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Sample records for acetyl xylan esterase

  1. Hydrolysis of Wheat Arabinoxylan by Two Acetyl Xylan Esterases from Chaetomium thermophilum

    DEFF Research Database (Denmark)

    Tong, Xiaoxue; Lange, Lene; Grell, Morten Nedergaard

    2015-01-01

    esterase genes, CtAxeA and CtAxeB, in Pichia pastoris. The recombinant proteins, rCtAxeA and rCtAxeB, released acetic acids from p-nitrophenyl acetate and water-insoluble wheat arabinoxylan. These results indicate that CtAxeA and CtAxeB are true acetyl xylan esterases. For both recombinant esterases, over...... 93 % of the initial activity was retained after 24 h of incubation at temperatures up to 60 °C, and over 90 % of the initial activity was retained after 24 h of incubation in different buffers from pH 4.0 to 9.0 at 4 and 50 °C. The overall xylose yield from wheat arabinoxylan hydrolysis was 8...

  2. Acetyl Xylan Esterase Axe1 (T. reesei, Carbohydrate Esterase Family 5) Supplemented to a (Hemi)cellulolytic Preparation Enhances Degradation of Recalcitrant Corn Silage Polysaccharides

    NARCIS (Netherlands)

    Neumüller, K.G.; Streekstra, H.; Gruppen, H.; Schols, H.A.

    2014-01-01

    The increase in hydrolytic activity towards corn silage water unextractable solids by supplementation of acetyl xylan esterase 1 (T. reesei, TrAxe1), belonging to arbohydrate esterase (CE) family 5, to an A. niger / T. emersonii enzyme preparation is presented. TrAxe1 was cloned and expressed in A

  3. An eleven amino acid residue deletion expands the substrate specificity of acetyl xylan esterase II (AXE II) from Penicillium purpurogenum

    Science.gov (United States)

    Colombres, Marcela; Garate, José A.; Lagos, Carlos F.; Araya-Secchi, Raúl; Norambuena, Patricia; Quiroz, Soledad; Larrondo, Luis; Pérez-Acle, Tomas; Eyzaguirre, Jaime

    2008-01-01

    The soft-rot fungus Penicillium purpurogenum secretes to the culture medium a variety of enzymes related to xylan biodegradation, among them three acetyl xylan esterases (AXE I, II and III). AXE II has 207 amino acids; it belongs to family 5 of the carbohydrate esterases and its structure has been determined by X-ray crystallography at 0.9 Å resolution (PDB 1G66). The enzyme possesses the α/β hydrolase fold and the catalytic triad typical of serine esterases (Ser90, His187 and Asp175). AXE II can hydrolyze esters of a large variety of alcohols, but it is restricted to short chain fatty acids. An analysis of its three-dimensional structure shows that a loop that covers the active site may be responsible for this strict specificity. Cutinase, an enzyme that hydrolyzes esters of long chain fatty acids and shows a structure similar to AXE II, lacks this loop. In order to generate an AXE II with this broader specificity, the preparation of a mutant lacking residues involving this loop (Gly104 to Ala114) was proposed. A set of molecular simulation experiments based on a comparative model of the mutant enzyme predicted a stable structure. Using site-directed mutagenesis, the loop's residues have been eliminated from the AXE II cDNA. The mutant protein has been expressed in Aspergillus nidulans A722 and Pichia pastoris, and it is active towards a range of fatty acid esters of up to at least 14 carbons. The availability of an esterase with broader specificity may have biotechnological applications for the synthesis of sugar esters.

  4. Determination of some significant batch culture conditions affecting acetyl-xylan esterase production by Penicillium notatum NRRL-1249

    Directory of Open Access Journals (Sweden)

    Akhtar MN

    2011-05-01

    Full Text Available Abstract Background Acetyl-xylan esterase (AXE, EC 3.1.1.72 hydrolyses acetate group from the linear chain of xylopyranose residues bound by β-1,4-linkage. The enzyme finds commercial applications in bio-bleaching of wood pulp, treating animal feed to increase digestibility, processing food to increase clarification and converting lignocellulosics to feedstock and fuel. In the present study, we report on the production of an extracellular AXE from Penicillium notatum NRRL-1249 by solid state fermentation (SSF. Results Wheat bran at a level of 10 g (with 4 cm bed height was optimized as the basal substrate for AXE production. An increase in enzyme activity was observed when 7.5 ml of mineral salt solution (MSS containing 0.1% KH2PO4, 0.05% KCl, 0.05% MgSO4.7H2O, 0.3% NaNO3, 0.001% FeSO4.2H2O and 0.1% (v/w Tween-80 as an initial moisture content was used. Various nitrogen sources including ammonium sulphate, urea, peptone and yeast extract were compared for enzyme production. Maximal enzyme activity of 760 U/g was accomplished which was found to be highly significant (p ≤ 0.05. A noticeable enhancement in enzyme activity was observed when the process parameters including incubation period (48 h, initial pH (5, 0.2% (w/w urea as nitrogen source and 0.5% (v/w Tween-80 as a stimulator were further optimized using a 2-factorial Plackett-Burman design. Conclusion From the results it is clear that an overall improvement of more than 35% in terms of net enzyme activity was achieved compared to previously reported studies. This is perhaps the first report dealing with the use of P. notatum for AXE production under batch culture SSF. The Plackett-Burman model terms were found highly significant (HS, suggesting the potential commercial utility of the culture used (df = 3, LSD = 0.126.

  5. Positional preferences of acetyl esterases from different CE families towards acetylated 4-O-methyl glucuronic acid-substituted xylo-oligosaccharides

    NARCIS (Netherlands)

    Neumüller, K.G.; Carvalho de Souza, A.; Rijn, van J.H.J.; Streekstra, H.; Gruppen, H.; Schols, H.A.

    2015-01-01

    Background Acetylation of the xylan backbone restricts the hydrolysis of plant poly- and oligosaccharides by hemicellulolytic enzyme preparations to constituent monosaccharides. The positional preferences and deacetylation efficiencies of acetyl esterases from seven different carbohydrate esterase

  6. Preliminary studies on the endo-xylanase and acetyl esterase of ...

    African Journals Online (AJOL)

    Clostridium thermohydrosulfuricum (JW102) produced low levels of xylanase in mineral medium containing yeast extract and beechwood xylan. Production of xylanase was highest at growth temperatures of 62 – 64oC. Both xylan and xylose supported acetyl esterase production. Xylose, as sole carbon source, supported ...

  7. Polypeptide having acetyl xylan esterase activity and uses thereof

    Energy Technology Data Exchange (ETDEWEB)

    Schoonneveld-Bergmans, Margot Elisabeth Francoise; Heijne, Wilbert Herman Marie; Los, Alrik Pieter

    2015-10-20

    The invention relates to a polypeptide comprising the amino acid sequence set out in SEQ ID NO: 2 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO: 1, or a variant polypeptide or variant polynucleotide thereof, wherein the variant polypeptide has at least 82% sequence identity with the sequence set out in SEQ ID NO: 2 or the variant polynucleotide encodes a polypeptide that has at least 82% sequence identity with the sequence set out in SEQ ID NO: 2. The invention features the full length coding sequence of the novel gene as well as the amino acid sequence of the full-length functional polypeptide and functional equivalents of the gene or the amino acid sequence. The invention also relates to methods for using the polypeptide in industrial processes. Also included in the invention are cells transformed with a polynucleotide according to the invention suitable for producing these proteins.

  8. Trichoderma reesei CE16 acetyl esterase and its role in enzymatic degradation of acetylated hemicellulose

    DEFF Research Database (Denmark)

    Biely, Peter; Cziszarava, Maria; Agger, Jane W.

    2014-01-01

    Results The combined action of GH10 xylanase and acetylxylan esterases (AcXEs) leads to formation of neutral and acidic xylooligosaccharides with a few resistant acetyl groups mainly at their non-reducing ends. We show here that these acetyl groups serve as targets for TrCE16 AcE. The most...... as substrate of the TrCE16 esterase. Conclusion Concerted action of CtGH10 xylanase, an AcXE and TrCE16 AcE resulted in close to complete deacetylation of neutral xylooligosaccharides, whereas substitution with MeGlcA prevents removal of acetyl groups from only a small fraction of the aldouronic acids....... Experiments with diacetyl derivatives of methyl β-d-xylopyranoside confirmed that the best substrate of TrCE16 AcE is 3-O-acetylated Xylp residue followed by 4-O-acetylated Xylp residue with a free vicinal hydroxyl group. General significance This study shows that CE16 acetyl esterases are crucial enzymes...

  9. Effects of enzymatic removal of plant cell wall acylation (acetylation, p-coumaroylation, and feruloylation) on accessibility of cellulose and xylan in natural (non-pretreated) sugar cane fractions.

    Science.gov (United States)

    Várnai, Anikó; Costa, Thales Hf; Faulds, Craig B; Milagres, Adriane Mf; Siika-Aho, Matti; Ferraz, André

    2014-01-01

    Sugar cane internodes can be divided diagonally into four fractions, of which the two innermost ones are the least recalcitrant pith and the moderately accessible pith-rind interface. These fractions differ in enzymatic hydrolyzability due to structural differences. In general, cellulose hydrolysis in plants is hindered by its physical interaction with hemicellulose and lignin. Lignin is believed to be linked covalently to hemicellulose through hydroxycinnamic acids, forming a compact matrix around the polysaccharides. Acetyl xylan esterase and three feruloyl esterases were evaluated for their potential to fragment the lignocellulosic network in sugar cane and to indirectly increase the accessibility of cellulose. The hydrolyzability of the pith and pith-rind interface fractions of a low-lignin-containing sugar cane clone (H58) was compared to that of a reference cultivar (RC). Acetyl xylan esterase enhanced the rate and overall yield of cellulose and xylan hydrolysis in all four substrates. Of the three feruloyl esterases tested, only TsFaeC was capable of releasing p-coumaric acid, while AnFaeA and NcFaeD released ferulic acid from both the pith and interface fractions. Ferulic acid release was higher from the less recalcitrant clone (H58)/fraction (pith), whereas more p-coumaric acid was released from the clone (RC)/fraction (interface) with a higher lignin content. In addition, a compositional analysis of the four fractions revealed that p-coumaroyl content correlated with lignin, while feruloyl content correlated with arabinose content, suggesting different esterification patterns of these two hydroxycinnamic acids. Despite the extensive release of phenolic acids, feruloyl esterases only moderately promoted enzyme access to cellulose or xylan. Acetyl xylan esterase TrAXE was more efficient in enhancing the overall saccharification of sugar cane, compared to the feruloyl esterases AnFaeA, TsFaeC, and NcFaeD. The hydroxycinnamic acid composition of sugar cane

  10. Downregulation of RWA genes in hybrid aspen affects xylan acetylation and wood saccharification.

    Science.gov (United States)

    Pawar, Prashant Mohan-Anupama; Ratke, Christine; Balasubramanian, Vimal K; Chong, Sun-Li; Gandla, Madhavi Latha; Adriasola, Mathilda; Sparrman, Tobias; Hedenström, Mattias; Szwaj, Klaudia; Derba-Maceluch, Marta; Gaertner, Cyril; Mouille, Gregory; Ezcurra, Ines; Tenkanen, Maija; Jönsson, Leif J; Mellerowicz, Ewa J

    2017-06-01

    High acetylation of angiosperm wood hinders its conversion to sugars by glycoside hydrolases, subsequent ethanol fermentation and (hence) its use for biofuel production. We studied the REDUCED WALL ACETYLATION (RWA) gene family of the hardwood model Populus to evaluate its potential for improving saccharification. The family has two clades, AB and CD, containing two genes each. All four genes are expressed in developing wood but only RWA-A and -B are activated by master switches of the secondary cell wall PtNST1 and PtMYB21. Histochemical analysis of promoter::GUS lines in hybrid aspen (Populus tremula × tremuloides) showed activation of RWA-A and -B promoters in the secondary wall formation zone, while RWA-C and -D promoter activity was diffuse. Ectopic downregulation of either clade reduced wood xylan and xyloglucan acetylation. Suppressing both clades simultaneously using the wood-specific promoter reduced wood acetylation by 25% and decreased acetylation at position 2 of Xylp in the dimethyl sulfoxide-extracted xylan. This did not affect plant growth but decreased xylose and increased glucose contents in the noncellulosic monosaccharide fraction, and increased glucose and xylose yields of wood enzymatic hydrolysis without pretreatment. Both RWA clades regulate wood xylan acetylation in aspen and are promising targets to improve wood saccharification. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  11. Functional and structural characterization of a thermostable acetyl esterase from Thermotoga maritima

    NARCIS (Netherlands)

    Levisson, M.; Han, G.W.; Deller, M.C.; Hendriks, S.N.A.; Oost, van der J.; Kengen, S.W.M.

    2012-01-01

    TM0077 from Thermotoga maritima is a member of the carbohydrate esterase family 7 and is active on a variety of acetylated compounds, including cephalosporin C. TM0077 esterase activity is confined to short-chain acyl esters (C2-C3), and is optimal around 100°C and pH 7.5. The positional specificity

  12. Expression of mung bean pectin acetyl esterase in potato tubers: effect on acetylation of cell wall polymers and tuber mechanical properties.

    Science.gov (United States)

    Orfila, Caroline; Dal Degan, Florence; Jørgensen, Bodil; Scheller, Henrik Vibe; Ray, Peter M; Ulvskov, Peter

    2012-07-01

    A mung bean (Vigna radiata) pectin acetyl esterase (CAA67728) was heterologously expressed in tubers of potato (Solanum tuberosum) under the control of the granule-bound starch synthase promoter or the patatin promoter in order to probe the significance of O-acetylation on cell wall and tissue properties. The recombinant tubers showed no apparent macroscopic phenotype. The enzyme was recovered from transgenic tubers using a high ionic strength buffer and the extract was active against a range of pectic substrates. Partial in vivo de-acetylation of cell wall polysaccharides occurred in the transformants, as shown by a 39% decrease in the degree of acetylation (DA) of tuber cell wall material (CWM). Treatment of CWM using a combination of endo-polygalacturonase and pectin methyl esterase extracted more pectin polymers from the transformed tissue compared to wild type. The largest effect of the pectin acetyl esterase (68% decrease in DA) was seen in the residue from this extraction, suggesting that the enzyme is preferentially active on acetylated pectin that is tightly bound to the cell wall. The effects of acetylation on tuber mechanical properties were investigated by tests of failure under compression and by determination of viscoelastic relaxation spectra. These tests suggested that de-acetylation resulted in a stiffer tuber tissue and a stronger cell wall matrix, as a result of changes to a rapidly relaxing viscoelastic component. These results are discussed in relation to the role of pectin acetylation in primary cell walls and its implications for industrial uses of potato fibres.

  13. Trichoderma longibrachiatum acetyl xylan esterase 1 enhances hemicellulolytic preparations to degrade corn silage polysaccharides

    NARCIS (Netherlands)

    Neumüller, K.G.; Streekstra, H.; Gruppen, H.; Schols, H.A.

    2014-01-01

    Supplementation of a Trichoderma longibrachiatum preparation to an industrial Aspergillus niger/Talaromyces emersonii enzyme mixture demonstrated synergy for the saccharification of corn silage water-unextractable solids (WUS). Sub-fractions of the crude T. longibrachiatum preparation obtained after

  14. The Four Arabidopsis Reduced Wall Acetylation Genes are Expressed in Secondary Wall-Containing Cells and Required for the Acetylation of Xylan

    Science.gov (United States)

    Xylan is one of the major polysaccharides in cellulosic biomass, and understanding the mechanisms underlying xylan biosynthesis will potentially help us design strategies to produce cellulosic biomass better suited for biofuel production. Although a number of genes have been show...

  15. Acetylation of woody lignocellulose: significance and regulation

    Directory of Open Access Journals (Sweden)

    Prashant Mohan-Anupama Pawar

    2013-05-01

    Full Text Available Non-cellulosic cell wall polysaccharides constitute approximately one quarter of usable biomass for human exploitation. In contrast to cellulose, these components are usually substituted by O-acetyl groups, which affect their properties and interactions with other polymers, thus affecting their solubility and extractability. However, details of these interactions are still largely obscure. Moreover, polysaccharide hydrolysis to constituent monosaccharides, is hampered by the presence of O-acetyl groups, necessitating either enzymatic (esterase or chemical de-acetylation, increasing the costs and chemical consumption. Reduction of polysaccharide acetyl content in planta is a way to modify lignocellulose towards improved saccharification. In this review we: 1 summarize literature on lignocellulose acetylation in different tree species, 2 present data and current hypotheses concerning the role of O-acetylation in determining woody lignocellulose properties, 3 describe plant proteins involved in lignocellulose O-acetylation, 4 give examples of microbial enzymes capable to de-acetylate lignocellulose, and 5 discuss prospects for exploiting these enzymes in planta to modify xylan acetylation.

  16. Evolution of Xylan Substitution Patterns in Gymnosperms and Angiosperms: Implications for Xylan Interaction with Cellulose.

    Science.gov (United States)

    Busse-Wicher, Marta; Li, An; Silveira, Rodrigo L; Pereira, Caroline S; Tryfona, Theodora; Gomes, Thiago C F; Skaf, Munir S; Dupree, Paul

    2016-08-01

    The interaction between cellulose and xylan is important for the load-bearing secondary cell wall of flowering plants. Based on the precise, evenly spaced pattern of acetyl and glucuronosyl (MeGlcA) xylan substitutions in eudicots, we recently proposed that an unsubstituted face of xylan in a 2-fold helical screw can hydrogen bond to the hydrophilic surfaces of cellulose microfibrils. In gymnosperm cell walls, any role for xylan is unclear, and glucomannan is thought to be the important cellulose-binding polysaccharide. Here, we analyzed xylan from the secondary cell walls of the four gymnosperm lineages (Conifer, Gingko, Cycad, and Gnetophyta). Conifer, Gingko, and Cycad xylan lacks acetylation but is modified by arabinose and MeGlcA. Interestingly, the arabinosyl substitutions are located two xylosyl residues from MeGlcA, which is itself placed precisely on every sixth xylosyl residue. Notably, the Gnetophyta xylan is more akin to early-branching angiosperms and eudicot xylan, lacking arabinose but possessing acetylation on alternate xylosyl residues. All these precise substitution patterns are compatible with gymnosperm xylan binding to hydrophilic surfaces of cellulose. Molecular dynamics simulations support the stable binding of 2-fold screw conifer xylan to the hydrophilic face of cellulose microfibrils. Moreover, the binding of multiple xylan chains to adjacent planes of the cellulose fibril stabilizes the interaction further. Our results show that the type of xylan substitution varies, but an even pattern of xylan substitution is maintained among vascular plants. This suggests that 2-fold screw xylan binds hydrophilic faces of cellulose in eudicots, early-branching angiosperm, and gymnosperm cell walls. © 2016 American Society of Plant Biologists. All Rights Reserved.

  17. Synergistic Enhancement of Cellobiohydrolase Performance on Pretreated Corn Stover by Addition of Xylanase and Esterase Activities

    Energy Technology Data Exchange (ETDEWEB)

    Selig, M. J.; Knoshaug E. P.; Adney, W. S.; Himmel, M. E.; Decker, S. R.

    2007-11-01

    Significant increases in the depolymerization of corn stover cellulose by cellobiohydrolase I (Cel7A) from Trichoderma reesei were observed using small quantities of non-cellulolytic cell wall-degrading enzymes. Purified endoxylanase (XynA), ferulic acid esterase (FaeA), and acetyl xylan esterase (Axe1) all enhanced Cel7A performance on corn stover subjected to hot water pretreatment. In all cases, the addition of these activities improved the effectiveness of the enzymatic hydrolysis in terms of the quantity of cellulose converted per milligram of total protein. Improvement in cellobiose release by the addition of the non-cellulolytic enzymes ranged from a 13-84% increase over Cel7A alone. The most effective combinations included the addition of both XynA and Axe1, which synergistically enhance xylan conversions resulting in additional synergistic improvements in glucan conversion. Additionally, we note a direct relationship between enzymatic xylan removal in the presence of XynA and the enhancement of cellulose hydrolysis by Cel7A.

  18. Spectroscopic study of drug-binding characteristics of unmodified and pNPA-based acetylated human serum albumin: Does esterase activity affect microenvironment of drug binding sites on the protein?

    International Nuclear Information System (INIS)

    Moradi, Nastaran; Ashrafi-Kooshk, Mohammad Reza; Ghobadi, Sirous; Shahlaei, Mohsen; Khodarahmi, Reza

    2015-01-01

    Human serum albumin (HSA) is the most prominent extracellular protein in blood plasma. There are several binding sites on the protein which provide accommodation for structurally-unrelated endogenous and exogenous ligands and a wide variety of drugs. “Esterase-like” activity (hydrolysis of p-nitrophenyl esters) by the protein has been also reported. In the current study, we set out to investigate the interaction of indomethacin and ibuprofen with the unmodified and modified HSA (pNPA-modified HSA) using various spectroscopic techniques. Fluorescence data showed that 1:1 binding of drug to HSA is associated with quenching of the protein intrinsic fluorescence. Decrease of protein surface hydrophobicity (PSH), alteration in drug binding affinity and change of the protein stability, after esterase-like activity and permanent acetylation of HSA, were also documented. Analysis of the quenching and thermodynamic parameters indicated that forces involved in drug–HSA interactions change upon the protein modification. - Highlights: • Binding propensity of indomethacin extremely decreased upon the protein acetylation. • There is no ibuprofen binding after protein acetylation. • Protein stability changes upon drug binding as well as protein acetylation. • Drug pharmacokinetics may be influenced under co-administration of HSA-modifier drugs

  19. Spectroscopic study of drug-binding characteristics of unmodified and pNPA-based acetylated human serum albumin: Does esterase activity affect microenvironment of drug binding sites on the protein?

    Energy Technology Data Exchange (ETDEWEB)

    Moradi, Nastaran [Medical Biology Research Center, Kermanshah University of Medical Sciences, Kermanshah (Iran, Islamic Republic of); Faculty of Pharmaceutical Sciences, Kermanshah University of Medical Sciences, Kermanshah (Iran, Islamic Republic of); Ashrafi-Kooshk, Mohammad Reza [Medical Biology Research Center, Kermanshah University of Medical Sciences, Kermanshah (Iran, Islamic Republic of); Ghobadi, Sirous [Department of Biology, Faculty of Sciences, Razi University, Kermanshah (Iran, Islamic Republic of); Shahlaei, Mohsen [Medical Biology Research Center, Kermanshah University of Medical Sciences, Kermanshah (Iran, Islamic Republic of); Faculty of Pharmaceutical Sciences, Kermanshah University of Medical Sciences, Kermanshah (Iran, Islamic Republic of); Khodarahmi, Reza, E-mail: rkhodarahmi@mbrc.ac.ir [Medical Biology Research Center, Kermanshah University of Medical Sciences, Kermanshah (Iran, Islamic Republic of); Faculty of Pharmaceutical Sciences, Kermanshah University of Medical Sciences, Kermanshah (Iran, Islamic Republic of)

    2015-04-15

    Human serum albumin (HSA) is the most prominent extracellular protein in blood plasma. There are several binding sites on the protein which provide accommodation for structurally-unrelated endogenous and exogenous ligands and a wide variety of drugs. “Esterase-like” activity (hydrolysis of p-nitrophenyl esters) by the protein has been also reported. In the current study, we set out to investigate the interaction of indomethacin and ibuprofen with the unmodified and modified HSA (pNPA-modified HSA) using various spectroscopic techniques. Fluorescence data showed that 1:1 binding of drug to HSA is associated with quenching of the protein intrinsic fluorescence. Decrease of protein surface hydrophobicity (PSH), alteration in drug binding affinity and change of the protein stability, after esterase-like activity and permanent acetylation of HSA, were also documented. Analysis of the quenching and thermodynamic parameters indicated that forces involved in drug–HSA interactions change upon the protein modification. - Highlights: • Binding propensity of indomethacin extremely decreased upon the protein acetylation. • There is no ibuprofen binding after protein acetylation. • Protein stability changes upon drug binding as well as protein acetylation. • Drug pharmacokinetics may be influenced under co-administration of HSA-modifier drugs.

  20. Biochemical Characterization and Relative Expression Levels of Multiple Carbohydrate Esterases of the Xylanolytic Rumen Bacterium Prevotella ruminicola 23 Grown on an Ester-Enriched Substrate ▿ †

    Science.gov (United States)

    Kabel, Mirjam A.; Yeoman, Carl J.; Han, Yejun; Dodd, Dylan; Abbas, Charles A.; de Bont, Jan A. M.; Morrison, Mark; Cann, Isaac K. O.; Mackie, Roderick I.

    2011-01-01

    We measured expression and used biochemical characterization of multiple carbohydrate esterases by the xylanolytic rumen bacterium Prevotella ruminicola 23 grown on an ester-enriched substrate to gain insight into the carbohydrate esterase activities of this hemicellulolytic rumen bacterium. The P. ruminicola 23 genome contains 16 genes predicted to encode carbohydrate esterase activity, and based on microarray data, four of these were upregulated >2-fold at the transcriptional level during growth on an ester-enriched oligosaccharide (XOSFA,Ac) from corn relative to a nonesterified fraction of corn oligosaccharides (AXOS). Four of the 16 esterases (Xyn10D-Fae1A, Axe1-6A, AxeA1, and Axe7A), including the two most highly induced esterases (Xyn10D-Fae1A and Axe1-6A), were heterologously expressed in Escherichia coli, purified, and biochemically characterized. All four enzymes showed the highest activity at physiologically relevant pH (6 to 7) and temperature (30 to 40°C) ranges. The P. ruminicola 23 Xyn10D-Fae1A (a carbohydrate esterase [CE] family 1 enzyme) released ferulic acid from methylferulate, wheat bran, corn fiber, and XOSFA,Ac, a corn fiber-derived substrate enriched in O-acetyl and ferulic acid esters, but exhibited negligible activity on sugar acetates. As expected, the P. ruminicola Axe1-6A enzyme, which was predicted to possess two distinct esterase family domains (CE1 and CE6), released ferulic acid from the same substrates as Xyn10D-Fae1 and was also able to cleave O-acetyl ester bonds from various acetylated oligosaccharides (AcXOS). The P. ruminicola 23 AxeA1, which is not assigned to a CE family, and Axe7A (CE7) were found to be acetyl esterases that had activity toward a broad range of mostly nonpolymeric acetylated substrates along with AcXOS. All enzymes were inhibited by the proximal location of other side groups like 4-O-methylglucuronic acid, ferulic acid, or acetyl groups. The unique diversity of carbohydrate esterases in P. ruminicola 23

  1. Regulation of the feruloyl esterase (faeA) gene from Aspergillus niger

    NARCIS (Netherlands)

    Vries, de R.P.; Visser, J.

    1999-01-01

    Feruloyl esterases can remove aromatic residues (e.g., ferulic acid) from plant cell wall polysaccharides (xylan, pectin) and are essential for complete degradation of these polysaccharides. Expression of the feruloyl esterase-encoding gene (faeA) from Aspergillus niger depends on D-xylose

  2. Structural and Enzymatic Characterization of NanS (YjhS) a 9-O-Acetyl N-acetylneuraminic Acid Esterase from Escherichia coli O157:H7

    Energy Technology Data Exchange (ETDEWEB)

    E Rangarajan; K Ruane; A Proteau; J Schrag; R Valladares; C Gonzalez; M Gilbert; A Yakunin; M Cygler

    2011-12-31

    There is a high prevalence of sialic acid in a number of different organisms, resulting in there being a myriad of different enzymes that can exploit it as a fermentable carbon source. One such enzyme is NanS, a carbohydrate esterase that we show here deacetylates the 9 position of 9-O-sialic acid so that it can be readily transported into the cell for catabolism. Through structural studies, we show that NanS adopts a SGNH hydrolase fold. Although the backbone of the structure is similar to previously characterized family members, sequence comparisons indicate that this family can be further subdivided into two subfamilies with somewhat different fingerprints. NanS is the founding member of group II. Its catalytic center contains Ser19 and His301 but no Asp/Glu is present to form the classical catalytic triad. The contribution of Ser19 and His301 to catalysis was confirmed by mutagenesis. In addition to structural characterization, we have mapped the specificity of NanS using a battery of substrates.

  3. Populus GT43 family members group into distinct sets required for primary and secondary wall xylan biosynthesis and include useful promoters for wood modification.

    Science.gov (United States)

    Ratke, Christine; Pawar, Prashant Mohan-Anupama; Balasubramanian, Vimal K; Naumann, Marcel; Duncranz, Mathilda Lönnäs; Derba-Maceluch, Marta; Gorzsás, András; Endo, Satoshi; Ezcurra, Ines; Mellerowicz, Ewa J

    2015-01-01

    The plant GT43 protein family includes xylosyltransferases that are known to be required for xylan backbone biosynthesis, but have incompletely understood specificities. RT-qPCR and histochemical (GUS) analyses of expression patterns of GT43 members in hybrid aspen, reported here, revealed that three clades of the family have markedly differing specificity towards secondary wall-forming cells (wood and extraxylary fibres). Intriguingly, GT43A and B genes (corresponding to the Arabidopsis IRX9 clade) showed higher specificity for secondary-walled cells than GT43C and D genes (IRX14 clade), although both IRX9 and IRX14 are required for xylosyltransferase activity. The remaining genes, GT43E, F and G (IRX9-L clade), showed broad expression patterns. Transient transactivation analyses of GT43A and B reporters demonstrated that they are activated by PtxtMYB021 and PNAC085 (master secondary wall switches), mediated in PtxtMYB021 activation by an AC element. The high observed secondary cell wall specificity of GT43B expression prompted tests of the efficiency of its promoter (pGT43B), relative to the CaMV 35S (35S) promoter, for overexpressing a xylan acetyl esterase (CE5) or downregulating REDUCED WALL ACETYLATION (RWA) family genes and thus engineering wood acetylation. CE5 expression was weaker when driven by pGT43B, but it reduced wood acetyl content substantially more efficiently than the 35S promoter. RNAi silencing of the RWA family, which was ineffective using 35S, was achieved when using GT43B promoter. These results show the utility of the GT43B promoter for genetically engineering properties of wood and fibres. © 2014 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  4. Growth advantage of Escherichia coli O104:H4 strains on 5-N-acetyl-9-O-acetyl neuraminic acid as a carbon source is dependent on heterogeneous phage-Borne nanS-p esterases.

    Science.gov (United States)

    Saile, Nadja; Schwarz, Lisa; Eißenberger, Kristina; Klumpp, Jochen; Fricke, Florian W; Schmidt, Herbert

    2018-03-26

    Enterohemorrhagic E. coli (EHEC) are serious bacterial pathogens which are able to cause a hemorrhagic colitis or the life-threatening hemolytic-uremic syndrome (HUS) in humans. EHEC strains can carry different numbers of phage-borne nanS-p alleles that are responsible for acetic acid release from mucin from bovine submaxillary gland and 5-N-acetyl-9-O-acetyl neuraminic acid (Neu5,9Ac 2 ), a carbohydrate present in mucin. Thus, Neu5,9Ac 2 can be transformed to 5-N-acetyl neuraminic acid, an energy source used by E. coli strains. We hypothesize that these NanS-p proteins are involved in competitive growth of EHEC in the gastrointestinal tract of humans and animals. The aim of the current study was to demonstrate and characterize the nanS-p alleles of the 2011 E. coli O104:H4 outbreak strain LB226692 and analyze whether the presence of multiple nanS-p alleles in the LB226692 genome causes a competitive growth advantage over a commensal E. coli strain. We detected and characterized five heterogeneous phage-borne nanS-p alleles in the genome of E. coli O104:H4 outbreak strain LB226692 by in silico analysis of its genome. Furthermore, successive deletion of all nanS-p alleles, subsequent complementation with recombinant NanS-p13-His, and in vitro co-culturing experiments with the commensal E. coli strain AMC 198 were conducted. We could show that nanS-p genes of E. coli O104:H4 are responsible for growth inhibition of strain AMC 198, when Neu5,9Ac 2 was used as sole carbon source in co-culture. The results of this study let us suggest that multiple nanS-p alleles may confer a growth advantage by outcompeting other E. coli strains in Neu5,9Ac 2 rich environments, such as mucus in animal and human gut. Copyright © 2018. Published by Elsevier GmbH.

  5. Characterization of O-acetil-(4-O-methylglucurono)xylans from Eucalyptus urograndis

    International Nuclear Information System (INIS)

    Magaton, Andreia da Silva; Pilo-Veloso, Dorila; Colodette, Jorge Luiz

    2008-01-01

    The O-acetyl- 4-O-methyl-(glucurono)xylans were isolated from E. urograndis by extraction with dimethyl sulfoxide, analysed for monosaccharide composition and structurally characterized by NMR spectroscopy. These xylans contained one 4-O-methyl-glucuronic acid substituent and 5.5 acetyl groups for approximately 10 xylose residues. About 10% of 4-O-methyl-glucuronic acid (MeGlcA) units were branched at O-2. The O-acetyl-4-O-methyl-(glucurono)-xylans were composed of the following (1→4)-linked β-D-xylopyranosyl structural elements: unsubstituted (51 mol%), 2-O-acetylated (12 mol%), 3-O-acetylated (20 mol%), 2,3-di-Oacetylated (6 mol%) and [MeGlcA α-(1→2)] [3-O-acetylated] (11 mol%). The weight-average molar mass and polydispersity of this xylan were 34.9 kDa and 1.16, respectively, as measured by size-exclusion chromatography. (author)

  6. Leukocyte esterase urine test

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003584.htm Leukocyte esterase urine test To use the sharing features on this page, please enable JavaScript. Leukocyte esterase is a urine test to look for ...

  7. EFFECTS OF XYLAN IN EUCALYPTUS PULP PRODUCTION

    Directory of Open Access Journals (Sweden)

    Bianca Moreira Barbosa

    2016-06-01

    Full Text Available The search for a better use of wood in the pulp industry has fuelled interest in a more rational use of its components, particularly xylans. The impact of xylans removal and of xylans redeposition on pulp properties for tissue and P&W paper grades are discussed in this paper. Kraft pulp (15.6% xylans treatment with 10-70 g.L-1 NaOH resulted in pulps of 14.5-5.9% xylans. The treatments decreased pulp lignin and HexA contents and caused significant positive impact on subsequent oxygen delignification and ECF bleaching. Xylan removal decreased pulp beatability, water retention value and tensile index but increased drainability, water absorption capacity, capillarity Klemm and bulk. Overall, xylan depleted pulps showed almost ideal properties for tissue paper grade pulps. In a second step of the research, xylans extracted from unbleached (BXL and bleached eucalyptus pulps (WXL by cold caustic extraction (CCE were added to a commercial brown pulp in the oxygen delignification (O-stage and further bleached. Xylans deposition occurred at variable degree (up to 7% on pulp weight depending upon the O-stage reaction pH. Pulp bleachability was not impaired by WXL xylan deposition but slightly negatively affected by BXL xylans. Pulp beatability was improved by xylan deposition. The deposited xylans were quite stable across bleaching and beating, with the WXL xylans being more stable than the BXL ones. At low energy consumption, the deposited xylans improved pulp physical and mechanical properties. Xylans extraction by CCE with subsequent deposition onto pulp in the O-stage proved attractive for manufacturing high xylan P&W paper grades.

  8. Delignification outperforms alkaline extraction for xylan fingerprinting of oil palm empty fruit bunch.

    Science.gov (United States)

    Murciano Martínez, Patricia; Kabel, Mirjam A; Gruppen, Harry

    2016-11-20

    Enzyme hydrolysed (hemi-)celluloses from oil palm empty fruit bunches (EFBs) are a source for production of bio-fuels or chemicals. In this study, after either peracetic acid delignification or alkaline extraction, EFB hemicellulose structures were described, aided by xylanase hydrolysis. Delignification of EFB facilitated the hydrolysis of EFB-xylan by a pure endo-β-1,4-xylanase. Up to 91% (w/w) of the non-extracted xylan in the delignified EFB was hydrolysed compared to less than 4% (w/w) of that in untreated EFB. Alkaline extraction of EFB, without prior delignification, yielded only 50% of the xylan. The xylan obtained was hydrolysed only for 40% by the endo-xylanase used. Hence, delignification alone outperformed alkaline extraction as pretreatment for enzymatic fingerprinting of EFB xylans. From the analysis of the oligosaccharide-fingerprint of the delignified endo-xylanase hydrolysed EFB xylan, the structure was proposed as acetylated 4-O-methylglucuronoarabinoxylan. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Hydrolysis of xylans by enzyme systems from solid cultures of Trichoderma harzianum strains.

    Science.gov (United States)

    Silveira, F Q; Ximenes, F A; Cacais, A O; Milagres, A M; Medeiros, C L; Puls, J; Filho, E X

    1999-08-01

    Xylanase activity was isolated from crude extracts of Trichoderma harzianum strains C and 4 grown at 28 degree C in a solid medium containing wheat bran as the carbon source. Enzyme activity was demonstrable in the permeate after ultrafiltration of the crude extracts using an Amicon system. The hydrolysis patterns of different xylans and paper pulps by xylanase activity ranged from xylose, xylobiose and xylotriose to higher xylooligosaccharides. A purified ss-xylosidase from the Trichoderma harzianum strain released xylose, xylobiose and xylotriose from seaweed, deacetylated, oat spelt and birchwood xylans. The purified enzyme was not active against acetylated xylan and catalyzed the hydrolysis of xylooligosaccharides, including xylotriose, xylotetraose and xylopentaose. However, the enzyme was not able to degrade xylohexaose. Xylanase pretreatment was effective for hardwood kraft pulp bleaching. Hardwood kraft pulp bleached in the XEOP sequence had its kappa number reduced from 13.2 to 8.9 and a viscosity of 20. 45 cp. The efficiency of delignification was 33%.

  10. Trimethyl Lock: A Stable Chromogenic Substrate for Esterases

    Directory of Open Access Journals (Sweden)

    Ronald T. Raines

    2008-01-01

    Full Text Available p-Nitrophenyl acetate is the most commonly used substrate for detecting thecatalytic activity of esterases, including those that activate prodrugs in human cells. Thissubstrate is unstable in aqueous solution, limiting its utility. Here, a stable chromogenicsubstrate for esterases is produced by the structural isolation of an acetyl ester andp-nitroaniline group using a trimethyl lock moiety. Upon ester hydrolysis, unfavorablesteric interactions between the three methyl groups of this o-hydroxycinnamic acidderivative encourage rapid lactonization to form a hydrocoumarin and releasep-nitroaniline. This “prochromophore” could find use in a variety of assays.

  11. Esterase screening using whole cells of Brazilian soil microorganisms

    Energy Technology Data Exchange (ETDEWEB)

    Mantovani, Simone M.; Oliveira, Luciana G. de; Marsaioli, Anita J., E-mail: anita@iqm.unicamp.b [Universidade Estadual de Campinas (IQ/UNICAMP), SP (Brazil). Inst. de Quimica

    2010-07-01

    A miniaturized enzymatic assay using fluorescent probes to reveal esterase producing microorganisms was optimized and applied to screen 64 soil bacterial strains. The best results were validated using traditional non-fluorogenic assays with acetyl and propanoyl phenylethanol to confirm the miniaturized results. The most active microorganisms belong to the genus Bacillus showing esterase activity and good enantiomeric ratios for the resolution of phenylethanol derivatives (E > 30). Part of the microorganisms are kept in our laboratory in glycerol or freezedried and the best microorganisms will be deposited in the CBMAI/CPQBA/UNICAMP culture collection. (author)

  12. Carboxymethylated-, hydroxypropylsulfonated- and quaternized xylan derivative films

    Czech Academy of Sciences Publication Activity Database

    Šimkovic, I.; Kelnar, Ivan; Uhliariková, I.; Mendichi, R.; Mandalika, A.; Elder, T.

    2014-01-01

    Roč. 110, 22 September (2014), s. 464-471 ISSN 0144-8617 Institutional support: RVO:61389013 Keywords : xylan * films * properties Subject RIV: CD - Macromolecular Chemistry Impact factor: 4.074, year: 2014

  13. Non-specific esterases and esterproteases in masticatory muscles from the muscular dystrophic mouse

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D; Vilmann, H

    1989-01-01

    With the aid of histochemical and electrophoretic techniques activities for esterase and esterprotease were investigated in the digastric and masseter muscles from normal and dystrophic mice. The substrates used were alpha-naphthyl acetate and N-acetyl-L-alanine alpha-naphthyl ester. According...

  14. Structural differences of xylans affect their interaction with cellulose

    NARCIS (Netherlands)

    Kabel, M.A.; Borne, van den H.; Vincken, J.P.; Voragen, A.G.J.; Schols, H.A.

    2007-01-01

    The affinity of xylan to cellulose is an important aspect of many industrial processes, e.g. production of cellulose, paper making and bio-ethanol production. However, little is known about the adsorption of structurally different xylans to cellulose. Therefore, the adsorption of various xylans to

  15. Esterase Isoenzyme Variants in Barley

    DEFF Research Database (Denmark)

    Hvid, S.; Nielsen, G.

    1977-01-01

    Gene symbols are proposed for 27 esterase isoenzyme alleles representing 10 loci in barley. Two new esterase loci, Est 9 and Est 10, each with an active and a silent allele, and three new alleles in previously described loci were found. A few chemical and physical characteristics of the different...... esterase isoenzyme systems were studied. The heat inactivation temperature differed for the isoenzymes coded by most of the loci, whereas the substrate and inhibitor specificity of the isoenzymes was less distinct. A possible relationship between some of the systems is discussed....

  16. Esterases as pesticide biomarkers in crayfish (Procambarus clarkii, Crustacea): tissue distribution, sensitivity to model compounds and recovery from inactivation.

    Science.gov (United States)

    Vioque-Fernández, A; de Almeida, E Alves; López-Barea, J

    2007-04-01

    The specific activities of acetyl- and butyrylcholinesterase and carboxylesterase were assayed in the digestive gland and in nervous and muscle tissues of the crayfish Procambarus clarkii. Since acetylcholinesterase prevails in nervous tissue and carboxylesterase in digestive gland, they are proposed as biomarkers. Muscle had negligible activities of all esterases, and all tissues had a low butyrylcholinesterase activity. Esterases were mostly cytosolic in digestive gland and muscle, but membrane-bound in nervous tissue; use of Triton X-100 is not recommended due to its widely diverging effects in esterase assays. Phenylmethylsulphonylfluoride inhibited acetyl- and butyrylcholinesterase in extracts from all tissues, and in digestive gland only carboxylesterase. In digestive gland, tetra[monoisopropyl]-pyrophosphorotetramide inhibited all esterases with different sensitivities, while in muscle and nervous tissue it only partially inhibited all esterases. Carbamates inhibited 100-fold more strongly than organophosphates acetyl- and butyrylcholinesterase activities. Carboxylesterase was inhibited by carbaryl and chlorpyrifos, but not by eserine and malathion. In vitro conditions to evaluate recovery from inactivation of esterases by model pesticides were established for acetylcholinesterase and carboxylesterase. The new reactivation protocol could be useful as a biomarker of pesticide exposure to differentiate between dilution-reversible inhibitions, indicating carbamate exposure, from dilution-irreversible effect, attributed to organophosphate exposure.

  17. The natural catalytic function of CuGE glucuronoyl esterase in hydrolysis of genuine lignin-carbohydrate complexes from birch

    DEFF Research Database (Denmark)

    Mosbech, Caroline; Holck, Jesper; Meyer, Anne S.

    2018-01-01

    Glucuronoyl esterases belong to carbohydrate esterase family 15 and catalyze de-esterification. Their natural function is presumed to be cleavage of ester linkages in lignin-carbohydrate complexes particularly those linking lignin and glucuronoyl residues in xylans in hardwood. Here, we show...... for the first time a detailed product profile of aldouronic acids released from birchwood lignin by a glucuronoyl esterase from the white-rot fungus Cerrena unicolor (CuGE). CuGE releases substrate for GH10 endo-xylanase which results in significantly increased product release compared to the action of endo...... with lignin and we propose that this is a direct result of enzymatic cleavage of the ester bonds connecting glucuronoxylan to lignin via 4-O-methyl glucuronoyl-ester linkages. This function appears important for the fungal organism's ability to effectively utilize all available carbohydrates...

  18. Carboxymethylated-, hydroxypropylsulfonated- and quaternized xylan derivative films

    Science.gov (United States)

    Ivan Simkovic; Ivan Kelnar; Iveta Uhliarikova; Raniero Mdndichi; Anurag Mandalika; Thomas Elder

    2014-01-01

    Under alkaline/water conditions carboxymethyl, 2-hydroxypropylsulfonate and trimethylammonium-2-hydroxypropyl groups were introduced into xylan in one step with the goal to prepare film specimens. The materials were characterized by NMR, SEC-MALS, TG/DTG/DTA, AFM and mechanical testing. The properties of triple, double and mono-substituted materials were compared. The...

  19. Enzymatic deconstruction of xylan for biofuel production

    Science.gov (United States)

    DODD, DYLAN; CANN, ISAAC K. O.

    2010-01-01

    The combustion of fossil-derived fuels has a significant impact on atmospheric carbon dioxide (CO2) levels and correspondingly is an important contributor to anthropogenic global climate change. Plants have evolved photosynthetic mechanisms in which solar energy is used to fix CO2 into carbohydrates. Thus, combustion of biofuels, derived from plant biomass, can be considered a potentially carbon neutral process. One of the major limitations for efficient conversion of plant biomass to biofuels is the recalcitrant nature of the plant cell wall, which is composed mostly of lignocellulosic materials (lignin, cellulose, and hemicellulose). The heteropolymer xylan represents the most abundant hemicellulosic polysaccharide and is composed primarily of xylose, arabinose, and glucuronic acid. Microbes have evolved a plethora of enzymatic strategies for hydrolyzing xylan into its constituent sugars for subsequent fermentation to biofuels. Therefore, microorganisms are considered an important source of biocatalysts in the emerging biofuel industry. To produce an optimized enzymatic cocktail for xylan deconstruction, it will be valuable to gain insight at the molecular level of the chemical linkages and the mechanisms by which these enzymes recognize their substrates and catalyze their reactions. Recent advances in genomics, proteomics, and structural biology have revolutionized our understanding of the microbial xylanolytic enzymes. This review focuses on current understanding of the molecular basis for substrate specificity and catalysis by enzymes involved in xylan deconstruction. PMID:20431716

  20. Maillard reaction products from chitosan-xylan ionic liquid solution.

    Science.gov (United States)

    Luo, Yuqiong; Ling, Yunzhi; Wang, Xiaoying; Han, Yang; Zeng, Xianjie; Sun, Runcang

    2013-10-15

    A facile method is reported to prepare Maillard reaction products (MRPs) from chitosan and xylan in co-solvent ionic liquid. UV absorbance and fluorescence changes were regarded as indicators of the occurrence of Maillard reaction. FT-IR, NMR, XRD and TG were used to investigate the structure of chitosan-xylan conjugate. The results revealed that when chitosan reacted with xylan in ionic liquid, the hydrogen bonds in chitosan were destroyed, the facts resulted in the formation of chitosan-xylan MRPs. Moreover, when the mass ratio of chitosan to xylan was 1:1, the Maillard reaction proceeded easily. In addition, relatively high antioxidant property was also noted for the chitosan-xylan conjugate with mass ratio 1:1. So the obtained chitosan-xylan MRP is a promising antioxidant agent for food industry. Copyright © 2013 Elsevier Ltd. All rights reserved.

  1. Hydrolysis of xylans by enzyme systems from solid cultures of Trichoderma harzianum strains

    Directory of Open Access Journals (Sweden)

    Silveira F.Q.P.

    1999-01-01

    Full Text Available Xylanase activity was isolated from crude extracts of Trichoderma harzianum strains C and 4 grown at 28oC in a solid medium containing wheat bran as the carbon source. Enzyme activity was demonstrable in the permeate after ultrafiltration of the crude extracts using an Amicon system. The hydrolysis patterns of different xylans and paper pulps by xylanase activity ranged from xylose, xylobiose and xylotriose to higher xylooligosaccharides. A purified ß-xylosidase from the Trichoderma harzianum strain released xylose, xylobiose and xylotriose from seaweed, deacetylated, oat spelt and birchwood xylans. The purified enzyme was not active against acetylated xylan and catalyzed the hydrolysis of xylooligosaccharides, including xylotriose, xylotetraose and xylopentaose. However, the enzyme was not able to degrade xylohexaose. Xylanase pretreatment was effective for hardwood kraft pulp bleaching. Hardwood kraft pulp bleached in the XEOP sequence had its kappa number reduced from 13.2 to 8.9 and a viscosity of 20.45 cp. The efficiency of delignification was 33%.

  2. Mechanistic Explanation of the Weak Carbonic Anhydrase’s Esterase Activity

    Directory of Open Access Journals (Sweden)

    Paolo Piazzetta

    2017-06-01

    Full Text Available In order to elucidate the elementary mechanism of the promiscuous esterase activity of human carbonic anhydrase (h-CA, we present an accurate theoretical investigation on the hydrolysis of fully-acetylated d-glucose functionalized as sulfamate. This h-CA’s inhibitor is of potential relevance in cancer therapy. The study has been performed within the framework of three-layer ONIOM (QM-high:QM’-medium:MM-low hybrid approach. The computations revealed that the hydrolysis process is not energetically favored, in agreement with the observed weak carbonic anhydrase’s esterase activity.

  3. Controlled enzyme catalyzed heteropolysaccharide degradation:Xylans

    OpenAIRE

    Rasmussen, Louise Enggaard; Meyer, Anne S.

    2011-01-01

    The work presented in this PhD thesis has provided a better understanding of the enzyme kinetics and quantitative phenomena of the hydrolysis of xylan substrates by selected pure enzyme preparations. Furthermore, the options for producing specific substituted xylooligosaccharides from selected substrates by specific xylanase treatment have been examined. The kinetics of the enzymatic degradation of water-extractable wheat arabinoxylan (WE-AX) during designed treatments with selected monocompo...

  4. Mannich reaction of polysaccharides: Xylan functionalization in aqueous basic medium.

    Science.gov (United States)

    Ünlü, Cüneyt H; Kutlu, Meltem; Atıcı, Oya Galioğlu

    2015-01-01

    In this study modification of xylan via Mannich reaction in aqueous basic solution to obtain dimethylaminomethylated products and characterization of modified xylan were examined. Components were xylan (obtained from corn cob and used without modification) as active hydrogen containing compound, formaldehyde as carbonyl compound having no α-hydrogen and dimethylamine. Mannich reaction was used with different parameters such as component concentration, reaction temperature, and time. The highest modification was observed about 35°C with a nitrogen content of 4.6% by weight indicating successive modification. Both 1D and 2D NMR measurements displayed new signals related with aminomethyl groups. Spectral characterizations indicated that aminomethylation took place on oxygen sites. Moreover modified xylan could form film while xylan could not without an auxiliary agent. Antimicrobial activity tests indicated that modified xylan acted as a bacteriostatic material. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Selection and production of insoluble xylan hydrolyzing enzyme by ...

    African Journals Online (AJOL)

    Forty-two strains of Thermomyces lanuginosus isolated from various sources in Thailand were divide into 4 groups based on the soluble xylan hydrolyzing (SXH) and insoluble xylan hydrolyzing (IXH) enzyme activities in the supernatant obtained from 5-day culture at 50°C in the liquid medium using corncob as substrate.

  6. Crosslinked xylan as an affinity adsorbent for endo-xylanases.

    NARCIS (Netherlands)

    Rozie, H.; Somers, W.; Bonte, A.; Rombouts, F.M.; Visser, J.

    1992-01-01

    In order to facilitate the purification of xylanases from Aspergillus niger, an affinity adsorbent has been developed from oat spelts xylan. A suitable adsorbent was only obtained by crosslinking oat spelts xylan with epichlorohydrin in water but not in ethanol or ethanol-water mixtures. After some

  7. Enzymatic degradation of lignin‐carbohydrate complexes (LCCs): Model studies using a fungal glucuronoyl esterase from Cerrena unicolor

    DEFF Research Database (Denmark)

    d'Errico, Clotilde; Jørgensen, Jonas O.; Krogh, Kristian B. R. M.

    2015-01-01

    Lignin‐carbohydrate complexes (LCCs) are believed to influence the recalcitrance of lignocellulosic plant material preventing optimal utilization of biomass in e.g. forestry, feed and biofuel applications. The recently emerged carbohydrate esterase (CE) 15 family of glucuronoyl esterases (GEs) has...... been proposed to degrade ester LCC bonds between glucuronic acids in xylans and lignin alcohols thereby potentially improving delignification of lignocellulosic biomass when applied in conjunction with other cellulases, hemicellulases and oxidoreductases. Herein, we report the synthesis of four new GE...... model substrates comprising α‐ and ɣ‐arylalkyl esters representative of the lignin part of naturally occurring ester LCCs as well as the cloning and purification of a novel GE from Cerrena unicolor (CuGE). Together with a known GE from Schizophyllum commune (ScGE), CuGE was biochemically characterized...

  8. Esterase profile of human masseter muscle

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D; Vilmann, H

    1988-01-01

    The esterase profile of fresh human masseter muscle was investigated by use of histochemistry and electrophoresis. The histochemical methods included reactions for alpha-naphthyl esterase, myofibrillar ATPase, reverse myofibrillar ATPase and succinic dehydrogenase. In frozen sections of the muscle...... the coloured reaction product for esterases was present both as a diffuse sarcoplasmic coloration and as distinct granules. The intensity of diffuse reaction was used to classify the muscle fibres as strongly, moderately and weakly reacting. The fibres with strong esterase activity belonged to Type I and ii......C. iM and Type II A fibres showed a moderate esterase reaction and Type II B fibres had a low activity. The electrophoretic gels stained for esterase activity showed that the human masseter muscle possesses a slow migrating double band with high enzyme activity and a cascade of faster migrating...

  9. A Bacillus licheniformis pectin acetylesterase is specific for homogalacturonans acetylated at O-3

    NARCIS (Netherlands)

    Remoroza, C.A.; Wagenknecht, M.; Buchholt, H.C.; Moerschbacher, B.M.; Schols, H.A.; Gruppen, H.

    2014-01-01

    A recombinant acetylesterase from Bacillus licheniformis DSM13, belonging to carbohydrate esterase family 12, was purified and biochemically characterized. The purified enzyme, termed BliPAE, was capable of deacetylating acetylated pectins, e.g. sugar beet pectin (SBP). Contrary to its provisional

  10. Correlation between serum esterase polymorphism and production ...

    African Journals Online (AJOL)

    The polymorphism of serum esterase (Es) of Henan Yuxi fat-tailed sheep was detected through polyacrylamide gel electrophoresis (PAGE), and the correlation between serum esterase and productivity was analyzed. The research result indicated that there are two alleles on the Es loci of Henan Yuxi fat-tailed sheep: Es+ ...

  11. aes, the gene encoding the esterase B in Escherichia coli, is a powerful phylogenetic marker of the species

    Directory of Open Access Journals (Sweden)

    Tuffery Pierre

    2009-12-01

    Full Text Available Abstract Background Previous studies have established a correlation between electrophoretic polymorphism of esterase B, and virulence and phylogeny of Escherichia coli. Strains belonging to the phylogenetic group B2 are more frequently implicated in extraintestinal infections and include esterase B2 variants, whereas phylogenetic groups A, B1 and D contain less virulent strains and include esterase B1 variants. We investigated esterase B as a marker of phylogeny and/or virulence, in a thorough analysis of the esterase B-encoding gene. Results We identified the gene encoding esterase B as the acetyl-esterase gene (aes using gene disruption. The analysis of aes nucleotide sequences in a panel of 78 reference strains, including the E. coli reference (ECOR strains, demonstrated that the gene is under purifying selection. The phylogenetic tree reconstructed from aes sequences showed a strong correlation with the species phylogenetic history, based on multi-locus sequence typing using six housekeeping genes. The unambiguous distinction between variants B1 and B2 by electrophoresis was consistent with Aes amino-acid sequence analysis and protein modelling, which showed that substituted amino acids in the two esterase B variants occurred mostly at different sites on the protein surface. Studies in an experimental mouse model of septicaemia using mutant strains did not reveal a direct link between aes and extraintestinal virulence. Moreover, we did not find any genes in the chromosomal region of aes to be associated with virulence. Conclusion Our findings suggest that aes does not play a direct role in the virulence of E. coli extraintestinal infection. However, this gene acts as a powerful marker of phylogeny, illustrating the extensive divergence of B2 phylogenetic group strains from the rest of the species.

  12. Laccase catalysed grafting of phenolic onto xylan to improve its applicability in films

    Science.gov (United States)

    Pei, Jicheng; Wang, Bing; Zhang, Fangdong; Li, Zhongyang; Yin, Yunbei; Zhang, Dongxu

    2015-07-01

    Xylan can be tailored for various value-added applications. However, its use in aqueous systems is hampered by its complex structure, and small molecular weight. This research aimed at improving the xylan molecular weight and changing its structure. Laccase-catalysed oxidation of 4-coumaric acid (PCA), ferulic acid (FA), syringaldehyde (SD), and vanillin (VA) onto xylan was grafted to study the changes in its structure, tensile properties, and antibacterial activities. A Fourier transform infrared (FTIR) spectrum analyser was used to observe the changes in functional groups of xylan. The results showed a band at 1635 cm-1 corresponding to the stretching vibration of conjugated carbonyl carboxy hemoglobin and a benzene ring structure were strengthened; the appearance of a new band between 1200 cm-1 and 1270 cm-1 corresponding to alkyl ethers on the aryl C-O stretching vibration was due to the fact that during the grafting process, the number of benzene ring structures increased and covalent connections occurred between phenols and xylan. The reaction mechanism for the laccase-catalysed oxidation of phenol compounds onto xylan was preliminary explored by 13C-NMR. The results showed that PCA-xylan, FA-xylan graft poly onto xylan by Cγ ester bond, SD-xylan graft poly onto xylan by ether bond and an ester bond, and VD-xylan graft poly onto xylan by ether bond. The film strength of xylan derivatives has been significantly increased, especially for the PCA-xylan derivative. The increases in tensile stress at break, tensile strength, tensile yield stress, and Young's modulus were: 24.04%, 31.30%, 55.56%, and 28.21%, respectively. After laccase/phenolics were modified, xylan had a good antibacterial effect to E. coli, Corynebacterium glutamicum, and Bacillus subtilis. The SD-xylan, FA-xylan, and PCA-xylan showed a greater efficacy against E. coli, Corynebacterium glutamicum, and Bacillus subtilis, respectively.

  13. Xylanolytic enzyme systems in Arthrobacter sp MTCC 5214 and Lactobacillus sp.

    Digital Repository Service at National Institute of Oceanography (India)

    Khandeparker, R.; Jalal, T.

    The production of extracellular xylanolytic enzymes such as xylanase, alfa-L-arabinofuranosidase (alfa-l-AFase), and acetyl xylan esterase (Axe) by marine Arthrobacter sp and Lactobacillus sp was investigated using different carbon sources Induction...

  14. Isolation and characterization of acetylated glucuronoarabinoxylan from sugarcane bagasse and straw.

    Science.gov (United States)

    Morais de Carvalho, Danila; Martínez-Abad, Antonio; Evtuguin, Dmitry V; Colodette, Jorge Luiz; Lindström, Mikael E; Vilaplana, Francisco; Sevastyanova, Olena

    2017-01-20

    Sugarcane bagasse and straw are generated in large volumes as by-products of agro-industrial production. They are an emerging valuable resource for the generation of hemicellulose-based materials and products, since they contain significant quantities of xylans (often twice as much as in hardwoods). Heteroxylans (yields of ca 20% based on xylose content in sugarcane bagasse and straw) were successfully isolated and purified using mild delignification followed by dimethyl sulfoxide (DMSO) extraction. Delignification with peracetic acid (PAA) was more efficient than traditional sodium chlorite (NaClO 2 ) delignification for xylan extraction from both biomasses, resulting in higher extraction yields and purity. We have shown that the heteroxylans isolated from sugarcane bagasse and straw are acetylated glucuronoarabinoxylans (GAX), with distinct molecular structures. Bagasse GAX had a slightly lower glycosyl substitution molar ratio of Araf to Xylp to (0.5:10) and (4-O-Me)GlpA to Xylp (0.1:10) than GAX from straw (0.8:10 and 0.1:10 respectively), but a higher degree of acetylation (0.33 and 0.10, respectively). A higher frequency of acetyl groups substitution at position α-(1→3) (Xyl-3Ac) than at position α-(1→2) (Xyl-2Ac) was confirmed for both bagasse and straw GAX, with a minor ratio of diacetylation (Xyl-2,3Ac). The size and molecular weight distributions for the acetylated GAX extracted from the sugarcane bagasse and straw were analyzed using multiple-detection size-exclusion chromatography (SEC-DRI-MALLS). Light scattering data provided absolute molar mass values for acetylated GAX with higher average values than did standard calibration. Moreover, the data highlighted differences in the molar mass distributions between the two isolation methods for both types of sugarcane GAX, which can be correlated with the different Araf and acetyl substitution patterns. We have developed an empirical model for the molecular structure of acetylated GAX extracted from

  15. Hydrolysis kinetics of tulip tree xylan in hot compressed water.

    Science.gov (United States)

    Yoon, Junho; Lee, Hun Wook; Sim, Seungjae; Myint, Aye Aye; Park, Hee Jeong; Lee, Youn-Woo

    2016-08-01

    Lignocellulosic biomass, a promising renewable resource, can be converted into numerous valuable chemicals post enzymatic saccharification. However, the efficacy of enzymatic saccharification of lignocellulosic biomass is low; therefore, pretreatment is necessary to improve the efficiency. Here, a kinetic analysis was carried out on xylan hydrolysis, after hot compressed water pretreatment of the lignocellulosic biomass conducted at 180-220°C for 5-30min, and on subsequent xylooligosaccharide hydrolysis. The weight ratio of fast-reacting xylan to slow-reacting xylan was 5.25 in tulip tree. Our kinetic results were applied to three different reaction systems to improve the pretreatment efficiency. We found that semi-continuous reactor is promising. Lower reaction temperatures and shorter space times in semi-continuous reactor are recommended for improving xylan conversion and xylooligosaccharide yield. In the theoretical calculation, 95% of xylooligosaccharide yield and xylan conversion were achieved simultaneously with high selectivity (desired product/undesired product) of 100 or more. Copyright © 2016. Published by Elsevier Ltd.

  16. Esterase resistant to inactivation by heavy metals

    KAUST Repository

    El, Dorry Hamza

    2014-09-25

    EstATII is an esterase that a halotolerant, thermophilic and resistant to a spectrum of heavy metals including toxic concentration of metals. It was isolated from the lowest convective layer of the Atlantis II Red Sea brine pool. The Atlantis II brine pool is an extreme environment that possesses multiple harsh conditions such as; high temperature, salinity, pH and high concentration of metals, including toxic heavy metals. A fosmid metagenomic library using DNA isolated from the lowest convective layer this pool was used to identify EstATII. Polynucleotides encoding EstATII and similar esterases are disclosed and can be used to make EstATII. EstATII or compositions or apparatuses that contain it may be used in various processes employing lipases/esterases especially when these processes are performed under harsh conditions that inactivate other kinds of lipases or esterases.

  17. Improved biomass degradation using fungal glucuronoyl-esterases-hydrolysis of natural corn fiber substrate.

    Science.gov (United States)

    d'Errico, Clotilde; Börjesson, Johan; Ding, Hanshu; Krogh, Kristian B R M; Spodsberg, Nikolaj; Madsen, Robert; Monrad, Rune Nygaard

    2016-02-10

    Lignin-carbohydrate complexes (LCCs) are in part responsible for the recalcitrance of lignocellulosics in relation to industrial utilization of biomass for biofuels. Glucuronoyl esterases (GEs) belonging to the carbohydrate esterase family 15 have been proposed to be able to degrade ester LCCs between glucuronic acids in xylans and lignin alcohols. By means of synthesized complex LCC model substrates we provide kinetic data suggesting a preference of fungal GEs for esters of bulky arylalkyl alcohols such as ester LCCs. Furthermore, using natural corn fiber substrate we report the first examples of improved degradation of lignocellulosic biomass by the use of GEs. Improved C5 sugar, glucose and glucuronic acid release was observed when heat pretreated corn fiber was incubated in the presence of GEs from Cerrena unicolor and Trichoderma reesei on top of different commercial cellulase/hemicellulase preparations. These results emphasize the potential of GEs for delignification of biomass thereby improving the overall yield of fermentable sugars for biofuel production. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. Penicillium brasilianum as an enzyme factory; the essential role of feruloyl esterases for the hydrolysis of the plant cell wall.

    Science.gov (United States)

    Panagiotou, Gianni; Olavarria, Reyes; Olsson, Lisbeth

    2007-06-30

    The production of arabinoxylan-degrading enzymes by the fungus Penicillium brasilianum, grown on different carbon and nitrogen sources as well as different environmental conditions was investigated. Highest feruloyl esterase (225 mU/ml) and alpha-L-arabinofuranosidase (211 mU/ml) activities were obtained when P. brasilianum was grown on sugar beet pulp, whereas maximum xylanase (17 U/ml) activity was found during growth on oat spelt xylan. Yeast extract was the preferable nitrogen source for the production of all the three enzymes. Further optimization of the production of the crude enzyme mixture was examined by experimental design using a D-optimal quadratic model. Investigation of the microbial regulation of enzyme production showed that the presence of free ferulic acid further stimulated the production and pointing to that the fungal regulatory mechanism involved a coordinated production and secretion of feruloyl esterase, xylanase and alpha-L-arabinofuranosidase. Since agroindustrial by-products are a potential source of phenolic acids, crude enzyme mixtures of P. brasilianum were tested for their hydrolysis abilities against eight complex or model substrates. While total release of phenolic acids and pentoses was not observed, the synergistic enhancement of hydrolysis in the presence of feruloyl esterase was clearly demonstrated.

  19. Xylan polysaccharides fabricated into nanofibrous substrate for myocardial infarction

    Energy Technology Data Exchange (ETDEWEB)

    Venugopal, J., E-mail: nnijrv@nus.edu.sg; Rajeswari, R.; Shayanti, M.; Sridhar, R.; Sundarrajan, S.; Balamurugan, R.; Ramakrishna, S.

    2013-04-01

    Myocardial infarction, a main cause of heart failure, leads to loss of cardiac tissue impairment of left ventricular function. Repair of diseased myocardium with in vitro engineered cardiac muscle patch/injectable biopolymers with cells may become a viable option for myocardial infarction. We attempted to solve these problems by in vitro study by selecting a plant based polysaccharides beech wood Xylan for the normal functioning of infarcted myocardium. The present study fabricated Xylan based nanofibrous scaffolds cross-linked with glutaraldehyde (Glu) vapors for 24 h, 48 h and 1% Glu blended fibers for the culture of neonatal rat cardiac cells for myocardial infarction. These nanofibers were characterized by SEM, FT-IR, tensile testing and cell culture studies for the normal expression of cardiac proteins. The observed results showed that the Xylan/polyvinyl alcohol (PVA) 24 h Glu vapor cross-linked nanofibers (427 nm) having mechanical strength of 2.43 MPa and Young modulus of 3.74 MPa are suitable for the culture of cardiac cells. Cardiac cells proliferation increased only by 11% in Xylan/PVA 24 h Glu cross-linked nanofibers compared to control tissue culture plate (TCP). The normal cardiac cell morphology was observed in 24 h cross-linked Xylan/PVA nanofibers but 48 h cross-linked fibers cell morphology was changed to flattened and elongated on the fibrous surfaces. Confocal analysis for cardiac expression proteins actinin, connexin 43 was observed normally in 24 h Glu cross-linked nanofibers compared to all other nanofibrous scaffolds. The fabricated Xylan/PVA nanofibrous scaffold may have good potential for the normal functioning of infarcted myocardium. - Highlights: ► Fabrication of polysaccharides Xylan/PVA nanofibers for cardiac tissue engineering ► Nanofibers characterized by SEM, FT-IR, tensile testing and cell culture studies ► Isolation of cardiac cells and cultured on Xylan/PVA nanofibrous scaffolds ► Cultured cells on 24 h Glu cross

  20. Xylan polysaccharides fabricated into nanofibrous substrate for myocardial infarction

    International Nuclear Information System (INIS)

    Venugopal, J.; Rajeswari, R.; Shayanti, M.; Sridhar, R.; Sundarrajan, S.; Balamurugan, R.; Ramakrishna, S.

    2013-01-01

    Myocardial infarction, a main cause of heart failure, leads to loss of cardiac tissue impairment of left ventricular function. Repair of diseased myocardium with in vitro engineered cardiac muscle patch/injectable biopolymers with cells may become a viable option for myocardial infarction. We attempted to solve these problems by in vitro study by selecting a plant based polysaccharides beech wood Xylan for the normal functioning of infarcted myocardium. The present study fabricated Xylan based nanofibrous scaffolds cross-linked with glutaraldehyde (Glu) vapors for 24 h, 48 h and 1% Glu blended fibers for the culture of neonatal rat cardiac cells for myocardial infarction. These nanofibers were characterized by SEM, FT-IR, tensile testing and cell culture studies for the normal expression of cardiac proteins. The observed results showed that the Xylan/polyvinyl alcohol (PVA) 24 h Glu vapor cross-linked nanofibers (427 nm) having mechanical strength of 2.43 MPa and Young modulus of 3.74 MPa are suitable for the culture of cardiac cells. Cardiac cells proliferation increased only by 11% in Xylan/PVA 24 h Glu cross-linked nanofibers compared to control tissue culture plate (TCP). The normal cardiac cell morphology was observed in 24 h cross-linked Xylan/PVA nanofibers but 48 h cross-linked fibers cell morphology was changed to flattened and elongated on the fibrous surfaces. Confocal analysis for cardiac expression proteins actinin, connexin 43 was observed normally in 24 h Glu cross-linked nanofibers compared to all other nanofibrous scaffolds. The fabricated Xylan/PVA nanofibrous scaffold may have good potential for the normal functioning of infarcted myocardium. - Highlights: ► Fabrication of polysaccharides Xylan/PVA nanofibers for cardiac tissue engineering ► Nanofibers characterized by SEM, FT-IR, tensile testing and cell culture studies ► Isolation of cardiac cells and cultured on Xylan/PVA nanofibrous scaffolds ► Cultured cells on 24 h Glu cross

  1. Esterase variation in Turkish white-toothed shrews (Crocidura: Record of a trimeric esterase

    Directory of Open Access Journals (Sweden)

    Tez C.

    2009-01-01

    Full Text Available This study focuses on esterase variation of the genus Crocidura in Turkey. A total of 248 white-toothed shrews were analyzed by means of cellulose acetate gel electrophoresis. Liver tissue and alfa naphthyl acetate were used to investigate esterase variation in Turkish white-toothed shrews. A different esterase banding pattern was found in one Crocidura individual. This phenotype had four anodally migrated bands on cellulose acetate gel. The Crocidura individual displaying the given phenotype was identified as Crocidura suaveolens. The different esterase banding pattern observed in this study is considered to be a result of the trimeric structure of esterase in the lesser white-toothed shrew (Crocidura suaveolens.

  2. Characterization of substituents in xylans from corn cobs and stover

    NARCIS (Netherlands)

    Dongen, van F.E.M.; Eylen, van D.; Kabel, M.A.

    2011-01-01

    Structural knowledge on hemicellulose from corn cobs and stover is helpful to better understand their position within the plant cell wall architecture as well as their enzymatic saccharification. In this research different extracts were prepared with water, 1 M and 4 M alkali. Most of the xylans

  3. Molecular Dissection of Xylan Biosynthesis During Wood Formation in Poplar

    Science.gov (United States)

    Xylan, being the second most abundant polysaccharide in dicot wood, is considered to be one of the factors contributing to wood biomass recalcitrance for biofuel production. To better utilize wood as biofuel feedstock, it is crucial to functionally characterize all the genes invo...

  4. [3H]Indole-3-acetyl-myo-inositol hydrolysis by extracts of Zea mays L. vegetative tissue

    Science.gov (United States)

    Hall, P. J.; Bandurski, R. S.

    1986-01-01

    [3H]Indole-3-acetyl-myo-inositol was hydrolyzed by buffered extracts of acetone powders prepared from 4 day shoots of dark grown Zea mays L. seedlings. The hydrolytic activity was proportional to the amount of extract added and was linear for up to 6 hours at 37 degrees C. Boiled or alcohol denatured extracts were inactive. Analysis of reaction mixtures by high performance liquid chromatography demonstrated that not all isomers of indole-3-acetyl-myo-inositol were hydrolyzed at the same rate. Buffered extracts of acetone powders were prepared from coleoptiles and mesocotyls. The rates of hydrolysis observed with coleoptile extracts were greater than those observed with mesocotyl extracts. Active extracts also catalyzed the hydrolysis of esterase substrates such as alpha-naphthyl acetate and the methyl esters of indoleacetic acid and naphthyleneacetic acid. Attempts to purify the indole-3-acetyl-myo-inositol hydrolyzing activity by chromatographic procedures resulted in only slight purification with large losses of activity. Chromatography over hydroxylapatite allowed separation of two enzymically active fractions, one of which catalyzed the hydrolysis of both indole-3-acetyl-myo-inositol and esterase substrates. With the other enzymic hydrolysis of esterase substrates was readily demonstrated, but no hydrolysis of indole-3-acetyl-myo-inositol was ever detected.

  5. Reconstitution of a thermostable xylan-degrading enzyme mixture from the bacterium Caldicellulosiruptor bescii.

    Science.gov (United States)

    Su, Xiaoyun; Han, Yejun; Dodd, Dylan; Moon, Young Hwan; Yoshida, Shosuke; Mackie, Roderick I; Cann, Isaac K O

    2013-03-01

    Xylose, the major constituent of xylans, as well as the side chain sugars, such as arabinose, can be metabolized by engineered yeasts into ethanol. Therefore, xylan-degrading enzymes that efficiently hydrolyze xylans will add value to cellulases used in hydrolysis of plant cell wall polysaccharides for conversion to biofuels. Heterogeneous xylan is a complex substrate, and it requires multiple enzymes to release its constituent sugars. However, the components of xylan-degrading enzymes are often individually characterized, leading to a dearth of research that analyzes synergistic actions of the components of xylan-degrading enzymes. In the present report, six genes predicted to encode components of the xylan-degrading enzymes of the thermophilic bacterium Caldicellulosiruptor bescii were expressed in Escherichia coli, and the recombinant proteins were investigated as individual enzymes and also as a xylan-degrading enzyme cocktail. Most of the component enzymes of the xylan-degrading enzyme mixture had similar optimal pH (5.5 to ∼6.5) and temperature (75 to ∼90°C), and this facilitated their investigation as an enzyme cocktail for deconstruction of xylans. The core enzymes (two endoxylanases and a β-xylosidase) exhibited high turnover numbers during catalysis, with the two endoxylanases yielding estimated k(cat) values of ∼8,000 and ∼4,500 s(-1), respectively, on soluble wheat arabinoxylan. Addition of side chain-cleaving enzymes to the core enzymes increased depolymerization of a more complex model substrate, oat spelt xylan. The C. bescii xylan-degrading enzyme mixture effectively hydrolyzes xylan at 65 to 80°C and can serve as a basal mixture for deconstruction of xylans in bioenergy feedstock at high temperatures.

  6. Characterization of oligomeric xylan structures from corn fiber resistant to pretreatment and simultaneous saccharification and fermentation.

    Science.gov (United States)

    Appeldoorn, Maaike M; Kabel, Mirjam A; Van Eylen, David; Gruppen, Harry; Schols, Henk A

    2010-11-10

    Corn fiber, a byproduct from the corn industry, would be a good source for bioethanol production if the hemicellulose, consisting of polymeric glucoronoarabinoxylans, can be degraded into fermentable sugars. Structural knowledge of the hemicellulose is needed to improve the enzymatic hydrolyses of corn fiber. Oligosaccharides that resisted a mild acid pretreatment and subsequent enzymatic hydrolysis, representing 50% of the starting material, were fractionated on reversed phase and size exclusion material and characterized. The oligosaccharides within each fraction were highly substituted by various compounds. Oligosaccharides containing uronic acid were accumulated in two polar fractions unless also a feruloyl group was present. Feruloylated oligosaccharides, containing mono- and/or diferulic acid, were accumulated within four more apolar fractions. All fractions contained high amounts of acetyl substituents. The data show that complex xylan oligomers are present in which ferulic acid, diferulates, acetic acid, galactose, arabinose, and uronic acids were combined within an oligomer. Hypothetical structures are discussed, demonstrating which enzyme activities are lacking to fully degrade corn glucuronoarabinoxylans.

  7. The mechanism of xylans removal during hydrothermal pretreatment of poplar fibers investigated by immunogold labeling.

    Science.gov (United States)

    Ma, Jing; Ji, Zhe; Chen, Jia C; Zhou, Xia; Kim, Yoon S; Xu, Feng

    2015-07-01

    Hydrothermal pretreatment initially removed the lignin-free xylan from the middle layer of secondary wall, followed by the lignin-bound xylan, but the cellulose-bound xylan was seldom removed by this pretreatment. An in-depth understanding of the mechanism of xylan removal during hydrothermal pretreatment (HTP) of wood is critical for cost-effective conversion of lignocellulosic biomass to biofuels. Several studies demonstrated the kinetics and mechanism of xylan removal during HTP on molecular scale, but the dissolution mechanism of xylan during HTP remains unclear at ultra-structural level. Our study investigated changes in the micro-distribution of xylan in poplar fiber cell walls during HTP by transmission electron microscopy (TEM) in combination with immunogold labeling. The study revealed that HTP caused greater decline in the density of xylan labeling in the S2 layer of fiber wall than in the S1 layer. There was a greater loss in the density of xylan labeling during HTP in the delignified and enzymatically treated fibers compared to untreated fibers. We propose that in the initial stages of HTP lignin-free xylan in the S2 layer was more readily hydrolyzed than in the S1 layer by hydronium ions. With increasing pretreatment time, the xylan covalently bound to lignin was also removed from the S2 layer due to the dissolution of lignin. The xylan tightly bound to cellulose was seldom removed during HTP, but was hydrolyzed in subsequent enzymatic treatment. This TEM-immunolabeling investigation reveals the manner in which different xylan fractions are removed from fiber cell wall during HTP, and we expect the information to be helpful in developing processes tailored for more effective conversion of cellulosic biomass into fermentable sugars.

  8. The effect of fixation on esterases

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D

    1984-01-01

    The localization of reaction product for non-specific esterase from fresh and aldehyde treated glandular tissue was examined. The electrophoretical studies showed a selective inhibition of certain isoenzymes and a change in mobility of some bands caused by aldehyde fixation. In sections a granular...

  9. x ORIGINAL ARTICLE ASSESSMENT OF LEUKOCYTE ESTERASE ...

    African Journals Online (AJOL)

    Dr Oboro VO

    ABSTRACT. This is a prospective study of urinary tract infection in 65 children (38 males and 27 females, M: F ratio 1: 0.7). Urine samples were evaluated by culture, microscopy and leukocyte esterase dipstick test. Positive urine culture, with significant bacteriuria was found in 19(29.2%). Urine microscopy for leukocyturia ...

  10. Phenol esterase activity of porcine skin

    Science.gov (United States)

    The alkyl esters of plant-derived phenols may serve as slow-release sources for cutaneous delivery of antioxidants. The ability of skin esterases to hydrolyze phenolic esters was examined. Esters of tyrosol and hydroxytyrosol were prepared from decanoic and lipoic acids. Ferulic acid was esterified ...

  11. Selection and production of insoluble xylan hydrolyzing enzyme by ...

    African Journals Online (AJOL)

    Jane

    2011-03-07

    Mar 7, 2011 ... The effect of pH and temperature on the enzyme activity and stability of crude enzyme produced by T. lanuginosus THKU 56 were investigated. To study the effect of pH on activity, the reaction mixture of 0.5 ml of enzyme and 0.5 ml of 1% insoluble oat spelt xylan in 50 mM buffers with various pH values ...

  12. Properties of plasticized composite films prepared from nanofibrillated cellulose and birch wood xylan

    DEFF Research Database (Denmark)

    Hansen, Natanya Majbritt Louie; Blomfeldt, Thomas O. J.; Hedenqvist, Mikael S.

    2012-01-01

    Xylans, an important sub-class of hemicelluloses, represent a largely untapped resource for new renewable materials derived from biomass. As with other carbohydrates, nanocellulose reinforcement of xylans is interesting as a route to new bio-materials. With this in mind, birch wood xylan was comb......Xylans, an important sub-class of hemicelluloses, represent a largely untapped resource for new renewable materials derived from biomass. As with other carbohydrates, nanocellulose reinforcement of xylans is interesting as a route to new bio-materials. With this in mind, birch wood xylan...... was combined with nanofibrillated cellulose (NFC) and films were cast with and without glycerol, sorbitol or methoxypolyethylene glycol (MPEG) as plasticizers. Microscopy revealed some NFC agglomeration in the composite films as well as a layered nanocellulose structure. Equilibrium moisture content...

  13. Study on the Modification of Bleached Eucalyptus Kraft Pulp Using Birch Xylan

    Science.gov (United States)

    Wenjia Han; Chuanshan Zhao; Thomas Elder; Rendang Yang; Dongho Kim; Yunqiao Pu; Jeffery Hsieh; Arthur J. Ragauskas

    2012-01-01

    In this study, birch xylan was deposited onto elementally chlorine free (ECF) bleached eucalyptus kraft pulp, and the corresponding changes in physical properties were determined. An aqueous 5% birch xylan solution at pH 9 was added to 5 wt% slurry of bleached kraft eucalyptus fibers, with stirring at 70 C for 15 min after which the pH was adjusted to 5–6. The xylan...

  14. Characterization of esterases from Cucurbita pepo cv. "Eskandrani".

    Science.gov (United States)

    Fahmy, Afaf S; Abo-Zeid, Amal Z; Mohamed, Tarek M; Ghanem, Hala M; Borai, Ibrahim H; Mohamed, Saleh A

    2008-01-01

    Two of the six esterases identified in Cucurbita pepo cv. "Eskandrani" were purified to homogeneity using two chromatography steps: anion exchange and gel filtration. The molecular weights of C. pepo esterases EIc and EII were 50,000 +/- 1500 and 68,000 +/- 1900 Da from gel filtration and 47,000 and 66,000 Da from SDS/PAGE, respectively, suggesting a monomeric structure for both enzymes. Esterases EIc and EII had K(m) values of 1.22 and 1.56 mM and pH optima at 9.0 and 8.0, respectively. The substrate specificity of C. pepo esterases EIc and EII were determined for a number of p-nitrophenyl esters, where their affinity toward these substrates were decreased as carbon atom number increased. Esterases EIc and EII had the same temperature optima, 40 degrees C. Thermal stability studies of esterases EIc and EII indicated that half maximal activities of EIc and EII esterases were reached at 55 degrees C and 50 degrees C, while they lost 45%, 51% and 70%, 77% of their activities after 30 and 90 min of incubation at 40 degrees C, respectively. The effect of different metal cations and inhibitors were examined. The inhibition studies revealed that the active sites of the two esterases contain serine and cysteine residues. The characteristics of C. pepo esterases are closely similar to those of microbial esterases used in food processing and food industry.

  15. Xylanase Activity of Streptomyces violascences BF 3.10 on Xylan Corncobs and its Xylooligosaccharide Production

    Directory of Open Access Journals (Sweden)

    W. Salupi

    2015-04-01

    Full Text Available Corn is one of the important carbohydrate sources in Indonesia that is mainly used for food and industrial materials. In addition, the byproducts of corn, such as corncobs, have been reported as xylan-containing materials that can be utilized as substrate in xylooligosaccharides (XOS production. XOS are natural prebiotic fibers that can enhance the performance of animal’s digestive system. The main objective of this study was to exploit xylan from corncobs to produce XOS. The research consisted of extraction and production of xylan from corncobs and the synthesis of XOS from corncob-produced xylan. The corncob and Streptomyces violascens BF 3.10 xylanase is a collection of PPSHB IPB Laboratory. Corncobs xylan extracted by using alkaline method and reducting sugar was analyzed by dinitrosalicylic acid method. The xylan extraction from corncobs could produce 7.93% (w/w of xylan. The activity of S. violascens BF 3.10 xylanase on the substrate of concorb-produced xylan was 6.4 U/mL at the optimum temperature of 60 °C in 50 mM phosphate buffer with pH 5.5. The thin layer chromatography analysis indicated that 1% (w/v corn-cob xylan could produce XOS with degree of polymerization (DP 3.92. XOS, with DP ranging from 2-4, could be used as a livestock feed mixture to stimulate the growth of normal microbes in the gastrointestinal tract of livestock.

  16. Biochemical and Structural Analyses of Two Cryptic Esterases in Bacteroides intestinalis and their Synergistic Activities with Cognate Xylanases.

    Science.gov (United States)

    Wefers, Daniel; Cavalcante, Janaina J V; Schendel, Rachel R; Deveryshetty, Jaigeeth; Wang, Kui; Wawrzak, Zdzislaw; Mackie, Roderick I; Koropatkin, Nicole M; Cann, Isaac

    2017-08-04

    Arabinoxylans are constituents of the human diet. Although not utilizable by the human host, they can be fermented by colonic bacteria. The arabinoxylan backbone is decorated with arabinose side chains that may be substituted with ferulic acid, thus limiting depolymerization to fermentable sugars. We investigated the polypeptides encoded by two genes upregulated during growth of the colonic bacterium Bacteroides intestinalis on wheat arabinoxylan. The recombinant proteins, designated BiFae1A and BiFae1B, were functionally assigned esterase activities. Both enzymes were active on acetylated substrates, although each showed a higher ferulic acid esterase activity on methyl-ferulate. BiFae1A showed a catalytic efficiency of 12mM s -1 on para-nitrophenyl-acetate, and on methyl-ferulate, the value was 27 times higher. BiFae1B showed low catalytic efficiencies for both substrates. Furthermore, the two enzymes released ferulic acid from various structural elements, and NMR spectroscopy indicated complete de-esterification of arabinoxylan oligosaccharides from wheat bran. BiFae1A is a tetramer based on the crystal structure, whereas BiFae1B is a dimer in solution based on size exclusion chromatography. The structure of BiFae1A was solved to 1.98Å resolution, and two tetramers were observed in the asymmetric unit. A flexible loop that may act as a hinge over the active site and likely coordinates critical interactions with the substrate was prominent in BiFae1A. Sequence alignments of the esterase domains in BiFae1B with the feruloyl esterase from Clostridium thermocellum suggest that both domains lack the flexible hinge in BiFae1A, an observation that may partly provide a molecular basis for the differences in activities in the two esterases. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. Esterase detoxification of acetylcholinesterase inhibitors by ...

    Science.gov (United States)

    Organophosphate (OP) and N-methylcarbamate pesticides inhibit acetylcholinesterase (AChE), but differences in metabolism and detoxification can influence potency of these pesticides across and within species. Carboxylesterase (CaE) and A-esterase (paraoxonase, PON) are considered factors underlying age-related sensitivity differences. We used an in vitro system to measure detoxification of AChE-inhibiting pesticides mediated via these esterases. Recombinant human AChE was used as a bioassay of inhibitor concentration following incubation with detoxifying tissue: liver plus Ca+2 (to stimulate PONs, measuring activity of both esterases) or EGTA (to inhibit PONs, thereby measuring CaE activity). Inhibitory concentrations of aldicarb, chlorpyrifos oxon, malaoxon, methamidophos, oxamyl, paraoxon, and methyl paraoxon were incubated with liver from adult male rat or one of 20 commercially provided human (11-83 years of age) liver samples. Detoxification was the difference in inhibition produced by the pesticide alone or in combination with liver plus Ca+2 or EGTA. Generally, rat liver produced more detoxification than did the human samples. There were large detoxification differences, which were not correlated with age or sex, across human samples for some pesticides (especially malaoxon, chlorpyrifos oxon) but not for others (e.g., aldicarb, methamidophos). Chlorpyrifos oxon was detoxified only in the presence of Ca+2 in both rat and human livers. Detoxification of pa

  18. Synthesis and characterization of birch wood xylan succinoylated in 1-n-butyl-3-methylimidazolium chloride

    DEFF Research Database (Denmark)

    Hansen, Natanya Majbritt Louie; Plackett, David

    2011-01-01

    analysis of the modified and native xylans showed a slight lowering of thermal stability with functionalization. Contact angle measurements on spin-coated surfaces of modified xylan films showed a significant increase in hydrophobicity with the introduction of the alkenyl-functionalized succinic anhydride...

  19. Hydrothermally treated xylan rich by-products yield different classes of xylo-oligosaccharides

    NARCIS (Netherlands)

    Kabel, M.A.; Carvalheiro, F.; Garrote, G.; Avgerinos, E.; Koukios, E.; Parajo, J.C.; Girio, M.

    2002-01-01

    Four xylan rich by-products, namely wheat bran, brewery's spent grain, corn cobs and Eucalyptus wood, were characterised and subjected to a mild hydrothermal treatment in order to release and degrade the xylan from the starting materials. The chemical characterisation of the feedstock materials,

  20. High oxygen nanocomposite barrier films based on xylan and nanocrystalline cellulose

    Science.gov (United States)

    Amit Saxena; Thomas J. Elder; Jeffrey Kenvin; Arthur J. Ragauskas

    2010-01-01

    The goal of this work is to produce nanocomposite film with low oxygen permeability by casting an aqueous solution containing xylan, sorbitol and nanocrystalline cellulose. The morphology of the resulting nanocomposite films was examined by scanning electron microscopy and atomic force microscopy which showed that control films containing xylan and sorbitol had a more...

  1. Engineering the Xylan Utilization System in Bacillus subtilis for Production of Acidic Xylooligosaccharides

    Science.gov (United States)

    Mun Su Rhee; Lusha Wei; Neha Sawhney; John D. Rice; Franz J. St. John; Jason C. Hurlbert; James F. Preston

    2014-01-01

    Xylans are the predominant polysaccharides in hemicelluloses and an important potential source of biofuels and chemicals. The ability of Bacillus subtilis subsp. subtilis strain 168 to utilize xylans has been ascribed to secreted glycoside hydrolase family 11 (GH11) and GH30 endoxylanases, encoded by the xynA and...

  2. Diversity in Production of Xylan-Degrading Enzymes Among Species Belonging to the Trichoderma Section Longibrachiatum

    NARCIS (Netherlands)

    Toth, K.; Gool, van M.P.; Schols, H.A.; Samuels, G.J.; Gruppen, H.; Szakacs, G.

    2013-01-01

    Xylan is an important part of plant biomass and represents a renewable raw material for biorefineries. Contrary to cellulose, the structure of hemicellulose is quite complex. Therefore, the biodegradation of xylan needs the cooperation of many enzymes. For industrial production of xylanase

  3. From plant biomass to bio-based chemicals: latest developments in xylan research.

    Science.gov (United States)

    Deutschmann, Rudolf; Dekker, Robert F H

    2012-01-01

    For a hundred years or more, oil and natural gas has supplied fuel and other raw chemicals to support economic growth. In the last decades their shrinking reservoirs and the increasing cost of production has become obvious, leading researchers to look for alternative substitutes of all the chemical materials presently derived from oil and gas. This review is focused on xylan, the second most abundant plant polysaccharide on our planet. Some xylan-derived products have already found commercial applications (ethanol, xylitol, xylo-oligosaccharides) while others could have a great future in a wide range of industries. The chemical and structural variations of xylans produced by different plants, and the concentration of xylan in various plant resources are summarized. This review discusses the latest research developments in extraction and purification methodologies, and chemical modification, as well as the analytical methods necessary for xylan related research. Crown Copyright © 2012. Published by Elsevier Inc. All rights reserved.

  4. Adsorption capacity of phenolic compounds onto cellulose and xylan

    Directory of Open Access Journals (Sweden)

    Telma dos Santos Costa

    2015-06-01

    Full Text Available The interaction between three phenolic compounds (catechin, caffeic acid and ferulic acid onto two dietary fibres (cellulose and xylan has been evaluated to inquire possible interferences on the biodisponibility of phenolic compounds. The adsorption kinetics were performed using solutions containing 100 mg/L of phenolic compounds during a contact time ranging between 10 and 120 minutes at pH 2.0, 4.5, and 7.0. After the kinetics, isotherms were obtained using phenolic compounds concentration ranging between 10 and 80 mg/L during 60 minutes, at pH 2.0 and 7.0 and temperature of 36 °C. Results indicate that adsorbed quantities mainly changed in function of pH, however the maximum adsorption was only of 0.978 mg of caffeic acid/g of xylan at pH 2 and after 60 min. Redlich-Peterson model were able to predict the adsorption isotherms of all phenolic compounds onto cellulose, except for caffeic acid at pH 7.0. The low adsorption capacities observed suggest that both dietary fibres are unable to compromise the biodisponibility of phenolic compounds, especially in the small intestine, where they are partially absorbed.

  5. Non-specific esterases in partly mineralized bovine enamel

    DEFF Research Database (Denmark)

    Moe, D; Kirkeby, S

    1990-01-01

    Activity for non-specific esterase was demonstrated in the matrix of developing bovine enamel with alpha-naphthyl acetate and 5-bromoindoxyl acetate as the esterase substrates. By use of high-performance liquid chromatography gel filtration, ion-exchange chromatography, and electrophoresis three ...

  6. Usefulness of C1 Esterase Inhibitor Protein Concentrate in the ...

    African Journals Online (AJOL)

    2018-04-04

    Apr 4, 2018 ... concentrate in a patient with mild‑to‑moderate dyspnea caused by swelling of the upper airway (larynx) and tongue. Keywords: C1 esterase inhibitor protein, hereditary angioedema, laryngeal edema, oropharyngeal swelling. Usefulness of C1 Esterase Inhibitor Protein Concentrate in the. Management of ...

  7. Non-specific esterases in partly mineralized bovine enamel

    DEFF Research Database (Denmark)

    Moe, D; Kirkeby, S

    1990-01-01

    esterases were shown to be present in the enamel matrix. The enzymes showed highest activity at pH 6.5-7.5. In sections a strong reaction was observed in the secretory ameloblasts. The esterases may be proteolytic enzymes that participate in the degradation of the matrix proteins....

  8. Xylooligosaccharide Production from Tobacco Stalk Xylan using Xylanase Streptomyces sp. BO 3.2

    Directory of Open Access Journals (Sweden)

    Muhammad Nur Kholis

    2015-06-01

    Full Text Available Tobacco stalk (TS, which is one type of lignocellulosic material, has a xylan content of up to 21.9%. Lignocellulose can be used to produce xylooligosaccharides (XOs. XOs are dietary fibers that have prebiotic activity. This study aimed to produce XOs from tobacco stalk xylan using xylanase from Streptomyces sp. BO 3.2. After the TS was delignified, the xylan was extracted using the alkali method. The delignification process, which used 1% natrium hypoclorite (NaOCl, decreased the lignins from 32.93% to 18.15%. Xylan extraction was conducted using 10% natrium hydoroxide (NaOH; this extraction produced xylan of 15.53% (w/w. The xylanase produced by Streptomyces sp. BO 3.2 on a 0.5% TS medium had 5.92 U/mL of activity, with the optimum condition occurring at pH 5.5 and a temperature of 60 °C. The xylanase was stable, at temperature 4 °C and 30 °C for 120 hours. The xylanase Streptomyces sp. BO 3.2 was capable of hydrolyzing 2% TS xylan and 2% beechwood xylan during the first, third, sixth, and twelfth hours of incubation time; it also produced XOs with degrees of polymerization (DP of 2.18 and 2.15, respectively. A Thin layer chomatography (TLC analysis indicated that the hydrolysis products were XOs with the absence of xylose, glucose, and arabinose.

  9. Xylan catabolism is improved by blending bioprospecting and metabolic pathway engineering in Saccharomyces cerevisiae.

    Science.gov (United States)

    Lee, Sun-Mi; Jellison, Taylor; Alper, Hal S

    2015-04-01

    Complete utilization of all available carbon sources in lignocellulosic biomass still remains a challenge in engineering Saccharomyces cerevisiae. Even with efficient heterologous xylose catabolic pathways, S. cerevisiae is unable to utilize xylose in lignocellulosic biomass unless xylan is depolymerized to xylose. Here we demonstrate that a blended bioprospecting approach along with pathway engineering and evolutionary engineering can be used to improve xylan catabolism in S. cerevisiae. Specifically, we perform whole genome sequencing-based bioprospecting of a strain with remarkable pentose catabolic potential that we isolated and named Ustilago bevomyces. The heterologous expression of xylan catabolic genes enabled S. cerevisiae to grow on xylan as a single carbon source in minimal medium. A combination of bioprospecting and metabolic pathway evolution demonstrated that the xylan catabolic pathway could be further improved. Ultimately, engineering efforts were able to achieve xylan conversion into ethanol of up to 0.22 g/L on minimal medium compositions with xylan. This pathway provides a novel starting point for improving lignocellulosic conversion by yeast. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Xylella fastidiosa esterase rather than hydroxynitrile lyase.

    Science.gov (United States)

    Torrelo, Guzman; Ribeiro de Souza, Fayene Zeferino; Carrilho, Emanuel; Hanefeld, Ulf

    2015-03-02

    In 2009, we reported that the product of the gene SCJ21.16 (XFa0032) from Xylella fastidiosa, a xylem-restricted plant pathogen that causes a range of diseases in several important crops, encodes a protein (XfHNL) with putative hydroxynitrile lyase activity. Sequence analysis and activity tests indicated that XfHNL exhibits an α/β-hydrolase fold and could be classified as a member of the family of FAD-independent HNLs. Here we provide a more detailed sequence analysis and new experimental data. Using pure heterologously expressed XfHNL we show that this enzyme cannot catalyse the cleavage/synthesis of mandelonitrile and that this protein is in fact a non-enantioselective esterase. Homology modelling and ligand docking simulations were used to study the active site and support these results. This finding could help elucidate the common ancestor of esterases and hydroxynitrile lyases with an α/β -hydrolase fold. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Fitness differences due to allelic variation at Esterase-4 locus in ...

    Indian Academy of Sciences (India)

    KAVITA KRISHNAMOORTI

    2017-08-31

    Aug 31, 2017 ... higher in Esterase-4 active larval haemolymph as well as in mature flies' homogenate than that of Esterase-4 null. Thus, Esterase-4 locus of D. ananassae has its role in fecundity, fertility and productivity of female, life span control and lipid metabolism. Keywords. esterases; null allele; reproductive fitness; ...

  12. Structural Investigation of Cell Wall Xylan Polysaccharides from the Leaves of Algerian Argania spinosa

    Directory of Open Access Journals (Sweden)

    Kadda Hachem

    2016-11-01

    Full Text Available Xylan-type polysaccharides were isolated from the leaves of Argania spinosa (L. Skeels collected in the Tindouf area (southwestern Algeria. Xylan fractions were obtained by sequential alkaline extractions and purified on Sepharose CL-4B. The xylan structure was investigated by enzymatic hydrolysis with an endo-β(1→4-xylanase followed by chromatography of the resulting fragments on Biogel P2, characterization by sugar analysis and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS . The results show that the A. spinosa xylan is composed of a β-(1→4-d-xylopyranose backbone substituted with 4-O-methyl-d-glucuronic acid and L-arabinose residues.

  13. Acetylation dynamics and stoichiometry in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Weinert, Brian Tate; Iesmantavicius, Vytautas; Moustafa, Tarek

    2014-01-01

    Lysine acetylation is a frequently occurring posttranslational modification; however, little is known about the origin and regulation of most sites. Here we used quantitative mass spectrometry to analyze acetylation dynamics and stoichiometry in Saccharomyces cerevisiae. We found that acetylation...

  14. Impact of the fouling mechanism on enzymatic depolymerization of xylan in different configurations of membrane reactors

    DEFF Research Database (Denmark)

    Mohd Sueb, Mohd Shafiq Bin; Luo, Jianquan; Meyer, Anne S.

    2017-01-01

    In order to maximize enzymatic xylan depolymerization while simultaneously purifying the resulting monosaccharide (xylose), different ultrafiltration (UF) membrane reactor configurations were evaluated. Initial results showed that the two hydrolytic enzymes required for complete depolymerization......) and the simultaneous reaction-filtration with both enzymes, respectively. This study thus confirmed that the reactor configuration has a crucial impact on the performance of both the reaction and the separation process of xylose during enzymatic xylan degradation, and that the type of fouling mechanism varies...

  15. Characterization and distribution of esterase activity in activated sludge

    NARCIS (Netherlands)

    Boczar, BA; Forney, LJ; Begley, WM; Larson, RJ; Federle, TW

    2001-01-01

    The location and activity of esterase enzymes in activated Sludge from three Municipal wastewater treatment plants were characterized using model Substrate, and denaturing and nondenaturing polyacrylamide gel electrophoresis (PAGE) Of particulate, freeze thaw (primarily periplasmic enzymes and those

  16. Esterase activity as a novel parameter of spore germination in Bacillus anthracis

    International Nuclear Information System (INIS)

    Ferencko, Linda; Cote, Mindy A.; Rotman, Boris

    2004-01-01

    Spores of Bacillus anthracis were shown to produce esterase activity about 4 min after exposure to conventional germinants such as combinations of amino acids and purine ribosides. Neither amino acids nor ribosides alone induce germination and esterase activity. Expression of esterase activity was chloramphenicol resistant, and correlated with loss of spore refractivity, a traditional parameter of early germination. Based on these observations, we hypothesized that esterase activity could be used as a novel parameter for quantifying early events during spore germination. To test this hypothesis, we measured expression of esterase activity under a variety of germinating conditions. Using diacetyl fluorescein as fluorogenic substrate of esterases, we demonstrated that esterase activity was invariably induced whenever spores were triggered by known germinants. Moreover, D-alanine, an inhibitor of L-alanine-mediated germination, was found to significantly inhibit expression of esterase activity. In terms of molecular mechanisms, esterase expression could represent activation of proteases at the onset of spore germination

  17. Xylan synthetase activity in differentiated xylem cells of sycamore trees (Acer pseudoplatanus).

    Science.gov (United States)

    Dalessandro, G; Northcote, D H

    1981-01-01

    Particulate enzymic preparations obtained from homogenates of differentiated xylem cells isolated from sycamore trees, catalyzed the formation of a radioactive xylan in the presence of UDP-D-[U-(14)C]xylose as substrate. The synthesized xylan was not dialyzable through Visking cellophane tubing. Successive extraction with cold water, hot water and 5% NaOH dissolved respectively 15, 5 and 80% of the radioactive polymer. Complete acid hydrolysis of the water-insoluble polysaccharide synthesized from UDP-D-[U-(14)C]xylose released all the radioactivity as xylose. β-1,4-Xylodextrins, degree of polymerization 2, 3, 4, 5 and 6, were obtained by partial acid hydrolysis (fuming HCl or 0.1 M HCl) of radioactive xylan. The polymer was hydrolysed to xylose, xylobiose and xylotriose by Driselase which contains 1,4-β xylanase activities. Methylation and then hydrolysis of the xylan released two methylated sugars which were identified as di-O-methyl[(14)C]xylose and tri-O-methyl-[(14)C]xylose, suggesting a 1→4-linked polymer. The linkage was confirmed by periodate oxidation studies. The apparent Km value of the synthetase for UDP-D-xylose was 0.4 mM. Xylan synthetase activity was not potentiated in the presence of a detergent. The enzymic activity was stimulated by Mg(2+) and Mn(2+) ions, although EDTA in the range of concentrations between 0.01 and 1 mM did not affect the reaction rate. It appears that the xylan synthetase system associated with membranes obtained from differentiated xylem cells of sycamore trees may serve for catalyzing the in vivo synthesis of the xylan main chain during the biogenesis of the plant cell wall.

  18. Direct xylan conversion into glycolipid biosurfactants, mannosylerythritol lipids, by Pseudozyma antarctica PYCC 5048(T).

    Science.gov (United States)

    Faria, Nuno Torres; Marques, Susana; Fonseca, César; Ferreira, Frederico Castelo

    2015-04-01

    Mannosylerythritol lipids (MEL) are glycolipid biosurfactants, produced by Pseudozyma spp., with increasing commercial interest. While MEL can be produced from d-glucose and d-xylose, the direct conversion of the respective lignocellulosic polysaccharides, cellulose and xylan, was not reported yet. The ability of Pseudozyma antarctica PYCC 5048(T) and Pseudozyma aphidis PYCC 5535(T) to use cellulose (Avicel(®)) and xylan (beechwood) as carbon and energy source has been assessed along with their capacity of producing cellulolytic and hemicellulolytic enzymes, toward a consolidated bioprocess (CBP) for MEL production. The yeasts assessed were neither able to grow in medium containing Avicel(®) nor produce cellulolytic enzymes under the conditions tested. On contrary, both yeasts were able to efficiently grow in xylan, but MEL production was only detected in P. antarctica PYCC 5048(T) cultures. MEL titers reached 1.3g/l after 10 days in batch cultures with 40g/l xylan, and 2.0g/l in fed-batch cultures with xylan feeding (additional 40g/l) at day 4. High levels of xylanase activities were detected in xylan cultures, reaching 47-62U/ml (31-32U/mg) at 50°C, and still exhibiting more than 10U/ml under physiological temperature (28°C). Total β-xylosidase activities, displayed mainly as wall-bounded and extracellular activity, accounted for 0.154 and 0.176U/ml in P. antarctica PYCC 5048(T) and P. aphidis PYCC 5535(T) cultures, respectively. The present results demonstrate the potential of Pseudozyma spp. for using directly a fraction of lignocellulosic biomass, xylan, and combining in the same bioprocess the production of xylanolytic enzymes with MEL production. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Determinants and Prediction of Esterase Substrate Promiscuity Patterns.

    Science.gov (United States)

    Martínez-Martínez, Mónica; Coscolín, Cristina; Santiago, Gerard; Chow, Jennifer; Stogios, Peter J; Bargiela, Rafael; Gertler, Christoph; Navarro-Fernández, José; Bollinger, Alexander; Thies, Stephan; Méndez-García, Celia; Popovic, Ana; Brown, Greg; Chernikova, Tatyana N; García-Moyano, Antonio; Bjerga, Gro E K; Pérez-García, Pablo; Hai, Tran; Del Pozo, Mercedes V; Stokke, Runar; Steen, Ida H; Cui, Hong; Xu, Xiaohui; Nocek, Boguslaw P; Alcaide, María; Distaso, Marco; Mesa, Victoria; Peláez, Ana I; Sánchez, Jesús; Buchholz, Patrick C F; Pleiss, Jürgen; Fernández-Guerra, Antonio; Glöckner, Frank O; Golyshina, Olga V; Yakimov, Michail M; Savchenko, Alexei; Jaeger, Karl-Erich; Yakunin, Alexander F; Streit, Wolfgang R; Golyshin, Peter N; Guallar, Víctor; Ferrer, Manuel; The Inmare Consortium

    2018-01-19

    Esterases receive special attention because of their wide distribution in biological systems and environments and their importance for physiology and chemical synthesis. The prediction of esterases' substrate promiscuity level from sequence data and the molecular reasons why certain such enzymes are more promiscuous than others remain to be elucidated. This limits the surveillance of the sequence space for esterases potentially leading to new versatile biocatalysts and new insights into their role in cellular function. Here, we performed an extensive analysis of the substrate spectra of 145 phylogenetically and environmentally diverse microbial esterases, when tested with 96 diverse esters. We determined the primary factors shaping their substrate range by analyzing substrate range patterns in combination with structural analysis and protein-ligand simulations. We found a structural parameter that helps rank (classify) the promiscuity level of esterases from sequence data at 94% accuracy. This parameter, the active site effective volume, exemplifies the topology of the catalytic environment by measuring the active site cavity volume corrected by the relative solvent accessible surface area (SASA) of the catalytic triad. Sequences encoding esterases with active site effective volumes (cavity volume/SASA) above a threshold show greater substrate spectra, which can be further extended in combination with phylogenetic data. This measure provides also a valuable tool for interrogating substrates capable of being converted. This measure, found to be transferred to phosphatases of the haloalkanoic acid dehalogenase superfamily and possibly other enzymatic systems, represents a powerful tool for low-cost bioprospecting for esterases with broad substrate ranges, in large scale sequence data sets.

  20. Biochemical characterization of xylan xylosyltransferases involved in wood formation in poplar.

    Science.gov (United States)

    Lee, Chanhui; Zhong, Ruiqin; Ye, Zheng-Hua

    2012-03-01

    The major polysaccharides in dicot wood biomass are cellulose and xylan. Although wood-associated cellulose synthase genes responsible for cellulose biosynthesis have been characterized, wood-associated xylan synthase genes have not been biochemically identified. A recent report by Lee et al. (2012) provides the first biochemical evidence that two functionally non-redundant Arabidopsis GT43 members are xylosyltransferases (XylTs) that function cooperatively in the elongation of the xylan backbone. We further extend this finding in the current report demonstrating that two poplar (Populus trichocarpa) GT43 glycosyltransferases, PtrGT43B and PtrGT43C, are xylan XylTs involved in wood formation. We show that microsomes from transgenic tobacco BY2 cells coexpressing PtrGT43B and PtrGT43C exhibited a high XylT activity capable of generating β-(1,4)-linked xylooligosaccharides, whereas little XylT activity was detected in microsomes with expression of PtrGT43B or PtrGT43C alone. These findings indicate that poplar GT43 members are XylTs that act cooperatively in catalyzing the successive transfer of xylosyl residues during xylan backbone biosynthesis, which provides further support of the hypothesis that the biochemical functions of GT43 members in vascular plants are evolutionarily conserved.

  1. Carboxymethylation of alkali extracted xylan for preparation of bio-based packaging films.

    Science.gov (United States)

    Alekhina, Marina; Mikkonen, Kirsi S; Alén, Raimo; Tenkanen, Maija; Sixta, Herbert

    2014-01-16

    This study describes the synthesis of carboxymethylxylan (CMX) and investigates its suitability as a film for packaging applications. High-purity polymeric xylan was extracted from commercial bleached birch kraft pulp and converted to CMX with three different degrees of substitution (DSs). The water vapor sorption, mechanical, and barrier properties of the films prepared from CMX were tested. Increasing DS of CMX films resulted in an increase in elongation at break and a decrease in tensile strength and Young's modulus. The DS also affected the barrier properties of the films. CMX films with higher DS showed improved (reduced) oxygen permeability (OP), and the water vapor permeability (WVP) increased with DS. It was demonstrated that the carboxymethylation of xylan recovered from industrial side-streams and its conversion to packaging films could be a viable option to valorize xylan. Copyright © 2013 Elsevier Ltd. All rights reserved.

  2. Biomimetic Inks Based on Cellulose Nanofibrils and Cross-Linkable Xylans for 3D Printing.

    Science.gov (United States)

    Markstedt, Kajsa; Escalante, Alfredo; Toriz, Guillermo; Gatenholm, Paul

    2017-11-22

    This paper presents a sustainable all-wood-based ink which can be used for 3D printing of constructs for a large variety of applications such as clothes, furniture, electronics, and health care products with a customized design and versatile gel properties. The 3D printing technologies where the material is dispensed in the form of liquids, so called inks, have proven suitable for 3D printing dispersions of cellulose nanofibrils (CNFs) because of their unique shear thinning properties. In this study, novel inks were developed with a biomimetic approach where the structural properties of cellulose and the cross-linking function of hemicelluloses that are found in the plant cell wall were utilized. The CNF was mixed with xylan, a hemicellulose extracted from spruce, to introduce cross-linking properties which are essential for the final stability of the printed ink. For xylan to be cross-linkable, it was functionalized with tyramine at different degrees. Evaluation of different ink compositions by rheology measurements and 3D printing tests showed that the degree of tyramine substitution and the ratio of CNFs to xylan-tyramine in the prepared inks influenced the printability and cross-linking density. Both two-layered gridded structures and more complex 3D constructs were printed. Similarly to conventional composites, the interactions between the components and their miscibility are important for the stability of the printed and cross-linked ink. Thus, the influence of tyramine on the adsorption of xylan to cellulose was studied with a quartz crystal microbalance to verify that the functionalization had little influence on xylan's adsorption to cellulose. Utilizing xylan-tyramine in the CNF dispersions resulted in all-wood-based inks which after 3D printing can be cross-linked to form freestanding gels while at the same time, the excellent printing properties of CNFs remain intact.

  3. Functional conservation and divergence of Miscanthus lutarioriparius GT43 gene family in xylan biosynthesis.

    Science.gov (United States)

    Wang, Xiaoyu; Tang, Qi; Zhao, Xun; Jia, Chunlin; Yang, Xuanwen; He, Guo; Wu, Aimin; Kong, Yingzhen; Hu, Ruibo; Zhou, Gongke

    2016-04-26

    Xylan is the most abundant un-cellulosic polysaccharides of plant cell walls. Much progress in xylan biosynthesis has been gained in the model plant species Arabidopsis. Two homologous pairs Irregular Xylem 9 (IRX9)/9L and IRX14/14L from glycosyltransferase (GT) family 43 have been proved to play crucial roles in xylan backbone biosynthesis. However, xylan biosynthesis in grass such as Miscanthus remains poorly understood. We characterized seven GT43 members in M. lutarioriparius, a promising bioenergy crop. Quantitative real-time RT-PCR (qRT-PCR) analysis revealed that the expression of MlGT43 genes was ubiquitously detected in the tissues examined. In-situ hybridization demonstrated that MlGT43A-B and MlGT43F-G were specifically expressed in sclerenchyma, while MlGT43C-E were expressed in both sclerenchyma and parenchyma. All seven MlGT43 proteins were localized to Golgi apparatus. Overexpression of MlGT43A-E but not MlGT43F and MlGT43G in Arabidopsis irx9 fully or partially rescued the mutant defects, including morphological changes, collapsed xylem and increased xylan contents, whereas overexpression of MlGT43F and MlGT43G but not MlGT43A-E complemented the defects of irx14, indicating that MlGT43A-E are functional orthologues of IRX9, while MlGT43F and MlGT43G are functional orthologues of IRX14. However, overexpression of all seven MlGT43 genes could not rescue the mucilage defects of irx14 seeds. Furthermore, transient transactivation analyses of MlGT43A-E reporters demonstrated that MlGT43A and MlGT43B but not MlGT43C-E were differentially activated by MlSND1, MlMYB46 or MlVND7. The results demonstrated that all seven MlGT43s are functionally conserved in xylan biosynthesis during secondary cell wall formation but diversify in seed coat mucilage xylan biosynthesis. The results obtained provide deeper insight into xylan biosynthesis in grass, which lay the foundation for genetic modification of grass cell wall components and structure to better suit for next

  4. Imidazole, a New Tunable Reagent for Producing Nanocellulose, Part I: Xylan-Coated CNCs and CNFs

    Directory of Open Access Journals (Sweden)

    Jia Mao

    2017-09-01

    Full Text Available Imidazole is reported to be an effective reactant for the production of nanocellulose from hardwood pulp. The morphologies and surface properties of the nanocellulose can be simply tailored according to the water content in the imidazole system: with pure imidazole, cellulose nanofibrils (CNFs in a yield of 10 wt % can be produced. With 25 wt % of water in imidazole, cellulose nanocrystals (CNCs are obtained in 20 wt % yield. Both nanocelluloses exhibit crystallinity indices in the order of 70%. Interestingly, they retain the original xylan from the pulp with ca. 9–10 wt % of residual xylan content.

  5. Fitness differences due to allelic variation at Esterase-4 Locus in ...

    Indian Academy of Sciences (India)

    Navya

    2017-01-04

    Jan 4, 2017 ... difference for the survivorship between the two genotypes. Triglycerides level was higher in. Esterase-4 active larval haemolymph as well as in mature flies' homogenate than that of. Esterase-4 null. Thus, Esterase-4 locus of D. ananassae has its role in fecundity, fertility and productivity of female, life span ...

  6. Differential bacterial capture and transport preferences facilitate co-growth on dietary xylan in the human gut

    DEFF Research Database (Denmark)

    Leth, Maria Louise; Ejby, Morten; Workman, Christopher

    2018-01-01

    and dynamic association to xylan via four xylan-binding modules. This xylanase operates in concert with an ATP-binding cassette transporter to mediate breakdown and selective internalization of xylan fragments. The transport protein of R. intestinalis prefers oligomers of 4-5 xylosyl units, whereas......Metabolism of dietary glycans is pivotal in shaping the human gut microbiota. However, the mechanisms that promote competition for glycans among gut commensals remain unclear. Roseburia intestinalis, an abundant butyrate-producing Firmicute, is a key degrader of the major dietary fibre xylan....... Despite the association of this taxon to a healthy microbiota, insight is lacking into its glycan utilization machinery. Here, we investigate the apparatus that confers R. intestinalis growth on different xylans. R. intestinalis displays a large cell-attached modular xylanase that promotes multivalent...

  7. Esterase SeE of Streptococcus equi ssp. equi is a novel nonspecific carboxylic ester hydrolase.

    Science.gov (United States)

    Xie, Gang; Liu, Mengyao; Zhu, Hui; Lei, Benfang

    2008-12-01

    Extracellular carboxylic ester hydrolases are produced by many bacterial pathogens and have been shown recently to be important for virulence of some pathogens. However, these hydrolases are poorly characterized in enzymatic activity. This study prepared and characterized the secreted ester hydrolase of Streptococcus equi ssp. equi (designated SeE for S. equi esterase). SeE hydrolyzes ethyl acetate, acetylsalicylic acid, and tributyrin but not ethyl butyrate. This substrate specificity pattern does not match those of the three conventional types of nonspecific carboxylic ester hydrolases (carboxylesterases, arylesterases, and acetylesterases). To determine whether SeE has lipase activity, a number of triglycerides and vinyl esters were tested in SeE-catalyzed hydrolysis. SeE does not hydrolyze triglycerides and vinyl esters of long-chain carboxylic acids nor display interfacial activation, indicating that SeE is not a lipase. Like the conventional carboxylesterases, SeE is inhibited by di-isopropylfluorophosphate. These findings indicate that SeE is a novel carboxylesterase with optimal activity for acetyl esters.

  8. Properties of polyvinyl alcohol/xylan composite films with citric acid.

    Science.gov (United States)

    Wang, Shuaiyang; Ren, Junli; Li, Weiying; Sun, Runcang; Liu, Shijie

    2014-03-15

    Composite films of xylan and polyvinyl alcohol were produced with citric acid as a new plasticizer or a cross-linking agent. The effects of citric acid content and polyvinyl alcohol/xylan weight ratio on the mechanical properties, thermal stability, solubility, degree of swelling and water vapor permeability of the composite films were investigated. The intermolecular interactions and morphology of composite films were characterized by FTIR spectroscopy and SEM. The results indicated that polyvinyl alcohol/xylan composite films had good compatibility. With an increase in citric acid content from 10% to 50%, the tensile strength reduced from 35.1 to 11.6 MPa. However, the elongation at break increased sharply from 15.1% to 249.5%. The values of water vapor permeability ranged from 2.35 to 2.95 × 10(-7)g/(mm(2)h). Interactions between xylan and polyvinyl alcohol in the presence of citric acid become stronger, which were caused by hydrogen bond and ester bond formation among the components during film forming. Copyright © 2013. Published by Elsevier Ltd.

  9. Arabidopsis GUX Proteins Are Glucuronyltransferases Responsible for the Addition of Glucuronic Acid Side Chains onto Xylan

    Science.gov (United States)

    Xylan, the second most abundant cell wall polysaccharide, is composed of a linear backbone of β-(1,4)-linked xylosyl residues that are often substituted with sugar side chains, such as glucuronic acid (GlcA) and methylglucuronic acid (MeGlcA). It has recently been shown that muta...

  10. Xylan hydrolysis in Populus trichocarpa × P. deltoides and model substrates during hydrothermal pretreatment.

    Science.gov (United States)

    Trajano, Heather L; Pattathil, Sivakumar; Tomkins, Bruce A; Tschaplinski, Timothy J; Hahn, Michael G; Van Berkel, Gary J; Wyman, Charles E

    2015-03-01

    Previous studies defined easy and difficult to hydrolyze fractions of hemicellulose that may result from bonds among cellulose, hemicellulose, and lignin. To understand how such bonds affect hydrolysis, Populus trichocarpa × Populus deltoides, holocellulose isolated from P. trichocarpa × P. deltoides and birchwood xylan were subjected to hydrothermal flow-through pretreatment. Samples were characterized by glycome profiling, HPLC, and UPLC-MS. Glycome profiling revealed steady fragmentation and removal of glycans from solids during hydrolysis. The extent of polysaccharide fragmentation, hydrolysis rate, and total xylose yield were lowest for P. trichocarpa × P. deltoides and greatest for birchwood xylan. Comparison of results from P. trichocarpa × P. deltoides and holocellulose suggested that lignin-carbohydrate complexes reduce hydrolysis rates and limit release of large xylooligomers. Smaller differences between results with holocellulose and birchwood xylan suggest xylan-cellulose hydrogen bonds limited hydrolysis, but to a lesser extent. These findings imply cell wall structure strongly influences hydrolysis. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Digestive and physiological effects of a wheat bran extract, arabino-xylan-oligosaccharide, in breakfast cereal

    Science.gov (United States)

    We assessed whether a wheat bran extract containing arabino-xylan-oligosaccharide (AXOS) elicited a prebiotic effect and influenced other physiologic parameters when consumed in ready-to-eat cereal at two dose levels. This double-blind, randomized, controlled, crossover trial evaluated the effects o...

  12. Usefulness of C1 Esterase Inhibitor Protein Concentrate in the ...

    African Journals Online (AJOL)

    2018-04-04

    Apr 4, 2018 ... of this case report is to describe the lifesaving use of a novel C1‑INH protein concentrate in a patient with mild‑to‑moderate dyspnea caused by swelling of the upper airway (larynx) and tongue. Keywords: C1 esterase inhibitor protein, hereditary angioedema, laryngeal edema, oropharyngeal swelling.

  13. Assessment Of Leukocyte Esterase Dipstick Test In Diagnosis Of ...

    African Journals Online (AJOL)

    This is a prospective study of urinary tract infection in 65 children (38 males and 27 females, M: F ratio 1: 0.7). Urine samples were evaluated by culture, microscopy and leukocyte esterase dipstick test. Positive urine culture, with significant bacteriuria was found in 19(29.2%). Urine microscopy for leukocyturia identified ...

  14. New cholesterol esterase inhibitors based on rhodanine and thiazolidinedione scaffolds

    DEFF Research Database (Denmark)

    Heng, Sabrina; Tieu, William; Hautmann, Stephanie

    2011-01-01

    We present a new class of inhibitors of pancreatic cholesterol esterase (CEase) based on 'priviledged' 5-benzylidenerhodanine and 5-benzylidene-2,4-thiazolidinedione structural scaffolds. The lead structures (5-benzylidenerhodanine 4a and 5-benzylidene-2,4-thiazolidinedione 4b) were identified...

  15. Usefulness of C1 esterase inhibitor protein concentrate in the ...

    African Journals Online (AJOL)

    Hereditary angioedema is an autosomal‑dominant disorder caused by mutation of the gene encoding the C1 esterase inhibitor (C1‑INH). It manifests as painless, nonpruritic, nonpitting episodic swelling of the subcutaneous tissues, gastrointestinal, and upper respiratory tracts in the absence of urticaria. An attack typically ...

  16. Esterase, total protein and seed storage protein diversity in Okra ...

    African Journals Online (AJOL)

    Twenty-two accessions of okra (Abelmoschus esculentus), maintained at the Plant Genetic Resources Centre, Bunso, Ghana, were assayed for diversity in esterases, and total and storage proteins. A total of 34 reproducible and easily scorable bands were exposed with the number of bands per accession ranging from one ...

  17. Hydrolytic and synthetic activities of esterases produced by Bacillus ...

    African Journals Online (AJOL)

    A novel esterase producer strain named Bacillus sp. A60 was isolated from a soil sample contaminated with hydrocarbons. It was found to belong to Bacillus subtilis species through morphological, biochemical and 16S rRNA gene sequence analyses. This strain which can tolerate 15% (w/v) NaCl and growth at 55°C, ...

  18. Protein engineering of esterases : climbing the protein fitness landscape

    NARCIS (Netherlands)

    Godinho, Luis Filipe da Silva

    2011-01-01

    Luis da Silva Godinho beschrijft in zijn proefschrift hoe esterases afkomstig vanBacillus subtilisenEscherichia coligebruikt kunnen worden voor de synthese van een zuiver chiraal synthon; hetgeen van groot belang is voor de farmaceutische industrie. De ontwikkeling van een enzymatisch proces voor de

  19. Third International Meeting on Esterases Reacting with Organophosphorus Compounds

    Science.gov (United States)

    1998-01-01

    ESTHER database (for esterases, oc/ß hydrolase enzymes and relatives) since 1994. This internet server is dedicated to several aspects of biology...Neurociencias University of Zagreb Facultad de Medicina Strossmayerov trg 14 Campus Universitario de San Juan 10000 Zagreb, Croatia 03050 Alicante, Spain

  20. Esterase polymorphism marking cultivars of Manihot esculenta, Crantz

    Directory of Open Access Journals (Sweden)

    Adriana Gazoli Resende

    2004-07-01

    Full Text Available Esterase isozymes were used to detected substrate-preference polymorphism in twenty cultivars of Manihot esculenta, and to show cultivar-specific variation of this species. A relatively complex extraction solution of proteins from leaves was needed to show a larger number of esterase isozymes. Similarity between cultivars from six groups ranged from 51 to 96%. The cultivars identified by the same name seemed to be biochemically different regarding esterase isozymes. Esterase isozyme electrophoretic patterns could, therefore, be used to discriminate the cultivars identified by the same name, and to monitor the vegetative propagation of cultivars maintained in the germplasm collection. In breeding strategies, isoesterase analysis could be used to avoid intercrossing between the similar genotypes.Isoenzimas esterases foram usadas no presente estudo, para detectar polimorfismos específicos para diferentes substratos em vinte cultivares de Manihot esculenta, e para mostrar variações específicas de cultivares nesta espécie. Os diferentes cultivares de M. esculenta tem sido mantidos na coleção de germoplasma do Departamento de Agronomia da Universidade Estadual de Maringá (Maringá, PR, e foram provenientes de cultivares tradicionais coletados nas regiões sudoeste e noroeste do Estado. Foi necessário a utilização de uma solução de extração de proteínas relativamente mais complexa, para evidenciar um maior número de isoenzimas esterases. A similaridade entre os cultivares variou de 51 a 96%. Cultivares identificados pelo mesmo nome parecem ser bioquimicamente diferentes para as isoenzimas esterases. Os padrões eletroforéticos das isoesterases podem, portanto, serem usados para discriminar os cultivares que são identificados pelo mesmo nome, e para monitorar a propagação vegetativa dos cultivares mantidos na coleção de germoplasma. A análise das isoesterases pode também ser usada para evitar cruzamentos entre genótipos mais

  1. Acetyl-Phosphate Is a Critical Determinant of Lysine Acetylation in E. coli

    DEFF Research Database (Denmark)

    Weinert, Brian T; Iesmantavicius, Vytautas; Wagner, Sebastian A

    2013-01-01

    Lysine acetylation is a frequently occurring posttranslational modification in bacteria; however, little is known about its origin and regulation. Using the model bacterium Escherichia coli (E. coli), we found that most acetylation occurred at a low level and accumulated in growth-arrested cells...... acetylate lysine residues in vitro and that AcP levels are correlated with acetylation levels in vivo, suggesting that AcP may acetylate proteins nonenzymatically in cells. These results uncover a critical role for AcP in bacterial acetylation and indicate that most acetylation in E. coli occurs at a low...

  2. Analysis of acetylation stoichiometry suggests that SIRT3 repairs nonenzymatic acetylation lesions

    DEFF Research Database (Denmark)

    Weinert, Brian T; Moustafa, Tarek; Iesmantavicius, Vytautas

    2015-01-01

    or suppresses acetylation. Using quantitative mass spectrometry, we measured acetylation stoichiometry in mouse liver tissue and found that SIRT3 suppressed acetylation to a very low stoichiometry at its target sites. By examining acetylation changes in the liver, heart, brain, and brown adipose tissue...... of fasted mice, we found that SIRT3-targeted sites were mostly unaffected by fasting, a dietary manipulation that is thought to regulate metabolism through SIRT3-dependent deacetylation. Globally increased mitochondrial acetylation in fasted liver tissue, higher stoichiometry at mitochondrial acetylation...... sites, and greater sensitivity of SIRT3-targeted sites to chemical acetylation in vitro and fasting-induced acetylation in vivo, suggest a nonenzymatic mechanism of acetylation. Our data indicate that most mitochondrial acetylation occurs as a low-level nonenzymatic protein lesion and that SIRT3...

  3. A Highly Efficient Xylan-Utilization System in Aspergillus niger An76: A Functional-Proteomics Study

    Directory of Open Access Journals (Sweden)

    Weili Gong

    2018-03-01

    Full Text Available Xylan constituted with β-1,4-D-xylose linked backbone and diverse substituted side-chains is the most abundant hemicellulose component of biomass, which can be completely and rapidly degraded into fermentable sugars by Aspergillus niger. This is of great value for obtaining renewable biofuels and biochemicals. To clarify the underlying mechanisms associated with highly efficient xylan degradation, assimilation, and metabolism by A. niger, we utilized functional proteomics to analyze the secreted proteins, sugar transporters, and intracellular proteins of A. niger An76 grown on xylan-based substrates. Results demonstrated that the complete xylanolytic enzyme system required for xylan degradation and composed of diverse isozymes was secreted in a sequential order. Xylan-backbone-degrading enzymes were preferentially induced by xylose or other soluble sugars, which efficiently produced large amounts of xylooligosaccharides (XOS and xylose; however, XOS was more efficient than xylose in triggering the expression of the key transcription activator XlnR, resulting in higher xylanase activity and shortening xylanase-production time. Moreover, the substituted XOS was responsible for improving the abundance of side-chain-degrading enzymes, specific transporters, and key reductases and dehydrogenases in the pentose catabolic pathway. Our findings indicated that industries might be able to improve the species and concentrations of xylan-degrading enzymes and shorten fermentation time by adding abundant intermediate products of natural xylan (XOS to cultures of filamentous fungi.

  4. A Highly Efficient Xylan-Utilization System in Aspergillus niger An76: A Functional-Proteomics Study.

    Science.gov (United States)

    Gong, Weili; Dai, Lin; Zhang, Huaiqiang; Zhang, Lili; Wang, Lushan

    2018-01-01

    Xylan constituted with β-1,4-D-xylose linked backbone and diverse substituted side-chains is the most abundant hemicellulose component of biomass, which can be completely and rapidly degraded into fermentable sugars by Aspergillus niger . This is of great value for obtaining renewable biofuels and biochemicals. To clarify the underlying mechanisms associated with highly efficient xylan degradation, assimilation, and metabolism by A. niger , we utilized functional proteomics to analyze the secreted proteins, sugar transporters, and intracellular proteins of A. niger An76 grown on xylan-based substrates. Results demonstrated that the complete xylanolytic enzyme system required for xylan degradation and composed of diverse isozymes was secreted in a sequential order. Xylan-backbone-degrading enzymes were preferentially induced by xylose or other soluble sugars, which efficiently produced large amounts of xylooligosaccharides (XOS) and xylose; however, XOS was more efficient than xylose in triggering the expression of the key transcription activator XlnR, resulting in higher xylanase activity and shortening xylanase-production time. Moreover, the substituted XOS was responsible for improving the abundance of side-chain-degrading enzymes, specific transporters, and key reductases and dehydrogenases in the pentose catabolic pathway. Our findings indicated that industries might be able to improve the species and concentrations of xylan-degrading enzymes and shorten fermentation time by adding abundant intermediate products of natural xylan (XOS) to cultures of filamentous fungi.

  5. In vitro comparison of rat and chicken brain neurotoxic esterase

    International Nuclear Information System (INIS)

    Novak, R.; Padilla, S.

    1986-01-01

    A systematic comparison was undertaken to characterize neurotoxic esterase (NTE) from rat and chicken brain in terms of inhibitor sensitivities, pH optima, and molecular weights. Paraoxon titration of phenyl valerate (PV)-hydrolyzing carboxylesterases showed that rat esterases were more sensitive than chicken to paraoxon inhibition at concentrations less than or equal to microM and superimposable with chicken esterases at concentrations of 2.5-1000 microM. Mipafox titration of the paraoxon-resistant esterases at a fixed paraoxon concentration of 100 microM (mipafox concentration: 0-1000 microM) resulted in a mipafox I50 of 7.3 microM for chicken brain NTE and 11.6 microM for rat brain NTE. NTE (i.e., paraoxon-resistant, mipafox-sensitive esterase activity) comprised 80% of chicken and 60% of rat brain paraoxon-resistant activity with the specific activity of chicken brain NTE approximately twice that of rat brain NTE. The pH maxima for NTE from both species was similar showing broad, slightly alkaline optima from pH 7.9 to 8.6. [ 3 H]Diisopropyl phosphorofluoridate (DFP)-labeled NTE from the brains of both species had an apparent mol wt of 160,000 measured by sodium dodecyl sulfate polyacrylamide gel electrophoresis. In conclusion, NTE from both species was very similar, with the mipafox I50 for rat NTE within the range of reported values for chicken and human NTE, and the inhibitor parameters of the chicken NTE assay were applicable for the rat NTE assay

  6. Solid-state 13C NMR spectroscopy studies of xylans in the cell wall of Palmaria palmata (L. Kuntze, Rhodophyta).

    Science.gov (United States)

    Lahaye, Marc; Rondeau-Mouro, Corinne; Deniaud, Estelle; Buléon, Alain

    2003-07-22

    The chemical structure and interactions of the cell wall polysaccharides from the red edible seaweed Palmaria palmata were studied by liquid-like magic-angle-spinning (MAS) and cross-polarization MAS (CPMAS) solid-state 13C NMR spectroscopy. The liquid-like MAS and CPMAS 13C NMR spectra of the rehydrated algal powder revealed the presence of beta-(1-->4)/beta-(1-->3)-linked D-xylan with chemical shifts close to those observed in the solution 13C NMR spectrum of the polysaccharide. Observation of mix-linked xylan in the liquid-like MAS 13C NMR spectrum indicated that part of this cell wall polysaccharide is loosely held in the alga. The CPMAS NMR spectrum of the dry algal powder alcohol insoluble residue (AIR) showed broad peaks most of which corresponded to the mix-linked xylan. Hydration of AIR induced a marked increase in the signal resolution also in the CPMAS NMR spectra together with a shift of the C-3 and C-4 signals of the (1-->3)- and (1-->4)-linked xylose, respectively. Such modifications were present in the spectrum of hydrated (1-->3)-linked xylan from the green seaweed Caulerpa taxifolia and absent in that of (1-->4)-linked xylan from P. palmata. This result emphasizes the important role of (1-->3) linkages on the mix-linked xylan hydration-induced conformational rearrangement. The mix-linked xylan signals were observed in the CPMAS NMR spectrum of hydrated residues obtained after extensive extractions by NaOH or strong chaotropic solutions indicating strong hydrogen bonds or covalent linkages. T(1 rho) relaxations were measured close or above 10 ms for the mix-linked xylan in the dry and hydrated state in AIR and indicated that the overall xylan chains likely remain rigid. Rehydration of the mix-linked xylan lead to a decrease in the motion of protons bounded to the C-1 and C-4 carbons of the (1-->4)-linked xylose supporting the re-organization of the xylan chains under hydration involving junction-zones held by hydrogen bonds between adjacent (1

  7. Mode of action of Bacillus licheniformis pectin methylesterase on highly methylesterified and acetylated pectins.

    Science.gov (United States)

    Remoroza, Connie; Wagenknecht, Martin; Buchholt, Hans Christian; Moerschbacher, Bruno M; Gruppen, Harry; Schols, Henk A

    2015-01-22

    A gene encoding a putative pectinesterase from Bacillus licheniformis DSM13 was cloned and expressed in Escherichia coli. The resulting recombinant enzyme (BliPME) was purified and characterized as a pectin methylesterase. The enzyme showed maximum activity at pH 8.0 and 50°C. BliPME is able to release up to 100% of the methylesters from lime pectin (DM 34-76→DM 0) and up to 73% of all methylesters from SBPs (DM 30-73→DM 14). BliPME efficiently de-methylesterifies lemon pectins and SBPs in a blockwise manner and is quite tolerant towards the acetyl groups present within the SBPs. Detailed analysis of the BliPME-modified pectins using HILIC-MSn and the classical calcium reactivity measurement showed that the enzyme generates pectins with low methylesterification (lime and SBP) and high acetyl content (SBP) while creating blocks of nonmethylesterified galacturonic acid residues. The high activity of BliPME towards highly methylesterified and acetylated pectins makes this novel esterase more efficient in removing methylesters from highly esterified beet pectin compared to other PMEs, e.g. Aspergillus niger PME. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Simultaneous catalytic conversion of cellulose and corncob xylan under temperature programming for enhanced sorbitol and xylitol production.

    Science.gov (United States)

    Ribeiro, Lucília Sousa; Órfão, José J de Melo; Pereira, Manuel Fernando Ribeiro

    2017-11-01

    Sorbitol and xylitol yields can be improved by converting cellulose and xylan simultaneously, due to a synergetic effect between both substrates. Furthermore, both yields can be greatly enhanced by simply adjusting the reaction conditions regarding the optimum for the production of each product, since xylitol (from xylan) and sorbitol (from cellulose) yields are maximized when the reaction is carried out at 170 and 205°C, respectively. Therefore, the combination of a simultaneous conversion of cellulose and xylan with a two-step temperature approach, which consists in the variation of the reaction temperature from 170 to 205°C after 2h, showed to be a good strategy for maximizing the production of sorbitol and xylitol directly from mixture of cellulose and xylan. Using this new and environmentally friendly approach, yields of sorbitol and xylitol of 75 and 77%, respectively, were obtained after 6h of reaction. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Overexpression of esterase D in kidney from trisomy 13 fetuses

    Energy Technology Data Exchange (ETDEWEB)

    Loughna, S.; Moore, G. (Institute of Obstetrics and Gynaecology, London (United Kingdom)); Gau, G.; Blunt, S. (Cytogenetics Lab., London (United Kingdom)); Nicolaides, K. (King' s College School of Medicine and Dentistry, London (United Kingdom))

    1993-10-01

    Human trisomy 13 (Patau syndrome) occurs in approximately 1 in 5,000 live births. It is compatible with life, but prolonged survival is rare. Anomalies often involve the urogenital, cardiac, craniofacial, and central nervous systems. It is possible that these abnormalities may be due to the overexpression of developmentally important genes on chromosome 13. The expression of esterase D (localized to chromosome 13q14.11) has been investigated in both muscle and kidney from trisomy 13 fetuses and has been compared with normal age- and sex-matched fetal tissues, by using northern analysis. More than a twofold increase in expression of esterase D was found in the kidney of two trisomy 13 fetuses, with normal levels in a third. Overexpression was not seen in the muscle tissues from these fetuses. 34 refs., 3 figs., 2 tabs.

  10. Leucocyte esterase in the rapid diagnosis of paediatric septic arthritis.

    LENUS (Irish Health Repository)

    Kelly, E G

    2013-02-01

    Septic arthritis may affect any age group but is more common in the paediatric population. Infection is generally bacterial in nature. Prompt diagnosis is crucial, as delayed treatment is associated with lifelong joint dysfunction. A clinical history and application of Kocher\\'s criteria may indicate that there is a septic arthritis. However, definitive diagnosis is made on culture of septic synovial fluid. The culture process can take over 24h for the initial culture to yield bacterial colonies. Leucocyte esterase is released by leucocytes at the site of an infection. We hypothesise that leucocyte esterase can be utilized in the rapid diagnosis of septic arthritis and shorten the time to decisive treatment whilst simultaneously decreasing unnecessary treatment of non-septic joints.

  11. Overexpression of esterase D in kidney from trisomy 13 fetuses.

    Science.gov (United States)

    Loughna, S; Bennett, P; Gau, G; Nicolaides, K; Blunt, S; Moore, G

    1993-01-01

    Human trisomy 13 (Patau syndrome) occurs in approximately 1 in 5,000 live births. It is compatible with life, but prolonged survival is rare. Anomalies often involve the urogenital, cardiac, craniofacial, and central nervous systems. It is possible that these abnormalities may be due to the overexpression of developmentally important genes on chromosome 13. The expression of esterase D (localized to chromosome 13q14.11) has been investigated in both muscle and kidney from trisomy 13 fetuses and has been compared with normal age- and sex-matched fetal tissues, by using northern analysis. More than a twofold increase in expression of esterase D was found in the kidney of two trisomy 13 fetuses, with normal levels in a third. Overexpression was not seen in the muscle tissues from these fetuses. Images Figure 1 Figure 2 Figure 3 PMID:8213811

  12. Down-regulation of glycosyltransferase 8D genes in Populus trichocarpa caused reduced mechanical strength and xylan content in wood.

    Science.gov (United States)

    Li, Quanzi; Min, Douyong; Wang, Jack Peng-Yu; Peszlen, Ilona; Horvath, Laszlo; Horvath, Balazs; Nishimura, Yufuko; Jameel, Hasan; Chang, Hou-Min; Chiang, Vincent L

    2011-02-01

    Members of glycosyltransferase protein families GT8, GT43 and GT47 are implicated in the biosynthesis of xylan in the secondary cell walls of Arabidopsis. The Arabidopsis mutant irx8 has a 60% reduction in xylan. However, over-expression of an ortholog of Arabidopsis IRX8, poplar PoGT8D, in Arabidopsis irx8 mutant could not restore xylan synthesis. The functions of tree GT8D genes remain unclear. We identified two GT8 gene homologs, PtrGT8D1 and PtrGT8D2, in Populus trichocarpa. They are the only two GT8D members and are abundantly and specifically expressed in the differentiating xylem of P. trichocarpa. PtrGT8D1 transcript abundance was >7 times that of PtrGT8D2. To elucidate the genetic function of GT8D in P. trichocarpa, the expression of PtrGT8D1 and PtrGT8D2 was simultaneously knocked down through RNAi. Four transgenic lines had 85-94% reduction in transcripts of PtrGT8D1 and PtrGT8D2, resulting in 29-36% reduction in stem wood xylan content. Xylan reduction had essentially no effect on cellulose quantity but caused an 11-25% increase in lignin. These transgenics exhibit a brittle wood phenotype, accompanied by increased vessel diameter and thinner fiber cell walls in stem xylem. Stem modulus of elasticity and modulus of rupture were reduced by 17-29% and 16-23%, respectively, and were positively correlated with xylan content but negatively correlated with lignin quantity. These results suggest that PtrGT8Ds play key roles in xylan biosynthesis in wood. Xylan may be a more important factor than lignin affecting the stiffness and fracture strength of wood.

  13. Catalytic Promiscuity of Ancestral Esterases and Hydroxynitrile Lyases.

    Science.gov (United States)

    Devamani, Titu; Rauwerdink, Alissa M; Lunzer, Mark; Jones, Bryan J; Mooney, Joanna L; Tan, Maxilmilien Alaric O; Zhang, Zhi-Jun; Xu, Jian-He; Dean, Antony M; Kazlauskas, Romas J

    2016-01-27

    Catalytic promiscuity is a useful, but accidental, enzyme property, so finding catalytically promiscuous enzymes in nature is inefficient. Some ancestral enzymes were branch points in the evolution of new enzymes and are hypothesized to have been promiscuous. To test the hypothesis that ancestral enzymes were more promiscuous than their modern descendants, we reconstructed ancestral enzymes at four branch points in the divergence hydroxynitrile lyases (HNL's) from esterases ∼ 100 million years ago. Both enzyme types are α/β-hydrolase-fold enzymes and have the same catalytic triad, but differ in reaction type and mechanism. Esterases catalyze hydrolysis via an acyl enzyme intermediate, while lyases catalyze an elimination without an intermediate. Screening ancestral enzymes and their modern descendants with six esterase substrates and six lyase substrates found higher catalytic promiscuity among the ancestral enzymes (P promiscuous and catalyzed both hydrolysis and lyase reactions with many substrates. A broader screen tested mechanistically related reactions that were not selected for by evolution: decarboxylation, Michael addition, γ-lactam hydrolysis and 1,5-diketone hydrolysis. The ancestral enzymes were more promiscuous than their modern descendants (P = 0.04). Thus, these reconstructed ancestral enzymes are catalytically promiscuous, but HNL1 is especially so.

  14. 3 Benzyl-6-chloropyrone: a suicide inhibitor of cholesterol esterase

    International Nuclear Information System (INIS)

    Saint, C.; Gallo, I.; Kantorow, M.; Bailey, J.M.

    1986-01-01

    Cholesterol, absorbed from the intestine, appears in lymph as the ester. Cholesterol esterase is essential for this process, since depletion of the enzyme blocks and repletion restores, absorption. Selective inhibitors of cholesterol esterase may thus prove useful in reducing cholesterol uptake. A series of potential suicide substrates were synthesized which, following cleavage by the enzyme, would attack the putative nucleophile in the active site. One of these, 3-benzyl-6-chloropyrone (3BCP), inhibited both synthesis and hydrolysis of 14 C-cholesteryl oleate with an I 50 of approximately 150 μM. The inactivation was time-dependent and characteristic of a suicide mechanism. The α pyrone structure (lactone analog) is cleaved by a serine-hydroxyl in the active site. This generates an enoyl chloride which inactivates the imidazole believed to play a part in the catalytic function of the enzyme. Inhibition by 3BCP is selective for cholesterol esterase. The activity of pancreatic lipase as not affected by concentrations up to 1 mM

  15. Synergistic effects of graft polymerization and polymer blending on the flexibility of xylan-based films.

    Science.gov (United States)

    Zhang, Xueqin; Liu, Chuanfu; Zhang, Aiping; Sun, Runcang

    2018-02-01

    To develop functional and sustainable films from xylan-based hemicelluloses, beechwood xylan was firstly modified with p-dioxanone (PDO) through ring-opening graft polymerization (ROGP) and then reinforced by poly(vinyl alcohol) (PVA) to fabricate xylan-graft-poly(p-dioxanone)/PVA (XGP/PVA) ternary composite films. FT-IR spectra proved the existence of intermolecular hydrogen bonding interactions between the hydroxyl groups of XGP and PVA. SEM analysis outlined the good compatibility between the XGP matrix and the PVA filler in blending films. From DSC data, the miscibility between XGP and PVA led to increase in the glass transition temperature (T g ) and the crystallinity (X c ) of XGP. In addition, XRD analysis also revealed the increased X c of XGP in the presence of PVA, which was consistent with the DSC results. TGA/DTG curves indicated that the addition of PVA improved the thermal stability of XGP. Tensile testing showed a dramatic increase in the elongation at break of films with the development of weight percent gain (WPG) of XGP. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Biochemical and structural insights into xylan utilization by the thermophilic bacterium Caldanaerobius polysaccharolyticus.

    Science.gov (United States)

    Han, Yejun; Agarwal, Vinayak; Dodd, Dylan; Kim, Jason; Bae, Brian; Mackie, Roderick I; Nair, Satish K; Cann, Isaac K O

    2012-10-12

    Hemicellulose is the next most abundant plant cell wall component after cellulose. The abundance of hemicellulose such as xylan suggests that their hydrolysis and conversion to biofuels can improve the economics of bioenergy production. In an effort to understand xylan hydrolysis at high temperatures, we sequenced the genome of the thermophilic bacterium Caldanaerobius polysaccharolyticus. Analysis of the partial genome sequence revealed a gene cluster that contained both hydrolytic enzymes and also enzymes key to the pentose-phosphate pathway. The hydrolytic enzymes in the gene cluster were demonstrated to convert products from a large endoxylanase (Xyn10A) predicted to anchor to the surface of the bacterium. We further use structural and calorimetric studies to demonstrate that the end products of Xyn10A hydrolysis of xylan are recognized and bound by XBP1, a putative solute-binding protein, likely for transport into the cell. The XBP1 protein showed preference for xylo-oligosaccharides as follows: xylotriose > xylobiose > xylotetraose. To elucidate the structural basis for the oligosaccharide preference, we solved the co-crystal structure of XBP1 complexed with xylotriose to a 1.8-Å resolution. Analysis of the biochemical data in the context of the co-crystal structure reveals the molecular underpinnings of oligosaccharide length specificity.

  17. Modification of pine pulp during oxygen delignification by xylan self-assembly.

    Science.gov (United States)

    Grigoray, Olga; Järnström, Joakim; Heikkilä, Elina; Fardim, Pedro; Heinze, Thomas

    2014-11-04

    Self-assembly is a technique of preparing functional materials based on targeted intermolecular interactions involving different macromolecules. In this work, hardwood xylan was disassembled from wood and birch bleached kraft pulp using pressurized hot water extraction (HWX) and cold alkali extraction (CAX), respectively. The extracted biopolymers were characterized using gas chromatography (GC), size exclusion chromatography (SEC) and Fourier transform infrared spectroscopy (FTIR), and subsequently added into an oxygen delignification reactor containing pine kraft pulp. The assembly of xylan-pulp fiber was characterized using advanced time-of-flight secondary ion mass spectrometry (ToF-SIMS) and imaging. The xylan-pine pulp assembly was not significantly removed during the whole elemental chlorine free bleaching sequence or during low consistency refining. Modified fibers had superior mechanical properties compared to the reference pulp. Our concept can be easily applied in the pulp and paper industry, and it opens new possibilities for the utilization of fully bio-based fibers in new materials. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Characterisation of a New Family of Carboxyl Esterases with an OsmC Domain.

    Directory of Open Access Journals (Sweden)

    Mai-Britt V Jensen

    Full Text Available Proteins in the serine esterase family are widely distributed in bacterial phyla and display activity against a range of biologically produced and chemically synthesized esters. A serine esterase from the psychrophilic bacterium Pseudoalteromonas arctica with a C-terminal OsmC-like domain was recently characterized; here we report on the identification and characterization of further putative esterases with OsmC-like domains constituting a new esterase family that is found in a variety of bacterial species from different environmental niches. All of these proteins contained the Ser-Asp-His motif common to serine esterases and a highly conserved pentapeptide nucleophilic elbow motif. We produced these proteins heterologously in Escherichia coli and demonstrated their activity against a range of esterase substrates. Two of the esterases characterized have activity of over two orders of magnitude higher than other members of the family, and are active over a wide temperature range. We determined the crystal structure of the esterase domain of the protein from Rhodothermus marinus and show that it conforms to the classical α/β hydrolase fold with an extended 'lid' region, which occludes the active site of the protein in the crystal. The expansion of characterized members of the esterase family and demonstration of activity over a wide-range of temperatures could be of use in biotechnological applications such as the pharmaceutical, detergent, bioremediation and dairy industries.

  19. A comparison between activities for non-specific esterases and esterproteases

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D

    1988-01-01

    Electrophoretic separation of non-specific esterases and esterproteases from kidney, lung, and liver have been carried out in polyacrylamide gels. By use of zone electrophoresis, isoelectric focusing, and 2-dimensional electrophoresis it was found that most of the esterprotease bands had the same...... localization in the gels as non-specific esterase bands. A number of esterase bands showed no activity towards the esterprotease substrates and a single kidney band possessed esterprotease activity only. Isozymes of the ES-6 and ES-9 zones showed sex dependent esterprotease reactions. In sections esterase...

  20. Hemicellulases from anaerobic thermophiles. Progress report

    Energy Technology Data Exchange (ETDEWEB)

    Wiegel, J.

    1994-05-01

    The longterm goal of this research effort is to obtain an anaerobic thermophilic bacterium that efficiently converts various hemicellulose-containing biomass to ethanol over a broad pH range. The strategy is to modify the outfit and regulation of the rate-limiting xylanases, glycosidases and xylan esterases in the ethanologenic, anaerobic thermophile Thermoanaerobacter ethanolicus, which grows between pH 4.5 and 9.5. Although it utilizes xylans, the xylanase, acetyl(xylan) esterase and O-methylglucuronidase activities in T. ethanolicus are barely measurable and regarded as the rate limiting steps in its xylan utilization. Thus, and also due to the presently limited knowledge of hemicellulases in anaerobic thermophiles, we characterize the hemicellulolytic enzymes from this and other anaerobic thermophiles as enzyme donors. Beside the active xylosidase/arabinosidase from T. ethanolicus, exhibiting the two different activities, we characterized 2 xylosidases, two acetyl(xylan) esterases, and an O-methylglucuronidase from Thermoanaerobacterium spec. We will continue with the characterization of xylanases from novel isolated slightly acidophilic, neutrophilic and slightly alkalophilic thermophiles. We have cloned, subcloned and partially sequenced the 165,000 Da (2 x 85,000) xylosidase/arabinosidase from T. ethanolicus and started with the cloning of the esterases from Thermoanaerobacterium spec. Consequently, we will develop a shuttle vector and continue to apply electroporation of autoplasts as a method for cloning into T. ethanolicus.

  1. Engineering of plants with improved properties as biofuels feedstocks by vessel-specific complementation of xylan biosynthesis mutants

    Directory of Open Access Journals (Sweden)

    Petersen Pia Damm

    2012-11-01

    Full Text Available Abstract Background Cost-efficient generation of second-generation biofuels requires plant biomass that can easily be degraded into sugars and further fermented into fuels. However, lignocellulosic biomass is inherently recalcitrant toward deconstruction technologies due to the abundant lignin and cross-linked hemicelluloses. Furthermore, lignocellulosic biomass has a high content of pentoses, which are more difficult to ferment into fuels than hexoses. Engineered plants with decreased amounts of xylan in their secondary walls have the potential to render plant biomass a more desirable feedstock for biofuel production. Results Xylan is the major non-cellulosic polysaccharide in secondary cell walls, and the xylan deficient irregular xylem (irx mutants irx7, irx8 and irx9 exhibit severe dwarf growth phenotypes. The main reason for the growth phenotype appears to be xylem vessel collapse and the resulting impaired transport of water and nutrients. We developed a xylan-engineering approach to reintroduce xylan biosynthesis specifically into the xylem vessels in the Arabidopsis irx7, irx8 and irx9 mutant backgrounds by driving the expression of the respective glycosyltransferases with the vessel-specific promoters of the VND6 and VND7 transcription factor genes. The growth phenotype, stem breaking strength, and irx morphology was recovered to varying degrees. Some of the plants even exhibited increased stem strength compared to the wild type. We obtained Arabidopsis plants with up to 23% reduction in xylose levels and 18% reduction in lignin content compared to wild-type plants, while exhibiting wild-type growth patterns and morphology, as well as normal xylem vessels. These plants showed a 42% increase in saccharification yield after hot water pretreatment. The VND7 promoter yielded a more complete complementation of the irx phenotype than the VND6 promoter. Conclusions Spatial and temporal deposition of xylan in the secondary cell wall of

  2. Towards the industrialization of new biosurfactants: Biotechnological opportunities for the lactone esterase gene from Starmerella bombicola.

    Science.gov (United States)

    Roelants, Sophie L K W; Ciesielska, Katarzyna; De Maeseneire, Sofie L; Moens, Helena; Everaert, Bernd; Verweire, Stijn; Denon, Quenten; Vanlerberghe, Brecht; Van Bogaert, Inge N A; Van der Meeren, Paul; Devreese, Bart; Soetaert, Wim

    2016-03-01

    Although sophorolipids (SLs) produced by S. bombicola are a real showcase for the industrialization of microbial biosurfactants, some important drawbacks are associated with this efficient biological process, e.g., the simultaneous production of acidic and lactonic SLs. Depending on the application, there is a requirement for the naturally produced mixture to be manipulated to give defined ratios of the components. Recently, the enzyme responsible for the lactonization of SLs was discovered. The discovery of the gene encoding this lactone esterase (sble) enabled the development of promising S. bombicola strains producing either solely lactonic (using a sble overexpression strain described in this paper: oe sble) or solely acidic SLs (using a sble deletion strain, which was recently described, but not characterized yet: Δsble). The new S. bombicola strains were used to investigate the production processes (fermentation and purification) of either lactonic or acidic SLs. The strains maintain the high inherent productivities of the wild-type or even perform slightly better and thus represent a realistic industrial opportunity. 100% acidic SLs with a mixed acetylation pattern were obtained for the Δsble strain, while the inherent capacity to selectively produce lactonic SLs was significantly increased (+42%) for the oe sble strain (99% lactonic SLs). Moreover, the regulatory effect of citrate on lactone SL formation for the wild-type was absent in this new strain, which indicates that it is more robust and better suited for the industrial production of lactonic SLs. Basic parameters were determined for the purified SLs, which confirm that the two new strains produce molecules with distinctive properties of which the application potential can now easily be investigated independently. © 2015 Wiley Periodicals, Inc.

  3. Non-enzymatic N-acetylation of Lysine Residues by AcetylCoA Often Occurs via a Proximal S-acetylated Thiol Intermediate Sensitive to Glyoxalase II

    Directory of Open Access Journals (Sweden)

    Andrew M. James

    2017-02-01

    Full Text Available Summary: Acetyl coenzyme A (AcCoA, a key intermediate in mitochondrial metabolism, N-acetylates lysine residues, disrupting and, in some cases, regulating protein function. The mitochondrial lysine deacetylase Sirtuin 3 (Sirt3 reverses this modification with benefits reported in diabetes, obesity, and aging. We show that non-enzymatic lysine N-acetylation by AcCoA is greatly enhanced by initial acetylation of a cysteine residue, followed by SN-transfer of the acetyl moiety to a nearby lysine on mitochondrial proteins and synthetic peptides. The frequent occurrence of an S-acetyl intermediate before lysine N-acetylation suggests that proximity to a thioester is a key determinant of lysine susceptibility to acetylation. The thioesterase glyoxalase II (Glo2 can limit protein S-acetylation, thereby preventing subsequent lysine N-acetylation. This suggests that the hitherto obscure role of Glo2 in mitochondria is to act upstream of Sirt3 in minimizing protein N-acetylation, thus limiting protein dysfunction when AcCoA accumulates. : James et al. show that the non-enzymatic N-acetylation of lysine residues in mitochondrial proteins frequently occurs via a proximal S-acetylated thiol intermediate. Glutathione equilibrates with this intermediate, allowing the thioesterase glyoxalase II to limit protein lysine N-acetylation. These findings expand our understanding of how protein acetylation arises. Keywords: AcetylCoA, lysine acetylation, glyoxalase

  4. Analysis of acetylated wood by electron microscopy

    NARCIS (Netherlands)

    Sander, C.; Beckers, E.P.J.; Militz, H.; Veenendaal, van W.

    2003-01-01

    The properties of acetylated solid wood were investigated earlier, in particular the anti-shrink efficiency and the resistance against decay. This study focuses on the possible changes and damage to the wood structure due to an acetylation process leading to weight per cent gains of up to 20%.

  5. The Acetylation of Starch by Reactive Extrusion

    NARCIS (Netherlands)

    Graaf, Robbert A. de; Broekroelofs, Annet; Janssen, Léon P.B.M.

    1998-01-01

    Potato starch has been acetylated in a counter rotating twin screw extruder using vinylacetate and sodium hydroxide. The desired starch acetylation reaction is accompanied by an undesired parallel base catalysed hydrolysis reaction of vinylacetate and a consecutive hydrolysis reaction of the

  6. 21 CFR 172.828 - Acetylated monoglycerides.

    Science.gov (United States)

    2010-04-01

    ... FOR HUMAN CONSUMPTION (CONTINUED) FOOD ADDITIVES PERMITTED FOR DIRECT ADDITION TO FOOD FOR HUMAN CONSUMPTION Multipurpose Additives § 172.828 Acetylated monoglycerides. The food additive acetylated... Reichert-Meissl value of 75-200 and an acid value of less than 6. (c) The food additive is used at a level...

  7. Alicyclobacillus acidocaldarius Thermophilic Esterase EST2's Activity in Milk and Cheese Models

    NARCIS (Netherlands)

    Mandrich, L.; Manco, M.; Rossie, M.; Floris, E.; Jansen-van den Bosch, T.; Smit, G.; Wouters, J.A.

    2006-01-01

    The aim of this work was to investigate the behavior of thermophilic esterase EST2 from Alicyclobacillus acidocaldarius in milk and cheese models. The pure enzyme was used to compare the EST2 hydrolytic activity to the activity of endogenous esterase EstA from Lactococcus lactis. The results

  8. Esterases in striated muscle from mice with the Chediak-Higashi syndrome

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D

    1981-01-01

    In this paper a localized strong reaction for non-specific esterase forming cylindric structures is described within skeletal muscle fibres from the beige mouse. It seems from zymograms and protein electrophoresis that this esterase is membrane bound, highly reactive and present in rather small...... amounts within the muscle fibres....

  9. Acetylation-mediated suppression of transcription-independent memory: bidirectional modulation of memory by acetylation.

    Directory of Open Access Journals (Sweden)

    Katja Merschbaecher

    Full Text Available Learning induced changes in protein acetylation, mediated by histone acetyl transferases (HATs, and the antagonistic histone deacetylases (HDACs play a critical role in memory formation. The status of histone acetylation affects the interaction between the transcription-complex and DNA and thus regulates transcription-dependent processes required for long-term memory (LTM. While the majority of studies report on the role of elevated acetylation in memory facilitation, we address the impact of both, increased and decreased acetylation on formation of appetitive olfactory memory in honeybees. We show that learning-induced changes in the acetylation of histone H3 at aminoacid-positions H3K9 and H3K18 exhibit distinct and different dynamics depending on the training strength. A strong training that induces LTM leads to an immediate increase in acetylation at H3K18 that stays elevated for hours. A weak training, not sufficient to trigger LTM, causes an initial increase in acetylation at H3K18, followed by a strong reduction in acetylation at H3K18 below the control group level. Acetylation at position H3K9 is not affected by associative conditioning, indicating specific learning-induced actions on the acetylation machinery. Elevating acetylation levels by blocking HDACs after conditioning leads to an improved memory. While memory after strong training is enhanced for at least 2 days, the enhancement after weak training is restricted to 1 day. Reducing acetylation levels by blocking HAT activity after strong training leads to a suppression of transcription-dependent LTM. The memory suppression is also observed in case of weak training, which does not require transcription processes. Thus, our findings demonstrate that acetylation-mediated processes act as bidirectional regulators of memory formation that facilitate or suppress memory independent of its transcription-requirement.

  10. Modifying solubility of polymeric xylan extracted from Eucalyptus grandis and sugarcane bagasse by suitable side chain removing enzymes.

    Science.gov (United States)

    Gomes, Katiana R; Chimphango, Annie F A; Görgens, Johann F

    2015-10-20

    α-l-Arabinofuranosidase (AbfB) and novel α-d-glucuronidase (Agu1B) enzymes were applied for selective hydrolysis of beechwood (Fagus sylvatica) xylan (Sigma-Aldrich) as well as xylans extracted from Eucalyptus grandis and sugarcane (Saccharum officinarum L.) bagasse, leading to precipitation of these carbohydrate biopolymers. Hemicellulose extraction was performed with two mild-alkali methods, Höije and Pinto. Precipitation occurred after removal of 67, 40 and 16% 4-O-methyl-d-glucuronic acid (MeGlcA) present in polymeric xylans from beechwood, E. grandis (Pinto) and E. grandis (Höije), respectively. Precipitation was maximized at Agu1B levels of 3.79-7.53mg/gsubstrate and hemicellulose concentrations of 4.5-5.0% (w/v). Polymeric xylan from sugarcane bagasse precipitated after removal of 48 and 22% of arabinose and MeGlcA, respectively, at optimal AbfB and Agu1B dosages of 9.0U/g and 6.4mg/g, respectively. Both the purity of polymeric xylans and structure thereof had a critical impact on the propensity for precipitation, and morphology of the resulting precipitate. Nano-to micro-meter precipitates were produced, with potential for carbohydrate nanotechnology applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Characterization of xylan in the early stages of secondary cell wall formation in tobacco bright yellow-2 cells.

    Science.gov (United States)

    Ishii, Tadashi; Matsuoka, Keita; Ono, Hiroshi; Ohnishi-Kameyama, Mayumi; Yaoi, Katsuro; Nakano, Yoshimi; Ohtani, Misato; Demura, Taku; Iwai, Hiroaki; Satoh, Shinobu

    2017-11-15

    The major polysaccharides present in the primary and secondary walls surrounding plant cells have been well characterized. However, our knowledge of the early stages of secondary wall formation is limited. To address this, cell walls were isolated from differentiating xylem vessel elements of tobacco bright yellow-2 (BY-2) cells induced by VASCULAR-RELATED NAC-DOMAIN7 (VND7). The walls of induced VND7-VP16-GR BY-2 cells consisted of cellulose, pectic polysaccharides, hemicelluloses, and lignin, and contained more xylan and cellulose compared with non-transformed BY-2 and uninduced VND7-VP16-GR BY-2 cells. A reducing end sequence of xylan containing rhamnose and galaturonic acid- residues is present in the walls of induced, uninduced, and non-transformed BY-2 cells. Glucuronic acid residues in xylan from walls of induced cells are O-methylated, while those of xylan in non-transformed BY-2 and uninduced cells are not. Our results show that xylan changes in chemical structure and amounts during the early stages of xylem differentiation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Effect of pH on production of xylanase by Trichoderma reesei on xylan- and cellulose-based media

    Energy Technology Data Exchange (ETDEWEB)

    Bailey, M.J. (VTT, Biotechnical Lab., Espoo (Finland)); Buchert, J. (VTT, Biotechnical Lab., Espoo (Finland)); Viikari, L. (VTT, Biotechnical Lab., Espoo (Finland))

    1993-11-01

    Trichoderma reesei VTT-D-86271 (Rut C-30) was cultivated on media based on cellulose and xylan as the main carbon source in fermentors with different pH minimum controls. Production of xylanase was favoured by a rather high pH minimum control between 6.0 and 7.0 on both cellulose- and xylan-based media. Although xylanase was produced efficiently on cellulose as well as on xylan as the carbon source, significant production of cellulase was observed only on the cellulose-based medium and best production was at lower pH (4.0 minimum). Production of xylanase at pH 7.0 was shown to be dependent on the nature of the xylan in the cultivation medium but was independent of other organic components. Best production of xylanase was observed on insoluble, unsubstituted beech xylan at pH 7.0. Similar results were obtained in laboratory and pilot (200-1) fermentors. Downstream processing of the xylanase-rich, low-cellulase culture filtrate presented no technical problems despite apparent autolysis of the fungus at the high pH. Enzyme produced in the 200-1 pilot fermentor was shown to be suitable for use in enzyme-aided bleaching of kraft pulp. Due to the high xylanase/cellulase ratio of enzyme activities in the culture filtrate, pretreatment for removal of cellulase activity prior to pulp bleaching was unnecessary. (orig.)

  13. Studies on the oxidizing system in Holt's medium for histochemical demonstration of esterase activity

    DEFF Research Database (Denmark)

    Kirkeby, S; Blecher, S R

    1978-01-01

    Esterase activity in guinea-pig thyroid and mouse epididymis epithelial cells has been studied using 5-bromoindoxyl acetate as substrate. The pattern of esterase activity in the thyroid of the guinea-pig is constant, irrespective of whether ferri-ferrocyanide (FFC) or certain copper compounds...... cells contain an esterase activity which is not inhibited by conventional SH blocking agents, nor by high concentrations of FFC. From these results it appears that the mode of action of FFC in Holt's medium is as follows. At low concentrations FFC appears to act primarily as a catalytic agent...... in oxidation of indoxyl to indigoid. At high concentration FFC acts as an inhibitor of guinea-pig thyroid esterase, by oxidation of SH groups in the active centre. The esterase of mouse epididymis cell type EH 1 is not subject to this inhibition by FFC, presumably because it does not contain accessible SH...

  14. A novel esterase from Saccharomyces carlsbergensis, a possible function for the yeast TIP1 gene

    DEFF Research Database (Denmark)

    Horsted, M W; Dey, E S; Holmberg, S

    1998-01-01

    An extracellular esterase was isolated from the brewer's yeast, Saccharomyces carlsbergensis. Inhibition by diisopropyl fluorophosphate shows that the enzyme has a serine active site. By mass spectrometry, the molecular weight of the enzyme was 16.9 kDa. The optimal pH for activity was in the range...... of four to five. Esterase activity was found in beer before pasteurization, and a low level of activity was still present after pasteurization. Caprylic acid, which is present in beer, competitively inhibited the esterase. The substrate preference towards esters of p-nitrophenol indicated that the enzyme...... prefers esters of fatty acids from four to 16 carbon atoms. The esterase has lipolytical activity; olive oil (C-18:1), which is a classical substrate for lipase, was hydrolysed. N-terminal sequence analysis of the esterase yielded a sequence which was identical to the deduced amino acid sequence of the S...

  15. A New Functional Classification of Glucuronoyl Esterases by Peptide Pattern Recognition

    DEFF Research Database (Denmark)

    Wittrup Agger, Jane; Busk, Peter Kamp; Pilgaard, Bo

    2017-01-01

    Glucuronoyl esterases are a novel type of enzymes believed to catalyze the hydrolysis of ester linkages between lignin and glucuronoxylan in lignocellulosic biomass, linkages known as lignin carbohydrate complexes. These complexes contribute to the recalcitrance of lignocellulose. Glucuronoyl...... esterases are a part of the microbial machinery for lignocellulose degradation and coupling their role to the occurrence of lignin carbohydrate complexes in biomass is a desired research goal. Glucuronoyl esterases have been assigned to CAZymes family 15 of carbohydrate esterases, but only few examples...... of characterized enzymes exist and the exact activity is still uncertain. Here peptide pattern recognition is used as a bioinformatic tool to identify and group new CE15 proteins that are likely to have glucuronoyl esterase activity. 1024 CE15-like sequences were drawn from GenBank and grouped into 24 groups...

  16. Profiling and functional classification of esterases in olive (Olea europaea) pollen during germination.

    Science.gov (United States)

    Rejón, Juan D; Zienkiewicz, Agnieszka; Rodríguez-García, María Isabel; Castro, Antonio J

    2012-10-01

    A pollen grain contains a number of esterases, many of which are released upon contact with the stigma surface. However, the identity and function of most of these esterases remain unknown. In this work, esterases from olive pollen during its germination were identifided and functionally characterized. The esterolytic capacity of olive (Olea europaea) pollen was examined using in vitro and in-gel enzymatic assays with different enzyme substrates. The functional analysis of pollen esterases was achieved by inhibition assays by using specific inhibitors. The cellular localization of esterase activities was performed using histochemical methods. Olive pollen showed high levels of non-specific esterase activity, which remained steady after hydration and germination. Up to 20 esterolytic bands were identified on polyacrylamide gels. All the inhibitors decreased pollen germinability, but only diisopropyl fluorophosphate (DIFP) hampered pollen tube growth. Non-specific esterase activity is localized on the surface of oil bodies (OBs) and small vesicles, in the pollen intine and in the callose layer of the pollen tube wall. Acetylcholinesterase (AChE) activity was mostly observed in the apertures, exine and pollen coat, and attached to the pollen tube wall surface and to small cytoplasmic vesicles. In this work, for the first time a systematic functional characterization of esterase enzymes in pollen from a plant species with wet stigma has been carried out. Olive pollen esterases belong to four different functional groups: carboxylesterases, acetylesterases, AChEs and lipases. The cellular localization of esterase activity indicates that the intine is a putative storage site for esterolytic enzymes in olive pollen. Based on inhibition assays and cellular localization of enzymatic activities, it can be concluded that these enzymes are likely to be involved in pollen germination, and pollen tube growth and penetration of the stigma.

  17. Acetylation of rice straw for thermoplastic applications.

    Science.gov (United States)

    Zhang, Guangzhi; Huang, Kai; Jiang, Xue; Huang, Dan; Yang, Yiqi

    2013-07-01

    An inexpensive and biodegradable thermoplastic was developed through acetylation of rice straw (RS) with acetic anhydride. Acetylation conditions were optimized. The structure and properties of acetylated RS were characterized by fourier transform infrared (FTIR), solid-state (13)C NMR spectroscopy, X-ray diffractometer (XRD), scanning electron microscope (SEM), differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA). The results showed that acetylation of RS has successfully taken place, and comparing with raw RS, the degree of crystallinity decreased and the decomposition rate was slow. The acetylated RS has got thermoplasticity when weight ratio of RS and acetic anhydride was 1:3, using sulphuric acid (9% to RS) as catalyst in glacial acetic acid 35°C for 12h, and the dosage of solvent was 9 times RS, in which weight percent gain (WPG) of the modified RS powder was 35.5% and its percent acetyl content was 36.1%. The acetylated RS could be formed into transparent thin films with different amount of plasticizer diethyl phthalate (DEP) using tape casting technology. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Pretreatment of wheat straw and conversion of xylose and xylan to ethanol by thermophilic anaerobic bacteria

    DEFF Research Database (Denmark)

    Ahring, Birgitte Kiær; Jensen, K.; Nielsen, P.

    1996-01-01

    Wheat straw was pretreated by wet oxidation (oxygen pressure, alkaline conditions, elevated temperature) or hydrothermal processing (without oxygen) in order to solubilize the hemicellulose, facilitating bio-conversion. The effect of oxygen pressure and sodium carbonate addition on hemicellulose....... Of five different thermophilic bacteria used in this study only two strains produced ethanol with xylan as substrate, one of them being the strain A3 isolated from an Icelandic hot-spring. Probably other degradation products formed in the presence of oxygen might act as inhibitors. Adaptation...

  19. Acetyl Fentanyl Toxicity: Two Case Reports.

    Science.gov (United States)

    Fort, Chelsea; Curtis, Byron; Nichols, Clay; Niblo, Cheryl

    2016-11-01

    Acetyl fentanyl is an illicit fentanyl analog recently appearing in forensic casework. A quantitative method was created for measuring acetyl fentanyl in various biological matrices acquired post-mortem due to recent positive screening results in casework. Initial detection by immunoassay and standard gas chromatography mass spectrometry (GC/MS) methods have been previously reported for acetyl fentanyl and are examined further here. A Selective Ion Monitoring (SIM) method was created using a GC/MS for quantitation. In two separate cases, acetyl fentanyl was found to be in similar concentrations to those previously reported and ruled to be the cause of death. Acetyl fentanyl concentrations were determined in blood samples, liver, brain, vitreous humor, and urine. Individual 1 had acetyl fentanyl concentrations as follows: heart blood-285 ng/mL, femoral blood-192 ng/mL, liver-1,100 ng/g, brain-620 ng/g, and urine-3,420 ng/mL. Individual 2 had acetyl fentanyl concentrations as follows: heart blood-210 ng/mL, femoral blood-255 ng/mL, urine-2,720 ng/mL and vitreous humor-140 ng/mL. Experimental conditions for screening and quantitation are provided, using immunoassay and GC/MS methods. Due to the recent emergence of acetyl fentanyl, more data will need to be generated to fully differentiate recreational and fatal concentrations of acetyl fentanyl to assist toxicologists accurately understanding its physiological impact. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  20. Engineering of plants with improved properties as biofuels feedstocks by vessel-specific complementation of xylan biosynthesis mutants

    DEFF Research Database (Denmark)

    Petersen, Pia; Lau, Jane; Ebert, Berit

    2012-01-01

    Background: Cost-efficient generation of second-generation biofuels requires plant biomass that can easily be degraded into sugars and further fermented into fuels. However, lignocellulosic biomass is inherently recalcitrant toward deconstruction technologies due to the abundant lignin and cross...... in the xylem vessels is sufficient to complement the irx phenotype of xylan deficient mutants, while maintaining low overall amounts of xylan and lignin in the cell wall. This engineering approach has the potential to yield bioenergy crop plants that are more easily deconstructed and fermented into biofuels.......-linked hemicelluloses. Furthermore, lignocellulosic biomass has a high content of pentoses, which are more difficult to ferment into fuels than hexoses. Engineered plants with decreased amounts of xylan in their secondary walls have the potential to render plant biomass a more desirable feedstock for biofuel production...

  1. Green synthesis of silver nanoparticles in xylan solution via Tollens reaction and their detection for Hg2+

    Science.gov (United States)

    Luo, Yuqiong; Shen, Suqin; Luo, Jiwen; Wang, Xiaoying; Sun, Runcang

    2014-12-01

    This work reported a facile and green method to prepare highly stable and uniformly distributed Ag nanoparticles (AgNPs), in which a biopolymer xylan was used as the stabilizing and reducing agent via the Tollens reaction under microwave irradiation. Different variables were evaluated to optimize the reaction conditions. Complete characterization was performed using UV-Vis, XRD, TEM, size distribution analysis and XPS. The results revealed that AgNPs were well dispersed with diameters of 20-35 nm due to the packing of xylan. The optimal conditions were as follows: microwave irradiation temperature was 60-70 °C, microwave power was 800 W, microwave time was 30 min, the ratio of xylan to AgNO3 was 50 mg: 0.13 mmol, and ammonia concentration was 2%. In addition, the AgNPs were collected via high-speed centrifugal separation, and the supernatant was tested by HPAEC, GPC, FT-IR, and NMR. By comparing the structure of xylan before and after the reaction, the reaction mechanism was discussed. It was noted that the xylan-AgNPs composites showed high selectivity and sensitivity for Hg2+ detection. The other 15 metal ions used had no obvious effect on the detection of Hg2+, and the limit of detection (LOD) was 4.6 nM, which is lower than the allowed maximum level of 30 nM for drinking water by WHO. In addition, the xylan-AgNPs composites can be applied for Hg2+ detection in real water samples. This study provides a novel way for the high-value utilization of a rich biomass resource, and a green method for the synthesis of AgNPs for the selective and sensitive detection of harmful heavy metals.This work reported a facile and green method to prepare highly stable and uniformly distributed Ag nanoparticles (AgNPs), in which a biopolymer xylan was used as the stabilizing and reducing agent via the Tollens reaction under microwave irradiation. Different variables were evaluated to optimize the reaction conditions. Complete characterization was performed using UV-Vis, XRD, TEM

  2. Production of xylan degrading endo-1, 4-β-xylanase from thermophilic Geobacillus stearothermophilus KIBGE-IB29

    Directory of Open Access Journals (Sweden)

    Zainab Bibi

    2014-10-01

    Full Text Available Xylan degrading bacterial strain was isolated from soil and identified as Geobacillus stearothermophilus KIBGE-IB29 on the basis of morphological, biochemical and 16S rDNA sequence analysis. Optimization of medium and culture conditions in submerged fermentation was investigated for maximum endo-1, 4-β-xylanase production. High yield of xylan degrading endo-1, 4-β-xylanase was achieved at 60 °C and pH-6.0 with 24 h of fermentation. Maximum enzyme was produced using 0.5% xylan as a carbon source, 0.5% peptone, 0.2% yeast extract and 0.1% meat extract as nitrogen sources. Di-potassium hydrogen phosphate (0.25%, calcium chloride (0.01%, potassium hydrogen phosphate (0.05% and ammonium sulfate (0.05% were also incorporated in the fermentation medium to enhance the enzyme production.

  3. Histone acetylation: molecular mnemonics on the chromatin.

    Science.gov (United States)

    Gräff, Johannes; Tsai, Li-Huei

    2013-02-01

    Long-lasting memories require specific gene expression programmes that are, in part, orchestrated by epigenetic mechanisms. Of the epigenetic modifications identified in cognitive processes, histone acetylation has spurred considerable interest. Whereas increments in histone acetylation have consistently been shown to favour learning and memory, a lack thereof has been causally implicated in cognitive impairments in neurodevelopmental disorders, neurodegeneration and ageing. As histone acetylation and cognitive functions can be pharmacologically restored by histone deacetylase inhibitors, this epigenetic modification might constitute a molecular memory aid on the chromatin and, by extension, a new template for therapeutic interventions against cognitive frailty.

  4. Green synthesis of silver nanoparticles in xylan solution via Tollens reaction and their detection for Hg(2+).

    Science.gov (United States)

    Luo, Yuqiong; Shen, Suqin; Luo, Jiwen; Wang, Xiaoying; Sun, Runcang

    2015-01-14

    This work reported a facile and green method to prepare highly stable and uniformly distributed Ag nanoparticles (AgNPs), in which a biopolymer xylan was used as the stabilizing and reducing agent via the Tollens reaction under microwave irradiation. Different variables were evaluated to optimize the reaction conditions. Complete characterization was performed using UV-Vis, XRD, TEM, size distribution analysis and XPS. The results revealed that AgNPs were well dispersed with diameters of 20-35 nm due to the packing of xylan. The optimal conditions were as follows: microwave irradiation temperature was 60-70 °C, microwave power was 800 W, microwave time was 30 min, the ratio of xylan to AgNO3 was 50 mg: 0.13 mmol, and ammonia concentration was 2%. In addition, the AgNPs were collected via high-speed centrifugal separation, and the supernatant was tested by HPAEC, GPC, FT-IR, and NMR. By comparing the structure of xylan before and after the reaction, the reaction mechanism was discussed. It was noted that the xylan-AgNPs composites showed high selectivity and sensitivity for Hg(2+) detection. The other 15 metal ions used had no obvious effect on the detection of Hg(2+), and the limit of detection (LOD) was 4.6 nM, which is lower than the allowed maximum level of 30 nM for drinking water by WHO. In addition, the xylan-AgNPs composites can be applied for Hg(2+) detection in real water samples. This study provides a novel way for the high-value utilization of a rich biomass resource, and a green method for the synthesis of AgNPs for the selective and sensitive detection of harmful heavy metals.

  5. Production of xylooligosaccharides from enzymatic hydrolysis of xylan by white-rot fungi Pleurotus - DOI: 10.4025/actascitechnol.v32i1.7648

    Directory of Open Access Journals (Sweden)

    Cristiano Ragagnin de Menezes

    2009-12-01

    Full Text Available Hemicellulose consists of non-cellulosic polysaccharides, with xylans and mannans as their main examples. In nature, xylan can be first degraded to xylooligosaccharides and finally to xylose by certain microorganisms. White-rot fungi basidiomycetes Pleurotus sp. BCCB068 and Pleurotus tailandia were used to degrade oat-spelts xylan under submerged fermentation for a period of 40 days. The study obtained activities of endo-1,4-ß-xylanase and ß-xylosidase and determination of xylan products by degradation. The fungi reached significant levels of xylan degradation by Pleurotus sp. BCCB068 (75.1% and P. tailandia (73.4%, following formations of xylooligosaccharides and sugar monomers. These Pleurotus strains proved to be a feasible alternative for biotechnological processes related to degradation of hemicellulose sources.

  6. Functional classification of esterases from leaves of Aspidosperma polyneuron M. Arg. (Apocynaceae

    Directory of Open Access Journals (Sweden)

    Carvalho Vanda Marilza de

    2003-01-01

    Full Text Available Polyacrylamide gel electrophoresis system (PAGE and inhibition tests for biochemical characterization of alpha- and beta-esterases were used to obtain a functional classification of esterases fromAspidosperma polyneuron. The characterization of alpha- and beta-esterases from young leaves of A. polyneuron by the PAGE system showed fourteen esterase isozymes. The differential staining pattern showed that Est-2 isozyme hydrolyzes beta-naphthyl acetate; Est-6, Est-7 and Est-8 isozymes hydrolyze alpha-naphthyl acetate, and Est-1, Est-3, Est-4, Est-5, Est-9, Est-10, Est-11, Est-12, Est-13, and Est-14 isozymes hydrolyze both alpha- and b-naphthyl acetate. Inhibition pattern of a- and beta-esterases showed that Folidol is a more potent inhibitor that Malathion, while Thiamethoxan (an insecticide with organophosphorus-like action acts as an Est-4 and Est-6 inhibitor and induces the appearance of Est-5 and Est-7 isozymes as more intensely stained bands. Inhibition tests showed that OPC insecticides inhibit or activate plant esterases. Thus, plant esterases may be used as bioindicators to detect the presence and toxicity of residues of topically applied insecticides in agriculture and may be valuable for monitoring pollutants in the environment.

  7. Study of new feruloyl esterases to understand lipase evolution: the case of Bacillus flexus.

    Science.gov (United States)

    Sánchez-González, Mónica; Blanco-Gámez, Allan; Parra-Saldívar, Roberto; Mateos-Díaz, Juan Carlos; Estrada-Alvarado, María Isabel

    2012-01-01

    Recently, the crystal structure of the feruloyl esterase A from Aspergillus niger (AnFaeA) was elucidated. This enzyme displays an α/β hydrolase fold and a catalytic triad similar to that found in fungal lipases (30-37% identity). Surprisingly, AnFaeA showed an overall fold similarity with the Rhizomucor miehei and other related fungal lipases. All these data strongly suggest that the ancestral function (lipase) had shifted, with molecular adaptation leading to a novel enzyme (type-A feruloyl esterase). The discovery of new feruloyl esterases could lead to get insight into the evolutionary pathways of these enzymes and into new possibilities of directed evolution of lipases. In this chapter, the production of Bacillus flexus NJY2 feruloyl esterases is described. Unlike the previously described feruloyl esterases, which mostly belong to eukaryotes (mainly fungus), this unique feruloyl esterases from a prokaryotic alkaliphile microorganism could be the starting point for new discoveries on lipase and feruloyl esterase evolutionary relationships.

  8. A Novel Cold Active Esterase from a Deep Sea Sponge Stelletta normani Metagenomic Library

    Directory of Open Access Journals (Sweden)

    Erik Borchert

    2017-09-01

    Full Text Available Esterases catalyze the hydrolysis of ester bonds in fatty acid esters with short-chain acyl groups. Due to the widespread applications of lipolytic enzymes in various industrial applications, there continues to be an interest in novel esterases with unique properties. Marine ecosystems have long been acknowledged as a significant reservoir of microbial biodiversity and in particular of bacterial enzymes with desirable characteristics for industrial use, such as for example cold adaptation and activity in the alkaline pH range. We employed a functional metagenomic approach to exploit the enzymatic potential of one particular marine ecosystem, namely the microbiome of the deep sea sponge Stelletta normani. Screening of a metagenomics library from this sponge resulted in the identification of a number of lipolytic active clones. One of these encoded a highly, cold-active esterase 7N9, and the recombinant esterase was subsequently heterologously expressed in Escherichia coli. The esterase was classified as a type IV lipolytic enzyme, belonging to the GDSAG subfamily of hormone sensitive lipases. Furthermore, the recombinant 7N9 esterase was biochemically characterized and was found to be most active at alkaline pH (8.0 and displays salt tolerance over a wide range of concentrations. In silico docking studies confirmed the enzyme's activity toward short-chain fatty acids while also highlighting the specificity toward certain inhibitors. Furthermore, structural differences to a closely related mesophilic E40 esterase isolated from a marine sediment metagenomics library are discussed.

  9. p53 Acetylation: Regulation and Consequences

    Energy Technology Data Exchange (ETDEWEB)

    Reed, Sara M. [Department of Pharmacology, The University of Iowa Carver College of Medicine, Iowa City, IA 52242 (United States); Medical Scientist Training Program, The University of Iowa Carver College of Medicine, Iowa City, IA 52242 (United States); Quelle, Dawn E., E-mail: dawn-quelle@uiowa.edu [Department of Pharmacology, The University of Iowa Carver College of Medicine, Iowa City, IA 52242 (United States); Medical Scientist Training Program, The University of Iowa Carver College of Medicine, Iowa City, IA 52242 (United States); Department of Pathology, The University of Iowa Carver College of Medicine, Iowa City, IA 52242 (United States)

    2014-12-23

    Post-translational modifications of p53 are critical in modulating its tumor suppressive functions. Ubiquitylation, for example, plays a major role in dictating p53 stability, subcellular localization and transcriptional vs. non-transcriptional activities. Less is known about p53 acetylation. It has been shown to govern p53 transcriptional activity, selection of growth inhibitory vs. apoptotic gene targets, and biological outcomes in response to diverse cellular insults. Yet recent in vivo evidence from mouse models questions the importance of p53 acetylation (at least at certain sites) as well as canonical p53 functions (cell cycle arrest, senescence and apoptosis) to tumor suppression. This review discusses the cumulative findings regarding p53 acetylation, with a focus on the acetyltransferases that modify p53 and the mechanisms regulating their activity. We also evaluate what is known regarding the influence of other post-translational modifications of p53 on its acetylation, and conclude with the current outlook on how p53 acetylation affects tumor suppression. Due to redundancies in p53 control and growing understanding that individual modifications largely fine-tune p53 activity rather than switch it on or off, many questions still remain about the physiological importance of p53 acetylation to its role in preventing cancer.

  10. Studies on esterase isozymes and mycelium growth speed of ganoderma lucidum carried by Shenzhou spaceship

    International Nuclear Information System (INIS)

    Qi Jianjun; Chen Xiangdong; Lan Jin

    2002-01-01

    The esterase isozymes and mycelium growth speed of four Ganoderma lucidum strains carried by Shenzhou spaceship were studied. The results showed that different effects occurred to esterase and mycelium growth speed. The SX, S3 esterase band had changed compared with their control CX, C3, respectively, but there were no differences between SH and CH, S4 and C4. The growth speed of S4 strain was faster than its control C4, SX strain lower than its control CX, and there were no difference between SH and CH, S3 and C3

  11. Heterologous Expression of Two Ferulic Acid Esterases from Penicillium Funiculosum

    Energy Technology Data Exchange (ETDEWEB)

    Knoshaug, E. P.; Selig, M. J.; Baker, J. O.; Decker, S. R.; Himmel, M. E.; Adney, W. S.

    2008-01-01

    Two recombinant ferulic acid esterases from Penicillium funiculosum produced in Aspergillus awamori were evaluated for their ability to improve the digestibility of pretreated corn stover. The genes, faeA and faeB, were cloned from P. funiculosum and expressed in A. awamori using their native signal sequences. Both enzymes contain a catalytic domain connected to a family 1 carbohydrate-binding module by a threonine-rich linker peptide. Interestingly, the carbohydrate binding-module is N-terminal in FaeA and C-terminal in FaeB. The enzymes were purified to homogeneity using column chromatography, and their thermal stability was characterized by differential scanning microcalorimetry. We evaluated both enzymes for their potential to enhance the cellulolytic activity of purified Trichoderma reesei Cel7A on pretreated corn stover.

  12. Heterologous Expression of Two Ferulic Acid Esterases from Penicillium funiculosum

    Science.gov (United States)

    Knoshaug, Eric P.; Selig, Michael J.; Baker, John O.; Decker, Stephen R.; Himmel, Michael E.; Adney, William S.

    Two recombinant ferulic acid esterases from Penicillium funiculosum produced in Aspergillus awamori were evaluated for their ability to improve the digestibility of pretreated corn stover. The genes, faeA and faeB, were cloned from P. funiculosum and expressed in A. awamori using their native signal sequences. Both enzymes contain a catalytic domain connected to a family 1 carbohydrate-binding module by a threonine-rich linker peptide. Interestingly, the carbohydrate binding-module is N-terminal in FaeA and C-terminal in FaeB. The enzymes were purified to homogeneity using column chromatography, and their thermal stability was characterized by differential scanning microcalorimetry. We evaluated both enzymes for their potential to enhance the cellulolytic activity of purified Trichoderma reesei Cel7A on pretreated corn stover.

  13. Paenibacillus tundrae sp. nov. and Paenibacillus xylanexedens sp. nov., Psychrotolerant, Xylan-Degrading, Bacteria from Alaskan Tundra

    Science.gov (United States)

    Psychrotolerant, xylan-degrading, strains of bacteria were isolated from soil beneath moist non-acidic and acidic tundra in northern Alaska. Phylogenetic analysis based on 16S rRNA gene sequences revealed that each strain belonged to the genus Paenibacillus. The highest levels of 16S rRNA gene sim...

  14. Discovery of the combined oxidative cleavage of plant xylan and cellulose by a new fungal polysaccharide monooxygenase

    NARCIS (Netherlands)

    Frommhagen, Matthias; Sforza, Stefano; Westphal, Adrie H.; Visser, Jaap; Hinz, Sandra W.A.; Koetsier, Martijn J.; Berkel, van Willem J.H.; Gruppen, Harry; Kabel, Mirjam A.

    2015-01-01

    Background: Many agricultural and industrial food by-products are rich in cellulose and xylan. Their enzymatic degradation into monosaccharides is seen as a basis for the production of biofuels and bio-based chemicals. Lytic polysaccharide monooxygenases (LPMOs) constitute a group of recently

  15. Deconstruction of lignin linked p-coumarates, ferulates and xylan by NaOH enhances the enzymatic conversion of glucan

    NARCIS (Netherlands)

    Murciano Martínez, Patricia; Punt, Arjen M.; Kabel, Mirjam A.; Gruppen, Harry

    2016-01-01

    Thermo-assisted NaOH pretreatment to deconstruct xylan and lignin in sugar cane bagasse (SCB) is poorly understood. Hence, in this research it is was aimed to study the effect of NaOH pretreatment on the insoluble remaining lignin structures. Hereto, SCB milled fibres were pretreated using

  16. Esterase-D and chromosome patterns in Central Amazon piranha (Serrasalmus rhombeus Linnaeus, 1766 from Lake Catalão

    Directory of Open Access Journals (Sweden)

    Aylton Saturnino Teixeira

    2006-01-01

    Full Text Available This study presents additional genetic data on piranha (Serrasalmus rhombeus Linnaeus, 1766 complex previously diagnosed due to the presence of distinct cytotypes 2n = 58 and 2n = 60. Three esterase-D enzyme loci (Est-D1, Est-D2 and Est-D3 were examined and complemented with chromosomal data from 66 piranha specimens collected from Lake Catalão. For all specimens the Est-D1 and Est-D2 loci were monomorphic. In contrast, the Est-D3 locus was polymorphic with genotypes and alleles being differentially distributed in the previously described cytotypes and served as the basis for detecting a new cytotype (2n = 60 B. In cytotype 2n = 58 the Est-D3 locus was also polymorphic and presented Mendelian allelic segregation with four genotypes (Est-D3(11, Est-D3(12, Est-D3(22 and Est-D3(33 out of six theoretically possible genotypes, presumably encoded by alleles Est-D3¹ (frequency = 0.237, EsT-D3² (0.710 and Est-D3³ (0.053. A Chi-squared (chi2 test for Hardy-Weinberg equilibrium was applied to the Est-D3 locus and revealed a genetic unbalance in cytotype 2n = 58, indicating the probable existence in the surveyed area of different stocks for that karyotypic structure. A silent null allele (Est-D3(0 with a high frequency (0.959 occurred exclusively in the 2n = 60 cytotype. On the other hand, the new cytotype 2n = 60 B described here for the first time was monomorphic for the presumably fixed Est-D3³ allele. The data as a whole should contribute to the better understanding the rhombeus complex taxonomic status definition in the Central Amazon.

  17. Esterase activity able to hydrolyze dietary antioxidant hydroxycinnamates is distributed along the intestine of mammals

    DEFF Research Database (Denmark)

    Andreasen, Mette Findal; Kroon, P A; Williamson, G

    2001-01-01

    Hydroxycinnamic acids are effective antioxidants and are abundant components of plant cell walls, especially in cereal bran. For example, wheat and rye brans are rich sources of the hydroxycinnamates ferulic acid, sinapic acid, and p-coumaric acid. These phenolics are part of human and animal diets...... hydroxycinnamates are distributed throughout the intestinal tract of mammals. In rats, the cinnamoyl esterase activity in the small intestine is derived mainly from the mucosa, whereas in the large intestine the esterase activity was found predominantly in the luminal microflora. Mucosa cell-free extracts obtained...... and wheat brans. Hydrolysis by intestinal esterase(s) is very likely the major route for release of antioxidant hydroxycinnamic acids in vivo. Udgivelsesdato: 2001-Nov...

  18. Physicochemical characterization and tissue distribution of esterases in two salamandridae species (Mertensiella luschani and Salamandra salamandra).

    Science.gov (United States)

    Tzannetatou-Polymeni, R; Haritos, A A

    1989-01-01

    1. Tissue- and species-specificity of the electrophoretic patterns of the multiple molecular forms of esterases were observed in the urodele amphibians Mertensiella luschani luschani, M.l. helverseni and Salamandra salamandra. All esterases--distributed into two electrophoretic mobility areas in gonads, muscles and brain and into four areas in liver, stomach and intestine--were characterized as carboxylesterases. 2. M. l. luschani and S. salamandra liver esterases were electrofocused into nine and eleven major bands with pIs ranging from 4.60 to 5.65 and from 4.40 to 6.20, respectively. 3. Two size groups of esterases were observed in liver extracts of the above three subspecies by Sephadex G-200 gel filtration. The mean values of their apparent molecular weights were 70,000 and 230,000 respectively.

  19. Multiple nucleophilic elbows leading to multiple active sites in a single module esterase from Sorangium cellulosum

    DEFF Research Database (Denmark)

    Udatha, D.B.R.K. Gupta; Madsen, Karina Marie; Panagiotou, Gianni

    2015-01-01

    The catalytic residues in carbohydrate esterase enzyme families constitute a highly conserved triad: serine, histidine and aspartic acid. This catalytic triad is generally located in a very sharp turn of the protein backbone structure, called the nucleophilic elbow and identified by the consensus...... sequence GXSXG. An esterase from Sorangium cellulosum Soce56 that contains five nucleophilic elbows was cloned and expressed in Escherichia coli and the function of each nucleophilic elbowed site was characterized. In order to elucidate the function of each nucleophilic elbow, site directed mutagenesis....... To our knowledge, this is the first report presenting the role of multiple nucleophilic elbows in the catalytic promiscuity of an esterase. Further structural analysis at protein unit level indicates the new evolutionary trajectories in emerging promiscuous esterases....

  20. Production and partial characterisation of feruloyl esterase by Sporotrichum thermophile in solid-state fermentation

    DEFF Research Database (Denmark)

    Topakas, E.; Kalogeris, E.; Kekos, D.

    2003-01-01

    A number of factors affecting production of feruloyl esterase an enzyme that hydrolyse ester linkages of ferulic acid (FA) in plant cell walls, by the thermophylic fungus Sporotrichum thermophile under solid state fermentation (SSF) were investigated. Initial moisture content and type of carbon...... source were consecutively optimised. SSF in a laboratory horizontal bioreactor using the optimised medium allowed the production of 156 mU g(-1) of carbon source, which compared favourably with those reported for the other micro-organisms. Optimal esterase activity was observed at pH 8 and 60 degrees......C. The activity of the esterase was measured on an insoluble feruloylated hemicellulose substrate (de-starched wheat bran (DSWB)). De-esterifcation of wheat straw yielded loss of feruloyl esterase production even though the supplementation of free FA comparable to the alkali-extractable levels of FA found...

  1. Esterase detoxification of acetylcholinesterase inhibitors by human or rat liver in vitro

    Science.gov (United States)

    Organophosphate (OP) and N-methylcarbamate pesticides inhibit acetylcholinesterase (AChE), but differences in metabolism and detoxification can influence potency of these pesticides across and within species. Carboxylesterase (CaE) and A-esterase (paraoxonase, PON) are considered...

  2. Esterase-sensitive sulfur dioxide prodrugs inspired by modified Julia olefination.

    Science.gov (United States)

    Wang, Wenyi; Wang, Binghe

    2017-09-12

    Sulfur dioxide (SO 2 ) is an endogenously produced gaseous molecule, and is emerging as a potential gasotransmitter. Herein, we describe the first series of esterase-sensitive prodrugs inspired by modified Julia olefination as SO 2 donors.

  3. Evaluation of leukocyte esterase and nitrite strip tests to detect spontaneous bacterial peritonitis in cirrhotic patients.

    Science.gov (United States)

    Torun, Serkan; Dolar, Enver; Yilmaz, Yusuf; Keskin, Murat; Kiyici, Murat; Sinirtas, Melda; Sarandol, Emre; Gurel, Selim; Nak, Selim-Giray; Gulten, Macit

    2007-12-07

    To investigate the diagnostic efficacy of leukocyte esterase and nitrite reagent strips for bedside diagnosis of spontaneous bacterial peritonitis (SBP). A total of 63 consecutive patients with cirrhotic ascites (38 male, 25 female) tested between April 2005 and July 2006 were included in the study. Bedside reagent strip testing was performed on ascitic fluid and the results compared to manual cell counting and ascitic fluid culture. SBP was defined as having a polymorphonuclear ascites count of >or= 250/mm(3). Fifteen samples showed SBP. The sensitivity, specificity, positive and negative predictive values of the leukocyte esterase reagent strips were; 93%, 100%, 100%, and 98%, respectively. The sensitivity, specificity, positive and negative predictive value of the nitrite reagent strips were 13%, 93%, 40%, and 77%, respectively. The combination of leukocyte esterase and nitrite reagents strips did not yield statistically significant effects on diagnostic accuracy. Leukocyte esterase reagent strips may provide a rapid, bedside diagnostic test for SBP.

  4. Esterases A5-B5 in organophosphate-resistant Culex pipiens from Italy.

    Science.gov (United States)

    Severini, C; Romi, R; Marinucci, M; Guillemaud, T; Raymond, M

    1997-04-01

    Culex pipiens mosquitos from Lignano city, Udine province, northeast Italy, were found to carry over-produced non-specific esterases A1, A2-B2 and A4-B4 or A5-B5, detected by starch gel electrophoresis, giving multiple resistance to organophosphorus insecticides. In order to differentiate between A4-B4 and A5-B5 esterases, the latter known only from Cyprus whereas the former is widespread in Italy and elsewhere, restriction fragment length polymorphism (RFLP) analysis was performed at the esterase B locus. Both B4 and B5 haplotypes were found. This is the first record of A5-B5 esterase-mediated resistance in continental Europe.

  5. Target size of neurotoxic esterase and acetylcholinesterase as determined by radiation inactivation.

    OpenAIRE

    Carrington, C D; Fluke, D J; Abou-Donia, M B

    1985-01-01

    The target size of neurotoxic esterase (NTE), the putative target site for the initiation of organophosphorus-compound-induced delayed neurotoxicity, and acetylcholinesterase (AChE) from hen brain were examined by determining the rate at which the activities of the esterases were destroyed by ionizing irradiation. Samples of hen brain were prepared by slowly drying a microsomal preparation under vacuum. The dried samples were then irradiated with electrons from a 1 MeV Van de Graaff generator...

  6. Extrusion of xylans extracted from corn cobs into biodegradable polymeric materials.

    Science.gov (United States)

    Bahcegul, Erinc; Akinalan, Busra; Toraman, Hilal E; Erdemir, Duygu; Ozkan, Necati; Bakir, Ufuk

    2013-12-01

    Solvent casting technique, which comprises multiple energy demanding steps including the dissolution of a polymer in a solvent followed by the evaporation of the solvent from the polymer solution, is currently the main technique for the production of xylan based polymeric materials. The present study shows that sufficient water content renders arabinoglucuronoxylan (AGX) polymers extrudable, enabling the production of AGX based polymeric materials in a single step via extrusion, which is economically advantageous to solvent casting process for mass production. AGX polymers with water content of 27% were found to yield extrudates at an extrusion temperature of 90°C. The extruded strips showed very good mechanical properties with an ultimate tensile strength of 76 ± 6 MPa and elongation at break value of 35 ± 8%, which were superior to the mechanical properties of the strips obtained from polylactic acid. Copyright © 2013 Elsevier Ltd. All rights reserved.

  7. Complete genome sequence of Thermus brockianus GE-1 reveals key enzymes of xylan/xylose metabolism.

    Science.gov (United States)

    Schäfers, Christian; Blank, Saskia; Wiebusch, Sigrid; Elleuche, Skander; Antranikian, Garabed

    2017-01-01

    Thermus brockianus strain GE-1 is a thermophilic, Gram-negative, rod-shaped and non-motile bacterium that was isolated from the Geysir geothermal area, Iceland. Like other thermophiles, Thermus species are often used as model organisms to understand the mechanism of action of extremozymes, especially focusing on their heat-activity and thermostability. Genome-specific features of T. brockianus GE-1 and their properties further help to explain processes of the adaption of extremophiles at elevated temperatures. Here we analyze the first whole genome sequence of T. brockianus strain GE-1. Insights of the genome sequence and the methodologies that were applied during de novo assembly and annotation are given in detail. The finished genome shows a phred quality value of QV50. The complete genome size is 2.38 Mb, comprising the chromosome (2,035,182 bp), the megaplasmid pTB1 (342,792 bp) and the smaller plasmid pTB2 (10,299 bp). Gene prediction revealed 2,511 genes in total, including 2,458 protein-encoding genes, 53 RNA and 66 pseudo genes. A unique genomic region on megaplasmid pTB1 was identified encoding key enzymes for xylan depolymerization and xylose metabolism. This is in agreement with the growth experiments in which xylan is utilized as sole source of carbon. Accordingly, we identified sequences encoding the xylanase Xyn10, an endoglucanase, the membrane ABC sugar transporter XylH, the xylose-binding protein XylF, the xylose isomerase XylA catalyzing the first step of xylose metabolism and the xylulokinase XylB, responsible for the second step of xylose metabolism. Our data indicate that an ancestor of T. brockianus obtained the ability to use xylose as alternative carbon source by horizontal gene transfer.

  8. Effectiveness of leucocyte esterase as a diagnostic test for acute septic arthritis.

    Science.gov (United States)

    Gautam, V K; Saini, Rishabh; Sharma, Siddharth

    2017-01-01

    We hypothesized that leucocyte esterase strip test can aid in diagnosing septic arthritis in native synovial fluid because leucocyte esterase concentrations would be elevated at the infection site because of secretion by recruited neutrophils. The cohort included 27 patients (suspected septic arthritis and normal subjects). A standard chemical test strip (graded as negative, trace, +, ++ or +++) was used to detect the presence of leucocyte esterase. Fluid leucocyte count, Gram staining, culture, erythrocyte sedimentation rate and C-reactive protein were also assessed. The leucocyte esterase test with a threshold of ++/+++ had a sensitivity of 79.2% (95% CI [confidence interval], 65.9% to 89.2%), specificity of 80.8% (95% CI, 73.3% to 87.1%), positive predictive value (PPV) of 61.8% (95% CI, 49.2% to 73.3%) and negative predictive value (NPV) of 90.1% (95% CI, 84.3% to 95.4%). The leucocyte esterase strip test yielded a high specificity, PPV, NPV, high sensitivity and high diagnostic accuracy. Leucocyte esterase is an accurate, quick and bedside test for septic arthritis and can be used effectively for diagnosing periprosthetic joint infections along with other battery of tests according to the Musculoskeletal Infection Society criteria.

  9. Esterase in imported fire ants, Solenopsis invicta and S. richteri (Hymenoptera: Formicidae): activity, kinetics and variation.

    Science.gov (United States)

    Chen, J; Rashid, T; Feng, G

    2014-11-19

    Solenopsis invicta and Solenopsis richteri are two closely related invasive ants native to South America. Despite their similarity in biology and behavior, S. invicta is a more successful invasive species. Toxic tolerance has been found to be important to the success of some invasive species. Esterases play a crucial role in toxic tolerance of insects. Hence, we hypothesized that the more invasive S. invicta would have a higher esterase activity than S. richteri. Esterase activities were measured for workers and male and female alates of both ant species using α-naphthyl acetate and β-naphthyl acetate as substrates. Esterase activities in S. invicta were always significantly higher than those in S. richteri supporting our hypothesis. In S. invicta, male alates had the highest esterase activities followed by workers then female alates for both substrates. In S. richetri, for α-naphthyl acetate, male alates had the highest activity followed by female alates then workers, while for β-naphthyl acetate, female alates had the highest activity followed by male alates then workers. For workers, S. richteri showed significantly higher levels of variation about the mean esterase activity than S. invicta. However, S. invicta showed significantly higher levels of variation in both female and male alates.

  10. Esterase inhibition by synergists in the western flower thrips Frankliniella occidentalis.

    Science.gov (United States)

    López-Soler, Neus; Cervera, Amelia; Quinto, Vicente; Abellán, Jaime; Bielza, Pablo; Martínez-Pardo, Rafael; Garcerá, Maria Dolores

    2011-12-01

    Western flower thrips (WFT), Frankliniella occidentalis (Pergande), is among the most important crop pests in the south-eastern region of Spain. Its increasing resistance to insecticides constitutes a serious problem, and understanding the mechanisms involved is therefore of great interest. Use of synergists to inhibit the enzymes involved in insecticide detoxification is widely used to determine their responsibility for insecticide resistance. However, they do not always act as intended or expected, and caution must be exercised when interpreting synergist results. Laboratory-selected strains of WFT were used to analyse the effects of the synergists piperonyl butoxide (PBO), S,S,S-tributyl phosphorotrithioate (DEF) and methiocarb on total esterase activity. Significant differences were found, indicating esterase activity inhibition by DEF, a lower effect for methiocarb and a small inhibition of the activity by PBO. Esterase isoenzyme inhibition by these compounds showed a similar result; this assay revealed an extreme sensitivity of Triplet A (resistance-associated esterases) to DEF. In an in vivo assay carried out with these compounds at different incubation times, only DEF caused posterior in vitro esterase activity inhibition, with a maximum effect 1 h after treatment. In this work, only DEF shows true synergistic inhibition of WFT esterases. Copyright © 2011 Society of Chemical Industry.

  11. Production of Nα-acetyl Tα1-HSA through in vitro acetylation by RimJ.

    Science.gov (United States)

    Chen, Jing; Li, Haibin; Wang, Tao; Sun, Shuyang; Liu, Jia; Chen, Jianhua

    2017-11-10

    Thymosin alpha 1 (Tα1) is an important immunomodulating agent with various clinical applications. The natural form of Tα1 is N α -acetylated, which was supposed to be related to in vivo stability of the hormone. In this study, fusion protein Tα1-HSA was constructed and expressed in Pichia pastoris . RimJ, a N α -acetyltransferase from E.coli , was also overexpressed and purified to homogeneity. In vitro acetylation of Tα1-HSA in the presence of RimJ and acetyl coenzyme A resulted in N α -acetyl Tα1-HSA. The N α -acetylation was determined by LC-MS/MS. Kinetic assay indicated that RimJ had a higher affinity to desacetyl Tα1 than to Tα1-HSA. Bioactivity assay revealed fully retained activity of Tα1 when the hormone was connected to the N-terminus of the fusion protein, while the activity was compromised in our previously constructed HSA-Tα1. With fully retained activity and N-terminal acetylation, N α -acetyl Tα1-HSA was expected to be a more promising pharmaceutical agent than Tα1.

  12. Switching Catalysis from Hydrolysis to Perhydrolysis in Pseudomonas fluorescens Esterase

    Energy Technology Data Exchange (ETDEWEB)

    Yin, D.; Bernhardt, P; Morley, K; Jiang, Y; Cheeseman, J; Purpero, V; Schrag, J; Kazlauskas, R

    2010-01-01

    Many serine hydrolases catalyze perhydrolysis, the reversible formation of peracids from carboxylic acids and hydrogen peroxide. Recently, we showed that a single amino acid substitution in the alcohol binding pocket, L29P, in Pseudomonas fluorescens (SIK WI) aryl esterase (PFE) increased the specificity constant of PFE for peracetic acid formation >100-fold [Bernhardt et al. (2005) Angew. Chem., Int. Ed. 44, 2742]. In this paper, we extend this work to address the three following questions. First, what is the molecular basis of the increase in perhydrolysis activity? We previously proposed that the L29P substitution creates a hydrogen bond between the enzyme and hydrogen peroxide in the transition state. Here we report two X-ray structures of L29P PFE that support this proposal. Both structures show a main chain carbonyl oxygen closer to the active site serine as expected. One structure further shows acetate in the active site in an orientation consistent with reaction by an acyl-enzyme mechanism. We also detected an acyl-enzyme intermediate in the hydrolysis of {var_epsilon}-caprolactone by mass spectrometry. Second, can we further increase perhydrolysis activity? We discovered that the reverse reaction, hydrolysis of peracetic acid to acetic acid and hydrogen peroxide, occurs at nearly the diffusion limited rate. Since the reverse reaction cannot increase further, neither can the forward reaction. Consistent with this prediction, two variants with additional amino acid substitutions showed 2-fold higher k{sub cat}, but K{sub m} also increased so the specificity constant, k{sub cat}/K{sub m}, remained similar. Third, how does the L29P substitution change the esterase activity? Ester hydrolysis decreased for most esters (75-fold for ethyl acetate) but not for methyl esters. In contrast, L29P PFE catalyzed hydrolysis of {var_epsilon}-caprolactone five times more efficiently than wild-type PFE. Molecular modeling suggests that moving the carbonyl group closer to the

  13. Non-enzymatic N-acetylation of Lysine Residues by AcetylCoA Often Occurs via a Proximal S-acetylated Thiol Intermediate Sensitive to Glyoxalase II.

    Science.gov (United States)

    James, Andrew M; Hoogewijs, Kurt; Logan, Angela; Hall, Andrew R; Ding, Shujing; Fearnley, Ian M; Murphy, Michael P

    2017-02-28

    Acetyl coenzyme A (AcCoA), a key intermediate in mitochondrial metabolism, N-acetylates lysine residues, disrupting and, in some cases, regulating protein function. The mitochondrial lysine deacetylase Sirtuin 3 (Sirt3) reverses this modification with benefits reported in diabetes, obesity, and aging. We show that non-enzymatic lysine N-acetylation by AcCoA is greatly enhanced by initial acetylation of a cysteine residue, followed by SN-transfer of the acetyl moiety to a nearby lysine on mitochondrial proteins and synthetic peptides. The frequent occurrence of an S-acetyl intermediate before lysine N-acetylation suggests that proximity to a thioester is a key determinant of lysine susceptibility to acetylation. The thioesterase glyoxalase II (Glo2) can limit protein S-acetylation, thereby preventing subsequent lysine N-acetylation. This suggests that the hitherto obscure role of Glo2 in mitochondria is to act upstream of Sirt3 in minimizing protein N-acetylation, thus limiting protein dysfunction when AcCoA accumulates. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  14. Nucleosome structure incorporated histone acetylation site prediction in Arabidopsis thaliana.

    Science.gov (United States)

    Zhao, Chen; Liu, Hui; Li, Jiang; Deng, Youping; Shi, Tieliu

    2010-11-02

    Acetylation is a crucial post-translational modification for histones, and plays a key role in gene expression regulation. Due to limited data and lack of a clear acetylation consensus sequence, a few researches have focused on prediction of lysine acetylation sites. Several systematic prediction studies have been conducted for human and yeast, but less for Arabidopsis thaliana. Concerning the insufficient observation on acetylation site, we analyzed contributions of the peptide-alignment-based distance definition and 3D structure factors in acetylation prediction. We found that traditional structure contributes little to acetylation site prediction. Identified acetylation sites of histones in Arabidopsis thaliana are conserved and cross predictable with that of human by peptide based methods. However, the predicted specificity is overestimated, because of the existence of non-observed acetylable site. Here, by performing a complete exploration on the factors that affect the acetylability of lysines in histones, we focused on the relative position of lysine at nucleosome level, and defined a new structure feature to promote the performance in predicting the acetylability of all the histone lysines in A. thaliana. We found a new spacial correlated acetylation factor, and defined a ε-N spacial location based feature, which contains five core spacial ellipsoid wired areas. By incorporating the new feature, the performance of predicting the acetylability of all the histone lysines in A. Thaliana was promoted, in which the previous mispredicted acetylable lysines were corrected by comparing to the peptide-based prediction.

  15. An important role for secreted esterase in disease establishment of the wheat powdery mildew fungus Blumeria graminis f. sp. tritici.

    Science.gov (United States)

    Feng, Jie; Wang, Feng; Hughes, Geoff R; Kaminskyj, Susan; Wei, Yangdou

    2011-03-01

    The activity of esterase secreted by conidia of wheat powdery mildew fungus, Blumeria graminis f. sp. tritici, was assayed using indoxyl acetate hydrolysis, which generates indigo blue crystals. Mature, ungerminated, and germinating conidia secrete esterase(s) on artificial media and on plant leaf surfaces. The activity of these esterases was inhibited by diisopropyl fluorophosphate, which is selective for serine esterases. When conidia were inoculated on wheat leaves pretreated with diisopropyl fluorophosphate, both appressorial germ tube differentiation and symptom development were significantly impaired, indicating an important role of secreted serine esterases in wheat powdery mildew disease establishment.

  16. Structural analysis of thermostabilizing mutations of cocaine esterase

    Energy Technology Data Exchange (ETDEWEB)

    Narasimhan, Diwahar; Nance, Mark R.; Gao, Daquan; Ko, Mei-Chuan; Macdonald, Joanne; Tamburi, Patricia; Yoon, Dan; Landry, Donald M.; Woods, James H.; Zhan, Chang-Guo; Tesmer, John J.G.; Sunahara, Roger K. (Michigan); (Columbia); (Kentucky)

    2010-09-03

    Cocaine is considered to be the most addictive of all substances of abuse and mediates its effects by inhibiting monoamine transporters, primarily the dopamine transporters. There are currently no small molecules that can be used to combat its toxic and addictive properties, in part because of the difficulty of developing compounds that inhibit cocaine binding without having intrinsic effects on dopamine transport. Most of the effective cocaine inhibitors also display addictive properties. We have recently reported the use of cocaine esterase (CocE) to accelerate the removal of systemic cocaine and to prevent cocaine-induced lethality. However, wild-type CocE is relatively unstable at physiological temperatures ({tau}{sub 1/2} {approx} 13 min at 37 C), presenting challenges for its development as a viable therapeutic agent. We applied computational approaches to predict mutations to stabilize CocE and showed that several of these have increased stability both in vitro and in vivo, with the most efficacious mutant (T172R/G173Q) extending half-life up to 370 min. Here we present novel X-ray crystallographic data on these mutants that provide a plausible model for the observed enhanced stability. We also more extensively characterize the previously reported variants and report on a new stabilizing mutant, L169K. The improved stability of these engineered CocE enzymes will have a profound influence on the use of this protein to combat cocaine-induced toxicity and addiction in humans.

  17. PENGGUNAAN XILANASE Streptomyces sp. 45 I-3 AMOBIL UNTUK HIDROLISIS XILAN TONGKOL JAGUNG [Immobilization of Extracellular Xylanase from Streptomyces sp. 45 I-3 for Hydrolysis of Corncob Xylan

    Directory of Open Access Journals (Sweden)

    Anja Meryandini1,2*

    2009-06-01

    Full Text Available Xylan extraction from corncob is done by using alkaline as solvent. Xylan extraction from corncob could give the yields as 10.9%. One percent of corncob xylan is used as substrate to produce the xylanase, compared to oatspelt xylan. Immobilization of xylanase was performed using 1% EudragitTM S100 solution (w/v, with 5:1 volume ratio of xylanase and 1 % EudragitTM S100 (w/v. Activity of the immobilized xylanase was decreased to 23.97% compared with free xylanase. Immobilized xylanase have optimum pH and temperature at 6.0 and 40C respectively, have also thermal stability at 30–40C for an hour. Immobilized xylanase could be reused, but its activity decreased to 52.38% after 3 times application.

  18. Acetylation and oxygenation transformations catalyzed by silica ...

    African Journals Online (AJOL)

    Acetylation of alcohols in refluxing ethyl acetate, and oxidation of aniline and cyclohexanol with 34 % H2O2 in the presence of H3PW12O40 and its supported forms on SiO2 (20 %, 40 %, and 60 % by weight) as active solid acid catalysts were performed under mild reaction conditions with moderate to good yields and with ...

  19. Lysine acetylation of major Chlamydia trachomatis antigens

    Directory of Open Access Journals (Sweden)

    Jelena Mihailovic

    2016-03-01

    Our data show that important Ct antigens could be post-translationally modified by acetylation of lysine residues at multiple sites. Further studies are needed to investigate total acetylome of Ct and the impact PTMs might have on Ct biology and pathogenicity.

  20. Characterization of esterase activity in the Bianchetta trevigiana grape variety under reducing conditions

    Directory of Open Access Journals (Sweden)

    Lomolino G

    2012-10-01

    Full Text Available Giovanna Lomolino, Anna LanteDepartment of Agronomy Food Natural Resources Animals and Environment, Agripolis, Università di Padova Viale dell'Università, Padova, ItalyBackground and methods: While extensive research has been carried out on the enzymes responsible for ester synthesis and hydrolysis by wine strains of Saccharomyces cerevisiae, grape esterase activity is limited. In this study, the autochthonous grape variety, Bianchetta trevigiana, widespread in the Prosecco wine production area of Treviso, Conegliano, and Asolo, Italy, was characterized according to its esterase activity. Because grape skin is very rich in compounds which impart qualitative characteristics to wine, the study of esterase was carried out on this part of the fruit.Results: During enzyme extraction from grape skin, the presence of the reducing agent, β-mercaptoethanol, allowed a better protein yield but reduced esterase activity. Further addition of increasing doses of reducing agents to grape skin protein extract, such as of K2S2O5 (used in winemaking and DTT, reduced or inhibited esterase activity. Even though the zymographic profiles of the extracts obtained with and without β-mercaptoethanol were qualitatively equal, the intensity of enzymatic bands, measured by densitometry, was different.Conclusion: The presence of reducing agents affected the activity of grape skin esterase, and given that this enzyme is involved in the hydrolysis and synthesis of esters, which are important compounds responsible for the flavor of wine, addition of reducing agents could affect the aromatic profile of wine.Keywords: esterase, grape, reducing agent, wine

  1. Enhanced biosurfactant production through cloning of three genes and role of esterase in biosurfactant release

    Directory of Open Access Journals (Sweden)

    Khanna Sunil

    2011-06-01

    Full Text Available Abstract Background Biosurfactants have been reported to utilize a number of immiscible substrates and thereby facilitate the biodegradation of panoply of polyaromatic hydrocarbons. Olive oil is one such carbon source which has been explored by many researchers. However, studying the concomitant production of biosurfactant and esterase enzyme in the presence of olive oil in the Bacillus species and its recombinants is a relatively novel approach. Results Bacillus species isolated from endosulfan sprayed cashew plantation soil was cultivated on a number of hydrophobic substrates. Olive oil was found to be the best inducer of biosurfactant activity. The protein associated with the release of the biosurfactant was found to be an esterase. There was a twofold increase in the biosurfactant and esterase activities after the successful cloning of the biosurfactant genes from Bacillus subtilis SK320 into E.coli. Multiple sequence alignment showed regions of similarity and conserved sequences between biosurfactant and esterase genes, further confirming the symbiotic correlation between the two. Biosurfactants produced by Bacillus subtilis SK320 and recombinant strains BioS a, BioS b, BioS c were found to be effective emulsifiers, reducing the surface tension of water from 72 dynes/cm to as low as 30.7 dynes/cm. Conclusion The attributes of enhanced biosurfactant and esterase production by hyper-producing recombinant strains have many utilities from industrial viewpoint. This study for the first time has shown a possible association between biosurfactant production and esterase activity in any Bacillus species. Biosurfactant-esterase complex has been found to have powerful emulsification properties, which shows promising bioremediation, hydrocarbon biodegradation and pharmaceutical applications.

  2. Enhanced biosurfactant production through cloning of three genes and role of esterase in biosurfactant release.

    Science.gov (United States)

    Sekhon, Kamaljeet Kaur; Khanna, Sunil; Cameotra, Swaranjit Singh

    2011-06-27

    Biosurfactants have been reported to utilize a number of immiscible substrates and thereby facilitate the biodegradation of panoply of polyaromatic hydrocarbons. Olive oil is one such carbon source which has been explored by many researchers. However, studying the concomitant production of biosurfactant and esterase enzyme in the presence of olive oil in the Bacillus species and its recombinants is a relatively novel approach. Bacillus species isolated from endosulfan sprayed cashew plantation soil was cultivated on a number of hydrophobic substrates. Olive oil was found to be the best inducer of biosurfactant activity. The protein associated with the release of the biosurfactant was found to be an esterase. There was a twofold increase in the biosurfactant and esterase activities after the successful cloning of the biosurfactant genes from Bacillus subtilis SK320 into E.coli. Multiple sequence alignment showed regions of similarity and conserved sequences between biosurfactant and esterase genes, further confirming the symbiotic correlation between the two. Biosurfactants produced by Bacillus subtilis SK320 and recombinant strains BioS a, BioS b, BioS c were found to be effective emulsifiers, reducing the surface tension of water from 72 dynes/cm to as low as 30.7 dynes/cm. The attributes of enhanced biosurfactant and esterase production by hyper-producing recombinant strains have many utilities from industrial viewpoint. This study for the first time has shown a possible association between biosurfactant production and esterase activity in any Bacillus species. Biosurfactant-esterase complex has been found to have powerful emulsification properties, which shows promising bioremediation, hydrocarbon biodegradation and pharmaceutical applications.

  3. Enhanced biosurfactant production through cloning of three genes and role of esterase in biosurfactant release

    Science.gov (United States)

    2011-01-01

    Background Biosurfactants have been reported to utilize a number of immiscible substrates and thereby facilitate the biodegradation of panoply of polyaromatic hydrocarbons. Olive oil is one such carbon source which has been explored by many researchers. However, studying the concomitant production of biosurfactant and esterase enzyme in the presence of olive oil in the Bacillus species and its recombinants is a relatively novel approach. Results Bacillus species isolated from endosulfan sprayed cashew plantation soil was cultivated on a number of hydrophobic substrates. Olive oil was found to be the best inducer of biosurfactant activity. The protein associated with the release of the biosurfactant was found to be an esterase. There was a twofold increase in the biosurfactant and esterase activities after the successful cloning of the biosurfactant genes from Bacillus subtilis SK320 into E.coli. Multiple sequence alignment showed regions of similarity and conserved sequences between biosurfactant and esterase genes, further confirming the symbiotic correlation between the two. Biosurfactants produced by Bacillus subtilis SK320 and recombinant strains BioS a, BioS b, BioS c were found to be effective emulsifiers, reducing the surface tension of water from 72 dynes/cm to as low as 30.7 dynes/cm. Conclusion The attributes of enhanced biosurfactant and esterase production by hyper-producing recombinant strains have many utilities from industrial viewpoint. This study for the first time has shown a possible association between biosurfactant production and esterase activity in any Bacillus species. Biosurfactant-esterase complex has been found to have powerful emulsification properties, which shows promising bioremediation, hydrocarbon biodegradation and pharmaceutical applications. PMID:21707984

  4. Gender differences in the activities of aspirin-esterases in rat tissues

    Directory of Open Access Journals (Sweden)

    Benedito M.A.C.

    1998-01-01

    Full Text Available The activities of aspirin (acetylsalicylic acid-esterases were measured in several tissues (liver, kidney, adrenal glands, brain and serum from adult male and female Wistar rats. In males, both aspirin-esterase I (assayed at pH 5.5 and II (assayed at pH 7.4 activities were higher in liver homogenates when compared to females (aspirin-esterase I: males 48.9 ± 4.8 (N = 8 and females 29.3 ± 4.2 (N = 8 nmol of salicylic acid formed min-1 mg protein-1; aspirin-esterase II: males 41.4 ± 4.1 (N = 8 and females 26.1 ± 4.5 (N = 8 nmol of salicylic acid formed min-1 mg protein-1, P<0.001. In serum, enzyme activity was higher in females than in males (aspirin-esterase I: males 0.85 ± 0.06 (N = 6 and females 1.18 ± 0.11 (N = 6 nmol of salicylic acid formed min-1 mg protein-1; aspirin-esterase II: males 1.03 ± 0.13 (N = 6 and females 1.34 ± 0.11 (N = 6 nmol of salicylic acid formed min-1 mg protein-1, P<0.001. In the other tissues assayed, no statistically significant difference between males and females was found. There were no statistically significant differences when the enzymes were assayed in different phases of the estrous cycle in liver and serum. These results show that the differences in aspirin-esterase activity observed between males and females are not due to the estrous cycle. The gender difference obtained in our study may indicate an involvement of gonadal hormones in the control of the hydrolysis of aspirin. This possibility is currently under investigation.

  5. Spinosad resistance, esterase isoenzymes and temporal synergism in Frankliniella occidentalis (Pergande) in Australia.

    Science.gov (United States)

    Herron, Grant A; Gunning, Robin V; Cottage, Emma L A; Borzatta, Valerio; Gobbi, Carlotta

    2014-09-01

    Spinosad has been widely used in Australia to control western flower thrips Frankliniella occidentalis (Pergande) but spinosad usefulness is now compromised by resistance. Here we studied a highly spinosad resistant strain of F. occidentalis to explore if esterases had a role in spinosad resistance. Enhanced esterase activity in pressured spinosad-resistant F. occidentalis was confirmed via PAGE electrophoresis and estimated to be approximately three times higher than that in a susceptible strain. Spinosad-esterase inhibition data in the resistant strain, showed a concentration effect with significant esterase-spinosad binding occurring at spinosad concentrations from 6.2× 10(-7) to 1.5× 10(-5) M. Similarly, a spinosad-piperonyl butoxide (PBO) inhibition curve showed a concentration effect, with significant esterase-PBO binding occurring in the resistant strain at PBO concentrations between 3.3× 10(-5) M and 8.4× 10(-4) M. No binding of esterase to spinosad or PBO occurred in the susceptible strain. Results of bioassays in which spinosad resistant F. occidentalis were sprayed with a 4h delayed release formulation of cyclodextrin-complexed spinosad with immediately available PBO demonstrated that spinosad resistance was significantly reduced from 577 to 72-fold. With further development the PBO synergism of spinosad using a delayed release formulation, similar to that used here, may provide effective control for spinosad resistant F. occidentalis. Temporal synergism of spinosad may prove to be effective tactic for the control of spinosad resistant F. occidentalis where the main resistance mechanism involved has been confirmed to be esterase based. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Three novel rice genes closely related to the Arabidopsis IRX9, IRX9L, and IRX14 genes and their roles in xylan biosynthesis

    Directory of Open Access Journals (Sweden)

    Dawn eChiniquy

    2013-04-01

    Full Text Available Xylan is the second most abundant polysaccharide on Earth, and represents a major component of both dicot wood and the cell walls of grasses. Much knowledge has been gained from studies of xylan biosynthesis in the model plant, Arabidopsis. In particular, the irregular xylem (irx mutants, named for their collapsed xylem cells, have been essential in gaining a greater understanding of the genes involved in xylan biosynthesis. In contrast, xylan biosynthesis in grass cell walls is poorly understood. We identified three rice genes Os07g49370 (OsIRX9, Os01g48440 (OsIRX9L, and Os06g47340 (OsIRX14, from glycosyltransferase family 43 as putative orthologs to the putative β-1,4-xylan backbone elongating Arabidopsis IRX9, IRX9L, and IRX14 genes, respectively. We demonstrate that the overexpression of the closely related rice genes, in full or partly complement the two well-characterized Arabidopsis irregular xylem (irx mutants: irx9 and irx14. Complementation was assessed by measuring dwarfed phenotypes, irregular xylem cells in stem cross sections, xylose content of stems, xylosyltransferase activity of stems, and stem strength. The expression of OsIRX9 in the irx9 mutant resulted in xylosyltransferase activity of stems that was over double that of wild type plants, and the stem strength of this line increased to 124% above that of wild type. Taken together, our results suggest that OsIRX9/OsIRX9L, and OsIRX14, have similar functions to the Arabidopsis IRX9 and IRX14 genes, respectively. Furthermore, our expression data indicate that OsIRX9 and OsIRX9L may function in building the xylan backbone in the secondary and primary cell walls, respectively. Our results provide insight into xylan biosynthesis in rice and how expression of a xylan synthesis gene may be modified to increase stem strength.

  7. Luxury at a Cost? Recombinant Mouse Hepatitis Viruses Expressing the Accessory Hemagglutinin Esterase Protein Display Reduced Fitness In Vitro

    Science.gov (United States)

    Lissenberg, A.; Vrolijk, M. M.; van Vliet, A. L. W.; Langereis, M. A.; de Groot-Mijnes, J. D. F.; Rottier, P. J. M.; de Groot, R. J.

    2005-01-01

    Group 2 coronaviruses encode an accessory envelope glycoprotein species, the hemagglutinin esterase (HE), which possesses sialate-O-acetylesterase activity and which, presumably, promotes virus spread and entry in vivo by facilitating reversible virion attachment to O-acetylated sialic acids. While HE may provide a strong selective advantage during natural infection, many laboratory strains of mouse hepatitis virus (MHV) fail to produce the protein. Apparently, their HE genes were inactivated during cell culture adaptation. For this report, we have studied the molecular basis of this phenomenon. By using targeted RNA recombination, we generated isogenic recombinant MHVs which differ exclusively in their expression of HE and produce either the wild-type protein (HE+), an enzymatically inactive HE protein (HE0), or no HE at all. HE expression or the lack thereof did not lead to gross differences in in vitro growth properties. Yet the expression of HE was rapidly lost during serial cell culture passaging. Competition experiments with mixed infections revealed that this was not due to the enzymatic activity: MHVs expressing HE+ or HE0 propagated with equal efficiencies. During the propagation of recombinant MHV-HE+, two types of spontaneous mutants accumulated. One produced an anchorless HE, while the other had a Gly-to-Trp substitution at the predicted C-terminal residue of the HE signal peptide. Neither mutant incorporated HE into virion particles, suggesting that wild-type HE reduces the in vitro propagation efficiency, either at the assembly stage or at a postassembly level. Our findings demonstrate that the expression of “luxury” proteins may come at a fitness penalty. Apparently, under natural conditions the costs of maintaining HE are outweighed by the benefits. PMID:16306576

  8. Determination of Isoniazid Acetylator Phenotype and its Clinical ...

    African Journals Online (AJOL)

    After a 2-month treatment of the TB patients, the sputum smear cultures were negative in about 81% independent of the acetylator or HIV status. Early side effects experienced were dominated by peripheral neuropathies mostly in slow and intermediate acetylators. Key words: Acetylator, isoniazid, phenotype, tuberculosis, ...

  9. Efficient acetylation of primary amines and amino acids in ...

    Indian Academy of Sciences (India)

    Abstract. Acetyl chloride is one of the most commonly available and cheap acylating agent but its high reactivity and concomitant instability in water precludes its use to carry out acetylation in aqueous medium. The present methodology illustrates the efficient acetylation of primary amines and amino acids in brine solution ...

  10. The growing landscape of lysine acetylation links metabolism and cell signalling

    DEFF Research Database (Denmark)

    Choudhary, Chuna Ram; Weinert, Brian Tate; Nishida, Yuya

    2014-01-01

    Lysine acetylation is a conserved protein post-translational modification that links acetyl-coenzyme A metabolism and cellular signalling. Recent advances in the identification and quantification of lysine acetylation by mass spectrometry have increased our understanding of lysine acetylation...

  11. Structural studies of the mix-linked beta-(1-->3)/beta-(1-->4)-D-xylans from the cell wall of Palmaria palmata (Rhodophyta).

    Science.gov (United States)

    Deniaud, Estelle; Quemener, Bernard; Fleurence, Joël; Lahaye, Marc

    2003-11-01

    The structure and organization of Palmaria palmata cell walls, which are largely involved in biological and physiological functions as well as in biotechnological and food applications of this red marine alga, are principally assumed by the interactions and linkages of major mix-linked beta-(1-->3)/beta-(1-->4)-D-xylans. These partly acidic polysaccharides are essentially held in the cell wall by H-bonds. The location of the acid groups and the distribution of 1-->3-linkage were studied following the endo-beta-(1,4)-xylanase hydrolysis of sequentially extracted xylans, and fine analysis of the oligosaccharides produced by anion exchange chromatography, high performance anion exchange chromatography (HPAEC)-PAD, nuclear magnetic resonance (NMR) and electrospray ion trap mass spectrometry (ESI-MS) techniques. The results indicate that the acidity of the xylans was related to potential linkages to sulfated and/or phosphorylated xylogalactoprotein complexes. H-bonding of the mix-linked xylans involved a regular 1,3-linkages distribution idealized in a pentameric repeating structure (one 1,3-linkage and four 1,4-linkages). Furthermore, MS analysis of the xylo-oligosaccharides revealed a substitution of the mix-linked xylans by a non-osidic component of 175 g mol(-1). The presence of this substituent and of the proposed covalent linkage between the mix-linked xylans and charged glycoproteins are discussed with regard to the polysaccharides interactions in P. palmata cell walls.

  12. Enzymic hydrolysis of xylans. I. A high xylanase and beta-xylosidase producing strain of Aspergillus niger

    Energy Technology Data Exchange (ETDEWEB)

    Conrad, D.

    1981-01-01

    Aspergillus niger, strain 110.42 (CBS) was selected as a producer of high xylanolytic activities. The time course of xylanase and beta-xylosidase production as well as the effect of pH and temperature on the activity of these enzymes were studied. High-performance liquid chromatography analysis of the enzymic degradation of arabinoxylan showed a nearly complete conversion to pentose sugars. Aspects of using crude xylanase preparations for enzymic saccharification of xylans are discussed.

  13. Xylan synthesized by Irregular Xylem 14 (IRX14) maintains the structure of seed coat mucilage in Arabidopsis

    OpenAIRE

    Hu, Ruibo; Li, Junling; Wang, Xiaoyu; Zhao, Xun; Yang, Xuanwen; Tang, Qi; He, Guo; Zhou, Gongke; Kong, Yingzhen

    2016-01-01

    During differentiation, the Arabidopsis seed coat epidermal cells synthesize and secrete large quantities of pectinaceous mucilage into the apoplast, which is then released to encapsulate the seed upon imbibition. In this study, we showed that mutation in Irregular Xylem 14 (IRX14) led to a mucilage cohesiveness defect due to a reduced xylan content. Expression of IRX14 was detected specifically in the seed coat epidermal cells, reaching peak expression at 13 days post-anthesis (DPA) when the...

  14. Esterase Active in Polar Organic Solvents from the Yeast Pseudozyma sp. NII 08165

    Directory of Open Access Journals (Sweden)

    Deepthy Alex

    2014-01-01

    Full Text Available Esterases/lipases active in water miscible solvents are highly desired in biocatalysis where substrate solubility is limited and also when the solvent is desired as an acyl acceptor in transesterification reactions, as with the case of biodiesel production. We have isolated an esterase from the glycolipid producing yeast-Pseudozyma sp. NII 08165 which in its crude form was alkali active, thermo stable, halo tolerant and also capable of acting in presence of high methanol concentration. The crude enzyme which maintained 90% of its original activity after being treated at 70°C was purified and the properties were characterized. The partially purified esterase preparation had temperature and pH optima of 60°C and 8.0 respectively. The enzyme retained almost complete activity in presence of 25% methanol and 80% activity in the same strength of ethanol. Conditions of enzyme production were optimized, which lead to 9 fold increase in the esterase yield. One of the isoforms of the enzyme LIP1 was purified to homogeneity and characterized. Purified LIP1 had a Km and Vmax of 0.01 and 1.12, respectively. The purified esterase lost its thermo and halo tolerance but interestingly, retained 97% activity in methanol.

  15. Organophosphate and pyrethroid hydrolase activities of mutant Esterases from the cotton bollworm Helicoverpa armigera.

    Science.gov (United States)

    Li, Yongqiang; Farnsworth, Claire A; Coppin, Chris W; Teese, Mark G; Liu, Jian-Wei; Scott, Colin; Zhang, Xing; Russell, Robyn J; Oakeshott, John G

    2013-01-01

    Two mutations have been found in five closely related insect esterases (from four higher Diptera and a hymenopteran) which each confer organophosphate (OP) hydrolase activity on the enzyme and OP resistance on the insect. One mutation converts a Glycine to an Aspartate, and the other converts a Tryptophan to a Leucine in the enzymes' active site. One of the dipteran enzymes with the Leucine mutation also shows enhanced activity against pyrethroids. Introduction of the two mutations in vitro into eight esterases from six other widely separated insect groups has also been reported to increase substantially the OP hydrolase activity of most of them. These data suggest that the two mutations could contribute to OP, and possibly pyrethroid, resistance in a variety of insects. We therefore introduced them in vitro into eight Helicoverpa armigera esterases from a clade that has already been implicated in OP and pyrethroid resistance. We found that they do not generally enhance either OP or pyrethroid hydrolysis in these esterases but the Aspartate mutation did increase OP hydrolysis in one enzyme by about 14 fold and the Leucine mutation caused a 4-6 fold increase in activity (more in one case) of another three against some of the most insecticidal isomers of fenvalerate and cypermethrin. The Aspartate enzyme and one of the Leucine enzymes occur in regions of the H. armigera esterase isozyme profile that have been previously implicated in OP and pyrethroid resistance, respectively.

  16. Organophosphate and pyrethroid hydrolase activities of mutant Esterases from the cotton bollworm Helicoverpa armigera.

    Directory of Open Access Journals (Sweden)

    Yongqiang Li

    Full Text Available Two mutations have been found in five closely related insect esterases (from four higher Diptera and a hymenopteran which each confer organophosphate (OP hydrolase activity on the enzyme and OP resistance on the insect. One mutation converts a Glycine to an Aspartate, and the other converts a Tryptophan to a Leucine in the enzymes' active site. One of the dipteran enzymes with the Leucine mutation also shows enhanced activity against pyrethroids. Introduction of the two mutations in vitro into eight esterases from six other widely separated insect groups has also been reported to increase substantially the OP hydrolase activity of most of them. These data suggest that the two mutations could contribute to OP, and possibly pyrethroid, resistance in a variety of insects. We therefore introduced them in vitro into eight Helicoverpa armigera esterases from a clade that has already been implicated in OP and pyrethroid resistance. We found that they do not generally enhance either OP or pyrethroid hydrolysis in these esterases but the Aspartate mutation did increase OP hydrolysis in one enzyme by about 14 fold and the Leucine mutation caused a 4-6 fold increase in activity (more in one case of another three against some of the most insecticidal isomers of fenvalerate and cypermethrin. The Aspartate enzyme and one of the Leucine enzymes occur in regions of the H. armigera esterase isozyme profile that have been previously implicated in OP and pyrethroid resistance, respectively.

  17. Relationships between Substrate Promiscuity and Chiral Selectivity of Esterases from Phylogenetically and Environmentally Diverse Microorganisms

    Directory of Open Access Journals (Sweden)

    Cristina Coscolín

    2018-01-01

    Full Text Available Substrate specificity and selectivity of a biocatalyst are determined by the protein sequence and structure of its active site. Finding versatile biocatalysts acting against multiple substrates while at the same time being chiral selective is of interest for the pharmaceutical and chemical industry. However, the relationships between these two properties in natural microbial enzymes remain underexplored. Here, we performed an experimental analysis of substrate promiscuity and chiral selectivity in a set of 145 purified esterases from phylogenetically and environmentally diverse microorganisms, which were assayed against 96 diverse esters, 20 of which were enantiomers. Our results revealed a negative correlation between substrate promiscuity and chiral selectivity in the evaluated enzymes. Esterases displaying prominent substrate promiscuity and large catalytic environments are characterized by low chiral selectivity, a feature that has limited commercial value. Although a low level of substrate promiscuity does not guarantee high chiral selectivity, the probability that esterases with smaller active sites possess chiral selectivity factors of interest for industry (>25 is significantly higher than for promiscuous enzymes. Together, the present study unambiguously demonstrates that promiscuous and selective esterases appear to be rare in nature and that substrate promiscuity can be used as an indicator of the chiral selectivity level of esterases, and vice versa.

  18. Facilitating the enzymatic saccharification of pulped bamboo residues by degrading the remained xylan and lignin-carbohydrates complexes.

    Science.gov (United States)

    Huang, Caoxing; He, Juan; Li, Xin; Min, Douyong; Yong, Qiang

    2015-09-01

    Kraft pulping was performed on bamboo residues and its impact on the chemical compositions and the enzymatic digestibility of the samples were investigated. To improve the digestibility of sample by degrading the xylan and lignin-carbohydrates complexes (LCCs), xylanase and α-L-arabinofuranosidase (AF) were supplemented with cellulase. The results showed more carbohydrates were remained in the samples pulped with low effective alkali (EA) charge, compared to conventional kraft pulping. When 120 IU/g xylanase and 15 IU/g AF were supplemented with 20 FPU/g cellulase, the xylan degradation yield of the sample pulped with 12% EA charge increased from 68.20% to 88.35%, resulting in an increased enzymatic saccharification efficiency from 58.98% to 83.23%. The amount of LCCs in this sample decreased from 8.63/100C9 to 2.99/100C9 after saccharification with these enzymes. The results indicated that degrading the remained xylan and LCCs in the pulp could improve its enzymatic digestibility. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Fungal treatment of cornstalks enhances the delignification and xylan loss during mild alkaline pretreatment and enzymatic digestibility of glucan.

    Science.gov (United States)

    Yu, Hongbo; Du, Wanqing; Zhang, Ji; Ma, Fuying; Zhang, Xiaoyu; Zhong, Weixin

    2010-09-01

    Fungal treatment with Irpex lacteus was used to enhance the delignification and xylan loss during mild alkaline pretreatment and subsequent enzymatic conversion in this research. The 15-day bio-treatment can modify the lignin structure and increase losses of lignin (from 75.67% to 80.00%) and xylan (from 40.68% to 51.37%) during alkaline pretreatment, making the enzymatic conversion more efficient. The high digestibility of glucan can be obtained after the bio-treatment and alkaline pretreatment at near room-temperature (30 degrees C), and the maximum digestibility increased 14% in comparison with that after the sole alkaline pretreatment. The bio-treatment enhanced delignification and glucan digestibility more significantly when the alkaline pretreatment was performed at lower severity. Additionally, Nuclei Growth model with a time-dependent rate constant can describe well the delignification and xylan loss. Results indicated that the bio-treatment increased the rate constant of initial reaction, but accelerated the decline of rate constant during alkaline pretreatment. (c) 2010 Elsevier Ltd. All rights reserved.

  20. Fusion of a xylan-binding module to gluco-oligosaccharide oxidase increases activity and promotes stable immobilization.

    Directory of Open Access Journals (Sweden)

    Thu V Vuong

    Full Text Available The xylan-binding module Clostridium thermocellum CBM22A was successfully fused to a gluco-oligosaccharide oxidase, GOOX-VN, from Sarocladium strictum via a short TP linker, allowing the fused protein to effectively bind different xylans. The presence of the CtCBM22A at the N-terminal of GOOX-VN increased catalytic activity on mono- and oligo-saccharides by 2-3 fold while not affecting binding affinity to these substrates. Notably, both GOOX-VN and its CBM fusion also showed oxidation of xylo-oligosaccharides with degrees of polymerization greater than six. Whereas fusion to CtCBM22A did not alter the thermostability of GOOX-VN or reduce substrate inhibition, CtCBM22A_GOOX-VN could be immobilized to insoluble oat spelt xylan while retaining wild-type activity. QCM-D analysis showed that the fused enzyme remained bound during oxidation. These features could be harnessed to generate hemicellulose-based biosensors that detect and quantify the presence of different oligosaccharides.

  1. Adhesives for Achieving Durable Bonds with Acetylated Wood

    Directory of Open Access Journals (Sweden)

    Charles R. Frihart

    2017-12-01

    Full Text Available Acetylation of wood imparts moisture durability, decay resistance, and dimensional stability to wood; however, making durable adhesive bonds with acetylated wood can be more difficult than with unmodified wood. The usual explanation is that the acetylated surface has fewer hydroxyl groups, resulting in a harder-to-wet surface and in fewer hydrogen bonds between wood and adhesive. This concept was evaluated using four different adhesives (resorcinol–formaldehyde, emulsion polymer isocyanate, epoxy, and melamine–formaldehyde with unmodified wood, acetylated wood, and acetylated wood that had been planed. Strikingly, acetylation did not hinder adhesive bonds with a waterborne resorcinol–formaldehyde adhesive that bonded equally well to both unmodified and acetylated yellow poplar. An epoxy adhesive bonded better to the acetylated wood than to the unmodified wood, in contrast to an emulsion polymer isocyanate, which gave less durable bonds to acetylated than to unmodified wood. Planing of the acetylated wood surface prior to bonding reduced bond durability for the epoxy adhesive and increased the amount of surface hydroxyl groups, as measured using X-ray photoelectron spectroscopic analysis of the trifluoroacetic anhydride-treated wood. These experiments showed that wood modification is useful in understanding wood-adhesive interactions, in addition to determining how to develop adhesives for acetylated woods.

  2. Identification of the active site serine in pancreatic cholesterol esterase by chemical modification and site-specific mutagenesis.

    Science.gov (United States)

    DiPersio, L P; Fontaine, R N; Hui, D Y

    1990-10-05

    Chemical modification and site-specific mutagenesis approaches were used in this study to identify the active site serine residue of pancreatic cholesterol esterase. In the first approach, purified porcine pancreatic cholesterol esterase was covalently modified by incubation with [3H]diisopropylfluorophosphate (DFP). The radiolabeled cholesterol esterase was digested with CNBr, and the peptides were separated by high performance liquid chromatography. A single 3H-containing peptide was obtained for sequence determination. The results revealed the binding of DFP to a serine residue within the serine esterase homologous domain of the protein. Furthermore, the DFP-labeled serine was shown to correspond to serine residue 194 of rat cholesterol esterase (Kissel, J. A., Fontaine, R. N., Turck, C. W., Brockman, H. L., and Hui, D. Y. (1989) Biochim. Biophys. Acta 1006, 227-236). The codon for serine 194 in rat cholesterol esterase cDNA was then mutagenized to ACT or GCT to yield mutagenized cholesterol esterase with either threonine or alanine, instead of serine, at position 194. Expression of the mutagenized cDNA in COS-1 cells demonstrated that substitution of serine 194 with threonine or alanine abolished enzyme activity in hydrolyzing the water-soluble substrate, p-nitrophenyl butyrate, and the lipid substrates cholesteryl [14C]oleate and [14C] lysophosphatidylcholine. These studies definitively identified serine 194 in the catalytic site of pancreatic cholesterol esterase.

  3. Role of the N-acetylation polymorphism in solithromycin metabolism.

    Science.gov (United States)

    Hein, David W; Doll, Mark A

    2017-06-01

    Solithromycin is a new macrolide antibiotic for the potential treatment of bacterial pneumonia. Solithromycin N-acetylation by human NAT1 and NAT2 was investigated following recombinant expression in yeast and in cryopreserved human hepatocytes from rapid, intermediate and slow acetylators. Solithromycin exhibited over twofold higher affinity for recombinant human NAT2 than NAT1. Apparent maximum velocities for the N-acetylation of solithromycin catalyzed by the NAT2 allozyme associated with rapid acetylators were significantly (p intermediate>slow acetylators) were exhibited in cryopreserved human hepatocytes in situ following incubation with 100 μM solithromycin. Solithromycin is N-acetylated by human NAT1 and NAT2 and the role of the NAT2 acetylation polymorphism on solithromycin metabolism may be concentration dependent.

  4. Enzymatic sequencing of partially acetylated chitosan oligomers.

    Science.gov (United States)

    Hamer, Stefanie Nicole; Moerschbacher, Bruno Maria; Kolkenbrock, Stephan

    2014-06-17

    Chitosan oligosaccharides have diverse biological activities with potentially valuable applications, for example, in the fields of medicine and agriculture. These functionalities are thought to depend on their degree of polymerization and acetylation, and possibly on specific patterns of acetylation. Chitosan oligomers with fully defined architecture are difficult to produce, and their complete analysis is demanding. Analysis is typically done using MS or NMR, requiring access to expensive infrastructure, and yielding unequivocal results only in the case of rather small oligomers. We here describe a simple and cost-efficient method for the sequencing of μg amounts of chitosan oligosaccharides which is based on the sequential action of two recombinant glycosidases, namely an exo-β-N-acetylhexosaminidase (GlcNAcase) from Bacillus subtilis 168 and an exo-β-d-glucosaminidase (GlcNase) from Thermococcus kodakarensis KOD1. Starting from the non-reducing end, GlcNAcase and GlcNase specifically remove N-acetyl glucosamine (A) and glucosamine (D) units, respectively. By the sequential addition and removal of these enzymes in an alternating way followed by analysis of the products using high-performance thin-layer chromatography, the sequence of chitosan oligosaccharides can be revealed. Importantly, both enzymes work under identical conditions so that no buffer exchange is required between steps, and the enzyme can be removed conveniently using simple ultra-filtration devices. As proof-of-principle, the method was used to sequence the product of enzymatic deacetylation of chitin pentamer using a recombinant chitin deacetylase from Vibrio cholerae which specifically removes the acetyl group from the second unit next to the non-reducing end of the substrate, yielding mono-deacetylated pentamer with the sequence ADAAA. Copyright © 2014 Elsevier Ltd. All rights reserved.

  5. Contribution of soil esterase to biodegradation of aliphatic polyester agricultural mulch film in cultivated soils.

    Science.gov (United States)

    Yamamoto-Tamura, Kimiko; Hiradate, Syuntaro; Watanabe, Takashi; Koitabashi, Motoo; Sameshima-Yamashita, Yuka; Yarimizu, Tohru; Kitamoto, Hiroko

    2015-01-01

    The relationship between degradation speed of soil-buried biodegradable polyester film in a farmland and the characteristics of the predominant polyester-degrading soil microorganisms and enzymes were investigated to determine the BP-degrading ability of cultivated soils through characterization of the basal microbial activities and their transition in soils during BP film degradation. Degradation of poly(butylene succinate-co-adipate) (PBSA) film was evaluated in soil samples from different cultivated fields in Japan for 4 weeks. Both the degradation speed of the PBSA film and the esterase activity were found to be correlated with the ratio of colonies that produced clear zone on fungal minimum medium-agarose plate with emulsified PBSA to the total number colonies counted. Time-dependent change in viable counts of the PBSA-degrading fungi and esterase activities were monitored in soils where buried films showed the most and the least degree of degradation. During the degradation of PBSA film, the viable counts of the PBSA-degrading fungi and the esterase activities in soils, which adhered to the PBSA film, increased with time. The soil, where the film was degraded the fastest, recorded large PBSA-degrading fungal population and showed high esterase activity compared with the other soil samples throughout the incubation period. Meanwhile, esterase activity and viable counts of PBSA-degrading fungi were found to be stable in soils without PBSA film. These results suggest that the higher the distribution ratio of native PBSA-degrading fungi in the soil, the faster the film degradation is. This could be due to the rapid accumulation of secreted esterases in these soils.

  6. Fragrance material review on acetyl carene.

    Science.gov (United States)

    Scognamiglio, J; Letizia, C S; Api, A M

    2013-12-01

    A toxicologic and dermatologic review of acetyl carene when used as a fragrance ingredient is presented. Acetyl carene is a member of the fragrance structural group Alkyl Cyclic Ketones. These fragrances can be described as being composed of an alkyl, R1, and various substituted and bicyclic saturated or unsaturated cyclic hydrocarbons, R2, in which one of the rings may include up to 12 carbons. Alternatively, R2 may be a carbon bridge of C2-C4 carbon chain length between the ketone and cyclic hydrocarbon. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for acetyl carene were evaluated then summarized and includes physical properties, acute toxicity, skin irritation, and skin sensitization data. A safety assessment of the entire Alkyl Cyclic Ketones will be published simultaneously with this document; please refer to Belsito et al. (Belsito, D., Bickers, D., Bruze, M., Calow, P., Dagli, M., Fryer, A.D., Greim, H., Miyachi, Y., Saurat, J.H., Sipes, I.G., 2013A Toxicologic and dermatologic assessment of alkyl cyclic ketones when used as fragrance ingredients. (submitted for publication).) for an overall assessment of the safe use of this material and all Alkyl Cyclic Ketones in fragrances. Copyright © 2013. Published by Elsevier Ltd.

  7. Zymography Detection of a Bacterial Extracellular Thermoalkaline Esterase/Lipase Activity.

    Science.gov (United States)

    Tapizquent, María; Fernández, Marleny; Barreto, Georgina; Hernández, Zully; Contreras, Lellys M; Kurz, Liliana; Wilkesman, Jeff

    2017-01-01

    Lipases are esterases that occur widely in nature, yet those with commercial relevance are exclusively from microbial origin. Glycerol and long-chain fatty acids are the products after hydrolysis of esters bonds in saponifiable lipids catalyzed by lipases. In this work, we describe lipase/esterase activity contained in cell-free fractions from thermophilic bacteria, cultured in medium containing olive oil. Analysis of the cell-free fractions by electrotransference zymography, using tributyrin as substrate, revealed bands corresponding to lipase activity. The method is simple, fast, and inexpensive.

  8. O-acetylation of Plant Cell Wall Polysaccharides

    Directory of Open Access Journals (Sweden)

    Sascha eGille

    2012-01-01

    Full Text Available Plant cell walls are composed of structurally diverse polymers, many of which are O-acetylated. How plants O-acetylate wall polymers and what its function is remained elusive until recently, when two protein families were identified in the model plant Arabidopsis that are involved in the O-acetylation of wall polysaccharides – the reduced wall acetylation (RWA and the trichome birefringence-like (TBL proteins. This review discusses the role of these two protein families in polysaccharide O-acetylation and outlines the differences and similarities of polymer acetylation mechanisms in plants, fungi, bacteria and mammals. Members of the TBL protein family had been shown to impact pathogen resistance, freezing tolerance, and cellulose biosynthesis. The connection of TBLs to polysaccharide O-acetylation thus gives crucial leads into the biological function of wall polymer O-acetylation.From a biotechnological point understanding the O-acetylation mechanism is important as acetyl-substituents inhibit the enzymatic degradation of wall polymers and released acetate can be a potent inhibitor in microbial fermentations, thus impacting the economic viability of e.g. lignocellulosic based biofuel production.

  9. Thermophilic acetylxylan esterase genes and enzymes from alicyclobacillus acidocaldarius and related organisms and methods

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, Vicki S.; Thompson, David N.; Reed, David W.; Lacey, Jeffrey A.; Apel, William A.

    2018-01-09

    A genetically modified organism comprising at least one nucleic acid sequence and/or at least one recombinant nucleic acid isolated from Alicyclobacillus acidocaldarius and encoding a polypeptide involved in at least partially degrading, cleaving, transporting, metabolizing, or removing polysaccharide, lignocellulose, hemicellulose, lignin, chitin, heteroxylan, and/or xylan-decorating group; and at least one nucleic acid sequence and/or at least one recombinant nucleic acid encoding a polypeptide involved in fermenting sugar molecules to a product. Additionally, enzymatic and/or proteinaceous extracts may be isolated from one or more genetically modified organisms. The extracts are utilized to convert biomass into a product. Further provided are methods of converting biomass into products comprising: placing the genetically modified organism and/or enzymatic extracts thereof in fluid contact with polysaccharides, cellulose, lignocellulose, hemicellulose, lignin, starch, sugars, sugar oligomers, carbohydrates, complex carbohydrates, chitin, heteroxylans, glycosides, and/or xylan-, glucan-, galactan-, or mannan-decorating groups.

  10. Protein acetylation involved in streptomycin biosynthesis in Streptomyces griseus.

    Science.gov (United States)

    Ishigaki, Yuji; Akanuma, Genki; Yoshida, Minoru; Horinouchi, Sueharu; Kosono, Saori; Ohnishi, Yasuo

    2017-02-23

    Protein acetylation, the reversible addition of an acetyl group to lysine residues, is a protein post-translational modification ubiquitous in living cells. Although the involvement of protein acetylation in the regulation of primary metabolism has been revealed, the function of protein acetylation is largely unknown in secondary metabolism. Here, we characterized protein acetylation in Streptomyces griseus, a streptomycin producer. Protein acetylation was induced in the stationary and sporulation phases in liquid and solid cultures, respectively, in S. griseus. By comprehensive acetylome analysis, we identified 134 acetylated proteins with 162 specific acetylated sites. Acetylation was found in proteins related to primary metabolism and translation, as in other bacteria. However, StrM, a deoxysugar epimerase involved in streptomycin biosynthesis, was identified as a highly acetylated protein by 2-DE-based proteomic analysis. The Lys70 residue, which is critical for the enzymatic activity of StrM, was the major acetylation site. Thus, acetylation of Lys70 was presumed to abolish enzymatic activity of StrM. In accordance with this notion, an S. griseus mutant producing the acetylation-mimic K70Q StrM hardly produced streptomycin, though the K70Q mutation apparently decreased the stability of StrM. A putative lysine acetyltransferase (KAT) SGR1683 in S. griseus, as well as the Escherichia coli KAT YfiQ, acetylated Lys70 of StrM in vitro. Furthermore, absolute quantification analysis estimated that 13% of StrM molecules were acetylated in mycelium grown in solid culture for 3days. These results indicate that StrM acetylation is of biological significance. We propose that StrM acetylation functions as a limiter of streptomycin biosynthesis in S. griseus. Protein acetylation has been extensively studied not only in eukaryotes, but also in prokaryotes. The acetylome has been analyzed in more than 14 bacterial species. Here, by comprehensive acetylome analysis, we showed

  11. Insights into the effect of N-acetyl-L-cysteine-capped CdTe quantum dots on the structure and activity of human serum albumin by spectroscopic techniques

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Haoyu; Yang, Xudan; Li, Meng; Han, Songlin; Liu, Yingxue [School of Environmental Science and Engineering, Shandong University, China-America CRC for Environment & Health, Shandong Province, 27# Shanda South Road, Jinan 250100 (China); Tan, Xuejie [School of Chemistry and Pharmaceutical Engineering, Qilu University of Technology, Jinan, Shandong Province 250353 (China); Liu, Chunguang, E-mail: chunguangliu2013@sdu.edu.cn [School of Environmental Science and Engineering, Shandong University, China-America CRC for Environment & Health, Shandong Province, 27# Shanda South Road, Jinan 250100 (China); Liu, Rutao [School of Environmental Science and Engineering, Shandong University, China-America CRC for Environment & Health, Shandong Province, 27# Shanda South Road, Jinan 250100 (China)

    2015-11-15

    Quantum dots (QDs) are a kind of nanostructured semiconductor crystals with the size range of 1–10 nm. Their unique photophysical properties and potential toxicity to human health have aroused wide concern of scientists and general public. However, the interaction mechanism of QDs on human serum albumin (HSA, the vital protein in human blood) from both structural and functional perspectives is rarely reported. In the present work, effects of N-acetyl-L-cysteine-capped CdTe quantum dots with fluorescence emission peak at 612 nm (QDs-612) on the conformation and function of HSA were investigated by spectroscopic methods, molecular docking study and esterase activity assay. The hydrophobic interaction between HSA and QDs-612 was spontaneous with the binding constants calculated to be 6.85×10{sup 5} L mol{sup −1} (298 K) and 8.89×10{sup 5} L mol{sup −1} (308 K). The binding of QDs-612 to HSA induced the static quenching of fluorescence and the changes of secondary structure and microenvironment of Tyr-411 residue, which resulted in serious decrease on the hydrolysis of substrate p-nitrophenylacetate in esterase activity assay of HSA. This work confirms the possibility on direct interaction of QDs-612 with HSA and obtains a possible mechanism of relationship between conformation and function of HSA. - Highlights: • The interaction between CdTe QDs (QDs-612) and HSA is spontaneous. • The predominant force of the binding is hydrophobic interaction. • The interaction changes the secondary structure of HSA. • Tyr-411 residue of HSA expose to a hydrophilic environment. • The esterase activity of HSA decreases by adding QDs-612.

  12. Xylans Provide the Structural Driving Force for Mucilage Adhesion to the Arabidopsis Seed Coat1

    Science.gov (United States)

    Crépeau, Marie-Jeanne; Vigouroux, Jacqueline; Berger, Adeline; Sallé, Christine; Botran, Lucy

    2016-01-01

    Arabidopsis (Arabidopsis thaliana) seed coat epidermal cells produce large amounts of mucilage that is released upon imbibition. This mucilage is structured into two domains: an outer diffuse layer that can be easily removed by agitation and an inner layer that remains attached to the outer seed coat. Both layers are composed primarily of pectic rhamnogalacturonan I (RG-I), the inner layer also containing rays of cellulose that extend from the top of each columella. Perturbation in cellulosic ray formation has systematically been associated with a redistribution of pectic mucilage from the inner to the outer layer, in agreement with cellulose-pectin interactions, the nature of which remained unknown. Here, by analyzing the outer layer composition of a series of mutant alleles, a tight proportionality of xylose, galacturonic acid, and rhamnose was evidenced, except for mucilage modified5-1 (mum5-1; a mutant showing a redistribution of mucilage pectin from the inner adherent layer to the outer soluble one), for which the rhamnose-xylose ratio was increased drastically. Biochemical and in vitro binding assay data demonstrated that xylan chains are attached to RG-I chains and mediate the adsorption of mucilage to cellulose microfibrils. mum5-1 mucilage exhibited very weak adsorption to cellulose. MUM5 was identified as a putative xylosyl transferase recently characterized as MUCI21. Together, these findings suggest that the binding affinity of xylose ramifications on RG-I to a cellulose scaffold is one of the factors involved in the formation of the adherent mucilage layer. PMID:26979331

  13. Preparation of Polyvinyl Alcohol/Xylan Blending Films with 1,2,3,4-Butane Tetracarboxylic Acid as a New Plasticizer

    Directory of Open Access Journals (Sweden)

    Cun-dian Gao

    2014-01-01

    Full Text Available Miscible, biodegradable polyvinyl alcohol (PVA/xylan blending films were firstly prepared in the range of the PVA/xylan weight ratio from 1 : 2 to 3 : 1 by casting method using 1,2,3,4-butane tetracarboxylic acid (BTCA as a new plasticizer. The properties of blending films as functions of PVA/xylan weight ratio and BTCA amount were discussed. XRD and FT-IR were applied to characterize the blending films. Experimental results indicated that tensile strength (TS and elongation at break (EAB of blending films decreased along with the decrease of the PVA/xylan weight ratio. Both of TS and EAB firstly increased and then decreased as the amount of BTCA was increased. More importantly, blending films were biodegraded almost by 41% with an addition of 10% BTCA in blending films within 30 days in soil. For all hydroxyl functionalized polymers (xylan and PVA, their molecular interactions and miscibility with BTCA endowed blending films with the biocompatibility and biodegradability. Therefore, these blending films are environmentally friendly materials which could be applied as biodegradable plastics for food packaging and agricultural applications.

  14. Fungal glucuronoyl esterases : Genome mining based enzyme discovery and biochemical characterization

    NARCIS (Netherlands)

    Dilokpimol, Adiphol; Mäkelä, Miia R; Cerullo, Gabriella; Zhou, Miaomiao; Varriale, Simona; Gidijala, Loknath; Brás, Joana L A; Jütten, Peter; Piechot, Alexander; Verhaert, Raymond; Faraco, Vincenza; Hilden, Kristiina S.; de Vries, Ronald P

    2018-01-01

    4-O-Methyl-d-glucuronic acid (MeGlcA) is a side-residue of glucuronoarabinoxylan and can form ester linkages to lignin, contributing significantly to the strength and rigidity of the plant cell wall. Glucuronoyl esterases (4-O-methyl-glucuronoyl methylesterases, GEs) can cleave this ester bond, and

  15. Characterization and histochemical localization of nonspecific esterase from ascocarps of desert truffle (Terfezia claveryi Chatin).

    Science.gov (United States)

    Pérez-Gilabert, Manuela; Morte, Asunción; Avila-González, Rizette; García-Carmona, Francisco

    2005-07-13

    An esterase activity from Terfezia claveryi Chatin ascocarps, a mycorrhizal hypogeous fungus, is described for the first time. The enzyme was partially purified using phase partitioning in Triton X-114 (TX-114), achieving a reduction of 87% in the triglyceride content and the removal of 63% of phenols. The enzyme showed maximum activity toward short-chain p-nitrophenyl esters, and no interfacial activation was observed, indicating that the enzyme responsible for this activity is an esterase and not a lipase. This esterase presented its maximum activity at pH 7.4 and 60 degrees C. The values obtained for Km at pH 7.4 were 0.3 mM for p-nitrophenyl butyrate and 0.6 mM for p-nitrophenyl acetate with catalytic efficiencies (Vmax/Km) of 0.23 and 0.32, respectively. T. claveryi esterase was inhibited by phenylboric acid, indicating that serine residues were involved in the enzyme activity. This activity was localized only in the hypothecium and was absent from the peridium and gleba.

  16. Chaperone-like activities of α-synuclein: α-Synuclein assists enzyme activities of esterases

    International Nuclear Information System (INIS)

    Ahn, Misun; Kim, SeungBum; Kang, Mira; Ryu, Yeonwoo; Doohun Kim, T.

    2006-01-01

    α-Synuclein, a major constituent of Lewy bodies (LBs), has been implicated to play a critical role in the pathogenesis of Parkinson's disease (PD), although the physiological function of α-synuclein has not yet been known. Here we have shown that α-synuclein, which has no well-defined secondary or tertiary structure, can protect the enzyme activity of microbial esterases against stress conditions such as heat, pH, and organic solvents. In particular, the flexibility of α-synuclein and its C-terminal region seems to be important for complex formation, but the structural integrity of the C-terminal region may not be required for stabilization of enzyme activity. In addition, atomic force microscopy (AFM) and in vivo enzyme assays showed highly specific interactions of esterases with α-synuclein. Our results indicate that α-synuclein not only protects the enzyme activity of microbial esterases in vitro, but also can stabilize the active conformation of microbial esterases in vivo

  17. Esterase profile of Rhipicephalus (Boophilus) microplus populations collected from Northern India exhibiting varied susceptibility to deltamethrin.

    Science.gov (United States)

    Abdullah, Swaid; Yadav, C L; Vatsya, Stuti

    2012-11-01

    Rhipicephalus (Boophilus) microplus is an economically important ectoparasite of cattle. Chemical acaricides remain the most practical method for control of these pests. During past two decades there have been increasing reports of resistance development against synthetic pyrethroids in tick populations of this species throughout the world. A study was conducted to determine the level of susceptibility of R. (B.) microplus to deltamethrin collected from different geographical locations of northern India. LPT bioassay results revealed LC(50) values of deltamethrin ranging from 0.035 to 0.00037 % A.I. Esterase profile of the tick larval extracts using native PAGE, revealed 5 bands of esterase activity designated EST-5 to EST-1A. Inhibitory tests recognized EST-1, EST-2 and EST-3 as Acetylcholinesterases (AchEs), EST-4 and EST-5 as Carboxylesterases (CaEs). The band intensity varied between tick populations of various locations, being more intense in case of the resistant populations. An extra band of esterase activity (EST-1A) was obtained in larval extracts of ticks from 3 locations. This increased esterase activity may be involved in the resistance development in these tick populations. Acaricide resistance is a multi-factorial phenomenon, thus other causes of increased resistance like sodium channel mutation and reduced drug penetration (e.g. cuticle thickening) and behavioural changes (e.g. avoiding the pesticides) are to be tested in future in order to confirm the basic cause of the resistance development in these acaricide resistant tick populations.

  18. 21 CFR 173.140 - Esterase-lipase derived from Mucor miehei.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Esterase-lipase derived from Mucor miehei. 173.140 Section 173.140 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES...; genus, Mucor; species, miehei; variety Cooney et Emerson. (b) The strain of Mucor miehei var. Cooney et...

  19. Cloning, expression, purification, and characterization of a novel esterase from Lactobacillus plantarum.

    Science.gov (United States)

    Brod, Fábio Cristiano Angonesi; Vernal, Javier; Bertoldo, Jean Borges; Terenzi, Hernán; Arisi, Ana Carolina Maisonnave

    2010-03-01

    Lactobacillus plantarum is an important lactic acid bacterium, usually found as natural inhabitant of food, such as fermented vegetables and meat products. However, little information about lactic acid bacteria, especially concerning L. plantarum, as a source of useful enzymes has been reported. The aim of this study was to clone, express in Escherichia coli, purify, and characterize an esterase from L. plantarum ATCC 8014. The esterase gene (1014 bp) was amplified and cloned in pET14b expression vector to express a His(6)-tagged protein in E. coli. Recombinant L. plantarum esterase was purified by Ni-NTA resin, presenting an apparent molecular mass of about 38 kDa. It presented highest activity at pH 6.0 and 40 degrees C. Also, it presented preference for p-nitrophenyl butyrate, but hydrolyzed more efficiently p-nitrophenyl acetate. Besides, this study shows, for the first time, CD data about secondary structure of an esterase from L. plantarum.

  20. Esterase and protease activities of Bacillus spp. from afitin, iru and ...

    African Journals Online (AJOL)

    The electrophoretic profiles of fermented African locust bean protein (ALBP), using strains presenting the highest protease activities in casein agar, were analyzed by SDS-PAGE to select strains with good ability to be used as starter cultures. All the Bacillus spp. tested showed esterase activity against tributyrin with high ...

  1. Respective importance of protein folding and glycosylation in the thermal stability of recombinant feruloyl esterase A

    NARCIS (Netherlands)

    Benoit, Isabelle; Asther, Michèle; Sulzenbacher, Gerlind; Record, Eric; Marmuse, Laurence; Parsiegla, Goetz; Gimbert, Isabelle; Asther, Marcel; Bignon, Christophe

    2006-01-01

    The thermal stability of four molecular forms (native, refolded, glycosylated, non-glycosylated) of feruloyl esterase A (FAEA) was studied. From the most to the least thermo-resistant, the four molecular species ranked as follows: (i) glycosylated form produced native, (ii) non-glycosylated form

  2. Cloning, Characterization, Controlled Overexpression, and Inactivation of the Major Tributyrin Esterase Gene of Lactococcus lactis

    NARCIS (Netherlands)

    Fernández, Leonides; Beerthuyzen, Marke M.; Brown, Julie; Siezen, Roland J.; Coolbear, Tim; Holland, Ross; Kuipers, Oscar P.

    2000-01-01

    The gene encoding the major intracellular tributyrin esterase of Lactococcus lactis was cloned using degenerate DNA probes based on 19 known N-terminal amino acid residues of the purified enzyme. The gene, named estA, was sequenced and found to encode a protein of 258 amino acid residues. The

  3. Esterase isozymes patterns of grape vine (Vitis vinifera L. are altered in response to fungicide exposure

    Directory of Open Access Journals (Sweden)

    Gleice Ribeiro Orasmo

    2015-10-01

    Full Text Available Current analysis characterizes the effect of different fungicides often applied for pest control on a-and b-esterase patterns of four economically important table-wine grape cultivars (Italia, Rubi, Benitaka and Brasil of Vitis vinifera. The a- and b-esterase patterns in bud leaves of the cultivars were assessed by native PAGE analysis. Cabrio Top® compound inhibited Est-2, Est-5, Est-6, Est-7, Est-8, Est-9 and Est-10 carboxylesterases, whereas Est-4, Est-11, Est-12, Est-13, Est-14 acetylesterases and Est-16 carboxylesterase were detected as weakly stained bands. Carboxylesterases and acetylesterases were also detected as weakly stained bands when exposed to fungicides Orthocide 500®, Positron Duo® and Folicur PM®. No changes in a- and b-esterase patterns were reported when the vines were exposed to the fungicides Rovral SC®, Kumulus DF®, Curzate M®, Score® or Cuprogarb 500®. The evidence of functional changes in carboxylesterase and acetylesterase levels in current study is a warning to grape producers on the dangers inherent in the indiscriminate use of potent and modern fungicides extensively used in agriculture. The inhibition effect of fungicides on esterase isozyme molecules seems to be independent of the fungicide chemical.

  4. Usefulness of Leukocyte Esterase Test Versus Rapid Strep Test for Diagnosis of Acute Strep Pharyngitis

    Directory of Open Access Journals (Sweden)

    Kumara V. Nibhanipudi MD

    2015-08-01

    Full Text Available Objective: A study to compare the usage of throat swab testing for leukocyte esterase on a test strip(urine dip stick-multi stick to rapid strep test for rapid diagnosis of Group A Beta hemolytic streptococci in cases of acute pharyngitis in children. Hypothesis: The testing of throat swab for leukocyte esterase on test strip currently used for urine testing may be used to detect throat infection and might be as useful as rapid strep. Methods: All patients who come with a complaint of sore throat and fever were examined clinically for erythema of pharynx, tonsils and also for any exudates. Informed consent was obtained from the parents and assent from the subjects. 3 swabs were taken from pharyngo-tonsillar region, testing for culture, rapid strep & Leukocyte Esterase. Results: Total number is 100. Cultures 9(+; for rapid strep== 84(- and16 (+; For LE== 80(- and 20(+ Statistics: From data configuration Rapid Strep versus LE test don’t seem to be a random (independent assignment but extremely aligned. The Statistical results show rapid and LE show very agreeable results. Calculated Value of Chi Squared Exceeds Tabulated under 1 Degree Of Freedom (P<.0.0001 reject Null Hypothesis and Conclude Alternative Conclusions: Leukocyte esterase on throat swab is as useful as rapid strep test for rapid diagnosis of strep pharyngitis on test strip currently used for urine dip stick causing acute pharyngitis in children.

  5. Preliminary X-ray Study of Naproxen Esterase from Bacillus subtilis

    NARCIS (Netherlands)

    van der Laan, Jan; Teplyakov, A.V.; Lammers, A.A.; Dijkstra, B.W.

    1993-01-01

    Single crystals of naproxen esterase from Bacillus subtilis have been obtained from PEG6000 solutions at pH 8.0 by liquid-liquid diffusion while applying a temperature gradient from 4°C to room temperature over a period of four weeks. The crystals belong to the trigonal space group P3121 or P3221

  6. Exceptional thermal stability and organic solvent tolerance of an esterase expressed from a thermophilic host

    DEFF Research Database (Denmark)

    Mei, Yuxia; Peng, Nan; Zhao, Shumiao

    2012-01-01

    A protein expression system recently developed for the thermophilic crenarchaeon Sulfolobus islandicus was employed to produce recombinant protein for EstA, a thermophilic esterase encoded in the same organism. Large amounts of protein were readily obtained by an affinity protein purification...

  7. Engineering Saccharomyces cerevisiae to produce feruloyl esterase for the release of ferulic acid from switchgrass

    Science.gov (United States)

    The Aspergillus niger ferulic acid esterase gene (faeA) was cloned into Saccharomyces cerevisiae via a yeast expression vector, resulting in efficient expression and secretion of the enzyme in the medium. The recombinant enzyme was purified to homogeneity by anion-exchange and hydrophobic interactio...

  8. Isolation and identification of an esterase from a Mexican strain of Boophilus microplus (Acari: Ixodidae).

    Science.gov (United States)

    Pruett, J H; Guerrero, F D; Hernandez, R

    2002-10-01

    A strain of Mexican Boophilus microplus (Cz) collected near Coatzacoalcos, Veracruz, Mexico, exhibits a moderate, but significant, level of permethrin resistance. Unlike other highly permethrin resistant strains, the Cz strain does not have a mutation within the sodium channel gene that results in target-site insensitivity. However, the Cz strain possesses a substantial increase in general and permethrin esterase activity relative to highly permethrin resistant and control strains suggesting the involvement of a metabolic esterase(s) in the expression of permethrin resistance. We report the isolation of a 62.8 kDa protein from Cz strain larvae that we think is the esterase previously reported as Cz EST9. In addition, internal amino acid sequence data obtained from the 62.8 kDa protein suggest that it is the gene product of a previously reported B. microplus carboxylesterase cDNA. We propose that the 62.8 kDa protein (Cz EST9) has permethrin hydrolytic activity and as a result plays an important role in Cz strain resistance to permethrin.

  9. Esterase detoxification of acetylcholinesterase inhibitors using human liver samples in vitro

    Science.gov (United States)

    Organophosphate (OP) and N-methylcarbamate pesticides inhibit acetylcholinesterase (AChE), but differences in metabolism and detoxification can influence potency of these pesticides across and within species. Carboxylesterase (CaE) and A-esterase (paraoxonase, PON1) are consider...

  10. Molecular characterization of a novel family VIII esterase from burkholderia multivorans UWC10

    CSIR Research Space (South Africa)

    Rashamuse, KJ

    2007-02-01

    Full Text Available ----- 398 Fig. 2: Multiple sequence alignment of B. multivorans esterase EstBL (AAV97951) and other re- lated proteins: Lip1 from Pseudomonas sp. nov. 109 (CAA43847), Lip8 from Ps. aeruginosa LST-03 (BAD69792), LipZM4 from Zygomonas mobilis ZM4 (AAG42401...

  11. Cholesterol esterase inhibitory activity of bioactives from leaves of Mangifera indica L.

    Science.gov (United States)

    Gururaja, G M; Mundkinajeddu, Deepak; Dethe, Shekhar M; Sangli, Gopala K; Abhilash, K; Agarwal, Amit

    2014-01-01

    In the earlier studies, methanolic extract of Mangifera indica L leaf was exhibited hypocholesterol activity. However, the bioactive compounds responsible for the same are not reported so far. To isolate the bioactive compounds with hypocholesterol activity from the leaf extract using cholesterol esterase inhibition assay which can be used for the standardization of extract. The leaf methanolic extract of M. indica (Sindoora variety) was partitioned with ethyl acetate and chromatographed on silica gel to yield twelve fractions and the activity was monitored by using cholesterol esterase inhibition assay. Active fractions were re-chromatographed to yield individual compounds. A major compound mangiferin present in the extract was screened along with other varieties of mango leaves for cholesterol esterase inhibition assay. However, the result indicates that compounds other than mangiferin may be active in the extract. Invitro pancreatic cholesterol esterase inhibition assay was used for bioactivity guided fractionation (BAGF) to yield bioactive compound for standardization of extract. Bioactivity guided fractionation afford the active fraction containing 3b-taraxerol with an IC50 value of 0.86μg/ml. This study demonstrates that M. indica methanol extract of leaf have significant hypocholesterol activity which is standardized with 3b-taraxerol, a standardized extract for hypocholesterol activity resulted in development of dietary supplement from leaves of Mangifera indica.

  12. Enrichment of maize and triticale bran with recombinant Aspergillus tubingensis ferulic acid esterase

    CSIR Research Space (South Africa)

    Zwane, EN

    2017-03-01

    Full Text Available acid, in particular for the enrichment of food substrates. A recombinant Aspergillus tubingensis ferulic acid esterase Type A (FAEA) was expressed in Aspergillus niger D15#26 and purified with anion-exchange chromatography (3487 U/mg, Km = 0.43 mM, Kcat...

  13. Fitness differences due to allelic variation at Esterase-4 locus in ...

    Indian Academy of Sciences (India)

    KAVITA KRISHNAMOORTI

    2017-08-31

    Aug 31, 2017 ... 2011 Ovariectomy in grasshoppers increases somatic storage, but proportional allocation of ingested nutri- ents to somatic tissues is unchanged. Aging Cell 10, 972–979. Kojima K. and Yarbrough K. M. 1967 Frequency dependent selection at the esterase 6 locus in Drosophila melanogaster. Proc. Nat.

  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. 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...

  16. Molecular population genetics of the β-esterase gene cluster of ...

    Indian Academy of Sciences (India)

    However there are some 'footprints' of directional and balancing selection shaping specific distribution of nucleotide polymorphism within the cluster. Intergenic epistatic selection between Est-6 and Est-6 may play an important role in the evolution of the -esterase gene cluster preserving the putative pseudogene from ...

  17. Molecular population genetics of the β-esterase gene cluster of ...

    Indian Academy of Sciences (India)

    We suggest that the demographic history (bottleneck and admixture of genetically differentiated populations) is the major factor shaping the pattern of nucleotide polymorphism in the -esterase gene cluster. However there are some 'footprints' of directional and balancing selection shaping specific distribution of nucleotide ...

  18. Molecular population genetics of the β-esterase gene cluster of ...

    Indian Academy of Sciences (India)

    Unknown

    neutrality with recombination are significant for the β−esterase gene cluster in the non-African samples but not signi- ficant in the African one. We suggest ...... I. Viability studies. Genetics 102,. 467–483. Selva E. M., New L., Crouse G. F. and Lahue R. S. 1995 Mis- match correction acts as a barrier to homologous recombina-.

  19. Fragrance material review on acetyl cedrene.

    Science.gov (United States)

    Scognamiglio, J; Letizia, C S; Politano, V T; Api, A M

    2013-12-01

    A toxicologic and dermatologic review of acetyl cedrene when used as a fragrance ingredient is presented. Acetyl cedrene is a member of the fragrance structural group Alkyl Cyclic Ketones. The generic formula for this group can be represented as (R1)(R2)CO. These fragrances can be described as being composed of an alkyl, R1, and various substituted and bicyclic saturated or unsaturated cyclic hydrocarbons, R2, in which one of the rings may include up to 12 carbons. Alternatively, R2 may be a carbon bridge of C2-C4 carbon chain length between the ketone and cyclic hydrocarbon. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for acetyl cedrene were evaluated then summarized and includes physical properties, acute toxicity, skin irritation, mucous membrane (eye) irritation, skin sensitization, elicitation, phototoxicity, photoallergy, toxicokinetics, repeated dose, reproductive toxicity, and genotoxicity data. A safety assessment of the entire Alkyl Cyclic Ketones will be published simultaneously with this document; please refer to Belsito et al. (2013) (Belsito, D., Bickers, D., Bruze, M., Calow, P., Dagli, M., Fryer, A.D., Greim, H., Miyachi, Y., Saurat, J.H., Sipes, I.G., 2013. A Toxicologic and Dermatologic Assessment of Alkyl Cyclic Ketones When Used as Fragrance Ingredients. Submitted with this manuscript.) for an overall assessment of the safe use of this material and all Alkyl Cyclic Ketones in fragrances. Copyright © 2013. Published by Elsevier Ltd.

  20. Angioedema as the first presentation of B-cell non-Hodgkin lymphoma--an unusual case with normal C1 esterase inhibitor level: a case report.

    Science.gov (United States)

    Gunatilake, Sonali Sihindi Chapa; Wimalaratna, Harith

    2014-08-07

    Acquired angioedema is a rare but recognized manifestation of lymphoproliferative disorders due to deficiency in C1 esterase inhibitor. Normal level of C1 esterase inhibitor proteins in association with angioedema due to lymphoproliferative disease is a rare and an uncommon finding caused by antibodies produced from the underlying disease. Antibodies cause inactivation of C1 esterase inhibitor, thus resulting in C1 esterase inhibitor dysfunction despite of normal quantity of C1 esterase inhibitor. A 50-year-old Sri Lankan male presented with first episode of angioedema without any family history. Physical examination revealed mild pallor with swelling of tongue, lips and perioral region. On investigations, erythrocyte sedimentation rate was persistently high and bone marrow with immunohistochemistry revealed infiltration with B-cell type low grade non-Hodgkin lymphoma. Computed tomography scan of the chest and abdomen showed paratracheal and subcarinal lymphadenopathy and splenomegaly, with the findings being compatible with lymphoma. He had normal C1 esterase inhibitor protein level with reduced activity and low C1q, C4 levels indicating antibodies against C1 esterase inhibitor causing dysfunctional C1 esterase inhibitor. Adult onset angioedema should prompt physicians to suspect underlying lymphoproliferative disorder despite of C1 esterase inhibitor protein level being normal. Though uncommon, presence of antibodies against C1 esterase inhibitor secondary to lymphoproliferative disorder should be considered in the presence of normal C1 esterase inhibitor protein levels with low functional capacity in the background of acquired angioedema.

  1. Xylan synthesized by Irregular Xylem 14 (IRX14) maintains the structure of seed coat mucilage in Arabidopsis.

    Science.gov (United States)

    Hu, Ruibo; Li, Junling; Wang, Xiaoyu; Zhao, Xun; Yang, Xuanwen; Tang, Qi; He, Guo; Zhou, Gongke; Kong, Yingzhen

    2016-03-01

    During differentiation, the Arabidopsis seed coat epidermal cells synthesize and secrete large quantities of pectinaceous mucilage into the apoplast, which is then released to encapsulate the seed upon imbibition. In this study, we showed that mutation in Irregular Xylem 14 (IRX14) led to a mucilage cohesiveness defect due to a reduced xylan content. Expression of IRX14 was detected specifically in the seed coat epidermal cells, reaching peak expression at 13 days post-anthesis (DPA) when the accumulation of mucilage polysaccharides has ceased. Sectioning of the irx14-1 seed coat revealed no visible structural change in mucilage secretory cell morphology. Although the total amount of mucilage was comparable with the wild type (WT), the partition between water-soluble and adherent layers was significantly altered in irx14-1, with redistribution from the adherent layer to the water-soluble layer. The monosaccharide composition analysis revealed that xylose content was significantly reduced in irx14-1 water-soluble and adherent mucilage compared with the WT. The macromolecular characteristics of the water-soluble mucilage were modified in irx14-1 with a loss of the larger polymeric components. In accordance, glycome profiling and dot immunoblotting of seed mucilage using antibodies specific for rhamnogalacturonan I (RG I) and xylan confirmed the ultra-structural alterations in the irx14-1 mucilage. Meanwhile, the crystalline cellulose content was reduced in the irx14-1 mucilage. These results demonstrated that IRX14 was required for the biosynthesis of seed mucilage xylan, which plays an essential role in maintaining mucilage architecture potentially through altering the crystallization and organization of cellulose. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  2. Recombinant Bacillus amyloliquefaciens xylanase A expressed in Pichia pastoris and generation of xylooligosaccharides from xylans and wheat bran.

    Science.gov (United States)

    Liu, Ming-Qi; Huo, Wen-Kang; Xu, Xin; Weng, Xiao-Yan

    2017-12-01

    In this study, BaxA (GenBank: KM624029), which encodes the Bacillus amyloliquefaciens xylanase A (BaxA), was highly expressed in Pichia pastoris GS115 under the control of the AOX1 promoter. The recombinant xylanase, namely rePBaxA, was purified to homogeneity by using Ni-affinity resin and its molecular weight was 35.0kDa. The optimum temperature and pH of rePBaxA were 50°C and 5.0, respectively. The kinetic parameters Michaelis-Menten constant (K m ) and maximum reaction rate (V max ) of rePBaxA were 5.41mg/mL and 22.42μmol/min/mL, respectively. High-performance liquid chromatography results showed that after 6h of hydrolysis, rePBaxA released xylose-xylohexaose (X1-X6) mixture from beechwood and birchwood xylan, with xylobiose (X2) and xylotriose (X3) as the major products, respectively. The hydrolyates from oat spelt, wheat bran insoluble xylan and pretreated wheat bran by rePBaxA included X2-X6, with X6 having the highest concentration. The mode of action analysis revealed that rePBaxA was an endo-acting xylanase with transglycosylation activity. X2 might be the minimum oligomer hydrolyzed by rePBaxA. The pretreated wheat bran and wheat bran insoluble xylan could be directly hydrolyzed by rePBaxA. This study provided a basis for using agricultural waste by-products as substrates for manufacting value-added probiotics, namely, xylooligosaccharides. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Predominance of N-acetyl transferase 2 slow acetylator alleles in ...

    African Journals Online (AJOL)

    Blood was spotted on filter paper prior to drug administration for DNA extraction by chelex method. Standard nested PCR followed by restriction enzyme analysis with KpnI, TaqI, and BamHI for detection of polymorphisms in the NAT2 was performed. Allelic frequencies and acetylator phenotypes were compared between ...

  4. Organophosphorous biocides reduce tenacity and cellular viability but not esterase activities in a non-target prosobranch (limpet)

    International Nuclear Information System (INIS)

    Browne, Mark Anthony; Dissanayake, Awantha; Galloway, Tamara S.

    2015-01-01

    Detecting impacts of organophosphorus biocides (OP) is facilitated by analysing “biomarkers” – biological responses to environmental insults. Understanding is hampered by studying biomarkers in isolation at different levels of biological response and limited work on ecologically-important species. We tested the relevance of esterases as biomarkers of OP-exposure in limpets (Patella vulgata), abundant prosobranchs that structure the assemblages on rocky shores through their grazing. We characterized esterases in haemolymph and tissue, and quantified their dose-dependent inhibition by chlorfenvinphos (0.1–3.0 mM) in vitro. To determine whether esterases are useful biomarkers we exposed limpets to chlorfenvinphos (0–10 μg L −1 ). Despite reduced tenacity (ability to stick to a surface) and haemocyte-viability, esterases remained unaffected. Tenacity was reduced by >50% at 5 μg L −1 and by 95% at 10 μg L −1 , whilst haemocyte-viability was more sensitive with >40% reductions at concentrations of 0.5 μg L −1 and above. We discuss results in relation to linking sub-lethal and ecological impacts at contaminated sites. - Highlights: • We investigated if esterases are useful biomarkers of chlorfenvinphos-exposure. • Esterases in tissues of limpets (Patella vulgata) were characterized. • The dose-dependent inhibition of esterases by chlorfenvinphos was shown in vitro. • In vivo, tenacity and haemocyte-viability were reduced, but not esterase activities. - Organophosphorous biocides reduce tenacity and cellular viability but not esterase activities in the limpet, Patella vulgata

  5. Differential patterns of histone acetylation in inflammatory bowel diseases

    Directory of Open Access Journals (Sweden)

    Adcock Ian M

    2011-01-01

    Full Text Available Abstract Post-translational modifications of histones, particularly acetylation, are associated with the regulation of inflammatory gene expression. We used two animal models of inflammation of the bowel and biopsy samples from patients with Crohn's disease (CD to study the expression of acetylated histones (H 3 and 4 in inflamed mucosa. Acetylation of histone H4 was significantly elevated in the inflamed mucosa in the trinitrobenzene sulfonic acid model of colitis particularly on lysine residues (K 8 and 12 in contrast to non-inflamed tissue. In addition, acetylated H4 was localised to inflamed tissue and to Peyer's patches (PP in dextran sulfate sodium (DSS-treated rat models. Within the PP, H3 acetylation was detected in the mantle zone whereas H4 acetylation was seen in both the periphery and the germinal centre. Finally, acetylation of H4 was significantly upregulated in inflamed biopsies and PP from patients with CD. Enhanced acetylation of H4K5 and K16 was seen in the PP. These results demonstrate that histone acetylation is associated with inflammation and may provide a novel therapeutic target for mucosal inflammation.

  6. Solvent-Free Synthesis of Some1-Acetyl Pyrazoles

    Energy Technology Data Exchange (ETDEWEB)

    Thirunarayanan, Ganesamoorthy [Annamalai Univ., Tamil Nadu (India); Sekar, Krishnamoorthy Guna [National College, Tiruchirappalli (India)

    2013-10-15

    Some N-acetyl pyrazoles including 1-(3-(3,4-dichlorophenyl)-5-(substituted phenyl)-4,5-dihydro-{sup 1}H-pyrazole-1-yl) ethanones have been synthesised by solvent free cyclization cum acetylation of chalcones like substituted styryl 3,4-dichlorophenyl ketones using hydrazine hydrate and acetic anhydride in presence of catalytic amount of fly-ash: H{sub 2}SO{sub 4} catalyst. The yield of these N-acetyl pyrazole derivatives are more than 75%. The synthesised N-acetyl pyrazoline derivatives were characterized by their physical constants and spectral data.

  7. Extração e caracterização de xilanas de sabugos de milho Extraction and characterization of xylans from corncobs

    Directory of Open Access Journals (Sweden)

    Simone S. Silva

    1998-06-01

    Full Text Available Neste trabalho, duas frações de xilana, denominadas xilana A e xilana B, foram isoladas a partir de sabugos de milho através de três processos diferentes, combinando métodos de extração aquosa, remoção de lipídeos, deslignificação e extração alcalina. Os produtos obtidos durante os processos foram analisados por termogravimetria. A etapa de deslignificação foi responsável por uma acentuada degradação dos polímeros, evidenciada por queda de rendimento e resistência térmica. Os espectros obtidos no infravermelho evidenciaram a ausência de ácidos urônicos na cadeia polimérica. As viscosidades intrínsecas obtidas para a xilana A (56 mL/g e xilana B (75 mL/g associadas aos resultados do infravermelho sugerem um número maior de grupos substituintes, constituídos basicamente por resíduos de L-arabinose, para a xilana B.In this work, two fractions of xylan, named xylan A and xylan B, were isolated from corncobs through three different processes using aqueous extraction, lipid removal, delignification, and alkaline extraction. The products obtained during the processes were analysed by thermogravimetry. The delignification step was responsible for the occurrence of an accentuated polymer degradation, evidenced by yield and thermal resistance decrease. Infrared spectra indicated the absence of uronic acids in the polymeric chains. Intrinsic viscosities obtained for xylan A (56 mL/g and xylan B (75 mL/g, associated to the results from i.r. analysis, suggested a higher number of substituents, basically constituted of L-arabinose residues, in the case of xylan B.

  8. Reduced Wall Acetylation Proteins Play Vital and Distinct Roles in Cell Wall O-Acetylation in Arabidopsis

    DEFF Research Database (Denmark)

    Manabe, Yuzuki; Verhertbruggen, Yves; Gille, Sascha

    2013-01-01

    The Reduced Wall Acetylation (RWA) proteins are involved in cell wall acetylation in plants. Previously, we described a single mutant, rwa2, which has about 20% lower level of O-acetylation in leaf cell walls and no obvious growth or developmental phenotype. In this study, we generated double......, triple, and quadruple loss-of-function mutants of all four members of the RWA family in Arabidopsis (Arabidopsis thaliana). In contrast to rwa2, the triple and quadruple rwa mutants display severe growth phenotypes revealing the importance of wall acetylation for plant growth and development...

  9. Bacteroides intestinalis DSM 17393, a member of the human colonic microbiome, upregulates multiple endoxylanases during growth on xylan.

    Science.gov (United States)

    Wang, Kui; Pereira, Gabriel V; Cavalcante, Janaina J V; Zhang, Meiling; Mackie, Roderick; Cann, Isaac

    2016-09-29

    Many human diets contain arabinoxylan, and the ease of genome sequencing coupled with reduced cost have led to unraveling the arsenal of genes utilized by the colonic Bacteroidetes to depolymerize this polysaccharide. The colonic Bacteroidetes with potential to ferment arabinoxylans include Bacteroides intestinalis. In this study, we analyzed the hydrolytic activities of members of a xylan degradation cluster encoded on the genome of Bacteroides intestinalis DSM 17393. Here, it is demonstrated that a cocktail of the xylanolytic enzymes completely hydrolyze arabinoxylans found in human diets. We show that this bacterium and relatives have evolved and secrete a unique bifunctional endoxylanase/arabinofuranosidase in the same polypeptide. The bifunctional enzyme and other secreted enzymes attack the polysaccharides extracellularly to remove the side-chains, exposing the xylan backbone for cleavage to xylo-oligosaccharides and xylose. These end products are transported into the cell where a β-xylosidase cleaves the oligosaccharides to fermentable sugars. While our experiments focused on B. intestinalis, it is likely that the extracellular enzymes also release nutrients to members of the colonic microbial community that practice cross-feeding. The presence of the genes characterized in this study in other colonic Bacteroidetes suggests a conserved strategy for energy acquisition from arabinoxylan, a component of human diets.

  10. Direct and efficient xylitol production from xylan by Saccharomyces cerevisiae through transcriptional level and fermentation processing optimizations.

    Science.gov (United States)

    Li, Zhe; Qu, Hongnan; Li, Chun; Zhou, Xiaohong

    2013-12-01

    In this study, four engineered Saccharomyces cerevisiae carrying xylanase, β-xylosidase and xylose reductase genes by different transcriptional regulations were constructed to directly convert xylan to xylitol. According to the results, the high-copy number plasmid required a rigid selection for promoter characteristics, on the contrast, the selection of promoters could be more flexible for low-copy number plasmid. For cell growth and xylitol production, glucose and galactose were found more efficient than other sugars. The semi-aerobic condition and feeding of co-substrates were taken to improve the yield of xylitol. It was found that the strain containing high-copy number plasmid had the highest xylitol yield, but it was sensitive to the change of fermentation. However, the strain carrying low-copy number plasmid was more adaptable to different processes. By optimization of the transcriptional regulation and fermentation processes, the xylitol concentration could be increased of 1.7 folds and the yield was 0.71 g xylitol/g xylan. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Dampak Hipoksia Sistemik terhadap Malondialdehida, Glial Fibrillary Acidic Protein dan Aktivitas Asetilkolin Esterase Otak Tikus

    Directory of Open Access Journals (Sweden)

    Andriani Andriani

    2016-09-01

    Full Text Available Hipoksia sistemik menyebabkan berkurangnya oksigen dan energi di otak sehingga memicupenglepasan neurotransmiter asetilkolin, meningkatkan radikal bebas dan glial fibrillary acidic protein (GFAPyang berfungsi menjaga kekuatan membran. Tujuan penelitian untuk melihat gambaran adaptasi otak padahipoksia sistemik terhadap fungsi asetilkolin esterase, kerusakan membran sel neuron dan astrosit. Penelitiandilakukan di Laboratorium Biokimia & Biologi Molekuler FK Universitas Indonesia, pada tahun 2013.Penelitian ekperimental ini menggunakan hewan coba tikus spraque dawley yang diinduksi hipoksia sistemikyang diambil jaringan otak bagian korteks dan plasma tikus. Kelompok tikus terdiri atas kelompok kontrol,kelompok perlakuan induksi hipoksia hari ke-1, 3 hari, 5 hari dan hari ke-7. Parameter yang diukur adalahkadar malondialdehida (MDA otak dan plasma, aktivitas spesifik enzim AChE jaringan otak serta kadar GFAPjaringan otak. Hasil menunjukkan bahwa hipoksia sistemik tidak meningkatkankadar MDA otak dan plasma.Induksi hipoksia sistemik meningkatkan aktivitas spesifik enzim AChE dan kadar GFAP jaringan otak secarabermakna. Pada plasma tidak terjadi peningkatan kadar GFAP. Hipoksia sistemik selama hari ke-7 belummenyebabkan kerusakan oksidatif, namun memperlihatkan peningkatan aktivitas AChe dan adaptasi astrositmelalui peningkatan GFAP. Kata kunci: hipoksia, astrosit, glial fibrillary acidic protein, malondialdehida, asetilkolin esterase   Systemic Hypoxia Effect on Rat Brain Malondialdehyde, Glial FibrillaryAcidic Protein, and Acetylcholine Esterase Activity Abstract Sistemic hypoxia causes lack of oxygen and energy in brain that trigger the release of acetylcholine,free radical and Glial fibrillary acidic protein (GFAP, a specific protein in astrocyte cells that act to strenghtenastrocite membrane. The aim of the research was to evaluate the damages of brain in systemic hypoxiathrough activity of acetylcholine esterase, neuron and astrocyte membran

  12. 21 CFR 172.372 - N-Acetyl-L-methionine.

    Science.gov (United States)

    2010-04-01

    ... HUMAN CONSUMPTION (CONTINUED) FOOD ADDITIVES PERMITTED FOR DIRECT ADDITION TO FOOD FOR HUMAN CONSUMPTION Special Dietary and Nutritional Additives § 172.372 N-Acetyl-L-methionine. The food additive N-acetyl-L... in paragraph (d) of this section. The minimum amount of the additive to achieve the desired effect...

  13. Influence of acetylation on the physicochemical properties of ...

    African Journals Online (AJOL)

    oni josiah

    Freeze-thaw stability of gels from composited starches was greatly enhanced as lower volume of exudate was generated from the acetylated starches in all the freeze-thaw cycles. The findings in this study have the potential of creating awareness among the food industry with respect to acetylated starch production from both.

  14. Influence of acetylation on the physicochemical properties of ...

    African Journals Online (AJOL)

    The study investigates the effect of acetylation on the physicochemical properties of composited starches from sweet potato and water yam. Starch was respectively isolated from both sources, dried and subjected to acetylation at different combination. The result shows that the modified starches were of low percentage of ...

  15. Discovery and characterization of Ku acetylation in Mycobacterium smegmatis.

    Science.gov (United States)

    Zhou, Ying; Chen, Tao; Zhou, Lin; Fleming, Joy; Deng, Jiaoyu; Wang, Xude; Wang, Liwei; Wang, Yingying; Zhang, Xiaoli; Wei, Wenjing; Bi, Lijun

    2015-03-01

    Lysine acetylation is an important post-translational modification and is known to regulate many eukaryotic cellular processes. Little, however, is known about acetylated proteins in prokaryotes. Here, using immunoblotting, mass spectrometry and mutagenesis studies, we investigate the acetylation dynamics of the DNA repair protein Ku and its relationship with the deacetylase protein Sir2 and the non-homologous end joining (NHEJ) pathway in Mycobacterium smegmatis. We report that acetylation of Ku increases with growth, while NHEJ activity decreases, providing support for the hypothesis that acetylation of Ku may be involved in the DNA damage response in bacteria. Ku has multiple lysine sites. Our results indicate that K29 is an important acetylation site and that deficiency of Sir2 or mutation of K29 affects the quantity of Ku and its acetylation dynamics. Our findings expand knowledge of acetylation targets in prokaryotes and indicate a new direction for further research on bacterial DNA repair mechanisms. © FEMS 2014. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  16. The kinetics of the acetylation of gelatinised potato starch

    NARCIS (Netherlands)

    de Graaf, R.A.; Broekroelofs, G.A.; Janssen, L.P.B.M.; Beenackers, A.A C M

    1995-01-01

    The reaction rates, in the base-catalysed acetylation of gelatinised aqueous starch (4 wt%), by vinylacetate (ViAc), were investigated in a semibatch reactor at temperatures ranging from 20 to 50 degrees C. The desired starch acetylation reaction is accompanied by an undesired parallel

  17. Efficient acetylation of primary amines and amino acids in ...

    Indian Academy of Sciences (India)

    The present methodology illustrates the efficient acetylation of primary amines and amino acids in brine solution by means of acetyl chloride under weakly basic condition in the presence of sodium acetate and/or triethyl amine followed by trituration with aqueous saturated bicarbonate solution. This effort represents the first ...

  18. Efficient acetylation of primary amines and amino acids in ...

    Indian Academy of Sciences (India)

    This effort represents the first efficient use of this most reactive but cheap acetylating agent to acetylate amines in excellent yields in aqueous medium. This is a potentially useful green chemical transformation where reaction takes place in environment-friendly brine solution leading to easy work-up and isolation of the ...

  19. Direct Kinetic Evidence for the Formation of an Acylpyridinium Intermediate in Synthetic p-Nitrophenyl Esterase-Catalyzed Hydrolysis Reactions

    National Research Council Canada - National Science Library

    Wang, Guang-Jia

    1996-01-01

    .... The deacylation rate was also found to exhibit a maximum for the same substrate 2 (n=6). These results are similar to those previously reported with cholesterol esterase as catalyst for the same hydrolysis reaction...

  20. Inactive methyl indole-3-acetic acid ester can be hydrolyzed and activated by several esterases belonging to the AtMES esterase family of Arabidopsis.

    Science.gov (United States)

    Yang, Yue; Xu, Richard; Ma, Choong-Je; Vlot, A Corina; Klessig, Daniel F; Pichersky, Eran

    2008-07-01

    The plant hormone auxin (indole-3-acetic acid [IAA]) is found both free and conjugated to a variety of carbohydrates, amino acids, and peptides. We have recently shown that IAA could be converted to its methyl ester (MeIAA) by the Arabidopsis (Arabidopsis thaliana) enzyme IAA carboxyl methyltransferase 1. However, the presence and function of MeIAA in vivo remains unclear. Recently, it has been shown that the tobacco (Nicotiana tabacum) protein SABP2 (salicylic acid binding protein 2) hydrolyzes methyl salicylate to salicylic acid. There are 20 homologs of SABP2 in the genome of Arabidopsis, which we have named AtMES (for methyl esterases). We tested 15 of the proteins encoded by these genes in biochemical assays with various substrates and identified several candidate MeIAA esterases that could hydrolyze MeIAA. MeIAA, like IAA, exerts inhibitory activity on the growth of wild-type roots when applied exogenously. However, the roots of Arabidopsis plants carrying T-DNA insertions in the putative MeIAA esterase gene AtMES17 (At3g10870) displayed significantly decreased sensitivity to MeIAA compared with wild-type roots while remaining as sensitive to free IAA as wild-type roots. Incubating seedlings in the presence of [(14)C]MeIAA for 30 min revealed that mes17 mutants hydrolyzed only 40% of the [(14)C]MeIAA taken up by plants, whereas wild-type plants hydrolyzed 100% of absorbed [(14)C]MeIAA. Roots of Arabidopsis plants overexpressing AtMES17 showed increased sensitivity to MeIAA but not to IAA. Additionally, mes17 plants have longer hypocotyls and display increased expression of the auxin-responsive DR5:beta-glucuronidase reporter gene, suggesting a perturbation in IAA homeostasis and/or transport. mes17-1/axr1-3 double mutant plants have the same phenotype as axr1-3, suggesting MES17 acts upstream of AXR1. The protein encoded by AtMES17 had a K(m) value of 13 microm and a K(cat) value of 0.18 s(-1) for MeIAA. AtMES17 was expressed at the highest levels in shoot

  1. Using a simple HPLC approach to identify the enzymatic products of UTL-5g, a small molecule TNF-α inhibitor, from porcine esterase and from rabbit esterase.

    Science.gov (United States)

    Swartz, Kenneth; Zhang, Yiguan; Valeriote, Frederick; Chen, Ben; Shaw, Jiajiu

    2013-12-01

    UTL-5g is a novel small-molecule chemoprotector that lowers hepatotoxicity, nephrotoxicity, and myelotoxicity induced by cisplatin through TNF-α inhibition among other factors. As a prelude to investigating the metabolites of UTL-5g, we set out to identify the enzymatic products of UTL-5g under the treatment of both porcine liver esterase (PLE) and rabbit liver esterase (RLE). First, a number of mixtures made by UTL-5g and PLE were incubated at 25°C. At predetermined time points, individual samples were quenched by acetonitrile, vortexed, and centrifuged. The supernatants were then analyzed by reversed-phase HPLC (using a C18 column). The retention times and UV/vis spectra of individual peaks were compared to those of UTL-5g and its two postulated enzymatic products; thus the enzymatic products of UTL-5g were tentatively identified. Secondly, a different HPLC method (providing different retentions times) was used to cross-check and to confirm the identities of the two enzymatic products. Based on the observations, it was concluded that under the treatment of PLE, the major enzymatic products of UTL-5g were 5-methyliosxazole-3-carboxylic acid (ISOX) and 2,4-dichloroaniline (DCA). Treatment of UTL-5g by RLE also provided the same enzymatic products of UTL-5g from esterase. These results indicate that the peptide bond in UTL-5g was cleaved by PLE/RLE. Michaelis-Menten kinetics showed that the Km values of UTL-5g were 2.07mM with PLE and 0.37mM with RLE indicating that UTL-5g had a higher affinity with RLE. In summary, by a simple HPLC approach, we have concluded that the peptide bond in UTL-5g was cleaved by esterase from either porcine liver or rabbit liver in vitro and afforded DCA (at a mole ratio of 1:1) and ISOX. However, further studies are needed in order to determine whether UTL-5g is metabolized by microsomal enzymes to produce ISOX and DCA. Copyright © 2013 Elsevier B.V. All rights reserved.

  2. Antemortem stress regulates protein acetylation and glycolysis in postmortem muscle.

    Science.gov (United States)

    Li, Zhongwen; Li, Xin; Wang, Zhenyu; Shen, Qingwu W; Zhang, Dequan

    2016-07-01

    Although exhaustive research has established that preslaughter stress is a major factor contributing to pale, soft, exudative (PSE) meat, questions remain regarding the biochemistry of postmortem glycolysis. In this study, the influence of preslaughter stress on protein acetylation in relationship to glycolysis was studied. The data show that antemortem swimming significantly enhanced glycolysis and the total acetylated proteins in postmortem longissimus dorsi (LD) muscle of mice. Inhibition of protein acetylation by histone acetyltransferase (HAT) inhibitors eliminated stress induced increase in glycolysis. Inversely, antemortem injection of histone deacetylase (HDAC) inhibitors, trichostatin A (TSA) and nicotinamide (NAM), further increased protein acetylation early postmortem and the glycolysis. These data provide new insight into the biochemistry of postmortem glycolysis by showing that protein acetylation regulates glycolysis, which may participate in the regulation of preslaughter stress on glycolysis in postmortem muscle. Copyright © 2016. Published by Elsevier Ltd.

  3. Unchanged acetylation of isoniazid by alcohol intake

    DEFF Research Database (Denmark)

    Wilcke, J T R; Døssing, M; Angelo, H R

    2004-01-01

    no impact on the conversion of INH to its metabolite acetylisoniazid, which is catalysed by the enzyme N-acetyltranferase. Accordingly, a metabolic effect of acute alcohol intake on INH metabolism probably contributes little to the therapeutic failure of anti-tuberculosis treatment among alcoholics.......SETTING: In 10 healthy subjects, the influence of acute alcohol intake on the pharmacokinetics of isoniazid (INH) was studied. OBJECTIVE: To test the hypothesis that alcohol increases the conversion of INH by acetylation into its metabolite acetylisoniazid. DESIGN: In a crossover design, an oral...... dose of 300 mg INH was administered on 2 separate days, 14 days apart, with or without alcohol to a serum alcohol of about 21 mmol/l (1 g/l) maintained for 12 h. RESULTS: Neither the metabolism of INH nor that of acetylisoniazid was changed by acute alcohol intake. CONCLUSION: Acute alcohol intake has...

  4. Unchanged acetylation of isoniazid by alcohol intake

    DEFF Research Database (Denmark)

    Wilcke, J T R; Døssing, M; Angelo, H R

    2004-01-01

    SETTING: In 10 healthy subjects, the influence of acute alcohol intake on the pharmacokinetics of isoniazid (INH) was studied. OBJECTIVE: To test the hypothesis that alcohol increases the conversion of INH by acetylation into its metabolite acetylisoniazid. DESIGN: In a crossover design, an oral...... dose of 300 mg INH was administered on 2 separate days, 14 days apart, with or without alcohol to a serum alcohol of about 21 mmol/l (1 g/l) maintained for 12 h. RESULTS: Neither the metabolism of INH nor that of acetylisoniazid was changed by acute alcohol intake. CONCLUSION: Acute alcohol intake has...... no impact on the conversion of INH to its metabolite acetylisoniazid, which is catalysed by the enzyme N-acetyltranferase. Accordingly, a metabolic effect of acute alcohol intake on INH metabolism probably contributes little to the therapeutic failure of anti-tuberculosis treatment among alcoholics....

  5. The biology of lysine acetylation integrates transcriptional programming and metabolism

    Directory of Open Access Journals (Sweden)

    Mujtaba Shiraz

    2011-03-01

    Full Text Available Abstract The biochemical landscape of lysine acetylation has expanded from a small number of proteins in the nucleus to a multitude of proteins in the cytoplasm. Since the first report confirming acetylation of the tumor suppressor protein p53 by a lysine acetyltransferase (KAT, there has been a surge in the identification of new, non-histone targets of KATs. Added to the known substrates of KATs are metabolic enzymes, cytoskeletal proteins, molecular chaperones, ribosomal proteins and nuclear import factors. Emerging studies demonstrate that no fewer than 2000 proteins in any particular cell type may undergo lysine acetylation. As described in this review, our analyses of cellular acetylated proteins using DAVID 6.7 bioinformatics resources have facilitated organization of acetylated proteins into functional clusters integral to cell signaling, the stress response, proteolysis, apoptosis, metabolism, and neuronal development. In addition, these clusters also depict association of acetylated proteins with human diseases. These findings not only support lysine acetylation as a widespread cellular phenomenon, but also impel questions to clarify the underlying molecular and cellular mechanisms governing target selectivity by KATs. Present challenges are to understand the molecular basis for the overlapping roles of KAT-containing co-activators, to differentiate between global versus dynamic acetylation marks, and to elucidate the physiological roles of acetylated proteins in biochemical pathways. In addition to discussing the cellular 'acetylome', a focus of this work is to present the widespread and dynamic nature of lysine acetylation and highlight the nexus that exists between epigenetic-directed transcriptional regulation and metabolism.

  6. Extracts of Edible Plants Inhibit Pancreatic Lipase, Cholesterol Esterase and Cholesterol Micellization, and Bind Bile Acids

    Directory of Open Access Journals (Sweden)

    Julnaryn Intrawangso

    2012-01-01

    Full Text Available The application of edible plants with more effective ability to inhibit fat digestion and absorption has recently been explored for possible treatment of hyperlipidaemia. The aim of the present study is to investigate the effect of nine edible plants on the inhibition of pancreatic lipase and pancreatic cholesterol esterase activities, as well as the inhibition of cholesterol micelle formation, and bile acid binding. Our findings have shown strong pancreatic lipase inhibitory activity and the inhibition of cholesterol micellization by mulberry leaf extract. Safflower extract was the most potent inhibitor of pancreatic cholesterol esterase. In addition, cat’s whiskers and safflower extracts had a potent bile acid binding activity. It is suggested that a daily intake of these edible plants may delay postprandial hypertriacylglycerolaemia and hypercholesterolaemia, and therefore may be applied for the prevention and treatment of hyperlipidaemia.

  7. Genetic diversity analysis of Capsicum spp germplasm bank accessions based on α/β-esterase polymorphism.

    Science.gov (United States)

    Monteiro, E R; Bronzato, A R; Orasmo, G R; Lopes, A C A; Gomes, R L F; Mangolin, C A; Machado, M F P S

    2013-04-12

    Genetic diversity and structure were analyzed in 10 accessions belonging to Banco Ativo de Germoplasma de Capsicum located at Federal University of Piauí in northwestern Brazil that receives pepper samples grown in community gardens in various regions and Brazilian states. Selections were made from seeds of C. chinense (4 accessions), C. annuum (5 accessions), and C. baccatum (1 accession). Samples consisting of leaves were collected from 4-10 plants of each accession (a total of 85 plants). Native polyacrylamide gel electrophoresis was used to identify α- and β-esterase polymorphisms. Polymorphism was clearly detected in 5 loci. Sixteen alleles were found at 5 α/β-esterase loci of the three Capsicum species. In the C. chinense samples, the highest HO and HE values were 0.3625 and 0.4395, respectively, whereas in C. annuum samples, HO and HE values were 0.2980 and 0.3310, respectively; the estimated HO and HE values in C. chinense samples were higher than those detected in C. annuum samples. A deficit of homozygous individuals was found in C. chinense (FIS = -0.6978) and C. annuum (FIS = 0.7750). Genetic differentiation between C. chinense and C. annuum at these loci was high (FST = 0.1867) indicating that C. chinense and C. annuum are genetically structured species for α/β- esterase isozymes. The esterase analysis showed high genetic diversity among the C. chinense and C. annuum samples and very high genetic differentiation (FST = 0.6321) among the C. chinense and C. annuum samples and the C. baccatum accession.

  8. A chlorogenic acid esterase with a unique substrate specificity from Ustilago maydis.

    Science.gov (United States)

    Nieter, Annabel; Haase-Aschoff, Paul; Kelle, Sebastian; Linke, Diana; Krings, Ulrich; Popper, Lutz; Berger, Ralf G

    2015-03-01

    An extracellular chlorogenic acid esterase from Ustilago maydis (UmChlE) was purified to homogeneity by using three separation steps, including anion-exchange chromatography on a Q Sepharose FF column, preparative isoelectric focusing (IEF), and, finally, a combination of affinity chromatography and hydrophobic interaction chromatography on polyamide. SDS-PAGE analysis suggested a monomeric protein of ∼71 kDa. The purified enzyme showed maximal activity at pH 7.5 and at 37°C and was active over a wide pH range (3.5 to 9.5). Previously described chlorogenic acid esterases exhibited a comparable affinity for chlorogenic acid, but the enzyme from Ustilago was also active on typical feruloyl esterase substrates. Kinetic constants for chlorogenic acid, methyl p-coumarate, methyl caffeate, and methyl ferulate were as follows: Km values of 19.6 μM, 64.1 μM, 72.5 μM, and 101.8 μM, respectively, and kcat/Km values of 25.83 mM(-1) s(-1), 7.63 mM(-1) s(-1), 3.83 mM(-1) s(-1) and 3.75 mM(-1) s(-1), respectively. UmChlE released ferulic, p-coumaric, and caffeic acids from natural substrates such as destarched wheat bran (DSWB) and coffee pulp (CP), confirming activity on complex plant biomass. The full-length gene encoding UmChlE consisted of 1,758 bp, corresponding to a protein of 585 amino acids, and was functionally produced in Pichia pastoris GS115. Sequence alignments with annotated chlorogenic acid and feruloyl esterases underlined the uniqueness of this enzyme. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  9. Biochemical characterization of an enantioselective esterase from Brevundimonas sp. LY-2.

    Science.gov (United States)

    Zhang, Jing; Zhao, Mengjun; Yu, Die; Yin, Jingang; Zhang, Hao; Huang, Xing

    2017-06-19

    Lactofen, a member of the diphenylether herbicides, has high activity and is commonly used to control broadleaf weeds. As a post-emergent herbicide, it is directly released to the environment, and easily caused the pollution. This herbicide is degraded in soil mainly by microbial activity, but the functional enzyme involved in the biodegradation of lactofen is still not clear now. A novel esterase gene lacH, involved in the degradation of lactofen, was cloned from the strain Brevundimonas sp. LY-2. The gene contained an open reading frame of 921 bp, and a putative signal peptide at the N-terminal was identified with the most likely cleavage site between Ala 28 and Ala 29. The encoded protein, LacH, could catalyze the hydrolysis of lactofen to form acifluorfen. Phylogenetic analysis showed that LacH belong to family V of bacterial lipolytic enzymes. Biochemical characterization analysis showed that LacH was a neutral esterase with an optimal pH of 7.0 and an optimal temperature of 40 °C toward lactofen. Besides, the activity of LacH was strongly inhibited by Hg 2+ and Zn 2+ . LacH preferred short chain p-nitrophenyl esters (C 2 -C 6 ), exhibited maximum activity toward p-nitrophenyl acetate. Furthermore, the enantioselectivity of LacH during lactofen hydrolysis was also studied, and the results show that R-(-)-lactofen was degraded faster than S-(+)-lactofen, indicating the occurrence of enantioselectivity in the enzymatic reaction. Our studies characterized a novel esterase involved in the biodegradation of diphenylether herbicide lactofen. The esterase showed enantioselectivity during lactofen degradation, which revealed the occurrence of enzyme-mediated enantioselective degradation of chiral herbicides.

  10. Biochemical Characterization of a Family 15 Carbohydrate Esterase from a Bacterial Marine Arctic Metagenome.

    Directory of Open Access Journals (Sweden)

    Concetta De Santi

    Full Text Available The glucuronoyl esterase enzymes of wood-degrading fungi (Carbohydrate Esterase family 15; CE15 form part of the hemicellulolytic and cellulolytic enzyme systems that break down plant biomass, and have possible applications in biotechnology. Homologous enzymes are predicted in the genomes of several bacteria, however these have been much less studied than their fungal counterparts. Here we describe the recombinant production and biochemical characterization of a bacterial CE15 enzyme denoted MZ0003, which was identified by in silico screening of a prokaryotic metagenome library derived from marine Arctic sediment. MZ0003 has high similarity to several uncharacterized gene products of polysaccharide-degrading bacterial species, and phylogenetic analysis indicates a deep evolutionary split between these CE15s and fungal homologs.MZ0003 appears to differ from previously-studied CE15s in some aspects. Some glucuronoyl esterase activity could be measured by qualitative thin-layer chromatography which confirms its assignment as a CE15, however MZ0003 can also hydrolyze a range of other esters, including p-nitrophenyl acetate, which is not acted upon by some fungal homologs. The structure of MZ0003 also appears to differ as it is predicted to have several large loop regions that are absent in previously studied CE15s, and a combination of homology-based modelling and site-directed mutagenesis indicate its catalytic residues deviate from the conserved Ser-His-Glu triad of many fungal CE15s. Taken together, these results indicate that potentially unexplored diversity exists among bacterial CE15s, and this may be accessed by investigation of the microbial metagenome. The combination of low activity on typical glucuronoyl esterase substrates, and the lack of glucuronic acid esters in the marine environment suggest that the physiological substrate of MZ0003 and its homologs is likely to be different from that of related fungal enzymes.

  11. Dipstik leukosit esterase untuk diagnosis servisitis mukopurulenta Kajian pada wanita pekerja seks

    OpenAIRE

    Meita Dewayani, Meita Dewayani

    2015-01-01

    Mucopurulent cervicitis (MPC) is an endocervical inflammation that causes a variety of complications, including infertility. The diagnosis of MPC is usually confirmed by counting the number of leucocytes using a Gram stain of an endocervical swab. This method requires trained personnel, microscopic equipment and time to read the result, so the detection of MPC in primary health care setting is often difficult. The leucocyte esterase dipstick (LED) is an easy and rapid method designed for dete...

  12. Interactions between resin monomers and commercial composite resins with human saliva derived esterases.

    Science.gov (United States)

    Jaffer, F; Finer, Y; Santerre, J P

    2002-04-01

    Cholesterol esterase (CE) and pseudocholinesterase (PCE) have been reported to degrade commercial and model composite resins containing bisphenylglycidyl dimethacrylate (BisGMA), triethylene glycol dimethacrylate (TEGDMA) or the latter in combination with urethane modified BisGMA monomer systems. In addition, human saliva has been shown to contain esterase like activities similar to CE and PCE. Hence, it was the aim of the current study to determine to what extent human saliva could degrade two common commercial composite resins (Z250 from 3M Inc. and Spectrum TPH from L.D. Caulk) which contain the above monomer systems. Saliva samples from different volunteers were collected, processed, pooled, and freeze-dried. TEGDMA and BisGMA monomers were incubated with human saliva derived esterase activity (HSDEA) and their respective hydrolysis was monitored using high performance liquid chromatography (HPLC). Both monomers were completely hydrolyzed within 25 h by HSDEA. Photopolymerized composites were incubated with buffer or human saliva (pH 7.0 and 37 C) for 2, 8 and 16 days. The incubation solutions were analyzed using HPLC and mass spectrometry. Surface morphology characterization was carried out using scanning electron microscopy. Upon biodegradation, the Z250 composite yielded higher amounts of BisGMA and TEGDMA related products relative to the TPH composite. However, there were higher amounts of ethoxylated bis-phenol A released from the TPH material. In terms of total mass of products released, human saliva demonstrated a greater ability to degrade Z250. In summary, HSDEA has been shown to contain esterase activities that can readily catalyze the biodegradation of current commercial composite resins.

  13. EVALUATION OF ESTERASE POLYMORPHISMS IN MATURE SEEDS OF RADISH (RAPHANUS SATIVUS L. ACCESSIONS OF VIR COLLECTION

    Directory of Open Access Journals (Sweden)

    A. S. Rudakova

    2017-01-01

    Full Text Available A biochemical evaluation of 25 radish accessions (Raphanus sativus L. on esterase isozymes of mature seeds has been carried out. The results of the experiments showed a wide range of diversity among the genotypes based on electrophoretic zones of esterase isoenzymes. The revealed isoenzyme complex of esterases was represented by eight isoforms with molecular weights from 37.7 kD to 57.6 kD. All accessions were divided into 13 electrophoretic zymotypes, differing from each other by the presence or absence of definite zones. The most often observed electrophoretic zymo-type is Gr. 1, which includes 24% of the total number of accessions evaluated. There are 8 zymotypes (Gr. 6 Gr. 13 with a frequency of occurrence 4%. Three groups (Gr. 2 – Gr. 4 had the same frequency of occurrence – 12%. Zimotype of Gr. 5 containes the maximum number of zones – 8. 2 zimotypes – Gr. 3 and Gr. 12 had the smallest number of 4 zones. Two zones of esterases – zones 7 and 8 (Мr 39.7кD and Мr 37.7 kD, respectively were monomorphic. The remaining six zones were polymorphic, i.e. could be absent in some zimotypes. The frequency of occurrence of each zone in different zymotypes has varied from 6.58% to 17.11%. As results of this research the accessions that were selected can become the most promising parent forms for future genetic and selection studies of this culture.

  14. Enhanced biosurfactant production through cloning of three genes and role of esterase in biosurfactant release

    OpenAIRE

    Sekhon, Kamaljeet Kaur; Khanna, Sunil; Cameotra, Swaranjit Singh

    2011-01-01

    Abstract Background Biosurfactants have been reported to utilize a number of immiscible substrates and thereby facilitate the biodegradation of panoply of polyaromatic hydrocarbons. Olive oil is one such carbon source which has been explored by many researchers. However, studying the concomitant production of biosurfactant and esterase enzyme in the presence of olive oil in the Bacillus species and its recombinants is a relatively novel approach. Results Bacillus species isolated from endosul...

  15. A Novel Halotolerant Thermoalkaliphilic Esterase from Marine Bacterium Erythrobacter seohaensis SW-135

    Directory of Open Access Journals (Sweden)

    Ying-Yi Huo

    2017-11-01

    Full Text Available A novel esterase gene, e69, was cloned from Erythrobacter seohaensis SW-135, which was isolated from a tidal flat sediment of the Yellow Sea in Korea. This gene is 825 bp in length and codes for a 29.54 kDa protein containing 274 amino acids. Phylogenetic analysis showed that E69 is a new member of the bacterial lipolytic enzyme family IV. This enzyme exhibited the highest level of activity toward p-nitrophenyl (NP butyrate but little or no activity toward the other p-NP esters tested. The optimum temperature and pH of the catalytic activity of E69 were 60°C and pH 10.5, respectively. The enzyme exhibited stable activity over a wide range of alkaline pH values (7.5–9.5. In addition, E69 was found to be a halotolerant esterase as it exhibited the highest hydrolytic activity in the presence of 0.5 M NaCl and was still active in the presence of 3 M NaCl. Moreover, it possessed some degree of tolerance to Triton X-100 and several organic solvents. Through homology modeling and comparison with other esterases, it was suggested that the absence of the cap domain and its narrow substrate-binding pocket might be responsible for its narrow substrate specificity. Sequence and structural analysis results suggested that its high ratio of negatively to positively charged residues, large hydrophobic surface area, and negative electrostatic potential on the surface may be responsible for its alkaline adaptation. The results of this study provide insight into marine alkaliphilic esterases, and the unique properties of E69 make it a promising candidate as a biocatalyst for industrial applications.

  16. Characterization of a cold-adapted esterase and mutants from a psychotolerant Pseudomonas sp. strain.

    Science.gov (United States)

    Dong, Juan; Gasmalla, Mohammed A A; Zhao, Wei; Sun, Jingtao; Liu, Wenyu; Wang, Mingming; Han, Liang; Yang, Ruijin

    2017-09-01

    A cold-adapted esterase-producing strain named T1-39 was isolated from Glacier No. 1, Tianshan, People's Republic of China and identified as Pseudomonas sp. from 16S rRNA sequence analysis. The esterase (EstT1-39) secreted by this strain preferentially hydrolyzed esters of glycerol with short- and medium-chain fatty acids. Mutants of T1-39 were generated by the atmospheric and room temperature plasma method and screened for enhanced esterase activity. Among all the mutants, strain TB11 had 4.45-fold higher esterase productivity than T1-39, with high genetic stability over 10 generations of continuous cultivation. Maximum activity of EstT1-39 and EstTB11 was observed at 30 ℃, pH 9.0 and 25 ℃, pH 8.5, respectively. EstTB11 was thermally more stable (50 ℃ for 1 H) and active over a broader pH range than EstT1-39. EstTB11 also retained 38% of its maximal activity at 0 ℃ and was found to be able to hydrolyze milk fats into short- and medium-chain fatty acids at 4 ℃. The characteristics of EstT1-39 made it a cold-adapted enzyme and the EstTB11 from the mutant, with its higher activity at lower temperatures, may be suitable for the production of aromas and flavors in the dairy industry. © 2016 International Union of Biochemistry and Molecular Biology, Inc.

  17. Phylogenetic Analyses of Taro (Colocasia esculenta (L.) Schott) and Related Species based on Esterase Isozymes

    OpenAIRE

    Nguyen, Viet Xuan; Yoshino, Hiromichi; Tahara, Makoto

    1998-01-01

    Phylogenetic relationships among the 84 accessions of taro (Colocasia esculenta (L) Schott), C gigantea Hook Alocasia macrorrhiza, A odora, Xanthosoma sagittifolium (L.) Schott and X. violaceum Schott were investigated using isozyme polymorphism of esterase. The phylogenetic tree estimated by the UPGMA analyses revealed that taro accessions formed a single cluster and C. gigantea was more closely related to Alocasia species than to taro. Taro accessions from Yunnan tended to share band patter...

  18. Insight into Aluminum Sulfate-Catalyzed Xylan Conversion into Furfural in a γ-Valerolactone/Water Biphasic Solvent under Microwave Conditions.

    Science.gov (United States)

    Yang, Tao; Zhou, Yi-Han; Zhu, Sheng-Zhen; Pan, Hui; Huang, Yao-Bing

    2017-10-23

    A simple and efficient biphasic system with an earth-abundant metal salt catalyst was used to produce furfural from xylan with a high yield of up to 87.8 % under microwave conditions. Strikingly, the metal salt Al 2 (SO 4 ) 3 exhibited excellent catalytic activity for xylan conversion, owing to a combination of Lewis and Brønsted acidity and its ability to promote good phase separation. The critical role of the SO 4 2- anion was first analyzed, which resulted in the aforementioned characteristics when combined with the Al 3+ cation. The mixed solvent system with γ-valerolactone (GVL) as the organic phase provided the highest furfural yield, resulting from its good dielectric properties and dissolving capacity, which facilitated the absorption of microwave energy and promoted mass transfer. Mechanistic studies suggested that the xylan-to-furfural conversion proceeded mainly through a hydrolysis-isomerization-dehydration pathway and the hexa-coordinated Lewis acidic [Al(OH) 2 (aq)] + species were the active sites for xylose-xylulose isomerization. Detailed kinetic studies of the subreaction for the xylan conversion revealed that GVL regulates the reaction rates and pathways by promoting the rates of the key steps involved for furfural production and suppressing the side reactions for humin production. Finally, the Al 2 (SO 4 ) 3 catalyst was used for the production of furfural from several lignocellulosic feedstocks, revealing its great potential for other biomass conversions. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Improved biomass degradation using fungal glucuronoyl-esterases-hydrolysis of natural corn fiber substrate

    DEFF Research Database (Denmark)

    d'Errico, Clotilde; Börjesson, Johan; Ding, Hanshu

    2016-01-01

    between glucuronic acids in xylans and lignin alcohols. By means of synthesized complex LCC model substrates we provide kinetic data suggesting a preference of fungal GEs for esters of bulky arylalkyl alcohols such as ester LCCs. Furthermore, using natural corn fiber substrate we report the first examples...... of improved degradation of lignocellulosic biomass by the use of GEs. Improved C5 sugar, glucose and glucuronic acid release was observed when heat pretreated corn fiber was incubated in the presence of GEs from Cerrena unicolor and Trichoderma reesei on top of different commercial cellulase...

  20. A New Esterase from Thermobifida halotolerans Hydrolyses Polyethylene Terephthalate (PET and Polylactic Acid (PLA

    Directory of Open Access Journals (Sweden)

    Georg Steinkellner

    2012-02-01

    Full Text Available A new esterase from Thermobifida halotolerans (Thh_Est was cloned and expressed in E. coli and investigated for surface hydrolysis of polylactic acid (PLA and polyethylene terephthalate (PET. Thh_Est is a member of the serine hydrolases superfamily containing the -GxSxG- motif with 85–87% homology to an esterase from T. alba, to an acetylxylan esterase from T. fusca and to various Thermobifida cutinases. Thh_Est hydrolyzed the PET model substrate bis(benzoyloxyethylterephthalate and PET releasing terephthalic acid and mono-(2-hydroxyethyl terephthalate in comparable amounts (19.8 and 21.5 mmol/mol of enzyme while no higher oligomers like bis-(2-hydroxyethyl terephthalate were detected. Similarly, PLA was hydrolyzed as indicated by the release of lactic acid. Enzymatic surface hydrolysis of PET and PLA led to a strong hydrophilicity increase, as quantified with a WCA decrease from 90.8° and 75.5° to 50.4° and to a complete spread of the water drop on the surface, respectively.

  1. Biochemical characterisation of the esterase activities of wine lactic acid bacteria.

    Science.gov (United States)

    Matthews, Angela; Grbin, Paul R; Jiranek, Vladimir

    2007-11-01

    Esters are an important group of volatile compounds that can contribute to wine flavour. Wine lactic acid bacteria (LAB) have been shown to produce esterases capable of hydrolysing ester substrates. This study aims to characterise the esterase activities of nine LAB strains under important wine conditions, namely, acidic conditions, low temperature (to 10 degrees C) and in the presence of ethanol (2-18% v/v). Esterase substrate specificity was also examined using seven different ester substrates. The bacteria were generally found to have a broad pH activity range, with the majority of strains showing maximum activity close to pH 6.0. Exceptions included an Oenococcus oeni strain that retained most activity even down to a pH of 4.0. Most strains exhibited highest activity across the range 30-40 degrees C. Increasing ethanol concentration stimulated activity in some of the strains. In particular, O. oeni showed an increase in activity up to a maximum ethanol concentration of around 16%. Generally, strains were found to have greater activity towards short-chained esters (C2-C8) compared to long-chained esters (C10-C18). Even though the optimal physicochemical conditions for enzyme activity differed from those found in wine, these findings are of potential importance to oenology because significant activities remained under wine-like conditions.

  2. Discovery of potential cholesterol esterase inhibitors using in silico docking studies

    Directory of Open Access Journals (Sweden)

    Thirumalaisamy Sivashanmugam

    2013-08-01

    Full Text Available New drug discovery is considered broadly in terms of two kinds of investiga-tional activities such as exploration and exploitation. This study deals with the evaluation of the cholesterol esterase inhibitory activity of flavonoids apigenin, biochanin, curcumin, diosmetin, epipervilline, glycitein, okanin, rhamnazin and tangeritin using in silico docking studies. In silico docking studies were carried out using AutoDock 4.2, based on the Lamarckian genetic algorithm principle. The results showed that all the selected flavonoids showed binding energy ranging between -7.08 kcal/mol to -5.64 kcal/mol when compared with that of the standard compound gallic acid (-4.11 kcal/mol. Intermolecular energy (-9.13 kcal/mol to -7.09 kcal/mol and inhibition constant (6.48 µM to 73.18 µM of the ligands also coincide with the binding energy. All the selected flavonoids contributed cholesterol esterase inhibitory activity, these molecular docking analyses could lead to the further develop-ment of potent cholesterol esterase inhibitors for the treatment of obesity.

  3. Eco-friendly surface modification on polyester fabrics by esterase treatment

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Jindan; Cai, Guoqiang; Liu, Jinqiang; Ge, Huayun; Wang, Jiping, E-mail: jipingwanghz@gmail.com

    2014-03-01

    Graphical abstract: - Highlights: • We used a simple and easy way to measure the enzyme activity. • We studied the mechanism by characterizing the chemical changes in the surface of fabric. • We studied the advantages in surface wettability, fiber integrity and mechanical performance of cutinase treated fabrics. • Cutinase pretreated fibers exhibited much improved fabric wicking and better fiber integrity comparing to alkali treated ones. • Cutinase pretreatment technology promotes energy conservation and emission reduction. - Abstract: Currently, traditional alkali deweighting technology is widely used to improve the hydrophilicity of polyester fabrics. However, the wastewater and heavy chemicals in the effluent cause enormous damage to the environment. Esterase treatment, which is feasible in mild conditions with high selectivity, can provide a clean and efficient way for polyester modification. Under the optimum conditions, the polyester fabric hydrolysis process of esterase had a linear kinetics. X-ray photoelectron spectrometry (XPS) results showed that hydroxyl and carboxyl groups were produced only on the surface of modified fiber without changing the chemical composition of the bulk. These fibers exhibited much improved fabric wicking, as well as greatly improved oily stain removal performance. Compared to the harsh alkali hydrolysis, the enzyme treatment led to smaller weight loss and better fiber integrity. The esterase treatment technology is promising to produce higher-quality polyester textiles with an environmental friendly approach.

  4. Production and characterization of a tributyrin esterase from Lactobacillus plantarum suitable for cheese lipolysis.

    Science.gov (United States)

    Esteban-Torres, M; Mancheño, J M; de las Rivas, B; Muñoz, R

    2014-11-01

    Lactobacillus plantarum is a lactic acid bacterium that can be found during cheese ripening. Lipolysis of milk triacylglycerols to free fatty acids during cheese ripening has fundamental consequences on cheese flavor. In the present study, the gene lp_1760, encoding a putative esterase or lipase, was cloned and expressed in Escherichia coli BL21 (DE3) and the overproduced Lp_1760 protein was biochemically characterized. Lp_1760 hydrolyzed p-nitrophenyl esters of fatty acids from C2 to C16, with a preference for p-nitrophenyl butyrate. On triglycerides, Lp_1760 showed higher activity on tributyrin than on triacetin. Although optimal conditions for activity were 45°C and pH 7, Lp_1760 retains activity under conditions commonly found during cheese making and ripening. The Lp_1760 showed more than 50% activity at 5°C and exhibited thermal stability at high temperatures. Enzymatic activity was strongly inhibited by sodium dodecyl sulfate and phenylmethylsulfonyl fluoride. The Lp_1760 tributyrin esterase showed high activity in the presence of NaCl, lactic acid, and calcium chloride. The results suggest that Lp_1760 might be a useful tributyrin esterase to be used in cheese manufacturing. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  5. Mechanism-Guided Discovery of an Esterase Scaffold with Promiscuous Amidase Activity

    Directory of Open Access Journals (Sweden)

    Charlotte Kürten

    2016-06-01

    Full Text Available The discovery and generation of biocatalysts with extended catalytic versatilities are of immense relevance in both chemistry and biotechnology. An enhanced atomistic understanding of enzyme promiscuity, a mechanism through which living systems acquire novel catalytic functions and specificities by evolution, would thus be of central interest. Using esterase-catalyzed amide bond hydrolysis as a model system, we pursued a simplistic in silico discovery program aiming for the identification of enzymes with an internal backbone hydrogen bond acceptor that could act as a reaction specificity shifter in hydrolytic enzymes. Focusing on stabilization of the rate limiting transition state of nitrogen inversion, our mechanism-guided approach predicted that the acyl hydrolase patatin of the α/β phospholipase fold would display reaction promiscuity. Experimental analysis confirmed previously unknown high amidase over esterase activity displayed by the first described esterase machinery with a protein backbone hydrogen bond acceptor to the reacting NH-group of amides. The present work highlights the importance of a fundamental understanding of enzymatic reactions and its potential for predicting enzyme scaffolds displaying alternative chemistries amenable to further evolution by enzyme engineering.

  6. Predicting post-translational lysine acetylation using support vector machines

    DEFF Research Database (Denmark)

    Gnad, Florian; Ren, Shubin; Choudhary, Chunaram

    2010-01-01

    Lysine acetylation is a post-translational protein modification and a primary regulatory mechanism that controls many cell signaling processes. Lysine acetylation sites are recognized by acetyltransferases and deacetylases through sequence patterns (motifs). Recently, we used high-resolution mass...... spectrometry to identify 3600 lysine acetylation sites on 1750 human proteins covering most of the previously annotated sites and providing the most comprehensive acetylome so far. This dataset should provide an excellent source to train support vector machines (SVMs) allowing the high accuracy in silico...

  7. Phosphorylation and Acetylation of Acyl-CoA Synthetase- I

    DEFF Research Database (Denmark)

    Frahm, Jennifer L; Li, Lei O; Grevengoed, Trisha J

    2011-01-01

    Long chain acyl-CoA synthetase 1 (ACSL1) contributes 50 to 90% of total ACSL activity in liver, adipose tissue, and heart and appears to direct the use of long chain fatty acids for energy. Although the functional importance of ACSL1 is becoming clear, little is understood about its post...... and acetylated amino acids by mass spectrometry. We then compared these results to the post-translational modifications observed in vivo in liver and brown adipose tissue after mice were fasted or exposed to a cold environment. We identified universal N-terminal acetylation, 15 acetylated lysines, and 25...

  8. Esterases of Varroa destructor (Acari: Varroidae), parasitic mite of the honeybee.

    Science.gov (United States)

    Dmitryjuk, Małgorzata; Żołtowska, Krystyna; Frączek, Regina; Lipiński, Zbigniew

    2014-04-01

    Varroa destructor is an ectoparasite that causes serious damage to the population of the honeybee. Increasing resistance of the parasite to acaricides is related, among others, to metabolic adaptations of its esterases to facilitate decomposition of the chemicals used. Esterases are a large heterogeneous group of enzymes that metabolize a number of endogenous and exogenous substrates with ester binding. The aim of the present study was to determine the activity of esterases in the body extracts (BE) and excretion/secretion products (E/SP) of the mite. The enzymes contained in the E/SP should originate mainly from the salivary glands and the alimentary system and they may play a particularly important role in the first line of defence of the mite against acaricides. Activity of cholinesterases (ChEs) [acetylcholinesterase (AChE) and butyrylcholinesterase], carboxylesterases (CEs) and phosphatases [alkaline phosphatase (AP) and acid phosphatase (AcP)] was investigated. The activity of all the enzymes except AChE was higher in the E/SP than in the BE. ChEs from the BE and from the E/SP reacted differently on eserine, a ChE inhibitor. Eserine inhibited both enzymes from the BE, increased decomposition of acetylcholine, but did not influence hydrolysis of butyrylcholine by the E/SP. Activity of the CEs from the BE in relation to the esters of carboxylic acids can be presented in the following series: C10 > C12 > C14 > C8 > C2 > C4 = C16, while activity of the CEs from the E/SP was: C4 > C8 > C2 > C14 > C10 > C12 > C16. The inhibitor of CEs, triphenyl phosphate, reduced the activity of esterases C2–C8 and C14–C16; however, it acted in the opposite way to CEs C10 and C12. The activity of both phosphatases was higher in the E/SP than in the BE (AcP about twofold and AP about 2.6-fold); the activities of AP and AcP in the same material were similar. Given the role of esterases in resistance to pesticides, further studies are necessary to obtain complete biochemical

  9. Xylan utilization in human gut commensal bacteria is orchestrated by unique modular organization of polysaccharide-degrading enzymes.

    Science.gov (United States)

    Zhang, Meiling; Chekan, Jonathan R; Dodd, Dylan; Hong, Pei-Ying; Radlinski, Lauren; Revindran, Vanessa; Nair, Satish K; Mackie, Roderick I; Cann, Isaac

    2014-09-02

    Enzymes that degrade dietary and host-derived glycans represent the most abundant functional activities encoded by genes unique to the human gut microbiome. However, the biochemical activities of a vast majority of the glycan-degrading enzymes are poorly understood. Here, we use transcriptome sequencing to understand the diversity of genes expressed by the human gut bacteria Bacteroides intestinalis and Bacteroides ovatus grown in monoculture with the abundant dietary polysaccharide xylan. The most highly induced carbohydrate active genes encode a unique glycoside hydrolase (GH) family 10 endoxylanase (BiXyn10A or BACINT_04215 and BACOVA_04390) that is highly conserved in the Bacteroidetes xylan utilization system. The BiXyn10A modular architecture consists of a GH10 catalytic module disrupted by a 250 amino acid sequence of unknown function. Biochemical analysis of BiXyn10A demonstrated that such insertion sequences encode a new family of carbohydrate-binding modules (CBMs) that binds to xylose-configured oligosaccharide/polysaccharide ligands, the substrate of the BiXyn10A enzymatic activity. The crystal structures of CBM1 from BiXyn10A (1.8 Å), a cocomplex of BiXyn10A CBM1 with xylohexaose (1.14 Å), and the CBM from its homolog in the Prevotella bryantii B14 Xyn10C (1.68 Å) reveal an unanticipated mode for ligand binding. A minimal enzyme mix, composed of the gene products of four of the most highly up-regulated genes during growth on wheat arabinoxylan, depolymerizes the polysaccharide into its component sugars. The combined biochemical and biophysical studies presented here provide a framework for understanding fiber metabolism by an important group within the commensal bacterial population known to influence human health.

  10. Xylan utilization in human gut commensal bacteria is orchestrated by unique modular organization of polysaccharide-degrading enzymes

    KAUST Repository

    Zhang, Meiling

    2014-08-18

    Enzymes that degrade dietary and host-derived glycans represent the most abundant functional activities encoded by genes unique to the human gut microbiome. However, the biochemical activities of a vast majority of the glycan-degrading enzymes are poorly understood. Here, we use transcriptome sequencing to understand the diversity of genes expressed by the human gut bacteria Bacteroides intestinalis and Bacteroides ovatus grown in monoculture with the abundant dietary polysaccharide xylan. The most highly induced carbohydrate active genes encode a unique glycoside hydrolase (GH) family 10 endoxylanase (BiXyn10A or BACINT-04215 and BACOVA-04390) that is highly conserved in the Bacteroidetes xylan utilization system. The BiXyn10A modular architecture consists of a GH10 catalytic module disrupted by a 250 amino acid sequence of unknown function. Biochemical analysis of BiXyn10A demonstrated that such insertion sequences encode a new family of carbohydrate-binding modules (CBMs) that binds to xy-lose- configured oligosaccharide/polysaccharide ligands, the substrate of the BiXyn10A enzymatic activity. The crystal structures of CBM1 from BiXyn10A (1.8 Å), a cocomplex of BiXyn10A CBM1 with xylohexaose (1.14 Å), and the CBM fromits homolog in the Prevotella bryantii B 14 Xyn10C (1.68 Å) reveal an unanticipated mode for ligand binding. Aminimal enzyme mix, composed of the gene products of four of the most highly up-regulated genes during growth on wheat arabinoxylan, depolymerizes the polysaccharide into its component sugars. The combined biochemical and biophysical studies presented here provide a framework for understanding fiber metabolism by an important group within the commensal bacterial population known to influence human health.

  11. Prospect for Developing a Consolidated Bioprocessing (CBP) Strain Using Xylan as the Substrate: the Case Study of Yarrowia lipolytica

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Wei; Wei, Hui; Alahuhta, Markus; Zhang, Min; Himmel, Michael E.

    2016-07-08

    To achieve the goal of developing a direct microbial sugar conversion platform for the production of lipids and drop-in fuels from cellulosic biomass substrate, Yarrowia lipolytica was used to investigate its potential for being developed as CBP strain by expressing cellulase and xylanase enzymes. Y. lipolytica is known to accumulate lipids intracellularly and is capable of metabolizing glucose and xylose to produce lipids; however, due to the lack of the biomass degrading enzymes, it cannot directly utilize lignocellulosic substrates as carbon sources. While research is continuing on the development of a Y. lipolytica strain able to degrade cellulose, in this study, we present successful expression of several xylanases in Y. lipolytica. To the best of our knowledge, this is the first study introducing heterologous hemicellulose genes into the genome of Y. lipolytica. SDS-PAGE and western blotting analysis showed that the endo-xylanase gene XynII and exo-xylosidase gene XlnD were successfully expressed and secreted, and the expressed xylanases were likely either not or sparsely glycosylated, which is advantageous for expression of heterologous proteins from any species. Enzymatic activity tests further demonstrated active expression of XynII and XlnD in Y. lipolytica. Furthermore, synergistic action on converting xylan to xylose was observed when XlnD worked in concert with XynII. XlnD was able to work on the xylo-oligomers generated by XynII, enhancing the xylan conversion to monomeric xylose. The successful expression of these xylanases in Yarrowia further advances us towards our goal to develop a direct microbial conversion process using this organism. and xylose to produce lipids; however, due to the lack of the biomass degrading enzymes, it cannot directly utilize lignocellulosic substrates as carbon sources. While research is continuing on the development of a Y. lipolytica strain able to degrade cellulose, in this study, we present successful expression of

  12. Obtaining cellulose binding and hydrolyzing activity of a family 11 hybrid xylanase by fusion with xylan binding domain.

    Science.gov (United States)

    Liu, Ming-Qi; Dai, Xian-Jun; Liu, Guang-Fu; Wang, Qian

    2013-03-01

    The xylan binding domain (XBD) and linker sequences (LS) from thermostable and thermophilic Thermomonospora fusca xylanase A (TfxA) was fused to the carboxyl-terminus of a family 11 hybrid xylanase ATx. The constructed chimera (ATxX) was successfully expressed in Pichia pastoris, partially purified to homogeneity, and then characterized in detail. After 96-h 0.25% methanol induction, the xylanase and cellulose activity of ATxX from pPATxX1 transformant culture medium supernatant were 452.1 U/mg and 19.3 U/mg, respectively. SDS-PAGE analysis revealed that the molecular mass of ATxX was about 33.01 kDa. 3.7% ATxX was bound after incubation with 1% microcrystal cellulose at 25 °C for 3 h, while the ATx did not show cellulose binding-hydrolyzing ability. These results suggested that ATx obtained cellulose binding and hydrolyzing ability by fusing with XBD and LS. Enzymatic studies showed that the temperature and pH optimum of the ATxX xylanase activity were 60 °C and pH 5.0, respectively, which were the same as that of ATx. The temperature and pH optimum of the ATxX cellulase activity were 60 °C and pH 6.0, respectively. The major hydrolytic products released by ATxX from birchwood xylan were xylotriose and xylohexaose. Xylooligosaccharides from xylobiose to xylohexaose could be hydrolyzed by ATxX. Mode of action analysis showed that the chimeric ATxX was an endo-acting enzyme. The XBD and LS plays an important role in the binding and hydrolyzing of xylanase to insoluble substrates. Copyright © 2012 Elsevier Inc. All rights reserved.

  13. Acetylation of Mammalian ADA3 Is Required for Its Functional Roles in Histone Acetylation and Cell Proliferation.

    Science.gov (United States)

    Mohibi, Shakur; Srivastava, Shashank; Bele, Aditya; Mirza, Sameer; Band, Hamid; Band, Vimla

    2016-10-01

    Alteration/deficiency in activation 3 (ADA3) is an essential component of specific histone acetyltransferase (HAT) complexes. We have previously shown that ADA3 is required for establishing global histone acetylation patterns and for normal cell cycle progression (S. Mohibi et al., J Biol Chem 287:29442-29456, 2012, http://dx.doi.org/10.1074/jbc.M112.378901). Here, we report that these functional roles of ADA3 require its acetylation. We show that ADA3 acetylation, which is dynamically regulated in a cell cycle-dependent manner, reflects a balance of coordinated actions of its associated HATs, GCN5, PCAF, and p300, and a new partner that we define, the deacetylase SIRT1. We use mass spectrometry and site-directed mutagenesis to identify major sites of ADA3 acetylated by GCN5 and p300. Acetylation-defective mutants are capable of interacting with HATs and other components of HAT complexes but are deficient in their ability to restore ADA3-dependent global or locus-specific histone acetylation marks and cell proliferation in Ada3-deleted murine embryonic fibroblasts (MEFs). Given the key importance of ADA3-containing HAT complexes in the regulation of various biological processes, including the cell cycle, our study presents a novel mechanism to regulate the function of these complexes through dynamic ADA3 acetylation. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  14. Generation of nitryl chloride from chlorotrimethylsilane-acetyl nitrate ...

    Indian Academy of Sciences (India)

    Administrator

    amyl nitrate does not yield NO2Cl with silicon reagent. However, acetyl nitrate reacts successfully with chlorotrimethylsi- lane to give nitryl chloride, which is characterized by its UV spectrum. If it is generated in presence of ketoximes ...

  15. Partially Acetylated Sugarcane Bagasse For Wicking Oil From Contaminated Wetlands

    Science.gov (United States)

    Sugarcane bagasse was partially acetylated to enhance its oil-wicking ability in saturated environments while holding moisture for hydrocarbon biodegradation. The water sorption capacity of raw bagasse was reduced fourfold after treatment, which indicated considerably increased ...

  16. A Non-Isotopic In Vitro Assay for Histone Acetylation

    Science.gov (United States)

    Kuninger, David; Lundblad, James; Semirale, Anthony; Rotwein, Peter

    2007-01-01

    We describe a simple, robust, and relatively inexpensive non-radioactive in vitro assay for measuring histone acetyl-transferase activity. The assay takes advantage of easy to purify recombinant E. coli-derived fusion proteins containing the NH2-terminal tails of histones H3 and H4 linked to epitope-tagged maltose binding protein (MBP), and immunoblotting with antibodies specific to acetylated H3 and H4. Here we show the specificity and dynamic range of this assay for the histone acetyl-transferases, p300 and PCAF. This assay may be adapted readily for other substrates by simply generating new fusion proteins and for other acetyl-transferases by modifying reaction conditions. PMID:17698235

  17. Synthesis of spiro [indolo-1, 5-benzodiazepines] from 3-acetyl ...

    Indian Academy of Sciences (India)

    3'-hydroxy-2'-oxo indolo) acetyl coumarins (3), which on dehydration afforded the corresponding ,-unsaturated ketones (4). Cyclocondensation of (4) with substituted -phenylene diamines resulted in novel 3-coumarinyl spiro[indolo-1 ...

  18. Mechanistic insights into the regulation of metabolic enzymes by acetylation

    Science.gov (United States)

    2012-01-01

    The activity of metabolic enzymes is controlled by three principle levels: the amount of enzyme, the catalytic activity, and the accessibility of substrates. Reversible lysine acetylation is emerging as a major regulatory mechanism in metabolism that is involved in all three levels of controlling metabolic enzymes and is altered frequently in human diseases. Acetylation rivals other common posttranslational modifications in cell regulation not only in the number of substrates it modifies, but also the variety of regulatory mechanisms it facilitates. PMID:22826120

  19. Acetyl radical generation in cigarette smoke: Quantification and simulations

    Science.gov (United States)

    Hu, Na; Green, Sarah A.

    2014-10-01

    Free radicals are present in cigarette smoke and can have a negative effect on human health. However, little is known about their formation mechanisms. Acetyl radicals were quantified in tobacco smoke and mechanisms for their generation were investigated by computer simulations. Acetyl radicals were trapped from the gas phase using 3-amino-2, 2, 5, 5-tetramethyl-proxyl (3AP) on solid support to form stable 3AP adducts for later analysis by high-performance liquid chromatography (HPLC), mass spectrometry/tandem mass spectrometry (MS-MS/MS) and liquid chromatography-mass spectrometry (LC-MS). Simulations were performed using the Master Chemical Mechanism (MCM). A range of 10-150 nmol/cigarette of acetyl radical was measured from gas phase tobacco smoke of both commercial and research cigarettes under several different smoking conditions. More radicals were detected from the puff smoking method compared to continuous flow sampling. Approximately twice as many acetyl radicals were trapped when a glass fiber particle filter (GF/F specifications) was placed before the trapping zone. Simulations showed that NO/NO2 reacts with isoprene, initiating chain reactions to produce hydroxyl radical, which abstracts hydrogen from acetaldehyde to generate acetyl radical. These mechanisms can account for the full amount of acetyl radical detected experimentally from cigarette smoke. Similar mechanisms may generate radicals in second hand smoke.

  20. Izoenzimas esterases para discriminar cultivares "sem nome" de mandioca (Manihot esculenta Esterase isozymes for the characterization of "unnamed" cassava cultivars (Manihot esculenta Crantz

    Directory of Open Access Journals (Sweden)

    Fábio Pablos de Souza

    2000-05-01

    Full Text Available Isoenzimas esterases foram usadas como marcadores moleculares para discriminar e agrupar sete cultivares "sem nomes" (acessos A-G de Manihot esculenta. Os cultivares "sem nomes" de mandioca foram comparados com 25 diferentes cultivares (BG que vêm sendo mantidos na coleção de germoplasma do Departamento de Agronomia, da Universidade Estadual de Maringá. Acetato e propionato de 4-metilumbeliferona e acetato de α–naftil, foram os substratos utilizados para a detecção e análise comparativa das isoesterases. A similaridade entre as plantas, usando o coeficiente de Jaccard, variou de 47,6% até 100%. O dendrograma produzido pela análise de agrupamento mostrou identidade entre as plantas do cultivar BG23 e as plantas do acesso D. As plantas dos acessos B e G também foram agrupadas com o cultivar BG 23, mostrando similaridade de 95% e 89%, respectivamente. As plantas dos acessos A e E foram similares às plantas BG 1, mostrando 95% e 90% de similaridade, respectivamente. As plantas do acesso F foram agrupadas com as plantas do cultivar BG 9, mostrando 94% de similaridade. O dendrograma mostrou também que a maioria dos cultivares foram agrupados com 85-90% de similaridade. Assim, concluímos que as isozimas esterases podem ser utilizadas como marcadores moleculares de genótipos de mandioca, para a caracterização dos cultivares sem nomes de M. esculentaEsterase isozymes were used as molecular markers to discriminate and cluster seven "unnamed" cultivars (accesses A-G of M. esculenta. The "unnamed" cassava cultivars were compared to 25 different M. esculenta cultivars (cultivars BG, which have been maintained in the germplasm collection of the Agronomy Department, State University of Maringá. 4-Methylumbelliferyl acetate, 4-methylumbelliferyl propionate and α–naphthyl acetate were utilized as substrates for isoesterase detection and comparative analysis. Similarity between plants, using Jaccard’s coefficient, ranged from 47.6% to 100

  1. Esterase isoenzymes and insecticide resistance in Frankliniella occidentalis populations from the south-east region of Spain.

    Science.gov (United States)

    López-Soler, Neus; Cervera, Amelia; Moores, Graham D; Martínez-Pardo, Rafael; Garcerá, M Dolores

    2008-12-01

    Frankliniella occidentalis (Pergande) is among the most important crop pests in the south-east region of Spain; its increasing resistance to insecticides constitutes a serious problem, and understanding the mechanisms involved is therefore of great interest. To this end, F. occidentalis populations, collected from the field at different locations in south-east Spain, were studied in terms of total esterase activity and esterase isoenzyme pattern. Individual thrips extracts were analysed by native polyacrylamide gel electrophoresis (PAGE) and stained for esterase activity with the model substrate alpha-naphthyl acetate. Significant correlations were found between resistance to the insecticides acrinathrin and methiocarb and the presence of a group of three intensely stained bands, named Triplet A. For each individual thrips extract, total esterase activity towards the substrates alpha-naphthyl acetate and alpha-naphthyl butyrate was also measured in a microplate reader. Insects possessing Triplet A showed a significantly higher alpha-naphthyl acetate specific activity and alpha-naphthyl acetate/alpha-naphthyl butyrate activity ratio. This observation allowed a reliable classification of susceptible or resistant insects either by PAGE analysis or by total esterase activity determination. The PAGE and microplate assays described can be used as a monitoring technique for detecting acrinathrin- and methiocarb-resistant individuals among F. occidentalis field populations.

  2. Seleção de bacillus spp. para produção de esterases e melhoramento de bacillus cereus (c124 Selection of bacillus spp. For esterase production and genetic improvement of bacillus cereus (c124

    Directory of Open Access Journals (Sweden)

    Analucia Longman Mendonça

    1998-06-01

    Full Text Available Forty-four Bacillus spp. strains obtained from sugar cane derivates and residues, six of them isolated in this work, were tested using Tween 80 as substrate (agar-Tween 80 medium, in order to determine their esterase activity through the enzymatic index averages. After statistic analysis, B. cereus (C124 strain, which presented better results, was submitted to genetic improvement by treatment with ultraviolet light (UV. The survival curve pointed out 28" as the time necessary to obtain 30% of survivors. Fifty survivors and the wild strain C124 were compared in relation to their esterase activity as mentioned previously. The wild strain and the mutant C124UV35, which showed enzymatic index average higher than C124, were characterized in polyacrilamide gel electrophoresis (PAGE. Eletrophoretic patterns for total proteins of wild and mutant strain showed different profiles according to number, position and intensity of bands. For esterase, the bands varied only in intensity.

  3. Molecular cloning and characterization of a novel pyrethroid-hydrolyzing esterase originating from the Metagenome

    Directory of Open Access Journals (Sweden)

    Liu Yu

    2008-12-01

    Full Text Available Abstract Background Pyrethroids and pyrethrins are widely used insecticides. Extensive applications not only result in pest resistance to these insecticides, but also may lead to environmental issues and human exposure. Numerous studies have shown that very high exposure to pyrethroids might cause potential problems to man and aquatic organisms. Therefore, it is important to develop a rapid and efficient disposal process to eliminate or minimize contamination of surface water, groundwater and agricultural products by pyrethroid insecticides. Bioremediation is considered to be a reliable and cost-effective technique for pesticides abatement and a major factor determining the fate of pyrethroid pesticides in the environment, and suitable esterase is expected to be useful for potential application for detoxification of pyrethroid residues. Soil is a complex environment considered as one of the main reservoirs of microbial diversity on the planet. However, most of the microorganisms in nature are inaccessible as they are uncultivable in the laboratory. Metagenomic approaches provide a powerful tool for accessing novel valuable genetic resources (novel enzymes and developing various biotechnological applications. Results The pyrethroid pesticides residues on foods and the environmental contamination are a public safety concern. Pretreatment with pyrethroid-hydrolyzing esterase has the potential to alleviate the conditions. To this end, a pyrethroid-hydrolyzing esterase gene was successfully cloned using metagenomic DNA combined with activity-based functional screening from soil, sequence analysis of the DNA responsible for the pye3 gene revealed an open reading frame of 819 bp encoding for a protein of 272 amino acid residues. Extensive multiple sequence alignments of the deduced amino acid of Pye3 with the most homologous carboxylesterases revealed moderate identity (45–49%. The recombinant Pye3 was heterologously expressed in E. coli BL21(DE3

  4. The application of digestive tract lactic acid bacteria with high esterase activity for zearalenone detoxification.

    Science.gov (United States)

    Chen, Shiau-Wei; Hsu, Jih-Tay; Chou, Yan-An; Wang, Han-Tsung

    2018-01-24

    Zearalenone (ZEA) is an estrogenic mycotoxin produced by several Fusarium species and frequently contaminates cereals used for food or animal feed. This study attempted to select lactic acid bacteria (LAB) with high esterase activity from the digestive tract, with the goal of using these bacteria for ZEA detoxification. No ZEA activity-related biotransformation products were observed in three isolates (B1, B2 and D10) during incubation in the presence of ZEA. All three LAB strains were Lactobacillus plantarum, but the API 50 CHL results suggested that the three isolates were different strains. Increased esterase activity was associated with an increase in cell growth, and the ZEA-detoxifying capabilities of isolates rely on the concentration of bacteria in the culture medium. The lipolytic activity and ZEA removal assay indicated that ZEA degradation by the supernatant fraction was dependent on esterase activity; the supernatant of B2 strain showed the highest ZEA degradation ability and did not release the binding ZEA back into the medium. The D10 strain showed fast ZEA binding ability during the late log phase but began to release the bound ZEA back into the medium after the early stationary phase. All isolates showed good acid and bile salt tolerance ability but all strains showed low adhesion ability to epithelial cells. Based on the ZEA removal characterization and ability of the isolates, it is suggested that the isolates could be applied to ZEA detoxification of contaminated feed, but the with the requirement of high cell number for ZEA binding and limited degradation time before absorption of ZEA in the digestive tract. © 2018 Society of Chemical Industry. © 2018 Society of Chemical Industry.

  5. 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...... (corn cobs) which compared favorably to those reported for the other microorganisms. Use of de-esterified corn cobs as carbon source decreased FAE production by 5.5-fold compared to untreated corn cobs even though ferulic acid (FA) was added to the concentration found in alkali-extracts of corn cobs...

  6. Esterase activity able to hydrolyze dietary antioxidant hydroxycinnamates is distributed along the intestine of mammals

    DEFF Research Database (Denmark)

    Andreasen, Mette Findal; Kroon, P A; Williamson, G

    2001-01-01

    and may contribute to the beneficial effects derived from consumption of cereal bran. However, these compounds are ester linked to the main polymers in the plant cell wall and cannot be absorbed in this complex form. The present work shows that esterases with activity toward esters of the major dietary......Hydroxycinnamic acids are effective antioxidants and are abundant components of plant cell walls, especially in cereal bran. For example, wheat and rye brans are rich sources of the hydroxycinnamates ferulic acid, sinapic acid, and p-coumaric acid. These phenolics are part of human and animal diets...

  7. 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...

  8. Acidic-alkaline ferulic acid esterase from Chaetomium thermophilum var. dissitum: Molecular cloning and characterization of recombinant enzyme expressed in Pichia pastoris

    DEFF Research Database (Denmark)

    Dotsenko, Gleb; Tong, Xiaoxue; Pilgaard, Bo

    2016-01-01

    A novel ferulic acid esterase encoding gene CtFae, was successfully cloned from a highly esterase active strain of the thermophile ascomycetous fungus Chaetomium thermophilum var. dissitum; the gene was heterologously expressed in Pichia pastoris KM71H. The recombinant enzyme (CtFae) was purified...

  9. Histochemical studies on genetical control of hormonal enzyme inducibility in the mouse. IV: Cellular localization of androgen sensitive nonspecific esterase in the epididymis

    DEFF Research Database (Denmark)

    Kirkeby, S; Blecher, S R

    1981-01-01

    Nonspecific esterase of mouse epididymis has previously been studied histochemically, using alpha naphthyl-acetate and 5-bromoindoxyl acetate techniques, as well as certain inhibitors. Epithelial cell types of the epididymis have been characterized, and certain esterase isozymes in a particular...

  10. Biochemical Characterization and Relative Expression Levels of Multiple Carbohydrate Esterases of the Xylanolytic Rumen Bacterium Prevotella ruminicola 23 Grown on an Ester-Enriched Substrate

    NARCIS (Netherlands)

    Kabel, M.A.; Yeoman, C.J.; Han, Y.; Dodd, D.; Abbas, C.A.; Bont, de J.A.M.; Morrison, M.; Cann, I.K.O.; Mackie, R.I.

    2011-01-01

    We measured expression and used biochemical characterization of multiple carbohydrate esterases by the xylanolytic rumen bacterium Prevotella ruminicola 23 grown on an ester-enriched substrate to gain insight into the carbohydrate esterase activities of this hemicellulolytic rumen bacterium. The P.

  11. Enzyme resistant feruloylated xylooligomer analogues from thermochemically treated corn fiber contain large side chains, ethyl glycosides and novel sites of acetylation.

    Science.gov (United States)

    Appeldoorn, Maaike M; de Waard, Pieter; Kabel, Mirjam A; Gruppen, Harry; Schols, Henk A

    2013-11-15

    In order to use corn fiber as a source for bioethanol production the enzymatic hydrolysis of the complex glucuronoarabinoxylans present has to be improved. Several oligosaccharides present in the supernatant of mild acid pretreated and enzymatically saccharified corn fiber that resist the current available enzymes were (semi)purified for structural analysis by NMR or ESI-MS(n). The structural features of 21 recalcitrant oligosaccharides are presented. A common feature of almost all these oligosaccharides is that they contain (part of) an α-l-galactopyranosyl-(1→2)-β-d-xylopyranosyl-(1→2)-5-O-trans-feruloyl-l-arabinofuranose side chain attached to the O-3 position of the β-1-4 linked xylose backbone. Several of the identified oligosaccharides contained an ethyl group at the reducing end hypothesized to be formed during SSF. The ethyl glycosides found are far more complex than previously described structures. A new feature present in more than half of the oligosaccharides is an acetyl group attached to the O-2 position of the same xylose to which the oligomeric side chain was attached to the O-3 position. Finding enzymes attacking these large side chains and the dense substituted xylan backbone will boost the hydrolysis of corn fiber glucuronoxylan. Copyright © 2013 Elsevier Ltd. All rights reserved.

  12. Furfural Production from d-Xylose and Xylan by Using Stable Nafion NR50 and NaCl in a Microwave-Assisted Biphasic Reaction

    Directory of Open Access Journals (Sweden)

    Sarah Le Guenic

    2016-08-01

    Full Text Available Pentose dehydration and direct transformation of xylan into furfural were performed in a water-cyclopentyl methyl ether (CPME biphasic system under microwave irradiation. Heated up between 170 and 190 °C in the presence of Nafion NR50 and NaCl, d-xylose, l-arabinose and xylan gave furfural with maximum yields of 80%, 42% and 55%, respectively. The influence of temperature and reaction time on the reaction kinetics was discussed. This study was also completed by the survey of different reactant ratios, such as organic layer-water or catalyst-inorganic salt ratios. The exchange between proton and cation induced by an excess of NaCl was monitored, and a synergetic effect between the remaining protons and the released HCl was also discovered.

  13. The effect of EDTA and metal cations on the 5-bromoindoxyl acetate esterase activity in the thyroid of the guinea pig

    DEFF Research Database (Denmark)

    Kirkeby, S

    1976-01-01

    Miscellaneous metal cations and EDTA have been used as activators and inhibitors of esterase activity in the thyroid of the guinea-pig. The results indicate that the 5-bromoiondoxyl acetate esterase in the epithelial cells probably consists of two different A-esterase isoenzymes, one present...... in group I cells (the para-, intra-, and inter-follicular cells) and the other in group II cells (the follicular cells proper). The first isoenzyme seems to be calcium-dependent whereas the other is activated by various metal ions. Ca2+ + Mn2+ and Ca2+ + Co2+ were found to activate the esterase activity...... in group I cells. EDTA and Mn2+, on the other hand, activated the esterase activity in group II cells....

  14. Asparagus IRX9, IRX10, and IRX14A Are Components of an Active Xylan Backbone Synthase Complex that Forms in the Golgi Apparatus1[OPEN

    Science.gov (United States)

    Zeng, Wei; Picard, Kelsey L.; Song, Lili; Wu, Ai-Min; Farion, Isabela M.; Zhao, Jia; Ford, Kris; Bacic, Antony

    2016-01-01

    Heteroxylans are abundant components of plant cell walls and provide important raw materials for the food, pharmaceutical, and biofuel industries. A number of studies in Arabidopsis (Arabidopsis thaliana) have suggested that the IRREGULAR XYLEM9 (IRX9), IRX10, and IRX14 proteins, as well as their homologs, are involved in xylan synthesis via a Golgi-localized complex termed the xylan synthase complex (XSC). However, both the biochemical and cell biological research lags the genetic and molecular evidence. In this study, we characterized garden asparagus (Asparagus officinalis) stem xylan biosynthesis genes (AoIRX9, AoIRX9L, AoIRX10, AoIRX14A, and AoIRX14B) by heterologous expression in Nicotiana benthamiana. We reconstituted and partially purified an active XSC and showed that three proteins, AoIRX9, AoIRX10, and AoIRX14A, are necessary for xylan xylosyltranferase activity in planta. To better understand the XSC structure and its composition, we carried out coimmunoprecipitation and bimolecular fluorescence complementation analysis to show the molecular interactions between these three IRX proteins. Using a site-directed mutagenesis approach, we showed that the DxD motifs of AoIRX10 and AoIRX14A are crucial for the catalytic activity. These data provide, to our knowledge, the first lines of biochemical and cell biological evidence that AoIRX9, AoIRX10, and AoIRX14A are core components of a Golgi-localized XSC, each with distinct roles for effective heteroxylan biosynthesis. PMID:26951434

  15. Asparagus IRX9, IRX10, and IRX14A Are Components of an Active Xylan Backbone Synthase Complex that Forms in the Golgi Apparatus.

    Science.gov (United States)

    Zeng, Wei; Lampugnani, Edwin R; Picard, Kelsey L; Song, Lili; Wu, Ai-Min; Farion, Isabela M; Zhao, Jia; Ford, Kris; Doblin, Monika S; Bacic, Antony

    2016-05-01

    Heteroxylans are abundant components of plant cell walls and provide important raw materials for the food, pharmaceutical, and biofuel industries. A number of studies in Arabidopsis (Arabidopsis thaliana) have suggested that the IRREGULAR XYLEM9 (IRX9), IRX10, and IRX14 proteins, as well as their homologs, are involved in xylan synthesis via a Golgi-localized complex termed the xylan synthase complex (XSC). However, both the biochemical and cell biological research lags the genetic and molecular evidence. In this study, we characterized garden asparagus (Asparagus officinalis) stem xylan biosynthesis genes (AoIRX9, AoIRX9L, AoIRX10, AoIRX14A, and AoIRX14B) by heterologous expression in Nicotiana benthamiana We reconstituted and partially purified an active XSC and showed that three proteins, AoIRX9, AoIRX10, and AoIRX14A, are necessary for xylan xylosyltranferase activity in planta. To better understand the XSC structure and its composition, we carried out coimmunoprecipitation and bimolecular fluorescence complementation analysis to show the molecular interactions between these three IRX proteins. Using a site-directed mutagenesis approach, we showed that the DxD motifs of AoIRX10 and AoIRX14A are crucial for the catalytic activity. These data provide, to our knowledge, the first lines of biochemical and cell biological evidence that AoIRX9, AoIRX10, and AoIRX14A are core components of a Golgi-localized XSC, each with distinct roles for effective heteroxylan biosynthesis. © 2016 American Society of Plant Biologists. All Rights Reserved.

  16. D-xylose isomerase from a marine bacterium, Vibrio sp. strain XY-214, and D-xylulose production from β-1,3-xylan.

    Science.gov (United States)

    Umemoto, Yoshiaki; Shibata, Toshiyuki; Araki, Toshiyoshi

    2012-02-01

    The xylA gene from a marine bacterium, Vibrio sp. strain XY-214, encoding D-xylose isomerase (XylA) was cloned and expressed in Escherichia coli. The xylA gene consisted of 1,320-bp nucleotides encoding a protein of 439 amino acids with a predicted molecular weight of 49,264. XylA was classified into group II xylose isomerases. The native XylA was estimated to be a homotetramer with a molecular mass of 190 kDa. The purified recombinant XylA exhibited maximal activity at 60°C and pH 7.5. Its apparent K (m) values for D-xylose and D-glucose were 7.93 and 187 mM, respectively. Furthermore, we carried out D-xylulose production from β-1,3-xylan, a major cell wall polysaccharide component of the killer alga Caulerpa taxifolia. The synergistic action of β-1,3-xylanase (TxyA) and β-1,3-xylosidase (XloA) from Vibrio sp. strain XY-214 enabled efficient saccharification of β-1,3-xylan to D-xylose. D-xylose was then converted to D-xylulose by using XylA from the strain XY-214. The conversion rate of D-xylose to D-xylulose by XylA was found to be approximately 40% in the presence of 4 mM sodium tetraborate after 2 h of incubation. These results demonstrated that TxyA, XloA, and XylA from Vibrio sp. strain XY-214 are useful tools for D-xylulose production from β-1,3-xylan. Because D-xylulose can be used as a source for ethanol fermentation by yeast Saccharomyces cerevisiae, the present study will provide a basis for ethanol production from β-1,3-xylan.

  17. Fibulin-1C, C1 esterase inhibitor and glucose regulated protein 75 interact with the CREC proteins, calumenin and reticulocalbin

    DEFF Research Database (Denmark)

    Hansen, Gry Aune Westergaard; Ludvigsen, Maja; Jacobsen, Christian

    2015-01-01

    Affinity purification, immunoprecipitation, gel electrophoresis and mass spectrometry were used to identify fibulin-1C, C1 esterase inhibitor and glucose regulated protein 75, grp75, as binding partners of the CREC proteins, calumenin and reticulocalbin. Surface plasmon resonance was used to verify...... the interaction of all three proteins with each of the CREC proteins. Fibulin-1C interacts with calumenin and reticulocalbin with an estimated dissociation constant around 50-60 nM. The interaction, at least for reticulocalbin, was not dependent upon the presence of Ca2+. C1 esterase inhibitor interacted...

  18. Synthesis of Acylated Xylan-Based Magnetic Fe3O4 Hydrogels and Their Application for H2O2 Detection

    Directory of Open Access Journals (Sweden)

    Qing-Qing Dai

    2016-08-01

    Full Text Available Acylated xylan-based magnetic Fe3O4 nanocomposite hydrogels (ACX-MNP-gels were prepared by fabricating Fe3O4 nanoctahedra in situ within a hydrogel matrix which was synthesized by the copolymerization of acylated xylan (ACX with acrylamide and N-isopropylacrylamide under ultraviolet irradiation. The size of the Fe3O4 fabricated within the hydrogel matrix could be adjusted through controlling the crosslinking concentrations (C. The magnetic hydrogels showed desirable magnetic and mechanical properties, which were confirmed by XRD, Raman spectroscopy, physical property measurement system, SEM, TGA, and compression test. Moreover, the catalytic performance of the magnetic hydrogels was explored. The magnetic hydrogels (C = 7.5 wt % presented excellent catalytic activity and provided a sensitive response to H2O2 detection even at a concentration level of 5 × 10−6 mol·L−1. This approach to preparing magnetic hydrogels loaded with Fe3O4 nanoparticles endows xylan-based hydrogels with new promising applications in biotechnology and environmental chemistry.

  19. Lysine Acetylation and Deacetylation in Brain Development and Neuropathies.

    Science.gov (United States)

    Tapias, Alicia; Wang, Zhao-Qi

    2017-02-01

    Embryonic development is critical for the final functionality and maintenance of the adult brain. Brain development is tightly regulated by intracellular and extracellular signaling. Lysine acetylation and deacetylation are posttranslational modifications that are able to link extracellular signals to intracellular responses. A wealth of evidence indicates that lysine acetylation and deacetylation are critical for brain development and functionality. Indeed, mutations of the enzymes and cofactors responsible for these processes are often associated with neurodevelopmental and psychiatric disorders. Lysine acetylation and deacetylation are involved in all levels of brain development, starting from neuroprogenitor survival and proliferation, cell fate decisions, neuronal maturation, migration, and synaptogenesis, as well as differentiation and maturation of astrocytes and oligodendrocytes, to the establishment of neuronal circuits. Hence, fluctuations in the balance between lysine acetylation and deacetylation contribute to the final shape and performance of the brain. In this review, we summarize the current basic knowledge on the specific roles of lysine acetyltransferase (KAT) and lysine deacetylase (KDAC) complexes in brain development and the different neurodevelopmental disorders that are associated with dysfunctional lysine (de)acetylation machineries. Copyright © 2017 The Authors. Production and hosting by Elsevier Ltd.. All rights reserved.

  20. Histones of Chlamydomonas reinhardtii. Synthesis, acetylation, and methylation

    International Nuclear Information System (INIS)

    Waterborg, J.H.; Robertson, A.J.; Tatar, D.L.; Borza, C.M.; Davie, J.R.

    1995-01-01

    Histones of the green alga Chlamydomonas reinhardtii were prepared by a new method and fractionated by reversed-phase high-performance liquid chromatography. Acid-urea-Triton gel analysis and tritiated acetate labeling demonstrated high levels of steady-state acetylation for the single histone H3 protein, in contrast to low levels on histones H4 and H2B. Twenty percent of histone H3 is subject to dynamic acetylation with, on average, three acetylated lysine residues per protein molecule. Histone synthesis in light-dark-synchronized cultures was biphasic with pattern differences between two histone H1 variants, between two H2A variants, and between H2B and ubiquitinated H2B. Automated protein sequence analysis of histone H3 demonstrated a site-specific pattern of steady-state acetylation between 7 and 17% at five of the six amino-terminal lysines and of monomethylation between 5 and 81% at five of the eight amino-terminal lysines in a pattern that may limit dynamic acetylation. An algal histone H3 sequence was confirmed by protein sequencing with a since threonine as residue 28 instead of the serine(28)-alanine(29) sequence, present in all other known plant and animal H3 histones

  1. Relationship of histone acetylation to DNA topology and transcription.

    Science.gov (United States)

    Krajewski, W A; Luchnik, A N

    1991-12-01

    An autonomously replicating plasmid constructed from bovine papiloma virus (BPV) and pBR322 was stably maintained as a nuclear episome in a mouse cell culture. Addition to a cell culture of sodium butyrate (5 mM) induced an increase in plasmid DNA supercoiling of 3-5 turns, an increase in acetylation of cellular histones, and a decrease in plasmid transcription by 2- to 4-fold. After withdrawal of butyrate, DNA supercoiling began to fluctuate in a wave-like manner with an amplitude of up to 3 turns and a period of 3-4 h. These waves gradually faded by 24 h. The transcription of the plasmid and acetylation of cellular histones also oscillated with the same period. The wave-like alterations were not correlated with the cell cycle, for there was no resumption of DNA replication after butyrate withdrawal for at least 24 h. In vitro chemical acetylation of histones with acetyl adenylate also led to an increase in the superhelical density of plasmid DNA. The parallel changes in transcription, histone acetylation, and DNA supercoiling in vivo may indicate a functional innerconnection. Also, the observed in vivo variation in the level of DNA supercoiling directly indicates the possibility of its natural regulation in eukaryotic cells.

  2. Characterisation of esterases as potential biomarkers of pesticide exposure in the lugworm Arenicola marina (Annelida: Polychaeta)

    International Nuclear Information System (INIS)

    Hannam, Marie L.; Hagger, Josephine A.; Jones, Malcolm B.; Galloway, Tamara S.

    2008-01-01

    Here, we identify and characterise cholinesterase (ChE) and carboxylesterase (CbE) activities in the body tissues of the sediment dwelling worm Arenicola marina. Exposure to the organophosphorus pesticide azamethiphos yielded an in vitro IC 50 of 5 μg l -1 for propionylcholinesterase (PChE). PChE was significantly inhibited in vivo after a 10 day exposure to 100 μg l -1 azamethiphos, equivalent to the recommended aquatic application rate (ANOVA; F = 2.75, P = 0.033). To determine sensitivity to environmental conditions, A. marina were exposed for 10 days to field collected sediments. PChE activity was significantly lower in worms exposed to sediments from an estuary classified to be at high risk from point source pollution by the UK Environment Agency (ANOVA; F = 15.33, P < 0.001). Whilst causality cannot be directly attributed from these latter exposures, they provide an important illustration of the potential utility of esterase activity as a biomarker of environmental quality in this ecologically relevant sentinel species. - This paper provides a preliminary characterisation of esterase enzyme activities in the tissues and body fluids of the sediment dwelling worm Arenicola marina and explores their potential use as biomarkers of organophosphorus pesticide exposure in the marine environment

  3. Manipulation of intracellular auxin in a single cell by light with esterase-resistant caged auxins.

    Science.gov (United States)

    Kusaka, Naoyuki; Maisch, Jan; Nick, Peter; Hayashi, Ken-ichiro; Nozaki, Hiroshi

    2009-09-04

    Auxin, a plant hormone, is polar transported from its site of production. This auxin polar transport system establishes an auxin gradient in plant tissue that is necessary for proper plant development. Therefore, the spatial effect of the auxin gradient on plant development is highly important for the understanding of plant auxin responses. Herein we report the design, syntheses and biological properties of esterase-resistant caged auxins. The conventional caging group, 2-nitrobenzyl ester, was found to be enzymatically hydrolyzed in plant cells and released original auxin without photolysis. The esterase-resistant caging group, (2,5-dimethoxyphenyl)(2-nitrobenzyl) ester, (DMPNB) was designed to improve the stability of caged auxins. Three auxins, indole 3-acetic acid, naphthalene 1-acetic acid and 2,4-dichlorophenoxy acetic acid were caged with the DMPNB caging group. DMPNB-caged auxins were inactive within a plant cell until photolysis, but they release auxins with photoirradiation to activate auxin-responsive gene expression. We demonstrated spatial and temporal control of intracellular auxin levels with photoirradiation by using this caged auxin system and were able to photocontrol the physiological auxin response in Arabidopsis plants. Additionally, the photoirradiation of DMPNB-caged auxin within a single cell can manipulate the intracellular auxin level and triggers auxin response.

  4. Analysis of esterase isozyme and SSR for mutagenic progenies induced by space mutation in mustard

    International Nuclear Information System (INIS)

    Shen Jinjuan; Liu Yihua; Zhang Zhaorong; Ran Guangkui; Zhao Shouzhong; Xiao Li

    2012-01-01

    Seeds of five mustard (Brassica juncea Coss) varieties were carried into outer space by 'Shijian No.8' satellite. After five years' consecutive planting and selection, ten relatively stable mutant lines were obtained, which had significant variation in agronomic and economic characters. The mutant lines and their original varieties without space mutation treatment as control were studied by esterase isozyme and SSR analyses. Electrophoresis analysis of esterase isozymes indicated that there were differences between mutant lines and their controls in enzyme types and enzyme activity. Different mustard varieties had different enzymographs, and so did the mutants induced by space mutation, which shows different sensitivity among different mustard varieties. The SSR analysis showed that large differences were found in the SSR loci between mutant lines and their original variety, the variation frequency was between 9.52% and 57.14% with an average frequency of 26.19% for all the mutant lines. Among the mutant SSR loci, about 56.36% showed changes in band number and 43.64% in molecular weight. These results indicated that the ten mutant lines had large genetic difference in phenotype, genomic sequence and gene expression, and the outer space mutation would be an effective method to develop new mustard germplasm and variety. (authors)

  5. Characterisation of esterases as potential biomarkers of pesticide exposure in the lugworm Arenicola marina (Annelida: Polychaeta)

    Energy Technology Data Exchange (ETDEWEB)

    Hannam, Marie L. [Ecotoxicology and Stress Biology Research Centre, School of Biological Sciences, University of Plymouth, Drake Circus, Plymouth, Devon, PL4 8AA (United Kingdom)], E-mail: marie.hannam@plymouth.ac.uk; Hagger, Josephine A.; Jones, Malcolm B.; Galloway, Tamara S. [Ecotoxicology and Stress Biology Research Centre, School of Biological Sciences, University of Plymouth, Drake Circus, Plymouth, Devon, PL4 8AA (United Kingdom)

    2008-03-15

    Here, we identify and characterise cholinesterase (ChE) and carboxylesterase (CbE) activities in the body tissues of the sediment dwelling worm Arenicola marina. Exposure to the organophosphorus pesticide azamethiphos yielded an in vitro IC{sub 50} of 5 {mu}g l{sup -1} for propionylcholinesterase (PChE). PChE was significantly inhibited in vivo after a 10 day exposure to 100 {mu}g l{sup -1} azamethiphos, equivalent to the recommended aquatic application rate (ANOVA; F = 2.75, P = 0.033). To determine sensitivity to environmental conditions, A. marina were exposed for 10 days to field collected sediments. PChE activity was significantly lower in worms exposed to sediments from an estuary classified to be at high risk from point source pollution by the UK Environment Agency (ANOVA; F = 15.33, P < 0.001). Whilst causality cannot be directly attributed from these latter exposures, they provide an important illustration of the potential utility of esterase activity as a biomarker of environmental quality in this ecologically relevant sentinel species. - This paper provides a preliminary characterisation of esterase enzyme activities in the tissues and body fluids of the sediment dwelling worm Arenicola marina and explores their potential use as biomarkers of organophosphorus pesticide exposure in the marine environment.

  6. Inhibition of Mitochondrial Bioenergetics by Esterase-Triggered COS/H2S Donors.

    Science.gov (United States)

    Steiger, Andrea K; Marcatti, Michela; Szabo, Csaba; Szczesny, Bartosz; Pluth, Michael D

    2017-08-18

    Hydrogen sulfide (H 2 S) is an important biological mediator, and synthetic H 2 S donating molecules provide an important class of investigative tools for H 2 S research. Here, we report esterase-activated H 2 S donors that function by first releasing carbonyl sulfide (COS), which is rapidly converted to H 2 S by the ubiquitous enzyme carbonic anhydrase (CA). We report the synthesis, self-immolative decomposition, and H 2 S release profiles of the developed scaffolds. In addition, the developed esterase-triggered COS/H 2 S donors exhibit higher levels of cytotoxicity than equivalent levels of Na 2 S or the common H 2 S donors GYY4137 and AP39. Using cellular bioenergetics measurements, we establish that the developed donors reduce cellular respiration and ATP synthesis in BEAS 2B human lung epithelial cells, which is consistent with COS/H 2 S inhibition of cytochrome c oxidase in the mitochondrial respiratory chain although not observed with common H 2 S donors at the same concentrations. Taken together, these results may suggest that COS functions differently than H 2 S in certain biological contexts or that the developed donors are more efficient at delivering H 2 S than other common H 2 S-releasing motifs.

  7. Hydrophobic-ionic chromatography: its application to microbial glucose oxidase, hyaluronidase, cholesterol oxidase, and cholesterol esterase.

    Science.gov (United States)

    Sasaki, I; Gotoh, H; Yamamoto, R; Tanaka, H; Takami, K; Yamashita, K; Yamashita, J; Horio, T

    1982-05-01

    Glucose oxidase from Aspergillus niger, hyaluronidase from Streptomyces hyalurolyticus, and cholesterol oxidase and cholesterol esterase from Pseudomonas fluorescens were effectively adsorbed on an Amberlite CG-50 column, when the cell-free cultured medium or the cultured medium with cell extract and without cell debris was applied without desalting but at pH less than or equal to 4.5. At the acidic pH, all the ion-exchange groups (-COOH) exist in the protonated form; the adsorption is not due to electrostatic attraction, but to hydrophobic interaction. The enzymes thus adsorbed were effectively eluted by increasing pH, at which the ion-exchange groups became dissociated. This type of adsorption-elution is called hydrophobic-ionic chromatography. By a single run of chromatography, glucose oxidase, hyaluronidase, cholesterol oxidase, and cholesterol esterase were purified 30-fold, 12-fold, 45-fold, and 20-fold with yields of 82%, 83%, 80%, and 90%, respectively. This indicates that hydrophobic-ionic chromatography on an Amberlite CG-50 column is effective for the purification of various enzymes, provided that they are stable at the acidic pH.

  8. Target size of neurotoxic esterase and acetylcholinesterase as determined by radiation inactivation.

    Science.gov (United States)

    Carrington, C D; Fluke, D J; Abou-Donia, M B

    1985-11-01

    The target size of neurotoxic esterase (NTE), the putative target site for the initiation of organophosphorus-compound-induced delayed neurotoxicity, and acetylcholinesterase (AChE) from hen brain were examined by determining the rate at which the activities of the esterases were destroyed by ionizing irradiation. Samples of hen brain were prepared by slowly drying a microsomal preparation under vacuum. The dried samples were then irradiated with electrons from a 1 MeV Van de Graaff generator. The doses ranged from 0 to 28 Mrad. The radiation doses were calibrated by the rate of inactivation of T1-bacteriophage plaque induction. Following the irradiation procedure, the samples were resuspended in buffer and enzymic activity was measured. The target size of NTE from hen brain was determined to be about 105 kDa, whereas hen brain AChE was found to have a target size of about 53 kDa. The target size of NTE was found to be similar in experiments with rat brain and cat brain. In addition, commercial preparations of electric-eel electric-organ AChE and horse serum butyrylcholinesterase were found to have target sizes that were identical with each other, and also were very similar to that of AChE from hen brain.

  9. Crystal structure of human esterase D: a potential genetic marker of retinoblastoma

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Dong; Li, Yang; Song, Gaojie; Zhang, David; Shaw, Neil; Liu, Zhi-Jie; (Chinese Aca. Sci.)

    2009-07-10

    Retinoblastoma (RB), a carcinoma of the retina, is caused by mutations in the long arm of chromosome 13, band 13q14. The esterase D (ESD) gene maps at a similar location as the RB gene locus and therefore serves as a potential marker for the prognosis of retinoblastoma. Because very little is known about the structure and function of ESD, we determined the 3-dimensional structure of the enzyme at 1.5 {angstrom} resolution using X-ray crystallography. ESD shows a single domain with an {alpha}/{beta}-hydrolase fold. A number of insertions are observed in the canonical {alpha}/{beta}-hydrolase fold. The active site is located in a positively charged, shallow cleft on the surface lined by a number of aromatic residues. Superimposition studies helped identify the typical catalytic triad residues -- Ser-153, His264, and Asp230 -- involved in catalysis. Mutagenesis of any of the catalytic triad residues to alanine abolished the enzyme activity. Backbone amides of Leu54 and Met150 are involved in the formation of the oxyanion hole. Interestingly, a M150A mutation increased the enzyme activity by 62%. The structure of human ESD determined in this study will aid the elucidation of the physiological role of the enzyme in the human body and will assist in the early diagnosis of retinoblastoma. Wu, D., Li, Y., Song, G., Zhang, D., Shaw, N., Liu, Z. J. Crystal structure of human esterase D: a potential genetic marker of retinoblastoma.

  10. 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

    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...

  11. Synthetic biology for engineering acetyl coenzyme a metabolism in yeast

    DEFF Research Database (Denmark)

    Nielsen, Jens

    2014-01-01

    The yeast Saccharomyces cerevisiae is a widely used cell factory for the production of fuels, chemicals, and pharmaceuticals. The use of this cell factory for cost-efficient production of novel fuels and chemicals requires high yields and low by-product production. Many industrially interesting...... chemicals are biosynthesized from acetyl coenzyme A (acetyl-CoA), which serves as a central precursor metabolite in yeast. To ensure high yields in production of these chemicals, it is necessary to engineer the central carbon metabolism so that ethanol production is minimized (or eliminated) and acetyl......-CoA can be formed from glucose in high yield. Here the perspective of generating yeast platform strains that have such properties is discussed in the context of a major breakthrough with expression of a functional pyruvate dehydrogenase complex in the cytosol....

  12. In-gel detection of esterase-like albumin activity: Characterization of esterase-free sera albumin and its putative role as non-invasive biomarker of hepatic fibrosis

    Directory of Open Access Journals (Sweden)

    Areeba Ahmad

    2017-07-01

    Full Text Available Albumin is a globular and un-glycosylated multifunctional plasma protein and thus correlated with several human diseases. Owing to esterase contamination, albumin levels are usually misleading. In this study, we propose methodical accuracy for albumin estimation taking healthy and fibrotic rats. Liver fibrosis in rats was generated by N′-Nitrosodimethylamine (NDMA (10 mg/kg body weight within three weeks followed by its confirmation through H&E and immunohistochemical staining for α-SMA expression. Animal sera were screened by native polyacrylamide gel electrophoresis (native-PAGE (7.5%. In-gel esterase-like albumin activity was detected using α- and β-naphthyl acetate (5.58 × 10−3 mM; pH 7.5 as substrate. Sera albumin was purified from unstained PA gel-slices through electroelution. Subsequent to conformation of albumin purity by its molecular weight determination using SDS–PAGE (10% and peptide mass fingerprinting by MALDI-TOF-MS, samples were treated with different concentrations of urea. Urea-treated albumins were screened for esterase activity, conformational change and, albumin levels by immunoblotting. Our results demonstrate that esterase-like albumin activity in rat sera albumin is located in domain-III. The esterase-like activity remains detectable up to 4 M urea, which diminishes with increasing urea concentrations. Further, immunoblotting of urea-treated albumin samples displays a significant decline in purified protein bands, indicating hypoalbuminemia during hepatic fibrosis in rats. In conclusion, the present approach of albumin separation and estimation is of potential interest and may be recommended for diagnostic purposes.

  13. Nebulized C1-Esterase Inhibitor does not Reduce Pulmonary Complement Activation in Rats with Severe Streptococcus Pneumoniae Pneumonia

    NARCIS (Netherlands)

    de Beer, Friso; Lagrand, Wim; Glas, Gerie J.; Beurskens, Charlotte J. P.; van Mierlo, Gerard; Wouters, Diana; Zeerleder, Sacha; Roelofs, Joris J. T. H.; Juffermans, Nicole P.; Horn, Janneke; Schultz, Marcus J.

    2016-01-01

    Complement activation plays an important role in the pathogenesis of pneumonia. We hypothesized that inhibition of the complement system in the lungs by repeated treatment with nebulized plasma-derived human C1-esterase inhibitor reduces pulmonary complement activation and subsequently attenuates

  14. Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm

    Energy Technology Data Exchange (ETDEWEB)

    Harholt, Jesper; Bach, Inga C; Lind-Bouquin, Solveig; Nunan, Kylie J.; Madrid, Susan M.; Brinch-Pedersen, Henrik; Holm, Preben B.; Scheller, Henrik V.

    2009-12-08

    Endo-xylanase (from Bacillus subtilis) or ferulic acid esterase (from Aspergillus niger) were expressed in wheat under the control of the endosperm specific 1DX5 glutenin promoter. Constructs both with and without the endoplasmic reticulum retention signal KDEL were used. Transgenic plants were recovered in all four cases but no qualitative differences could be observed whether KDEL was added or not. Endo-xylanase activity in transgenic grains was increased between two and three fold relative to wild type. The grains were shriveled and had a 25-33% decrease in mass. Extensive analysis of the cell walls showed a 10-15% increase in arabinose to xylose ratio, a 50% increase in the proportion of water extractable arabinoxylan, and a shift in the MW of the water extractable arabinoxylan from being mainly larger than 85 kD to being between 2 kD and 85 kD. Ferulic acid esterase expressing grains were also shriveled and the seed weight was decreased by 20-50%. No ferulic acid esterase activity could be detected in wild type grains whereas ferulic acid esterase activity was detected in transgenic lines. The grain cell walls had 15-40% increase in water unextractable arabinoxylan and a decrease in monomeric ferulic acid between 13 and 34%. In all the plants the observed changes are consistent with a plant response that serves to minimize the effect of the heterologously expressed enzymes by increasing arabinoxylan biosynthesis and cross-linking.

  15. Statistical optimization of medium components and physicochemical parameters to simultaneously enhance bacterial growth and esterase production by Bacillus thuringiensis.

    Science.gov (United States)

    Mazzucotelli, Cintia Anabela; Moreira, María del Rosario; Ansorena, María Roberta

    2016-01-01

    Bacillus thuringiensis is a genus extensively studied because of its high potential for biotechnological application, principally in biocontrol techniques. However, the optimization of esterase production by this strain has been scarcely studied. The aim of this work was to select and optimize the physicochemical and nutritional parameters that significantly influence the growth and esterase production of B. thuringiensis. To this purpose, 6 nutritional factors and 2 physicochemical parameters were evaluated using a Plackett-Burman design. Significant variables were optimized using a Box-Behnken design and through the desirability function to select the levels of the variables that simultaneously maximize microbial growth and esterase production. The optimum conditions resulting from simultaneous optimization of the responses under study were found to be 1 g/L glucose, 15 g/L peptone, and 3.25 g/L NaCl. Under these optimal conditions, it was possible to achieve a 2.5 log CFU/mL increase in bacterial growth and a 113-fold increase in esterase productivity, compared with minimal medium without agitation.

  16. Influence of ammonium salts on the lipase/esterase activity assay using p-nitrophenyl esters as substrates.

    Science.gov (United States)

    De Yan, Hong; Zhang, Yin Jun; Liu, Hong Cai; Zheng, Jian Yong; Wang, Zhao

    2013-01-01

    p-Nitrophenyl esters with a short-chain carboxylic group, such as p-nitrophenyl acetate (p-NPA) and p-nitrophenyl butyrate (p-NPB), could be effectively hydrolyzed by ammonium salts. p-Nitrophenyl esters were usually used as substrates to assay the lipase/esterase activity. Ammonium sulfate precipitation was often used to purify proteins, and some ammonium salts were usually used as nitrogen sources or inorganic salts for the lipase/esterase production. To study the effect of ammonium salts on the assay of the lipase/esterase activity, the contributing factors of hydrolysis of p-NPA/p-NPB catalyzed by ammonium salts were investigated. The lipase activities were compared in the presence and absence of ammonium sulfate. The hydrolysis reaction could be catalyzed under neutral and alkaline circumstances. The hydrolysis rate increased with the increase in the reaction temperature or the concentration of ammonium ion. When p-NPA was employed as the substrate for the analysis of the lipase/esterase activity, the effect of ammonium sulfate on the analysis could be neutralized by setting a control when the concentration of ammonium sulfate was less than 40% saturation. However, when the concentration of ammonium sulfate increased from 40% to 100% saturation, the enzyme activities decreased about 13-40%, which could not be ignored for accurate analysis of the enzyme activity. © 2013 International Union of Biochemistry and Molecular Biology, Inc.

  17. Design and production in Aspergillus niger of a chimeric protein associating a fungal feruloyl esterase and a clostridial dockerin domain

    NARCIS (Netherlands)

    Levasseur, A.; Pagès, S.; Fierobe, H.-P.; Navarro, D.; Punt, P.; Belaïch, J.-P.; Asther, M.; Record, E.

    2004-01-01

    A chimeric enzyme associating feruloyl esterase A (FAEA) from Aspergilhis niger and dockerin from Clostridium thermocellum was produced in A. niger. A completely truncated form was produced when the dockerin domain was located downstream of the FAEA (FAEA-Doc), whereas no chimeric protein was

  18. Crystallization and preliminary crystallographic analysis of an esterase with a novel domain from the hyperthermophile Thermotoga maritima

    NARCIS (Netherlands)

    Sun, Lei; Levisson, Mark; Hendriks, Sjon; Akveld, Twan; Kengen, Serve W. M.; Dijkstra, Bauke W.; van der Oost, John

    A predicted esterase ( EstA) with an unusual new domain from the hyperthermophilic bacterium Thermotoga maritima has been cloned and overexpressed in Escherichia coli. The purified protein was crystallized by the hanging-drop vapour-diffusion technique in the presence of lithium sulfate and

  19. Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm

    DEFF Research Database (Denmark)

    Harholt, Jesper; Bach, Inga Christensen; Lind Bouquin, Solveig

    2010-01-01

    Endo-xylanase (from Bacillus subtilis) or ferulic acid esterase (from Aspergillus niger) were expressed in wheat under the control of the endosperm-specific 1DX5 glutenin promoter. Constructs both with and without the endoplasmic reticulum retention signal (Lys-Asp-Glu-Leu) KDEL were used....... Extensive analysis of the cell walls showed a 10%-15% increase in arabinose to xylose ratio, a 50% increase in the proportion of water-extractable arabinoxylan, and a shift in the MW of the water-extractable arabinoxylan from being mainly larger than 85 kD to being between 2 and 85 kD. Ferulic acid esterase......-expressing grains were also shrivelled, and the seed weight was decreased by 20%-50%. No ferulic acid esterase activity could be detected in wild-type grains whereas ferulic acid esterase activity was detected in transgenic lines. The grain cell walls had 15%-40% increase in water-unextractable arabinoxylan...

  20. Evaluation of the nitrite and leukocyte esterase activity tests for the diagnosis of acute symptomatic urinary tract infection in men.

    NARCIS (Netherlands)

    Koeijers, J.J.; Kessels, A.G.H.; Nys, S.; Bartelds, A.; Donker, G.; Stobberingh, E.; Verbon, A.

    2007-01-01

    For 422 male patients with symptoms indicative of a urinary tract infection, nitrite and leukocyte esterase activity dipstick test results were compared with results of culture of urine samples. The positive predictive value of a positive nitrite test result was 96%. Addition of results of the

  1. Spatial distribution and esterase activity in populations of Aedes (Stegomyia aegypti (Linnaeus (Diptera: Culicidae resistant to temephos

    Directory of Open Access Journals (Sweden)

    Wanessa Porto Tito Gambarra

    2013-04-01

    Full Text Available INTRODUCTION: The need for studies that describe the resistance patterns in populations of Aedes aegypti (Linnaeus in function of their region of origin justified this research, which aimed to characterize the resistance to temephos and to obtain information on esterase activity in populations of Aedes aegypti collected in municipalities of the State of Paraíba. METHODS: Resistance to temephos was evaluated and characterized from the diagnostic dose of 0.352mg i.a./L and multiple concentrations that caused mortalities between 5% and 99%. Electrophoresis of isoenzymes was used to verify the patterns of esterase activity among populations of the vector. RESULTS: All populations of Aedes aegypti were resistant to temephos, presenting a resistance rate (RR greater than 20. The greatest lethal dose 50% of the sample (CL50 was found for the municipality of Lagoa Seca, approximately forty-one times the value of CL50 for the Rockefeller population. The populations characterized as resistant showed two to six regions of α and β-esterase, called EST-1 to EST-6, while the susceptible population was only seen in one region of activity. CONCLUSIONS: Aedes aegypti is widely distributed and shows a high degree of resistance to temephos in all municipalities studied. In all cases, esterases are involved in the metabolism and, consequently, in the resistance to temephos.

  2. The ferulic acid esterases of Chrysosporium lucknowense C1: Purification, characterization and their potential application in biorefinery

    NARCIS (Netherlands)

    Kuhnel, S.; Pouvreau, L.A.M.; Appeldoorn, M.M.; Hinz, S.W.A.; Schols, H.A.; Gruppen, H.

    2012-01-01

    Three ferulic acid esterases from the filamentous fungus Chrysosporium lucknowense C1 were purified and characterized. The enzymes were most active at neutral pH and temperatures up to 45 °C. All enzymes released ferulic acid and p-coumaric acid from a soluble corn fibre fraction. Ferulic acid

  3. Cloning and sequence analysis of cDNA encoding a putative juvenile hormone esterase from the Colorado potato beetle.

    NARCIS (Netherlands)

    Vermunt, A.M.W.; Koopmanschap, A.B.; Vlak, J.M.; Kort, de C.A.D.

    1998-01-01

    In the Colorado potato beetle, Leptinotarsa decemlineata, reproduction and diapause are mediated by the juvenile hormone (JH) titer in the hemolymph. This titer is controlled by JH synthesis in the corpora allata and by JH degradation. The main pathway of JH degradation is by JH esterase in the

  4. 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

  5. Pig Liver Esterase (PLE) as Biocatalyst in Organic Synthesis: From Nature to Cloning and to Practical Applications

    NARCIS (Netherlands)

    Dominguez de Maria, Pablo; Garcia-Burgos, Carlos A.; Bargeman, Gerrald; van Gemert, Robert W.

    2007-01-01

    Pig liver esterase (PLE, EC 3.1.1.1) has been employed extensively for research purposes during the last three decades, especially in kinetic resolutions, in desymmetrizations of prochiral substrates, and in the synthesis of nucleosides. Its practical use, however, has been traditionally hampered

  6. Novel ferulate esterase from Gram-positive lactic acid bacteria and analyses of the recombinant enzyme produced in E. coli

    Science.gov (United States)

    Using a plate containing ethyl ferulate as sole carbon source, various bacteria cultures were screened for ferulate esterase (FAE). Among a dozen of species showing positive FAE, one Lactobacillus fermentum strain NRRL 1932 demonstrated the strongest activity. Using a published sequence of ferulate ...

  7. 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...

  8. ISOLATION OF JUVENILE HORMONES ESTERASE AND ITS PARTIAL CDNA CLONE FROM THE BEETLE, TENEBRIO MOLITOR. (R825433)

    Science.gov (United States)

    Juvenile hormone esterase (JHE) plays an essential role in insect development. It is partially responsible for the clearance of juvenile hormone (JH) which regulates various aspects of insect development and reproduction. Because of its role in regulating JH titer, this enzyme...

  9. An Esterase with Superior Activity and Enantioselectivity towards 1,2-O-Isopropylideneglycerol Esters Obtained by Protein Design

    NARCIS (Netherlands)

    Godinho, Luis F.; Reis, C.R.; van Merkerk, Ronald; Poelarends, Gerrit J.; Quax, Wim J.

    2012-01-01

    The Escherichia coli esterase YbfF displays high activity towards 1,2-O-isopropylideneglycerol (IPG) butyrate and IPG caprylate, and prefers the R-enantiomer of these substrates, producing the S-enantiomer of the IPG product in excess. To improve the potential of the enzyme for the kinetic

  10. Effects of N-acetylation degree on N-acetylated chitosan hydrolysis with commercially available and modified pectinases.

    Science.gov (United States)

    Shin-ya; Lee; Hinode; Kajiuchi

    2001-01-01

    Three types of N-acetylated chitosans (NACs) with different degrees of acetylation (DA) were prepared and used as a substrate for enzymatic hydrolysis with a commercially available pectinase and a modified one. Pectinase modification was conducted using polyalkyleneoxide-maleic anhydride copolymer (PEO-MA copolymer). The effects of DA on enzymatic reaction with native and modified pectinases were investigated experimentally. Initial hydrolysis rate and Michaelis-Menten kinetic parameters were measured by analysis of reducing sugars. DA of NAC strongly affected the hydrolytic characteristics of native and modified pectinases. N-acetylation of chitosan increased the initial hydrolysis rate and the enzyme-substrate affinity with respect to both pectinases: NACs with DA over 0.3 showed high initial hydrolysis rate and strong affinity between enzyme and substrate. Especially, when NAC with DA over 0.3 was treated with modified pectinase, the affinity became much stronger than the native pectinase.

  11. EVALUATION OF LEUKOCYTE ESTERASE REAGENT STRIPS TEST IN THE DIAGNOSIS OF SPONTANEOUS BACTERIAL PERITONITIS IN CHILDREN WITH CIRRHOSIS

    Directory of Open Access Journals (Sweden)

    Naser HONAR

    2015-09-01

    Full Text Available BackgroundSpontaneous bacterial peritonitis is defined as an ascetic fluid infection without an evident intra-abdominal surgically treatable source. Spontaneous bacterial peritonitis is one of the severe complications in patients with cirrhosis and ascites. Without early antibiotic treatment, this complication is associated with high mortality rate; therefore, early diagnosis and treatment of spontaneous bacterial peritonitis is necessary for survival. Leukocyte esterase reagent can rapidly diagnose the spontaneous bacterial peritonitis.ObjectiveThis study aimed to find out the diagnostic accuracy of leukocyte esterase dipstick test for the diagnosis of spontaneous bacterial peritonitis.MethodsA single centered hospital-based cross-sectional study was conducted during July 2013 to August 2014 on children with cirrhotic liver disease and ascites who were admitted in the Department of Pediatric Gastroenterology in Nemazee Hospital affiliated to Shiraz University of Medical Sciences (Iran. All patients underwent abdominal paracentesis, and the ascitic fluid was processed for cell count, leukocyte esterase reagent strip test (Combiscreen SL10 and culture. Spontaneous bacterial peritonitis was defined as having a polymorphonuclear count (PMN ≥250/m3 in ascitic fluid. Sensitivity, specificity, positive predictive value and negative predictive value of leukocyte esterase test were calculated according to the formula.ResultsTotally, 150 ascitic fluid sample of cirrhotic male patients (53.2% and their mean age (4.33±1.88 years were analyzed. Biliary atresia (n=44, 29.4% and idiopathic neonatal hepatitis (n=29, 19.3% were the most frequent etiology of cirrhosis. Also, abdominal pain (68.6% and distension (64% were the most common presenting complaint. Of all cases, 41patients (27.35% were diagnosed to have spontaneous bacterial peritonitis (PMN ≥250/mm3. Sensitivity and specificity of leukocyte esterase reagent test according to PMNs ≥250mm3 were

  12. The active site of a carbohydrate esterase displays divergent catalytic and noncatalytic binding functions.

    Directory of Open Access Journals (Sweden)

    Cedric Montanier

    2009-03-01

    Full Text Available Multifunctional proteins, which play a critical role in many biological processes, have typically evolved through the recruitment of different domains that have the required functional diversity. Thus the different activities displayed by these proteins are mediated by spatially distinct domains, consistent with the specific chemical requirements of each activity. Indeed, current evolutionary theory argues that the colocalization of diverse activities within an enzyme is likely to be a rare event, because it would compromise the existing activity of the protein. In contrast to this view, a potential example of multifunctional recruitment into a single protein domain is provided by CtCel5C-CE2, which contains an N-terminal module that displays cellulase activity and a C-terminal module, CtCE2, which exhibits a noncatalytic cellulose-binding function but also shares sequence identity with the CE2 family of esterases. Here we show that, unlike other CE2 members, the CtCE2 domain displays divergent catalytic esterase and noncatalytic carbohydrate binding functions. Intriguingly, these diverse activities are housed within the same site on the protein. Thus, a critical component of the active site of CtCE2, the catalytic Ser-His dyad, in harness with inserted aromatic residues, confers noncatalytic binding to cellulose whilst the active site of the domain retains its esterase activity. CtCE2 catalyses deacetylation of noncellulosic plant structural polysaccharides to deprotect these substrates for attack by other enzymes. Yet it also acts as a cellulose-binding domain, which promotes the activity of the appended cellulase on recalcitrant substrates. The CE2 family encapsulates the requirement for multiple activities by biocatalysts that attack challenging macromolecular substrates, including the grafting of a second, powerful and discrete noncatalytic binding functionality into the active site of an enzyme. This article provides a rare example of

  13. Analysing deltamethrin susceptibility and pyrethroid esterase activity variations in sylvatic and domestic Triatoma infestans at the embryonic stage

    Science.gov (United States)

    Santo-Orihuela, Pablo Luis; Carvajal, Guillermo; Picollo, María Inés; Vassena, Claudia Viviana

    2013-01-01

    The aim of the present work was to study the deltamethrin susceptibility of eggs from Triatoma infestans populations and the contribution of pyrethroid esterases to deltamethrin degradation. Insects were collected from sylvatic areas, including Veinte de Octubre and Kirus-Mayu (Bolivia) and from domiciliary areas, including El Palmar (Bolivia) and La Pista (Argentina). Deltamethrin susceptibility was determined by dose-response bioassays. Serial dilutions of deltamethrin (0.0005-1 mg/mL) were topically applied to 12-day-old eggs. Samples from El Palmar had the highest lethal dose ratio (LDR) value (44.90) compared to the susceptible reference strain (NFS), whereas the Veinte de Octubre samples had the lowest value (0.50). Pyrethroid esterases were evaluated using 7-coumaryl permethrate (7-CP) on individually homogenised eggs from each population and from NFS. The El Palmar and La Pista samples contained 40.11 and 36.64 pmol/min/mg protein, respectively, and these values were statistically similar to NFS (34.92 pmol/min/mg protein) and different from Kirus-Mayu and Veinte de Octubre (27.49 and 22.69 pmol/min/mg protein, respectively). The toxicological data indicate that the domestic populations were resistant to deltamethrin, but no statistical contribution of 7-CP esterases was observed. The sylvatic populations had similar LDR values to NFS, but lower 7-CP esterase activities. Moreover, this is the first study of the pyrethroid esterases on T. infestans eggs employing a specific substrate (7-CP). PMID:24402155

  14. Analysing deltamethrin susceptibility and pyrethroid esterase activity variations in sylvatic and domestic Triatoma infestans at the embryonic stage

    Directory of Open Access Journals (Sweden)

    Pablo Luis Santo-Orihuela

    2013-12-01

    Full Text Available The aim of the present work was to study the deltamethrin susceptibility of eggs from Triatoma infestans populations and the contribution of pyrethroid esterases to deltamethrin degradation. Insects were collected from sylvatic areas, including Veinte de Octubre and Kirus-Mayu (Bolivia and from domiciliary areas, including El Palmar (Bolivia and La Pista (Argentina. Deltamethrin susceptibility was determined by dose-response bioassays. Serial dilutions of deltamethrin (0.0005-1 mg/mL were topically applied to 12-day-old eggs. Samples from El Palmar had the highest lethal dose ratio (LDR value (44.90 compared to the susceptible reference strain (NFS, whereas the Veinte de Octubre samples had the lowest value (0.50. Pyrethroid esterases were evaluated using 7-coumaryl permethrate (7-CP on individually homogenised eggs from each population and from NFS. The El Palmar and La Pista samples contained 40.11 and 36.64 pmol/min/mg protein, respectively, and these values were statistically similar to NFS (34.92 pmol/min/mg protein and different from Kirus-Mayu and Veinte de Octubre (27.49 and 22.69 pmol/min/mg protein, respectively. The toxicological data indicate that the domestic populations were resistant to deltamethrin, but no statistical contribution of 7-CP esterases was observed. The sylvatic populations had similar LDR values to NFS, but lower 7-CP esterase activities. Moreover, this is the first study of the pyrethroid esterases on T. infestans eggs employing a specific substrate (7-CP.

  15. CLOCK Acetylates ASS1 to Drive Circadian Rhythm of Ureagenesis.

    Science.gov (United States)

    Lin, Ran; Mo, Yan; Zha, Haihong; Qu, Zhipeng; Xie, Pancheng; Zhu, Zheng-Jiang; Xu, Ying; Xiong, Yue; Guan, Kun-Liang

    2017-10-05

    In addition to responding to environmental entrainment with diurnal variation, metabolism is also tightly controlled by cell-autonomous circadian clock. Extensive studies have revealed key roles of transcription in circadian control. Post-transcriptional regulation for the rhythmic gating of metabolic enzymes remains elusive. Here, we show that arginine biosynthesis and subsequent ureagenesis are collectively regulated by CLOCK (circadian locomotor output cycles kaput) in circadian rhythms. Facilitated by BMAL1 (brain and muscle Arnt-like protein), CLOCK directly acetylates K165 and K176 of argininosuccinate synthase (ASS1) to inactivate ASS1, which catalyzes the rate-limiting step of arginine biosynthesis. ASS1 acetylation by CLOCK exhibits circadian oscillation in human cells and mouse liver, possibly caused by rhythmic interaction between CLOCK and ASS1, leading to the circadian regulation of ASS1 and ureagenesis. Furthermore, we also identified NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 9 (NDUFA9) and inosine-5'-monophosphate dehydrogenase 2 (IMPDH2) as acetylation substrates of CLOCK. Taken together, CLOCK modulates metabolic rhythmicity by acting as a rhythmic acetyl-transferase for metabolic enzymes. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Acetylated starch of Ofada rice as a sustained polymer in ...

    African Journals Online (AJOL)

    Background: Acetylated starches with degrees of substitution (DS) of > 2 have been found suitable for sustained release applications because of their hydrophobic nature and thermoplasticity. The short half-life and high dosing frequency of repaglinide make it an ideal candidate for sustained release. Objectives: To ...

  17. Acetylation of wood components and fourier transform infra-red ...

    African Journals Online (AJOL)

    Acetylation of Turkish pine or cedar wood flour with acetic anhydride was significantly improved in the presence of potassium carbonate at 100°C. Maximum of about 20 and 18% weight percentage gain (WPG) values were obtained with Turkish pine (Pinus brutia) and cedar (Cedrus libani) wood flour after 3 h reaction at ...

  18. Surface effects in the acetylation of granular potato starch

    NARCIS (Netherlands)

    Steeneken, P.A.M.; Woortman, A.J.J.

    2008-01-01

    The occurrence of surface effects in the acetylation of granular potato starch with acetic anhydride to degrees of substitution 0.04-0.2 was studied by two different approaches. The first approach involved the fractionation of granular starch acetates into five different size classes and analysis of

  19. Potentiometric studies of Nickel (II) and copper (II) acetyl acetonato ...

    African Journals Online (AJOL)

    Potentiometric studies of Nickel (II) and copper (II) acetyl acetonato complexes. HN Aliyu, A Mustapha. Abstract. The dissociation constant pKa of acetylacetone has been determined potentiometrically. The pKa value obtained is 9.40, indicating a weak acid. The stability constants of the complex compounds formed from the ...

  20. Effect Of Nicotine And Tobacco Consumption On Brain Acetyl ...

    African Journals Online (AJOL)

    The effect of nicotine and tobacco consumption on brain acetyl cholinesterase and serum alkaline phosphatase in rats was studied. Rats were divided into three groups and the first group was fed rat chow and water ad libitum and an oral administration of 2ml of 0.1%(v/v) nicotine per 100g body weight of rats per day.

  1. Effect of acetylation and varietal differences on the pasting ...

    African Journals Online (AJOL)

    The pasting properties of starch from eight varieties of corn; Okomasa, Obatanpa, Dodzi, Mamaba, Dadaba, Dorke, Golden crystal, and CIDA-ba were studied to establish the effects of acetylation and varietal differences on the pasting properties. Native starches extracted from the corn varieties were modified with 10% v/v ...

  2. The potential role of wood acetylation in climate change mitigation

    NARCIS (Netherlands)

    Van der Lugt, P.; Vogtländer, J.G.; Alexander, J.; Bongers, F.; Stebbins, H.

    2014-01-01

    In a carbon footprint assessment, the greenhouse gas emissions during the life cycle of a material can be measured, and compared to alternative products in terms of kg CO2 equivalent. If applied correctly, wood acetylation opens up a range of new innovative applications in which high performance yet

  3. Synthesis activity-based zymography for detection of lipases and esterases.

    Science.gov (United States)

    Kwon, Min-A; Kim, Hyun Suk; Hahm, Dae-Hyun; Song, Jae Kwang

    2011-04-01

    A new zymography method for lipases and esterases was developed on the basis of the esterification reaction between fatty acids and alcohols. The enzymes were separated by SDS-PAGE and native PAGE. The gel was washed and then incubated in an aqueous solution containing fatty acids (oleic acid 18:1 or caprylic acid 8:0) and dodecanol. Synthesis was visualized by in situ precipitation of water-insoluble and non-diffusible fatty acid esters, such as dodecyl oleate and dodecyl octanoate. The synthesis activity-based zymography was confirmed with different enzyme samples, including commercial lipase preparations, purified recombinant lipase and cutinase, and crude culture supernatants of lipolytic enzyme-producing soil bacteria.

  4. Effects of temperature, ultraviolet radiation and pectin methyl esterase on aerobic methane release from plant material

    DEFF Research Database (Denmark)

    Bruhn, Dan; Mikkelsen, Teis Nørgaard; Øbro, J.

    2009-01-01

    This study examines the effects of different irradiance types on aerobic methane (CH4) efflux rates from terrestrial plant material. Furthermore, the role of the enzyme pectin methyl esterase (PME) on CH4 efflux potential was also examined. Different types of plant tissue and purified pectin were...... incubated in glass vials with different combinations of irradiation and/or temperature. Purified dry pectin was incubated in solution, and with or without PME. Before and after incubation, the concentration of CH4 was measured with a gas chromatograph. Rates of CH4 emission were found to depend...... exponentially on temperature and linearly on UV-B irradiance. UV-B had a greater stimulating effect than UV-A, while visible light had no effect on emission rates. PME was found to substantially reduce the potential for aerobic CH4 emissions upon demethylation of pectin....

  5. Cytological localization of adenosine kinase, nucleoside phosphorylase-1, and esterase-10 genes on mouse chromosome 14

    International Nuclear Information System (INIS)

    Samuelson, L.C.; Farber, R.A.

    1985-01-01

    The authors have determined the regional locations on mouse chromosome 14 of the genes for mouse adenosine kinase (ADK), nucleoside phosphorylase- 1 (NP-1), and esterase-10 (ES-10) by analysis of rearranged mouse chromosomes in gamma-irradiated Chinese hamster X mouse hybrid cell lines. Irradiated clones were screened for expression of the murine forms of these enzymes; segregant clones that expressed only one or two of the three markers were karyotyped. The patterns of enzyme expression in these segregants were correlated with the presence of rearranged chromosomes. The Adk gene was localized to bands A2 to B, Np-1 to bands B to C1, and Es-10 to bands D2 to E2

  6. Effect of xylan oligosaccharides generated from corncobs on food acceptability, growth performance, haematology and immunological parameters of Dicentrarchus labrax fingerlings.

    Science.gov (United States)

    Abdelmalek, Baha Eddine; Driss, Dorra; Kallel, Fatma; Guargouri, Molka; Missaoui, Hechmi; Chaabouni, Semia Ellouz; Ayadi, Mohamed Ali; Bougatef, Ali

    2015-12-01

    The objective of this study was to determine the effect of two levels of inclusion of xylan oligosaccharides (XOS) extracted from corncob on growth, feed utilization, immune status and disease resistance of Mediterranean sea bass (Dicentrarchus labrax) fingerlings. Specimens of 4.75 ± 0.69 g at initial density of 2.7 ± 0.13 kg/m(3) were fed during 12 weeks at 0 g kg(-1) diet, 5 g kg(-1) diet and 10 g kg(-1) diet, dietary XOS level of inclusion in a commercial sea bass diet. Feeding the fish at both XOS dietary inclusion levels significantly increased weight gain, protein efficiency ratio and feed conversion ratio. Feeding of supplemented diets to fish led to reducing mortalities after challenging with A. hydrophila. The haematological and immunological parameters were assayed in both pre-challenged and post-challenged groups. There was an increased trend in red blood corpuscles, white blood corpuscles, pack cell volume, haemoglobin (Hb %) and serum protein content in treated groups over the control as time elapsed with the feeding trials. The serum immunoglobulin level and lysozyme activity showed an increased trend in the fed groups. Histological features of the liver showed lower lipid vacuolization and regular-shaped morphology of hepatocytes around the sinusoidal spaces denoting a better utilization of dietary nutrients supported with the morphometric data. In conclusion, XOS added at a designated dose (5 g kg(-1) diet) in the diet improves growth and stimulates the immunity and makes D. labrax fingerlings more resistant to infection by A. hydrophila.

  7. GDSL esterase/lipase genes in Brassica rapa L.: genome-wide identification and expression analysis.

    Science.gov (United States)

    Dong, Xiangshu; Yi, Hankuil; Han, Ching-Tack; Nou, Ill-Sup; Hur, Yoonkang

    2016-04-01

    GDSL esterase/lipase proteins (GELPs), a very large subfamily of lipolytic enzymes, have been identified in microbes and many plants, but only a few have been characterized with respect to their roles in growth, development, and stress responses. In Brassica crops, as in many other species, genome-wide systematic analysis and functional studies of these genes are still lacking. As a first step to study their function in B. rapa ssp. pekinensis (Chinese cabbage), we comprehensively identified all GELP genes in the genome. We found a total of 121 Brassica rapa GDSL esterase/lipase protein genes (BrGELPs), forming three clades in the phylogenetic analysis (two major and one minor), with an asymmetrical chromosomal distribution. Most BrGELPs possess four strictly conserved residues (Ser-Gly-Asn-His) in four separate conserved regions, along with short conserved and clade-specific blocks, suggesting functional diversification of these proteins. Detailed expression profiling revealed that BrGELPs were expressed in various tissues, including floral organs, implying that BrGELPs play diverse roles in various tissues and during development. Ten percent of BrGELPs were specifically expressed in fertile buds, rather than male-sterile buds, implying their involvement in pollen development. Analyses of EXL6 (extracellular lipase 6) expression and its co-expressed genes in both B. rapa and Arabidopsis, as well as knockdown of this gene in Arabidopsis, revealed that this gene plays an important role in pollen development in both species. The data described in this study will facilitate future investigations of other BrGELP functions.

  8. Tissue distribution, characterization and in vitro inhibition of B-esterases in the earwig Forficula auricularia.

    Science.gov (United States)

    Malagnoux, Laure; Capowiez, Yvan; Rault, Magali

    2014-10-01

    Earwigs are important natural enemies of numerous pests in pome fruit orchards worldwide. Studying the effects of agricultural practices on these biological control agents is important for understanding its vulnerability in the field. The aim of this study was to characterize the B-esterase activities in the European earwig Forficula auricularia and to evaluate in vitro its sensitivity to organophosphate and carbamate pesticides. Acetylcholinesterase (AChE) activity was mainly measured with 1.5 mM acetylthiocholine as the substrate in the microsomal fraction of earwig heads (70% of total AChE activity). Carboxylesterase (CbE) activities were measured with three substrates [5 mM 4-nitrophenyl acetate (4-NPA), 1mM 4-nitrophenyl valerate (4-NPV), and 2 mM α-naphtyl acetate (α-NA)] to examine different isoenzymes, which were present mainly in the cytosolic fraction (about 70-88% of total activities) of all earwig tissues. CbE activity was higher than AChE activity, especially with α-NA, then 4-NPA and lastly 4-NPV. Chlorpyrifos-oxon an organophosphate, and carbaryl a carbamate pesticide, inhibited AChE and CbE activities in a concentration-dependent manner. Earwig CbE activities showed a stronger sensitivity to organophosphate than AChE, with the strongest effect for chlorpyrifos-oxon on male carboxylesterase activities. CbE and AChE showed about the same sensitivity to carbamate pesticides regardless of sex. These results suggest that B-type esterases in the European earwig F.auricularia are suitable biomarkers of pesticide exposure. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. Solution behavior and activity of a halophilic esterase under high salt concentration.

    Science.gov (United States)

    Rao, Lang; Zhao, Xiubo; Pan, Fang; Li, Yin; Xue, Yanfen; Ma, Yanhe; Lu, Jian R

    2009-09-14

    Halophiles are extremophiles that thrive in environments with very high concentrations of salt. Although the salt reliance and physiology of these extremophiles have been widely investigated, the molecular working mechanisms of their enzymes under salty conditions have been little explored. A halophilic esterolytic enzyme LipC derived from archeaon Haloarcula marismortui was overexpressed from Escherichia coli BL21. The purified enzyme showed a range of hydrolytic activity towards the substrates of p-nitrophenyl esters with different alkyl chains (n = 2-16), with the highest activity being observed for p-nitrophenyl acetate, consistent with the basic character of an esterase. The optimal esterase activities were found to be at pH 9.5 and [NaCl] = 3.4 M or [KCl] = 3.0 M and at around 45 degrees C. Interestingly, the hydrolysis activity showed a clear reversibility against changes in salt concentration. At the ambient temperature of 22 degrees C, enzyme systems working under the optimal salt concentrations were very stable against time. Increase in temperature increased the activity but reduced its stability. Circular dichroism (CD), dynamic light scattering (DLS) and small angle neutron scattering (SANS) were deployed to determine the physical states of LipC in solution. As the salt concentration increased, DLS revealed substantial increase in aggregate sizes, but CD measurements revealed the maximal retention of the alpha-helical structure at the salt concentration matching the optimal activity. These observations were supported by SANS analysis that revealed the highest proportion of unimers and dimers around the optimal salt concentration, although the coexistent larger aggregates showed a trend of increasing size with salt concentration, consistent with the DLS data. The solution alpha-helical structure and activity relation also matched the highest proportion of enzyme unimers and dimers. Given that all the solutions studied were structurally inhomogeneous, it

  10. Feruloyl Esterase Activity from Coffee Pulp in Solid-State Fermentation

    Directory of Open Access Journals (Sweden)

    Gerardo Saucedo-Castañeda

    2011-01-01

    Full Text Available Hydroxycinnamic acids (HAs have a potential application in the food and pharmaceutical industry because they are rich in phenolics. Feruloyl esterases release phenolic compounds from plant cell walls. Coffee pulp is rich in HAs linked to polysaccharides. A solvent extraction of free HAs was performed with aqueous methanol (80 %. A response surface methodology was applied to optimise the extraction of these compounds from coffee pulp, and the best results were obtained at 56 °C for 34 min. Alkaline and acid hydrolyses were performed to evaluate the content of linked HAs. Treated (extracted coffee pulp was used to produce feruloyl esterases in solid-state fermentation by Aspergillus tamarii V12307, previously selected by a hydrolysis plate assay. Different dilutions of a culture medium were added to the coffee pulp, and the diluted medium with half the nutrients allowed for higher CO2 production. A specific growth rate (μCO2 of 0.25 h^–1 and a lag phase (tlag of 14.3 h were observed under the selected conditions. Finally, enzymatic activities were 14.0 and 10.8 nkat per g of dried matter when methyl and ethyl ferulate were used as substrates, respectively. Productivities (9.3 and 7.2 nkat per g of dried matter per day, respectively were higher when compared to other studies carried out in solid-state fermentation. Utilisation of coffee pulp for enzyme production improves the added value of this abundant by-product of the coffee industry.

  11. Proteomic analysis of lysine acetylation sites in rat tissues reveals organ specificity and subcellular patterns

    DEFF Research Database (Denmark)

    Lundby, Alicia; Hansen, Kasper Lage; Weinert, Brian Tate

    2012-01-01

    that the subcellular acetylation distribution is tissue-type dependent and that acetylation targets tissue-specific pathways involved in fundamental physiological processes. We compare lysine acetylation patterns for rat as well as human skeletal muscle biopsies and demonstrate its general involvement in muscle...

  12. Comprehensive Proteomic Analysis of Lysine Acetylation in the Foodborne Pathogen Trichinella spiralis

    Directory of Open Access Journals (Sweden)

    Yong Yang

    2018-01-01

    Full Text Available Lysine acetylation is a dynamic and highly conserved post-translational modification that plays a critical role in regulating diverse cellular processes. Trichinella spiralis is a foodborne parasite with a considerable socio-economic impact. However, to date, little is known regarding the role of lysine acetylation in this parasitic nematode. In this study, we utilized a proteomic approach involving anti-acetyl lysine-based enrichment and highly sensitive mass spectrometry to identify the global acetylated proteome and investigate lysine acetylation in T. spiralis. In total, 3872 lysine modification sites were identified in 1592 proteins that are involved in a wide variety of biological processes. Consistent with the results of previous studies, a large number of the acetylated proteins appear to be involved in metabolic and biosynthetic processes. Interestingly, according to the functional enrichment analysis, 29 acetylated proteins were associated with phagocytosis, suggesting an important role of lysine acetylation in this process. Among the identified proteins, 15 putative acetylation motifs were detected. The presence of serine downstream of the lysine acetylation site was commonly observed in the regions surrounding the sites. Moreover, protein interaction network analysis revealed that various interactions are regulated by protein acetylation. These data represent the first report of the acetylome of T. spiralis and provide an important resource for further explorations of the role of lysine acetylation in this foodborne pathogen.

  13. DMPD: Acetylation of MKP-1 and the control of inflammation. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18922786 Acetylation of MKP-1 and the control of inflammation. Chi H, Flavell RA. S...ci Signal. 2008 Oct 14;1(41):pe44. (.png) (.svg) (.html) (.csml) Show Acetylation of MKP-1 and the control of inflammation.... PubmedID 18922786 Title Acetylation of MKP-1 and the control of inflammation. Authors Chi H,

  14. Relativistic Density Functional Theory Calculations of the Electron Paramagnetic Resonance Parameters for Vanadyl Acetyl Acetonate and Copper Acetyl Acetonate

    Science.gov (United States)

    Mainali, Laxman; Sahu, Indra; Earle, Keith

    2008-03-01

    Relativistic density functional theory calculations of electron paramagnetic resonance (EPR) parameters using a variety of basis sets have been computed for the model systems Vanadyl acetyl acetonate and Copper acetyl acetonate using the ORCA program. The basis set dependence of g and A tensor calculations for Vanadyl acetyl acetonate and Copper acetyl acetonate were studied using Pople Style and Ahlrichs basis sets in Local and gradient corrected functionals (BP86 and PWP) and Hybrid functionals (B3LYP and PW1PW). The PW1PW hybrid functional gives the best values for VO(acac)2 using the TZV basis set and for Cu(acac)2 using the 6-311G(d) basis set. The calculated A values with PW1PW hybrid functional for VO(acac)2 and Local and gradient corrected functional (BP86) for Cu(acac)2 with same basis set (DZ) give better results than previously reported values using the Amsterdam Density Functional Theory (ADF) Software. Our calculated g and A tensor values are in good agreement with the values determined from experiment.

  15. The Acetyl Group Buffering Action of Carnitine Acetyltransferase Offsets Macronutrient-Induced Lysine Acetylation of Mitochondrial Proteins

    Directory of Open Access Journals (Sweden)

    Michael N. Davies

    2016-01-01

    Full Text Available Lysine acetylation (AcK, a posttranslational modification wherein a two-carbon acetyl group binds covalently to a lysine residue, occurs prominently on mitochondrial proteins and has been linked to metabolic dysfunction. An emergent theory suggests mitochondrial AcK occurs via mass action rather than targeted catalysis. To test this hypothesis, we performed mass spectrometry-based acetylproteomic analyses of quadriceps muscles from mice with skeletal muscle-specific deficiency of carnitine acetyltransferase (CrAT, an enzyme that buffers the mitochondrial acetyl-CoA pool by converting short-chain acyl-CoAs to their membrane permeant acylcarnitine counterparts. CrAT deficiency increased tissue acetyl-CoA levels and susceptibility to diet-induced AcK of broad-ranging mitochondrial proteins, coincident with diminished whole body glucose control. Sub-compartment acetylproteome analyses of muscles from obese mice and humans showed remarkable overrepresentation of mitochondrial matrix proteins. These findings reveal roles for CrAT and L-carnitine in modulating the muscle acetylproteome and provide strong experimental evidence favoring the nonenzymatic carbon pressure model of mitochondrial AcK.

  16. Differently sized granules from acetylated potato and sweet potato starches differ in the acetyl substitution pattern of their amylose polulations.

    NARCIS (Netherlands)

    Chen Zenghong,; Schols, H.A.; Voragen, A.G.J.

    2004-01-01

    Acetylated potato and sweet potato starches were fractionated according to granule size. From the fractions obtained amylose and amylopectin were isolated and characterized with respect to degree of substitution (DS) and degradability with -amylase, -amylase and amyloglucosidase. The DS of the

  17. Studies investigating the excretion of acetyl urea in the milk of dairy cows receiving oral doses of 14C acetyl urea

    International Nuclear Information System (INIS)

    Bergner, H.; Kijora, C.; Goersch, R.

    1976-01-01

    2 experimental cows were fed acetyl urea several weeks before the trial was started. The first cow received a daily amount of 200 g and the second cow 855 g. On the first day of experiment both cows were given 5 mCi of 14 C acetyl urea intraruminally. Up to 6 hrs after the beginning of the experiment acetyl urea in blood plasma was shown to contain a higher proportion of 14 C activity than urea. 0.21 g urea and 0.18 g acetyl urea were contained in 1 kg of milk from cow No 1 while 1 kg of milk from cow No 2 contained 0.18 g urea and 0.12 g acetyl urea. The feeding of acetyl urea to dairy cows is not recommended on the basis of the fact that any further contamination of human nutrition with foreign substances should be possibly avoided. (author)

  18. Sequential Dy(OTf)3 -Catalyzed Solvent-Free Per-O-Acetylation and Regioselective Anomeric De-O-Acetylation of Carbohydrates.

    Science.gov (United States)

    Yan, Yi-Ling; Guo, Jiun-Rung; Liang, Chien-Fu

    2017-09-19

    Dysprosium(III) trifluoromethanesulfonate-catalyzed per-O-acetylation and regioselective anomeric de-O-acetylation of carbohydrates can be tuned by adjusting the reaction medium. In this study, the per-O-acetylation of unprotected sugars by using a near-stoichiometric amount of acetic anhydride under solvent-free conditions resulted in the exclusive formation of acetylated saccharides as anomeric mixtures, whereas anomeric de-O-acetylation in methanol resulted in a moderate-to-excellent yield. Reactions with various unprotected monosaccharides or disaccharides followed by a semi-one-pot sequential conversion into the corresponding acetylated glycosyl hemiacetal also resulted in high yields. Furthermore, the obtained hemiacetals could be successfully transformed into trichloroimidates after Dy(OTf) 3 -catalyzed glycosylation. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Heterodisaccharide 4-O-(N-acetyl-β-D-glucosaminyl)-D-glucosamine is an effective chemotactic attractant for Vibrio bacteria that produce chitin oligosaccharide deacetylase.

    Science.gov (United States)

    Hirano, T; Aoki, M; Kadokura, K; Kumaki, Y; Hakamata, W; Oku, T; Nishio, T

    2011-08-01

    To investigate the attractant effect of 4-O-(N-acetyl-β-D-glucosaminyl)-D-glucosamine (GlcNAc-GlcN) in the chemotaxis of Vibrio bacteria that produce carbohydrate esterase (CE) family 4 chitin oligosaccharide deacetylase (COD), an enzyme that catalyzes the production of GlcNAc-GlcN from N,N'-diacetylchitobiose (GlcNAc)(2). The chemotactic effect of disaccharides from chitin on several strains of Vibrio bacteria was investigated using an agar gel lane-migration method. The results demonstrated that GlcNAc-GlcN functions as an effective chemoattractant in the CE family 4 COD-producing vibrios, Vibrio parahaemolyticus and Vibrio alginolyticus. In contrast, this phenomenon was not observed in Vibrio nereis or Vibrio furnissii, which lack genes encoding this enzyme. From transmission electron microscope observation of V. parahaemolyticus cells following the chemotaxis assay, GlcNAc-GlcN appears to stimulate polar flagellum rotation. GlcNAc-GlcN is a specific chemoattractant for the CE family 4 COD-producing vibrios, V. parahaemolyticus and V. alginolyticus. It was clarified for the first time that GlcNAc-GlcN functions as a signalling molecule in the chemotaxis of Vibrio bacteria that have an ability to produce CE family 4 COD, which generate GlcNAc-GlcN from (GlcNAc)(2). © 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.

  20. Two-dimensional NMR evidence for cleavage of lignin and xylan substituents in wheat straw through hydrothermal pretreatment and enzymatic hydrolysis

    DEFF Research Database (Denmark)

    Yelle, Daniel J.; Kaparaju, Laxmi-Narasimha Prasad; Hunt, Christopher G.

    2013-01-01

    correlation spectroscopy, via an heteronuclear single quantum coherence experiment, revealed substantial lignin β-aryl ether cleavage, deacetylation via cleavage of the natural acetates at the 2-O- and 3-O-positions of xylan, and uronic acid depletion via cleavage of the (1 → 2)-linked 4-O....... g., further deacylation revealed by the depletion in ferulate and p-coumarate structures). Supplementary chemical analyses showed that the hydrothermal pretreatment increased the cellulose and lignin concentration with partial removal of extractives and hemicelluloses. The subsequent enzymatic...

  1. Esterase activity in the guinea pig thyroid under normal and pathological conditions (vitamin A deficiency) with special regard to cyst-like structures

    DEFF Research Database (Denmark)

    Kirkeby, S

    1977-01-01

    By use of different activators and inhibitors, TOCP(tri-o-cresyl phosphate), PCMB (parachloromercury benzoate), NiCl2, Pb(NO3)2, HgCl2, Hg(NO3)2, eserine and sodium taurocholate, it is shown that the esterase in the cyst cells and in group I cells of the guinea pig thyroid probably are A-esterase...... isoenzymes. The activity in the majority of cyst cells is considerably stronger than in the other thyroid epithelial cells, and it is resistant to Hg inactivation. Neither esterase type nor intensity of the reaction product is altered in group I-II or cyst cells during vitamin A-deficiency. When a normal...... diet is given, the esterase in all thyroid epithelial cells is very sensitive to sodium taurocholate, while in cyst cells it is rather resistant to this inhibitor at vitamin-A-deficiency....

  2. Histochemical studies on genetical control of hormonal enzyme inducibility in the mouse. I. Non-specific esterase activity and regional histology of the epididymis

    DEFF Research Database (Denmark)

    Blecher, S R; Kirkeby, S

    1978-01-01

    As a base line for future cell genetical studies the authors record the distribution of non-specific esterase reaction in the various histologically distinguishable cell types of the mouse epididymis. The findings are correlated with previous descriptions of the lobar structure of the organ....... Assuming the sequence of lobes of the head to be as implied in these classical descriptions, the esterase activity of the epithelial cells gradates between strong to weak several times along the length of the epididymal duct. The relationship of the lobes to each other, as seen in transverse sections......, is described. Methodological studies using different fixatives indicate that apparent similarity of esterase reaction at different sites may camouflage an underlying difference in the nature of the esterases at these sites....

  3. Proteome-wide analysis of lysine acetylation suggests its broad regulatory scope in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Henriksen, Peter; Wagner, Sebastian Alexander; Weinert, Brian Tate

    2012-01-01

    acetyltransferases and deacetylases. However, only a few dozen acetylation sites in S. cerevisiae are known, presenting a major obstacle for further understanding the regulatory roles of acetylation in this organism. Here we use high resolution mass spectrometry to identify about 4000 lysine acetylation sites in S......-containing histone H2B deubiquitylase complex. Our data provides the first global survey of acetylation in budding yeast, and suggests a wide-ranging regulatory scope of this modification. The provided dataset may serve as an important resource for the functional analysis of lysine acetylation in eukaryotes....

  4. Lipids Reprogram Metabolism to Become a Major Carbon Source for Histone Acetylation

    DEFF Research Database (Denmark)

    McDonnell, Eoin; Crown, Scott B; Fox, Douglas B

    2016-01-01

    Cells integrate nutrient sensing and metabolism to coordinate proper cellular responses to a particular nutrient source. For example, glucose drives a gene expression program characterized by activating genes involved in its metabolism, in part by increasing glucose-derived histone acetylation....... Here, we find that lipid-derived acetyl-CoA is a major source of carbon for histone acetylation. Using (13)C-carbon tracing combined with acetyl-proteomics, we show that up to 90% of acetylation on certain histone lysines can be derived from fatty acid carbon, even in the presence of excess glucose...

  5. The activity of non-specific esterase in the thyroid epithelial cells of the guinea pig as influenced by various inhibitors and activators. A histochemical study

    DEFF Research Database (Denmark)

    Kirkeby, S

    1976-01-01

    The action of various inhibitors and activators upon esterase activity in the thyroid epithelial cells is demonstrated. The agents used were triorthocresylphosphate (TOCP), parachloromercuribenzoate (PCMB), Arsanillic acid, p-nitrophenyl dimethyl carbamate and bis p-nitrophenyl phosphate. TOCP...... in the para-, inter- and intrafollicular cells was unchanged. The results obtained are related to previous biochemical and histochemical observations and the nature of esterases in the thyroid is discussed....

  6. Histochemical studies on genetical control of hormonal enzyme inducibility in the mouse. IV: Cellular localization of androgen sensitive nonspecific esterase in the epididymis

    DEFF Research Database (Denmark)

    Kirkeby, S; Blecher, S R

    1981-01-01

    Nonspecific esterase of mouse epididymis has previously been studied histochemically, using alpha naphthyl-acetate and 5-bromoindoxyl acetate techniques, as well as certain inhibitors. Epithelial cell types of the epididymis have been characterized, and certain esterase isozymes in a particular c...... to normal. This method can now be applied to the study of epididymides of genetically sex-reversed chromosomal female, to analyze genetic control of X-chromosomal activation....

  7. Acetylation Increases EWS-FLI1 DNA Binding and Transcriptional Activity

    International Nuclear Information System (INIS)

    Schlottmann, Silke; Erkizan, Hayriye V.; Barber-Rotenberg, Julie S.; Knights, Chad; Cheema, Amrita; Üren, Aykut; Avantaggiati, Maria L.; Toretsky, Jeffrey A.

    2012-01-01

    Ewing Sarcoma (ES) is associated with a balanced chromosomal translocation that in most cases leads to the expression of the oncogenic fusion protein and transcription factor EWS-FLI1. EWS-FLI1 has been shown to be crucial for ES cell survival and tumor growth. However, its regulation is still enigmatic. To date, no functionally significant post-translational modifications of EWS-FLI1 have been shown. Since ES are sensitive to histone deacetylase inhibitors (HDI), and these inhibitors are advancing in clinical trials, we sought to identify if EWS-FLI1 is directly acetylated. We convincingly show acetylation of the C-terminal FLI1 (FLI1-CTD) domain, which is the DNA binding domain of EWS-FLI1. In vitro acetylation studies showed that acetylated FLI1-CTD has higher DNA binding activity than the non-acetylated protein. Over-expression of PCAF or treatment with HDI increased the transcriptional activity of EWS-FLI1, when co-expressed in Cos7 cells. However, our data that evaluates the acetylation of full-length EWS-FLI1 in ES cells remains unclear, despite creating acetylation specific antibodies to four potential acetylation sites. We conclude that EWS-FLI1 may either gain access to chromatin as a result of histone acetylation or undergo regulation by direct acetylation. These data should be considered when patients are treated with HDAC inhibitors. Further investigation of this phenomenon will reveal if this potential acetylation has an impact on tumor response.

  8. Esterase activity in the guinea pig thyroid under normal and pathological conditions (vitamin A deficiency) with special regard to cyst-like structures

    DEFF Research Database (Denmark)

    Kirkeby, S

    1977-01-01

    By use of different activators and inhibitors, TOCP(tri-o-cresyl phosphate), PCMB (parachloromercury benzoate), NiCl2, Pb(NO3)2, HgCl2, Hg(NO3)2, eserine and sodium taurocholate, it is shown that the esterase in the cyst cells and in group I cells of the guinea pig thyroid probably are A-esterase......By use of different activators and inhibitors, TOCP(tri-o-cresyl phosphate), PCMB (parachloromercury benzoate), NiCl2, Pb(NO3)2, HgCl2, Hg(NO3)2, eserine and sodium taurocholate, it is shown that the esterase in the cyst cells and in group I cells of the guinea pig thyroid probably are A......-esterase isoenzymes. The activity in the majority of cyst cells is considerably stronger than in the other thyroid epithelial cells, and it is resistant to Hg inactivation. Neither esterase type nor intensity of the reaction product is altered in group I-II or cyst cells during vitamin A-deficiency. When a normal...... diet is given, the esterase in all thyroid epithelial cells is very sensitive to sodium taurocholate, while in cyst cells it is rather resistant to this inhibitor at vitamin-A-deficiency....

  9. Flexibility of backbone fibrils in α-chitin crystals with different degree of acetylation.

    Science.gov (United States)

    Yu, Zechuan; Lau, Denvid

    2017-10-15

    Acetyl groups are backbone outreaches that enhance inter-fibril connection in chitin and chitosan fibril bundle. Removal of acetyl groups affects flexibility of chitosan fibril bundle, thereby affecting mechanical strength of chitosan-based products. Understandings of relationship between degree of acetylation and flexibility of chitin fibril bundle conduce to optimization of synthetic chitin materials. Here, the relationship is examined by performing molecular dynamics simulations. Coiling of chitin and chitosan fibril bundle with different degree of acetylation is observed and flexibility of fibrils is measured. Number and alignment of acetyl groups are found to be important factors determining the flexibility of chitin and chitosan fibril bundle. Structural instability can be caused by incompatible alignment of acetyl groups. Our findings on synthetic chitin-based materials indicate that adding a small amount of acetyl groups to chitosan can significantly enhance the integrity of fibril bundle. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Snail acetylation by histone acetyltransferase p300 in lung cancer

    OpenAIRE

    Chang, Rui; Zhang, Yinjie; Zhang, Peng; Zhou, Qinghua

    2017-01-01

    Background Epithelial to mesenchymal transition (EMT) is a complex and dynamic molecular event in lung cancer metastasis that has not yet been thoroughly investigated. EMT transcriptional factors, such as Snail, play a central role in regulation of the EMT process. In this study, we sought to identify an association between p300 and Snail in lung cancer, as well as the engagement of p300 in Snail acetylation. Methods We transfected p300 small interfering RNA into lung cancer cells to detect S...

  11. Acetylated DNA-damaging clerodane diterpenes from Casearia sylvestris.

    Science.gov (United States)

    de Carvalho, Paulo Roberto F.; Furlan, Maysa; Young, Maria Claudia M.; Kingston, David G. I.; Bolzani, Vanderlan da S.

    1998-11-20

    In addition to the known diterpene casearin G (1), two new clerodane diterpene casearins type, casearin S (2) and casearin T (3), were isolated from an acetylated bioactive CH(2)Cl(2)/MeOH extract from leaves of Casearia sylvestris. The diterpenes 1-3 exhibited moderate but selective activity towards the DNA-repair deficient yeast Saccharomyces cerevisiae mutants RAD 52YK and RS 321. The structures of 1-3 were established on the basis of NMR spectroscopic experiments

  12. Acetylation site specificities of lysine deacetylase inhibitors in human cells

    DEFF Research Database (Denmark)

    Schölz, Christian; Weinert, Brian Tate; Wagner, Sebastian A

    2015-01-01

    Lysine deacetylases inhibitors (KDACIs) are used in basic research, and many are being investigated in clinical trials for treatment of cancer and other diseases. However, their specificities in cells are incompletely characterized. Here we used quantitative mass spectrometry (MS) to obtain acety......1-α, providing a possible mechanistic explanation of its adverse, pro-inflammatory effects. Our results offer a systems view of KDACI specificities, providing a framework for studying function of acetylation and deacetylases....

  13. Geographic and climatic differentiation of electrophoretic forms of esterase, glutamate dehydrogenase and peroxidase in Scots pine tissues

    Directory of Open Access Journals (Sweden)

    Barbara Kieliszewska-Rokicka

    2014-01-01

    Full Text Available The enzymatic systems have been investigated in individual trees derived from 17 distant localities in Europe and Asia. Esterase and glutamate dehydrogenase polymorphism was determined in extracts from mixture of 50 macrogametophytes, iperoxidase polymorphism was investigated in needle extracts from 3-month-old seedlings. While testing by disc electrophoresis needle extracts of Scots pine deriving from specimens of different geographic ranges it was found that those from the South displayed mare isoperoxidases than those from the North. Most pronounced differences in number and relative activity of esterase and glutamate dehydrogenase isozymes were found in the specimens originating from isolated stands. It is suggested that hight temperatures and dry summer periods could influence the number and activity of the isozymes.

  14. Whole-Cell Biocatalytic Synthesis of Cinnamyl Acetate with a Novel Esterase from the DNA Library of Acinetobacter hemolyticus.

    Science.gov (United States)

    Dong, Hao; Secundo, Francesco; Xue, Changhu; Mao, Xiangzhao

    2017-03-15

    Cinnamyl acetate has a wide application in the flavor and fragrance industry because of its sweet, balsamic, and floral odor. Up to now, lipases have been mainly used in enzyme-mediated synthesis of cinnamyl acetate, whereas esterases are used in only a few cases. Moreover, the use of purified enzymes is often a disadvantage, which leads to increases of the production costs. In this paper, a genomic DNA library of Acinetobacter hemolyticus was constructed, and a novel esterase (EstK1) was identified. After expression in Escherichia coli, the whole-cell catalyst of EstK1 displayed high transesterification activity to produce cinnamyl acetate in nonaqueous systems. Furthermore, under optimal conditions (vinyl acetate as acyl donor, isooctane as solvent, molar ratio 1:4, temperature 40 °C), the conversion ratio of cinnamyl alcohol could be up to 94.1% at 1 h, and it reached an even higher level (97.1%) at 2 h.

  15. Isolation and characterization of a heavy metal-resistant, thermophilic esterase from a Red Sea Brine Pool

    KAUST Repository

    Mohamed, Yasmine M.

    2013-11-28

    The Red Sea Atlantis II brine pool is an extreme environment that displays multiple harsh conditions such as high temperature, high salinity and high concentrations of multiple, toxic heavy metals. The survival of microbes in such an environment by utilizing resistant enzymes makes them an excellent source of extremophilic enzymes. We constructed a fosmid metagenomic library using DNA isolated from the deepest and most secluded layer of this pool. We report the isolation and biochemical characterization of an unusual esterase: EstATII. EstATII is thermophilic (optimum temperature, 65 C), halotolerant (maintains its activity in up to 4.5â€...M NaCl) and maintains at least 60% of its activity in the presence of a wide spectrum of heavy metals. The combination of biochemical characteristics of the Red Sea Atlantis II brine pool esterase, i.e., halotolerance, thermophilicity and resistance to heavy metals, makes it a potentially useful biocatalyst.

  16. Functional characterization of a juvenile hormone esterase related gene in the moth Sesamia nonagrioides through RNA interference.

    Directory of Open Access Journals (Sweden)

    Dimitrios Kontogiannatos

    Full Text Available Juvenile hormone esterase (JHE is a carboxylesterase that has attracted great interest because of its critical role in regulating larval to adult transition in insects and other arthropods. Previously, we characterized an ecdysteroid sensitive and juvenile hormone non-susceptible juvenile hormone esterase related gene (SnJHER in the corn stalk borer, Sesamia nonagrioides. SnJHER was rhythmically up-regulated close to each molt during the corn stalk borer's larval development. In this paper we attempted to functionally characterize SnJHER using several reverse genetics techniques. To functionally characterize SnJHER, we experimented with different dsRNA administration methods, including hemolymph, bacterial or baculovirus-mediated RNA interference, (RNAi. Our findings indicate the potential implication of SnJHER in the developmental programming of Sesamia nonagrioides. It is still unclear whether SnJHER is closely related to the authentic JHE gene, with different or similar biological functions.

  17. The effect of rabbit age on in vitro caecal fermentation of starch, pectin, xylan, cellulose, compound feed and its fibre.

    Science.gov (United States)

    Lavrenčič, A

    2007-03-01

    In vitro gas production kinetics of six different substrates, pectin (PEC), xylan (XYL), starch (STA), cellulose (CEL), commercial compound feed (FEED; 201 g crude protein per kg, 155 g crude fibre per kg, 334 g neutral-detergent fibre (NDF) per kg and 190 g acid-detergent fibre (ADF) per kg) and an NDF prepared from commercial compound feed (NDFFEED) were determined using the caecum contents of weaned rabbits (36 days of age) and of rabbits at slaughter age (78 days of age) as inoculums. The cumulated gas production over 96 h of incubation was modelled with Gompertz model, and the kinetic parameters compared. The total potential gas production (parameter 'B' of the Gompertz model) was not affected (P>0.05) by the inoculum source, except with STA, where rabbits at slaughter weight had significantly higher total potential fermentability (314 ml/g dry matter (DM)) than those at weaning age (189 ml/g DM). Intensities of fermentation (maximum fermentation rate; MFR) of PEC (32.2 ml/h) and XYL (24.4 ml/h) were significantly greater in rabbits at weaning, while that of STA (45 ml/h) was significantly lower than at slaughter age (23.0, 14.3 and 14.0 ml/h for PEC, XYL and STA, respectively). The MFRs of CEL and NDFFEED were very similar between inoculum sources. In the first 10 h of fermentation which correspond to the normal retention time of the substrates in the caecum, the highest amount of gas was produced from PEC, followed by FEED and XYL. These substrates had a time of maximum fermentation rate (TMFR) at both rabbit ages short enough (8.0 and 9.5 h for PEC, 9.5 and 6.6 h for FEED, 13.7 and 14.2 h for XYL at weaning and at slaughter age, respectively) to be almost completely fermented in vivo.

  18. EWSR1 regulates mitosis by dynamically influencing microtubule acetylation.

    Science.gov (United States)

    Wang, Yi-Long; Chen, Hui; Zhan, Yi-Qun; Yin, Rong-Hua; Li, Chang-Yan; Ge, Chang-Hui; Yu, Miao; Yang, Xiao-Ming

    2016-08-17

    EWSR1, participating in transcription and splicing, has been identified as a translocation partner for various transcription factors, resulting in translocation, which in turn plays crucial roles in tumorigenesis. Recent studies have investigated the role of EWSR1 in mitosis. However, the effect of EWSR1 on mitosis is poorly understood. Here, we observed that depletion of EWSR1 resulted in cell cycle arrest in the mitotic phase, mainly due to an increase in the time from nuclear envelope breakdown to metaphase, resulting in a high percentage of unaligned chromosomes and multipolar spindles. We also demonstrated that EWSR1 is a spindle-associated protein that interacts with α-tubulin during mitosis. EWSR1 depletion increased the cold-sensitivity of spindle microtubules, and decreased the rate of spindle assembly. EWSR1 regulated the level of microtubule acetylation in the mitotic spindle; microtubule acetylation was rescued in EWSR1-depleted mitotic cells following suppression of HDAC6 activity by its specific inhibitor or siRNA treatment. In summary, these results suggest that EWSR1 regulates the acetylation of microtubules in a cell cycle-dependent manner through its dynamic location on spindle MTs, and may be a novel regulator for mitosis progress independent of its translocation.

  19. Autoradiographic study of nuclear protein acetylation during Locust spermiogenesis

    International Nuclear Information System (INIS)

    Bouvier, D.; Chevaillier, P.

    1975-01-01

    Autoradiographic studies, at the light and electron microscope level, demonstrate that spermatid nuclei of the Locust Locusta migratoria incorporate 3 H-acetate, especially during the first stages of spermiogenesis. The highest level of acetate incorporation is observed during stage II of spermiogenesis. During this stage and the following, the spermatid nucleus undergoes a number of structural and chemical modifications: chromatin decondenses and somatic histones are progressively replaced by newly synthesized arginine-rich proteins. Therefore, the higher degree of acetylation of nuclear components coincides with chromatin decondensation and precedes the protein transition occurring in later stages. Cytochemical and autoradiographic tests have been realized so as to localize 3 H-acetate in the nuclear components. Trichloracetic acid was used at various concentrations: the action of hydrochloric acid, pronase and DNase was also tested. The results support the idea that proteins, and among them histones, are the only nuclear components to be acetylated during spermiogenesis. Thus, histone acetylation seems to play an important role in modulating histone-DNA interactions and allowing histone replacement [fr

  20. Substituent-specific antibody against glucuronoxylan reveals close association of glucuronic acid and acetyl substituents and distinct labeling patterns in tree species

    DEFF Research Database (Denmark)

    Koutaniemi, Sanna; Guillon, Fabienne; Tranquet, Olivier

    2012-01-01

    Immunolabeling can be used to locate plant cell wall carbohydrates or other components to specific cell types or to specific regions of the wall. Some antibodies against xylans exist; however, many partly react with the xylan backbone and thus provide limited information on the type of substituen...

  1. Esterase inhibition in tadpoles of Scinax fuscovarius (Anura, Hylidae) as a biomarker for exposure to organophosphate pesticides.

    Science.gov (United States)

    Leite, Patricia Zazeri; Margarido, Tatiana Cristina Stefani; de Lima, Daína; Rossa-Feres, Denise de Cerqueira; de Almeida, Eduardo Alves

    2010-09-01

    Organophosphate pesticides (OPs) are among the most used insecticides in agriculture, causing the inhibition of esterases like acetylcholinesterase (AChE), butyrylcholinesterase (BChE) and carboxylesterase (CbE). Pesticides can reach the aquatic environment, posing risks to non-target organisms, including tadpoles. In this work, we characterized the activities of AChE, BChE and CbE in tadpoles of the snouted treefrog Scinax fuscovarius, and verified their in vitro sensibility to different inhibitors [phenylmethane sulfonyl fluoride (PMSF), tetra-isopropylpyrophosphamide (iso-OMPA) and the OP diazinon]. In vivo effects of diazinon and esterase recovery after 2-pyridine-aldoxime (2-PAM) treatment of the protein extract were also studied in tadpoles with distinct stages of development exposed to 1 and 3 mg/l for 2 and 7 days. Optimal conditions were established for AChE and CbE; BChE activity was negligible. PMSF affected esterase activities and is not recommended for homogenization buffers. Iso-OMPA treatment caused no changes in AChE and CbE activities, but diazinon inhibited these enzymes in a dose-responsive manner. In vivo, CbE activity was insensitive to diazinon in younger tadpoles, but inhibited after 2 days of exposure in more developed tadpoles. AChE activity was inhibited after 2 and 7 days of exposure, in a dose-responsive manner. Esterase reactivation by 2-PAM was obtained both in vitro and in vivo. (1) Tadpoles can be adequate sentinel organisms in biomonitoring studies of OP contamination; (2) AChE was more sensitive than CbE to diazinon; (3) tadpoles from earlier developmental stages seems to be less responsive to OPs; (4) AChE activity was sensitive to diazinon in both development stages, being a better OP biomarker.

  2. Dehydrogenases, Acid and Alkaline Phosphatases, and Esterases for Chemotaxonomy of Selected Meloidogyne, Ditylenchus, Heterodera and Aphelenchus spp.

    Science.gov (United States)

    Dickson, D W; Huisingh, D; Sasser, J N

    1971-01-01

    Various taxonomically useful profiles of four dehydrogenases (lactate, malate, glucose-6-phosphate, and a-glycerophosphate) and three hydrolases (acid and alkaline phosphatase and esterase) were detected in whole nematode homogenates of Meloidogynejavanica, M. hapla, M. incognita, M. arenaria, Ditylenchus dipsaci, D. triformis, Heterodera glycines, and Aphelenchus avenae. The enzyme profiles were stable in populations cultured on several different hosts. A tentative enzymically-determined phylogeny of Meloidogyne is given.

  3. Enhanced Agarose and Xylan Degradation for Production of Polyhydroxyalkanoates by Co-Culture of Marine Bacterium, Saccharophagus degradans and Its Contaminant, Bacillus cereus

    Directory of Open Access Journals (Sweden)

    Shailesh S. Sawant

    2017-02-01

    Full Text Available Over reliance on energy or petroleum products has raised concerns both in regards to the depletion of their associated natural resources as well as their increasing costs. Bioplastics derived from microbes are emerging as promising alternatives to fossil fuel derived petroleum plastics. The development of a simple and eco-friendly strategy for bioplastic production with high productivity and yield, which is produced in a cost effective manner utilising abundantly available renewable carbon sources, would have the potential to result in an inexhaustible global energy source. Here we report the biosynthesis of bioplastic polyhydroxyalkanoates (PHAs in pure cultures of marine bacterium, Saccharophagus degradans 2-40 (Sde 2-40, its contaminant, Bacillus cereus, and a co-culture of these bacteria (Sde 2-40 and B. cereus degrading plant and algae derived complex polysaccharides. Sde 2-40 degraded the complex polysaccharides agarose and xylan as sole carbon sources for biosynthesis of PHAs. The ability of Sde 2-40 to degrade agarose increased after co-culturing with B. cereus. The association of Sde 2-40 with B. cereus resulted in increased cell growth and higher PHA production (34.5% of dry cell weight from xylan as a carbon source in comparison to Sde 2-40 alone (22.7% of dry cell weight. The present study offers an innovative prototype for production of PHA through consolidated bioprocessing of complex carbon sources by pure and co-culture of microorganisms.

  4. Effects of the cellulose, xylan and lignin constituents on biomass pyrolysis characteristics and bio-oil composition using the Simplex Lattice Mixture Design method

    International Nuclear Information System (INIS)

    Fan, Yongsheng; Cai, Yixi; Li, Xiaohua; Jiao, Lihua; Xia, Jisheng; Deng, Xiuli

    2017-01-01

    Highlights: • Simplex Lattice Mixture Design was firstly applied to study biomass pyrolysis process. • Interactions between the constituents had effects on the biomass pyrolysis behavior. • Biomass pyrolysis behavior can be predicted based on the ratios of three constituents. • Bio-oil composition was affected by the constituents and their pyrolysis products. - Abstract: In order to clarify the relationships between biomass pyrolysis mechanism and its main constituents. The effects of main constituents on biomass pyrolysis characteristics were firstly determined by thermo-gravimetric analysis based on the Simplex Lattice Mixture Design to investigate that whether the prediction of the pyrolysis behavior of a certain lignocellulosic biomass is possible when its main constituent contents are known. The results showed that there are constituent interactions in the pyrolysis process, which can be intuitively reflected through the change laws of kinetics parameters. The mathematical models for calculating kinetics values were established, and the models were proved to be valid for predicting lignocellulosic biomass pyrolysis behavior. In addition, the effects of biomass constituents on bio-oil compositions were explored by subsequent vacuum pyrolysis experiments. The xylan pyrolysis had a certain inhibitory effect on the pyrolysis of cellulose, and the pyrolysis products of lignin might promote the further decomposition of sugars from cellulose pyrolysis, while the interaction between xylan and lignin had a little effect on the bio-oil composition.

  5. Production of xylitol and tetrahydrofurfuryl alcohol from xylan in napier grass by a hydrothermal process with phosphorus oxoacids followed by aqueous phase hydrogenation.

    Science.gov (United States)

    Takata, Eri; Tsuruoka, Tatsushi; Tsutsumi, Ken; Tsutsumi, Yuji; Tabata, Kenji

    2014-09-01

    The production of xylitol and tetrahydrofurfuryl alcohol (THFA) from napier grass was studied using two steps: a hydrothermal process with phosphorus oxoacids followed by aqueous phase hydrogenation with Pd/C. Xylose obtained from the napier grass by the hydrothermal treatment with 3.0 wt% phosphorous acid was subsequently converted into xylitol at 51.6% yield of the xylan in napier grass by hydrogenation with 5.0 wt% Pd/C. The furfural produced from napier grass with a 3.0 wt% phosphoric acid treatment was also directly subjected to the hydrogenation as a hydrolysate to yield 41.4% THFA based on the xylan in napier grass. The yields of xylitol and THFA obtained by hydrogenation using the napier grass hydrolysate containing xylose or furfural were almost the same as those of hydrogenation using commercial materials. To our knowledge, this is the first report on the production of THFA in high yield by hydrogenation directly from biomass hydrolysate. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Endophytic fungi producing of esterases: evaluation in vitro of the enzymatic activity using pH indicator.

    Science.gov (United States)

    Lisboa, Helen Cristina Fávero; Biasetto, Carolina Rabal; de Medeiros, João Batista; Âraújo, Angela Regina; Silva, Dulce Helena Siqueira; Teles, Helder Lopes; Trevisan, Henrique Celso

    2013-01-01

    A sensitive and efficient colorimetric method was optimized for detection of esterase enzymes produced by endophytic fungi for development of High-Throughput Screening (HTS). The fungi were isolated and obtained previously from plant species of Cerrado and Atlantic Forest located in areas of environmental preservation in the State of Sao Paulo / Brazil, as part of the project "Chemical and biological prospecting endophytic fungi associated to plant species of Cerrado and Atlantic Forest". The compounds ethyl butyrate, ethyl acetate and methyl propionate were used as standards esters which were hydrolyzed by extracellular enzyme from endophytic fungi (EC. 3.1.1.1--carboxyl-esterases) for production of carboxylic acids. Thus, the reduction of the pH increases the protonated indicator concentration (bromothymol blue), changing the color of the reaction medium (from blue to yellow), that can be observed and measured by spectrophotometry at 616 nm. The methodology with acid-base indicator was performed on 13 microorganisms, aiming Periconia atropurpurea as a potential source of esterase for biotransformation of short chain esters. The results also evidenced that this methodology showed to be efficient, fast, cheap, having low consumption of reagents and easy development, and can be applied to screen carboxylic-ester hydrolases in a large number of microorganisms.

  7. Captopril/enalapril inhibit promiscuous esterase activity of carbonic anhydrase at micromolar concentrations: An in vitro study.

    Science.gov (United States)

    Esmaeili, Sajjad; Ashrafi-Kooshk, Mohammad Reza; Adibi, Hadi; Khodarahmi, Reza

    2017-03-01

    The inhibitory activity of captopril, a thiol-containing competitive inhibitor of the angiotensin-converting enzyme, ACE, against esterase activity of carbonic anhydrase, CA was investigated. This small molecule, as well as enalapril, was selected in order to represents both thiol and carboxylate, as two well-known metal binding functional groups of metalloprotein inhibitors. Since captopril, has also been observed to inhibit other metalloenzymes such as tyrosinase and metallo-beta lactamase through binding to the catalytic metal ions and regarding CA as a zinc-containing metallo-enzyme, in the current study, we set out to determine whether captopril/enalapril inhibit CA esterase activity of the purified human CA II or not? Then, we revealed the inhibitors' potencies (IC 50 , K i and K diss values) and also mode of inhibition. Our results also showed that enalapril is more potent CA inhibitor than captopril. Since enalapril represents no sulfhydryl moiety, thus carboxylate group may have a determinant role in inhibiting of CA esterase activity, the conclusion confirmed by molecular docking studies. Additionally, since CA inhibitory potencies of captopril/enalapril were much lower than those of classic sulfonamide drugs, the findings of the current study may explain why these drugs exhibit no effective CA inhibition at the concentrations reached in vivo and also may shed light on the way of generating new class of inhibitors that will discriminately inhibit various CA isoforms. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Contribution of lactic acid bacteria esterases to the release of fatty acids in miniature ewe's milk cheese models.

    Science.gov (United States)

    Abeijón Mukdsi, María C; Medina, Roxana B; Katz, Marta B; Pivotto, Rodolfo; Gatti, Patricia; González, Silvia N

    2009-02-11

    The present work evaluates the contribution of esterase activities of lactic acid bacteria isolated from ewe's dairy products to the release of free fatty acids (FFA) in ewe's milk cheese models. At 60 days of ripening, single-strain cheeses Ov 409 and Ov 421 showed high levels of total FFA (3075 and 2494.62 mg/kg, respectively). Cheeses Ov 227-Ov 409 and Ov 421-Ov 409 presented high percentages of short-chain fatty acids (SCFA). The highest levels of volatile free fatty acids (VFFA) were detected in cheeses Ov 409, Ov 421-Ov 409, and Ov 421-Ov 227. Studies on esterase activities showed that these strains hydrolyzed alpha-naphthyl derivatives of fatty acids from C2 to C6, mainly associated with the wall-membrane fraction. The results showed that the strains studied contributed to the release of FFA during ripening of ewe's milk cheese models. The increase of SCFA throughout ripening involves the action of esterases of starter strains.

  9. Est16, a New Esterase Isolated from a Metagenomic Library of a Microbial Consortium Specializing in Diesel Oil Degradation.

    Science.gov (United States)

    Pereira, Mariana Rangel; Mercaldi, Gustavo Fernando; Maester, Thaís Carvalho; Balan, Andrea; Lemos, Eliana Gertrudes de Macedo

    2015-01-01

    Lipolytic enzymes have attracted attention from a global market because they show enormous biotechnological potential for applications such as detergent production, leather processing, cosmetics production, and use in perfumes and biodiesel. Due to the intense demand for biocatalysts, a metagenomic approach provides methods of identifying new enzymes. In this study, an esterase designated as Est16 was selected from 4224 clones of a fosmid metagenomic library, revealing an 87% amino acid identity with an esterase/lipase (accession number ADM63076.1) from an uncultured bacterium. Phylogenetic studies showed that the enzyme belongs to family V of bacterial lipolytic enzymes and has sequence and structural similarities with an aryl-esterase from Pseudomonas fluorescens and a patented Anti-Kazlauskas lipase (patent number US20050153404). The protein was expressed and purified as a highly soluble, thermally stable enzyme that showed a preference for basic pH. Est16 exhibited activity toward a wide range of substrates and the highest catalytic efficiency against p-nitrophenyl butyrate and p-nitrophenyl valerate. Est16 also showed tolerance to the presence of organic solvents, detergents and metals. Based on molecular modeling, we showed that the large alpha-beta domain is conserved in the patented enzymes but not the substrate pocket. Here, it was demonstrated that a metagenomic approach is suitable for discovering the lipolytic enzyme diversity and that Est16 has the biotechnological potential for use in industrial processes.

  10. A cold active (2R,3R)-(-)-di-O-benzoyl-tartrate hydrolyzing esterase from Rhodotorula mucilaginosa.

    Science.gov (United States)

    Zimmer, Christian; Platz, Tanja; Cadez, Neza; Giffhorn, Friedrich; Kohring, Gert-Wieland

    2006-11-01

    In a screening procedure a pink-colored yeast was isolated from enrichment cultures with (2R,3R)-(-)-di-O-benzoyl-tartrate (benzoyl-tartrate) as the sole carbon source. The organism saar1 was identified by morphological, physiological, and 18S ribosomal DNA/internal transcribed spacer analysis as Rhodotorula mucilaginosa, a basidiomycetous yeast. During growth the yeast hydrolyzed the dibenzoyl ester stoichiometrically to the monoester using the separated benzoate as the growth substrate, before the monoester was further cleaved into benzoate and tartrate, which were both metabolized. The corresponding benzoyl esterase was purified from the culture supernatant and characterized as a monomeric glycosylated 86-kDa protein with an optimum pH of 7.5 and an optimum temperature of 45 degrees C. At 0 degrees C the esterase still exhibited 20% of the corresponding activity at 30 degrees C, which correlates it to psychrophilic enzymes. The esterase could hydrolyze short chain p-nitrophenyl-alkyl esters and several benzoyl esters like benzoyl-methyl ester, ethylene-glycol-dibenzoyl ester, phenyl-benzoyl ester, cocaine, and 1,5-anhydro-D: -fructose-tribenzoyl ester. However feruloyl-ethyl ester was not hydrolyzed. The activity characteristics let the enzyme appear as a promising tool for synthesis of benzoylated compounds for pharmaceutical, cosmetic, or fine chemical applications, even at low temperatures.

  11. Protein Phosphatase Methyl-Esterase PME-1 Protects Protein Phosphatase 2A from Ubiquitin/Proteasome Degradation.

    Science.gov (United States)

    Yabe, Ryotaro; Miura, Akane; Usui, Tatsuya; Mudrak, Ingrid; Ogris, Egon; Ohama, Takashi; Sato, Koichi

    2015-01-01

    Protein phosphatase 2A (PP2A) is a conserved essential enzyme that is implicated as a tumor suppressor based on its central role in phosphorylation-dependent signaling pathways. Protein phosphatase methyl esterase (PME-1) catalyzes specifically the demethylation of the C-terminal Leu309 residue of PP2A catalytic subunit (PP2Ac). It has been shown that PME-1 affects the activity of PP2A by demethylating PP2Ac, but also by directly binding to the phosphatase active site, suggesting loss of PME-1 in cells would enhance PP2A activity. However, here we show that PME-1 knockout mouse embryonic fibroblasts (MEFs) exhibit lower PP2A activity than wild type MEFs. Loss of PME-1 enhanced poly-ubiquitination of PP2Ac and shortened the half-life of PP2Ac protein resulting in reduced PP2Ac levels. Chemical inhibition of PME-1 and rescue experiments with wild type and mutated PME-1 revealed methyl-esterase activity was necessary to maintain PP2Ac protein levels. Our data demonstrate that PME-1 methyl-esterase activity protects PP2Ac from ubiquitin/proteasome degradation.

  12. Est16, a New Esterase Isolated from a Metagenomic Library of a Microbial Consortium Specializing in Diesel Oil Degradation.

    Directory of Open Access Journals (Sweden)

    Mariana Rangel Pereira

    Full Text Available Lipolytic enzymes have attracted attention from a global market because they show enormous biotechnological potential for applications such as detergent production, leather processing, cosmetics production, and use in perfumes and biodiesel. Due to the intense demand for biocatalysts, a metagenomic approach provides methods of identifying new enzymes. In this study, an esterase designated as Est16 was selected from 4224 clones of a fosmid metagenomic library, revealing an 87% amino acid identity with an esterase/lipase (accession number ADM63076.1 from an uncultured bacterium. Phylogenetic studies showed that the enzyme belongs to family V of bacterial lipolytic enzymes and has sequence and structural similarities with an aryl-esterase from Pseudomonas fluorescens and a patented Anti-Kazlauskas lipase (patent number US20050153404. The protein was expressed and purified as a highly soluble, thermally stable enzyme that showed a preference for basic pH. Est16 exhibited activity toward a wide range of substrates and the highest catalytic efficiency against p-nitrophenyl butyrate and p-nitrophenyl valerate. Est16 also showed tolerance to the presence of organic solvents, detergents and metals. Based on molecular modeling, we showed that the large alpha-beta domain is conserved in the patented enzymes but not the substrate pocket. Here, it was demonstrated that a metagenomic approach is suitable for discovering the lipolytic enzyme diversity and that Est16 has the biotechnological potential for use in industrial processes.

  13. Solution behavior and activity of a halophilic esterase under high salt concentration.

    Directory of Open Access Journals (Sweden)

    Lang Rao

    Full Text Available BACKGROUND: Halophiles are extremophiles that thrive in environments with very high concentrations of salt. Although the salt reliance and physiology of these extremophiles have been widely investigated, the molecular working mechanisms of their enzymes under salty conditions have been little explored. METHODOLOGY/PRINCIPAL FINDINGS: A halophilic esterolytic enzyme LipC derived from archeaon Haloarcula marismortui was overexpressed from Escherichia coli BL21. The purified enzyme showed a range of hydrolytic activity towards the substrates of p-nitrophenyl esters with different alkyl chains (n = 2-16, with the highest activity being observed for p-nitrophenyl acetate, consistent with the basic character of an esterase. The optimal esterase activities were found to be at pH 9.5 and [NaCl] = 3.4 M or [KCl] = 3.0 M and at around 45 degrees C. Interestingly, the hydrolysis activity showed a clear reversibility against changes in salt concentration. At the ambient temperature of 22 degrees C, enzyme systems working under the optimal salt concentrations were very stable against time. Increase in temperature increased the activity but reduced its stability. Circular dichroism (CD, dynamic light scattering (DLS and small angle neutron scattering (SANS were deployed to determine the physical states of LipC in solution. As the salt concentration increased, DLS revealed substantial increase in aggregate sizes, but CD measurements revealed the maximal retention of the alpha-helical structure at the salt concentration matching the optimal activity. These observations were supported by SANS analysis that revealed the highest proportion of unimers and dimers around the optimal salt concentration, although the coexistent larger aggregates showed a trend of increasing size with salt concentration, consistent with the DLS data. CONCLUSIONS/SIGNIFICANCE: The solution alpha-helical structure and activity relation also matched the highest proportion of enzyme unimers

  14. 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...

  15. Rabbit N-acetyltransferase 2 genotyping method to investigate role of acetylation polymorphism on N- and O-acetylation of aromatic and heterocyclic amine carcinogens.

    Science.gov (United States)

    Hein, David W; Doll, Mark A

    2017-09-01

    The rabbit was the initial animal model to investigate the acetylation polymorphism expressed in humans. Use of the rabbit model is compromised by lack of a rapid non-invasive method for determining acetylator phenotype. Slow acetylator phenotype in the rabbit results from deletion of the N-acetyltransferase 2 (NAT2) gene. A relatively quick and non-invasive method for identifying the gene deletion was developed and acetylator phenotypes confirmed by measurement of N- and O-acetyltransferase activities in hepatic cytosols. Rabbit liver cytosols catalyzed the N-acetylation of sulfamethazine (p = 0.0014), benzidine (p = 0.0257), 4-aminobiphenyl (p = 0.0012), and the O-acetylation of N-hydroxy-2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N-OH-PhIP; p = 0.002) at rates significantly higher in rabbits possessing NAT2 gene than rabbits with NAT2 gene deleted. In contrast, hepatic cytosols catalyzed the N-acetylation of p-aminobenzoic acid (an N-acetyltransferase 1 selective substrate) at rates that did not differ significantly (p > 0.05) between rabbits positive and negative for NAT2. The new NAT2 genotyping method facilitates use of the rabbit model to investigate the role of acetylator polymorphism in the metabolism of aromatic and heterocyclic amine drugs and carcinogens.

  16. Global proteomic analysis of lysine acetylation in zebrafish (Danio rerio) embryos.

    Science.gov (United States)

    Kwon, Oh Kwang; Kim, Sunjoo; Lee, Sangkyu

    2016-12-01

    Lysine acetylation is an important post-translational modification (PTM). Since the development of MS-based proteomics technology, important roles of lysine acetylation beyond histones have focused on chromatin remodeling during the cell cycle and regulation of nuclear transport, metabolism, and translation. Zebrafish (Danio rerio) is a widely used vertebrate model in genetics and biologic studies. Although studies in several mammalian species have been performed, the mechanism of lysine acetylation in D. rerio embryos is incompletely understood. Here, we investigated the global acetylome in D. rerio embryos by using an MS-based proteomics approach. We identified 351 acetylated peptides and 377 nonredundant acetylation sites on 189 lysine-acetylated proteins in 5-day postfertilization (hpf) embryos of D. rerio. Among lysine-acetylated peptides, 40.2% indicated three motifs: (ac)KxxxK, (ac)KxxxxK, and Lx(ac)K. Of 190 acetylated proteins, 81 (42.6%) were mainly distributed in the cytoplasm. Gene ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway analyses showed that lysine acetylation in D. rerio was enriched in metabolic pathways. Additionally, 17 of 30 acetylated ribosomal proteins were evolutionarily conserved between zebrafish and humans. Our results indicate that acetyllysine might have regulatory effects on ribosomal proteins involved in protein biosynthesis. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Inhibitory effects of four neonicotinoid active ingredients on acetylcholine esterase activity.

    Science.gov (United States)

    Győri, János; Farkas, Anna; Stolyar, Oksana; Székács, András; Mörtl, Mária; Vehovszky, Ágnes

    2017-12-01

    There is a great concern about the decline of pollinators, and neonicotinoids emerging bee disorders are assumed to play a significant role. Since changes in learning ability has been observed in honey bees exposed to some acetylcholine esterase (AChE) inhibitors, we therefore, tested in vitro the effect of four neonicotinoids on purified eel AChE. AChE activity was inhibited in a concentration-dependent manner, and calculated IC 50 values for thiamethoxam (IC 50 = 414 μM) and clothianidin (IC 50 = 160 μM) were found to be much higher compared to acetamiprid (IC 50 = 75.2 μM) and thiacloprid (IC 50 = 87.8 μM). The Lineweaver-Burk reciprocal plots for acetamiprid shows unchanged V max and increased K m values with inhibitor concentrations, while analysis of Michaelis-Menten plots shows predominantly competitive mechanism. The inhibition constant value (K i = 24.3 μM) indicates strong binding of the acetamiprid complex to AChE. Finally, the four tested neonicotinoids are not a uniform group regarding their blocking ability. Our results suggest a previously not established, direct AChE blocking mechanism of neonicotinoids tested, thus the neuronal AChE enzyme is likely among the direct targets of the neonicotinoid insecticides. We conclude, that these AChE inhibitory effects may also contribute to toxic effects on the whole exposed animal.

  18. Glucuronoyl Esterase Screening and Characterization Assays Utilizing Commercially Available Benzyl Glucuronic Acid Ester

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    Hampus Sunner

    2015-09-01

    Full Text Available Research on glucuronoyl esterases (GEs has been hampered by the lack of enzyme assays based on easily obtainable substrates. While benzyl d-glucuronic acid ester (BnGlcA is a commercially available substrate that can be used for GE assays, several considerations regarding substrate instability, limited solubility and low apparent affinities should be made. In this work we discuss the factors that are important when using BnGlcA for assaying GE activity and show how these can be applied when designing BnGlcA-based GE assays for different applications: a thin-layer chromatography assay for qualitative activity detection, a coupled-enzyme spectrophotometric assay that can be used for high-throughput screening or general activity determinations and a HPLC-based detection method allowing kinetic determinations. The three-level experimental procedure not merely facilitates routine, fast and simple biochemical characterizations but it can also give rise to the discovery of different GEs through an extensive screening of heterologous Genomic and Metagenomic expression libraries.

  19. Addition of feruloyl esterase and xylanase produced on-site improves sugarcane bagasse hydrolysis.

    Science.gov (United States)

    Braga, Cleiton Márcio Pinto; Delabona, Priscila da Silva; Lima, Deise Juliana da Silva; Paixão, Douglas Antônio Alvaredo; Pradella, José Geraldo da Cruz; Farinas, Cristiane Sanchez

    2014-10-01

    Accessory enzymes that assist biomass degradation could be used to improve the recovery of fermentable sugar for use in biorefineries. In this study, different fungal strains isolated from the Amazon rainforest were evaluated in terms of their ability to produce feruloyl esterase (FAE) and xylanase enzymes, and an assessment was made of the contributions of the enzymes in the hydrolysis of pretreated sugarcane bagasse. In the selection step, screening using plate assays was followed by shake flask submerged cultivations. After carbon source selection and cultivation in a stirred-tank bioreactor, Aspergillusoryzae P21C3 proved to be a promising strain for production of the enzymes. Supplementation of a commercial enzyme preparation with 30% (v/v) crude enzymatic complex from A. oryzae P21C3 increased the conversion of cellulose derived from pretreated sugarcane bagasse by 36%. Supplementation with FAE and xylanase enzymes produced on-site can therefore be used to improve the hydrolysis of sugarcane bagasse. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. [Anaesthesic management of vaginal delivery in a parturient with C1 esterase deficiency].

    Science.gov (United States)

    Libert, N; Schérier, S; Dubost, C; Franck, L; Rouquette, I; Tortosa, J-C; Rousseau, J-M

    2009-04-01

    Hereditary and acquired angioedema (HAE/AAE) are the clinical translation of a qualitative or a quantitative deficit of C1 esterase inhibitor (C1 INH). The frequency and severity of clinical manifestations vary greatly, ranging from a moderate swelling of the extremities to obstruction of upper airway. Anaesthesiologists and intensivists must be prepared to manage acute manifestations of this disease in case of life-threatening laryngeal edema. Surgery, physical trauma and labour are classical triggers of the disease. The anaesthesiologists should be aware of the drugs used as prophylaxis and treatment of acute attacks when considering labour and caesarean section. Androgens are contraindicated during pregnancy. If prophylaxis is required, tranexamic acid may be used with caution. The safest obstetric approach appears to be to administer a predelivery infusion of C1 INH concentrate. It is important to avoid manipulation of the airway as much as possible by relying on regional techniques. We report the case of a patient suffering from an HAE discovered during pregnancy. The management included administration of C1 INH during labor and early epidural analgesia for pain relief. A short review of the pathophysiology and therapeutic options follows.

  1. Subunit Stabilization and Polyethylene Glycolation of Cocaine Esterase Improves In Vivo Residence Time

    Energy Technology Data Exchange (ETDEWEB)

    Narasimhan, Diwahar; Collins, Gregory T.; Nance, Mark R.; Nichols, Joseph; Edwald, Elin; Chan, Jimmy; Ko, Mei-Chuan; Woods, James H.; Tesmer, John J.G.; Sunahara, Roger K. (Michigan)

    2012-03-15

    No small-molecule therapeutic is available to treat cocaine addiction, but enzyme-based therapy to accelerate cocaine hydrolysis in serum has gained momentum. Bacterial cocaine esterase (CocE) is the fastest known native enzyme that hydrolyzes cocaine. However, its lability at 37 C has limited its therapeutic potential. Cross-linking subunits through disulfide bridging is commonly used to stabilize multimeric enzymes. Herein we use structural methods to guide the introduction of two cysteine residues within dimer interface of CocE to facilitate intermolecular disulfide bond formation. The disulfide-crosslinked enzyme displays improved thermostability, particularly when combined with previously described mutations that enhance stability (T172R-G173Q). The newly modified enzyme yielded an extremely stable form of CocE (CCRQ-CocE) that retained greater than 90% of its activity after 41 days at 37 C, representing an improvement of more than 4700-fold over the wild-type enzyme. CCRQ-CocE could also be modified by polyethylene glycol (PEG) polymers, which improved its in vivo residence time from 24 to 72 h, as measured by a cocaine lethality assay, by self-administration in rodents, and by measurement of inhibition of cocaine-induced cardiovascular effects in rhesus monkeys. PEG-CCRQ elicited negligible immune response in rodents. Subunit stabilization and PEGylation has thus produced a potential protein therapeutic with markedly higher stability both in vitro and in vivo.

  2. HIGHLY METHYL ESTERIFIED SEEDS is a pectin methyl esterase involved in embryo development.

    Science.gov (United States)

    Levesque-Tremblay, Gabriel; Müller, Kerstin; Mansfield, Shawn D; Haughn, George W

    2015-03-01

    Homogalacturonan pectin domains are synthesized in a highly methyl-esterified form that later can be differentially demethyl esterified by pectin methyl esterase (PME) to strengthen or loosen plant cell walls that contain pectin, including seed coat mucilage, a specialized secondary cell wall of seed coat epidermal cells. As a means to identify the active PMEs in seed coat mucilage, we identified seven PMEs expressed during seed coat development. One of these, HIGHLY METHYL ESTERIFIED SEEDS (HMS), is abundant during mucilage secretion, peaking at 7 d postanthesis in both the seed coat and the embryo. We have determined that this gene is required for normal levels of PME activity and homogalacturonan methyl esterification in the seed. The hms-1 mutant displays altered embryo morphology and mucilage extrusion, both of which are a consequence of defects in embryo development. A significant decrease in the size of cells in the embryo suggests that the changes in embryo morphology are a consequence of lack of cell expansion. Progeny from a cross between hms-1 and the previously characterized PME inhibitor5 overexpression line suggest that HMS acts independently from other cell wall-modifying enzymes in the embryo. We propose that HMS is required for cell wall loosening in the embryo to facilitate cell expansion during the accumulation of storage reserves and that its role in the seed coat is masked by redundancy. © 2015 American Society of Plant Biologists. All Rights Reserved.

  3. Acetylcholine esterase activity in mild cognitive impairment and Alzheimer's disease

    Energy Technology Data Exchange (ETDEWEB)

    Herholz, Karl [University of Manchester, Wolfson Molecular Imaging Centre, Clinical Neuroscience, Manchester (United Kingdom); University of Cologne, Cologne (Germany)

    2008-03-15

    Impairment of cholinergic neurotransmission is a well-established fact in Alzheimer's disease (AD), but there is controversy about its relevance at the early stages of the disease and in mild cognitive impairment (MCI). In vivo positron emission tomography imaging of cortical acetylcholine esterase (AChE) activity as a marker of cholinergic innervation that is expressed by cholinergic axons and cholinoceptive neurons has demonstrated a reduction of this enzyme activity in manifest AD. The technique is also useful to measure the inhibition of cerebral AChE induced by cholinesterase inhibitors for treatment of dementia symptoms. A reduction of cortical AchE activity was found consistently in all studies of AD and in few cases of MCI who later concerted to AD. The in vivo findings in MCI and very mild AD are still preliminary, and studies seem to suggest that cholinergic innervation and AChE as the main degrading enzyme are both reduced, which might result in partial compensation of their effect. (orig.)

  4. A cold-adapted, solvent and salt tolerant esterase from marine bacterium Psychrobacter pacificensis.

    Science.gov (United States)

    Wu, Gaobing; Zhang, Xiangnan; Wei, Lu; Wu, Guojie; Kumar, Ashok; Mao, Tao; Liu, Ziduo

    2015-11-01

    Lipolytic enzymes with unique physico-chemical characteristics are gaining more attention for their immense industrial importance. In this study, a novel lipolytic enzyme (Est11) was cloned from the genomic library of a marine bacterium Psychrobacter pacificensis. The enzyme was expressed in Escherichia coli and purified to homogeneity with molecular mass of 32.9kDa. The recombinant Est11 was able to hydrolyze short chain esters (C2-C8) and displayed an optimum activity against butyrate ester (C4). The optimal temperature and pH were 25°C and 7.5, respectively. Est11 retained more than 70% of its original activity at 10°C, suggesting that it was a cold-active esterase. The enzyme was highly active and stable at high concentration of NaCl (5M). Further, incubation with ethanol, isopropanol, propanediol, DMSO, acetonitrile, and glycerol rendered remarkable positive effects on Est11 activity. Typically, even at the concentration of 30% (v/v), ethanol, DMSO, and propanediol increased Est11 activity by 1.3, 2.0, and 2.4-folds, respectively. This new robust enzyme with remarkable properties like cold-adaptability, exceptional tolerance to salt and organic solvents provides us a promising candidate to meet the needs of some harsh industrial processes. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. Pectin Methyl Esterase Activity Change in Intermediate Moisture Sun-Dried Figs after Storage

    Directory of Open Access Journals (Sweden)

    Dilek Demirbüker Kavak

    2015-12-01

    Full Text Available Intermediate moisture fruits can be obtained by rehydrating dried fruits. Intermediate moisture fruits are suitable for direct consumption compared to dry fruits and can be directly used in the production of various products such as bakery products, dairy products and candies. Aim of this study is to compare the pectin methyl esterase (PME activity of intermediate moisture figs which causes softening of the texture and to compare their microbial stability after 3 months storage period. For this purpose, dried figs were rehydrated in 30 and 80° C water until they reach 30% moisture content. Rehydrated samples were stored for 3 months at +4°C. Results showed that there was no statistically significant difference between the control samples and the samples rehydrated at 80°C according to the total viable counts. At the end of the storage period, results of residual PME activity in control samples was 24.1 μmol COOH min-1g-1, while it was found 17.4 μmol COOH min-1g-1 in samples rehydrated at 80°C. As a result rehydration conducted at 80°C provided 28% reduction in PME activity compared to the control samples rehydrated at 30°C, although it did not affect the microbial load significantly after storage.

  6. Separating esterase targets of organophosphorus compounds in the brain by preparative chromatography.

    Science.gov (United States)

    Mangas, I; Vilanova, E; Benabent, M; Estévez, J

    2014-02-10

    Low level exposure to organophosphorus esters (OPs) may cause long-term neurological effects and affect specific cognition domains in experimental animals and humans. Action on known targets cannot explain most of these effects by. Soluble carboxylesterases (EC 3.1.1.1) of chicken brain have been kinetically discriminated using paraoxon, mipafox and phenylmethyl sulfonylfluoride as inhibitors and phenyl valerate as a substrate. Three different enzymatic components were discriminated and called Eα, Eβ and Eγ. In this work, a fractionation procedure with various steps was developed using protein native separation methods by preparative HPLC. Gel permeation chromatography followed by ion exchange chromatography allowed enriched fractions with different kinetic behaviors. The soluble chicken brain fraction was fractionated, while total esterase activity, proteins and enzymatic components Eα, Eβ and Eγ were monitored in each subfraction. After the analysis, 13 fractions were pooled and conserved. Preincubation of the soluble chicken brain fraction of with the organophosphorus mipafox gave rise to a major change in the ion exchange chromatography profile, but not in the molecular exchanged chromatography profile, which suggest that mipafox permanently modifies the ionic properties of numerous proteins. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  7. The role of low levels of juvenile hormone Esterase in the metamorphosis of Manduca sexta

    Directory of Open Access Journals (Sweden)

    M.H. Browder

    2001-10-01

    Full Text Available The activity of juvenile hormone esterase (JHE in feeding fifth instar larvae of Manduca sexta increases gradually with larval weight and rises to a peak after larvae pass the critical weight when juvenile hormone secretion ceases. Starvation of larvae of Manduca sexta (L. that had exceeded the critical weight inhibited peak levels of JHE, but did not delay entry into the wandering stage when larvae leave the plant in search of a pupation site. This suggests that peak levels of JHE may not be essential for the normal timing of metamorphosis. Starved larvae pupated normally, indicating the peak of JHE was not necessary for a morphologically normal pupation. Treatments of larvae with the selective JHE inhibitor O-ethyl-S-phenyl phosphoramidothiolate (EPPAT that began immediately after larvae achieved the critical weight (6.0 to 6.5 grams for our strain of Manduca delayed entry into the wandering stage. By contrast, EPPAT treatment of larvae at weights above 8.0g had no effect on the subsequent timing of the onset of wandering. Therefore, although the normal timing of the onset of wandering does not require peak levels of JHE, it requires low to moderate levels of JHE to be present until larvae reach a weight of about 8.0g.

  8. Characterization and purification of a bacterial chlorogenic acid esterase detected during the extraction of chlorogenic acid from arbuscular mycorrhizal tomato roots.

    Science.gov (United States)

    Negrel, Jonathan; Javelle, Francine; Morandi, Dominique; Lucchi, Géraldine

    2016-12-01

    A Gram-negative bacterium able to grow using chlorogenic acid (5-caffeoylquinic acid) as sole carbon source has been isolated from the roots of tomato plants inoculated with the arbuscular mycorrhizal fungus Rhizophagus irregularis. An intracellular esterase exhibiting very high affinity (K m  = 2 μM) for chlorogenic acid has been extracted and purified by FPLC from the chlorogenate-grown cultures of this bacterium. The molecular mass of the purified esterase determined by SDS-PAGE was 61 kDa and its isoelectric point determined by chromatofocusing was 7.75. The esterase hydrolysed chlorogenic acid analogues (caffeoylshikimate, and the 4- and 3-caffeoylquinic acid isomers), feruloyl esterases substrates (methyl caffeate and methyl ferulate), and even caffeoyl-CoA in vitro but all of them were less active than chlorogenic acid, demonstrating that the esterase is a genuine chlorogenic acid esterase. It was also induced when the bacterial strain was cultured in the presence of hydroxycinnamic acids (caffeic, p-coumaric or ferulic acid) as sole carbon source, but not in the presence of simple phenolics such as catechol or protocatechuic acid, nor in the presence of organic acids such as succinic or quinic acids. The purified esterase was remarkably stable in the presence of methanol, rapid formation of methyl caffeate occurring when its activity was measured in aqueous solutions containing 10-60% methanol. Our results therefore show that this bacterial chlorogenase can catalyse the transesterification reaction previously detected during the methanolic extraction of chlorogenic acid from arbuscular mycorrhizal tomato roots. Data are presented suggesting that colonisation by Rhizophagus irregularis could increase chlorogenic acid exudation from tomato roots, especially in nutrient-deprived plants, and thus favour the growth of chlorogenate-metabolizing bacteria on the root surface or in the mycorhizosphere. Copyright © 2016 Elsevier Masson SAS. All rights

  9. PRODUCTION AND CHARACTERIZATION OF AN ALKALOTHERMOSTABLE, ORGANIC SOLVENT TOLERANT AND SURFACTANT TOLERANT ESTERASE PRODUCED BY A THERMOPHILIC BACTERIUM GEOBACILLUS SP. AGP-04, ISOLATED FROM BAKRESHWAR HOT SPRING, INDIA

    Directory of Open Access Journals (Sweden)

    Amit Ghati

    2013-10-01

    Full Text Available A thermophilic bacteria, Geobacillus sp. AGP-04, isolated from Surya Kund hot spring, Bakreshwar, West Bengal, India was studied in terms of capability of tributyrin hydrolysis and characterization of its thermostable esterase activity using p-nitrophenyl butyrate (PNPB as substrate. The extracellular crude preparation was characterized in terms of pH and temperature optima and stability, organic solvent tolerance capacity and stability, substrate specificity, surfactant tolerance capacity, kinetic parameters and activation/inhibition behavior towards some metal ions and chemicals. Tributyrin agar assay exhibited that Geobacillus sp. AGP-04 secretes an extracellular esterase. The Vmax and Km values of the esterase were found to be 5099 U/Land 103.5µM, respectively in the presence of PNPB as substrate. The optimum temperature and pH, for Geobacillus sp. AGP-04 esterase was 60oC and 8.0, respectively. Although the enzyme activity was not significantly altered by incubating crude extract solution at 20-70oC for 1 hour, the enzyme activity was fully lost at 90oC for same incubation period. The pH stability profile showed that original crude esterase activity is stable at a broad range (pH 5.0-10.0. Moreover, the enzyme was highly organic solvent and surfactant tolerant. The effect of some chemical on crude esterase activity indicated that Geobacillus sp. AGP-04 produce an esterase which contains a serine residue in active site and for its activity -SH groups are essential. Besides, enzyme production was highly induced if fermentation medium contain polysaccharides and oil as carbon source.

  10. Novel Redox-Dependent Esterase Activity (EC 3.1.1.2 for DJ-1: Implications for Parkinson’s Disease

    Directory of Open Access Journals (Sweden)

    Emmanuel Vázquez-Mayorga

    2016-08-01

    Full Text Available Mutations the in human DJ-1 (hDJ-1 gene are associated with early-onset autosomal recessive forms of Parkinson’s disease (PD. hDJ-1/parkinsonism associated deglycase (PARK7 is a cytoprotective multi-functional protein that contains a conserved cysteine-protease domain. Given that cysteine-proteases can act on both amide and ester substrates, we surmised that hDJ-1 possessed cysteine-mediated esterase activity. To test this hypothesis, hDJ-1 was overexpressed, purified and tested for activity towards 4-nitrophenyl acetate (pNPA as µmol of pNPA hydrolyzed/min/mg·protein (U/mg protein. hDJ-1 showed maximum reaction velocity esterase activity (Vmax = 235.10 ± 12.00 U/mg protein, with a sigmoidal fit (S0.5 = 0.55 ± 0.040 mM and apparent positive cooperativity (Hill coefficient of 2.05 ± 0.28. A PD-associated mutant of DJ-1 (M26I lacked activity. Unlike its protease activity which is inactivated by reactive oxygen species (ROS, esterase activity of hDJ-1 is enhanced upon exposure to low concentrations of hydrogen peroxide (<10 µM and plateaus at elevated concentrations (>100 µM suggesting that its activity is resistant to oxidative stress. Esterase activity of DJ-1 requires oxidation of catalytic cysteines, as chemically protecting cysteines blocked its activity whereas an oxido-mimetic mutant of DJ-1 (C106D exhibited robust esterase activity. Molecular docking studies suggest that C106 and L126 within its catalytic site interact with esterase substrates. Overall, our data show that hDJ-1 contains intrinsic redox-sensitive esterase activity that is abolished in a PD-associated mutant form of the hDJ-1 protein.

  11. Evaluation of gels obtained from acetylation of chitosan in heterogeneous medium

    International Nuclear Information System (INIS)

    Garcia, Rosangela Balaban; Silva, Dayse Luzia Pinheiro da; Costa, Marta

    2008-01-01

    Chitosan was acetylated during 2, 5 and 10 h and physical gels were obtained at different polymer concentrations in N,N-dimethylacetamide containing 5% of LiCl. Acetylation was confirmed by infrared spectroscopy and 13 C NMR, and degrees of acetylation in the range of 0.82-0.91 were determined by NMR. The O-acetylation degree (0.12-0.15) was exclusively determined by a volumetric method. Rheological studies showed that the storage modulus values were smaller for the more acetylated samples and increased with the temperature and the polymer concentration. All the gels presented storage modulus superior to loss modulus, evidencing more elastic than viscous characteristics. The results obtained in this work suggest a gelation process based on a balance between O and N-acetylation and intermolecular bonds. (author)

  12. Kinetic Modelling and Experimental Studies for the Effects of Fe2+ Ions on Xylan Hydrolysis with Dilute-Acid Pretreatment and Subsequent Enzymatic Hydrolysis

    Directory of Open Access Journals (Sweden)

    Hui Wei

    2018-01-01

    Full Text Available High-temperature (150–170 °C pretreatment of lignocellulosic biomass with mineral acids is well established for xylan breakdown. Fe2+ is known to be a cocatalyst of this process although kinetics of its action remains unknown. The present work addresses the effect of ferrous ion concentration on sugar yield and degradation product formation from corn stover for the entire two-step treatment, including the subsequent enzymatic cellulose hydrolysis. The feedstock was impregnated with 0.5% acid and 0.75 mM iron cocatalyst, which was found to be optimal in preliminary experiments. The detailed kinetic data of acid pretreatment, with and without iron, was satisfactorily modelled with a four-step linear sequence of first-order irreversible reactions accounting for the formation of xylooligomers, xylose and furfural as intermediates to provide the values of Arrhenius activation energy. Based on this kinetic modelling, Fe2+ turned out to accelerate all four reactions, with a significant alteration of the last two steps, that is, xylose degradation. Consistent with this model, the greatest xylan conversion occurred at the highest severity tested under 170 °C/30 min with 0.75 mM Fe2+, with a total of 8% xylan remaining in the pretreated solids, whereas the operational conditions leading to the highest xylose monomer yield, 63%, were milder, 150 °C with 0.75 mM Fe2+ for 20 min. Furthermore, the subsequent enzymatic hydrolysis with the prior addition of 0.75 mM of iron(II increased the glucose production to 56.3% from 46.3% in the control (iron-free acid. The detailed analysis indicated that conducting the process at lower temperatures yet long residence times benefits the yield of sugars. The above kinetic modelling results of Fe2+ accelerating all four reactions are in line with our previous mechanistic research showing that the pretreatment likely targets multiple chemistries in plant cell wall polymer networks, including those represented by the C

  13. (1-Acetyl-2,6-diphenylpiperidin-4-ylidene(phenylacetonitrile

    Directory of Open Access Journals (Sweden)

    R. J. Butcher

    2008-05-01

    Full Text Available In the title molecule, C27H24N2O, the piperidine ring adopts a boat conformation. The acetyl group at position 1 has a bisectional orientation. The two phenyl rings attached to the piperidine ring at positions 2 and 6 have bisectional and axial orientations, respectively, and make a dihedral angle of 75.27 (10°. The phenylacetonitrile group at position 4 has an equatorial orientation. Molecules are linked by C—H...N, C—H...O intermolecular and C—H...π interactions. A C—H...O intramolecular interaction is also found in the molecule.

  14. Epigenetic Readers of Lysine Acetylation Regulate Cocaine-Induced Plasticity

    Science.gov (United States)

    Sartor, Gregory C.; Powell, Samuel K.; Brothers, Shaun P.

    2015-01-01

    Epigenetic processes that regulate histone acetylation play an essential role in behavioral and molecular responses to cocaine. To date, however, only a small fraction of the mechanisms involved in the addiction-associated acetylome have been investigated. Members of the bromodomain and extraterminal (BET) family of epigenetic “reader” proteins (BRD2, BRD3, BRD4, and BRDT) bind acetylated histones and serve as a scaffold for the recruitment of macromolecular complexes to modify chromatin accessibility and transcriptional activity. The role of BET proteins in cocaine-induced plasticity, however, remains elusive. Here, we used behavioral, pharmacological, and molecular techniques to examine the involvement of BET bromodomains in cocaine reward. Of the BET proteins, BRD4, but not BRD2 or BRD3, was significantly elevated in the nucleus accumbens (NAc) of mice and rats following repeated cocaine injections and self-administration. Systemic and intra-accumbal inhibition of BRD4 with the BET inhibitor, JQ1, attenuated the rewarding effects of cocaine in a conditioned place preference procedure but did not affect conditioned place aversion, nor did JQ1 alone induce conditioned aversion or preference. Investigating the underlying mechanisms, we found that repeated cocaine injections enhanced the binding of BRD4, but not BRD3, to the promoter region of Bdnf in the NAc, whereas systemic injection of JQ1 attenuated cocaine-induced expression of Bdnf in the NAc. JQ1 and siRNA-mediated knockdown of BRD4 in vitro also reduced expression of Bdnf. These findings indicate that disrupting the interaction between BET proteins and their acetylated lysine substrates may provide a new therapeutic avenue for the treatment of drug addiction. SIGNIFICANCE STATEMENT Proteins involved in the “readout” of lysine acetylation marks, referred to as BET bromodomain proteins (including BRD2, BRD3, BRD4, and BRDT), have been shown to be key regulators of chromatin dynamics and disease, and

  15. Epigenetic Readers of Lysine Acetylation Regulate Cocaine-Induced Plasticity.

    Science.gov (United States)

    Sartor, Gregory C; Powell, Samuel K; Brothers, Shaun P; Wahlestedt, Claes

    2015-11-11

    Epigenetic processes that regulate histone acetylation play an essential role in behavioral and molecular responses to cocaine. To date, however, only a small fraction of the mechanisms involved in the addiction-associated acetylome have been investigated. Members of the bromodomain and extraterminal (BET) family of epigenetic "reader" proteins (BRD2, BRD3, BRD4, and BRDT) bind acetylated histones and serve as a scaffold for the recruitment of macromolecular complexes to modify chromatin accessibility and transcriptional activity. The role of BET proteins in cocaine-induced plasticity, however, remains elusive. Here, we used behavioral, pharmacological, and molecular techniques to examine the involvement of BET bromodomains in cocaine reward. Of the BET proteins, BRD4, but not BRD2 or BRD3, was significantly elevated in the nucleus accumbens (NAc) of mice and rats following repeated cocaine injections and self-administration. Systemic and intra-accumbal inhibition of BRD4 with the BET inhibitor, JQ1, attenuated the rewarding effects of cocaine in a conditioned place preference procedure but did not affect conditioned place aversion, nor did JQ1 alone induce conditioned aversion or preference. Investigating the underlying mechanisms, we found that repeated cocaine injections enhanced the binding of BRD4, but not BRD3, to the promoter region of Bdnf in the NAc, whereas systemic injection of JQ1 attenuated cocaine-induced expression of Bdnf in the NAc. JQ1 and siRNA-mediated knockdown of BRD4 in vitro also reduced expression of Bdnf. These findings indicate that disrupting the interaction between BET proteins and their acetylated lysine substrates may provide a new therapeutic avenue for the treatment of drug addiction. Proteins involved in the "readout" of lysine acetylation marks, referred to as BET bromodomain proteins (including BRD2, BRD3, BRD4, and BRDT), have been shown to be key regulators of chromatin dynamics and disease, and BET inhibitors are currently

  16. 2-acetyl-1-pyrroline - key aroma compound in Mediterranean dried sausages

    DEFF Research Database (Denmark)

    Stahnke, Marie Louise Heller

    2000-01-01

    and Southern types were attributed to a burned coffee odour from smoke in the smoked sausages and a popcorn note in the Mediterranean products covered with mould. The two compounds were 2-furfurylthiol and 2-acetyl-1-pyrroline, respectively. An analysis of five dried, moulded sausages showed that the surface...... edge of the sausages contained higher amounts of 2-acetyl-1-pyrroline than the core, indicating that the mould growing on the surface of Mediterranean products produces 2-acetyl-1-pyrroline....

  17. Lysine acetylation targets protein complexes and co-regulates major cellular functions

    DEFF Research Database (Denmark)

    Choudhary, Chuna Ram; Kumar, Chanchal; Gnad, Florian

    2009-01-01

    Lysine acetylation is a reversible posttranslational modification of proteins and plays a key role in regulating gene expression. Technological limitations have so far prevented a global analysis of lysine acetylation's cellular roles. We used high-resolution mass spectrometry to identify 3600......, cell cycle, splicing, nuclear transport, and actin nucleation. Acetylation impaired phosphorylation-dependent interactions of 14-3-3 and regulated the yeast cyclin-dependent kinase Cdc28. Our data demonstrate that the regulatory scope of lysine acetylation is broad and comparable with that of other...

  18. Lysine Acetylation of CREBH Regulates Fasting-Induced Hepatic Lipid Metabolism

    Science.gov (United States)

    Kim, Hyunbae; Mendez, Roberto; Chen, Xuequn; Fang, Deyu

    2015-01-01

    Cyclic AMP-responsive element-binding protein 3-like 3, hepatocyte specific (CREBH), is a hepatic transcription factor that functions as a key regulator of energy homeostasis. Here, we defined a regulatory CREBH posttranslational modification process, namely, lysine-specific acetylation, and its functional involvement in fasting-induced hepatic lipid metabolism. Fasting induces CREBH acetylation in mouse livers in a time-dependent manner, and this event is critical for CREBH transcriptional activity in regulating hepatic lipid homeostasis. The histone acetyltransferase PCAF-mediated acetylation and the deacetylase sirtuin-1-mediated deacetylation coexist to maintain CREBH acetylation states under fasting conditions. Site-directed mutagenesis and functional analyses revealed that the lysine (K) residue at position 294 (K294) within the bZIP domain of the CREBH protein is the site where fasting-induced acetylation/deacetylation occurs. Introduction of the acetylation-deficient (K294R) or acetylation-mimicking (K294Q) mutation inhibited or enhanced CREBH transcriptional activity, respectively. Importantly, CREBH acetylation at lysine 294 was required for the interaction and synergy between CREBH and peroxisome proliferator-activated receptor α (PPARα) in activating their target genes upon fasting or glucagon stimulation. Introduction of the CREBH lysine 294 mutation in the liver leads to hepatic steatosis and hyperlipidemia in animals under prolonged fasting. In summary, our study reveals a molecular mechanism by which fasting or glucagon stimulation modulates lipid homeostasis through acetylation of CREBH. PMID:26438600

  19. Metabolism of 1-acyl-2-acetyl-sn-glycero-3-phosphocholine in the human neutrophil

    International Nuclear Information System (INIS)

    Triggiani, M.; D'Souza, D.M.; Chilton, F.H.

    1991-01-01

    The biosynthesis of 1-acyl-2-acetyl-sn-glycero-3-phosphocholine (1-acyl-2-acetyl-GPC) together with that of 1-alkyl-2-acetyl-GPC (platelet-activating factor) has been demonstrated in a variety of inflammatory cells and tissues. It has been hypothesized that the relative proportion of these phospholipids produced upon cell activation may be influenced by their rates of catabolism. We studied the catabolism of 1-acyl-2-acetyl-GPC in resting and activated human neutrophils and compared it to that of 1-alkyl-2-acetyl-GPC. Neutrophils rapidly catabolize both 1-alkyl-2-acetyl-GPC and 1-acyl-2-acetyl-GPC; however, the rate of catabolism of 1-acyl-2-acetyl-GPC is approximately 2-fold higher than that of 1-alkyl-2-acetyl-GPC. In addition, most of 1-acyl-2-acetyl-GPC is catabolized through a pathway different from that of 1-alkyl-2-acetyl-GPC. The main step in the catabolism of 1-acyl-2-acetyl-GPC is the removal of the long chain at the sn-1 position; the long chain residue is subsequently incorporated either into triglycerides or into phosphatidylcholine. The 1-lyso-2-acetyl-GPC formed in this reaction is then further degraded to glycerophosphocholine, choline, or phosphocholine. 1-Acyl-2-acetyl-GPC is also catabolized, to a lesser extent, through deacetylation at the sn-2 position and reacylation with a long chain fatty acid. Stimulation of neutrophils by A23187 results in a higher rate of catabolism of 1-acyl-2-acetyl-GPC by increasing both the removal of the long chain at the sn-1 position and the deacetylation-reacylation at the sn-2 position. In a broken cell preparation, the cytosolic fraction of the neutrophil was shown to contain an enzyme activity which cleaved the sn-1 position of 1-acyl-2-acetyl-GPC and 1-acyl-2-lyso-GPC but not of 1,2-diacyl-GPC

  20. Aspirin-mediated acetylation induces structural alteration and aggregation of bovine pancreatic insulin.

    Science.gov (United States)

    Yousefi, Reza; Taheri, Behnaz; Alavi, Parnian; Shahsavani, Mohammad Bagher; Asadi, Zahra; Ghahramani, Maryam; Niazi, Ali; Alavianmehr, Mohammad Mehdi; Moosavi-Movahedi, Ali Akbar

    2016-01-01

    The simple aggregation of insulin under various chemical and physical stresses is still an important challenge for both pharmaceutical production and clinical formulation. In the storage form, this protein is subjected to various chemical modifications which alter its physicochemical and aggregation properties. Aspirin (acetylsalicylic acid) which is the most widely used medicine worldwide has been indicated to acetylate a large number of proteins both in vitro and in vivo. In this study, as insulin treated with aspirin at 37°C, a significant level of acetylation was observed by flourescamine and o-phthalaldehyde assay. Also, different spectroscopic techniques, gel electrophoresis, and microscopic assessment were applied to compare the structural variation and aggregation/fibrillation propensity among acetylated and non-acetylated insulin samples. The results of spectroscopic assessments elucidate that acetylation induces insulin unfolding which is accompanied with the exposure of protein hydrophobic patches, a transition from alpha-helix to beta-sheet and increased propensity of the protein for aggregation. The kinetic studies propose that acetylation increases aggregation rate of insulin under both thermal and chemical stresses. Also, gel electrophoresis and dynamic light scattering experiments suggest that acetylation induces insulin oligomerization. Additionally, the results of Thioflavin T fluorescence study, Congo red absorption assessment, and microscopic analysis suggest that acetylation with aspirin enhances the process of insulin fibrillation. Overall, the increased susceptibility of acetylated insulin for aggregation may reflect the fact that this type of modification has significant structural destabilizing effect which finally makes the protein more vulnerable for pathogenic aggregation/fibrillation.

  1. Treadmill exercise induces selective changes in hippocampal histone acetylation during the aging process in rats.

    Science.gov (United States)

    de Meireles, Louisiana Carolina Ferreira; Bertoldi, Karine; Cechinel, Laura Reck; Schallenberger, Bruna Luisa; da Silva, Vanessa Kappel; Schröder, Nadja; Siqueira, Ionara Rodrigues

    2016-11-10

    Physical exercise and the aging process have been shown to induce opposite effects on epigenetic marks, such as histone acetylation. The impact of exercise on hippocampal histone acetylation on specific lysine residues, especially during the aging process, is rarely studied. The aim of this study was to investigate the effect of treadmill exercise (20min/day during 2 weeks) on H3K9, H4K5 and H4K12 acetylation levels in hippocampi of young adult and aged rats. Male Wistar rats aged 3 or 20-21 months were assigned to sedentary and exercise groups. Single-trial step-down inhibitory avoidance conditioning was employed as an aversive memory paradigm. Hippocampal H3K9, H4K5 and H4K12 acetylation was determined by Western blotting. The daily moderate exercise protocol improved the aversive memory performance and increased hipocampal H4K12 acetylation levels in both tested ages. Exercise was also able to increase H3K9 acetylation levels in aged rats. An age-related decline in memory performance was observed, without any effect of the aging process on histone acetylation state. Our data suggest that treadmill exercise can impact hippocampal the histone acetylation profile in an age- and lysine-dependent manner. In addition, higher hippocampal H4K12 acetylation levels at both ages may be related to improvement of aversive memory performance. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  2. Production of xylooligosaccharides from enzymatic hydrolysis of xylan by white-rot fungi Pleurotus = Produção de xilooligossacarídeos pela hidrólise enzimática de xylana por fungos Pleurotus

    Directory of Open Access Journals (Sweden)

    Cristiano Ragagnin de Menezes

    2010-01-01

    Full Text Available Hemicellulose consists of non-cellulosic polysaccharides, with xylans and mannans as their main examples. In nature, xylan can be first degraded to xylooligosaccharides and finally to xylose by certain microorganisms. White-rot fungi basidiomycetes Pleurotus sp. BCCB068 and Pleurotus tailandia were used to degrade oat-spelts xylan under submerged fermentation for a period of 40 days. The study obtained activities of endo-1,4-β-xylanase and β-xylosidase and determination of xylan products by degradation. The fungi reached significant levels of xylan degradation by Pleurotus sp. BCCB068 (75.1% and P. tailandia (73.4%, following formations of xylooligosaccharides and sugar monomers. These Pleurotus strains proved to be a feasible alternative for biotechnological processes related to degradation of hemicellulose sources. A hemicelulose é um polissacarídeo não-celulósico, tendo como exemplos principais as xilanas e mananas. Na natureza, as xilanas podem ser degradadas por microrganismos, primeiramente a xilooligossacarídeos e finalmente a xilose. Fungos basidiomicetos Pleurotus sp. BCCB068 e Pleurotus tailandia foram utilizados para degradar xilana de aveia em fermentação submersa durante o período de 40 dias. Foram obtidas as atividades de endo-1,4-β-xilanase e β-xilosidase e a determinação dos produtos de degradação da xilana. Os fungos atingiram níveis significativos de degradação da xilana porPleurotus sp. BCCB068 (75.1% and P. tailandia (73.4%, seguido da formação de xilooligossacarídeos e monômeros de açúcar. Essas cepas de Pleurotus demonstraram ser uma alternativa viável para os processos biotecnológicos relacionados à degradação de fontes dehemicelulose.

  3. β-D-(1→4), β-D-(1→3) 'mixed linkage' xylans from red seaweeds of the order Nemaliales and Palmariales.

    Science.gov (United States)

    Viana, Adriano G; Noseda, Miguel D; Gonçalves, Alan G; Duarte, Maria Eugênia R; Yokoya, Nair; Matulewicz, Maria C; Cerezo, Alberto S

    2011-06-01

    Xylans from five seaweeds belonging to the order Nemaliales (Galaxaura marginata, Galaxaura obtusata, Tricleocarpacylindrica, Tricleocarpa fragilis, and Scinaia halliae) and one of the order Palmariales (Palmaria palmata) collected on the Brazilian coasts were extracted with hot water and purified from acid xylomannans and/or xylogalactans through Cetavlon precipitation of the acid polysaccharides. The β-D-(1→4), β-D-(1→3) 'mixed linkage' structures were determined using methylation analysis and 1D and 2D NMR spectroscopy. The presence of large sequences of β-(1→4)-linked units suggests transient aggregates of ribbon- or helical-ordered structures that would explain the low optical rotations. Copyright © 2011 Elsevier Ltd. All rights reserved.

  4. Deletion of the pflA gene in Escherichia coli LS5218 and its effects on the production of polyhydroxyalkanoates using beechwood xylan as a feedstock.

    Science.gov (United States)

    Salamanca-Cardona, Lucia; Scheel, Ryan A; Lundgren, Benjamin R; Stipanovic, Arthur J; Matsumoto, Ken'ichiro; Taguchi, Seiichi; Nomura, Christopher T

    2014-01-01

    Engineering of microorganisms to directly utilize plant biomass as a feedstock for the biosynthesis of value-added products such as bioplastics is the aim of consolidated bioprocessing. In previous research we successfully engineered E. coli LS5218 to produce polyhydroxyalkanoates (PHAs) from xylan. In this study we report further genetic modifications to Escherichia coli LS5218 in order to increase the lactic acid (LA) fraction in poly(lactic acid-co-3-hydroxyalkanoate) P(LA-co-HA) copolymers. Deletion of the pflA gene resulted in increased content of LA repeating units in the copolymers by over 3-fold compared with the wild type; however, this increase was offset by reduced yields in cell mass. Additionally, when acetate was used as a feedstock LA monomer incorporation reached 18.5 (mol%), which suggests that acetate can be used as a feedstock for the production of P(LA-co-HA) copolymers by E. coli.

  5. The structure of the complex between a branched pentasaccharide and Thermobacillus xylanilyticus GH-51 arabinofuranosidase reveals xylan-binding determinants and induced fit

    DEFF Research Database (Denmark)

    Paës, Gabriel; Skov, Lars K; O'Donohue, Michael J

    2008-01-01

    substrate xylan. The overall structure shows the two characteristic GH-51 domains: a catalytic domain that is folded into a (beta/alpha) 8-barrel and a C-terminal domain that displays jelly roll architecture. The pentasaccharide is bound in a groove on the surface of the enzyme, with the mono arabinosyl....... In the absence of substrate, the enzyme adopts an open conformation. In the substrate-bound form, the long loop connecting beta-strand 2 to alpha-helix 2 closes the active site and interacts with the substrate through residues His98 and Trp99. The results of kinetic and fluorescence titration studies using......The crystal structure of the family GH-51 alpha- l-arabinofuranosidase from Thermobacillus xylanilyticus has been solved as a seleno-methionyl derivative. In addition, the structure of an inactive mutant Glu176Gln is presented in complex with a branched pentasaccharide, a fragment of its natural...

  6. The Metabolic Fate of Deoxynivalenol and Its Acetylated Derivatives in a Wheat Suspension Culture: Identification and Detection of DON-15-O-Glucoside, 15-Acetyl-DON-3-O-Glucoside and 15-Acetyl-DON-3-Sulfate

    Directory of Open Access Journals (Sweden)

    Clemens Schmeitzl

    2015-08-01

    Full Text Available Deoxynivalenol (DON is a protein synthesis inhibitor produced by the Fusarium species, which frequently contaminates grains used for human or animal consumption. We treated a wheat suspension culture with DON or one of its acetylated derivatives, 3-acetyl-DON (3-ADON, 15-acetyl-DON (15-ADON and 3,15-diacetyl-DON (3,15-diADON, and monitored the metabolization over a course of 96 h. Supernatant and cell extract samples were analyzed using a tailored LC-MS/MS method for the quantification of DON metabolites. We report the formation of tentatively identified DON-15-O-β-D-glucoside (D15G and of 15-acetyl-DON-3-sulfate (15-ADON3S as novel deoxynivalenol metabolites in wheat. Furthermore, we found that the recently identified 15-acetyl-DON-3-O-β-D-glucoside (15-ADON3G is the major metabolite produced after 15-ADON challenge. 3-ADON treatment led to a higher intracellular content of toxic metabolites after six hours compared to all other treatments. 3-ADON was exclusively metabolized into DON before phase II reactions occurred. In contrast, we found that 15-ADON was directly converted into 15-ADON3G and 15-ADON3S in addition to metabolization into deoxynivalenol-3-O-β-D-glucoside (D3G. This study highlights significant differences in the metabolization of DON and its acetylated derivatives.

  7. Organophosphates induce distal axonal damage, but not brain oedema, by inactivating neuropathy target esterase

    International Nuclear Information System (INIS)

    Read, David J.; Li Yong; Chao, Moses V.; Cavanagh, John B.; Glynn, Paul

    2010-01-01

    Single doses of organophosphorus compounds (OP) which covalently inhibit neuropathy target esterase (NTE) can induce lower-limb paralysis and distal damage in long nerve axons. Clinical signs of neuropathy are evident 3 weeks post-OP dose in humans, cats and chickens. By contrast, clinical neuropathy in mice following acute dosing with OPs or any other toxic compound has never been reported. Moreover, dosing mice with ethyloctylphosphonofluoridate (EOPF) - an extremely potent NTE inhibitor - causes a different (subacute) neurotoxicity with brain oedema. These observations have raised the possibility that mice are intrinsically resistant to neuropathies induced by acute toxic insult, but may incur brain oedema, rather than distal axonal damage, when NTE is inactivated. Here we provide the first report that hind-limb dysfunction and extensive axonal damage can occur in mice 3 weeks after acute dosing with a toxic compound, bromophenylacetylurea. Three weeks after acutely dosing mice with neuropathic OPs no clinical signs were observed, but distal lesions were present in the longest spinal sensory axons. Similar lesions were evident in undosed nestin-cre:NTEfl/fl mice in which NTE had been genetically-deleted from neural tissue. The extent of OP-induced axonal damage in mice was related to the duration of NTE inactivation and, as reported in chickens, was promoted by post-dosing with phenylmethanesulfonylfluoride. However, phenyldipentylphosphinate, another promoting compound in chickens, itself induced in mice lesions different from the neuropathic OP type. Finally, EOPF induced subacute neurotoxicity with brain oedema in both wild-type and nestin-cre:NTEfl/fl mice indicating that the molecular target for this effect is not neural NTE.

  8. Effect of C1-Esterase-inhibitor in angiotensin-converting enzyme inhibitor-induced angioedema.

    Science.gov (United States)

    Greve, Jens; Bas, Murat; Hoffmann, Thomas K; Schuler, Patrick J; Weller, Patrick; Kojda, Georg; Strassen, Ulrich

    2015-06-01

    The study objective was to generate pilot data to evaluate the effectiveness and safety of C1-esterase-inhibitor concentrate (C1-INH) compared to standard treatment in patients with angiotensin-converting enzyme inhibitor (ACEi)-induced angioedema affecting the upper aerodigestive tract. Proof-of-concept case series with historical control. Adult patients with angioedema in the upper aerodigestive tract presenting to the emergency department were included. After establishing the diagnosis of ACEi-induced angioedema based on patient history and thorough clinical examination, all patients were administered 1,000 international units (IU) of C1-INH intravenously. A historical control group consisting of adult patients with ACEi-induced angioedema who had been treated with intravenous corticosteroids and antihistamines at the same institution over the past 8 years was used for comparison. The most important parameters assessed were the time to complete resolution of symptoms and the need for intubation or tracheotomy. Ten patients were included in the C1-INH group and 47 in the corticosteroid/antihistamine group. The time to complete resolution of symptoms was considerably longer in the historical control group (33.1 ± 19.4 hours) than in the C1-INH group (10.1 ± 3.0 hours). No intubation or tracheotomy was needed in the C1-INH group (0/10 patients), whereas three out of the 47 historical controls required tracheotomy and two were intubated (5/47). The results suggest a role for C1-INH as an effective and safe therapeutic option in patients with ACEi-induced angioedema, which needs to be confirmed by further larger and double-blinded studies. 4. © 2015 The American Laryngological, Rhinological and Otological Society, Inc.

  9. A stereospecific carboxyl esterase from Bacillus coagulans hosting nonlipase activity within a lipase-like fold.

    Science.gov (United States)

    De Vitis, Valerio; Nakhnoukh, Cristina; Pinto, Andrea; Contente, Martina L; Barbiroli, Alberto; Milani, Mario; Bolognesi, Martino; Molinari, Francesco; Gourlay, Louise J; Romano, Diego

    2018-03-01

    Microbial carboxylesterases are important biocatalysts that selectively hydrolyze an extensive range of esters. Here, we report the biochemical and structural characterization of an atypical carboxylesterase from Bacillus coagulans (BCE), endowed with high enantioselectivity toward different 1,2-O-isopropylideneglycerol (IPG or solketal) esters. BCE efficiently catalyzes the production of enantiopure (S)-IPG, a chiral building block for the synthesis of β-blockers, glycerophospholipids, and prostaglandins; efficient hydrolysis was observed up to 65 °C. To gain insight into the mechanistic bases of such enantioselectivity, we solved the crystal structures of BCE in apo- and glycerol-bound forms at resolutions of 1.9 and 1.8 Å, respectively. In silico docking studies on the BCE structure confirmed that IPG esters with small acyl chains (≤ C6) were easily accommodated in the active site pocket, indicating that small conformational changes are necessary to accept longer substrates. Furthermore, docking studies suggested that enantioselectivity may be due to an improved stabilization of the tetrahedral reaction intermediate for the S-enantiomer. Contrary to the above functional data implying nonlipolytic functions, BCE displays a lipase-like 3D structure that hosts a "lid" domain capping the main entrance to the active site. In lipases the lid mediates catalysis through interfacial activation, a process that we did not observe for BCE. Overall, we present the functional-structural properties of an atypical carboxyl esterase that has nonlipase-like functions, yet possesses a lipase-like 3D fold. Our data provide original enzymatic information in view of BCE applications as an inexpensive, efficient biocatalyst for the production of enantiopure (S)-IPG. Coordinates and structure factors have been deposited in the Protein Data Bank (www.rcsb.org) under accession numbers 5O7G (apo-BCE) and 5OLU (glycerol-bound BCE). © 2017 Federation of European Biochemical

  10. Immobilization of cholesterol esterase in mesoporous silica materials and its hydrolytic activity toward diethyl phthalate

    Energy Technology Data Exchange (ETDEWEB)

    Orita, Toru, E-mail: nqj45366@nifty.com [Division of Chemistry for Materials, Graduate School of Engineering, Mie University, 1577 Kurimamachiya-cho, Tsu, Mie 514-8570 (Japan); Taiyo Kagaku Co. Ltd., 800 Yamada-cho, Yokkaichi, Mie 512-1111 (Japan); Tomita, Masahiro [Division of Chemistry for Materials, Graduate School of Engineering, Mie University, 1577 Kurimamachiya-cho, Tsu, Mie 514-8570 (Japan); Saito, Takao; Nishida, Nasakazu; Kato, Katsuya [National Institute of Advanced Industrial Science and Technology, 2266-78 Anagahora, Moriyamaku, Nagoya, Aichi 463-8560 (Japan)

    2012-05-01

    Cholesterol esterase (CE, cholesteryl ester hydrolase, EC 3.1.1.13) from porcine pancreas (molecular weight 400-500 kDa) exhibits hydrolytic activity toward various toxic organic phthalate esters. CE was confined in the nanospace (diameter 3-30 nm) of five types of mesoporous silica (MPS) that differ in structural properties such as pore diameter, pore volume, and particle morphology. These structural properties were characterized by transmission electron microscopy, small-angle X-ray diffraction, N{sub 2} adsorption-desorption experiments, solid-state {sup 13}C nuclear magnetic resonance (NMR), and solid-state {sup 29}Si NMR. Catalytic activities of immobilized and free CE were evaluated by the hydrolysis of diethyl phthalate in phosphate buffer solutions containing an organic cosolvent. Optimal activity recovery was achieved when CE was immobilized in n-decane-functionalized MPS, which had a large pore size (22.5 nm). The immobilization also protected against effects of temperature within the range 30 Degree-Sign C-60 Degree-Sign C; CE immobilized in n-decyl-functionalized MPS exhibited better thermal stability than in non-functionalized MPS or free CE. Moreover, it retained approximately 60% of its catalytic activity even after six catalytic cycles. - Highlights: Black-Right-Pointing-Pointer The highest activity of immobilized CE was shown in MPS with a pore size of 22.5 nm. Black-Right-Pointing-Pointer Catalytic efficiency improved when MPS was functionalized by n-decyl substitution. Black-Right-Pointing-Pointer Immobilized CE exhibited good thermal stability and reusability. Black-Right-Pointing-Pointer Organic co-solvent and the substrate structures affected enzyme activities.

  11. Functional tuning of the catalytic residue pKa in a de novo designed esterase.

    Science.gov (United States)

    Hiebler, Katharina; Lengyel, Zsófia; Castañeda, Carlos A; Makhlynets, Olga V

    2017-09-01

    AlleyCatE is a de novo designed esterase that can be allosterically regulated by calcium ions. This artificial enzyme has been shown to hydrolyze p-nitrophenyl acetate (pNPA) and 4-nitrophenyl-(2-phenyl)-propanoate (pNPP) with high catalytic efficiency. AlleyCatE was created by introducing a single-histidine residue (His 144 ) into a hydrophobic pocket of calmodulin. In this work, we explore the determinants of catalytic properties of AlleyCatE. We obtained the pK a value of the catalytic histidine using experimental measurements by NMR and pH rate profile and compared these values to those predicted from electrostatics pK a calculations (from both empirical and continuum electrostatics calculations). Surprisingly, the pK a value of the catalytic histidine inside the hydrophobic pocket of calmodulin is elevated as compared to the model compound pK a value of this residue in water. We determined that a short-range favorable interaction with Glu 127 contributes to the elevated pK a of His 144 . We have rationally modulated local electrostatic potential in AlleyCatE to decrease the pK a of its active nucleophile, His 144 , by 0.7 units. As a direct result of the decrease in the His 144 pK a value, catalytic efficiency of the enzyme increased by 45% at pH 6. This work shows that a series of simple NMR experiments that can be performed using low field spectrometers, combined with straightforward computational analysis, provide rapid and accurate guidance to rationally improve catalytic efficiency of histidine-promoted catalysis. Proteins 2017; 85:1656-1665. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  12. Vaginal Fornix Discharge Cellularity and Its Leukocyte Esterase Activity for Diagnosis of Endometritis in Dairy Cows

    Directory of Open Access Journals (Sweden)

    Abolfazl HAJIBEMANI

    2016-01-01

    Full Text Available The objective of the present study was to evaluate the application of some strip test markers (i.e., leukocyte esterase (LE activity, protein, nitrate and pH for diagnosis of endometritis in dairy cows using vaginal fornix discharge. Also, the total white blood cell count (t-WBC/l of this secretion and degenerative changes of neutrophils in cervical cytology were used as alternative methods to predict progression of the endometritis severity. Holstein cows (n=215 between 30-40 days in milk (DIM were included and examined. Giemsa-stained smear was prepared from cervical mucus. Cervical cytology test was considered as reference screening method for the detection of subclinical endometritis. The LE activity and t-WBC in the vaginal fornix discharge of subclinical endometritis cows were significantly higher than those from healthy cows. Sensitivity and specificity were 78% and 73% for LE10 activity (10 minutes after contacting with discharges and 60% and 69% for t-WBC (cut off point=210 cells/l for diagnosis of subclinical endometritis, respectively. There was a good agreement between LE10 activity, t-WBC and cervical cytology test with a Kappa coefficient of 0.4 and 0.42, respectively (P<0.0001. Total WBC count in discharge and degenerative neutrophils (DN percentages increase simultaneously with the degree and severity of endometritis. There was a highly significant (P<0.01 correlation between t-WBC and some reagent strip test markers (LE activity, protein and nitrate in clear discharge of studied cows. In conclusion, the present results suggest the LE activity and t-WBC in vaginal fornix discharge could be used as non-invasive reliable and valid methods for screening of subclinical endometritis in postpartum dairy herds.

  13. C1-esterase inhibitor protects against early vein graft remodeling under arterial blood pressure.

    Science.gov (United States)

    Krijnen, Paul A J; Kupreishvili, Koba; de Vries, Margreet R; Schepers, Abbey; Stooker, Wim; Vonk, Alexander B A; Eijsman, Leon; Van Hinsbergh, Victor W M; Zeerleder, Sacha; Wouters, Diana; van Ham, Marieke; Quax, Paul H A; Niessen, Hans W M

    2012-01-01

    Arterial pressure induced vein graft injury can result in endothelial loss, accelerated atherosclerosis and vein graft failure. Inflammation, including complement activation, is assumed to play a pivotal role herein. Here, we analyzed the effects of C1-esterase inhibitor (C1inh) on early vein graft remodeling. Human saphenous vein graft segments (n=8) were perfused in vitro with autologous blood either supplemented or not with purified human C1inh at arterial pressure for 6h. The vein segments and perfusion blood were analyzed for cell damage and complement activation. In addition, the effect of purified C1inh on vein graft remodeling was analyzed in vivo in atherosclerotic C57Bl6/ApoE3 Leiden mice, wherein donor caval veins were interpositioned in the common carotid artery. Application of C1inh in the in vitro perfusion model resulted in significantly higher blood levels and significantly more depositions of C1inh in the vein wall. This coincided with a significant reduction in endothelial loss and deposition of C3d and C4d in the vein wall, especially in the circular layer, compared to vein segments perfused without supplemented C1inh. Administration of purified C1inh significantly inhibited vein graft intimal thickening in vivo in atherosclerotic C57Bl6/ApoE3 Leiden mice, wherein donor caval veins were interpositioned in the common carotid artery. C1inh significantly protects against early vein graft remodeling, including loss of endothelium and intimal thickening. These data suggest that it may be worth considering its use in patients undergoing coronary artery bypass grafting. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  14. The dynamic organization of fungal acetyl-CoA carboxylase

    Science.gov (United States)

    Hunkeler, Moritz; Stuttfeld, Edward; Hagmann, Anna; Imseng, Stefan; Maier, Timm

    2016-04-01

    Acetyl-CoA carboxylases (ACCs) catalyse the committed step in fatty-acid biosynthesis: the ATP-dependent carboxylation of acetyl-CoA to malonyl-CoA. They are important regulatory hubs for metabolic control and relevant drug targets for the treatment of the metabolic syndrome and cancer. Eukaryotic ACCs are single-chain multienzymes characterized by a large, non-catalytic central domain (CD), whose role in ACC regulation remains poorly characterized. Here we report the crystal structure of the yeast ACC CD, revealing a unique four-domain organization. A regulatory loop, which is phosphorylated at the key functional phosphorylation site of fungal ACC, wedges into a crevice between two domains of CD. Combining the yeast CD structure with intermediate and low-resolution data of larger fragments up to intact ACCs provides a comprehensive characterization of the dynamic fungal ACC architecture. In contrast to related carboxylases, large-scale conformational changes are required for substrate turnover, and are mediated by the CD under phosphorylation control.

  15. Protein acetylation sites mediated by Schistosoma mansoni GCN5

    International Nuclear Information System (INIS)

    Moraes Maciel, Renata de; Furtado Madeiro da Costa, Rodrigo; Meirelles Bastosde Oliveira, Francisco; Rumjanek, Franklin David; Fantappie, Marcelo Rosado

    2008-01-01

    The transcriptional co-activator GCN5, a histone acetyltransferase (HAT), is part of large multimeric complexes that are required for chromatin remodeling and transcription activation. As in other eukaryotes, the DNA from the parasite Schistosome mansoni is organized into nucleosomes and the genome encodes components of chromatin-remodeling complexes. Using a series of synthetic peptides we determined that Lys-14 of histone H3 was acetylated by the recombinant SmGCN5-HAT domain. SmGCN5 was also able to acetylate schistosome non-histone proteins, such as the nuclear receptors SmRXR1 and SmNR1, and the co-activator SmNCoA-62. Electron microscopy revealed the presence of SmGCN5 protein in the nuclei of vitelline cells. Within the nucleus, SmGCN5 was found to be located in interchromatin granule clusters (IGCs), which are transcriptionally active structures. The data suggest that SmGCN5 is involved in transcription activation

  16. Identification and characterization of a novel cold-adapted esterase from a metagenomic library of mountain soil.

    Science.gov (United States)

    Ko, Kyong-Cheol; Rim, Soon-Ok; Han, Yunjon; Shin, Bong Seok; Kim, Geun-Joong; Choi, Jong Hyun; Song, Jae Jun

    2012-05-01

    A novel lipolytic enzyme was isolated from a metagenomic library after demonstration of lipolytic activity on an LB agar plate containing 1% (w/v) tributyrin. A novel esterase gene (estIM1), encoding a lipolytic enzyme (EstIM1), was cloned using a shotgun method from a pFosEstIM1 clone of the metagenomic library, and the enzyme was characterized. The estIM1 gene had an open reading frame (ORF) of 936 base pairs and encoded a protein of 311 amino acids with a molecular mass 34 kDa and a pI value of 4.32. The deduced amino acid sequence was 62% identical to that of an esterase from an uncultured bacterium (ABQ11271). The amino acid sequence indicated that EstIM1 was a member of the family IV of lipolytic enzymes, all of which contain a GDSAG motif shared with similar enzymes of lactic acid microorganisms. EstIM1 was active over a temperature range of 1-50°C, at alkaline pH. The activation energy for hydrolysis of p-nitrophenyl propionate was 1.04 kcal/mol, within a temperature range of 1-40°C. The activity of EstIM1 was about 60% of maximal even at 1°C, suggesting that EstIM1 is efficiently cold-adapted. Further characterization of this cold-adapted enzyme indicated that the esterase may be very valuable in industrial applications.

  17. A new cold-adapted, alkali-stable and highly salt-tolerant esterase from Bacillus licheniformis.

    Science.gov (United States)

    Zhang, Weijia; Xu, Hui; Wu, Yingqiang; Zeng, Jie; Guo, Ziwei; Wang, Lu; Shen, Cheng; Qiao, Dairong; Cao, Yi

    2018-05-01

    Bacterial esterases and lipases, especially extremozymes attract increasing attention due to various advantages both in good properties and wide applications. In the present study, a cold-adapted, alkali-stable and highly salt-tolerant esterase (Est700) was cloned from Bacillus licheniformis, expressed and purified with a molecular mass of 25 kDa. The optimal temperature of Est700 was 30 °C, with 35% maximal activity retaining at 0 °C. Its optimal pH was 8.0 and showed high stability at pH 5.0-11.0. Noticeably, Est700 was highly activated by 3.5 M NaCl and the extent of this activation is much stronger than that of currently reported halophilic ones. It was also stable in 5 M NaCl with no activity loss. High salt concentrations changed the secondary structure and folding properties of Est700 with formation of more α-helix and less β-sheet domains. With salt incubation, its melting temperature was estimated to be 57.2 °C, which is 12.8 °C higher than that of native one. Interestingly, Est700 lacks the acidic surface that is considered essential for enzyme stability at high salinity. However, it has a mainly positive surface electrostatic potential, which is probably different from most reported halotolerant esterases. These multiple properties make Est700 a valuable candidate in both basic research and industrial applications. Copyright © 2018. Published by Elsevier B.V.

  18. A novel esterase gene cloned from a metagenomic library from neritic sediments of the South China Sea

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    Peng Qing

    2011-11-01

    Full Text Available Abstract Background Marine microbes are a large and diverse group, which are exposed to a wide variety of pressure, temperature, salinity, nutrient availability and other environmental conditions. They provide a huge potential source of novel enzymes with unique properties that may be useful in industry and biotechnology. To explore the lipolytic genetic resources in the South China Sea, 23 sediment samples were collected in the depth Results A metagenomic library of South China Sea sediments assemblage in plasmid vector containing about 194 Mb of community DNA was prepared. Screening of a part of the unamplified library resulted in isolation of 15 unique lipolytic clones with the ability to hydrolyze tributyrin. A positive recombinant clone (pNLE1, containing a novel esterase (Est_p1, was successfully expressed in E. coli and purified. In a series of assays, Est_p1 displayed maximal activity at pH 8.57, 40°C, with ρ-Nitrophenyl butyrate (C4 as substrate. Compared to other metagenomic esterases, Est_p1 played a notable role in specificity for substrate C4 (kcat/Km value 11,500 S-1m M-1 and showed no inhibited by phenylmethylsulfonyl fluoride, suggested that the substrate binding pocket was suitable for substrate C4 and the serine active-site residue was buried at the bottom of substrate binding pocket which sheltered by a lid structure. Conclusions Esterase, which specificity towards short chain fatty acids, especially butanoic acid, is commercially available as potent flavoring tools. According the outstanding activity and specificity for substrate C4, Est_p1 has potential application in flavor industries requiring hydrolysis of short chain esters.

  19. Crystal structure of hyperthermophilic esterase EstE1 and the relationship between its dimerization and thermostability properties

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    Koh Eunhee

    2007-07-01

    Full Text Available Abstract Background EstE1 is a hyperthermophilic esterase belonging to the hormone-sensitive lipase family and was originally isolated by functional screening of a metagenomic library constructed from a thermal environmental sample. Dimers and oligomers may have been evolutionally selected in thermophiles because intersubunit interactions can confer thermostability on the proteins. The molecular mechanisms of thermostabilization of this extremely thermostable esterase are not well understood due to the lack of structural information. Results Here we report for the first time the 2.1-Å resolution crystal structure of EstE1. The three-dimensional structure of EstE1 exhibits a classic α/β hydrolase fold with a central parallel-stranded beta sheet surrounded by alpha helices on both sides. The residues Ser154, Asp251, and His281 form the catalytic triad motif commonly found in other α/β hydrolases. EstE1 exists as a dimer that is formed by hydrophobic interactions and salt bridges. Circular dichroism spectroscopy and heat inactivation kinetic analysis of EstE1 mutants, which were generated by structure-based site-directed mutagenesis of amino acid residues participating in EstE1 dimerization, revealed that hydrophobic interactions through Val274 and Phe276 on the β8 strand of each monomer play a major role in the dimerization of EstE1. In contrast, the intermolecular salt bridges contribute less significantly to the dimerization and thermostability of EstE1. Conclusion Our results suggest that intermolecular hydrophobic interactions are essential for the hyperthermostability of EstE1. The molecular mechanism that allows EstE1 to endure high temperature will provide guideline for rational design of a thermostable esterase/lipase using the lipolytic enzymes showing structural similarity to EstE1.

  20. Comparative Analysis of Proteome-Wide Lysine Acetylation in Juvenile and Adult Schistosoma japonicum

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    Qing Li

    2017-11-01

    Full Text Available Schistosomiasis is a devastating parasitic disease caused by tremotodes of the genus Schistosoma. Eggs produced by sexually mature schistosomes are the causative agents of for pathogenesis and transmission. Elucidating the molecular mechanism of schistosome development and sexual maturation would facilitate the prevention and control of schistosomiasis. Acetylation of lysine is a dynamic and reversible post-translational modification playing keys role in many biological processes including development in both eukaryotes and prokaryotes. To investigate the impacts of lysine acetylation on Schistosoma japonicum (S. japonicum development and sexual maturation, we used immunoaffinity-based acetyllysine peptide enrichment combined with mass spectrometry (MS, to perform the first comparative analysis of proteome-wide lysine acetylation in both female and male, juvenile (18 days post infection, 18 dpi and adult (28 dpi schistosome samples. In total, we identified 874 unique acetylated sites in 494 acetylated proteins. The four samples shared 47 acetylated sites and 46 proteins. More acetylated sites and proteins shared by both females and males were identified in 28 dpi adults (189 and 143, respectively than in 18 dpi schistosomula (76 and 59, respectively. More stage-unique acetylated sites and proteins were also identified in 28 dpi adults (494 and 210, respectively than in 18 dpi schistosomula (73 and 44, respectively. Functional annotation showed that in different developmental stages and genders, a number of proteins involving in muscle movement, glycometabolism, lipid metabolism, energy metabolism, environmental stress resistance, antioxidation, etc., displayed distinct acetylation profiles, which was in accordance with the changes of their biological functions during schistosome development, suggesting that lysine acetylation modification exerted important regulatory roles in schistosome development. Taken together, our data provided the first

  1. Estrogenic and esterase-inhibiting potency in rainwater in relation to pesticide concentrations, sampling season and location.

    Science.gov (United States)

    Hamers, Timo; van den Brink, Paul J; Mos, Lizzy; van der Linden, Sander C; Legler, Juliette; Koeman, Jan H; Murk, Albertinka J

    2003-01-01

    In a year-round monitoring program (1998), pesticide composition and toxic potency of the mix of pollutants present in rainwater were measured. The goal of the study was to relate atmospheric deposition of toxic potency and pesticide composition to each other and to sampling period and local agricultural activity. Rainwater was collected in 26 consecutive periods of 14 days in a background location (BACK) and in two locations representative for different agricultural practices, i.e. intensive greenhouse horticulture (HORT) and flower bulb culture (BULB). Samples were chemically analyzed for carbamate (CARB), organophosphate (OP) and organochlorine (OC) pesticides and metabolites. Esterase inhibiting potency of rainwater extracts was measured in a specially developed bio-assay with honeybee esterases and was expressed as an equivalent concentration of the model inhibitor dichlorvos. Estrogenic potency of the extracts was measured in the ER-CALUX reporter gene assay and was expressed as an equivalent concentration of estradiol. Multivariate principal component analysis (PCA) techniques proved to be valuable tools to analyze the numerous pesticide concentrations in relation to toxic potency, sampling location, and sampling season. Pesticide composition in rainwater depended much more on sampling season than on sampling location, but differences between and were mainly attributed to local differences in agricultural practice. On average, the esterase inhibiting potency exceeded the maximum permissible concentration set for dichlorvos in The Netherlands, and was significantly higher in than in and . Esterase inhibition correlated significantly with OP and CARB concentrations, as expected given the working mechanism of these insecticides. The estrogenic potency incidentally exceeded NOEC levels reported for aquatic organisms and was highest in . Although estrogenic potency of rainwater correlated with OC concentrations, the ER-CALUX responses could not be attributed to

  2. The Alpha-Defensin Immunoassay and Leukocyte Esterase Colorimetric Strip Test for the Diagnosis of Periprosthetic Infection

    Science.gov (United States)

    Wyatt, M.C.; Beswick, A.D.; Kunutsor, S.K.; Wilson, M.J.; Whitehouse, M.R.; Blom, A.W.

    2016-01-01

    Background: Synovial biomarkers have recently been adopted as diagnostic tools for periprosthetic joint infection (PJI), but their utility is uncertain. The purpose of this systematic review and meta-analysis was to synthesize the evidence on the accuracy of the alpha-defensin immunoassay and leukocyte esterase colorimetric strip test for the diagnosis of PJI compared with the Musculoskeletal Infection Society diagnostic criteria. Methods: We performed a systematic review to identify diagnostic technique studies evaluating the accuracy of alpha-defensin or leukocyte esterase in the diagnosis of PJI. MEDLINE and Embase on Ovid, ACM, ADS, arXiv, CERN DS (Conseil Européen pour la Recherche Nucléaire Document Server), CrossRef DOI (Digital Object Identifier), DBLP (Digital Bibliography & Library Project), Espacenet, Google Scholar, Gutenberg, HighWire, IEEE Xplore (Institute of Electrical and Electronics Engineers digital library), INSPIRE, JSTOR (Journal Storage), OAlster (Open Archives Initiative Protocol for Metadata Harvesting), Open Content, Pubget, PubMed, and Web of Science were searched for appropriate studies indexed from inception until May 30, 2015, along with unpublished or gray literature. The classification of studies and data extraction were performed independently by 2 reviewers. Data extraction permitted meta-analysis of sensitivity and specificity with construction of receiver operating characteristic curves for each test. Results: We included 11 eligible studies. The pooled diagnostic sensitivity and specificity of alpha-defensin (6 studies) for PJI were 1.00 (95% confidence interval [CI], 0.82 to 1.00) and 0.96 (95% CI, 0.89 to 0.99), respectively. The area under the curve (AUC) for alpha-defensin and PJI was 0.99 (95% CI, 0.98 to 1.00). The pooled diagnostic sensitivity and specificity of leukocyte esterase (5 studies) for PJI were 0.81 (95% CI, 0.49 to 0.95) and 0.97 (95% CI, 0.82 to 0.99), respectively. The AUC for leukocyte esterase and PJI

  3. Estrogenic and esterase-inhibiting potency in rainwater in relation to pesticide concentrations, sampling season and location

    International Nuclear Information System (INIS)

    Hamers, T.; Brink, P.J. van den; Mos, L.; Linden, S.C. van der; Legler, J.; Koeman, J.H.; Murk, A.J.

    2003-01-01

    Estrogenic potency of rainwater correlated well with organochlorine concentrations, but could not be attributed to specific pesticides. - In a year-round monitoring program (1998), pesticide composition and toxic potency of the mix of pollutants present in rainwater were measured. The goal of the study was to relate atmospheric deposition of toxic potency and pesticide composition to each other and to sampling period and local agricultural activity. Rainwater was collected in 26 consecutive periods of 14 days in a background location (BACK) and in two locations representative for different agricultural practices, i.e. intensive greenhouse horticulture (HORT) and flower bulb culture (BULB). Samples were chemically analyzed for carbamate (CARB), organophosphate (OP) and organochlorine (OC) pesticides and metabolites. Esterase inhibiting potency of rainwater extracts was measured in a specially developed bio-assay with honeybee esterases and was expressed as an equivalent concentration of the model inhibitor dichlorvos. Estrogenic potency of the extracts was measured in the ER-CALUX reporter gene assay and was expressed as an equivalent concentration of estradiol. Multivariate principal component analysis (PCA) techniques proved to be valuable tools to analyze the numerous pesticide concentrations in relation to toxic potency, sampling location, and sampling season. Pesticide composition in rainwater depended much more on sampling season than on sampling location, but differences between SPRING and SUMMER were mainly attributed to local differences in agricultural practice. On average, the esterase inhibiting potency exceeded the maximum permissible concentration set for dichlorvos in The Netherlands, and was significantly higher in HORT than in BACK and BULB. Esterase inhibition correlated significantly with OP and CARB concentrations, as expected given the working mechanism of these insecticides. The estrogenic potency incidentally exceeded NOEC levels reported for

  4. Relationship between esterase activity and acrinathrin and methiocarb resistance in field populations of western flower thrips, Frankliniella occidentalis.

    Science.gov (United States)

    Maymó, Ana C; Cervera, Amelia; Dolores Garcerá, M; Bielza, Pablo; Martínez-Pardo, Rafael

    2006-12-01

    The western flower thrips, Frankliniella occidentalis (Pergande), is a serious pest in the south-east of Spain owing to its direct feeding on crops, transmission of the tomato spotted wilt virus and its very high level of resistance to insecticides. Mechanisms of resistance were examined using field populations of F. occidentalis with different susceptibilities to acrinathrin, methiocarb (selective insecticides), endosulfan, metamidophos and deltamethrin (broad-spectrum insecticides). Esterase activity towards alpha-naphthyl acetate and p-nitrophenyl acetate in resistant strains was significantly higher than in the reference strain (MLFOM) for both model substrates. This higher activity was significantly correlated with acrinathrin and methiocarb resistance. Copyright 2006 Society of Chemical Industry.

  5. Comparative study of esterases in deltamethrin and diazinon resistant Rhipicephalus microplus and Hyalomma anatolicum ticks collected from the Trans-Gangetic plains of India.

    Science.gov (United States)

    Gaur, Ruchi Singh; Sangwan, Arun Kumar; Sangwan, Nirmal; Ghosh, Mayukh; Kumar, Sachin

    2017-09-01

    A comparative analysis of esterases in susceptible and resistant ticks revealed six types of esterases (EST-1b, EST-2b, EST-3b, EST-4b, EST-5b and EST-6b) in Rhipicephalus microplus and four types (EST-1h, EST-2h, EST-3h, EST-4h) in Hyalomma anatolicum using α-naphthyl acetate substrate. Inhibition studies with eserine sulfate, p-chloromercuribenzoate, copper sulphate and phenylmethylsulfonyl fluoride revealed a marked variation in band intensity between susceptible and resistant ticks, with the latter being more intense. Qualitative expression of EST-4b along with an extra band of EST-5b and EST-6b were indicative of deltamethrin and diazinon resistance in R. microplus, whereas qualitative expression of EST-4h was probably responsible for diazinon resistance in H. anatolicum. The data suggest that increased esterase activity may represent a detoxification strategy leading to the development of resistance in these tick populations.

  6. Esterases no exame da estrutura populacional de Camu-camu (Myrciaria dubia (Kunth McVaugh-Myrtaceae Esterases for examining the population structure of Camu-camu (Myrciaria dubia (Kunth McVaugh-Myrtaceae

    Directory of Open Access Journals (Sweden)

    Aylton Saturnino Teixeira

    2004-01-01

    Full Text Available Dois sistemas enzimáticos (esterase e esterase-D, analisados pela técnica de eletroforese em gel de amido, em folhas jovens de plantas cultivadas em terra firme, de sementes provenientes de três amostras de populações naturais de camu-camu, Myrciaria dubia (Kunth McVaugh-Myrtaceae, procedentes de Iquitos, Boa Vista e Uatumã, revelaram a presença de 6 locos: Est-1, Est-2, Est-3, Est-4, Est-D1 e Est-D2. Dois dos seis locos gênicos examinados no presente estudo (Est-3 e Est-D2 mostraram-se polimórficos, sendo desse modo considerados valiosos no estudo de caracterização da estrutura populacional da espécie. Os padrões de polimorfismo revelados nos locos Est-3 e Est-D2 de camu-camu, são típicos de enzimas monoméricas e diméricas, respectivamente. O loco Est-3 apresentou um grande desbalanço genético dentro e entre as amostras populacionais examinadas, devido ao excessivo número observado de plantas heterozigóticas em relação ao número esperado. O loco Est-D2 apresentou um polimorfismo exclusivo para os alelos Est-D2¹,Est-D2² e Est-D2³, e um bom balanço genético na amostra populacional de Uatumã. Em função disso, dentre os demais locos gênicos aqui investigados, o loco Est-D2 parece ser o mais adequado para identificação e delimitação de prováveis estoques de camu-camu. Portanto, recomenda-se que esse loco esteja presente na lista dos marcadores isoenzimáticos a serem usados em futuras prospecções sobre genética populacional dessa espécie na região amazônica. Dados sobre a distribuição das freqüências alélicas, estimativas das distâncias genéticas, e estimativas de variação genética nos 6 locos de esterases examinados, foram eficazes na demonstração de diferenças genéticas entre as amostras populacionais examinadas da espécie. Os maiores valores de heterozigozidade média (0,1353; proporção de locos polimórficos (0,33 e número médio de alelos por loco (1,33 revelados na amostra

  7. The Use of Plasma-Derived Complement C1-Esterase Inhibitor Concentrate (Berinert®) in the Treatment of Angiotensin Converting Enzyme-Inhibitor Related Angioedema

    DEFF Research Database (Denmark)

    Hermanrud, Thorbjørn; Duus, Nicolaj; Bygum, Anette

    2016-01-01

    Angioedema of the upper airways is a severe and potentially life-threatening condition. The incidence has been increasing in the past two decades, primarily due to pharmaceuticals influencing the generation or degradation of the vasoactive molecule bradykinin. Plasma-derived C1-esterase inhibitor...... concentrate is a well-established treatment option of hereditary and acquired complement C1-esterase inhibitor deficiency, which are also mediated by an increased level of bradykinin resulting in recurrent angioedema. We here present a case of severe angiotensin converting enzyme-inhibitor related angioedema...

  8. Prevalence of positive urinary dipstick analysis (leucocyte esterase, nitrite, haemoglobin, or glucose) in a population of 3645 adult subjects--consequence for measurement of urinary albumin excretion rate

    DEFF Research Database (Denmark)

    Clausen, P; Jensen, J S; Borch-Johnsen, K

    1998-01-01

    OBJECTIVES: To assess prevalence of positive urinary dipstick analysis for leucocyte esterase, nitrite, haemoglobin, or glucose in the general population and measure the urinary albumin excretion rate (UAER) in subjects with or without a positive dipstick analysis. DESIGN: A cross-sectional study...... of 3645 subjects. SETTING: An unselected urban population study. MAIN OUTCOME MEASURES: Prevalence data of positive dipstick analyses and UAER values. RESULTS: Prevalence data of a positive dipstick analysis were 12%, 4%, 3% and 6%, respectively, for leucocyte esterase, nitrite, haemoglobin, and glucose...

  9. Prediction of Nepsilon-acetylation on internal lysines implemented in Bayesian Discriminant Method.

    Science.gov (United States)

    Li, Ao; Xue, Yu; Jin, Changjiang; Wang, Minghui; Yao, Xuebiao

    2006-12-01

    Protein acetylation is an important and reversible post-translational modification (PTM), and it governs a variety of cellular dynamics and plasticity. Experimental identification of acetylation sites is labor-intensive and often limited by the availability of reagents such as acetyl-specific antibodies and optimization of enzymatic reactions. Computational analyses may facilitate the identification of potential acetylation sites and provide insights into further experimentation. In this manuscript, we present a novel protein acetylation prediction program named PAIL, prediction of acetylation on internal lysines, implemented in a BDM (Bayesian Discriminant Method) algorithm. The accuracies of PAIL are 85.13%, 87.97%, and 89.21% at low, medium, and high thresholds, respectively. Both Jack-Knife validation and n-fold cross-validation have been performed to show that PAIL is accurate and robust. Taken together, we propose that PAIL is a novel predictor for identification of protein acetylation sites and may serve as an important tool to study the function of protein acetylation. PAIL has been implemented in PHP and is freely available on a web server at: http://bioinformatics.lcd-ustc.org/pail.

  10. Prediction of Nε-acetylation on internal lysines implemented in Bayesian Discriminant Method

    Science.gov (United States)

    Li, Ao; Xue, Yu; Jin, Changjiang; Wang, Minghui; Yao, Xuebiao

    2007-01-01

    Protein acetylation is an important and reversible post-translational modification (PTM), and it governs a variety of cellular dynamics and plasticity. Experimental identification of acetylation sites is labor-intensive and often limited by the availability reagents such as acetyl-specific antibodies and optimization of enzymatic reactions. Computational analyses may facilitate the identification of potential acetylation sites and provide insights into further experimentation. In this manuscript, we present a novel protein acetylation prediction program named PAIL, prediction of acetylation on internal lysines, implemented in a BDM (Bayesian Discriminant Method) algorithm. The accuracies of PAIL are 85.13%, 87.97% and 89.21% at low, medium and high thresholds, respectively. Both Jack-Knife validation and n-fold cross validation have been performed to show that PAIL is accurate and robust. Taken together, we propose that PAIL is a novel predictor for identification of protein acetylation sites and may serve as an important tool to study the function of protein acetylation. PAIL has been implemented in PHP and is freely available on a web server at: http://bioinformatics.lcd-ustc.org/pail. PMID:17045240

  11. The Effect of Hypochlorite Oxidation and Acetylation on Some of the ...

    African Journals Online (AJOL)

    The study evaluated the effect of hypochlorite oxidation and acetylation on some physicochemical properties of Icacina trichantha starch. The native and modified (oxidized and acetylated) starches were studied with respect to Infrared spectroscopy(IR), microscopy, gelatinization, swelling power, solubility index, amylose ...

  12. Coactivator-dependent acetylation stabilizes members of the SREBP family of transcription factors.

    Science.gov (United States)

    Giandomenico, Valeria; Simonsson, Maria; Grönroos, Eva; Ericsson, Johan

    2003-04-01

    Members of the SREBP family of transcription factors control cholesterol and lipid homeostasis and play important roles during adipocyte differentiation. The transcriptional activity of SREBPs is dependent on the coactivators p300 and CBP. We now present evidence that SREBPs are acetylated by the intrinsic acetyltransferase activity of p300 and CBP. In SREBP1a, the acetylated lysine residue resides in the DNA-binding domain of the protein. Coexpression with p300 dramatically increases the expression of both SREBP1a and SREBP2, and this effect is dependent on the acetyltransferase activity of p300, indicating that acetylation of SREBPs regulates their stability. Indeed, acetylation or mutation of the acetylated lysine residue in SREBP1a stabilizes the protein. We demonstrate that the acetylated residue in SREBP1a is also targeted by ubiquitination and that acetylation inhibits this process. Thus, our studies define acetylation-dependent stabilization of transcription factors as a novel mechanism for coactivators to regulate gene expression.

  13. Protective Effects of Acetylation on the Pathological Reactions of the Lens Crystallins with Homocysteine Thiolactone.

    Directory of Open Access Journals (Sweden)

    Zeinab Moafian

    Full Text Available Various post-translational lens crystallins modifications result in structural and functional insults, contributing to the development of lens opacity and cataract disorders. Lens crystallins are potential targets of homocysteinylation, particularly under hyperhomocysteinemia which has been indicated in various eye diseases. Since both homocysteinylation and acetylation primarily occur on protein free amino groups, we applied different spectroscopic methods and gel mobility shift analysis to examine the possible preventive role of acetylation against homocysteinylation. Lens crystallins were extensively acetylated in the presence of acetic anhydride and then subjected to homocysteinylation in the presence of homocysteine thiolactone (HCTL. Extensive acetylation of the lens crystallins results in partial structural alteration and enhancement of their stability, as well as improvement of α-crystallin chaperone-like activity. In addition, acetylation partially prevents HCTL-induced structural alteration and aggregation of lens crystallins. Also, acetylation protects against HCTL-induced loss of α-crystallin chaperone activity. Additionally, subsequent acetylation and homocysteinylation cause significant proteolytic degradation of crystallins. Therefore, further experimentation is required in order to judge effectively the preventative role of acetylation on the structural and functional insults induced by homocysteinylation of lens crystallins.

  14. Requirements for Carnitine Shuttle-Mediated Translocation of Mitochondrial Acetyl Moieties to the Yeast Cytosol

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    Harmen M. van Rossum

    2016-05-01

    Full Text Available In many eukaryotes, the carnitine shuttle plays a key role in intracellular transport of acyl moieties. Fatty acid-grown Saccharomyces cerevisiae cells employ this shuttle to translocate acetyl units into their mitochondria. Mechanistically, the carnitine shuttle should be reversible, but previous studies indicate that carnitine shuttle-mediated export of mitochondrial acetyl units to the yeast cytosol does not occur in vivo. This apparent unidirectionality was investigated by constitutively expressing genes encoding carnitine shuttle-related proteins in an engineered S. cerevisiae strain, in which cytosolic acetyl coenzyme A (acetyl-CoA synthesis could be switched off by omitting lipoic acid from growth media. Laboratory evolution of this strain yielded mutants whose growth on glucose, in the absence of lipoic acid, was l-carnitine dependent, indicating that in vivo export of mitochondrial acetyl units to the cytosol occurred via the carnitine shuttle. The mitochondrial pyruvate dehydrogenase complex was identified as the predominant source of acetyl-CoA in the evolved strains. Whole-genome sequencing revealed mutations in genes involved in mitochondrial fatty acid synthesis (MCT1, nuclear-mitochondrial communication (RTG2, and encoding a carnitine acetyltransferase (YAT2. Introduction of these mutations into the nonevolved parental strain enabled l-carnitine-dependent growth on glucose. This study indicates intramitochondrial acetyl-CoA concentration and constitutive expression of carnitine shuttle genes as key factors in enabling in vivo export of mitochondrial acetyl units via the carnitine shuttle.

  15. Novel myelin penta- and hexa-acetyl-galactosyl-ceramides: structural characterization and immunoreactivity in cerebrospinal fluid

    DEFF Research Database (Denmark)

    Podbielska, Maria; Dasgupta, Somsankar; Levery, Steven B

    2010-01-01

    -acetylation of the 2-hydroxy-fatty acid. The immuno-reactivity in human cerebrospinal fluid (CSF) to these acetylated glycolipids was examined in central nervous system (CNS) infectious disease, noninflammatory disorders, and multiple sclerosis (MS). Screening for lipid binding in MS and other neurological disease...

  16. Total levels of hippocampal histone acetylation predict normal variability in mouse behavior.

    Directory of Open Access Journals (Sweden)

    Addie May I Nesbitt

    Full Text Available Genetic, pharmacological, and environmental interventions that alter total levels of histone acetylation in specific brain regions can modulate behaviors and treatment responses. Efforts have been made to identify specific genes that are affected by alterations in total histone acetylation and to propose that such gene specific modulation could explain the effects of total histone acetylation levels on behavior - the implication being that under naturalistic conditions variability in histone acetylation occurs primarily around the promoters of specific genes.Here we challenge this hypothesis by demonstrating with a novel flow cytometry based technique that normal variability in open field exploration, a hippocampus-related behavior, was associated with total levels of histone acetylation in the hippocampus but not in other brain regions.Results suggest that modulation of total levels of histone acetylation may play a role in regulating biological processes. We speculate in the discussion that endogenous regulation of total levels of histone acetylation may be a mechanism through which organisms regulate cellular plasticity. Flow cytometry provides a useful approach to measure total levels of histone acetylation at the single cell level. Relating such information to behavioral measures and treatment responses could inform drug delivery strategies to target histone deacetylase inhibitors and other chromatin modulators to places where they may be of benefit while avoiding areas where correction is not needed and could be harmful.

  17. Global profiling of lysine acetylation in human histoplasmosis pathogen Histoplasma capsulatum.

    Science.gov (United States)

    Xie, Longxiang; Fang, Wenjie; Deng, Wanyan; Yu, Zhaoxiao; Li, Juan; Chen, Min; Liao, Wanqing; Xie, Jianping; Pan, Weihua

    2016-04-01

    Histoplasma capsulatum is the causative agent of human histoplasmosis, which can cause respiratory and systemic mycosis in immune-compromised individuals. Lysine acetylation, a protein posttranslational protein modification, is widespread in both eukaryotes and prokaryotes. Although increasing evidence suggests that lysine acetylation may play critical roles in fungus physiology, very little is known about its extent and function in H. capsulatum. To comprehensively profile protein lysine acetylation in H. capsulatum, we performed a global acetylome analysis through peptide prefractionation, antibody enrichment, and LC-MS/MS analysis, identifying 775 acetylation sites on 456 acetylated proteins; and functionally analysis showing their involvement in different biological processes. We defined six types of acetylation site motifs, and the results imply that lysine residue of polypeptide with tyrosine at the -1 and +1 positions, histidine at the +1 position, and phenylalanine (F) at the +1 and +2 position is a preferred substrate of lysine acetyltransferase. Moreover, some virulence factors candidates including calmodulin and DnaK are acetylated. In conclusion, our data set may serve as an important resource for the elucidation of associations between functional protein lysine acetylation and virulence in H. capsulatum. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Acetylome analysis reveals the involvement of lysine acetylation in biosynthesis of antibiotics in Bacillus amyloliquefaciens.

    Science.gov (United States)

    Liu, Lin; Wang, Guangyuan; Song, Limin; Lv, Binna; Liang, Wenxing

    2016-01-29

    Lysine acetylation is a major post-translational modification that plays an important regulatory role in almost every aspects in both eukaryotes and prokaryotes. Bacillus amyloliquefaciens, a Gram-positive bacterium, is very effective for the control of plant pathogens. However, very little is known about the function of lysine acetylation in this organism. Here, we conducted the first lysine acetylome in B. amyloliquefaciens through a combination of highly sensitive immune-affinity purification and high-resolution LC-MS/MS. Overall, we identified 3268 lysine acetylation sites in 1254 proteins, which account for 32.9% of the total proteins in this bacterium. Till date, this is the highest ratio of acetylated proteins that have been identified in bacteria. Acetylated proteins are associated with a variety of biological processes and a large fraction of these proteins are involved in metabolism. Interestingly, for the first time, we found that about 71.1% (27/38) and 78.6% (22/28) of all the proteins tightly related to the synthesis of three types of pepketides and five families of lipopeptides were acetylated, respectively. These findings suggest that lysine acetylation plays a critical role in the regulation of antibiotics biosynthesis. These data serves as an important resource for further elucidation of the physiological role of lysine acetylation in B. amyloliquefaciens.

  19. Histone Acetylation is Recruited in Consolidation as a Molecular Feature of Stronger Memories

    Science.gov (United States)

    Federman, Noel; Fustinana, Maria Sol; Romano, Arturo

    2009-01-01

    Gene expression is a key process for memory consolidation. Recently, the participation of epigenetic mechanisms like histone acetylation was evidenced in long-term memories. However, until now the training strength required and the persistence of the chromatin acetylation recruited are not well characterized. Here we studied whether histone…

  20. Discovery and characterization of sialic acid O-acetylation in group B Streptococcus.

    Science.gov (United States)

    Lewis, Amanda L; Nizet, Victor; Varki, Ajit

    2004-07-27

    Group B Streptococcus (GBS) is the leading cause of human neonatal sepsis and meningitis. The GBS capsular polysaccharide is a major virulence factor and the active principle of vaccines in phase II trials. All GBS capsules have a terminal alpha 2-3-linked sialic acid [N-acetylneuraminic acid (Neu5Ac)], which interferes with complement-mediated killing. We show here that some of the Neu5Ac residues of the GBS type III capsule are O-acetylated at carbon position 7, 8, or 9, a major modification evidently missed in previous studies. Data are consistent with initial O-acetylation at position 7, and subsequent migration of the O-acetyl ester at positions 8 and 9. O-acetylation was also present on several other GBS serotypes (Ia, Ib, II, V, and VI). Deletion of the CMP-Neu5Ac synthase gene neuA by precise, in-frame allelic replacement gave intracellular accumulation of O-acetylated Neu5Ac, whereas overexpression markedly decreased O-acetylation. Given the known GBS Neu5Ac biosynthesis pathway, these data indicate that O-acetylation occurs on free Neu5Ac, competing with the CMP-Neu5Ac synthase. O-acetylation often generates immunogenic epitopes on bacterial capsular polysaccharides and can modulate human alternate pathway complement activation. Thus, our discovery has important implications for GBS pathogenicity, immunogenicity, and vaccine design.