine and uric acid). Red blood cell concentrations of hypoxanthine, adenosine diphosphate, ATP and guanosine triphosphate were lower in dolphins than in humans; adenosine monophosphate and nicotinamide adenine dinucleotide concentrations were higher in dolphins. HGPRT activity in red blood cells was higher in humans than in dolphins. The lower concentrations of purine catabolism and recycling by-products in plasma from dolphins could be beneficial in providing substrates for recovery of ATP depleted during diving or vigorous swimming. These results suggest that purine salvage in dolphins could be a mechanism for delivering nucleotide precursors to tissues with high ATP and guanosine triphosphate requirements. PMID:27375492

  6. Effect of Arsenic and Chromium on the Serum Amino-Transferases Activity in Indian Major Carp, Labeo rohita

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    Anjaneyulu Yerramilli

    2007-09-01

    Full Text Available Arsenic and hexavalent chromium toxicity results from their ability to interact with sulfahydryl groups of proteins and enzymes, and to substitute phosphorus in a variety of biochemical reactions. Alanine aminotransferase (ALT; E.C: 2.6.1.2 and Aspartate amino transferase (AST; EC 2.6.1.1 play a crucial role in transamination reactions and can be used as potential biomarkers to indicate hepatotoxicity and cellular damage. While histopathological studies in liver tissue require more time and expertise, simple and reliable biochemical analysis of ALT and AST can be used for a rapid assessment of tissue and cellular damage within 96 h. The main objective of this study was to determine the acute effects of arsenic and hexavalent chromium on the activity of ALT and AST in the Indian major carp, Labeo rohita for 24 h and 96 h. Significant increase in the activity of ALT (P < 0.01 from controls in arsenic exposed fish indicates serious hepatic damage and distress condition to the fish. However, no such significant changes were observed in chromium-exposed fish suggesting that arsenic is more toxic to the fish. These findings indicate that ALT and AST are candidate biomarkers for arsenic-induced hepatotoxicity in Labeo rohita.

  7. Potential use of acetylcholinesterase, glutathione-S-transferase and metallothionein for assessment of contaminated sediment in tropical chironomid, Chironomus javanus.

    Science.gov (United States)

    Somparn, A; Iwai, C B; Noller, B

    2015-11-01

    Heavy metals and organophosphorus insecticide is known to act as disruptors for the enzyme system, leading to physiologic disorders. The present study was conducted to investigate the potential use of these enzymes as biomarkers in assessment of contaminated sediments on tropical chironomid species. Acetylcholinesterase (AChE), glutathione-S-transferase (GST) and metallothionein (MT) activity was measured in the fourth-instar chironomid larvae, Chironomus javanus, Kieffer, after either 48-hr or 96-hr exposure to organophosphorus insecticide, chlorpyrifos (0.01- 0.25 mg kg(-1)) or heavy metal cadmium (0.1-25 mg kg(-1)). Exposure to chlorpyrifos (0.01 mg kg(-1)) at 48 and 96 hr significantly of AChE activity (64.2%-85.9%) and induced GST activity (33.9-63.8%) when compared with control (P GST activity (11.7-40%) and MT level (9.0%-70.5%) when compared with control (P impact of enzyme activity on chlorpyrifos and cadmium contamination. Activity of AChE, GST and MT could serve as potential biomarkers for assessment and biomonitoring the effects of insecticide and heavy metal contamination in tropical aquatic ecosystems. PMID:26688973

  8. Proteomic Profiling of Cytosolic Glutathione Transferases from Three Bivalve Species: Corbicula fluminea, Mytilus galloprovincialis and Anodonta cygnea

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    José Carlos Martins

    2014-01-01

    Full Text Available Suspension-feeding bivalves are considered efficient toxin vectors with a relative insensitivity to toxicants compared to other aquatic organisms. This fact highlights the potential role of detoxification enzymes, such as glutathione transferases (GSTs, in this bivalve resistance. Nevertheless, the GST system has not been extensively described in these organisms. In the present study, cytosolic GSTs isoforms (cGST were surveyed in three bivalves with different habitats and life strategies: Corbicula fluminea, Anodonta cygnea and Mytilus galloprovincialis. GSTs were purified by glutathione-agarose affinity chromatography, and the collection of expressed cGST classes of each bivalve were identified using a proteomic approach. All the purified extracts were also characterized kinetically. Results reveal variations in cGST subunits collection (diversity and properties between the three tested bivalves. Using proteomics, four pi-class and two sigma-class GST subunits were identified in M. galloprovincialis. C. fluminea also yielded four pi-class and one sigma-class GST subunits. For A. cygnea, two mu-class and one pi-class GST subunits were identified, these being the first record of GSTs from these freshwater mussels. The affinity purified extracts also show differences regarding enzymatic behavior among species. The variations found in cGST collection and kinetics might justify diverse selective advantages for each bivalve organism.

  9. Conformational change of glutathione-S-transferase by its co-expression with prion domain of yeast Ure2p

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    The Ure2 protein from Saccharomyces cerevisisae has a changeable structure similar to that ofrnammalian prion protein. Its N-terminal is the prion domain (PrD) consisting of 65 amino acids which plays a critical role in yeast prion development. In this study, PrD gene was recombinated with glutathione-S-transferase(GST) gene, and a soluble GST-PrD(sGST-PrD) fusion protein was expressed in E. coli. sGST-PrD could spontaneously polymerize into amyloid fibrils in vitro, displaying typical β-sheet-type structure; it had increased resistance to proteinase K and exhibited amvloid-like optical properties. Moreover, the aggregated GST-PrD(aGST-PrD) could induce sGST-PrD to aggregate into fibrils. These results indicate that PrD could change the conformation of GST moiety in a recombinant protein with PrD to form a prion-like chimeric protein, which proves that PrD has the ability to mediate a prion-like conversion of other proteins fused with it.

  10. Glutathione S-Transferase P1 (GSTP1 gene polymorphism increases age-related susceptibility to hepatocellular carcinoma

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    Kuo Wu-Hsien

    2010-03-01

    Full Text Available Abstract Background Hepatocellular carcinoma (HCC is one of the most frequent malignant neoplasms in the world. Genetic polymorphism has been reported to be a factor increasing the risk of HCC. Phase II enzymes such as glutathione s-transferases (GSTP1, GSTA1 play important roles in protecting cells against damage induced by carcinogens. The aim of this study was to estimate the relationship of the GSTP1 and GSTA1 gene polymorphisms to HCC risk and clinico-pathological status. Methods Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP was used to measure GSTP1 (A→G and GSTA1 (C→T gene polymorphisms in 386 healthy controls and 177 patients with HCC. Results Neither gene polymorphism was associated with the clinico-pathological status of HCC and serum expression of liver-related clinico-pathological markers. No association between the GSTA1 gene polymorphism and HCC susceptibility was found. However, in the younger group, aged ≤ 57 years, individuals with AG or GG alleles of GSTP1 had a 2.18-fold (95%CI = 1.09-4.36; p = 0.02 and 5.64-fold (95%CI = 1.02-31.18; p = 0.04 risk, respectively, of developing HCC compared to individuals with AA alleles, after adjusting for other confounders. Conclusion AG and GG alleles of GSTP1 gene polymorphisms may be considered as factors increasing the susceptibility to and risk of HCC in Taiwanese aged ≤ 57 years.

  11. Non-enzymatic roles for the URE2 glutathione S-transferase in the response of Saccharomyces cerevisiae to arsenic.

    Science.gov (United States)

    Todorova, Tatina T; Kujumdzieva, Anna V; Vuilleumier, Stéphane

    2010-11-01

    The response of Saccharomyces cerevisiae to arsenic involves a large ensemble of genes, many of which are associated with glutathione-related metabolism. The role of the glutathione S-transferase (GST) product of the URE2 gene involved in resistance of S. cerevisiae to a broad range of heavy metals was investigated. Glutathione peroxidase activity, previously reported for the Ure2p protein, was unaffected in cell-free extracts of an ure2Δ mutant of S. cerevisiae. Glutathione levels in the ure2Δ mutant were lowered about threefold compared to the isogenic wild-type strain but, as in the wild-type strain, increased 2-2.5-fold upon addition of either arsenate (As(V)) or arsenite (As(III)). However, lack of URE2 specifically caused sensitivity to arsenite but not to arsenate. The protective role of URE2 against arsenite depended solely on the GST-encoding 3'-end portion of the gene. The nitrogen source used for growth was suggested to be an important determinant of arsenite toxicity, in keeping with non-enzymatic roles of the URE2 gene product in GATA-type regulation. PMID:20740275

  12. Inhibition of carnitine-acyl transferase I by oxfenicine studied in vivo with [{sup 11}C]-labeled fatty acids

    Energy Technology Data Exchange (ETDEWEB)

    Angsten, Gertrud [Department of Pediatric Surgery, University Children' s Hospital, S-751 85 Uppsala (Sweden)]. E-mail: gertrud.angsten@surgsci.uu.se; Valind, Sven [Uppsala University PET Centre, Uppsala University, S-751 05 Uppsala (Sweden); Department of Clinical Physiology, University Hospital, S-751 85 Uppsala (Sweden); Takalo, Reijo [Uppsala University PET Centre, Uppsala University, S-751 05 Uppsala (Sweden); Department of Clinical Physiology, University Hospital, S-751 85 Uppsala (Sweden); Neu, Henrik [Uppsala University PET Centre, Uppsala University, S-751 05 Uppsala (Sweden); Department of Organic Chemistry, Uppsala University, S-751 24 Uppsala (Sweden); Meurling, Staffan [Department of Pediatric Surgery, University Children' s Hospital, S-751 85 Uppsala (Sweden); Langstroem, Bengt [Uppsala University PET Centre, Uppsala University, S-751 05 Uppsala (Sweden); Department of Organic Chemistry, Uppsala University, S-751 24 Uppsala (Sweden)

    2005-07-01

    Methods: Anesthetized pigs were studied with [{sup 11}C]-labeled fatty acids (FAs) with carbon chain length ranging from 8 to 16 carbon atoms, during control conditions and during inhibition of carnitine-palmitoyl transferase I (CPT I) with oxfenicine. The myocardial uptake of [{sup 11}C]-FAs from blood was measured together with the relative distribution of [{sup 11}C]-acyl-CoA between rapid mitochondrial oxidation and incorporation into slow turnover lipid pools in the heart. Results: During baseline conditions, the fractional oxidative utilization of palmitate was almost as high as that of carnitine-independent short-chain FAs, unless the carnitine shuttle was inhibited by high levels of lactate. Inhibition of CPT I almost completely blocked the oxidative pathway for palmitic acid and reduced the fractional oxidative utilization, while the rate of oxidative metabolism of acyl-CoA was unaffected. Conclusions: [{sup 11}C]-Labeled FAs allow rapid oxidation to be well separated from esterification into slow turnover lipid pools in the heart of anaesthetized pigs. The fractional oxidative utilization of [{sup 11}C]-palmitate serves well to characterize, in vivo, the carnitine-dependent transfer of long-chain FAs.

  13. Inhibition of carnitine-acyl transferase I by oxfenicine studied in vivo with [11C]-labeled fatty acids

    International Nuclear Information System (INIS)

    Methods: Anesthetized pigs were studied with [11C]-labeled fatty acids (FAs) with carbon chain length ranging from 8 to 16 carbon atoms, during control conditions and during inhibition of carnitine-palmitoyl transferase I (CPT I) with oxfenicine. The myocardial uptake of [11C]-FAs from blood was measured together with the relative distribution of [11C]-acyl-CoA between rapid mitochondrial oxidation and incorporation into slow turnover lipid pools in the heart. Results: During baseline conditions, the fractional oxidative utilization of palmitate was almost as high as that of carnitine-independent short-chain FAs, unless the carnitine shuttle was inhibited by high levels of lactate. Inhibition of CPT I almost completely blocked the oxidative pathway for palmitic acid and reduced the fractional oxidative utilization, while the rate of oxidative metabolism of acyl-CoA was unaffected. Conclusions: [11C]-Labeled FAs allow rapid oxidation to be well separated from esterification into slow turnover lipid pools in the heart of anaesthetized pigs. The fractional oxidative utilization of [11C]-palmitate serves well to characterize, in vivo, the carnitine-dependent transfer of long-chain FAs

  14. Carnitine palmitoyl transferase activity in Morris Hepatoma 7777 mitochondria and its sensitivity to malonyl CoA inhibition

    International Nuclear Information System (INIS)

    Earlier reports in the literature have indicated no detectable Carnitine Palymitoyl Transferase (CPT) activity in homogenates prepared from Morris Hepatoma 7777. In its study CPT activity in isolated mitochondria (mito) was measured by butanol extraction of the [3H]palmitoyl carnitine formed as outlined by Bremer et al. Contrary to the earlier work where no appreciable activity of CPT was observed the authors find significant levels of CPT (2.6 nMol/min/mg protein) in isolated mito from Morris Hepatoma 7777 (MH 7777). The level of CPT activity observed in MH 7777 mito was, however, 36% lower compared to the host liver CPT activity (4.1 nMol/min/mg protein). The enzyme in MH 7777 mito showed 83% inhibition in the presence of 10 μM malonyl CoA, in agreement with the degree of sensitivity observed with the host liver isolated mito. On freeze thawing host mito, total CPT activity increased and the sensitivity of the enzyme to malonyl CoA decreased. Frozen thawed MH 7777 mito showed a similar response to malonyl CoA but no change in the total CPT level was observed. The authors results establish for the first time the presence of a malonyl CoA sensitive CPT in MH 7777 mito, which may have slightly different properties from normal due to the membrane environment of the enzyme

  15. Terminal deoxynucleotidyl transferase is down-regulated by AP-1-like regulatory elements in human lymphoid cells.

    Science.gov (United States)

    Peralta-Zaragoza, Oscar; Recillas-Targa, Félix; Madrid-Marina, Vicente

    2004-02-01

    Terminal deoxynucleotidyl transferase (TdT) is a template-independent DNA polymerase that catalyses the incorporation of deoxyribonucleotides into the 3'-hydroxyl end of DNA templates and is thought to increase junctional diversity of antigen receptor genes. TdT is expressed only on immature lymphocytes and acute lymphoblastic leukaemia cells and its transcriptional expression is tightly regulated. We had previously found that protein kinase C (PKC) activation down-regulates TdT expression. PKC-activation induces the synthesis of the Fos and Jun proteins, known as the major components of activation protein 1 (AP-1) transcriptional factor implicated in transcriptional control. Here we report the identification of several DNA-protein interactions within the TdT promoter region in non-TdT expressing human cells. Sequence analysis revealed the presence of a putative AP-1-like DNA-binding site, suggesting that AP-1 may play a relevant role in TdT transcriptional regulation. Using a different source of nuclear extracts and the AP-1-TdT motif as a probe we identified several DNA-protein retarded complexes in electrophoretic mobility shift assays. Super-band shifting analysis using an antibody against c-Jun protein confirmed that the main interaction is produced by a nuclear factor that belongs to the AP-1 family transcription factors. Our findings suggest that the TdT gene expression is down-regulated, at least in part, through AP-1-like transcription factors. PMID:15027905

  16. Membrane-bound catechol-O-methyl transferase in cortical neurons and glial cells is intracellularly oriented

    Directory of Open Access Journals (Sweden)

    Björn H Schott

    2010-10-01

    Full Text Available Catechol-O-methyl transferase (COMT is involved in the inactivation of dopamine in brain regions in which the dopamine transporter (DAT1 is sparsely expressed. The membrane-bound isoform of COMT (MB-COMT is the predominantly expressed form in the mammalian central nervous system (CNS. It has been a matter of debate whether in neural cells of the CNS the enzymatic domain of MB-COMT is oriented towards the cytoplasmic or the extracellular compartment. Here we used live immunocytochemistry on cultured neocortical neurons and glial cells to investigate the expression and membrane orientation of native COMT and of transfected MB-COMT fused to green fluorescent protein (GFP. After live staining, COMT immunoreactivity was reliably detected in both neurons and glial cells after permeabilization, but not on unpermeabilized cells. Similarly, autofluorescence of COMT-GFP fusion protein and antibody fluorescence showed overlap only in permeabilized neurons. Our data provide converging evidence for an intracellular membrane orientation of MB-COMT in neurons and glial cells, suggesting the presence of a DAT1-independent postsynaptic uptake mechanism for dopamine, prior to its degradation via COMT.

  17. Immunocytochemical studies of the distribution of alpha and pi isoforms of glutathione S-transferase in cystic renal diseases.

    Science.gov (United States)

    Hiley, C G; Otter, M; Bell, J; Strange, R C; Keeling, J W

    1994-01-01

    We describe immunohistochemical studies of the expression of alpha and pi class glutathione S-transferases (GSTs) in normal fetal kidneys. These define, in greater detail, changes in expression of alpha isoforms in the proximal tubule. At about 36 weeks of gestation expression of alpha isoforms was down-regulated in the distal tubules and collecting ducts while pi was expressed throughout the nephron. Tubular expression of alpha isoforms was restricted to the part adjacent to the glomerulus; cells farthest from the glomerulus were negative. After 40 weeks of gestation, alpha isoforms were expressed along the entire proximal tubule, while pi was restricted to the distal tubule and collecting ducts. GST expression was also studied in multicystic renal dysplasia, autosomal recessive polycystic kidney disease, and autosomal dominant polycystic kidney disease to determine whether the patterns of expression of alpha and pi isoforms allow identification of the origin of the cysts that characterize these diseases. Cysts were lined by epithelia that were strongly positive for alpha and pi isoforms. The epithelia of noncystic nephrons in renal cystic dysplasia demonstrated delayed maturity, suggesting that GST expression was dependent on the stage of development and not length of gestation. PMID:8066005

  18. Activity Based High-Throughput Screening for Novel O-GlcNAc Transferase Substrates Using a Dynamic Peptide Microarray.

    Directory of Open Access Journals (Sweden)

    Jie Shi

    Full Text Available O-GlcNAcylation is a reversible and dynamic protein post-translational modification in mammalian cells. The O-GlcNAc cycle is catalyzed by O-GlcNAc transferase (OGT and O-GlcNAcase (OGA. O-GlcNAcylation plays important role in many vital cellular events including transcription, cell cycle regulation, stress response and protein degradation, and altered O-GlcNAcylation has long been implicated in cancer, diabetes and neurodegenerative diseases. Recently, numerous approaches have been developed to identify OGT substrates and study their function, but there is still a strong demand for highly efficient techniques. Here we demonstrated the utility of the peptide microarray approach to discover novel OGT substrates and study its specificity. Interestingly, the protein RBL-2, which is a key regulator of entry into cell division and may function as a tumor suppressor, was identified as a substrate for three isoforms of OGT. Using peptide Ala scanning, we found Ser 420 is one possible O-GlcNAc site in RBL-2. Moreover, substitution of Ser 420, on its own, inhibited OGT activity, raising the possibility of mechanism-based development for selective OGT inhibitors. This approach will prove useful for both discovery of novel OGT substrates and studying OGT specificity.

  19. Proteomic profiling of cytosolic glutathione transferases from three bivalve species: Corbicula fluminea, Mytilus galloprovincialis and Anodonta cygnea.

    Science.gov (United States)

    Martins, José Carlos; Campos, Alexandre; Osório, Hugo; da Fonseca, Rute; Vasconcelos, Vítor

    2014-01-01

    Suspension-feeding bivalves are considered efficient toxin vectors with a relative insensitivity to toxicants compared to other aquatic organisms. This fact highlights the potential role of detoxification enzymes, such as glutathione transferases (GSTs), in this bivalve resistance. Nevertheless, the GST system has not been extensively described in these organisms. In the present study, cytosolic GSTs isoforms (cGST) were surveyed in three bivalves with different habitats and life strategies: Corbicula fluminea, Anodonta cygnea and Mytilus galloprovincialis. GSTs were purified by glutathione-agarose affinity chromatography, and the collection of expressed cGST classes of each bivalve were identified using a proteomic approach. All the purified extracts were also characterized kinetically. Results reveal variations in cGST subunits collection (diversity and properties) between the three tested bivalves. Using proteomics, four pi-class and two sigma-class GST subunits were identified in M. galloprovincialis. C. fluminea also yielded four pi-class and one sigma-class GST subunits. For A. cygnea, two mu-class and one pi-class GST subunits were identified, these being the first record of GSTs from these freshwater mussels. The affinity purified extracts also show differences regarding enzymatic behavior among species. The variations found in cGST collection and kinetics might justify diverse selective advantages for each bivalve organism. PMID:24473139

  20. Proanthocyanidins inhibit Ascaris suum glutathione-S-transferase activity and increase susceptibility of larvae to levamisole in vitro.

    Science.gov (United States)

    Hansen, Tina V A; Fryganas, Christos; Acevedo, Nathalie; Caraballo, Luis; Thamsborg, Stig M; Mueller-Harvey, Irene; Williams, Andrew R

    2016-08-01

    Proanthocyanidins (PAC) are a class of plant secondary metabolites commonly found in the diet that have shown potential to control gastrointestinal nematode infections. The anti-parasitic mechanism(s) of PAC remain obscure, however the protein-binding properties of PAC suggest that disturbance of key enzyme functions may be a potential mode of action. Glutathione-S-transferases (GSTs) are essential for parasite detoxification and have been investigated as drug and vaccine targets. Here, we show that purified PAC strongly inhibit the activity of both recombinant and native GSTs from the parasitic nematode Ascaris suum. As GSTs are involved in detoxifying xenobiotic substances within the parasite, we hypothesised that this inhibition may render parasites hyper-susceptible to anthelmintic drugs. Migration inhibition assays with A. suum larvae demonstrated that the potency of levamisole (LEV) and ivermectin (IVM) were significantly increased in the presence of PAC purified from pine bark (4.6-fold and 3.2-fold reduction in IC50 value for LEV and IVM, respectively). Synergy analysis revealed that the relationship between PAC and LEV appeared to be synergistic in nature, suggesting a specific enhancement of LEV activity, whilst the relationship between PAC and IVM was additive rather than synergistic, suggesting independent actions. Our results demonstrate that these common dietary compounds may increase the efficacy of synthetic anthelmintic drugs in vitro, and also suggest one possible mechanism for their well-known anti-parasitic activity. PMID:27094225

  1. N-acetylglucosamine-1-Phosphate Transferase Suppresses Lysosomal Hydrolases in Dysfunctional Osteoclasts: A Potential Mechanism for Vascular Calcification

    Directory of Open Access Journals (Sweden)

    Yang Lei

    2015-04-01

    Full Text Available In addition to increased differentiation of vascular smooth muscle cells into osteoblast-like phenotypes, the limited accumulation of osteoclasts in atherosclerotic plaques or their dysfunction may participate in potential mechanisms for vascular calcification. N-acetylglucosamine-1-phosphate transferase containing alpha and beta subunits (GNPTAB is a transmembrane enzyme complex that mediates the vesicular transport of lysosomal hydrolases. GNPTAB may also regulate the biogenesis of lysosomal hydrolases from bone-marrow derived osteoclasts. In this study, the areas surrounding calcification in human atherosclerotic plaques contained high levels of GNPTAB and low levels of lysosomal hydrolases such as cathepsin K (CTSK and tartrate-resistant acid phosphatase (TRAP, as demonstrated by immunohistochemistry and laser-capture microdissection-assisted mRNA expression analysis. We therefore hypothesized that GNPTAB secretion may suppress the release of CTSK and TRAP by vascular osteoclast-like cells, thus causing their dysfunction and reducing the resorption of calcification. We used human primary macrophages derived from peripheral blood mononuclear cells, an established osteoclast differentiation model. GNPTAB siRNA silencing accelerated the formation of functional osteoclasts as detected by increased secretion of CTSK and TRAP and increased their bone resorption activity as gauged by resorption pits assay. We concluded that high levels of GNPTAB inhibit secretion of lysosomal hydrolases in dysfunctional osteoclasts, thereby affecting their resorption potential in cardiovascular calcification.

  2. Substrate specificity combined with stereopromiscuity in glutathione transferase A4-4-dependent metabolism of 4-hydroxynonenal.

    Science.gov (United States)

    Balogh, Larissa M; Le Trong, Isolde; Kripps, Kimberly A; Shireman, Laura M; Stenkamp, Ronald E; Zhang, Wei; Mannervik, Bengt; Atkins, William M

    2010-02-23

    Conjugation to glutathione (GSH) by glutathione transferase A4-4 (GSTA4-4) is a major route of elimination for the lipid peroxidation product 4-hydroxynonenal (HNE), a toxic compound that contributes to numerous diseases. Both enantiomers of HNE are presumed to be toxic, and GSTA4-4 has negligible stereoselectivity toward them, despite its high catalytic chemospecificity for alkenals. In contrast to the highly flexible, and substrate promiscuous, GSTA1-1 isoform that has poor catalytic efficiency with HNE, GSTA4-4 has been postulated to be a rigid template that is preorganized for HNE metabolism. However, the combination of high substrate chemoselectivity and low substrate stereoselectivity is intriguing. The mechanism by which GSTA4-4 achieves this combination is important, because it must metabolize both enantiomers of HNE to efficiently detoxify the biologically formed mixture. The crystal structures of GSTA4-4 and an engineered variant of GSTA1-1 with high catalytic efficiency toward HNE, cocrystallized with a GSH-HNE conjugate analogue, demonstrate that GSTA4-4 undergoes no enantiospecific induced fit; instead, the active site residue Arg15 is ideally located to interact with the 4-hydroxyl group of either HNE enantiomer. The results reveal an evolutionary strategy for achieving biologically useful stereopromiscuity toward a toxic racemate, concomitant with high catalytic efficiency and substrate specificity toward an endogenously formed toxin. PMID:20085333

  3. Designer xanthone: an inhibitor scaffold for MDR-involved human glutathione transferase isoenzyme A1-1.

    Science.gov (United States)

    Zoi, Ourania G; Thireou, Trias N; Rinotas, Vagelis E; Tsoungas, Petros G; Eliopoulos, Elias E; Douni, Eleni K; Labrou, Nikolaos E; Clonis, Yannis D

    2013-10-01

    Glutathione transferases (GSTs) are cell detoxifiers involved in multiple drug resistance (MDR), hampering the effectiveness of certain anticancer drugs. To our knowledge, this is the first report on well-defined synthetic xanthones as GST inhibitors. Screening 18 xanthones revealed three derivatives bearing a bromomethyl and a methyl group (7) or two bromomethyl groups (8) or an aldehyde group (17), with high inhibition potency (>85%), manifested by low IC(50) values (7: 1.59 ± 0.25 µM, 8: 5.30 ± 0.30 µM, and 17: 8.56 ± 0.14 µM) and a competitive modality of inhibition versus CDNB (Ki(7) = 0.76 ± 0.18 and Ki(17) = 1.69 ± 0.08 µM). Of them, derivative 17 readily inhibited hGSTA1-1 in colon cancer cell lysate (IC(50) = 10.54 ± 2.41 µM). Furthermore, all three derivatives were cytotoxic to Caco-2 intact cells, with 17 being the least cytotoxic (LC(50) = 151.3 ± 16.3 µM). The xanthone scaffold may be regarded as a pharmacophore for hGSTA1-1 and the three derivatives, especially 17, as potent precursors for the synthesis of new inhibitors and conjugate prodrugs for human GSTs. PMID:23749766

  4. Influence of genetic polymorphisms of glutathione S-transferases T1 and M1 on serum lipid parameters

    International Nuclear Information System (INIS)

    To determine the effect of genetic polymorphisms of glutathione S-transferase theta1 (GSTT1) and GSTM1 on serum levels of lipid parameters. We conducted this cross-sectional study on 152 adult healthy subjects (54 females and 98 males) from January 2004 to September 2004. The participants in our study were recruited from the Research Clinic in Abarku (Yazd province, central part of Iran). There were unrelated Iranian Muslims. The genotypes of GSTT1 and GSTM1 were determined using polymerase chain reaction based method. After an overnight fasting, serum lipid indices including triglyceride (TG), total cholesterol (TC) and high density lipoprotein cholesterol (HDL-C) were measured. There were significant partial correlation coefficients between levels of TG (r= -0.48333, df=48, p<0.0001) and TG/HDL-C ratio (r=-0.4041, df=48, p=0.004) and numbers of active GST GST genotypes in females after controlling for age and body mass index (BMI). In males, the level of TG increased as a function of numbers of active GST genotypes after controlling for age and BMI (r=+0.2082, df=94, p=0.042). There were significant differences between females and males. Data show that genetic polymorphisms of GSTM1 and GSTTT1 modulate levels of TG and TG/HDL-C in females. (author)

  5. The Glutathione-S-Transferase, Cytochrome P450 and Carboxyl/Cholinesterase Gene Superfamilies in Predatory Mite Metaseiulus occidentalis

    Science.gov (United States)

    Hoy, Marjorie A.

    2016-01-01

    Pesticide-resistant populations of the predatory mite Metaseiulus (= Typhlodromus or Galendromus) occidentalis (Arthropoda: Chelicerata: Acari: Phytoseiidae) have been used in the biological control of pest mites such as phytophagous Tetranychus urticae. However, the pesticide resistance mechanisms in M. occidentalis remain largely unknown. In other arthropods, members of the glutathione-S-transferase (GST), cytochrome P450 (CYP) and carboxyl/cholinesterase (CCE) gene superfamilies are involved in the diverse biological pathways such as the metabolism of xenobiotics (e.g. pesticides) in addition to hormonal and chemosensory processes. In the current study, we report the identification and initial characterization of 123 genes in the GST, CYP and CCE superfamilies in the recently sequenced M. occidentalis genome. The gene count represents a reduction of 35% compared to T. urticae. The distribution of genes in the GST and CCE superfamilies in M. occidentalis differs significantly from those of insects and resembles that of T. urticae. Specifically, we report the presence of the Mu class GSTs, and the J’ and J” clade CCEs that, within the Arthropoda, appear unique to Acari. Interestingly, the majority of CCEs in the J’ and J” clades contain a catalytic triad, suggesting that they are catalytically active. They likely represent two Acari-specific CCE clades that may participate in detoxification of xenobiotics. The current study of genes in these superfamilies provides preliminary insights into the potential molecular components that may be involved in pesticide metabolism as well as hormonal/chemosensory processes in the agriculturally important M. occidentalis. PMID:27467523

  6. Cloning and expression of alpha class glutathione S-transferase gene from the small hermaphroditic fish Rivulus marmoratus (Cyprinodontiformes, Rivulidae).

    Science.gov (United States)

    Lee, Young-Mi; Chang, Sung Yeoul; Jung, Sang-Oun; Kweon, Hee-Seok; Lee, Jae-Seong

    2005-01-01

    In order to assess its potential as a biomarker of aquatic pollution, an alpha class glutathione S-transferase gene (GSTalpha gene) was cloned from the small hermaphroditic fish Rivulus marmoratus. The R. marmoratus GSTalpha gene spanned 1.3 kb, consisting of 6 exons encoding 221 amino acid residues. It showed high similarity to zebrafish GST. We named this R. marmoratus GSTalpha gene as rm-GSTalpha. The cDNA of the rm-GSTalpha gene was also investigated for its phylogeny, tissue-specific and chemical-induced expression. Rm-GSTalpha was subcloned into a 6 x His-tagged pCRT7 TOPO TA expression vector to produce the recombinant 6 x His-tagged rm-GST protein. This will be used in future to raise an rm-GSTalpha antibody for use in the study of phase II metabolism involved in detoxification. We also exposed R. marmoratus to 300 microg/l of 4-nonylphenol in water, and found approximately 4-fold induction of R. marmoratus GSTalpha mRNA in the treated animals. In this paper, we discuss the characteristics of the R. marmoratus GSTalpha gene as well as its potential use in relation to environmental pollution. PMID:16081109

  7. Purification and properties of an O-acetyl-transferase from Escherichia coli that can O-acetylate polysialic acid sequences

    International Nuclear Information System (INIS)

    Certain strains of bacteria synthesize an outer polysialic acid (K1) capsule. Some strains of K1+ E.coli are also capable of adding O-acetyl-esters to the exocyclic hydroxyl groups of the sialic acid residues. Both the capsule and the O-acetyl modification have been correlated with differences in antigenicity and pathogenicity. The authors have developed an assay for an O-acetyl-transferase in E.coli that transfers O-[3H]acetyl groups from [3H]acetyl-Coenzyme A to colominic acid (fragments of the polysialic acid capsule). Using this assay, the enzyme was solubilized, and purified ∼ 600-fold using a single affinity chromatography step with Procion Red-A Agarose. The enzyme also binds to Coenzyme A Sepharose, and can be eluted with high salt or Coenzyme A. The partially purified enzyme has a pH optimum of 7.0 - 7.5, is unaffected by divalent cations, is inhibited by high salt concentrations, is inhibited by Coenzyme A (50% inhibition at 100 μM), and shows an apparent Km for colominic acid of 3.7 mM (sialic acid concentration). This enzyme could be involved in the O-acetyl +/- form variation seen in some strains of K1+ E.coli

  8. Pigs fed camelina meal increase hepatic gene expression of cytochrome 8b1, aldehyde dehydrogenase, and thiosulfate transferase.

    Science.gov (United States)

    Meadus, William Jon; Duff, Pascale; McDonald, Tanya; Caine, William R

    2014-01-01

    Camelina sativa is an oil seed crop which can be grown on marginal lands. Camelina seed oil is rich in omega-3 fatty acids (>35%) and γ-tocopherol but is also high in erucic acid and glucosinolates. Camelina meal, is the by-product after the oil has been extracted. Camelina meal was fed to 28 d old weaned pigs at 3.7% and 7.4% until age 56 d. The camelina meal supplements in the soy based diets, improved feed efficiency but also significantly increased the liver weights. Gene expression analyses of the livers, using intra-species microarrays, identified increased expression of phase 1 and phase 2 drug metabolism enzymes. The porcine versions of the enzymes were confirmed by real time PCR. Cytochrome 8b1 (CYP8B1), aldehyde dehydrogenase 2 (Aldh2), and thiosulfate transferase (TST) were all significantly stimulated. Collectively, these genes implicate the camelina glucosinolate metabolite, methyl-sulfinyldecyl isothiocyanate, as the main xeniobiotic, causing increased hepatic metabolism and increased liver weight. PMID:24383433

  9. Pummelo Protects Doxorubicin-Induced Cardiac Cell Death by Reducing Oxidative Stress, Modifying Glutathione Transferase Expression, and Preventing Cellular Senescence

    Directory of Open Access Journals (Sweden)

    L. Chularojmontri

    2013-01-01

    Full Text Available Citrus flavonoids have been shown to reduce cardiovascular disease (CVD risks prominently due to their antioxidant effects. Here we investigated the protective effect of pummelo (Citrus maxima, CM fruit juice in rat cardiac H9c2 cells against doxorubicin (DOX- induced cytotoxicity. Four antioxidant compositions (ascorbic acid, hesperidin, naringin, and gallic acid were determined by HPLC. CM significantly increased cardiac cell survival from DOX toxicity as evaluated by MTT assay. Reduction of cellular oxidative stress was monitored by the formation of DCF fluorescent product and total glutathione (GSH levels. The changes in glutathione-S-transferase (GST activity and expression were determined by enzyme activity assay and Western blot analysis, respectively. Influence of CM on senescence-associated β-galactosidase activity (SA-β-gal was also determined. The mechanisms of cytoprotection involved reduction of intracellular oxidative stress, maintaining GSH availability, and enhanced GST enzyme activity and expression. DOX-induced cellular senescence was also attenuated by long-term CM treatment. Thus, CM fruit juice can be promoted as functional fruit to protect cells from oxidative cell death, enhance the phase II GSTP enzyme activity, and decrease senescence phenotype population induced by cardiotoxic agent such as DOX.

  10. The Putative O-Linked N-Acetylglucosamine Transferase SPINDLY Inhibits Class I TCP Proteolysis to Promote Sensitivity to Cytokinin.

    Science.gov (United States)

    Steiner, Evyatar; Livne, Sivan; Kobinson-Katz, Tammy; Tal, Lior; Pri-Tal, Oded; Mosquna, Assaf; Tarkowská, Danuše; Mueller, Bruno; Tarkowski, Petr; Weiss, David

    2016-06-01

    Arabidopsis (Arabidopsis thaliana) SPINDLY (SPY) is a putative serine and threonine O-linked N-acetylglucosamine transferase (OGT). While SPY has been shown to suppress gibberellin signaling and to promote cytokinin (CK) responses, its catalytic OGT activity was never demonstrated and its effect on protein fate is not known. We previously showed that SPY interacts physically and functionally with TCP14 and TCP15 to promote CK responses. Here, we aimed to identify how SPY regulates TCP14/15 activities and how these TCPs promote CK responses. We show that SPY activity is required for TCP14 stability. Mutation in the putative OGT domain of SPY (spy-3) stimulated TCP14 proteolysis by the 26S proteasome, which was reversed by mutation in CULLIN1 (CUL1), suggesting a role for SKP, CUL1, F-box E3 ubiquitin ligase in TCP14 proteolysis. TCP14 proteolysis in spy-3 suppressed all TCP14 misexpression phenotypes, including the enhanced CK responses. The increased CK activity in TCP14/15-overexpressing flowers resulted from increased sensitivity to the hormone and not from higher CK levels. TCP15 overexpression enhanced the response of the CK-induced synthetic promoter pTCS to CK, suggesting that TCP14/15 affect early steps in CK signaling. We propose that posttranslational modification of TCP14/15 by SPY inhibits their proteolysis and that the accumulated proteins promote the activity of the CK phosphorelay cascade in developing Arabidopsis leaves and flowers. PMID:27208284

  11. Genome-Wide Analysis of the Glutathione S-Transferase Gene Family in Capsella rubella: Identification, Expression, and Biochemical Functions

    Science.gov (United States)

    He, Gang; Guan, Chao-Nan; Chen, Qiang-Xin; Gou, Xiao-Jun; Liu, Wei; Zeng, Qing-Yin; Lan, Ting

    2016-01-01

    Extensive subfunctionalization might explain why so many genes have been maintained after gene duplication, which provides the engine for gene family expansion. However, it is still a particular challenge to trace the evolutionary dynamics and features of functional divergences in a supergene family over the course of evolution. In this study, we identified 49 Glutathione S-transferase (GST) genes from the Capsella rubella, a close relative of Arabidopsis thaliana and a member of the mustard family. Capsella GSTs can be categorized into eight classes, with tau and phi GSTs being the most numerous. The expansion of the two classes mainly occurs through tandem gene duplication, which results in tandem-arrayed gene clusters on chromosomes. By integrating phylogenetic analysis, expression patterns, and biochemical functions of Capsella and Arabidopsis GSTs, functional divergence, both in gene expression and enzymatic properties, were clearly observed in paralogous gene pairs in Capsella (even the most recent duplicates), and orthologous GSTs in Arabidopsis/Capsella. This study provides functional evidence for the expansion and organization of a large gene family in closely related species.

  12. Prognostic value of serum γ-glutamyl transferase in unresectable hepatocellular carcinoma patients treated with transcatheter arterial chemoembolization combined with conformal radiotherapy

    OpenAIRE

    Chen, Dong; Wang, Renben; Meng, Xiangjiao; YAN, HONGJIANG; JIANG, SHUMEI; Feng, Rui; ZHU, KUNLI; Xu, Xiaoqing; Dou, Xue; JIN, LINZHI

    2014-01-01

    The detection of γ-glutamyl transferase (GGT) has previously been reported to be useful in the diagnosis in hepatocellular carcinoma (HCC). The aim of the present study was to investigate the baseline serum GGT levels in patients with intermediate HCC (Barcelona Clinic Liver Cancer stage B) following treatment with transcatheter arterial chemoembolization (TACE) combined with three-dimensional conformal radiotherapy (3DCRT). A total of 154 intermediate HCC patients with Child-Pugh grade A wer...

  13. Gamma-glutamyl transferase and C-reactive protein as alternative markers of metabolic abnormalities and their associated comorbidites: a prospective cohort study

    OpenAIRE

    Melvin, Jennifer C; Rodrigues, Crystal; Holmberg, Lars; Garmo, Hans; Hammar, Niklas; Jungner, Ingmar; Walldius, Göran; Lambe, Mats; Jassem, Wayel; Van Hemelrijck, Mieke

    2012-01-01

    Background: Recent studies suggested that gamma-glutamyl transferase (GGT) and C-reactive protein (CRP) are good markers of metabolic abnormalities. We assessed the link between GGT, CRP and common metabolic abnormalities, as well their link to related diseases, such as cancer and cardiovascular disease (CVD). Methods: We selected 333,313 subjects with baseline measurements of triglycerides (TG), total cholesterol (TC), glucose, GGT and CRP in the Swedish AMORIS study. Baseline measurement of...

  14. Purification, molecular cloning, and characterization of glutathione S-transferases (GSTs) from pigmented Vitis vinifera L. cell suspension cultures as putative anthocyanin transport proteins

    OpenAIRE

    Conn, Simon; Curtin, Chris; Bézier, Annie; Franco, Chris; Zhang, Wei

    2008-01-01

    The ligandin activity of specific glutathione S-transferases (GSTs) is necessary for the transport of anthocyanins from the cytosol to the plant vacuole. Five GSTs were purified from Vitis vinifera L. cv. Gamay Fréaux cell suspension cultures by glutathione affinity chromatography. These proteins underwent Edman sequencing and mass spectrometry fingerprinting, with the resultant fragments aligned with predicted GSTs within public databases. The corresponding coding sequences were cloned, with...

  15. Elevated Levels of Urinary 8-Hydroxy-2′-deoxyguanosine, Lymphocytic Micronuclei, and Serum Glutathione S-Transferase in Workers Exposed to Coke Oven Emissions

    OpenAIRE

    Liu, Ai-Lin; Lu, Wen-Qing; Wang, Zeng-Zhen; Chen, Wei-Hong; Lu, Wen-Hong; Yuan, Jing; Nan, Pei-Hong; Sun, Jian-Ya; Zou, Ya-Lin; Zhou, Li-Hong; Zhang, Chi; Wu, Tang-chun

    2005-01-01

    To investigate associations among occupational exposure to coke oven emissions (COEs), oxidative stress, cytogenotoxic effects, change in the metabolizing enzyme glutathione S-transferase (GST), and internal levels of polycyclic aromatic hydrocarbons (PAHs) in coke oven workers, we recruited 47 male coke oven workers and 31 male control subjects from a coke oven plant in northern China. We measured the levels of 1-hydroxypyrene (1-OHP) and 8-hydroxy-2′-deoxyguanosine (8-OHdG) in urine, micron...

  16. Glutathione-S-transferase subtypes α and π as a tool to predict and monitor graft failure or regeneration in a pilot study of living donor liver transplantation

    OpenAIRE

    Jochum C; Beste M; Sowa J-P; Farahani MS; Penndorf V; Nadalin S; Saner F; Canbay A; Gerken G

    2011-01-01

    Abstract Objective Glutathione-S-Transferase (GST) subtype α and π are differentially expressed in adult liver tissue. Objective of the study was if GST α and p may serve as predictive markers for liver surgery, especially transplantations. Methods 13 patients receiving living donor liver transplantation (LDLT) and their corresponding donors were analyzed for standard serum parameters (ALT, AST, gGT, bilirubin) as well as GST-α and -π before LDLT and daily for 10 days after LDLT. Patients (R)...

  17. 28-Homobrassinolide Alters Protein Content and Activities of Glutathione-S-Transferase and Polyphenol Oxidase in Raphanus Sativus L. Plants Under Heavy Metal Stress

    OpenAIRE

    Sharma, Neha; Hundal, Gurjinder Singh; Sharma, Indu; Bhardwaj, Renu

    2014-01-01

    Objectives: The application of brassinosteroids (BRs), the plant steroidal hormones, results in an increased tolerance toward stress and thus helps improving the yield of crop plants. The present study was carried out to investigate the effect of 28-homobrassinolide (28-HBL) on the protein content as well as activities of antioxidant enzymes viz., glutathione-s-transferase (GST) and polyphenol oxidase (PPO) in radish plants grown under Cadmium (Cd) and Mercury (Hg) metal stress. Materials and...

  18. Analysis of Common Mutations in the Galactose-1-Phosphate Uridyl Transferase Gene : New Assays to Increase the Sensitivity and Specificity of Newborn Screening for Galactosemia

    OpenAIRE

    Dobrowolski, Steven F.; Banas, Richard A.; Suzow, Joseph G.; Berkley, Michelle; Naylor, Edwin W.

    2003-01-01

    Classical galactosemia is a genetic disease caused by mutations in the galactose-1-phosphate uridyl transferase (GALT) gene. Prospective newborn screening for galactosemia is routine and utilizes the universally collected newborn dried blood specimen on filter paper. Screening for galactosemia is achieved through analysis of total galactose (galactose and galactose-1-phosphate) and/or determining the activity of the GALT enzyme. While this approach is effective, en vironmental factors and the...

  19. Glutathione S-transferase activity and isoenzyme composition in benign ovarian tumours, untreated malignant ovarian tumours, and malignant ovarian tumours after platinum/cyclophosphamide chemotherapy.

    OpenAIRE

    Zee, A.G. Van der; van Ommen, B.; Meijer, C; Hollema, H; Bladeren, P.J. van; de Vries, E. G.

    1992-01-01

    Glutathione S-transferase (GST) isoenzyme composition, isoenzyme quantities and enzymatic activity were investigated in benign (n = 4) ovarian tumours and malignant ovarian tumours, before (n = 20) and after (n = 16) chemotherapy. Enzymatic activity of GST in cytosols was measured by determining 1-chloro-2,4-dinitrobenzene conjugation with glutathione, cytosolic GST subunits were determined by wide pore reversed phase HPLC, using a S-hexylglutathione-agarose affinity column, and isoelectric f...

  20. Tissue and Life Stage Specificity of Glutathione S-Transferase Expression in the Hessian Fly, Mayetiola destructor: Implications for Resistance to Host Allelochemicals

    OpenAIRE

    Mittapalli, Omprakash; Neal, Jonathan J.; Shukle, Richard H

    2007-01-01

    Two new Delta and Sigma glutathione S-transferases (GSTs) in the Hessian fly, Mayetiola destructor (Diptera: Cecidomyiidae), were characterized and transcription profiles described. The deduced amino acid sequences for the two M. destructor Delta GSTs (MdesGST-1 and MdesGST-3) showed high similarity with other insect Delta GSTs including the conserved catalytic serine residue. The deduced amino acid sequence for the M. destructor Sigma GST (MdesGST-2) showed high similarity with other insect ...

  1. Loss of ICG uptake in the process of rat hepatocarcinogenesis correlates to the disappearance of glutathione-S-transferase alpha subunit.

    OpenAIRE

    Ling, Liu; Higashi,Toshihiro; Tsuchida, Shigeki; Sato, Kiyomi; Tsuji, Takao

    1993-01-01

    Reduced indocyanine green (ICG) uptake is one of the functional changes of human hepatocellular carcinoma (HCC). To clarify the mechanisms of loss of ICG uptake, and determine which subunit of glutathione-S-transferase (GST), alpha or pi, plays a role in ICG transport in hepatocytes, an experimental HCC model was developed that used nodules induced by 2-acetylamino-fluorene (2-AAF) administration. Many of the ICG stained nodules, which consisted of benign and borderline lesions, were GST-alph...

  2. G9a, a putative histone methyl-transferase in Drosophila interacts with Tungus, a protein associated with α-Actinin

    OpenAIRE

    2005-01-01

    Histone lysine methylation is considered to be a relatively stable modification associated with important functions in epigenetic gene control and for organizing chromatin domains. Genes encoding mammalian homologues of the Drosophila suppressor of PEV Su(var)3-9 were the first shown to encode proteins with histone lysine methyl-transferase (HKMT) activity. A hallmark signature of this class of proteins is the evolutionary conserved SET-domain found in numerous chromatin regulators, and was n...

  3. Functional Characterization of the Tau Class Glutathione-S-Transferases Gene (SbGSTU) Promoter of Salicornia brachiata under Salinity and Osmotic Stress

    OpenAIRE

    Tiwari, Vivekanand; Patel, Manish Kumar; Chaturvedi, Amit Kumar; Mishra, Avinash; Jha, Bhavanath

    2016-01-01

    Reactive oxygen or nitrogen species are generated in the plant cell during the extreme stress condition, which produces toxic compounds after reacting with the organic molecules. The glutathione-S-transferase (GST) enzymes play a significant role to detoxify these toxins and help in excretion or sequestration of them. In the present study, we have cloned 1023 bp long promoter region of tau class GST from an extreme halophyte Salicornia brachiata and functionally characterized using the transg...

  4. Transcriptional and Functional Analysis of Oxalyl-Coenzyme A (CoA) Decarboxylase and Formyl-CoA Transferase Genes from Lactobacillus acidophilus

    OpenAIRE

    Azcarate-Peril, M. Andrea; Bruno-Bárcena, Jose M.; Hassan, Hosni M.; Klaenhammer, Todd R.

    2006-01-01

    Oxalic acid is found in dietary sources (such as coffee, tea, and chocolate) or is produced by the intestinal microflora from metabolic precursors, like ascorbic acid. In the human intestine, oxalate may combine with calcium, sodium, magnesium, or potassium to form less soluble salts, which can cause pathological disorders such as hyperoxaluria, urolithiasis, and renal failure in humans. In this study, an operon containing genes homologous to a formyl coenzyme A transferase gene (frc) and an ...

  5. Synthetic fragments of antigenic lipophosphoglycans from Leishmania major and Leishmania mexicana and their use for characterisation of the Leishmania elongating alpha-D-mannopyranosylphosphate transferase.

    Science.gov (United States)

    Higson, Adrian P; Ross, Andrew J; Tsvetkov, Yury E; Routier, Françoise H; Sizova, Olga V; Ferguson, Michael A J; Nikolaev, Andrei V

    2005-03-18

    The phosphorylated branched heptasaccharides 7 and 8, the octasaccharide 9 and the phosphorylated trisaccharides 5 and 6, which are fragments of the phosphoglycan portion of the surface lipophosphoglycans from Leishmania mexicana (5) or L. major (6-9), were synthesised by using the glycosyl hydrogenphosphonate method for the preparation of phosphodiester bridges. The compounds were tested as acceptor substrates/putative inhibitors for the Leishmania elongating alpha-D-mannosylphosphate transferase. PMID:15685582

  6. Role of glutathione S-transferase M1, T1 and P1 gene polymorphisms in childhood acute lymphoblastic leukemia susceptibility in a Turkish population

    OpenAIRE

    Mehmet Guven; Selin Unal; Duygu Erhan; Nihal Ozdemir; Safa Baris; Tiraje Celkan; Merve Bostancı; Bahadir Batar

    2015-01-01

    The variations between different individuals in the xenobiotic metabolizing enzymes' activity were shown to modify susceptibility to childhood acute lymphoblastic leukemia (ALL). Polymorphisms associated with genes coding for the glutathione S-transferase (GST) enzyme were known to affect the metabolism of different carcinogens. The aim of this study was to evaluate the influence of the GSTM1 and GSTT1 deletion polymorphisms, and the GSTP1 Ile105Val single nucleotide polymorphism (SNP) on the...

  7. Minor Modifications of the C-terminal Helix Reschedule the Favored Chemical Reactions Catalyzed by Theta Class Glutathione Transferase T1-1*

    OpenAIRE

    Shokeer, Abeer; Mannervik, Bengt

    2009-01-01

    Adaptive responses to novel toxic challenges provide selective advantages to organisms in evolution. Glutathione transferases (GSTs) play a pivotal role in the cellular defense because they are main contributors to the inactivation of genotoxic compounds of exogenous as well as of endogenous origins. GSTs are promiscuous enzymes catalyzing a variety of chemical reactions with numerous alternative substrates. Despite broad substrate acceptance, individual GSTs display pronounced selectivities ...

  8. Controlled ribonucleotide tailing of cDNA ends (CRTC) by terminal deoxynucleotidyl transferase: a new approach in PCR-mediated analysis of mRNA sequences.

    OpenAIRE

    Schmidt, W. M.; Mueller, M W

    1996-01-01

    Controlled ribonucleotide tailing of cDNA ends (CRTC) by terminal deoxynucleotidyl transferase is a polymerase chain reaction (PCR)-mediated technique that was developed to facilitate cloning and direct sequence analysis of complete 5'-terminal unknown coding regions of rare RNA molecules. In contrast with standard tailing protocols using dNTPs as the substrate, ribo-tailing of cDNA ends is easily controllable, self-limited (from two to four rNMP incorporations) and highly efficient (>98%). B...

  9. Decreased glutathione content and glutathione S-transferase activity in red blood cells of coal miners with early stages of pneumoconiosis.

    OpenAIRE

    Evelo, C T; Bos, R P; Borm, P J

    1993-01-01

    Blood samples of miners heavily exposed to coal dust were examined for changes in glutathione S-transferase (GST) activity. Decreased GST activity was found in red blood cells of subjects with early stages of coal workers' pneumoconiosis (International Labour Office classification 0/1-1/2) when compared with control miners. At further progression of coal workers' pneumoconiosis (> or = 2/1), the activity of GST was not different from controls. In the same group with moderate coal workers' pne...

  10. Immunohistochemical expression of pi class glutathione S-transferase in the basal cell layer of benign prostate tissue following chronic treatment with finasteride.

    OpenAIRE

    Montironi, R; Mazzucchelli, R; Pomante, R; Thompson, D.; Duval da Silva, V; Vaught, L; Bartels, P H

    1999-01-01

    BACKGROUND: Glutathione S-transferases (GST) may prevent carcinogenesis through inactivation of reactive electrophiles by conjugation to reduced glutathione. Treatment directed at the induction or preservation of GST-pi expression in normal epithelium could have a profound impact on the prevention of prostate neoplasia. Finasteride, a 5-alpha-reductase inhibitor, is used as a chemopreventive agent because it blocks the conversion of testosterone to its byproduct which promotes prostate tumour...

  11. Glutathione-S-transferase genes and asthma phenotypes: a Human Genome Epidemiology (HuGE) systematic review and meta-analysis including unpublished data

    OpenAIRE

    Minelli, Cosetta; Granell, Raquel; Newson, Roger; Rose-Zerilli, Matthew J; Torrent, Maties; Ring, Sue M; Holloway, John W; Shaheen, Seif O.; Henderson, John A.

    2010-01-01

    Background: oxidative stress is thought to be involved in the pathogenesis of asthma. Glutathione-S-transferase (GST) enzymes, which play an important role in antioxidant defences, may therefore influence asthma risk. Two common deletion polymorphisms of GSTM1 and GSTT1 genes and the GSTP1 Ile105Val polymorphism have been associated with asthma in children and adults, but results are inconsistent across studies. Methods: systematic review and meta-analysis of the effects of GST genes on a...

  12. Glutathione S-transferase class mu regulation of apoptosis signal-related kinase 1 protein during VCD-induced ovotoxicity in neonatal rat ovaries

    OpenAIRE

    Bhattacharya, Poulomi; Madden, Jill A.; Sen, Nivedita; Hoyer, Patricia B.; Keating, Aileen F.

    2012-01-01

    4-vinylcyclohexene diepoxide (VCD) destroys ovarian primordial and small primary follicles via apoptosis. In mice, VCD exposure induces ovarian mRNA expression of glutathione S-transferase (GST) family members, including isoform mu (Gstm). Extra-ovarian GSTM negatively regulates pro-apoptotic apoptosis signal-related kinase 1 (ASK1) through protein complex formation, which dissociates during stress, thereby initiating ASK1-induced apoptosis. The present study investigated the ovarian response...

  13. Two pear glutathione S-transferases genes are regulated during fruit development and involved in response to salicylic acid, auxin, and glucose signaling.

    Directory of Open Access Journals (Sweden)

    Hai-Yan Shi

    Full Text Available Two genes encoding putative glutathione S-transferase proteins were isolated from pear (Pyrus pyrifolia and designated PpGST1 and PpGST2. The deduced PpGST1 and PpGST2 proteins contain conserved Glutathione S-transferase N-terminal domain (GST_N and Glutathione S-transferase, C-terminal domain (GST_C. Using PCR amplification technique, the genomic clones corresponding to PpGST1 and PpGST2 were isolated and shown to contain two introns and a singal intron respectively with typical GT/AG boundaries defining the splice junctions. Phylogenetic analysis clearly demonstrated that PpGST1 belonged to Phi class of GST superfamilies and had high homology with apple MdGST, while PpGST2 was classified into the Tau class of GST superfamilies. The expression of PpGST1 and PpGST2 genes was developmentally regulated in fruit. Further study demonstrated that PpGST1 and PpGST2 expression was remarkably induced by glucose, salicylic acid (SA and indole-3-aceticacid (IAA treatments in pear fruit, and in diseased fruit. These data suggested that PpGST1 and PpGST2 might be involved in response to sugar, SA, and IAA signaling during fruit development of pear.

  14. Parâmetros cinéticos da Glutationa S-Transferase e sua ativação por extratos de vegetais Kinetics parameters of Glutathione S-Transferase and its activation by vegetable extracts

    Directory of Open Access Journals (Sweden)

    Maria Célia Lopes Torres

    2004-06-01

    Full Text Available Este estudo teve como objetivos avaliar a indução da Glutationa S-Transferase, com extratos de vegetais, e caracterizar os parâmetros cinéticos desta enzima. Foram obtidos os extratos aquoso, etanólico e hexanólico de vegetais, amplamente consumidos no Brasil, como berinjela (Solanum melongena L., couve-flor (Brassica oleracea L., couve (Brassica oleracea L., brócolis (Brassica oleracea L., couve-de-bruxelas (Brassicaoleraea L., cebola (Allium cepa L., alho (Allium sativum L.; vegetais que apresentam gosto amargo, como jiló (Solanum gilo Raddi, guariroba (Syagrus oleracea Becc., mostarda (Brassica nigra L., carqueja (Cacalia spp., e de plantas relacionadas, na cultura popular, como curadoras de determinadas doenças, como a babosa (Aloe vera L.. A atividade da enzima foi determinada usando como substrato o 1 cloro 2, 4 dinitrobenzeno, na presença dos extratos vegetais. A mistura da reação, sem a presença do extrato, foi considerada controle. Das amostras de vegetais avaliadas, a berinjela, a couve e o brócolis apresentaram maior indução na atividade da GST, sendo o extrato etanólico o mais eficaz. A enzima apresentou um Vmax de 0,016 abs. min-1/unidade da enzima e um Km de 0,323mM. O baixo valor de Km encontrado indica uma alta especificidade da enzima pelo substrato 1 cloro 2, 4 dinitrobenzeno e a atividade máxima da enzima foi na faixa de pH entre 6,5 e 7,0.This study was done to evaluate induction Glutathione S-Transferase, with vegetable extracts, and characterize its kinetics parameters. The aqueous, alcoholic, and hexanoic extracts were obtained from vegetables widely consumed in Brazil: eggplant (Solanum melongena L., cauliflower (Brassica oleracea L., cauli leaves (Brassica oleracea L., broccoli (Brassica oleracea L., Brussels sprout (Brassicaoleraea L., onions (Allium cepa L., garlic (Allium sativum L.; and bitter tasting vegetable such as jiló (Solanum gilo Raddi, guariroba (Syagrus oleracea Becc., black mustard

  15. Directed evolution of Tau class glutathione transferases reveals a site that regulates catalytic efficiency and masks co-operativity.

    Science.gov (United States)

    Axarli, Irine; Muleta, Abdi W; Vlachakis, Dimitrios; Kossida, Sophia; Kotzia, Georgia; Maltezos, Anastasios; Dhavala, Prathusha; Papageorgiou, Anastassios C; Labrou, Nikolaos E

    2016-03-01

    A library of Tau class GSTs (glutathione transferases) was constructed by DNA shuffling using the DNA encoding the Glycine max GSTs GmGSTU2-2, GmGSTU4-4 and GmGSTU10-10. The parental GSTs are >88% identical at the sequence level; however, their specificity varies towards different substrates. The DNA library contained chimaeric structures of alternated segments of the parental sequences and point mutations. Chimaeric GST sequences were expressed in Escherichia coli and their enzymatic activities towards CDNB (1-chloro-2,4-dinitrobenzene) and the herbicide fluorodifen (4-nitrophenyl α,α,α-trifluoro-2-nitro-p-tolyl ether) were determined. A chimaeric clone (Sh14) with enhanced CDNB- and fluorodifen-detoxifying activities, and unusual co-operative kinetics towards CDNB and fluorodifen, but not towards GSH, was identified. The structure of Sh14 was determined at 1.75 Å (1 Å=0.1 nm) resolution in complex with S-(p-nitrobenzyl)-glutathione. Analysis of the Sh14 structure showed that a W114C point mutation is responsible for the altered kinetic properties. This was confirmed by the kinetic properties of the Sh14 C114W mutant. It is suggested that the replacement of the bulky tryptophan residue by a smaller amino acid (cysteine) results in conformational changes of the active-site cavity, leading to enhanced catalytic activity of Sh14. Moreover, the structural changes allow the strengthening of the two salt bridges between Glu(66) and Lys(104) at the dimer interface that triggers an allosteric effect and the communication between the hydrophobic sites. PMID:26637269

  16. Genetic polymorphisms in glutathione S-transferase (GST) superfamily and arsenic metabolism in residents of the Red River Delta, Vietnam

    International Nuclear Information System (INIS)

    To elucidate the role of genetic factors in arsenic metabolism, we investigated associations of genetic polymorphisms in the members of glutathione S-transferase (GST) superfamily with the arsenic concentrations in hair and urine, and urinary arsenic profile in residents in the Red River Delta, Vietnam. Genotyping was conducted for GST ω1 (GSTO1) Ala140Asp, Glu155del, Glu208Lys, Thr217Asn, and Ala236Val, GST ω2 (GSTO2) Asn142Asp, GST π1 (GSTP1) Ile105Val, GST μ1 (GSTM1) wild/null, and GST θ1 (GSTT1) wild/null. There were no mutation alleles for GSTO1 Glu208Lys, Thr217Asn, and Ala236Val in this population. GSTO1 Glu155del hetero type showed higher urinary concentration of AsV than the wild homo type. Higher percentage of DMAV in urine of GSTM1 wild type was observed compared with that of the null type. Strong correlations between GSTP1 Ile105Val and arsenic exposure level and profile were observed in this study. Especially, heterozygote of GSTP1 Ile105Val had a higher metabolic capacity from inorganic arsenic to monomethyl arsenic, while the opposite trend was observed for ability of metabolism from AsV to AsIII. Furthermore, other factors including sex, age, body mass index, arsenic level in drinking water, and genotypes of As (+ 3 oxidation state) methyltransferase (AS3MT) were also significantly co-associated with arsenic level and profile in the Vietnamese. To our knowledge, this is the first study indicating the associations of genetic factors of GST superfamily with arsenic metabolism in a Vietnamese population.

  17. Purification and partial characterization of glutathione S-transferases from three field populations of Panonychus citri (Acari: Tetranychidae).

    Science.gov (United States)

    Niu, Jin-Zhi; Dou, Wei; Wang, Bao-Jun; Zhang, Guo-Na; Zhang, Rui; Yin, Yi; Wang, Jin-Jun

    2012-02-01

    Glutathione S-transferases (GSTs) play central roles in phase II detoxification of both xenobiotics (drugs, insecticides, and herbicides) and endogenous compounds in almost all living organisms. In this study, we successfully purified the GSTs from the citrus red mite, Panonychus citri, by affinity chromatography on Glutathione Sepharose 4B and compared the biochemical characterizations of the purified GSTs from three field populations [beibei (BB), wanzhou (WZ), and zhongxian (ZX)]. SDS-PAGE revealed that the molecular weight of GSTs from three populations consisted of two subunits of 27.3 and 26.1 kDa. The specific activity of the purified GSTs from the WZ and ZX populations was increased 1.5- and 3.8-fold, respectively, compared with the BB population. Accordingly, the pyridaben susceptibility of WZ and ZX populations was less compared with BB population. Kinetic analyses showed that the WZ and ZX populations had higher substrate specificity compared with the BB population based on the values of k (cat) and k (cat) /K (m) to both reduced glutathione (GSH) and 1-chloro-2,4-dinitrobenzene (CDNB). The in vitro inhibition studies of GSTs indicated that the I (50) values of pyridaben from WZ and ZX populations of P. citri expressed 1.6- and 4.4-fold decreases, respectively, compared to the I (50) value of pyridaben from the BB population. In conclusion, all evidence suggested that the purified GSTs may partially contribute to the susceptibility of acaricide pyridaben in field populations of P. citri. PMID:21979304

  18. Effects of high-intensity intermittent training on carnitine palmitoyl transferase activity in the gastrocnemius muscle of rats

    Energy Technology Data Exchange (ETDEWEB)

    Carnevali, L.C. Jr. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Centro Universitário Ítalo-Brasileiro (Unítalo), São Paulo SP (Brazil); Eder, R.; Lira, F.S. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Lima, W.P. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Instituto Federal de Educação,Ciência e Tecnologia de São Paulo, São Paulo SP (Brazil); Gonçalves, D.C. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Zanchi, N.E. [Laboratorio de Nutrição e Metabolismo Aplicado à Atividade Motora, Escola de Educação Física e Esporte, Universidade de São Paulo, São Paulo SP (Brazil); Centro de Pesquisa do Genoma Humano, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Nicastro, H. [Laboratorio de Nutrição e Metabolismo Aplicado à Atividade Motora, Escola de Educação Física e Esporte, Universidade de São Paulo, São Paulo SP (Brazil); Lavoie, J.M. [Department of Kinesiology, University of Montreal, Montreal (Canada); Seelaender, M.C.L. [Grupo de Biologia Molecular da Célula, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil)

    2012-06-29

    We examined the capacity of high-intensity intermittent training (HI-IT) to facilitate the delivery of lipids to enzymes responsible for oxidation, a task performed by the carnitine palmitoyl transferase (CPT) system in the rat gastrocnemius muscle. Male adult Wistar rats (160-250 g) were randomly distributed into 3 groups: sedentary (Sed, N = 5), HI-IT (N = 10), and moderate-intensity continuous training (MI-CT, N = 10). The trained groups were exercised for 8 weeks with a 10% (HI-IT) and a 5% (MI-CT) overload. The HI-IT group presented 11.8% decreased weight gain compared to the Sed group. The maximal activities of CPT-I, CPT-II, and citrate synthase were all increased in the HI-IT group compared to the Sed group (P < 0.01), as also was gene expression, measured by RT-PCR, of fatty acid binding protein (FABP; P < 0.01) and lipoprotein lipase (LPL; P < 0.05). Lactate dehydrogenase also presented a higher maximal activity (nmol·min{sup −1}·mg protein{sup −1}) in HI-IT (around 83%). We suggest that 8 weeks of HI-IT enhance mitochondrial lipid transport capacity thus facilitating the oxidation process in the gastrocnemius muscle. This adaptation may also be associated with the decrease in weight gain observed in the animals and was concomitant to a higher gene expression of both FABP and LPL in HI-IT, suggesting that intermittent exercise is a “time-efficient” strategy inducing metabolic adaptation.

  19. Comprehensive expression analysis suggests overlapping and specific roles of rice glutathione S-transferase genes during development and stress responses

    Directory of Open Access Journals (Sweden)

    Bhattacharjee Annapurna

    2010-01-01

    Full Text Available Abstract Background Glutathione S-transferases (GSTs are the ubiquitous enzymes that play a key role in cellular detoxification. Although several GSTs have been identified and characterized in various plant species, the knowledge about their role in developmental processes and response to various stimuli is still very limited. In this study, we report genome-wide identification, characterization and comprehensive expression analysis of members of GST gene family in crop plant rice, to reveal their function(s. Results A systematic analysis revealed the presence of at least 79 GST genes in the rice genome. Phylogenetic analysis grouped GST proteins into seven classes. Sequence analysis together with the organization of putative motifs indicated the potential diverse functions of GST gene family members in rice. The tandem gene duplications have contributed a major role in expansion of this gene family. Microarray data analysis revealed tissue-/organ- and developmental stage-specific expression patterns of several rice GST genes. At least 31 GST genes showed response to plant hormones auxin and cytokinin. Furthermore, expression analysis showed the differential expression of quite a large number of GST genes during various abiotic stress (20, arsenate stress (32 and biotic stress (48 conditions. Many of the GST genes were commonly regulated by developmental processes, hormones, abiotic and biotic stresses. Conclusion The transcript profiling suggests overlapping and specific role(s of GSTs during various stages of development in rice. Further, the study provides evidence for the role of GSTs in mediating crosstalk between various stress and hormone response pathways and represents a very useful resource for functional analysis of selected members of this family in rice.

  20. Glutathione, glutathione S-transferases, and related redox enzymes in Adriamycin-resistant cell lines with a multidrug resistant phenotype.

    Science.gov (United States)

    Schisselbauer, J C; Crescimanno, M; D'Alessandro, N; Clapper, M; Toulmond, S; Tapiero, H; Tew, K D

    1989-01-01

    Friend erythroleukemia cells (FLC) selected by exposure to Adriamycin (doxorubicin) express an approximate 2.5-fold (ARN1) or 13-fold (ARN2) resistance to the drug with various degrees of cross-resistance to other anthracyclines, vinca alkaloids, and epipodophyllotoxins. Because the redox cycling of the quinone moiety of Adriamycin is known to produce oxidative stress, however, an analysis of glutathione (GSH) and related enzyme systems was undertaken in the wild-type and selected resistant cells. In ARN1 and ARN2, superoxide dismutase (SOD) and catalase activities were slightly decreased, intracellular GSH and GSH reductase were essentially unchanged, and total GSH peroxidase, glutathione S-transferase (GST), and DT-diaphorase activities were slightly elevated. In each case there was no stoichiometric relationship between degree of resistance and level of activity. GST isozymes were purified from each cell line by HPLC GSH affinity column chromatography. Two-dimensional gel electrophoresis and western blot immunoreactivity against a battery of GST isozyme polyclonal antibodies determined that both the resistant and sensitive cells expressed isozymes of the alpha, pi, and mu classes (alternative murine nomenclature: M1, M2, M3). Of significance, both ARN1 and ARN2 cell lines expressed a unique alpha subunit which was absent from the parent FLC cell line. This isozyme presumably accounted for the increased GSH peroxidase activity (cumene hydroperoxide as substrate) found in ARN1 and ARN2 and may play a role in the small incremental resistance to melphalan found for both resistant lines. Expression of the isozyme was not stoichiometric with respect to degree of resistance. The presence of this isozyme may contribute to the resistant phenotype or may be the consequence of a more general cellular response to oxidative stress. PMID:2639724