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Sample records for carboxyl-terminal esterase l1

  1. Conserved expression of ubiquitin carboxyl-terminal esterase L1 (UCHL1) in mammalian testes.

    Science.gov (United States)

    Devi, Lalitha; Pawar, Rahul Mohanchandra; Makala, Himesh; Goel, Sandeep

    2015-05-01

    Spermatogonia, the adult germ cells that initiate spermatogenesis in mammalian testis, are capable of dividing both mitotically and meiotically. Isolation and preservation of spermatogonia helps in preserving genetic pool of endangered animals. In this context, identification of marker(s) that can distinguish spermatogonia from other cells in testis gains significance. Here, we examined the expression of ubiquitin carboxyl-terminal esterase L1 (UCHL1) gene and protein in the testes of several mammals, including highly endangered species. Semi-quantitative-reverse transcriptase-polymerase chain reaction (RT-PCR) analysis showed presence of UCHL1 amplicon of 442 bp in all the 18 mammals studied. Nucleotide sequence analysis of these amplicons and their predicted protein sequences revealed 88-99% and 95-100% homology with available human UCHL1 and UCHL1 sequences of other available species in the GenBank, respectively. Western blot analysis showed that UCHL1 protein size was unique in all wild mammals. Immunohistology results confirmed UCHL1 expression in the spermatogonia/gonocytes in testes of several mammals belonging to eight distinct families including highly endangered Felidae, Canidae and Cercopithecoidae. These findings suggest that UCHL1 expression is conserved in the mammalian testis, and could be used as a specific marker for gonocytes/spermatogonia for developing male germ-cell based conservation techniques.

  2. Ubiquitin carboxyl terminal hydrolase L1 negatively regulates TNF{alpha}-mediated vascular smooth muscle cell proliferation via suppressing ERK activation

    Energy Technology Data Exchange (ETDEWEB)

    Ichikawa, Tomonaga; Li, Jinqing; Dong, Xiaoyu; Potts, Jay D. [Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States); Tang, Dong-Qi [Department of Pathology, Immunology, and Laboratory Medicine, University of Florida College of Medicine, Gainesville, FL 32610-0275 (United States); Li, Dong-Sheng, E-mail: dsli@yymc.edu.cn [Hubei Key Laboratory of Embryonic Stem Cell Research, Tai He Hospital, Yunyang Medical College, 32 S. Renmin Rd., Shiyan, Hubei 442000 (China); Cui, Taixing, E-mail: taixing.cui@uscmed.sc.edu [Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States)

    2010-01-01

    Deubiquitinating enzymes (DUBs) appear to be critical regulators of a multitude of processes such as proliferation, apoptosis, differentiation, and inflammation. We have recently demonstrated that a DUB of ubiquitin carboxyl terminal hydrolase L1 (UCH-L1) inhibits vascular lesion formation via suppressing inflammatory responses in vasculature. However, the precise underlying mechanism remains to be defined. Herein, we report that a posttranscriptional up-regulation of UCH-L1 provides a negative feedback to tumor necrosis factor alpha (TNF{alpha})-mediated activation of extracellular signal-regulated kinases (ERK) and proliferation in vascular smooth muscle cells (VSMCs). In rat adult VSMCs, adenoviral over-expression of UCH-L1 inhibited TNF{alpha}-induced activation of ERK and DNA synthesis. In contrast, over-expression of UCH-L1 did not affect platelet derived growth factor (PDGF)-induced VSMC proliferation and activation of growth stimulating cascades including ERK. TNF{alpha} hardly altered UCH-L1 mRNA expression and stability; however, up-regulated UCH-L1 protein expression via increasing UCH-L1 translation. These results uncover a novel mechanism by which UCH-L1 suppresses vascular inflammation.

  3. Downregulation of miR-181b in mouse brain following ischemic stroke induces neuroprotection against ischemic injury through targeting heat shock protein A5 and ubiquitin carboxyl-terminal hydrolase isozyme L1.

    Science.gov (United States)

    Peng, Zhifeng; Li, Jiefei; Li, Yun; Yang, Xuan; Feng, Sujuan; Han, Song; Li, Junfa

    2013-10-01

    Understanding the molecular mechanism of cerebral hypoxic preconditioning (HPC)-induced endogenous neuroprotection may provide potential therapeutic targets for ischemic stroke. By using bioinformatics analysis, we found that miR-181b, one of 19 differentially expressed miRNAs, may target aconitate hydratase (ACO2), heat shock protein A5 (HSPA5), and ubiquitin carboxyl-terminal hydrolase isozyme L1 (UCHL1) among 26 changed protein kinase C isoform-specific interacting proteins in HPC mouse brain. In this study, the role of miR-181b in oxygen-glucose deprivation (OGD)-induced N2A cell ischemic injury in vitro and mouse middle cerebral artery occlusion (MCAO)-induced cerebral ischemic injury in vivo, and its regulation of ACO2, HSPA5, and UCHL1 were further determined. We found that miR-181b expression levels significantly decreased in mouse brain following MCAO and in OGD-treated N2A cells. Up- and downregulation of miR-181b by transfection of pre- or anti-miR-181b could negatively regulate HSPA5 and UCHL1 (but not ACO2) protein levels as well as N2A cell death and programmed cell death in OGD-treated N2A cells. By using a T7 promoter-driven control dual luciferase assay, we confirmed that miR-181b could bind to the 3'-untranslated rergions of HSPA5 and UCHL1 mRNAs and repress their translations. miR-181b antagomir reduced caspase-3 cleavage and neural cell loss in cerebral ischemic cortex and improved neurological deficit of mice after MCAO. In addition, HSPA5 and UCHL1 short interfering RNAs (siRNAs) blocked anti-miR-181b-mediated neuroprotection against OGD-induced N2A cell injury in vitro. These results suggest that the downregulated miR-181b induces neuroprotection against ischemic injury through negatively regulating HSPA5 and UCHL1 protein levels, providing a potential therapeutic target for ischemic stroke.

  4. EPOXY RESINS TOUGHENED WITH CARBOXYL TERMINATED POLYETHERS

    Institute of Scientific and Technical Information of China (English)

    YU Yunchao; LI Yiming

    1983-01-01

    Carboxyl terminated polyethers, the adducts of hydroxyl terminated polytetrahydrofuran and maleic anhydride, were used as toughener for epoxy resins. The morphology of the toughened resins was investigated by means of turbidity measurement, dynamic mechanical testing and scanning electron microscope observation. It turned out that the molecular weight and the carboxyl content of the polyether and the cure conditions are important factors, which affect the particle size of the polyether-rich domains and, in turn, the mechanical properties of the cured resin. Carboxyl terminated polytetrahydrofurans have a low glass transition temperature, and in appropriate amount they do not affect the thermal resistance of the resin. These advantages make them preferable as toughener for epoxy resins.

  5. Affinity chromatography using protein immobilized via arginine residues: purification of ubiquitin carboxyl-terminal hydrolases.

    Science.gov (United States)

    Duerksen-Hughes, P J; Williamson, M M; Wilkinson, K D

    1989-10-17

    4-(Oxoacetyl)phenoxyacetic acid (OAPA) forms a stable, covalent bond between its glyoxal group and the guanidino group of arginine and arginine derivatives [Duerksen, P. J., & Wilkinson, K. D. (1987) Anal. Biochem. 160, 444-454]. Studies were carried out to determine the chemical nature of this linkage, and the structure of the stable adduct between OAPA and methylguanidine was elucidated. The stable product results from an internal oxidation-reduction of the Schiff base adduct to form a cyclic alpha-aminoamide, 4-[4-(carboxymethoxy)phenyl]-2-(methylimino)-5-oxoimidazolidine. OAPA coupled to polyacrylamide beads was used to immobilize ubiquitin via its arginine residues, and the resulting affinity support was shown to specifically and reversibly bind a previously described enzyme, ubiquitin carboxyl-terminal hydrolase [Pickart, C. M., & Rose, I. A. (1985) J. Biol. Chem. 260, 7903-7910]. The resin was then used to isolate three newly identified ubiquitin carboxyl-terminal hydrolytic activities, which did not bind to ubiquitin immobilized via lysine residues. Significant purification was achieved in each case, and one isozyme was further purified to homogeneity.

  6. Carboxyl-terminal multi-site phosphorylation regulates internalization and desensitization of the human sst2 somatostatin receptor.

    Science.gov (United States)

    Lehmann, Andreas; Kliewer, Andrea; Schütz, Dagmar; Nagel, Falko; Stumm, Ralf; Schulz, Stefan

    2014-04-25

    The somatostatin receptor 2 (sst2) is the pharmacological target of somatostatin analogs that are widely used in the diagnosis and treatment of human neuroendocrine tumors. We have recently shown that the stable somatostatin analogs octreotide and pasireotide (SOM230) stimulate distinct patterns of sst2 receptor phosphorylation and internalization. Like somatostatin, octreotide promotes the phosphorylation of at least six carboxyl-terminal serine and threonine residues namely S341, S343, T353, T354, T356 and T359, which in turn leads to a robust receptor endocytosis. Unlike somatostatin, pasireotide stimulates a selective phosphorylation of S341 and S343 of the human sst2 receptor followed by a partial receptor internalization. Here, we show that exchange of S341 and S343 by alanine is sufficient to block pasireotide-driven internalization, whereas mutation of T353, T354, T356 and T359 to alanine is required to strongly inhibited both octreotide- and somatostatin-induced internalization. Yet, combined mutation of T353, T354, T356 and T359 is not sufficient to prevent somatostatin-driven β-arrestin mobilization and receptor desensitization. Replacement of all fourteen carboxyl-terminal serine and threonine residues by alanine completely abrogates sst2 receptor internalization and β-arrestin mobilization in HEK293 cells. Together, our findings demonstrate for the first time that agonist-selective sst2 receptor internalization is regulated by multi-site phosphorylation of its carboxyl-terminal tail.

  7. A sensitive chemiluminescent enzyme immunoassay for the bioanalysis of carboxyl-terminal B-chain analogues of human insulin.

    Science.gov (United States)

    Cao, Y; Smith, W C; Bowsher, R R

    2001-08-01

    Quantification of analogues of human insulin in biological matrices is complicated by differences in their immunoreactivity and the presence of both the analogue and endogenous concentrations of insulin in test samples. To facilitate pharmacokinetic comparisons of carboxyl-terminal B-chain analogues of human insulin, we undertook development of a sensitive ELISA. The ELISA detection method was optimized systematically to permit routine analysis of 10-microl serum samples. Accordingly, a noncompetitive 'sandwich' chemiluminescent ELISA was validated for the quantification of carboxyl-terminal B-chain insulin analogues in human serum over a concentration range from 5 to 3125 pM. The mean bias (RE%) within the validated range varied from -10.3 to 4.3%, with an intermediate precision (inter-assay CV%) from 4.2 to 11.5%. The two-sided 90% expectation tolerance interval for total measurement error was within +/-25% of the nominal concentration for all levels of validation samples. Insulin lispro, human insulin, proinsulin, despentapeptide insulin (DPI) and porcine insulin displayed comparable crossreactivity in the ELISA. Potential utility of the new assay for insulin bioanalysis in nonhuman species was investigated by assessing the pharmacokinetic profile of DPI in rats following administration of a single subcutaneous dose. The sensitive chemiluminescent detection method is simple to perform and should be readily adaptable for ELISAs of other therapeutic proteins.

  8. A novel ubiquitin carboxyl terminal hydrolase is involved in toad oocyte maturation

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    p28, a 28kD protein from toad (Bufo bufo gargarizans) oocytes, was identified by using p13suc1-agaroseaffinity chromatography. Sequence homology analysis of the full-length cDNA of p28 (Gene Bank accessionnumber: AF 314091) indicated that it encodes a protein containing 224 amino-acids with about 55% iden-tities and more than 70% positives to human, rat or mouse UCH-L1, and contains homological functionaldomains of UCH family. Anti-p28 monoclonal antibody, on injecting into the oocytes, could inhibit theprogesterone-induced resumption of meiotic division in a dose-dependent manner. The recombinant proteinp28 showed similar SDS/PAGE behaviors to the native one, and promoted ubiquitin ethyl ester hydrolysis,a classical catalytic reaction for ubiquitin carboxyl terminai hydrolases (UCHs). The results in this paperreveal that a novel protein, p28, exists in the toad oocytes, is a UCH L1 homolog, was engaged in theprocess of progesterone-induced oocyte maturation possibly through an involvement in protein turnover anddegradation.

  9. A novel carboxyl-terminal protease derived from Paenibacillus lautus CHN26 exhibiting high activities at multiple sites of substrates

    DEFF Research Database (Denmark)

    Li, Yunxia; Pan, Yingjie; She, Qunxin;

    2013-01-01

    Carboxyl-terminal protease (CtpA) plays essential functions in posttranslational protein processing in prokaryotic and eukaryotic cells. To date, only a few bacterial ctpA genes have been characterized. Here we cloned and characterized a novel CtpA. The encoding gene, ctpAp (ctpA of Paenibacillus...

  10. An actin monomer binding activity localizes to the carboxyl-terminal half of the Saccharomyces cerevisiae cyclase-associated protein.

    Science.gov (United States)

    Freeman, N L; Chen, Z; Horenstein, J; Weber, A; Field, J

    1995-03-10

    The Saccharomyces cerevisiae adenylyl cyclase complex contains at least two subunits, a 200-kDa catalytic subunit and a 70-kDa cyclase-associated protein, CAP (also called Srv2p). Genetic studies suggested two roles for CAP, one as a positive regulator of cAMP levels in yeast and a second role as a cytoskeletal regulator. We present evidence showing that CAP sequesters monomeric actin (Kd in the range of 0.5-5 microM), decreasing actin incorporation into actin filaments. Anti-CAP monoclonal antibodies co-immunoprecipitate a protein with a molecular size of about 46 kDa. When CAP was purified from yeast using an anti-CAP monoclonal antibody column, the 46-kDa protein co-purified with a stoichiometry of about 1:1 with CAP. Western blots identified the 46-kDa protein as yeast actin. CAP also bound to muscle actin in vitro in immunoprecipitation assays and falling ball viscometry assays. Experiments with pyrene-labeled actin demonstrated that CAP sequesters actin monomers. The actin monomer binding activity is localized to the carboxyl-terminal half of CAP. Together, these data suggest that yeast CAP regulates the yeast cytoskeleton by sequestering actin monomers.

  11. Carboxyl-terminated butadiene-acrylonitrile-toughened epoxy/carboxyl-modified carbon nanotube nanocomposites: Thermal and mechanical properties

    Directory of Open Access Journals (Sweden)

    H. F. Xie

    2012-09-01

    Full Text Available Carboxyl-modified multi-walled carbon nanotubes (MWCNT–COOHs as nanofillers were incorporated into diglycidyl ether of bisphenol A (DGEBA toughened with carboxyl-terminated butadiene-acrylonitrile (CTBN. The carboxyl functional carbon nanotubes were characterized by Fourier-transform infrared spectroscopy and thermogravimetric analysis. Furthermore, cure kinetics, glass transition temperature (Tg, mechanical properties, thermal stability and morphology of DGEBA/CTBN/MWCNT–COOHs nanocomposites were investigated by differential scanning calorimetry (DSC, dynamic mechanical analysis (DMA, universal test machine, thermogravimetric analysis and scanning electron microscopy (SEM. DSC kinetic studies showed that the addition of MWCNT–COOHs accelerated the curing reaction of the rubber-toughened epoxy resin. DMA results revealed that Tg of rubber-toughened epoxy nanocomposites lowered with MWCNT–COOH contents. The tensile strength, elongation at break, flexural strength and flexural modulus of DGEBA/CTBN/MWCNT-COOHs nanocomposites were increased at lower MWCNT-COOH concentration. A homogenous dispersion of nanocomposites at lower MWCNT–COOH concentration was observed by SEM.

  12. Characterization of five new mutants in the carboxyl-terminal domain of human apolipoprotein E: No cosegregation with severe hyperlipidemia

    Energy Technology Data Exchange (ETDEWEB)

    Maagdenberg, A.M.J.M. van den; Bruijn, I.H. de; Hofker, M.H.; Frants, R.R. (Leiden Univ. (Netherlands)); Knijff, P. de; Smelt, A.H.M.; Leuven, J.A.G.; van' t Hooft, F.; Assmann, G.; Havekes, L.M. (Univ. Hospital, Leiden (Netherlands)); Weng, Wei; Funke, H. (Westfalische Wilhelms-Universitaet, Muester (Germany))

    1993-05-01

    Assessment of the apolipoprotein E (apoE) phenotype by isoelectric focusing of both hyperlipidemic and normolipidemic individuals identified five new variants. All mutations were confined to the downstream part of the APOE gene by using denaturing gradient gel electrophoresis (DGGE). Sequence analysis revealed five new mutations causing unique amino acid substitutions in the carboxyl-terminal part of the protein containing the putative lipid-binding domain. Three hyperlipoproteinemic probands were carriers of the APOE*2(Va1236[r arrow]Glu) allele, the APOE*3(Cys112-Arg; Arg251[r arrow]Gly) allele, or the APOE*1(Arg158[r arrow]Cys; Leu252[r arrow]Glu) allele. DGGE of the region encoding the receptor-binding domain was useful for haplotyping the mutations at codons 112 and 158. Family studies failed to demonstrate cosegregation between the new mutations and severe hyperlipoproteinemia, although a number of carriers for the APOE*3(Cys112[r arrow]Arg; Arg251[r arrow]Gly) allele and the APOE*1(Arg158-Cys; Leu252[r arrow]Glu) allele expressed hypertriglyceridemia and/ or hypercholesterolemia. Two other mutant alleles, APOE*4[sup [minus

  13. Isolation and characterization of Exodus-2, a novel C-C chemokine with a unique 37-amino acid carboxyl-terminal extension.

    Science.gov (United States)

    Hromas, R; Kim, C H; Klemsz, M; Krathwohl, M; Fife, K; Cooper, S; Schnizlein-Bick, C; Broxmeyer, H E

    1997-09-15

    Chemokines are a group of small, homologous proteins that regulate leukocyte migration, hemopoiesis, and HIV-1 absorption. We report here the cloning and characterization of a novel murine and human C-C chemokine termed Exodus-2 for its similarity to Exodus-1/MIP-3alpha/LARC, and its chemotactic ability. This novel chemokine has a unique 36 or 37 (murine and human, respectively) amino acid carboxyl-terminal extension not seen in any other chemokine family member. Purified recombinant Exodus-2 was found to have two activities classically associated with chemokines: inhibiting hemopoiesis and stimulating chemotaxis. However, Exodus-2 also had unusual characteristics for C-C chemokines. It selectively stimulated the chemotaxis of T-lymphocytes and was preferentially expressed in lymph node tissue. The combination of these characteristics may be a functional correlate for the unique carboxyl-terminal structure of Exodus-2.

  14. Ubiquitin C-Terminal Hydrolase L1 in Tumorigenesis

    Directory of Open Access Journals (Sweden)

    Jennifer Hurst-Kennedy

    2012-01-01

    Full Text Available Ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1, aka PGP9.5 is an abundant, neuronal deubiquitinating enzyme that has also been suggested to possess E3 ubiquitin-protein ligase activity and/or stabilize ubiquitin monomers in vivo. Recent evidence implicates dysregulation of UCH-L1 in the pathogenesis and progression of human cancers. Although typically only expressed in neurons, high levels of UCH-L1 have been found in many nonneuronal tumors, including breast, colorectal, and pancreatic carcinomas. UCH-L1 has also been implicated in the regulation of metastasis and cell growth during the progression of nonsmall cell lung carcinoma, colorectal cancer, and lymphoma. Together these studies suggest UCH-L1 has a potent oncogenic role and drives tumor development. Conversely, others have observed promoter methylation-mediated silencing of UCH-L1 in certain tumor subtypes, suggesting a potential tumor suppressor role for UCH-L1. In this paper, we provide an overview of the evidence supporting the involvement of UCH-L1 in tumor development and discuss the potential mechanisms of action of UCH-L1 in oncogenesis.

  15. Structures and bonding on a colloidal silver surface of the various length carboxyl terminal fragments of bombesin.

    Science.gov (United States)

    Podstawka, Edyta; Ozaki, Yukihiro; Proniewicz, Leonard M

    2008-10-07

    Raman (RS) and surface-enhanced Raman scattering spectra (SERS) were measured for various length carboxyl terminal fragments (X-14 of amino acid sequence) of bombesin ( BN): BN13-14, BN12-14, BN11-14, BN10-14, BN9-14, and BN8-14 in silver colloidal solutions. Density functional theory (DFT) calculations of Raman wavenumbers and intensities with extended basis sets (B3LYP/6-31++G**) were performed with the aim of providing the definitive band allocations to the normal coordinates. The proposed band assignment is consistent with the assignment for similar compounds reported in the literature. The nonadsorbed and adsorbed molecular structures were deducted by detailed spectral analysis of the RS and SERS spectra, respectively. This analysis also allowed us to propose the particular surface geometry and orientation of these peptides on silver surface, and their specific interaction with the surface. For example, a SERS spectrum of BN8-14 indicates that the interaction of a thioether atom and Trp8 with the silver surface is favorable and may dictate the orientation and conformation of adsorbed peptide. One of the most prominent and common features in all of the fragments' SERS spectra is a approximately 692 cm (-1) band due to nu(C-S) accompanied by two or three bands of different C-S conformers for all, except BN8-14, which suggests that all of the above-mentioned compounds adsorb on the silver surface through the thioether atom and that the attachment of Trp8 produces limitation in a number of possible C-S conformers adopted on this surface. Our results also show clearly that His12 and CO do not interact with the colloid surface, which supports our earlier results.

  16. Identification of a nonsense mutation in the carboxyl-terminal region of DNA-dependent protein kinase catalytic subunit in the scid mouse.

    Science.gov (United States)

    Blunt, T; Gell, D; Fox, M; Taccioli, G E; Lehmann, A R; Jackson, S P; Jeggo, P A

    1996-01-01

    DNA-dependent protein kinase (DNA-PK) consists of a heterodimeric protein (Ku) and a large catalytic subunit (DNA-PKcs). The Ku protein has double-stranded DNA end-binding activity that serves to recruit the complex to DNA ends. Despite having serine/threonine protein kinase activity, DNA-PKcs falls into the phosphatidylinositol 3-kinase superfamily. DNA-PK functions in DNA double-strand break repair and V(D)J recombination, and recent evidence has shown that mouse scid cells are defective in DNA-PKcs. In this study we have cloned the cDNA for the carboxyl-terminal region of DNA-PKcs in rodent cells and identified the existence of two differently spliced products in human cells. We show that DNA-PKcs maps to the same chromosomal region as the mouse scid gene. scid cells contain approximately wild-type levels of DNA-PKcs transcripts, whereas the V-3 cell line, which is also defective in DNA-PKcs, contains very reduced transcript levels. Sequence comparison of the carboxyl-terminal region of scid and wild-type mouse cells enabled us to identify a nonsense mutation within a highly conserved region of the gene in mouse scid cells. This represents a strong candidate for the inactivating mutation in DNA-PKcs in the scid mouse. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:8816792

  17. Two separate functions are encoded by the carboxyl-terminal domains of the yeast cyclase-associated protein and its mammalian homologs. Dimerization and actin binding.

    Science.gov (United States)

    Zelicof, A; Protopopov, V; David, D; Lin, X Y; Lustgarten, V; Gerst, J E

    1996-07-26

    The yeast adenylyl cyclase-associated protein, CAP, was identified as a component of the RAS-activated cyclase complex. CAP consists of two functional domains separated by a proline-rich region. One domain, which localizes to the amino terminus, mediates RAS signaling through adenylyl cyclase, while a domain at the carboxyl terminus is involved in the regulation of cell growth and morphogenesis. Recently, the carboxyl terminus of yeast CAP was shown to sequester actin, but whether this function has been conserved, and is the sole function of this domain, is unclear. Here, we demonstrate that the carboxyl-terminal domains of CAP and CAP homologs have two separate functions. We show that carboxyl-terminals of both yeast CAP and a mammalian CAP homolog, MCH1, bind to actin. We also show that this domain contains a signal for dimerization, allowing both CAP and MCH1 to form homodimers and heterodimers. The properties of actin binding and dimerization are mediated by separate regions on the carboxyl terminus; the last 27 amino acids of CAP being critical for actin binding. Finally, we present evidence that links a segment of the proline-rich region of CAP to its localization in yeast. Together, these results suggest that all three domains of CAP proteins are functional.

  18. Involvement of the carboxyl-terminal region of the yeast peroxisomal half ABC transporter Pxa2p in its interaction with Pxa1p and in transporter function.

    Directory of Open Access Journals (Sweden)

    Cheng-Yi Chuang

    Full Text Available BACKGROUND: The peroxisome is a single membrane-bound organelle in eukaryotic cells involved in lipid metabolism, including β-oxidation of fatty acids. The human genetic disorder X-linked adrenoleukodystrophy (X-ALD is caused by mutations in the ABCD1 gene (encoding ALDP, a peroxisomal half ATP-binding cassette [ABC] transporter. This disease is characterized by defective peroxisomal β-oxidation and a large accumulation of very long-chain fatty acids in brain white matter, adrenal cortex, and testis. ALDP forms a homodimer proposed to be the functional transporter, whereas the peroxisomal transporter in yeast is a heterodimer comprising two half ABC transporters, Pxa1p and Pxa2p, both orthologs of human ALDP. While the carboxyl-terminal domain of ALDP is engaged in dimerization, it remains unknown whether the same region is involved in the interaction between Pxa1p and Pxa2p. METHODS/PRINCIPAL FINDINGS: Using a yeast two-hybrid assay, we found that the carboxyl-terminal region (CT of Pxa2p, but not of Pxa1p, is required for their interaction. Further analysis indicated that the central part of the CT (designated CT2 of Pxa2p was indispensable for its interaction with the carboxyl terminally truncated Pxa1_NBD. An interaction between the CT of Pxa2p and Pxa1_NBD was not detected, but could be identified in the presence of Pxa2_NBD-CT1. A single mutation of two conserved residues (aligned with X-ALD-associated mutations at the same positions in ALDP in the CT2 of the Pxa2_NBD-CT protein impaired its interaction with Pxa1_NBD or Pxa1_NBD-CT, resulting in a mutant protein that exhibited a proteinase K digestion profile different from that of the wild-type protein. Functional analysis of these mutant proteins on oleate plates indicated that they were defective in transporter function. CONCLUSIONS/SIGNIFICANCE: The CT of Pxa2p is involved in its interaction with Pxa1p and in transporter function. This concept may be applied to human ALDP studies

  19. Molecular cloning and recombinant expression of the VP28 carboxyl-terminal hydrophilic region from a brazilian white spot syndrome virus isolate

    Directory of Open Access Journals (Sweden)

    Patricia Braunig

    2011-04-01

    Full Text Available In the present study, a fragment of the VP28 coding sequence from a Brazilian WSSV isolate (BrVP28 was cloned, sequenced and expressed in E. coli BL21(DE3 pLysS strain in order to produce the VP28 carboxyl-terminal hydrophilic region. The expression resulted in a protein of about 21 kDa, which was purified under denaturing conditions, resulting in a final highly purified BrVP28 preparation. The recombinant protein obtained can be used in several biotechnology applications, such as the production of monoclonal antibodies which could be used in the development of diagnostic tools as well as in the studies on the characterization of white spot syndrome virus (WSSV isolated in Brazil.

  20. Water-COOH Composite Structure with Enhanced Hydrophobicity Formed by Water Molecules Embedded into Carboxyl-Terminated Self-Assembled Monolayers.

    Science.gov (United States)

    Guo, Pan; Tu, Yusong; Yang, Jinrong; Wang, Chunlei; Sheng, Nan; Fang, Haiping

    2015-10-30

    By combining molecular dynamics simulations and quantum mechanics calculations, we show the formation of a composite structure composed of embedded water molecules and the COOH matrix on carboxyl-terminated self-assembled monolayers (COOH SAMs) with appropriate packing densities. This composite structure with an integrated hydrogen bond network inside reduces the hydrogen bonds with the water above. This explains the seeming contradiction on the stability of the surface water on COOH SAMs observed in experiments. The existence of the composite structure at appropriate packing densities results in the two-step distribution of contact angles of water droplets on COOH SAMs, around 0° and 35°, which compares favorably to the experimental measurements of contact angles collected from forty research articles over the past 25 years. These findings provide a molecular-level understanding of water on surfaces (including surfaces on biomolecules) with hydrophilic functional groups.

  1. Studies on blends of cycloaliphatic epoxy resin with varying concentrations of carboxyl terminated butadiene acrylonitrile copolymer I: Thermal and morphological properties

    Indian Academy of Sciences (India)

    Garima Tripathi; Deepak Srivastava

    2009-04-01

    Differential scanning calorimetric (DSC), thermogravimetric analysis (TGA) and dynamic mechanical analysis (DMA) of the blends of cycloaliphatic epoxy (CAE) resin toughened with liquid elastomer such as carboxyl terminated butadiene acrylonitrile copolymer (CTBN) have been carried out. Exothermal heat of reaction due to cross linking of the resin in the presence of diamino diphenyl sulphone (DDS, an amine hardener) showed a decreasing trend with increasing rubber concentration. Enhancement of thermal stability as well as lower mass loss of the epoxy–rubber blends with increasing rubber concentration have been observed in thermogravimetric analysis (TGA). Dynamic mechanical properties reflected a monotonic decrease in the storage modulus (′) with increasing rubber concentration. The loss modulus (″) and the loss tangent (tan ) values, however, showed an increasing trend with rise of temperature up to a maximum (peak) followed by a gradual fall in both cases.

  2. Structure and carboxyl-terminal domain (CTD) binding of the Set2 SRI domain that couples histone H3 Lys36 methylation to transcription.

    Science.gov (United States)

    Vojnic, Erika; Simon, Bernd; Strahl, Brian D; Sattler, Michael; Cramer, Patrick

    2006-01-06

    During mRNA elongation, the SRI domain of the histone H3 methyltransferase Set2 binds to the phosphorylated carboxyl-terminal domain (CTD) of RNA polymerase II. The solution structure of the yeast Set2 SRI domain reveals a novel CTD-binding fold consisting of a left-handed three-helix bundle. NMR titration shows that the SRI domain binds an Ser2/Ser5-phosphorylated CTD peptide comprising two heptapeptide repeats and three flanking NH2-terminal residues, whereas a single CTD repeat is insufficient for binding. Residues that show strong chemical shift perturbations upon CTD binding cluster in two regions. Both CTD tyrosine side chains contact the SRI domain. One of the tyrosines binds in the region with the strongest chemical shift perturbations, formed by the two NH2-terminal helices. Unexpectedly, the SRI domain fold resembles the structure of an RNA polymerase-interacting domain in bacterial sigma factors (domain sigma2 in sigma70).

  3. The phenotypic features of osteogenesis imperfecta resulting from a mutation of the carboxyl-terminal pro alpha 1(I) propeptide that impairs the assembly of type I procollagen and formation of the extracellular matrix

    NARCIS (Netherlands)

    Cole, WG; Chow, CW; Bateman, JF; Sillence, DO

    1996-01-01

    The features of a baby with lethal perinatal osteogenesis imperfecta (OI-II), resulting from the substitution of tryptophan 94 by cysteine in the carboxyl-terminal propeptide of pro alpha 1(I) chains of type I procollagen, were studied. The limbs and torso were of normal length, shape, and proportio

  4. Influence of Isoforms and Carboxyl-Terminal Truncations on the Capacity of Apolipoprotein E To Associate with and Activate Phospholipid Transfer Protein.

    Science.gov (United States)

    Dafnis, Ioannis; Metso, Jari; Zannis, Vassilis I; Jauhiainen, Matti; Chroni, Angeliki

    2015-09-29

    Phospholipid transfer protein (PLTP), a main protein in lipid and lipoprotein metabolism, exists in high-activity (HA-PLTP) and low-activity (LA-PLTP) forms in human plasma. Proper phospholipid transfer activity of PLTP is modulated by interactions with various apolipoproteins (apo) including apoE. The domains of apoE involved in interactions with PLTP are not known. Here we analyzed the capacity of recombinant apoE isoforms and apoE4 mutants with progressive carboxyl-terminal deletions to bind to and activate HA-PLTP and LA-PLTP. Our analyses demonstrated that lipid-free apoE isoforms bind to both HA-PLTP and LA-PLTP, resulting in phospholipid transfer activation, with apoE3 inducing the highest PLTP activation. The isoform-specific differences in apoE/PLTP binding and PLTP activation were abolished following apoE lipidation. Lipid-free apoE4[Δ(260-299)], apoE4[Δ(230-299)], apoE4[Δ(203-299)], and apoE4[Δ(186-299)] activated HA-PLTP by 120-160% compared to full-length apoE4. Lipid-free apoE4[Δ(186-299)] also activated LA-PLTP by 85% compared to full-length apoE4. All lipidated truncated apoE4 forms displayed a similar effect on HA-PLTP and LA-PLTP activity as full-length apoE4. Strikingly, lipid-free or lipidated full-length apoE4 and apoE4[Δ(186-299)] demonstrated similar binding capacity to LA-PLTP and HA-PLTP. Biophysical studies showed that the carboxyl-terminal truncations of apoE4 resulted in small changes of the structural or thermodynamic properties of lipidated apoE4, that were much less pronounced compared to changes observed previously for lipid-free apoE4. Overall, our findings show an isoform-dependent binding to and activation of PLTP by lipid-free apoE. Furthermore, the domain of apoE4 required for PLTP activation resides within its amino-terminal 1-185 region. The apoE/PLTP interactions can be modulated by the conformation and lipidation state of apoE.

  5. Molecular mechanism of the intramembrane cleavage of the β-carboxyl terminal fragment of amyloid precursor protein by γ-secretase

    Directory of Open Access Journals (Sweden)

    Maho eMorishima-Kawashima

    2014-11-01

    Full Text Available Amyloid β-protein (Aβ plays a central role in the pathogenesis of Alzheimer’s disease, the most common age-associated neurodegenerative disorder. Aβ is generated through intramembrane proteolysis of the β-carboxyl terminal fragment (βCTF of β-amyloid precursor protein (APP by γ-secretase. The initial cleavage by γ-secretase occurs in the membrane/cytoplasm boundary of the βCTF, liberating the APP intracellular domain (AICD. The remaining βCTFs, which are truncated at the C-terminus (longer Aβs, are then cropped sequentially in a stepwise manner, predominantly at three residue intervals, to generate Aβ. There are two major Aβ product lines which generate Aβ40 and Aβ42 with concomitant release of three and two tripeptides, respectively. Additionally, many alternative cleavages occur, releasing peptides with three to six residues. These modulate the Aβ product lines and define the species and quantity of Aβ generated. Here, we review our current understanding of the intramembrane cleavage of the βCTF by γ-secretase, which may contribute to the future goal of developing an efficient therapeutic strategy for Alzheimer’s disease.

  6. Preparation and Characterization of Carboxyl-terminated Poly (butadiene-co-acrylonitrile)-epoxy Resin Prepolymers for Fusion-bonded-epoxy Powder Coating

    Institute of Scientific and Technical Information of China (English)

    LIU Jingcheng; JIA Xiuli; ZHANG Shengwen; LIU Ren; LIU Xiaoya

    2012-01-01

    Liquid carboxyl-terminated poly(butadiene-co-acrylonitrile)(CTBN)-epoxy resin(EP)prepolymers were prepared with different contents of CTBN.The chemical reactions between EP and CTBN were characterized by Fourier ransform infrared (FTIR) spectroscopy and gel permeation chromatography (GPC).The scanning electron micrograph (SEM) and dynamic mechanical analysis (DMA) of curing films showed phase separation,and the rubber particles were finely dispersed in the epoxy matrix.Mechanical properties analysis of curing films showed that impact strength and elongation at break increased significantly upon the addition of CTBN,indicating good toughness of the modified epoxy resins.Thermogravimetric analysis (TGA) showed that the incorporation of CTBN had little effect on the thermal stability of EP.Fusion-bondedepoxy (FBE) powder coatings modified with CTBN-EP prepolymers were prepared.The experimental results demonstrate the ability of CTBN-EP prepolymers,toughening technology to dramatically enhance the flexibility and impact resistance of FBE coatings without compromising other key properties such as corrosion protection.

  7. Expression of the Gene Encoding the Tetraploid of Carboxyl-terminal Peptide of β-hCG Containing Thirty-seven Amino Acid Residues in E. coli

    Institute of Scientific and Technical Information of China (English)

    王健; 沈卫英; 周清平; 申庆祥

    2000-01-01

    Objective This study was carried out to investigate the possible enhancement of immunogenicity of the carboxyl-terminal peptide of β-hCG which is made up of 37 amino acid residues (109~145) and contains the specific epitope (antigenic determinant) of hCG.Materials & Methods hCGβ-CTP37 tetraploid cDNA was constructed by linking four hCGβ-CTP37 cDNAs together. The product was then subcloned into the E. coli expression vector pQE60 to construct the expression vector pQE60/ (hCGβ-CTP37)4. Recombinant (hCGβ-CTP37 ) 4 was expressed in E. coil-X-blue.Results Western blot analysis showed that the tetraploid of hCGβ-CTP37 had an apparent molecular weight of 20 kD and had relatively stronger anti-hCG antibody-binding activity compared with the diploid from.Conclusion The tetraploid of hCGβ-CTP37 may be a more potent immunogen for raising anti-hCG vaccines for fertility regulation or suppression of tumor.

  8. Improvement of in vitro stability and pharmacokinetics of hIFN-α by fusing the carboxyl-terminal peptide of hCG β-subunit.

    Science.gov (United States)

    Ceaglio, Natalia; Gugliotta, Agustina; Tardivo, María Belén; Cravero, Dianela; Etcheverrigaray, Marina; Kratje, Ricardo; Oggero, Marcos

    2016-03-10

    Improving in vivo half-life and in vitro stability of protein-based therapeutics is a current challenge for the biopharmaceutical industry. In particular, recombinant human interferon alpha-2b (rhIFN-α2b), which belongs to a group of cytokines extensively used for the treatment of viral diseases and cancers, shows a poor stability in solution and an extremely short plasma half-life which determines a strict therapeutic regimen comprising high and repeated doses. In this work, we have used a strategy based on the fusion of the carboxyl-terminal peptide (CTP) of human chorionic gonadotropin (hCG) β-subunit, bearing four O-linked oligosaccharide recognition sites, to each or both N- and C-terminal ends of rhIFN-α2b. Molecules containing from 5 (CTP-IFN and IFN-CTP) to 9 (CTP-IFN-CTP) O-glycosylation sites were efficiently expressed and secreted to CHO cells supernatants, and exhibited antiviral and antiproliferative bioactivities in vitro. Significant improvements in pharmacokinetics in rats were achieved through this approach, since the doubly CTP-modified IFN variant showed a 10-fold longer elimination half-life and a 19-fold decreased plasma apparent clearance compared to the wild-type cytokine. Moreover, CTP-IFN-CTP demonstrated a significant increase in in vitro thermal resistance and a higher stability against plasma protease inactivation, both features attributed to the stabilizing effects of the O-glycans provided by the CTP moiety. These results constitute the first report that postulates CTP as a tag for improving both the in vitro and in vivo stability of rhIFN-α2b which, in turn, would positively influence its in vivo bioactivity.

  9. Epoxy Resin Modified Carboxyl-terminated Nitrile Rubber%端羧基丁腈橡胶改性环氧树脂的研究

    Institute of Scientific and Technical Information of China (English)

    陈青; 宫大军; 魏伯荣; 柳丛辉

    2011-01-01

    Using 2-ethyl-4-methyl imidazole as curing agent, pre-polymerization and curing reaction of epoxy resin modified by carboxyl-terminated nitrile rubber(CTBN) were analyzed by infrared spectroscopy.By testing the impact strength, flexural strength, tensile shear strength of modified cured epoxy resin, the toughening effects were evaluated and the toughening mechanism was investigated.The thermal stability of the cured resin was studied by thermogravimetric analysis(TG).The results show that the impact strength of CTBN modified EP increases greatly, the tensile shear strength increases and the thermal stability improves, but the bending strength and bending modulus decrease.%以2-乙基-4-甲基咪唑作为固化剂,运用红外光谱对端羧基丁腈橡胶(CTBN)增韧改性环氧树脂(EP)的预聚反应和固化反应进行分析.通过测试改性环氧树脂固化物的冲击强度、弯曲强度和拉伸剪切强度评价其增韧效果,探讨其增韧机理,并通过热失重(TG)分析固化物的热稳定性能.结果表明:CTBN改性EP后,树脂的冲击强度明显提高,拉伸剪切强度有所提高,热稳定性能提高,但弯曲强度和弯曲模量有所降低.

  10. Technology in L1

    DEFF Research Database (Denmark)

    Elf, Nikolaj Frydensbjerg; Hanghøj, Thorkild; Skaar, Håvard

    2015-01-01

    In recent decades, several Scandinavian research projects have had an explicit focus on how technology intervenes in L1 (or so-called Mother Tongue Education) practices in Swedish, Norwegian and Danish educational contexts, and how this may impact on understanding of the subject. There is currently...... of empirical stud-ies, what characterizes the research field?; and 3) for discussion, which broader implications does the review suggest for a rethinking of L1 in terms of practice and research? Introducing the notion of educa-tional boundary objects, a theoretical framework is developed, which suggests four...... metaphors for un-derstanding technology within L1: as a tool, as media, as socialization, and as literacy practices. These are found useful for analyzing and comparing both theoretical perspectives and empirical research on L1. A key finding of the study is that, although the included research...

  11. Association of the beta-1 adrenergic receptor carboxyl terminal variants with left ventricular hypertrophy among diabetic and non-diabetic survivors of acute myocardial infarction

    Directory of Open Access Journals (Sweden)

    Hakalahti Anna E

    2010-08-01

    Full Text Available Abstract Background The beta-1 adrenergic receptor (β1AR plays a fundamental role in the regulation of cardiovascular functions. It carries a nonsynonymous single nucleotide polymorphism in its carboxyl terminal tail (Arg389Gly, which has been shown to associate with various echocardiographic parameters linked to left ventricular hypertrophy (LVH. Diabetes mellitus (DM, on the other hand, represents a risk factor for LVH. We investigated the possible association between the Arg389Gly polymorphism and LVH among non-diabetic and diabetic acute myocardial infarction (AMI survivors. Methods The study population consisted of 452 AMI survivors, 20.6% of whom had diagnosed DM. Left ventricular parameters were measured with two-dimensional guided M-mode echocardiography 2-7 days after AMI, and the Arg389Gly polymorphism was determined using a polymerase chain reaction-restriction fragment length polymorphism assay. Results The Arg389 homozygotes in the whole study population had a significantly increased left ventricular mass index (LVMI when compared to the Gly389 carriers (either Gly389 homozygotes or Arg389/Gly389 heterozygotes [62.7 vs. 58.4, respectively (p = 0.023]. In particular, the Arg389 homozygotes displayed thicker diastolic interventricular septal (IVSd measures when compared to the Gly389 carriers [13.2 vs. 12.3 mm, respectively (p = 0.004]. When the euglycemic and diabetic patients were analyzed separately, the latter had significantly increased LVMI and diastolic left ventricular posterior wall (LVPWd values compared to the euglycemic patients [LVMI = 69.1 vs. 58.8 (p = 0.001 and LVPWd = 14.2 vs. 12.3 mm (p Conclusions The data suggest an association between the β1AR Arg389Gly polymorphism and LVH, particularly the septal hypertrophy. The Arg389 variant appears to confer a higher risk of developing LVH than the corresponding Gly389 variant among patients who have suffered AMI. This association cannot be considered to be universal

  12. ATG16L1

    DEFF Research Database (Denmark)

    Salem, Mohammad; Ammitzboell, Mette; Nys, Kris;

    2015-01-01

    Genetic variations in the autophagic pathway influence genetic predispositions to Crohn disease. Autophagy, the major lysosomal pathway for degrading and recycling cytoplasmic material, constitutes an important homeostatic cellular process. Of interest, single-nucleotide polymorphisms in ATG16L1...... (autophagy-related 16-like 1 [S. cerevisiae]), a key component in the autophagic response to invading pathogens, have been associated with an increased risk of developing Crohn disease. The most common and well-studied genetic variant of ATG16L1 (rs2241880; leading to a T300A conversion) exhibits a strong...

  13. Electrochemical biosensor for carbofuran pesticide based on esterases from Eupenicillium shearii FREI-39 endophytic fungus.

    Science.gov (United States)

    Grawe, Gregory Ferreira; de Oliveira, Tássia Regina; de Andrade Narciso, Esther; Moccelini, Sally Katiuce; Terezo, Ailton José; Soares, Marcos Antonio; Castilho, Marilza

    2015-01-15

    In this work, a biosensor was constructed by physical adsorption of the isolated endophytic fungus Eupenicillium shearii FREI-39 esterase on halloysite, using graphite powder, multi-walled carbon nanotubes and mineral oil for the determination of carbofuran pesticide by inhibition of the esterase using square-wave voltammetry (SWV). Specific esterase activities were determined each 2 days over a period of 15 days of growth in four different inoculation media. The highest specific activity was found on 6th day, with 33.08 U on PDA broth. The best performance of the proposed biosensor was obtained using 0.5 U esterase activity. The carbofuran concentration response was linear in the range from 5.0 to 100.0 µg L(-1) (r=0.9986) with detection and quantification limits of 1.69 µg L(-1) and 5.13 µg L(-1), respectively. A recovery study of carbofuran in spiked water samples showed values ranging from 103.8±6.7% to 106.7±9.7%. The biosensor showed good repeatability and reproducibility and remained stable for a period of 20 weeks. The determination of carbofuran in spiked water samples using the proposed biosensor was satisfactory when compared to the chromatographic reference method. The results showed no significant difference at the 95% confidence level with t-test statistics. The application of enzymes from endophytic fungi in constructing biosensors broadens the biotechnological importance of these microorganisms.

  14. Studies on the physico-mechanical and thermal characteristics of blends of DGEBA epoxy, 3,4 epoxy cyclohexylmethyl, 3',4'-epoxycylohexane carboxylate and carboxyl terminated butadiene co-acrylonitrile (CTBN)

    Energy Technology Data Exchange (ETDEWEB)

    Tripathi, Garima [Department of Plastic Technology, H. B. Technological Institute, Kanpur 208002 (India); Srivastava, Deepak [Department of Plastic Technology, H. B. Technological Institute, Kanpur 208002 (India)], E-mail: deepak_sri92@rediffmail.com

    2008-11-25

    Toughening of blend of diglycidyl ether of bisphenol-A (DGEBA) and 3,4 epoxy cyclohexylmethyl, 3',4'-epoxycylohexane carboxylate, i.e. cycloaliphatic epoxy resin (CAE) with varying weight ratios (0-25 wt%) of carboxyl terminated butadiene acrylonitrile (CTBN) copolymer have been investigated. Fourier transform infrared (FTIR) spectroscopic analysis established that the interaction between oxirane groups of DGEBA, CAE and CTBN were responsible for characteristics peak shifts in the blends compared to their counterparts. Physico-mechanical properties of the prepared samples, e.g. tensile, flexural and impact strengths showed an optimum concentration of CTBN (15 wt%) into epoxy matrix, which offered maximum toughening. Thermal stability of the prepared samples was analyzed by dynamic thermogravimetric runs. Cross-sections of the cured samples which failed during impact testing have been critically studied through scanning electron microscopic (SEM) analysis to gain insight into the phase morphology.

  15. Amino acid substitutions of conserved residues in the carboxyl-terminal domain of the [alpha]I(X) chain of type X collagen occur in two unrelated families with metaphyseal chondrodysplasia type Schmid

    Energy Technology Data Exchange (ETDEWEB)

    Wallis, G.A.; Rash, B.; Sweetman, W.A.; Thomas, J.T.; Grant, M.E.; Boot-Handford, R.P. (Univ. of Manchester (United Kingdom)); Super, M. (Royal Manchester Children' s Hospital, Manchester (United Kingdom)); Evans, G. (Robert Jones Orthopaedic Hospital, Oswestry (United Kingdom))

    1994-02-01

    Type X collagen is a homotrimeric, short-chain, nonfibrillar extracellular-matrix component that is specifically and transiently synthesized by hypertrophic chondrocytes at the site of endochondral ossification. The precise function of type X collagen is not known, but its specific pattern of expression suggests that mutations within the encoding gene (COL10A1) that alter the structure or synthesis of the protein may cause heritable forms of chondrodysplasia. The authors used the PCR and the SSCP techniques to analyze the coding and upstream promoter regions of the COL10A1 gene in a number of individuals with forms of chondrodysplasia. Using this approach, they identified two individuals with metaphyseal chondrodysplasia type Schmid (MCDS) with SSCP changes in the region of the gene encoding the carboxyl-terminal domain. Sequence analysis demonstrated that the individuals were heterozygous for two unique single-base-pair transitions that led to the substitution of the highly conserved amino acid residue tyrosine at position 598 by aspartic acid in one person and of leucine at position 614 by proline in the other. The substitution at residue 598 segregated with the phenotype in a family of eight (five affected and three unaffected) related persons. The substitutions at residue 614 occurred in a sporadically affected individual but not in her unaffected mother and brother. Additional members of this family were not available for further study. These results suggest that certain amino acid substitutions within the carboxyl-terminal domain of the chains of the type X collagen molecule cause MCDS. These amino acid substitutions are likely to alter either chain recognition or assembly of the type X collagen molecule, thereby depleting the amount of normal type X collagen deposited in the extracellular matrix, with consequent aberrations in bone growth and development. 36 refs., 5 figs.

  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. 多臂端羧基聚硅氧烷的可控合成及表征%Controlled Synthesis and Characteristics of Multi-branched Carboxyl Terminated Polydimethylsiloxane

    Institute of Scientific and Technical Information of China (English)

    程柳军; 刘琼琼; 杨林; 张安强; 林雅铃

    2014-01-01

    以八甲基环四硅氧烷(D4)、苯基三(二甲基硅氧烷基)硅烷(PTDMS,俗称三氢封头剂)或四(二甲基硅氧基)硅烷(TDMS,俗称四氢封头剂)为原料,通过开环聚合法合成了三端氢基聚硅氧烷(PDMS-H3)和四端氢基聚硅氧烷(PDMS-H4);再将其与甲基丙烯酸叔丁酯(t-BMA)在氯铂酸-四氢呋喃(THF)溶液催化下通过硅氢加成反应得到三端酯基聚硅氧烷(PDMS-t-BMA3)和四端酯基聚硅氧烷(PDMS-t-BMA4);最后在浓硫酸催化下水解制得三端羧基聚硅氧烷(PDMS-COOH3)和四端羧基聚硅氧烷(PDMS-COOH4)。采用FT-IR、1 H NMR和GPC对各阶段产物的结构和摩尔质量及其分布进行表征。结果表明,通过开环聚合、硅氢加成、酸性水解三步反应能够获得结构清晰、摩尔质量可控的多臂端羧基聚硅氧烷,且叔丁酯基聚硅氧烷酸性水解反应效率高,后处理简单,对硅氧烷主链的影响较小。%With octamethylcyclotetrasiloxane (D4 ),tris (dimethylsiloxy) phenylsilane (PTDMS ) or tetrakis (dimethylsiloxy)-silane (TDMS )as raw materials,hydrogen terminated polydimethylsilox-anes (PDMS-Hx,x=3,4)were synthesized via ring-opening polymerization.Then tert-butyl methacry-late (t-BMA)was grafted to PDMS-Hx via hydrosilylation to prepare ester terminated polydimethylsilox-anes (PDMS-t-BMAx,x=3,4)with H2PtCl6 as catalyst,and the multi-branched carboxyl terminated polydimethylsiloxanes (PDMS-COOHx,x=3,4)were prepared by subsequent hydrolysis reaction under acid.The structures and molecular weights of the products were characterized by FT-IR,1 H NMR and GPC.Results show that multi-branched carboxyl terminated polydimethylsiloxanes with certain structures and molecular weights can be synthesized under control via a three-step procedure:ring-opening polymeri-zation,hydrosilylation reaction and acid hydrolysis reaction.The acid hydrolysis of polydimethylsiloxanes with a tertiary

  18. Connexins in the early development of the African clawed frog Xenopus laevis (Amphibia: The role of the connexin43 carboxyl terminal tail in the establishment of the dorso-ventral axis

    Directory of Open Access Journals (Sweden)

    Jaime Cofre

    2007-03-01

    Full Text Available Connexins are a family of related proteins identified in vertebrate forming gap junctions, which mediate cell-to-cell communication in early embryos, with an important role in establishing embryonic asymmetry and ‘communication compartments’. By in situ hybridization, immunocytochemistry, reverse transcriptase PCR (RT-PCR and western blotting we show that a Cx43-like molecule is present in oocytes and embryos of the African clawed frog Xenopus laevis, with specific localization in the animal-vegetal axis. This specific distribution is suggestive for an important role for this protein in the establishment of the dorso-ventral axis. Antisense RNA and antibodies directed against rat carboxyl terminal tail of the Cx43 (CT-Cx43 and injected in 1-cell stage Xenopus embryos, induced pronounced alterations in nervous system development, with a severe ventralization phenotype. Coherently, the overexpression of CT-Cx43 produced a dorsalization of the embryos. In antisense treated embryos, the expression of the beta-catenin gene is eliminated from the Nieuwkoop center, the pattern expression of the Chordin, Xnot and Xbra is modified, with no effect in expression of the Goosecoid gene. In CT-Cx43 mRNA treated embryos the pattern of expression of the beta-catenin, Chordin, Goosecoid, Xnot and engrailed-2 genes is modified. The expression of beta-catenin is increased in the Nieuwkoop center, the expression pattern of Chordin and Goosecoid is expanded to the posterior neural plate and engrailed-2 presents ectopic expression in the ventral region. Taken together our data suggest a role for CT-Cx43 as a maternal determinant with a critical function in the formation of the dorso-ventral axis in Xenopus laevis. The Cx43 may be one of the earliest markers of the dorso-ventral axis in these embryos and could possibly be acting through regionalization of factors responsible for the establishment of this axis.

  19. Identification of genes encoding microbial glucuronoyl esterases

    NARCIS (Netherlands)

    Li, Xin-Liang; Spániková, Silvia; de Vries, Ronald P; Biely, Peter

    2007-01-01

    One type of covalent linkages connecting lignin and hemicellulose in plant cell walls is the ester linkage between 4-O-methyl-D-glucuronic acid of glucuronoxylan and lignin alcohols. An enzyme that could hydrolyze such linkages, named glucuronoyl esterase, occurs in the cellulolytic system of the wo

  20. 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 ...

  1. Phenol esterase activity of porcine skin.

    Science.gov (United States)

    Laszlo, Joseph A; Smith, Leslie J; Evans, Kervin O; Compton, David L

    2015-01-01

    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 with octadecanol, glycerol, and dioleoylglycerol. These phenolic derivatives were treated in taurodeoxycholate microemulsion and unilamellar liposomes with ex vivo porcine skin and an aqueous extract of the skin. Extracted esterases hydrolyzed the microemulsions at rates in the order: tyrosyl lipoate > tyrosyl decanoate > hydroxytyrosyl lipoate > hydroxytyrosyl decanoate. The tyrosyl decanoate was subject to comparatively little hydrolysis (10-30% after 24h) when incorporated into liposomes, while hydroxytyrosyl decanoate in liposomes was not hydrolyzed at all by the skin extract. Ferulate esters were not hydrolyzed by the extract in aqueous buffer, microemulsion, nor liposomes. Tyrosyl decanoate applied topically to skin explants in microemulsion were readily hydrolyzed within 4h, while hydrolysis was minimal when applied in liposomes. These findings indicate that porcine skin displays a general esterase activity toward medium-chain esters of tyrosol and hydroxytyrosol, which can be moderated by the physiochemical properties of the lipid vehicle, but no feruloyl esterase activity.

  2. Sparse Recovery via l1 and L1 Optimization

    Science.gov (United States)

    2014-11-01

    Sparse Recovery via `1 and L1 Optimization Stanley Osher and Wotao Yin Department of Mathematics , UCLA sjo@math.ucla.edu and wotaoyin@math.ucla.edu 1...WORK UNIT NUMBER 7. PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) University of California, Los Angeles,Department of Mathematics ,Los Angeles...ϕi, ϕj〉 = δjk, for i, j = 1, . . . , N . We get densely supported ϕj , (think of sines and cosines when V = 0). Physicists and chemists want short

  3. A comparative study on esterases from three species of Raillietina.

    Science.gov (United States)

    Balasubramanian, M P; Dhandayuthapani, S; Nellaiappan, K; Ramalingam, K

    1984-06-01

    The multiplicity of soluble esterases in Raillietina tetragona, R. echinobothrida and R. cesticillus was studied by use of slab polyacrylamide gel electrophoresis. Five fractions of esterase activity were observed in R. tetragona, seven in R. echinobothrida and three in R. cesticillus. The various fractions of esterase activity of closely related species of Raillietina showed differential behaviour towards various chemicals. Based on the inhibitory effect of inhibitors p-CMB, EDTA, malathion, silver nitrate and eserine sulphate, the various esterases have been classified into arylesterase, carboxylesterase, acetylesterase and cholinesterase.

  4. Main: L1DCPAL1 [PLACE

    Lifescience Database Archive (English)

    Full Text Available L1DCPAL1 S000504 15-September-2006 (last modified) kehi L1 element, found in PAL1 p...romoter in carrot (Daucus carota), is a protoplastization (dilution) responsive element; L1 contains Box L-l...ike sequence (ACCTACCC); see also S000492 (BOXL CORE of DC PAL1); L1 Daucus carota (carrot) ATTCACCTACCC ...

  5. Studies on the Purification and Characterization of Soybean Esterase,and Its Sensitivity to Organophosphate and Carbamate Pesticides

    Institute of Scientific and Technical Information of China (English)

    LI Jian-ke; ZHOU Yan-li; WEN Yan-xia; WANG Jian-hua; HU Qiu-hui

    2009-01-01

    Soybean esterase,a cholinesterase-like enzyme,was purified by differential centrifugation firstly,then,ammonium sulfate precipitation,dialysis,and finally,DEAE-cellulose-32 ion-exchange chromatography after extracting it from soybean seeds with phosphate buffer(0.3 mol L-1,pH 7.0).The extract recovery rate of the purified enzyme was 8.18% and purification fold was 91.58.The soybean esterase appeared as two bands on the denaturing SDS-PAGE with molecular weights of 24 and 37.2 kDa,respectively,which proved that it is a dimer protein consisting of two subunits.The result of nondenaturing PAGE revealed that the soybean esterase is a single band with cholinesterase-like activity using α-naphthyl acetate as the substrate and fast blue B salt as coloring agent.The esterase showed very high sensitivity to 18 kinds of organophosphate pesticides and 6 kinds of carbamate pesticides with the lowest detective limits of 0.03125-0.0625 and 0.03 125-0.25 mg kg-1,respectively,and can meet the demands of MRL specified by the most countries.

  6. 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 ...

  7. Main: L1BOXATPDF1 [PLACE

    Lifescience Database Archive (English)

    Full Text Available L1BOXATPDF1 S000386 05-November-2005 (last modified) kehi L1 box found in promoter ...of Arabidopsis thaliana (A.t.) PROTODERMAL FACTOR1 (PDF1) gene; Located between -134 and -127; Involved in L...ding motif (Wang et al., 2004); HDZip IV; See also S000371; PDF1; L1 box; L1 layer-specific expression; Shoo

  8. 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

  9. [Purification and characterization of esterase from Morganella morganii ZJB-09203].

    Science.gov (United States)

    Zheng, Renchao; Wang, Tianzhen; Li, Xiaojun; Zheng, Yuguo

    2014-01-01

    Enantioselective hydrolysis of 2-carboxyethyl-3-cyano-5-methylhexanoic acid (CNDE) is the key step in chemoenzymatic synthesis of pregabalin. We purified an intracellular carboxyl esterase from Morganella morganii ZJB-09203, which exhibited high enantioselectivity and activity towards CNDE. The carboxyl esterase was purified to electrophoretic homogeneity by ammonium sulfate fraction precipitation, Phenyl Sepharose 6 FF hydrophobic interaction chromatography, anion exchange with DEAE Sephadex A-50 and Bio-Scale CHT column. The purified enzyme was a monomer with molecular mass of 68 kDa determined by SDS-PAGE and gel chromatography. Substrate specificity of the enzyme towards p-nitrophenyl esters suggested that the purified enzyme was an esterase. The optimal reaction pH for CNDE hydrolysis was 9.0, and optimal temperature was 45 degrees C. The esterase was stable between pH 7.0 and 9.0, and at 40 degrees C. The enzyme activity was enhanced by Ca2+, Cu2+ and Mn2+, whereas strongly inhibited by Co2+, Fe3+, Ni2+ and EDTA. Meanwhile, we investigated the kinetic parameters of the esterase towards p-nitrophenyl esters and effect of CNDE concentration on conversion. The present study reported the esterase capable of stereospecific hydrolysis of CNDE for the first time. Our research will provide foundations for industrial production of Pregabalin using the new biocatalyst.

  10. PD-L1-specific T cells

    DEFF Research Database (Denmark)

    Ahmad, Shamaila Munir; Borch, Troels Holz; Hansen, Morten

    2016-01-01

    for targeting the tumor microenvironment and for boosting the clinical effects of additional anticancer immunotherapy. This review summarizes present information about PD-L1 as a T cell antigen, depicts the initial findings about the function of PD-L1-specific T cells in the adjustment of immune responses...

  11. Kinetic interactions of a neuropathy potentiator (phenylmethylsulfonyl fluoride) with the neuropathy target esterase and other membrane bound esterases.

    Science.gov (United States)

    Mangas, Iris; Vilanova, Eugenio; Estévez, Jorge

    2014-02-01

    Phenylmethylsulfonyl fluoride (PMSF) is a protease and esterase inhibitor that causes protection, or potentiation/"promotion," of organophosphorus delayed neuropathy (OPIDN), depending on whether it is dosed before or after an inducer of delayed neuropathy, such as mipafox. The molecular target of the potentiation/promotion of OPIDN has not yet been identified. The kinetic data of phenyl valerate esterase inhibition by PMSF were obtained with membrane chicken brain fractions, the animal model and tissue in which neuropathy target esterase (NTE) was first described. Data were analyzed using a kinetic model with a multienzymatic system in which inhibition, simultaneous chemical hydrolysis of the inhibitor and "ongoing inhibition" (inhibition during the substrate reaction) were considered. Three main esterase components were discriminated: two sensitive enzymatic entities representing 44 and 41 %, with I 50 (20 min) of 35 and 198 μM at 37 °C, respectively, and a resistant fraction of 15 % of activity. The estimated constant of the chemical hydrolysis of PMSF was also calculated (kh = 0.28 min(-1)). Four esterase components were globally identified considering also previously data with paraoxon and mipafox: EPα (4-8 %), highly sensitive to paraoxon and mipafox, spontaneously reactivates after inhibition with paraoxon, and resistant to PMSF; EPβ (38-41 %), sensitive to paraoxon and PMSF, but practically resistant to mipafox, this esterase component has the kinetic characteristics expected for the PMSF potentiator target, even though paraoxon cannot be a potentiator in vivo due to high AChE inhibition; EPγ (NTE) (39-48 %), paraoxon-resistant and sensitive to the micromolar concentration of mipafox and PMSF; and EPδ (10 %), resistant to all the inhibitors assayed. This kinetic characterization study is needed for further isolation and molecular characterization studies, and these PMSF phenyl valerate esterase components will have to be considered in further studies

  12. L1-norm minimization for quaternion signals

    CERN Document Server

    Wu, Jiasong; Wang, Xiaoqing; Senhadji, Lotfi; Shu, Huazhong

    2012-01-01

    The l1-norm minimization problem plays an important role in the compressed sensing (CS) theory. We present in this letter an algorithm for solving the problem of l1-norm minimization for quaternion signals by converting it to second-order cone programming. An application example of the proposed algorithm is also given for practical guidelines of perfect recovery of quaternion signals. The proposed algorithm may find its potential application when CS theory meets the quaternion signal processing.

  13. Canonical Sets of Best L1-Approximation

    Directory of Open Access Journals (Sweden)

    Dimiter Dryanov

    2012-01-01

    Full Text Available In mathematics, the term approximation usually means either interpolation on a point set or approximation with respect to a given distance. There is a concept, which joins the two approaches together, and this is the concept of characterization of the best approximants via interpolation. It turns out that for some large classes of functions the best approximants with respect to a certain distance can be constructed by interpolation on a point set that does not depend on the choice of the function to be approximated. Such point sets are called canonical sets of best approximation. The present paper summarizes results on canonical sets of best L1-approximation with emphasis on multivariate interpolation and best L1-approximation by blending functions. The best L1-approximants are characterized as transfinite interpolants on canonical sets. The notion of a Haar-Chebyshev system in the multivariate case is discussed also. In this context, it is shown that some multivariate interpolation spaces share properties of univariate Haar-Chebyshev systems. We study also the problem of best one-sided multivariate L1-approximation by sums of univariate functions. Explicit constructions of best one-sided L1-approximants give rise to well-known and new inequalities.

  14. Characterization of an acetyl esterase from Myceliophthorathermophila C1 able to deacetylate xanthan

    NARCIS (Netherlands)

    Kool, M.M.; Schols, H.A.; Wagenknecht, M.; Hinz, S.W.A.; Moerschbacher, B.M.; Gruppen, H.

    2014-01-01

    Screening of eight carbohydrate acetyl esterases for their activity towards xanthan resulted in the recogni-tion of one active esterase. AXE3, a CAZy family CE1 acetyl xylan esterase originating from Myceliophthorathermophila C1, removed 31% of all acetyl groups present in xanthan after a 48 h incub

  15. Feruloyl Esterases as Biotechnological Tools: Current and Future Perspectives

    Institute of Scientific and Technical Information of China (English)

    Ahmed E FAZARY; Yi-Hsu JU

    2007-01-01

    Feruloyl esterases represent a diverse group of hydrolases catalyzing the cleavage and formation of ester bonds between plant cell wall polysaccharide and phenolic acid. They are widely distributed in plants and microorganisms. Besides lipases, a considerable number of microbial feruloyl esterases have also been discovered and overexpressed. This review summarizes the latest research on their classification,production, and biophysicochemical properties. Special emphasis is given to the importance of that type of enzyme and their related phenolic ferulic acid compound in biotechnological processes, and industrial and medicinal applications.

  16. Face recognition with L1-norm subspaces

    Science.gov (United States)

    Maritato, Federica; Liu, Ying; Colonnese, Stefania; Pados, Dimitris A.

    2016-05-01

    We consider the problem of representing individual faces by maximum L1-norm projection subspaces calculated from available face-image ensembles. In contrast to conventional L2-norm subspaces, L1-norm subspaces are seen to offer significant robustness to image variations, disturbances, and rank selection. Face recognition becomes then the problem of associating a new unknown face image to the "closest," in some sense, L1 subspace in the database. In this work, we also introduce the concept of adaptively allocating the available number of principal components to different face image classes, subject to a given total number/budget of principal components. Experimental studies included in this paper illustrate and support the theoretical developments.

  17. Voice and Narrative in L1 Writing

    DEFF Research Database (Denmark)

    Krogh, Ellen; Piekut, Anke

    2015-01-01

    in lower secondary L1, she found that her previous writing strategies were not rewarded in upper secondary school. In the second empiri-cal study, two upper-secondary exam papers are investigated, with a focus on their approaches to exam genres and their use of narrative resources to address issues...

  18. Reference: L1DCPAL1 [PLACE

    Lifescience Database Archive (English)

    Full Text Available L1DCPAL1 Takeda J, Ito Y, Maeda K, Ozeki Y. Assignment of UVB-responsive cis-elemen...anine ammonia-lyase gene (gDcPAL1). Photochem Photobiol. 76:232-238 (2002). PubMed: 12194222 ...

  19. Activity of pectin methyl esterase during blanching of peaches

    NARCIS (Netherlands)

    Tijskens, L.M.M.; Rodis, P.S.; Hertog, M.L.A.T.M.; Proxenia, N.; Dijk, van C.

    1999-01-01

    The activity of pectin methyl esterase (PE) in peaches during blanching treatments was modelled and analyzed. It was postulated that the enzyme exists in two configurations, one bound and one soluble. The bound configuration can be converted into the soluble configuration. These two configurations h

  20. 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...

  1. 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

  2. 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

  3. High-throughput screening method for lipases/esterases.

    Science.gov (United States)

    Mateos-Díaz, Eduardo; Rodríguez, Jorge Alberto; de Los Ángeles Camacho-Ruiz, María; Mateos-Díaz, Juan Carlos

    2012-01-01

    High-throughput screening (HTS) methods for lipases and esterases are generally performed by using synthetic chromogenic substrates (e.g., p-nitrophenyl, resorufin, and umbelliferyl esters) which may be misleading since they are not their natural substrates (e.g., partially or insoluble triglycerides). In previous works, we have shown that soluble nonchromogenic substrates and p-nitrophenol (as a pH indicator) can be used to quantify the hydrolysis and estimate the substrate selectivity of lipases and esterases from several sources. However, in order to implement a spectrophotometric HTS method using partially or insoluble triglycerides, it is necessary to find particular conditions which allow a quantitative detection of the enzymatic activity. In this work, we used Triton X-100, CHAPS, and N-lauroyl sarcosine as emulsifiers, β-cyclodextrin as a fatty acid captor, and two substrate concentrations, 1 mM of tributyrin (TC4) and 5 mM of trioctanoin (TC8), to improve the test conditions. To demonstrate the utility of this method, we screened 12 enzymes (commercial preparations and culture broth extracts) for the hydrolysis of TC4 and TC8, which are both classical substrates for lipases and esterases (for esterases, only TC4 may be hydrolyzed). Subsequent pH-stat experiments were performed to confirm the preference of substrate hydrolysis with the hydrolases tested. We have shown that this method is very useful for screening a high number of lipases (hydrolysis of TC4 and TC8) or esterases (only hydrolysis of TC4) from wild isolates or variants generated by directed evolution using nonchromogenic triglycerides directly in the test.

  4. L1-norm packings from function fields

    Institute of Scientific and Technical Information of China (English)

    LI Hongli

    2005-01-01

    In this paper, we study some packings in a cube, namely, how to pack n points in a cube so as to maximize the minimal distance. The distance is induced by the L1-norm which is analogous to the Hamming distance in coding theory. Two constructions with reasonable parameters are obtained, by using some results from a function field including divisor class group, narrow ray class group, and so on. We also present some asymptotic results of the two packings.

  5. 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.

  6. 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.

  7. Esterase D and ABO Polymorphisms in Turkish Population

    OpenAIRE

    Perçin, E. Ferda; Sezgin, Ilhan; Çolak, Ahmet; ÇINAR, Ziynet

    2014-01-01

    This study is intended to be a contribution to the determination of the genetic structure of the Turkish population. For this purpose, esterase D (ESD), an erythrocyte enzyme, and ABO blood group loci phenotypes from the blood samples of two hundred healthy and unrelated individuals from different regions of Turkey were determined. ESD enzyme phenotypes were determined by cellulose acetate electrophoresis, and ABO blood group phenotypes were determined by the aglutination test; in this w...

  8. Biochemical Characterization of a First Fungal Esterase from Rhizomucor miehei Showing High Efficiency of Ester Synthesis

    OpenAIRE

    Yu Liu; Haibo Xu; Qiaojuan Yan; Shaoqing Yang; Xiaojie Duan; Zhengqiang Jiang

    2013-01-01

    BACKGROUND: Esterases with excellent merits suitable for commercial use in ester production field are still insufficient. The aim of this research is to advance our understanding by seeking for more unusual esterases and revealing their characterizations for ester synthesis. METHODOLOGY/PRINCIPAL FINDINGS: A novel esterase-encoding gene from Rhizomucor miehei (RmEstA) was cloned and expressed in Escherichia coli. Sequence analysis revealed a 975-bp ORF encoding a 324-amino-acid polypeptide be...

  9. Contribution of L1 in EFL Teaching

    Directory of Open Access Journals (Sweden)

    Wahjuningsih Usadiati

    2009-01-01

    Full Text Available This study is conducted in a classroom action research to improve the students’ achievement in writing English sentences in Present Perfect Tense in Structure 1 lessons. The subject consisted of 20 Semester II students who took Structure I lessons in English Education Department of Palangka Raya University, Central Kalimantan, Indonesia. The data were taken from the results of pre test and post test after the action was done. The results show that in cycle 1, in which the explanations were fully in English, only 40% of the students got a good achievement; 5-7 out of 20 test items were correct. After cycle 2 was done using L1 interchangeably with English in the explanations, the students’ achievement of writing English sentences in Present Perfect Tense increased to 75%, in which 15-18 out 20 test items were correct.

  10. Extended Lagrange interpolation in L1 spaces

    Science.gov (United States)

    Occorsio, Donatella; Russo, Maria Grazia

    2016-10-01

    Let w (x )=e-xβxα , w ¯(x )=x w (x ) and denote by {pm(w)}m,{pn(w¯)}n the corresponding sequences of orthonormal polynomials. The zeros of the polynomial Q2 m +1=pm +1(w )pm(w ¯) are simple and are sufficiently far among them. Therefore it is possible to construct an interpolation process essentially based on the zeros of Q2m+1, which is called "Extended Lagrange Interpolation". Here we study the convergence of this interpolation process in suitable weighted L1 spaces. This study completes the results given by the authors in previous papers in weighted Lup((0 ,+∞ )) , for 1≤p≤∞. Moreover an application of the proposed interpolation process in order to construct an e cient product quadrature scheme for weakly singular integrals is given.

  11. The effect of liver esterases and temperature on remifentanil degradation in vitro.

    Science.gov (United States)

    Piazza, Ornella; Cascone, Sara; Sessa, Linda; De Robertis, Edoardo; Lamberti, Gaetano

    2016-08-20

    Remifentanil is a potent opioid metabolized by serum and tissue esterases; it is routinely administered to patients with liver failure as anaesthetic and analgo-sedative without variation in doses, even if prolonged clinical effects and respiratory depression have been observed in these patients. The aim of this study was to determine remifentanil enzymatic degradation kinetics bearing in mind the effect of liver esterases in order to trace a more accurate pharmacokinetic profile of the drug. Solution samples were taken over time and analysed to measure remifentanil concentration by HPLC. We reproduced the physiological settings, varying temperature and pH in vitro and evaluated the kinetics of degradation of remifentanil in the presence of Rhizopus Oryzae esterases, equine liver esterases and porcine liver esterases. Remifentanil kinetics of degradation was accelerated by porcine liver esterases. Remifentanil in vitro half-life decreases with increasing temperatures in the presence of porcine liver esterases. A drug model simulation considering the effect of temperature in the presence of liver esterases was developed. Remifentanil in vitro half-life decreases with increasing temperatures when porcine liver esterases are present. In this paper we propose a model for describing remifentanil degradation kinetics at various temperatures.

  12. Isozymic variations in specific and nonspecific esterase and its thermostability in silkworm, Bombyx mori L.

    Science.gov (United States)

    Patnaik, Bharat Bhusan; Biswas, Tapati Datta; Nayak, Sandeepta Kumar; Saha, A K; Majumdar, M K

    2012-09-01

    Esterase isozymic variations were documented in the haemolymph of developed multivoltine and bivoltine silkworm breeds during unfavorable seed crop seasons of May - September using á- and â- napthylacetate separately to identify specific and nonspecific esterase having thermotolerant potentiality. Variations existed in the isozyme pattern with three bands (Est-2, 3 and 4) in pure Nistari race and other developed multivoltine and bivoltine breeds. Est-2 and Est-3 were non-specific esterases as they were observed when both á- and â-napthylacetate was used as substrates separately. Est-4 band was observed only with á-napthylacetate as substrate and was therefore confirmed to be specific á-esterase band in the haemolymph of silkworm, Bombyx mori L. Zymograms showed that the non-specific esterase band (Est-3) with R1 of 0.43 and specific á-esterase band (Est-4) with R(f) of 0.32 predominately withstood a temperature of 70 +/- 2 degrees C for a duration of 10 min and were confirmed as thermostable esterases in haemolymph of silkworm, Bombyx mori L. This also categorized the presence of thermostable esterases in developed multivoltine and bivoltine breeds of silkworm, even though the qualitative activity was more in the former than the latter. The qualitative presence of thermostable esterases and their activity could be adopted as an indicative biochemical marker in relation to thermotolerance in silkworm.

  13. Kinetic measurement of esterase-mediated hydrolysis for methacrylate monomers used in dental composite biomaterials

    Science.gov (United States)

    Russo, Karen Ann

    Methacrylate-based monomers are routinely used in medical biomaterials. Monomers undergo polymerization reactions to form the solid resin. These polymerization reactions can be incomplete thus making unpolymerized monomer available for possible biodistribution. Understanding the fate of these monomers is essential not only for their toxicological profile but also for development of future biomaterials. Aromatic methacrylate-based monomers included in this study were bisphenol A dimethacrylate and bisphenol A diglycidyl dimethathacrylate; aliphatic methacrylate monomers were 2-hydroxyethyl methacrylate and triethyleneglycol dimethacrylate. These compounds contain ester moieties thought to be susceptible to esterase-mediated hydrolysis. The hypothesis was that the ester bond of the methacrylate monomers can be hydrolyzed by esterases and these reactions would occur in a measurable, time-dependent manner confirmed by specific Michaelis-Menten kinetic relationships. Including aliphatic and aromatic methacrylate monomers in this work allowed for structure-based comparisons. In vitro enzymolysis of the test compounds by acetylcholinesterase and cholesterol esterase was performed in buffered solutions. The hydrolysis reactions were monitored by high performance liquid chromatography with ultraviolet detection. The disappearance of parent compound and appearance of hydrolysis products were quantitated. The aromatic methacrylate monomers, bisphenol A dimethacrylate and bisphenol A diglycidyl dimethacrylate, were resistant to acetylcholine esterase hydrolysis but were converted by cholesterol esterase. The putative xenoestrogen, bisphenol A, was identified as a hydrolysis product from bisphenol A dimethacrylate conversion. Cholesterol esterase induced hydrolysis of bisphenol A diglycidyl dimethacrylate yielded a Km value of 1584 muM and Vmax of 14 muM min-1. Triethyleneglycol was converted by both esterases with calculated Km values of 394 and 1311 muM for acetylcholine

  14. Production and characterization of esterase in Lantinus tigrinus for degradation of polystyrene.

    Science.gov (United States)

    Tahir, Lubna; Ishtiaq Ali, Muhammad; Zia, Muhammad; Atiq, Naima; Hasan, Fariha; Ahmed, Safia

    2013-01-01

    Polystyrene is considered stable to biological degradation. Lantinus tigrinus isolated from wood sample produced esterase in growth medium under normal conditions. However, acidic medium, 37 degrees C temperature, presence of tween 80; and urea and yeast extract in mineral salt medium enhance the production of esterase and specific activity. Purified esterase was active at broad pH range and 45 degrees C. FTIR analysis confirmed that esterase produced by Lantinus tigrinus effectively degraded polystyrene film and broke macromolecules down to non-toxic molecules. This study concludes that the presence of Lantinus tigrinus at dumping sites can be exploited for waste management containing high molecular weight synthetic polymers.

  15. Analysis list: Tcf7l1 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Tcf7l1 Embryonic fibroblast + mm9 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Tcf7l1....1.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Tcf7l1.5.tsv http://dbarchive.biosci...encedbc.jp/kyushu-u/mm9/target/Tcf7l1.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Tcf7l1.Embr

  16. The vulnerability of gender on determiners in L1, 2L1 and L2 acquisition

    NARCIS (Netherlands)

    van der Linden, E.; Hulk, A.

    2009-01-01

    The acquisition of gender has been reported to be problematic for some groups of learners acquiring Germanic or Romance languages. It has been shown that L1 learners do better in gender acquisition than others, like bilingual children, child L2 learners and adult L2 learners. The reason for these di

  17. Acylation of Quercetin with a Novel Thermophilic Esterase as Biocatalyst

    Institute of Scientific and Technical Information of China (English)

    XIE Xiao-na; ZHANG Chun-li; XUN Er-na; WANG Jia-xin; ZHANG Hong; WANG Lei; WANG Zhi

    2012-01-01

    The regioselective acylation of quercetin catalyzed by a novel thermophilic esterase(APE1547)from the archaeon Aeropyrum pernix K1 was successfully conducted in organic solvents.The effects of acyl donor,substrate ratio,organic solvent,temperature,and water activity were investigated.Under the optimum conditions,a yield of 74% for its mono ester could be achieved in the reaction for about 6 h.With the reaction time extending to about 30 h,the final conversion of quercetin was about 100% and three products were synthesized.

  18. Extraction systems for isolating esterases having interfacial adsorption

    Directory of Open Access Journals (Sweden)

    Alberto del Monte Martínez

    2009-06-01

    Full Text Available Resumen: En el presente trabajo se optimizaron las condiciones de extracción de esterasas con actividad en interfaces, a partir de la anémona marina Stichodactyla helianthus y del camarón peneido Litopenaeus vannamei. Las esterasas interfaciales, cuya presencia en estas especies había sido informada previamente, presentan características funcionales que las hacen muy atractivas para su empleo industrial. Los homogenados de los animales se trataron con los detergentes Tritón X-100, Tween 20 y Tween 80 en dos concentraciones cada uno: la Concentración Micelar Crítica (CMC y la mitad de ésta. Además se empleó NaCl 0,5 mol/L y n-butanol a las proporciones 5, 10 y 20%. Cada variante fue comparada con el método tradicional de extracción con agua destilada, que fue tomado como control. Los mejores resultados se obtuvieron empleando n-butanol al 20%, para recuperar las actividades esterasa y fosfolipasa, y al 10%, en el aislamiento de la actividad lipasa. La efectividad de este solvente en el aislamiento de estas enzimas con afinidad por las interfaces lípido/agua, pudiera estar dada por su capacidad para romper los agregados entre estas moléculas y causar la desorción de las mismas a los restos de membrana y tejidos presentes en la preparación.Palabras clave: activación interfacial, esterasas interfaciales, lipasas, Stichodactyla helianthus, Litopenaeus vannamei.interfacial activation, interfacial esterase, lipase, Stichodactyla helianthus, Litopenaeus vannamei.Abstract: Interfacial esterases present great functional versatility, making them very attractive molecules for industrial applications. The conditions for extracting interfacial esterases previously detected in the sea anemone Stichodactyla helianthus and the shrimp Litopenaeus vannamei were optimised in this work. Animal homogenates were treated with Triton X-100, Tween 20 and Tween 80 detergents at two different concentrations: critical micellar concentration (CMC and half

  19. Rendezvous missions with minimoons from L1

    Science.gov (United States)

    Chyba, M.; Haberkorn, T.; Patterson, G.

    2014-07-01

    We propose to present asteroid capture missions with the so-called minimoons. Minimoons are small asteroids that are temporarily captured objects on orbits in the Earth-Moon system. It has been suggested that, despite their small capture probability, at any time there are one or two meter diameter minimoons, and progressively greater numbers at smaller diameters. The minimoons orbits differ significantly from elliptical orbits which renders a rendezvous mission more challenging, however they offer many advantages for such missions that overcome this fact. First, they are already on geocentric orbits which results in short duration missions with low Delta-v, this translates in cost efficiency and low-risk targets. Second, beside their close proximity to Earth, an advantage is their small size since it provides us with the luxury to retrieve the entire asteroid and not only a sample of material. Accessing the interior structure of a near-Earth satellite in its morphological context is crucial to an in-depth analysis of the structure of the asteroid. Historically, 2006 RH120 is the only minimoon that has been detected but work is ongoing to determine which modifications to current observation facilities is necessary to provide detection algorithm capabilities. In the event that detection is successful, an efficient algorithm to produce a space mission to rendezvous with the detected minimoon is highly desirable to take advantage of this opportunity. This is the main focus of our work. For the design of the mission we propose the following. The spacecraft is first placed in hibernation on a Lissajoux orbit around the liberation point L1 of the Earth-Moon system. We focus on eight-shaped Lissajoux orbits to take advantage of the stability properties of their invariant manifolds for our transfers since the cost to minimize is the spacecraft fuel consumption. Once a minimoon has been detected we must choose a point on its orbit to rendezvous (in position and velocities

  20. 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

    been sequenced, its carbohydrate esterases are not annotated yet. We applied peptide pattern recognition (PPR) tool for sequence analysis of the C. thermophilum genome, and 11 carbohydrate esterase genes were discovered. Furthermore, we cloned and heterologously expressed two putative acetyl xylan...

  1. 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

    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...

  2. Organophosphate acetylcholine esterase inhibitor poisoning from a home-made shampoo.

    Science.gov (United States)

    Sadaka, Yair; Broides, Arnon; Tzion, Raffi Lev; Lifshitz, Matitiahu

    2011-07-01

    Organophosphate acetylcholine esterase inhibitor poisoning is a major health problem in children. We report an unusual cause of organophosphate acetylcholine esterase inhibitor poisoning. Two children were admitted to the pediatric intensive care unit due to organophosphate acetylcholine esterase inhibitor poisoning after exposure from a home-made shampoo that was used for the treatment of head lice. Owing to no obvious source of poisoning, the diagnosis of organophosphate acetylcholine esterase inhibitor poisoning in one of these patients was delayed. Both patients had an uneventful recovery. Organophosphate acetylcholine esterase inhibitor poisoning from home-made shampoo is possible. In cases where the mode of poisoning is unclear, direct questioning about the use of home-made shampoo is warranted, in these cases the skin and particularly the scalp should be rinsed thoroughly as soon as possible.

  3. Analysis list: Whsc1l1 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Whsc1l1 Blood + mm9 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Whsc1l1.1....tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Whsc1l1.5.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Whsc1...l1.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Whsc1l1.Blood.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Blood.gml ...

  4. PD-L1 Expression in Lung Cancer

    Science.gov (United States)

    Yu, Hui; Boyle, Theresa A.; Zhou, Caicun; Rimm, David L.; Hirsch, Fred R.

    2017-01-01

    Immunotherapies targeted against programmed death ligand 1 (PD-L1) and its receptor (PD-1) have improved survival in a subset of patients with advanced lung cancer. PD-L1 protein expression has emerged as a biomarker that predicts which patients are more likely to respond to immunotherapy. The understanding of PD-L1 as a biomarker is complicated by the history of use of different immunohistochemistry platforms with different PD-L1 antibodies, scoring systems, and positivity cut-offs for immunotherapy clinical trials with different anti-PD-L1 and anti-PD-1 drugs. Herein, we summarize the brief history of PD-L1 as a biomarker and describe the challenges remaining to harmonize PD-L1 detection and interpretation for best patient care. PMID:27117833

  5. 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.

  6. 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.

  7. Three multidomain esterases from the cellulolytic rumen anaerobe Ruminococcus flavefaciens 17 that carry divergent dockerin sequences.

    Science.gov (United States)

    Aurilia, V; Martin, J C; McCrae, S I; Scott, K P; Rincon, M T; Flint, H J

    2000-06-01

    Three enzymes carrying esterase domains have been identified in the rumen cellulolytic anaerobe Ruminococcus flavefaciens 17. The newly characterized CesA gene product (768 amino acids) includes an N-terminal acetylesterase domain and an unidentified C-terminal domain, while the previously characterized XynB enzyme (781 amino acids) includes an internal acetylesterase domain in addition to its N-terminal xylanase catalytic domain. A third gene, xynE, is predicted to encode a multidomain enzyme of 792 amino acids including a family 11 xylanase domain and a C-terminal esterase domain. The esterase domains from CesA and XynB share significant sequence identity (44%) and belong to carbohydrate esterase family 3; both domains are shown here to be capable of deacetylating acetylated xylans, but no evidence was found for ferulic acid esterase activity. The esterase domain of XynE, however, shares 42% amino acid identity with a family 1 phenolic acid esterase domain identified from Clostridum thermocellum XynZ. XynB, XynE and CesA all contain dockerin-like regions in addition to their catalytic domains, suggesting that these enzymes form part of a cellulosome-like multienzyme complex. The dockerin sequences of CesA and XynE differ significantly from those previously described in R. flavefaciens polysaccharidases, including XynB, suggesting that they might represent distinct dockerin specificities.

  8. Interactions of neuropathy inducers and potentiators/promoters with soluble esterases.

    Science.gov (United States)

    Estévez, Jorge; Mangas, Iris; Sogorb, Miguel Ángel; Vilanova, Eugenio

    2013-03-25

    Organophosphorus compounds (OPs) cause neurotoxic disorders through interactions with well-known target esterases, such as acetylcholinesterase and neuropathy target esterase (NTE). However, the OPs can potentially interact with other esterases of unknown significance. Therefore, identifying, characterizing and elucidating the nature and functional significance of the OP-sensitive pool of esterases in the central and peripheral nervous systems need to be investigated. Kinetic models have been developed and applied by considering multi-enzymatic systems, inhibition, spontaneous reactivation, the chemical hydrolysis of the inhibitor and "ongoing inhibition" (inhibition during the substrate reaction time). These models have been applied to discriminate enzymatic components among the esterases in nerve tissues of adult chicken, this being the experimental model for delayed neuropathy and to identify different modes of interactions between OPs and soluble brain esterases. The covalent interaction with the substrate catalytic site has been demonstrated by time-progressive inhibition during ongoing inhibition. The interaction of sequential exposure to an esterase inhibitor has been tested in brain soluble fraction where exposure to one inhibitor at a non inhibitory concentration has been seen to modify sensitivity to further exposure to others. The effect has been suggested to be caused by interaction with sites other than the inhibition site at the substrate catalytic site. This kind of interaction among esterase inhibitors should be considered to study the potentiation/promotion phenomenon, which is observed when some esterase inhibitors enhance the severity of the OP induced neuropathy if they are dosed after a non neuropathic low dose of a neuropathy inducer.

  9. 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

    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...... different esterase activity. Hence, the CE15 family is likely to comprise other enzyme functions than glucuronoyl esterase alone. Gene annotation in a variety of fungal and bacterial microorganisms showed that coprophilic fungi are rich and diverse sources of CE15 proteins. Combined with the lifestyle...

  10. β-Glucuronidase-coupled assays of glucuronoyl esterases.

    Science.gov (United States)

    Fraňová, Lucia; Puchart, Vladimír; Biely, Peter

    2016-10-01

    Glucuronoyl esterases (GEs) are microbial enzymes with potential to cleave the ester bonds between lignin alcohols and xylan-bound 4-O-methyl-d-glucuronic acid in plant cell walls. This activity renders GEs attractive research targets for biotechnological applications. One of the factors impeding the progress in GE research is the lack of suitable substrates. In this work, we report a facile preparation of methyl esters of chromogenic 4-nitrophenyl and 5-bromo-4-chloro-3-indolyl β-D-glucuronides for qualitative and quantitative GE assay coupled with β-glucuronidase as the auxiliary enzyme. The indolyl derivative affording a blue indigo-type product is suitable for rapid and sensitive assay of GE in commercial preparations as well as for high throughput screening of microorganisms and genomic and metagenomic libraries.

  11. Plasma B-esterase activities in European raptors.

    Science.gov (United States)

    Roy, Claudie; Grolleau, Gérard; Chamoulaud, Serge; Rivière, Jean-Louis

    2005-01-01

    B-esterases are serine hydrolases composed of cholinesterases, including acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), and carboxylesterase (CbE). These esterases, found in blood plasma, are inhibited by organophosphorus (OP) and carbamate (CB) insecticides and can be used as nondestructive biomarkers of exposure to anticholinesterase insecticides. Furthermore, B-esterases are involved in detoxification of these insecticides. In order to establish the level of these enzymes and to have reference values for their normal activities, total plasma cholinesterase (ChE), AChE and BChE activities, and plasma CbE activity were determined in 729 European raptors representing 20 species, four families, and two orders. The diurnal families of the Falconiforme order were represented by Accipitridae and Falconidae and the nocturnal families of the Strigiforme order by Tytonidae and Strigidae. Intraspecies differences in cholinesterase activities according to sex and/or age were investigated in buzzards (Buteo buteo), sparrowhawks (Accipiter nisus), kestrels (Falco tinnunculus), barn owls (Tyto alba), and tawny owls (Strix aluco). Sex-related differences affecting ChE and AChE activities were observed in young kestrels (2-3-mo-old) and age-related differences in kestrels (ChE and AChE), sparrowhawks (AChE), and tawny owls (ChE, AChE, and BChE). The interspecies analysis yielded a negative correlation between ChE activity and body mass taking into account the relative contribution of AChE and BChE to ChE activity, with the exception of the honey buzzard (Pernis apivorus). The lowest ChE activities were found in the two largest species, Bonelli's eagle (Hieraaetus fasciatus) and Egyptian vulture (Neophron percnopterus) belonging to the Accipitridae family. The highest ChE activities were found in the relatively small species belonging to the Tytonidae and Strigidae families and in honey buzzard of the Accipitridae family. Species of the Accipitridae, Tytonidae, and

  12. Phylogeny, classification and metagenomic bioprospecting of microbial acetyl xylan esterases.

    Science.gov (United States)

    Adesioye, Fiyinfoluwa A; Makhalanyane, Thulani P; Biely, Peter; Cowan, Don A

    2016-11-01

    Acetyl xylan esterases (AcXEs), also termed xylan deacetylases, are broad specificity Carbohydrate-Active Enzymes (CAZymes) that hydrolyse ester bonds to liberate acetic acid from acetylated hemicellulose (typically polymeric xylan and xylooligosaccharides). They belong to eight families within the Carbohydrate Esterase (CE) class of the CAZy database. AcXE classification is largely based on sequence-dependent phylogenetic relationships, supported in some instances with substrate specificity data. However, some sequence-based predictions of AcXE-encoding gene identity have proved to be functionally incorrect. Such ambiguities can lead to mis-assignment of genes and enzymes during sequence data-mining, reinforcing the necessity for the experimental confirmation of the functional properties of putative AcXE-encoding gene products. Although one-third of all characterized CEs within CAZy families 1-7 and 16 are AcXEs, there is a need to expand the sequence database in order to strengthen the link between AcXE gene sequence and specificity. Currently, most AcXEs are derived from a limited range of (mostly microbial) sources and have been identified via culture-based bioprospecting methods, restricting current knowledge of AcXEs to data from relatively few microbial species. More recently, the successful identification of AcXEs via genome and metagenome mining has emphasised the huge potential of culture-independent bioprospecting strategies. We note, however, that the functional metagenomics approach is still hampered by screening bottlenecks. The most relevant recent reviews of AcXEs have focused primarily on the biochemical and functional properties of these enzymes. In this review, we focus on AcXE phylogeny, classification and the future of metagenomic bioprospecting for novel AcXEs.

  13. The Facilitating Role of L1 in ESL Classes

    Directory of Open Access Journals (Sweden)

    Çağrı Tuğrul Mart

    2013-01-01

    Full Text Available It has been widely advocated that insistence on English-only policy and the complete prohibition of L1 would maximize the effect of learning L2 but recent studies demonstrate that the appropriate use of L1 has a facilitating role in L2 classroom. This study attempts to demonstrate that the use of L1 in ESL classes does not hinder foreign language learning.

  14. L1TV Computes the Flat Norm for Boundaries

    Directory of Open Access Journals (Sweden)

    Simon P. Morgan

    2007-01-01

    Full Text Available We show that the recently introduced L1TV functional can be used to explicitly compute the flat norm for codimension one boundaries. Furthermore, using L1TV, we also obtain the flat norm decomposition. Conversely, using the flat norm as the precise generalization of L1TV functional, we obtain a method for denoising nonboundary or higher codimension sets. The flat norm decomposition of differences can made to depend on scale using the flat norm with scale which we define in direct analogy to the L1TV functional. We illustrate the results and implications with examples and figures.

  15. On L1-convergence of Walsh-Fourier series

    Directory of Open Access Journals (Sweden)

    C. W. Onneweer

    1978-01-01

    Full Text Available Let G denote the dyadic group, which has as its dual group the Walsh(-Paley functions. In this paper we formulate a condition for functions in L1(G which implies that their Walsh-Fourier series converges in L1(G-norm. As a corollary we obtain a Dini-Lipschitz-type theorem for L1(G convergence and we prove that the assumption on the L1(G modulus of continuity in this theorem cannot be weakened. Similar results also hold for functions on the circle group T and their (trigonometric Fourier series.

  16. Insight into Substrate Preference of Two Chimeric Esterases by Combining Experiment and Molecular Simulation

    Institute of Scientific and Technical Information of China (English)

    ZHOU Xiao-li; HAN Wei-wei; ZHENG Bai-song; FENG Yan

    2013-01-01

    Better understanding of the relationship between the substrate preference and structural module of esterases is helpful to novel enzyme development.For this purpose,two chimeric esterases AAM7 and PAR,constructed via domain swapping between two ancient thermophilic esterases,were investigated on their molecular simulation(including homology modeling,substrates docking and substrate binding affinity validation) and enzymatic assay(specific activities and activation energies calculating).Our results indicate that the factors contributing to the substrate preference of many enzymes especially the broad-specificity enzymes like esterases are multiple and complicated,the substrate binding domains or binding pockets are important but not the only factor for substrate preference.

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

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    AIM: To investigate the diagnostic efficacy of leukocyte esterase and nitrite reagent strips for bedside diagnosis of spontaneous bacterial peritonitis (SBP).METHODS: 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 olymorphonuclear ascites count of ≥ 250/mm3.RESULTS: 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. CONCLUSION: Leukocyte esterase reagent strips may provide a rapid, bedside diagnostic test for SBP.

  18. Production, partial purification and characterization of feruloyl esterase by Aspergillus awamori in submerged fermentation.

    Science.gov (United States)

    Fazary, Ahmed Eid; Ju, Yi-Hsu

    2008-10-01

    Microbial feruloyl esterases acting on plant cell wall polymers represent key tools for the degradation of plant cell wall. In this paper, we describe in detail the microbial production, partial purification and characterization of feruloyl esterase from a culture medium of Aspergillus awamori strain IFO4033 obtained from a crude hemicellulose preparation of wheat straw, corncobs and wheat germ. Feruloyl esterase was extracted using centrifugation and dialysis, and then purified by ion exchange chromatography and microfiltration to homogeneity, which was checked by SDSPAGE and isoelectric focusing-PAGE. Protein content and activity of the enzyme were measured in each step of extraction and purification. Biomass was determined by the dry weight method. pH and temperature optima of feruloyl esterase enzyme were also determined. The effects of culturing time, and carbon and nitrogen sources on enzyme production were systematically investigated. Finally, enzyme activities under different storage conditions were examined.

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

    DEFF Research Database (Denmark)

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

    2012-01-01

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

  20. Esterase as molecular marker for salt tolerance in regenerated plants of rice, Oryza sativa L.

    Science.gov (United States)

    Swapna, T S

    2002-09-01

    Esterase variation was studied in plants regenerated from callus cultures of four rice (Oryza sativa) varieties, viz. pokkali, which is a moderately salt tolerant variety and three salt sensitive varieties MI 48, annapoorna and jyothi. Variation was studied at tillering stage of plants regenerated from callus culture and germinated from seeds. Somaclonal variants for salt tolerance could be detected using variation in esterase banding pattern and activity.

  1. Teachers' Language: L1 Attrition in Russian-English Bilinguals

    Science.gov (United States)

    Isurin, Ludmila

    2007-01-01

    The present study reports on the evidence of first language (L1) attrition in a population that may appear to be the most resistant to L1 changes. Russian monolinguals (n=3) and Russian-English bilinguals (n=10) participated in the study. The bilinguals were graduate students teaching Russian as a foreign language at a U.S. university. The data…

  2. ROBOTIC SYNTHESIS OF L-[1-C-11]TYROSINE

    NARCIS (Netherlands)

    LUURTSEMA, G; MEDEMA, J; ELSINGA, PH; VISSER, GM; VAALBURG, W

    1994-01-01

    L-[1-C-11]tyrosine promises to become an important tracer for determination of the protein synthesis rate (PSR) in tumor tissue and brain. The commercially available Anatech RB-86 robotic system is utilized for the automation of the L-[1-C-11]tyrosine production via the isocyanide method as reported

  3. Cell Divisions Are Required for L1 Retrotransposition▿

    Science.gov (United States)

    Shi, Xi; Seluanov, Andrei; Gorbunova, Vera

    2007-01-01

    LINE-1 (L1) retrotransposons comprise a large fraction of genomic DNAs of many organisms. Many L1 elements are active and may generate potentially deleterious mutations by inserting into genes, yet little is known about the control of retrotransposition by the host. Here we examined whether retrotransposition depends on the cell cycle by using a retrotransposition assay with cultured human cells. We show that in both cancer cells and primary human fibroblasts, retrotransposition was strongly inhibited in the cells arrested in the G1, S, G2, or M stage of the cell cycle. Retrotransposition was also inhibited during cellular senescence in primary human fibroblasts. The levels of L1 transcripts were strongly reduced in arrested cells, suggesting that the reduction in L1 transcript abundance limits retrotransposition in nondividing cells. We hypothesize that inhibition of retrotransposition in nondividing cells protects somatic tissues from accumulation of deleterious mutations caused by L1 elements. PMID:17145770

  4. Effect of non-enzymatic glycation on esterase activities of hemoglobin and myoglobin.

    Science.gov (United States)

    Sen, Subhrojit; Bose, Tania; Roy, Anjana; Chakraborti, Abhay Sankar

    2007-07-01

    Heme proteins--hemoglobin and myoglobin possess esterase activities. Studies with purified hemoglobin from normal individuals and diabetic patients revealed that the esterase activity as measured from hydrolysis of p-nitrophenyl acetate (p-NPA) was higher in diabetic condition and increased progressively with extent of the disease. HbA(1c), the major glycated hemoglobin, which increases proportionately with blood glucose level in diabetes mellitus, exhibited more esterase activity than the non-glycated hemoglobin fraction, HbA(0), as demonstrated spectrophotometrically as well as by activity staining. Glycation influenced esterase activity of hemoglobin by increasing the affinity for the substrate and the rate of the reaction. Both HbA(0) and HbA(1c)-mediated catalysis of p-NPA hydrolysis was pH-dependent. Esterase activity of in vitro-glycated myoglobin (GMb) was also higher than that of its non-glycated analog (Mb). The amplified esterase activities of hemoglobin and myoglobin might be associated with glycation-induced structural modifications of the proteins.

  5. Do L1 Reading Achievement and L1 Print Exposure Contribute to the Prediction of L2 Proficiency?

    Science.gov (United States)

    Sparks, Richard L.; Patton, Jon; Ganschow, Leonore; Humbach, Nancy

    2012-01-01

    The study examined whether individual differences in high school first language (L1) reading achievement and print exposure would account for unique variance in second language (L2) written (word decoding, spelling, writing, reading comprehension) and oral (listening/speaking) proficiency after adjusting for the effects of early L1 literacy and…

  6. 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

  7. Accessing PBeast and Monitoring the L1 Trigger

    CERN Document Server

    Thompson, Emily

    2015-01-01

    During the summer student program, I worked in the Trigger and Data Acquisition (TDAQ) group of ATLAS, more specifically in the Level 1 (L1) Trigger group. My first project was to design and implement a better way to access a file based system called 'PBeast', which stores all the conditions data of ATLAS. My second project was to improve the monitoring of the L1 trigger by making useful plots to display on the Central Trigger busy monitoring webpage. In this report, I will first briefly describe how the L1 trigger works. Then, I will describe the motivation for both of my projects and detail the progress I made this summer.

  8. 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.

  9. Hydrolysis of synthetic polyesters by Clostridium botulinum esterases.

    Science.gov (United States)

    Perz, Veronika; Baumschlager, Armin; Bleymaier, Klaus; Zitzenbacher, Sabine; Hromic, Altijana; Steinkellner, Georg; Pairitsch, Andris; Łyskowski, Andrzej; Gruber, Karl; Sinkel, Carsten; Küper, Ulf; Ribitsch, Doris; Guebitz, Georg M

    2016-05-01

    Two novel esterases from the anaerobe Clostridium botulinum ATCC 3502 (Cbotu_EstA and Cbotu_EstB) were expressed in Escherichia coli BL21-Gold(DE3) and were found to hydrolyze the polyester poly(butylene adipate-co-butylene terephthalate) (PBAT). The active site residues (triad Ser, Asp, His) are present in both enzymes at the same location only with some amino acid variations near the active site at the surrounding of aspartate. Yet, Cbotu_EstA showed higher kcat values on para-nitrophenyl butyrate and para-nitrophenyl acetate and was considerably more active (sixfold) on PBAT. The entrance to the active site of the modeled Cbotu_EstB appears more narrowed compared to the crystal structure of Cbotu_EstA and the N-terminus is shorter which could explain its lower activity on PBAT. The Cbotu_EstA crystal structure consists of two regions that may act as movable cap domains and a zinc metal binding site.

  10. TES/Aura L1B Spectra Limb V002

    Data.gov (United States)

    National Aeronautics and Space Administration — The L1B Limb granule consists of radiometrically calibrated spectra & associated NESR, observed at 0.025 cm-1 resolution for an entire Global Survey &...

  11. Nonmonotone Spectral Gradient Method for l_1-regularized Least Squares

    Directory of Open Access Journals (Sweden)

    Wanyou Cheng

    2016-08-01

    Full Text Available In the paper, we investigate a linear constraint optimization reformulation to a more general form of the l_1 regularization problem and give some good properties of it. We first show that the equivalence between the linear constraint optimization problem and the l_1 regularization problem. Second, the KKT point of the linear constraint problem always exists since the constraints are linear; we show that the half constraints must be active at any KKT point. In addition, we show that the KKT points of the linear constraint problem are the same as the stationary points of the l_1 regularization problem. Based on the linear constraint optimization problem, we propose a nonomotone spectral gradient method and establish its global convergence. Numerical experiments with compressive sense problems show that our approach is competitive with several known methods for standard l_2-l_1 problem.

  12. TES/Aura L1B Spectra Nadir V003

    Data.gov (United States)

    National Aeronautics and Space Administration — The L1B Nadir granule consists of radiometrically calibrated spectra & associated NESR, observed at 0.1 cm-1 resolution for an entire Global Survey &...

  13. TES/Aura L1B Spectra Nadir V004

    Data.gov (United States)

    National Aeronautics and Space Administration — The L1B Nadir granule consists of radiometrically calibrated spectra & associated NESR, observed at 0.1 cm-1 resolution for an entire Global Survey &...

  14. Phosphatidylcholine induces apoptosis of 3T3-L1 adipocytes

    Directory of Open Access Journals (Sweden)

    Li Hailan

    2011-12-01

    Full Text Available Abstract Background Phosphatidylcholine (PPC formulation is used for lipolytic injection, even though its mechanism of action is not well understood. Methods The viability of 3T3-L1 pre-adipocytes and differentiated 3T3-L1 cells was measured after treatment of PPC alone, its vehicle sodium deoxycholate (SD, and a PPC formulation. Western blot analysis was performed to examine PPC-induced signaling pathways. Results PPC, SD, and PPC formulation significantly decreased 3T3-L1 cell viability in a concentration-dependent manner. PPC alone was not cytotoxic to CCD-25Sk human fibroblasts at concentrations Conclusions PPC results in apoptosis of 3T3-L1 cells.

  15. 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.

  16. 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.

  17. The effect of surfactants on the aggregation behavior of phthalocyanine zinc (Ⅱ) bearing poly(aryl benzyl ether)dendritic substituents with carboxylic terminal%表面活性剂对以羧基为端基的芳基苄醚树枝配体取代酞菁锌(Ⅱ)配合物聚集行为的影响

    Institute of Scientific and Technical Information of China (English)

    陈婉玲; 彭亦如; 贺丹丹; 马冬冬; 张甜甜; 魏珍珍; 吴雪蓉

    2013-01-01

    通过紫外光谱法和荧光光谱法比较,研究了阳离子表面活性剂十六烷基三甲基溴化铵(CTAB)对0~2代以羧基为端基的芳基苄醚树枝配体取代酞菁锌(Ⅱ)配合物(ZnPc(COOH)4、G1-ZnPc(COOH)8和G2-ZnPc (COOH )16)聚集行为的影响。ZnPc (COOH)4、G1-ZnPc(COOH)8和 G2-ZnPc(COOH)16在水溶液中主要以二聚体形式存在。加入 CTAB 后, ZnPc (COOH )4、G1-ZnPc (COOH )8和 G2-ZnPc (COOH)16的单体吸收峰强度均增强,二聚体吸收峰强度均逐渐减弱;荧光光谱均明显增强,这表明 CTAB对树枝酞菁体系具有明显的解聚作用。这是因为 Zn-Pc (COOH )4、G1-ZnPc (COOH )8和 G2-ZnPc (COOH)16表面带负电荷的羧基与带正电荷的 CTAB通过静电作用形成纳米胶束,破坏了羧基酞菁聚集体之间的氢键。通过Zetasize粒度分析仪研究了不同浓度的CTAB与羧基酞菁形成纳米胶束的粒径分布情况,平均粒径范围约在5~30 nm,并随着 CTAB 浓度的增加,纳米胶束的平均粒径增大;随着树枝代数增加,纳米胶束的平均粒径逐渐减小。因此,阳离子表面活性剂CTAB可以有效抑制酞菁在水溶液中的聚集行为,在开展酞菁的光化学和物理研究方面具有很好的应用前景。%The interaction between the phthalocyanine zinc(Ⅱ)bearing poly(aryl benzyl ether)dendritic substit-uents with carboxylic terminal (ZnPc(COOH)4 ,G1-ZnPc(COOH)8 and G2-ZnPc(COOH)16 )and cationic sur-factants(cetyltrimethyl ammonium bromide(CTAB))were studied by fluorescence and UV-Vis spectroscopic methods.ZnPc(COOH)4 ,G1-ZnPc(COOH)8 and G2-ZnPc(COOH)16 mainly existed as a dimer at 625 nm in aqueous media,but they mainly exhibited as monomers at 685 nm with addition of CTAB.As the concentra-tion of CTAB increased,the intensity of the monomeric absorption peak increased,while that of the dimer peaks decreased gradually.Meanwhile,the fluorescence intensity of ZnPc(COOH)4 ,G1-ZnPc(COOH)8 and G2-ZnPc(COOH)16 also markedly enhanced

  18. Research into communication between L1DDC and FELIX

    CERN Document Server

    Keser, Marceline

    2016-01-01

    This report describes the summer student project of Marceline Keser, performed with the team for the New Small Wheel electronics upgrade of ATLAS. The main goal of the project is to create communication between the FELIX and L1DDC, in order to create a bidirectional data transmission. During this project, research has been done and has resulted in a GBT link between the L1DDC and FELIX.

  19. Gold nanoparticles functionalized with a fragment of the neural cell adhesion molecule L1 stimulate L1-mediated functions

    Science.gov (United States)

    Schulz, Florian; Lutz, David; Rusche, Norman; Bastús, Neus G.; Stieben, Martin; Höltig, Michael; Grüner, Florian; Weller, Horst; Schachner, Melitta; Vossmeyer, Tobias; Loers, Gabriele

    2013-10-01

    The neural cell adhesion molecule L1 is involved in nervous system development and promotes regeneration in animal models of acute and chronic injury of the adult nervous system. To translate these conducive functions into therapeutic approaches, a 22-mer peptide that encompasses a minimal and functional L1 sequence of the third fibronectin type III domain of murine L1 was identified and conjugated to gold nanoparticles (AuNPs) to obtain constructs that interact homophilically with the extracellular domain of L1 and trigger the cognate beneficial L1-mediated functions. Covalent conjugation was achieved by reacting mixtures of two cysteine-terminated forms of this L1 peptide and thiolated poly(ethylene) glycol (PEG) ligands (~2.1 kDa) with citrate stabilized AuNPs of two different sizes (~14 and 40 nm in diameter). By varying the ratio of the L1 peptide-PEG mixtures, an optimized layer composition was achieved that resulted in the expected homophilic interaction of the AuNPs. These AuNPs were stable as tested over a time period of 30 days in artificial cerebrospinal fluid and interacted with the extracellular domain of L1 on neurons and Schwann cells, as could be shown by using cells from wild-type and L1-deficient mice. In vitro, the L1-derivatized particles promoted neurite outgrowth and survival of neurons from the central and peripheral nervous system and stimulated Schwann cell process formation and proliferation. These observations raise the hope that, in combination with other therapeutic approaches, L1 peptide-functionalized AuNPs may become a useful tool to ameliorate the deficits resulting from acute and chronic injuries of the mammalian nervous system.The neural cell adhesion molecule L1 is involved in nervous system development and promotes regeneration in animal models of acute and chronic injury of the adult nervous system. To translate these conducive functions into therapeutic approaches, a 22-mer peptide that encompasses a minimal and functional L1

  20. Using L1 in Teaching English to Advanced Students

    Directory of Open Access Journals (Sweden)

    Khalid Al Hariri

    2015-11-01

    Full Text Available Using L1 in the English classroom has long been considered as an unforgivable wrongdoing. Many English teachers who use L1 in the classroom feel guilty about it and in many cases do not admit it. Recent researchers started to question the validity of this claim suggesting contexts and situations in which using L1 can be very helpful in ELT and refuting, on the other hand, the claims upon which the idea of using L2 solely were built.  My paper will examine the attitudes of advanced English language learners toward using L1 in the class of English where the students and the teacher speak the same first language and will also examine the attitudes of a group of EFL and ESL teachers toward the same practice. The same questionnaire given to the students will be redesigned and given to a number of teachers so that the study will look at the viewpoints from both perspectives. Keywords:  L1, Code Switching, Interference of L1, ELT, TESOL

  1. The Pleiotropic Role of L1CAM in Tumor Vasculature

    Science.gov (United States)

    Angiolini, Francesca; Cavallaro, Ugo

    2017-01-01

    Angiogenesis, the formation of new vessels, is a key step in the development, invasion, and dissemination of solid tumors and, therefore, represents a viable target in the context of antitumor therapy. Indeed, antiangiogenic approaches have given promising results in preclinical models and entered the clinical practice. However, in patients, the results obtained so far with antiangiogenic drugs have not completely fulfilled expectations, especially because their effect has been transient with tumors developing resistance and evasion mechanisms. A better understanding of the mechanisms that underlie tumor vascularization and the functional regulation of cancer vessels is a prerequisite for the development of novel and alternative antiangiogenic treatments. The L1 cell adhesion molecule (L1CAM), a cell surface glycoprotein previously implicated in the development and plasticity of the nervous system, is aberrantly expressed in the vasculature of various cancer types. L1CAM plays multiple pro-angiogenic roles in the endothelial cells of tumor-associated vessels, thus emerging as a potential therapeutic target. In addition, L1CAM prevents the maturation of cancer vasculature and its inhibition promotes vessel normalization, a process that is thought to improve the therapeutic response of tumors to cytotoxic drugs. We here provide an overview on tumor angiogenesis and antiangiogenic therapies and summarize the current knowledge on the biological role of L1CAM in cancer vasculature. Finally, we highlight the clinical implications of targeting L1CAM as a novel antiangiogenic and vessel-normalizing approach. PMID:28134764

  2. Dose L1 Retrotransposition Cause Neuronal Loss in Neurodegenerative Disorders?

    Directory of Open Access Journals (Sweden)

    Laleh Habibi

    2010-10-01

    Full Text Available "nNeurodegenerative disorders are among debilitating diseases that could affect many aspects of patient's life. Several mechanisms were shown to be involved in neuronal degeneration. However, the direct role of genomic instability is little considered in such disorders. L1 retrotransposons could cause genomic instability in different ways. Studies have shown increasing in L1 retrotransposition due to some reagents like heavy metals, stressors and the ones that may cause neuronal degeneration; Therefore cause cell to die. On the other hand, L1s retrotransposition was shown in neuronal precursor cells (NPCs providing the first evidence for movement of theses elements in nervous system."nHere, we propose that stimulation of L1 retrotransposition by environmental and genetic factors in neurons of central nervous system may lead them to apoptosis and result in neurodegenerative disorders. This hypothesis will be verified using L1-RP vector transfecting to definite neuronal cell line. By adding toxic agents including oxidative stress reagents and heavy metals to cell culture, we may track L1 retrotransposition and effects of this movement on cell physiology. Finding the involvement of mechanism in neurodegeneration may result in inventing new drugs for these disorders.

  3. 反胶束萃取阿魏酸酯酶%Purification of Feruloyl Esterase Using Reverse Micelles Extraction

    Institute of Scientific and Technical Information of China (English)

    王镇发; 陈培钦; 邓秩韬; 谢晨; 李夏兰

    2013-01-01

    利用十六烷基三甲基溴化铵(CTAB)/正己醇/异辛烷反胶束萃取阿魏酸酯酶.考察pH值、离子强度、CTAB浓度对阿魏酸酯酶的CTAB/正己醇/异辛烷反胶束萃取及其反萃取的影响.结果表明:萃取的最佳条件为酶液透析24 h,pH值为12,萃取液为25 mmol· L-1 CTAB的正己醇/异辛烷溶液(体积比为1∶5),萃取率接近100%;反萃取的最佳条件为pH值为7的0.20 mol·L-1 KCl溶液,反萃取率可达79%.纯化后比活为356.7μkat·g-1,纯化倍数为6.9,SDS-PAGE电泳显示两条带.%The cetrimonium bromide/hexanol/isooctane reverse micelles extraction and backward extraction were used to purify feruloyl esterase. The effects of pH, ionic strength and cetrimonium bromide (CTAB) concentration on the cetrimonium bromide/hexanol/isooctane reverse micelles extraction and backward extraction were studied. The results showed that the extraction efficiency was nearly 100% when the optimal extraction conditions were used as followed: enzyme dialysis time 24 h, pH 12 and CTAB concentration in hexanol/isooctane solution (volume ratio 1:5) 25 mmol · L-1. The optimal condition of backward extraction was 0. 20 mol · L-1 KCl with pH 7, and the efficiency of backward extraction was 79%. After extraction and backward extraction of the enzyme, specific activity was 356. 7 μkat · g-1, purification fold was 6. 9, and SDS-PAGE electrophoresis showed two bands.

  4. Distinction between esterases and lipases: a kinetic study with vinyl esters and TAG.

    Science.gov (United States)

    Chahinian, Henri; Nini, Lylia; Boitard, Elisabeth; Dubès, Jean-Paul; Comeau, Louis-Claude; Sarda, Louis

    2002-07-01

    The better to characterize enzymes hydrolyzing carboxyl ester bonds (carboxyl ester hydrolases), we have compared the kinetic behavior of various lipases and esterases against solutions and emulsions of vinyl esters and TAG. Short-chain vinyl esters are hydrolyzed at comparable rates by esterases and lipases and have higher limits of solubility in water than corresponding TAG. Therefore, they are suited to study the influence of the physical state of the substrate on carboxyl ester hydrolase activity within a large concentration range. Enzymes used in this study are TAG lipases from microorganisms, lipases from human and guinea pig pancreas, pig liver esterase, and acetylcholinesterase. This study also includes cutinase, a fungal enzyme that displays functional properties between esterases and lipases. Esterases display maximal activity against solutions of short-chain vinyl esters (vinyl acetate, vinyl propionate, and vinyl butyrate) and TAG (triacetin, tripropionin, and tributyrin). Half-maximal activity is reached at ester concentrations far below the solubility limit. The transition from solution to emulsion at substrate concentrations exceeding the solubility limit has no effect on esterase activity. Lipases are active on solutions of short-chain vinyl esters and TAG but, in contrast to esterases, they all display maximal activity against emulsified substrates and half-maximal activity is reached at substrate concentrations near the solubility limit of the esters. The kinetics of hydrolysis of soluble substrates by lipases are either hyperbolic or deviate from the Michaelis-Menten model and show no or weak interfacial activation. The presence of molecular aggregates in solutions of short-chain substrates, as evidenced by a spectral dye method, likely accounts for the activity of lipases against soluble esters. Unlike esterases, lipases hydrolyze emulsions of water-insoluble medium- and long-chain vinyl esters and TAG such as vinyl laurate, trioctanoin, and

  5. Biochemical characterization of a first fungal esterase from Rhizomucor miehei showing high efficiency of ester synthesis.

    Directory of Open Access Journals (Sweden)

    Yu Liu

    Full Text Available BACKGROUND: Esterases with excellent merits suitable for commercial use in ester production field are still insufficient. The aim of this research is to advance our understanding by seeking for more unusual esterases and revealing their characterizations for ester synthesis. METHODOLOGY/PRINCIPAL FINDINGS: A novel esterase-encoding gene from Rhizomucor miehei (RmEstA was cloned and expressed in Escherichia coli. Sequence analysis revealed a 975-bp ORF encoding a 324-amino-acid polypeptide belonging to the hormone-sensitive lipase (HSL family IV and showing highest similarity (44% to the Paenibacillus mucilaginosus esterase/lipase. Recombinant RmEstA was purified to homogeneity: it was 34 kDa by SDS-PAGE and showed optimal pH and temperature of 6.5 and 45°C, respectively. The enzyme was stable to 50°C, under a broad pH range (5.0-10.6. RmEstA exhibited broad substrate specificity toward p-nitrophenol esters and short-acyl-chain triglycerols, with highest activities (1,480 U mg(-1 and 228 U mg(-1 for p-nitrophenyl hexanoate and tributyrin, respectively. RmEstA efficiently synthesized butyl butyrate (92% conversion yield when immobilized on AOT-based organogel. CONCLUSION: RmEstA has great potential for industrial applications. RmEstA is the first reported esterase from Rhizomucor miehei.

  6. Activity of esterases from different tissues of freshwater fish and responses of their isoenzymes to inhibitors.

    Science.gov (United States)

    Li, S N; Fan, D F

    1997-06-06

    Activity of nonspecific esterase from different tissues (i.e., liver, gallbladder, heart, intestine, and muscle) of five species of freshwater fish, namely, topmouth gudgeon (Pseudorasbora parva), goldfish (Carassius auratus), nile tilapia (Tilapia nilotica), mosquitofish (Gambusia affinis), and rainbow trout (Salmo gairdneri) was tested using alpha-naphthyl acetate as substrate. The results indicated that activity of the enzyme was mainly concentrated in the digestive system (i.e., intestine, liver, bile). The overall activity was highest in nile tilapia, followed by mosquitofish, topmouth gudgeon, goldfish, and lowest in rainbow trout. Electrophoresis and the following in vitro treatment of the isoenzymes with triphenol phosphate (TPP, an inhibitor of carboxylesterase) indicated the TPP-sensitive esterase was mainly distributed in liver of the five species. The enzyme was not found in the other five tissues (including gill) except in gallbladder of topmouth gudgeon and goldfish. The correlation was obviously improved between susceptibility and detoxification capacity if activity of the TPP-sensitive esterase was employed instead of that of the nonspecific esterase to make the comparison. In vitro treatment of nonspecific esterase in liver with malaoxon proved that the active metabolite of malathion inhibited a different isoenzyme from the TPP-sensitive one.

  7. 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.

  8. Study on the production of acetyl esterase and side-group cleaving glycosidases of ammonia fungi.

    Science.gov (United States)

    Soponsathien, Siriphan

    1998-12-01

    Acetyl esterase was found to be widely distributed in ammonia fungi in a screen comprising 26 species (71 strains). No great differences appeared in enzyme production for acetyl esterase, beta-xylosidase, alpha-arabinosidase, or beta-glucosidase between different strains of the same species, but differences were detected between different genera. Acetyl esterase of Coprinus phlyctidosporus and Lyophyllum tylicolor may act cooperatively with beta-glucosidase. An increase in urea concentration significantly affected enzyme activity. It was supposed that urea used as 20 mg/g litter may solubilize leaf nutrients. At 20 mg urea added/g litter, a sizable increase in beta-glucosidase activity of C. phlyctidosporus and L. tylicolor was found, whereas a decrease in enzyme production of alpha-arabinosidase and beta-xylosidase was detected in some strains. Acetyl esterase and beta-glucosidase of C. phlyctidosporus, L. tylicolor, C. leucocephala 589, and C. rhombisperma 248 were most active in acidic conditions (pH 5.3-6.3), whereas acetyl esterase of L. nuda 561 and L. tarda 564 was most active in alkaline conditions (pH 8.3).

  9. Molecular cloning and characterization of a new and highly thermostable esterase from Geobacillus sp. JM6.

    Science.gov (United States)

    Zhu, Yanbing; Zheng, Wenguang; Ni, Hui; Liu, Han; Xiao, Anfeng; Cai, Huinong

    2015-10-01

    A new lipolytic enzyme gene was cloned from a thermophile Geobacillus sp. JM6. The gene contained 750 bp and encoded a 249-amino acid protein. The recombinant enzyme was expressed and purified from Escherichia coli BL21 (DE3) with a molecular mass of 33.6 kDa. Enzyme assays using p-nitrophenyl esters with different acyl chain lengths as the substrates confirmed its esterase activity, yielding the highest activity with p-nitrophenyl butyrate. When p-nitrophenyl butyrate was used as a substrate, the optimum reaction temperature and pH for the enzyme were 60 °C and pH 7.5, respectively. Geobacillus sp. JM6 esterase showed excellent thermostability with 68% residual activity after incubation at 100 °C for 18 h. A theoretical structural model of strain JM6 esterase was developed with a monoacylglycerol lipase from Bacillus sp. H-257 as a template. The predicted core structure exhibits an α/β hydrolase fold, and a putative catalytic triad (Ser97, Asp196, and His226) was identified. Inhibition assays with PMSF indicated that serine residue is involved in the catalytic activity of strain JM6 esterase. The recombinant esterase showed a relatively good tolerance to the detected detergents and denaturants, such as SDS, Chaps, Tween 20, Tween 80, Triton X-100, sodium deoxycholate, urea, and guanidine hydrochloride.

  10. Solid-state fermentation as a potential technique for esterase/lipase production by halophilic archaea.

    Science.gov (United States)

    Martin del Campo, Martha; Camacho, Rosa M; Mateos-Díaz, Juan C; Müller-Santos, Marcelo; Córdova, Jesus; Rodríguez, Jorge A

    2015-11-01

    Halophilic archaea are extremophiles, adapted to high-salt environments, showing a big biotechnological potential as enzyme, lipids and pigments producers. Four inert supports (perlite, vermiculite, polyurethane foam and glass fiber) were employed for solid-state fermentation (SSF) of the halophilic archaeon Natronococcus sp. TC6 to investigate biomass and esterase production. A very low esterase activity and high water activity were observed when perlite, vermiculite and polyurethane were used as supports. When glass fiber was employed, an important moisture loss was observed (8.6%). Moreover, moisture retention was improved by mixing polyurethane and glass fiber, resulting in maximal biomass and esterase production. Three halophilic archaea: Natronococcus sp. TC6, Halobacterium sp. NRC-1 and Haloarcula marismortui were cultured by submerged fermentation (SmF) and by SSF; an improvement of 1.3- to 6.2-fold was observed in the biomass and esterase production when SSF was used. Growth was not homogeneous in the mixture, but was predominant in the glass fiber thus was probably because the glass fiber provides a holder to the cells, while the polyurethane acts as an impregnation medium reservoir. To the best of our knowledge, this work is the first report on haloarchaea cultivation by SSF aiming biomass and esterase/lipase activity production.

  11. Isolation and characterization of novel lipases/esterases from a bovine rumen metagenome.

    Science.gov (United States)

    Privé, Florence; Newbold, C Jamie; Kaderbhai, Naheed N; Girdwood, Susan G; Golyshina, Olga V; Golyshin, Peter N; Scollan, Nigel D; Huws, Sharon A

    2015-07-01

    Improving the health beneficial fatty acid content of meat and milk is a major challenge requiring an increased understanding of rumen lipid metabolism. In this study, we isolated and characterized rumen bacterial lipases/esterases using functional metagenomics. Metagenomic libraries were constructed from DNA extracted from strained rumen fluid (SRF), solid-attached bacteria (SAB) and liquid-associated rumen bacteria (LAB), ligated into a fosmid vector and subsequently transformed into an Escherichia coli host. Fosmid libraries consisted of 7,744; 8,448; and 7,680 clones with an average insert size of 30 to 35 kbp for SRF, SAB and LAB, respectively. Transformants were screened on spirit blue agar plates containing tributyrin for lipase/esterase activity. Five SAB and four LAB clones exhibited lipolytic activity, and no positive clones were found in the SRF library. Fosmids from positive clones were pyrosequenced and twelve putative lipase/esterase genes and two phospholipase genes retrieved. Although the derived proteins clustered into diverse esterase and lipase families, a degree of novelty was seen, with homology ranging from 40 to 78% following BlastP searches. Isolated lipases/esterases exhibited activity against mostly short- to medium-chain substrates across a range of temperatures and pH. The function of these novel enzymes recovered in ruminal metabolism needs further investigation, alongside their potential industrial uses.

  12. Statistical Optimization of Medium Components for Improved Product Ion of Recombinant Hyperthermophilic Esterase

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The optimization of nutrient levels for the production of recombinant hyperthermophilic esterase by E. coli was carried out with response surface methodology(RSM) based on the central composite rotatable design(CCRD). A 24central composite rotatable design was used to study the combined effect of the nutritional constituents like yeast extract, peptone, mineral salt and trace metals. The P-value of the coefficient for the linear effect of peptone concentration was 0. 0081 and trace metals solution was less than 0. 0001, suggesting that these were the principal variables with significant effect on the hyperthermophilic esterase production. The predicted optimal hyperthermophilic esterase yield was 269. 17 U/mL, whereas an actual experimental value of 284. 58 U/mL was obtained.

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

    Science.gov (United States)

    Wu, Jindan; Cai, Guoqiang; Liu, Jinqiang; Ge, Huayun; Wang, Jiping

    2014-03-01

    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.

  14. Role of an esterase in flavor volatile variation within the tomato clade.

    Science.gov (United States)

    Goulet, Charles; Mageroy, Melissa H; Lam, Nghi B; Floystad, Abbye; Tieman, Denise M; Klee, Harry J

    2012-11-13

    Tomato flavor is dependent upon a complex mixture of volatiles including multiple acetate esters. Red-fruited species of the tomato clade accumulate a relatively low content of acetate esters in comparison with the green-fruited species. We show that the difference in volatile ester content between the red- and green-fruited species is associated with insertion of a retrotransposon adjacent to the most enzymatically active member of a family of esterases. This insertion causes higher expression of the esterase, resulting in the reduced levels of multiple esters that are negatively correlated with human preferences for tomato. The insertion was evolutionarily fixed in the red-fruited species, suggesting that high expression of the esterase and consequent low ester content may provide an adaptive advantage in the ancestor of the red-fruited species. These results illustrate at a molecular level how closely related species exhibit major differences in volatile production by altering a volatile-associated catabolic activity.

  15. 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. EDTA and Mn2+, on the other hand, activated the esterase activity in group II cells....

  16. Learners’ L1 Use in a Task-based Classroom

    DEFF Research Database (Denmark)

    Bao, Rui; Du, Xiangyun

    2015-01-01

    In the past two decades, strong theoretical and pedagogical arguments have been made advocating for task-based activities in the language-learning context. However, many teachers have been reluctant to in- corporate task-based activities into their teaching practices due to concerns about learners......, but with only a very small amount oc- curring for off-task talk across tasks. L1 use mainly occurred in learners’ efforts to mediate completion of the tasks. The findings highlight the role of L1 in foreign language learning and suggest that L1 use is associated with a number of factors, such as task types......, learners’ proficiency, and learning context. Implications for lan- guage teachers and task designers are also discussed....

  17. TV-L1 Optical Flow for Vector Valued Images

    DEFF Research Database (Denmark)

    Rakêt, Lars Lau; Roholm, Lars; Nielsen, Mads;

    2011-01-01

    a generalized algorithm that works on vector valued images, by means of a generalized projection step. We give examples of calculations of flows for a number of multi- dimensional constancy assumptions, e.g. gradient and RGB, and show how the developed methodology expands to any kind of vector valued images......The variational TV-L1 framework has become one of the most popular and successful approaches for calculating optical flow. One reason for the popularity is the very appealing properties of the two terms in the energy formulation of the problem, the robust L1-norm of the data fidelity term combined...... with the total variation (TV) regular- ization that smoothes the flow, but preserve strong discontinuities such as edges. Specifically the approach of Zach et al. [1] has provided a very clean and efficient algorithm for calculating TV-L1 optical flows between grayscale images. In this paper we propose...

  18. Parallel Coordinate Descent for L1-Regularized Loss Minimization

    CERN Document Server

    Bradley, Joseph K; Bickson, Danny; Guestrin, Carlos

    2011-01-01

    We propose Shotgun, a parallel coordinate descent algorithm for minimizing L1-regularized losses. Though coordinate descent seems inherently sequential, we prove convergence bounds for Shotgun which predict linear speedups, up to a problem-dependent limit. We present a comprehensive empirical study of Shotgun for Lasso and sparse logistic regression. Our theoretical predictions on the potential for parallelism closely match behavior on real data. Shotgun outperforms other published solvers on a range of large problems, proving to be one of the most scalable algorithms for L1.

  19. The Total Variation Regularized L1 Model for Multiscale Decomposition

    Science.gov (United States)

    2006-01-01

    N00014-03-1- 0514, and DOE Grant GE-FG01-92ER-25126. †Department of Industrial Engineering and Operations Research, Columbia University, New York, NY...5f. WORK UNIT NUMBER 7. PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) Columbia University,Department of Industrial Engineering and Operations...2r . THE TV-L1 MODEL FOR MULTISCALE DECOMPOSITION 5 In general the minimizer of the TV-L1 is nonunique . In the above disk example, if λ = 2/r

  20. Study of TAMe (p-tosyl-L-arginine methyl ester) esterase activity of bovine plasma.

    Science.gov (United States)

    Claxton, J; Black, W D; Gentry, P A

    1978-07-01

    The TAMe (p-tosyl-L-arginine methyl ester) esterase activity of mature and immature bovine plasma was studied and compared with the activity of this enzyme in human plasma. Kaolin activation of 2 minutes was required to produce maximal activation in cattle, as compared with 1 minute activation in man. The kaolin-activated TAMe esterase values in bovine plasma were approximately one-half the values found in human plasma. The activity of this enzyme was statistically greater in immature than in mature cattle (P less than 0.05) at kaolin activation times of 1, 2, 15, and 20 minutes.

  1. Molecular Basis for Stereospecific Hydrolysis of Ethyl Mandelate by Thermophilic Esterase

    Institute of Scientific and Technical Information of China (English)

    ZHANG Guo-yan; TAO Jin; ZHENG Liang-yu; CAO Shu-gui

    2011-01-01

    The stereospecific hydrolysis of mandelate can be effectively catalyzed by hyperthermophilic acylpeptide esterase APE 1547(Aeropyrum pernix esterase 1547).APE 1547 used in this reaction showed a remarkable stereodiscrimination in favour of R-mandelic acid(99% e.e.) with an enantiomeric ratio E>200.The results of computer simulation are consistent with the experimental results.It can be inferred that the R-substrate adopted a binding mode productive of the reaction due to the formation of the hydrogen bond at the active site of APE 1547.

  2. Lexical Bundles in L1 and L2 Academic Writing

    Science.gov (United States)

    Chen, Yu-Hua; Baker, Paul

    2010-01-01

    This paper adopts an automated frequency-driven approach to identify frequently-used word combinations (i.e., "lexical bundles") in academic writing. Lexical bundles retrieved from one corpus of published academic texts and two corpora of student academic writing (one L1, the other L2), were investigated both quantitatively and qualitatively.…

  3. On the L1 Attrition of the Spanish Present Tense

    Science.gov (United States)

    Cuza, Alejandro

    2010-01-01

    This study examines the potential native language (L1) attrition of the ongoing value of the Spanish present tense among long-term Spanish immigrants. Based on the assumption of second-language (L2) transfer and proposals on the permeability of interface-conditioned structures, it is hypothesized that long-term Spanish immigrants will show…

  4. Adaptation is Unnecessary in L1-"Adaptive" Control

    OpenAIRE

    2014-01-01

    We show in the paper that the, so--called, "new architecture of L_1-Adaptive Control" is, indeed, different from classical model reference adaptive control. Alas, it is not new, since it exactly coincides with a full--state feedback, linear time--invariant proportional plus integral controller (with a decaying additive disturbance).

  5. L1-L2 Sentence Translation in Classroom Grammar Tests

    Science.gov (United States)

    Salem, Ilana

    2012-01-01

    L1-L2 translation of separate sentences is one kind of task format used by mainstream EFL teachers to assess their learners' grammatical accuracy. Aimed at improving teacher-written translation items, this study analyses linguistic features potentially causing such decontextualized cues (and their target responses) to sound odd or untypical of…

  6. Greedy vs. L1 convex optimization in sparse coding

    DEFF Research Database (Denmark)

    Ren, Huamin; Pan, Hong; Olsen, Søren Ingvor;

    2015-01-01

    , such as face and action recognition, a comparative study of codes in abnormal event detection is less studied and hence no conclusion is gained on the effect of codes in detecting abnormalities. We constrict our comparison in two types of the above L0-norm solutions: greedy algorithms and convex L1-norm...

  7. Conceptualising the Potential Role of L1 in CLIL

    Science.gov (United States)

    Lin, Angel M. Y.

    2015-01-01

    Content and language integrated learning (CLIL) is a rapidly growing area of both research and practice in all parts of the world, especially in Europe and Asia. As a young discipline, CLIL has a good potential of distinguishing itself from monolingual L2 immersion education models by becoming more flexible and balanced about the role of L1 in…

  8. Distribution and substrate specificity of esterases in the housefly, Musca domestica L.

    NARCIS (Netherlands)

    Asperen, K. van

    1959-01-01

    Housefly homogenates perform high cholinesterase and ali-esterase activity. Warburg-manometric studies show that acetylcholine, acetyl-β-methylcholine, butyrylcholine, and benzoylcholine are exclusively hydrolysed by a cholinesterase, the properties of which are more or less comparable to those of t

  9. 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.

  10. 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...

  11. Purification, crystallization and preliminary X-ray analysis of an acetylxylan esterase from Bacillus pumilus.

    Science.gov (United States)

    Benini, S; Degrassi, G; Krastanova, I; Lamba, D; Venturi, V

    2001-12-01

    The gene encoding for acetylxylan esterase from Bacillus pumilus has been cloned and expressed in Escherichia coli. The recombinant protein has been purified to homogeneity and crystallized. The crystals obtained are of regular shape of dimensions 0.05 x 0.05 x 0.05 mm with R32 symmetry and diffract to 2.0 A using synchrotron radiation.

  12. 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.

  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 to...

  14. 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...

  15. Probing the enantioselectivity of Bacillus subtilis esterase BS2 for tert. alcohols

    NARCIS (Netherlands)

    Wiggers, Michiel; Holt, Jarle; Kourist, Robert; Bartsch, Sebastian; Arends, Isabel W. C. E.; Minnaard, Adriaan J.; Bornscheuer, Uwe T.; Hanefeld, Ulf

    2009-01-01

    The activity and enantioselectivity of several mutants of the esterase BS2 from Bacillus subtilis have been investigated. In the enzymatic hydrolysis of alpha,alpha-disubstituted cyanohydrin acetates, a class of tert. alcohol esters, they were active but not selective. In contrast to this result sim

  16. Performance and Applications of L1B2 Ultrasonic Motors

    OpenAIRE

    Gal Peled; Roman Yasinov; Nir Karasikov

    2016-01-01

    Piezoelectric ultrasonic motors offer important advantages for motion applications where high speed is coupled with high precision. The advances made in the recent decades in the field of ultrasonic motor based motion solutions allow the construction of complete motion platforms in the fields of semiconductors, aerospace and electro-optics. Among the various motor designs, the L1B2 motor type has been successful in industrial applications, offering high precision, effective control and operat...

  17. On $L^1$ extremal problem for entire functions

    CERN Document Server

    Yuditskii, Peter

    2012-01-01

    We generalized the Korkin-Zolotarev theorem to the case of entire functions having the smallest $L^1$ norm on a system of intervals $E$. If $\\bbC\\setminus E$ is a domain of Widom type with the Direct Cauchy Theorem we give an explicit formula for the minimal deviation. Important relations between the problem and the theory of canonical systems with reflectionless resolvent functions are shown.

  18. Molecular population genetics of the -esterase gene cluster of Drosophila melanogaster

    Indian Academy of Sciences (India)

    Evgeniy S. Balakirev; Francisco J. Ayala

    2003-12-01

    We have investigated nucleotide polymorphism at the -esterase gene cluster including the Est-6 gene and Est-6 putative pseudogene in four samples of Drosophila melanogaster derived from natural populations of southern Africa (Zimbabwe), Europe (Spain), North America (USA: California), and South America (Venezuela). A complex haplo-type structure is revealed in both Est-6 and Est-6. Total nucleotide diversity is twice in Est-6 as in Est-6; diversity is higher in the African sample than in the non-African ones. Strong linkage disequilibrium occurs within the -esterase gene cluster in non-African samples, but not in the African one. Intragenic gene conversion events are detected within Est-6 and, to a much greater extent, within Est-6; intergenic gene conversion events are rare. Tests of neutrality with recombination are significant for the -esterase gene cluster in the non-African samples but not significant in the African one. 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 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 degenerative destruction and reflecting possible functional interaction between the functional gene and the putative pseudogene. Est-6 and Est-6 may represent an indivisible intergenic complex (‘intergene’) in which each single component (Est-6 or Est-6) cannot separately carry out the full functional role.

  19. Serum cholesterol concentration associated with aspirin esterase activity in older people: preliminary data

    Directory of Open Access Journals (Sweden)

    Kazuhiko Kotani, Russell Caccavello, Ricardo Hermo, Toshiyuki Yamada, Nobuyuki Taniguchi, Alejandro Gugliucci

    2010-01-01

    Full Text Available OBJECTIVE: Metabolism of aspirin (acetylsalicylic acid, commonly used in older people for the prevention of cardiovascular disease, is important to the effectiveness of this drug. Whereas part of aspirin hydrolysis occurs in blood, there is a paucity of information in regards to circulating aspirin esterase activity in various physiological and pathological conditions. High aspirin esterase activity, corresponding to faster aspirin hydrolysis (thus aspirin non-responsiveness, may occur in cardiovascular disease-prone states. The objective of this study was to investigate the effects of cardio-metabolic variables such as cholesterol on serum aspirin esterase activity in older people who participated in an intervention study on physical activity. METHODS: A total of 18 non-medicated subjects (7 men/11 women, mean age 67.8 years, body mass index = 23.4 ± 3.3 kg/m2, who completed a 3-month interventional program for a mild-to-moderate increase in physical activity, were analyzed. The body mass index, plasma glucose, serum total cholesterol and aspirin esterase activity were measured in the pre- and post-interventional phases of the study. RESULTS: During the interventional period, the changes in aspirin esterase activity correlated significantly and positively with those of total cholesterol concentrations (r = 0.542, P = 0.020; β = 0.609, P = 0.035 in a multiple linear regression analysis after adjusting for all the measured variables. CONCLUSION: The results suggest that cholesterol metabolism alterations may be associated with aspirin metabolism in older people.

  20. Salmonella induces PD-L1 expression in B cells.

    Science.gov (United States)

    Lopez-Medina, Marcela; Perez-Lopez, Araceli; Alpuche-Aranda, Celia; Ortiz-Navarrete, Vianney

    2015-10-01

    Salmonella persists for a long time in B cells; however, the mechanism(s) through which infected B cells avoid effector CD8 T cell responses has not been characterized. In this study, we show that Salmonella infects and survives within all B1 and B2 cell subpopulations. B cells are infected with a Salmonella typhimurium strain expressing an ovalbumin (OVA) peptide (SIINFEKL) to evaluate whether B cells process and present Salmonella antigens in the context of MHC-I molecules. Our data showed that OVA peptides are presented by MHC class I K(b)-restricted molecules and the presented antigen is generated through proteasomal degradation and vacuolar processing. In addition, Salmonella-infected B cells express co-stimulatory molecules such as CD40, CD80, and CD86 as well as inhibitory molecules such as PD-L1. Thus, the cross-presentation of Salmonella antigens and the expression of activation molecules suggest that infected B cells are able to prime and activate specific CD8(+) T cells. However, the Salmonella infection-stimulated expression of PD-L1 suggests that the PD-1/PD-L1 pathway may be involved in turning off the cytotoxic effector response during Salmonella persistent infection, thereby allowing B cells to become a reservoir for the bacteria.

  1. L1 elements, processed pseudogenes and retrogenes in mammalian genomes.

    Science.gov (United States)

    Ding, Wenyong; Lin, Lin; Chen, Bing; Dai, Jianwu

    2006-12-01

    Long interspersed nuclear elements 1 (L1 elements or LINE1) are the most active autonomous retrotransposons in mammalian genomes. In addition to L1 elements themselves, other protein-coding mRNAs can also be reverse transcribed and integrated into the genome through the L1-mediated retrotransposition, leading to the formation of processed pseudogenes (PPs) and retrogenes, both of which are characterized by the lack of introns and the presence of a 3' polyA tract and flanking direct repeats. PPs are unable to encode a functional protein and have accumulated frameshift mutations and premature stop codons during evolution. A few of PPs are transcriptionally active. Retrogenes preserve undisrupted coding frames and are capable of encoding a functional protein that is identical or nearly identical to that of the progenitor gene. There is a significant excess of retrogenes that originate from the X chromosome and are retrotransposed into autosomes, and most of these retrogenes are specially expressed in male germ cells, suggesting the inactivation of X-linked genes during male meiosis provides a strong selection pressure on retrogenes originating from the X chromosome.

  2. Least-squares RTM with L1 norm regularisation

    Science.gov (United States)

    Wu, Di; Yao, Gang; Cao, Jingjie; Wang, Yanghua

    2016-10-01

    Reverse time migration (RTM), for imaging complex Earth models, is a reversal procedure of the forward modelling of seismic wavefields, and hence can be formulated as an inverse problem. The least-squares RTM method attempts to minimise the difference between the observed field data and the synthetic data generated by the migration image. It can reduce the artefacts in the images of a conventional RTM which uses an adjoint operator, instead of an inverse operator, for the migration. However, as the least-squares inversion provides an average solution with minimal variation, the resolution of the reflectivity image is compromised. This paper presents the least-squares RTM method with a model constraint defined by an L1-norm of the reflectivity image. For solving the least-squares RTM with L1 norm regularisation, the inversion is reformulated as a ‘basis pursuit de-noise (BPDN)’ problem, and is solved directly using an algorithm called ‘spectral projected gradient for L1 minimisation (SPGL1)’. Three numerical examples demonstrate the effectiveness of the method which can mitigate artefacts and produce clean images with significantly higher resolution than the least-squares RTM without such a constraint.

  3. Detection of L1, infectious virions and anti-L1 antibody in domestic rabbits infected with cottontail rabbit papillomavirus.

    Science.gov (United States)

    Hu, Jiafen; Budgeon, Lynn R; Cladel, Nancy M; Culp, Timothy D; Balogh, Karla K; Christensen, Neil D

    2007-12-01

    Shope papillomavirus or cottontail rabbit papillomavirus (CRPV) is one of the first small DNA tumour viruses to be characterized. Although the natural host for CRPV is the cottontail rabbit (Sylvilagus floridanus), CRPV can infect domestic laboratory rabbits (Oryctolagus cuniculus) and induce tumour outgrowth and cancer development. In previous studies, investigators attempted to passage CRPV in domestic rabbits, but achieved very limited success, leading to the suggestion that CRPV infection in domestic rabbits was abortive. The persistence of specific anti-L1 antibody in sera from rabbits infected with either virus or viral DNA led us to revisit the questions as to whether L1 and infectious CRPV can be produced in domestic rabbit tissues. We detected various levels of L1 protein in most papillomas from CRPV-infected rabbits using recently developed monoclonal antibodies. Sensitive in vitro infectivity assays additionally confirmed that extracts from these papillomas were infectious. These studies demonstrated that the CRPV/New Zealand White rabbit model could be used as an in vivo model to study natural virus infection and viral life cycle of CRPV and not be limited to studies on abortive infections.

  4. Improved enantioselectivity of thermostable esterase from Archaeoglobus fulgidus toward (S)-ketoprofen ethyl ester by directed evolution and characterization of mutant esterases.

    Science.gov (United States)

    Kim, Jinyeong; Kim, Seungbum; Yoon, Sangyoung; Hong, Eunsoo; Ryu, Yeonwoo

    2015-08-01

    Thermostable esterases have potential applications in various biotechnology industries because of their resistance to high temperature and organic solvents. In a previous study, we isolated an esterase from Archaeoglobus fulgidus DSM 4304 (Est-AF), which showed high thermostability but low enantioselectivity toward (S)-ketoprofen ethyl ester. (R)-ketoprofenor (S)-ketoprofenis produced by esterase hydrolysis of the ester bond of (R,S)-ketoprofen ethyl ester and (S)-ketoprofen has better pharmaceutical activity and lower side effects than (R)-ketoprofen. Therefore, we have generated mutants of Est-AF that retained high thermostability whilst improving enantioselectivity. A library of Est-AF mutants was created by error-prone polymerase chain reaction, and mutants with improved enantioselectivity were isolated by site-saturation mutagenesis. The regions of Est-AF containing amino acid mutations were analyzed by homology modeling of its three-dimensional structure, and structure-based explanations for the changes in enantioselectivity are proposed. Finally, we isolated two mutants showing improved enantioselectivity over Est-AF (ee% = -16.2 ± 0.2 and E = 0.7 ± 0.0): V138G (ee% = 35.9 ± 1.0 and E = 3.0 ± 0.1) and V138G/L200R (ee% = 89.2 ± 0.2 and E = 19.5 ± 0.5). We also investigated various characteristics of these mutants and found that the mutants showed similar thermostability and resistance to additives or organic solvents to Est-AF, without a significant trade-off between activity and stability.

  5. The FPGA based L1 track finding Tracklet approach

    Science.gov (United States)

    Kyriacou, Savvas; CMS Collaboration

    2017-01-01

    The High Luminosity upgraded LHC is expected to deliver proton-proton collisions per 25ns with an estimated 140-200 pile up interactions per bunch crossing. Ultrafast track finding is vital for handling trigger rates in such conditions. An FPGA based road search algorithm is developed, the Tracklet approach one of a few currently under consideration, for the CMS L1 trigger system. Based on low/high transverse momentum track discrimination and designed for the HL upgraded outer tracker, the algorithm achieves microsecond scale track reconstruction in the expected high track multiplicity environment. The Tracklet method overview, implementation, hardware demonstrator and performance results are presented and discussed.

  6. RENEWAL THEOREM FOR (L, 1)-RANDOM WALK IN RANDOM ENVIRONMENT

    Institute of Scientific and Technical Information of China (English)

    洪文明; 孙鸿雁

    2013-01-01

    We consider a random walk on Z in random environment with possible jumps{-L, · · · ,-1, 1}, in the case that the environment{ωi: i∈Z}are i.i.d.. We establish the renewal theorem for the Markov chain of “the environment viewed from the particle” in both annealed probability and quenched probability, which generalize partially the results of Kesten (1977) and Lalley (1986) for the nearest random walk in random environment on Z, respectively. Our method is based on the intrinsic branching structure within the (L, 1)-RWRE formulated in Hong and Wang (2013).

  7. 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.

  8. Balance of Activities of Alcohol Acetyltransferase and Esterase in Saccharomyces cerevisiae Is Important for Production of Isoamyl Acetate

    OpenAIRE

    Fukuda, Kiyoshi; Yamamoto, Nagi; Kiyokawa, Yoshifumi; Yanagiuchi, Toshiyasu; Wakai, Yoshinori; Kitamoto, Katsuhiko; Inoue, Yoshiharu; Kimura, Akira

    1998-01-01

    Isoamyl acetate is synthesized from isoamyl alcohol and acetyl coenzyme A by alcohol acetyltransferase (AATFase) in Saccharomyces cerevisiae and is hydrolyzed by esterases at the same time. We hypothesized that the balance of both enzyme activities was important for optimum production of isoamyl acetate in sake brewing. To test this hypothesis, we constructed yeast strains with different numbers of copies of the AATFase gene (ATF1) and the isoamyl acetate-hydrolyzing esterase gene (IAH1) and ...

  9. L1 track finding for a time multiplexed trigger

    Science.gov (United States)

    Cieri, D.; Brooke, J.; Grimes, M.; Newbold, D.; Harder, K.; Shepherd-Themistocleous, C.; Tomalin, I.; Vichoudis, P.; Reid, I.; Iles, G.; Hall, G.; James, T.; Pesaresi, M.; Rose, A.; Tapper, A.; Uchida, K.

    2016-07-01

    At the HL-LHC, proton bunches will cross each other every 25 ns, producing an average of 140 pp-collisions per bunch crossing. To operate in such an environment, the CMS experiment will need a L1 hardware trigger able to identify interesting events within a latency of 12.5 μs. The future L1 trigger will make use also of data coming from the silicon tracker to control the trigger rate. The architecture that will be used in future to process tracker data is still under discussion. One interesting proposal makes use of the Time Multiplexed Trigger concept, already implemented in the CMS calorimeter trigger for the Phase I trigger upgrade. The proposed track finding algorithm is based on the Hough Transform method. The algorithm has been tested using simulated pp-collision data. Results show a very good tracking efficiency. The algorithm will be demonstrated in hardware in the coming months using the MP7, which is a μTCA board with a powerful FPGA capable of handling data rates approaching 1 Tb/s.

  10. Cannabidiol promotes browning in 3T3-L1 adipocytes.

    Science.gov (United States)

    Parray, Hilal Ahmad; Yun, Jong Won

    2016-05-01

    Recruitment of the brown-like phenotype in white adipocytes (browning) and activation of existing brown adipocytes are currently being investigated as a means to combat obesity. Thus, a wide variety of dietary agents that contribute to browning of white adipocytes have been identified. The present study was designed to investigate the effects of cannabidiol (CBD), a major nonpsychotropic phytocannabinoid of Cannabis sativa, on induction of browning in 3T3-L1 adipocytes. CBD enhanced expression of a core set of brown fat-specific marker genes (Ucp1, Cited1, Tmem26, Prdm16, Cidea, Tbx1, Fgf21, and Pgc-1α) and proteins (UCP1, PRDM16, and PGC-1α). Increased expression of UCP1 and other brown fat-specific markers contributed to the browning of 3T3-L1 adipocytes possibly via activation of PPARγ and PI3K. In addition, CBD increased protein expression levels of CPT1, ACSL, SIRT1, and PLIN while down-regulating JNK2, SREBP1, and LPL. These data suggest possible roles for CBD in browning of white adipocytes, augmentation of lipolysis, thermogenesis, and reduction of lipogenesis. In conclusion, the current data suggest that CBD plays dual modulatory roles in the form of inducing the brown-like phenotype as well as promoting lipid metabolism. Thus, CBD may be explored as a potentially promising therapeutic agent for the prevention of obesity.

  11. Sparse SAR imaging based on L1/2 regularization

    Institute of Scientific and Technical Information of China (English)

    ZENG JinShan; FANG Jian; XU ZongBen

    2012-01-01

    In this paper,a novel method for synthetic aperture radar (SAR) imaging is proposed.The approach is based on L1/2 regularization to reconstruct the scattering field,which optimizes a quadratic error term of the SAR observation process subject to the interested scene sparsity. Compared to the conventional SAR imaging technique,the new method implements SAR imaging effectively at much lower sampling rate than the Nyquist rate,and produces high-quality images with reduced sidelobes and increased resolution. Also,over the prevalent greedy pursuit and L1 regularization based SAR imaging methods,there are remarkable performance improvements of the new method.On one hand,the new method significantly reduces the number of measurements needed for reconstruction,as supported by a phase transition diagram study.On the other hand,the new method is more robust to the observation noise.These fundamental properties of the new method are supported and demonstrated both by simulations and real SAR data experiments.

  12. Aspartame downregulates 3T3-L1 differentiation.

    Science.gov (United States)

    Pandurangan, Muthuraman; Park, Jeongeun; Kim, Eunjung

    2014-10-01

    Aspartame is an artificial sweetener used as an alternate for sugar in several foods and beverages. Since aspartame is 200 times sweeter than traditional sugar, it can give the same level of sweetness with less substance, which leads to lower-calorie food intake. There are reports that consumption of aspartame-containing products can help obese people lose weight. However, the potential role of aspartame in obesity is not clear. The present study investigated whether aspartame suppresses 3T3-L1 differentiation, by downregulating phosphorylated peroxisome proliferator-activated receptor γ (p-PPARγ), peroxisome proliferator-activated receptor γ (PPARγ), fatty acid-binding protein 4 (FABP4), CCAAT/enhancer-binding protein α (C/EBPα), and sterol regulatory element-binding protein 1 (SREBP1), which are critical for adipogenesis. The 3T3-L1 adipocytes were cultured and differentiated for 6 d in the absence and presence of 10 μg/ml of aspartame. Aspartame reduced lipid accumulation in differentiated adipocytes as evidenced by Oil Red O staining. qRT-PCR analysis showed that the PPARγ, FABP4, and C/EBPα mRNA expression was significantly reduced in the aspartame-treated adipocytes. Western blot analysis showed that the induction of p-PPARγ, PPARγ, SREBP1, and adipsin was markedly reduced in the aspartame-treated adipocytes. Taken together, these data suggest that aspartame may be a potent substance to alter adipocyte differentiation and control obesity.

  13. Acquisition of Japanese contracted sounds in L1 phonology

    Science.gov (United States)

    Tsurutani, Chiharu

    2002-05-01

    Japanese possesses a group of palatalized consonants, known to Japanese scholars as the contracted sounds, [CjV]. English learners of Japanese appear to treat them initially as consonant + glide clusters, where there is an equivalent [Cj] cluster in English, or otherwise tend to insert an epenthetic vowel [CVjV]. The acquisition of the Japanese contracted sounds by first language (L1) learners has not been widely studied compared with the consonant clusters in English with which they bear a close phonetic resemblance but have quite a different phonological status. This is a study to investigate the L1 acquisition process of the Japanese contracted sounds (a) in order to observe how the palatalization gesture is acquired in Japanese and (b) to investigate differences in the sound acquisition processes of first and second language (L2) learners: Japanese children compared with English learners. To do this, the productions of Japanese children ranging in age from 2.5 to 3.5 years were transcribed and the pattern of misproduction was observed.

  14. L1 track finding for a time multiplexed trigger

    Energy Technology Data Exchange (ETDEWEB)

    Cieri, D., E-mail: davide.cieri@bristol.ac.uk [University of Bristol, Bristol (United Kingdom); Rutherford Appleton Laboratory, Didcot (United Kingdom); Brooke, J.; Grimes, M. [University of Bristol, Bristol (United Kingdom); Newbold, D. [University of Bristol, Bristol (United Kingdom); Rutherford Appleton Laboratory, Didcot (United Kingdom); Harder, K.; Shepherd-Themistocleous, C.; Tomalin, I. [Rutherford Appleton Laboratory, Didcot (United Kingdom); Vichoudis, P. [CERN, Geneva (Switzerland); Reid, I. [Brunel University, London (United Kingdom); Iles, G.; Hall, G.; James, T.; Pesaresi, M.; Rose, A.; Tapper, A.; Uchida, K. [Imperial College, London (United Kingdom)

    2016-07-11

    At the HL-LHC, proton bunches will cross each other every 25 ns, producing an average of 140 pp-collisions per bunch crossing. To operate in such an environment, the CMS experiment will need a L1 hardware trigger able to identify interesting events within a latency of 12.5 μs. The future L1 trigger will make use also of data coming from the silicon tracker to control the trigger rate. The architecture that will be used in future to process tracker data is still under discussion. One interesting proposal makes use of the Time Multiplexed Trigger concept, already implemented in the CMS calorimeter trigger for the Phase I trigger upgrade. The proposed track finding algorithm is based on the Hough Transform method. The algorithm has been tested using simulated pp-collision data. Results show a very good tracking efficiency. The algorithm will be demonstrated in hardware in the coming months using the MP7, which is a μTCA board with a powerful FPGA capable of handling data rates approaching 1 Tb/s.

  15. Handling L2 Input in Phonological STM: The Effect of Non-L1 Phonetic Segments and Non-L1 Phonotactics on Nonword Repetition

    Science.gov (United States)

    Kovacs, Gabor; Racsmany, Mihaly

    2008-01-01

    This article reports on an experiment comparing the effects of three discrete types of deviance from native language (L1) phonetics and phonology on verbal short-term memory performance. A nonword repetition task was used to measure the recall of four stimulus types: (a) high-probability L1-sounding nonwords, (b) low-probability L1-sounding…

  16. Gene cloning and nucleotide sequencing and properties of a cocaine esterase from Rhodococcus sp. strain MB1.

    Science.gov (United States)

    Bresler, M M; Rosser, S J; Basran, A; Bruce, N C

    2000-03-01

    A strain of Rhodococcus designated MB1, which was capable of utilizing cocaine as a sole source of carbon and nitrogen for growth, was isolated from rhizosphere soil of the tropane alkaloid-producing plant Erythroxylum coca. A cocaine esterase was found to initiate degradation of cocaine, which was hydrolyzed to ecgonine methyl ester and benzoate; both of these esterolytic products were further metabolized by Rhodococcus sp. strain MB1. The structural gene encoding a cocaine esterase, designated cocE, was cloned from Rhodococcus sp. strain MB1 genomic libraries by screening recombinant strains of Rhodococcus erythropolis CW25 for growth on cocaine. The nucleotide sequence of cocE corresponded to an open reading frame of 1,724 bp that codes for a protein of 574 amino acids. The amino acid sequence of cocaine esterase has a region of similarity with the active serine consensus of X-prolyl dipeptidyl aminopeptidases, suggesting that the cocaine esterase is a serine esterase. The cocE coding sequence was subcloned into the pCFX1 expression plasmid and expressed in Escherichia coli. The recombinant cocaine esterase was purified to apparent homogeneity and was found to be monomeric, with an M(r) of approximately 65,000. The apparent K(m) of the enzyme (mean +/- standard deviation) for cocaine was measured as 1.33 +/- 0.085 mM. These findings are of potential use in the development of a linked assay for the detection of illicit cocaine.

  17. Theoretical studies of interaction models of human acetylcholine esterase with different inhibitors

    Institute of Scientific and Technical Information of China (English)

    ZHENG QingChuan; CHU HuiYing; NIU RuiJuan; SUN ChiaChung

    2009-01-01

    Alzheimer's disease (AD) is a progressive neurodegenerative disorder and one of the most common causes of dementia in the elderly.Acetyicholine esterase inhibitors (AChEl) are the main drugs used in the treatment of AD.In this work,docking studies have been performed in order to understand the interaction between a number of inhibitors (tacrine,rivastigmine,huperzine A,TV-3326 (ladostigil),donepezil and anseculin) and acetylcholine esterase (AChE).The calculated binding affinities between inhibitors and AChE increase in the order tacrine<rivastigmine<huperzine A<TV-3326<donepezil<anseculin,which reflects the experimental inhibitory activity expressed in terms of the half maximal inhibitory concentration (the IC50 value).Of the above inhibitors,anseculin is the most useful drug for the treatment of dementia.

  18. Theoretical studies of interaction models of human acetylcholine esterase with different inhibitors

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Alzheimer’s disease(AD) is a progressive neurodegenerative disorder and one of the most common causes of dementia in the elderly.Acetylcholine esterase inhibitors(AChEI) are the main drugs used in the treatment of AD.In this work,docking studies have been performed in order to understand the interaction between a number of inhibitors(tacrine,rivastigmine,huperzine A,TV-3326(ladostigil),donepezil and anseculin) and acetylcholine esterase(AChE).The calculated binding affinities between inhibitors and AChE increase in the order tacrine

  19. Pseudomonas aeruginosa biofilm growth inhibition on medical plastic materials by immobilized esterases and acylase.

    Science.gov (United States)

    Kisch, Johannes Martin; Utpatel, Christian; Hilterhaus, Lutz; Streit, Wolfgang R; Liese, Andreas

    2014-09-01

    Biofilms are matrix-encapsulated cell aggregates that cause problems in technical and health-related areas; for example, 65 % of all human infections are biofilm associated. This is mainly due to their ameliorated resistance against antimicrobials and immune systems. Pseudomonas aeruginosa, a biofilm-forming organism, is commonly responsible for nosocomial infections. Biofilm development is partly mediated by signal molecules, such as acyl-homoserine lactones (AHLs) in Gram-negative bacteria. We applied horse liver esterase, porcine kidney acylase, and porcine liver esterase; these can hydrolyze AHLs, thereby inhibiting biofilm formation. As biofilm infections are often related to foreign material introduced into the human body, we immobilized the enzymes on medical plastic materials. Biofilm formation was quantified by Crystal Violet staining and confocal laser scanning microscopy, revealing up to 97 % (on silicone), 54 % (on polyvinyl chloride), and 77 % (on polyurethane) reduced biomass after 68 h growth.

  20. Heterozygosity of the sheep: Polymorphism of 'malic enzyme', isocitrate dehydrogenase (NADP+), catalase and esterase.

    Science.gov (United States)

    Baker, C M; Manwell, C

    1977-04-01

    In contrast to other reports, it is found that the sheep has approximately as much enzyme variation as man. Most of the genetically interpretable enzyme variation in heart, liver, kidney and muscle from 52 sheep (Merinos or Merino crosses) is in the NADP-dependent dehydrogenases [two 'malic enzymes' and the supernatant isocitrate dehydrogenase (NADP+)] and in the esterases. Ten different loci for NAD-dependent dehydrogenases are electrophoretically monomorphic, as are five different NADH diaphorases from heart muscle and 15 different major proteins from skeletal muscle. It is highly statistically significant that NADP-dependent dehydrogenases and esterases are polymorphic but representatives of several other major classes of enzymes are not. The physiological significance of this polymorphism may be related to the role of these enzymes in growth and detoxication, sheep having been selected by man for faster growth, of wool or of carcass, and for grazing a wide variety of plants.

  1. 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...... was used to generate variants with deactivated nucleophilic elbows and the functional promiscuity was analyzed. In silico analysis together with enzymological characterization interestingly showed that each nucleophilic elbow formed a local active site with varied substrate specificities and affinities...

  2. Acute promyelocytic leukemia, hypogranular variant, with uncharacteristic staining with chloroacetate esterase.

    Science.gov (United States)

    Dunphy, C H; Polski, J M; Johns, G; Evans, H L; Gardner, L J

    2001-06-01

    A diagnosis of the hypogranular variant of acute promyelocytic leukemia (APLv) may be difficult to establish based on cytomorphology alone. However, the great majority of cases have a classical immunophenotype by flow cytometric immunophenotyping (FCI) (CD13+, CD33+, dim CD64+, HLA-DR-, and CD34-) and a classical enzyme cytochemical (EC) staining pattern. [intensely staining with myeloperoxidase, Sudan Black B, and chloroacetate esterase (CAE) and negative with alpha'-naphthyl acetate and butyrate esterases]. Although the immunophenotype of APLv by FCI has varied in the literature (HLA-DR +/- and CD34 +/-), the EC staining pattern has remained constant. We report a case of APLv with characteristic cytomorphology, compatible FCI data (CD13+, CD33+, dim CD64+, HLA-DR +/-, and CD34-), chromosomal detection of t(15; 17), and molecular detection of the PML/RAR alpha fusion gene; however, staining of the leukemic cells with CAE was quite uncharacteristic. We describe our findings.

  3. Regulation of polyisoprenylated methylated protein methyl esterase by polyunsaturated fatty acids and prostaglandins

    OpenAIRE

    Amissah, Felix; Taylor, Shalina; Duverna, Randolph; Ayuk-Takem, Lambert T.; Lamango, Nazarius S

    2011-01-01

    Polyisoprenylation is a set of secondary modifications involving proteins whose aberrant activities are implicated in cancers and degenerative disorders. The last step of the pathway involves an ester-forming polyisoprenylated protein methyl transferase- and hydrolytic polyisoprenylated methylated protein methyl esterase (PMPMEase)-catalyzed reactions. Omega-3 and omega-6 polyunsaturated fatty acids (PUFAs) have been linked with antitumorigeneis and tumorigenesis, respectively. PUFAs are stru...

  4. Exocellular esterase and emulsan release from the cell surface of Acinetobacter calcoaceticus.

    OpenAIRE

    Shabtai, Y; Gutnick, D L

    1985-01-01

    An esterase activity has been found, both in the cell-free growth medium and on the cell surface of the hydrocarbon-degrading Acinetobacter calcoaceticus RAG-1. The enzyme catalyzed the hydrolysis of acetyl and other acyl groups from triglycerides and aryl and alkyl esters. Emulsan, the extracellular heteropolysaccharide bioemulsifier produced by strain RAG-1, was also a substrate for the enzyme. Gel filtration showed that the cell-free enzyme was released from the cell surface either emulsan...

  5. 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.

  6. Mycobacteriocins produced by rapidly growing mycobacteria are Tween-hydrolyzing esterases.

    OpenAIRE

    Saito, H; Tomioka, H.; Watanabe, T; YONEYAMA, T.

    1983-01-01

    Smegmatocin, a protein produced by Mycobacterium smegmatis ATCC 14468, was found to have an esterase activity, hydrolyzing Tween 80, polyoxyethylene sorbitan monooleate, added to the assay medium for various "bacteriocins" from mycobacteria. Because M. diernhoferi ATCC 19340 (indicator strain for smegmatocin) is highly susceptible to oleic acid and smegmatocin requires Tween 80 for manifestation of its anti-M. diernhoferi activity, it is likely that smegmatocin-mediated antimicrobial action i...

  7. Purification of neuropathy target esterase from avian brain after prelabelling with [3H]diisopropyl phosphorofluoridate.

    Science.gov (United States)

    Rüffer-Turner, M E; Read, D J; Johnson, M K

    1992-01-01

    Neuropathy target esterase from hen brains was radiolabelled at the active site with [3H]diisopropyl phosphorofluoridate. The labelled protein was purified by differential centrifugation and Nonidet P40 solubilization, detergent phase partitioning, anion exchange, and preparative sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The volatilizable counts assay and analytical SDS-PAGE were used to monitor the protein. The 150-kDa subunit polypeptide appears as a single band on analytical SDS-PAGE.

  8. Characterization of patatin esterase activity in AOT-isooctane reverse micelles.

    Science.gov (United States)

    Jiménez, M; Escribano, J; Gandía-Herrero, F; Chazarra, S; Cabanes, J; García-Carmona, F; Pérez-Gilabert, M

    2002-01-01

    Patatin is a family of glycoproteins that accounts for 30-40% of the total soluble protein in potato (Solanum tuberosum L.) tubers. This protein has been reported not only to serve as a storage protein but also to exhibit lipid acyl hydrolase (LAH) activity. In this study patatin is characterized in AOT-isooctane reverse micelles. The influence on the enzymatic activity of characteristic parameters of reverse micelles, w(o) (= H(2)O/AOT), and the percentage of H(2)O, theta, were investigated. The results obtained show that patatin esterase activity varies with w(o) but remains constant throughout the range of theta values studied. The variation with w(o) showed that the activity follows an S-shaped behavior pattern, reaching a maximum at about w(o) = 20 for 2% H(2)O. Patatin esterase activity was compared with p-nitrophenyl (PNP) fatty acid esters of different chain lengths. The activity was much higher for PNP-caprylate. The pH optimum was 6.0, different from the value obtained when patatin esterase activity was measured in mixed micelle systems. The optimal temperature was 35 degrees C, above which the activity decreased to almost zero. The kinetic parameters were also evaluated (K(m) = 10 mM, V(m) = 158 microM/min, V(m)/K(m) = 15.8 x 10(-3) min(-1)). This paper shows the suitability of reverse micelles for measuring patatin esterase activity, since it allows the study of the enzyme in similar conditions to that prevailing in vivo.

  9. 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.

  10. Structural and functional profile of the carbohydrate esterase gene complement in Phytophthora infestans.

    Science.gov (United States)

    Ospina-Giraldo, Manuel D; McWalters, Jessica; Seyer, Lauren

    2010-12-01

    The plant cell cuticle is the first obstacle for penetration of the host by plant pathogens. To breach this barrier, most pathogenic fungi employ a complex assortment of cell wall-degrading enzymes including carbohydrate esterases, glycoside hydrolases, and polysaccharide lyases. We characterized the full complement of carbohydrate esterase-coding genes in three Phytophthora species and analyzed the expression of cutinase in vitro and in planta; we also determined the cutinase allele distribution in multiple isolates of P. infestans. Our investigations revealed that there are 49, 21, and 37 esterase homologs in the P. infestans, P. ramorum, and P. sojae genomes, respectively, with a considerable number predicted to be extracellular. Four cutinase gene copies were found in both the P. infestans and P. ramorum genomes, while 16 copies were found in P. sojae. Transcriptional analyses of cutinase in P. infestans revealed that its expression level during infection is significantly upregulated at all time points compared to that of the same gene in mycelium grown in vitro. Expression achieves maximum values at 15 hpi, declining at subsequent time points. These results may suggest, therefore, that cutinase most likely plays a role in P. infestans pathogenicity.

  11. 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.

  12. 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.

  13. B-esterase determination and organophosphate insecticide inhibitory effects in JEG-3 trophoblasts.

    Science.gov (United States)

    Espinoza, Marlon; Rivero Osimani, Valeria; Sánchez, Victoria; Rosenbaum, Enrique; Guiñazú, Natalia

    2016-04-01

    The placenta and trophoblasts express several B-esterases. This family includes acethylcholinesterase (AChE), carboxylesterase (CES) and butyrylcholinesterase (BChE), which are important targets of organophosphate insecticide (OP) toxicity. To better understand OP effects on trophoblasts, B-esterase basal activity and kinetic behavior were studied in JEG-3 choriocarcinoma cell cultures. Effects of the OP azinphos-methyl (Am) and chlorpyrifos (Cp) on cellular enzyme activity were also evaluated. JEG-3 cells showed measurable activity levels of AChE and CES, while BChE was undetected. Recorded Km for AChE and CES were 0.33 and 0.26 mM respectively. Native gel electrophoresis and RT-PCR analysis demonstrated CES1 and CES2 isoform expression. Cells exposed for 4 and 24 h to the OP Am or Cp, showed a differential CES and AChE inhibition profiles. Am inhibited CES and AChE at 4 h treatment while Cp showed the highest inhibition profile at 24 h. Interestingly, both insecticides differentially affected CES1 and CES2 activities. Results demonstrated that JEG-3 trophoblasts express AChE, CES1 and CES2. B-esterase enzymes were inhibited by in vitro OP exposure, indicating that JEG-3 cells metabolization capabilities include phase I enzymes, able to bioactivate OP. In addition, since CES enzymes are important for medicinal drug activation/deactivation, OP exposure may interfere with trophoblast CES metabolization, probably being relevant in a co-exposure scenario during pregnancy.

  14. 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.

  15. Conversion of a Rhizopus chinensis lipase into an esterase by lid swapping.

    Science.gov (United States)

    Yu, Xiao-Wei; Zhu, Shan-Shan; Xiao, Rong; Xu, Yan

    2014-06-01

    In an effort to explore the feasibility of converting a lipase into an esterase by modifying the lid region, we designed and characterized two novel Rhizopus chinensis lipase variants by lid swapping. The substrate specificity of an R. chinensis lipase was successfully modified toward water-soluble substrates, that is, turned into an esterase, by replacing the hydrophobic lid with a hydrophilic lid from ferulic acid esterase from Aspergillus niger Meanwhile, as a comparison, the lid of R. chinensis lipase was replaced by a hydrophobic lid from Rhizomucor miehei lipase, which did not alter its substrate specificity but led to a 5.4-fold higher catalytic efficiency (k*cat/K*m) toward p-nitrophenyl laurate. Based on the analysis of structure-function relationships, it suggests that the amphipathic nature of the lid is very important for the substrate specificity. This study provides new insight into the structural basis of lipase specificities and a way to tune the substrate preference of lipases.

  16. Cloning, expression and characterization of a feruloyl esterase C from Penicillium chrysogenum

    Institute of Scientific and Technical Information of China (English)

    LV Shan-shan; LI Gui-Iong; YANG Shao-Iong

    2016-01-01

    Objective: To clone feruloyl esterase gene C from Penicillium chrysogenum and characterize the general properties of the enzyme. Methods:The feruloyl esterase C gene was amplified by PCR based on the Penicillium chrysogenum feruloyl esterase C gene sequence and cloned into the expression vector pPIC9K, resulting the recombinant plasmid pPIC9K-PcfaeC. The recombiant plasmid was linerized and transformed into P. pastoris by electroporation. The transformants was screened based on the transparent zone technology. The screened transformants was then induced by methanol. the enzymatic properties of the protein were then measured. Results:SDS-PAGE analysis showed that the molecular mass of the enzyme was about 30 kD. The length of the gene was 762 bp. It comprised one open reading framwork(ORF) and annotated to encode 249 amino acid. The optimal temperature and pH was found to be 40℃and 6, respectively. Moreover, the recombinant enzyme was stable at 40-50℃and pH 5-7. Conclusion:The enzyme successfully expressed in P. pastoris could laid theoretical foundation in food, fodder and paper making industry.

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

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    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.

  18. Cloning, expression and characterization of a novel esterase from a South China Sea sediment metagenome

    Science.gov (United States)

    Zhang, Hao; Li, Fuchao; Chen, Huaxin; Zhao, Jin; Yan, Jinfei; Jiang, Peng; Li, Ronggui; Zhu, Baoli

    2015-07-01

    Lipolytic enzymes, including esterases and lipases, represent a group of hydrolases that catalyze the cleavage and formation of ester bonds. A novel esterase gene, scsEst01, was cloned from a South China Sea sediment metagenome. The scsEst01 gene consisted of 921 bp encoding 307 amino acid residues. The predicted amino acid sequence shared less than 90% identity with other lipolytic enzymes in the NCBI nonredundant protein database. ScsEst01 was successfully co-expressed in Escherichia coli BL21 (DE3) with chaperones (dnaK-dnaJ-grpE) to prevent the formation of inclusion bodies. The recombinant protein was purified on an immobilized metal ion affinity column containing chelating Sepharose charged with Ni2+. The enzyme was characterized using p -nitrophenol butyrate as a substrate. ScsEst01 had the highest lipolytic activity at 35°C and pH 8.0, indicative of a meso-thermophilic alkaline esterase. ScsEst01 was thermostable at 20°C. The lipolytic activity of scsEst01 was strongly increased by Fe2+, Mn2+ and 1% Tween 80 or Tween 20.

  19. Greedy vs. L1 Convex Optimization in Sparse Coding

    DEFF Research Database (Denmark)

    Ren, Huamin; Pan, Hong; Olsen, Søren Ingvor

    2015-01-01

    Sparse representation has been applied successfully in many image analysis applications, including abnormal event detection, in which a baseline is to learn a dictionary from the training data and detect anomalies from its sparse codes. During this procedure, sparse codes which can be achieved...... and action recognition, a comparative study of codes in abnormal event detection is less studied and hence no conclusion is gained on the effect of codes in detecting abnormalities. We constrict our comparison in two types of the above L0-norm solutions: greedy algorithms and convex L1-norm solutions....... Considering the property of abnormal event detection, i.e., only normal videos are used as training data due to practical reasons, effective codes in classification application may not perform well in abnormality detection. Therefore, we compare the sparse codes and comprehensively evaluate their performance...

  20. Exact recoverability from dense corrupted observations via $L_1$ minimization

    CERN Document Server

    Nguyen, Nam H

    2011-01-01

    This paper presents a surprising phenomenon: given $m$ highly corrupted measurements $y = A_{\\Omega \\bullet} x^{\\star} + e^{\\star}$, where $A_{\\Omega \\bullet}$ is a submatrix selected uniformly at random from an orthogonal matrix $A$ and $e^{\\star}$ is an unknown sparse error vector whose nonzero entries may be unbounded, we show that with high probability $l_1$-minimization can recover $x^{\\star}$ exactly from only $m = C \\mu^2 k (\\log n)^2$ where $k$ is the number of nonzero components of $x^{\\star}$ and $\\mu = n \\max_{ij} A_{ij}^2$, even if nearly $100 %$ measurements are corrupted. We further guarantee that stable recovery is possible when measurements are polluted by both gross sparse and small dense errors: $y = A_{\\Omega \\bullet} x^{\\star} + e^{\\star}+ \

  1. Performance and Applications of L1B2 Ultrasonic Motors

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    Gal Peled

    2016-06-01

    Full Text Available Piezoelectric ultrasonic motors offer important advantages for motion applications where high speed is coupled with high precision. The advances made in the recent decades in the field of ultrasonic motor based motion solutions allow the construction of complete motion platforms in the fields of semiconductors, aerospace and electro-optics. Among the various motor designs, the L1B2 motor type has been successful in industrial applications, offering high precision, effective control and operational robustness. This paper reviews the design of high precision motion solutions based on L1B2 ultrasonic motors—from the basic motor structure to the complete motion solution architecture, including motor drive and control, material considerations and performance envelope. The performance is demonstrated, via constructed motion stages, to exhibit fast move and settle, a repeatability window of tens of nanometers, lifetime into the tens of millions of operational cycles, and compatibility with clean room and aerospace environments. Example stages and modules for semiconductor, aerospace, electro-optical and biomedical applications are presented. The described semiconductor and aerospace solutions are powered by Nanomotion HR type motors, driven by a sine wave up to 80 V/mm rms, having a driving frequency of 39.6 kHz, providing a maximum force up to 4 N per driving element (at 5 W power consumption per element and a maximum linear velocity above 300 mm/s. The described electro-optical modules are powered by small Nanomotion Edge motors driven by voltages up to 11 V AC, providing stall forces up to 0.35 N (power consumption up to 0.75 W and maximum linear velocity above 200 mm/s.

  2. 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

  3. The degrees of freedom of penalized l1 minimization

    CERN Document Server

    Dossal, Charles; Fadili, Jalal M; Peyré, Gabriel; Chesneau, Christophe

    2011-01-01

    In this paper, we study the degrees of freedom (df) of penalized l1 minimization (also known as the Lasso) for linear regression models. We give a closed-form expression of the degrees of freedom of the Lasso response. Namely, we show that for any given Lasso regularization parameter \\lambda and any observed data y belongs to a set of full measure, the cardinal of the support of a particular solution of the Lasso problem is an unbiased estimator of the degrees of freedom of the Lasso response. This is achieved without any assumption on the uniqueness of the Lasso solution. Thus, our result remains true for both the underdetermined and the overdetermined case studied originally in Zou et al.. We also prove that a key result in Zou et al. is not true by providing a simple counterexample. An effective estimator of the number of degrees of freedom may have several applications including an objectively guided choice of the regularization parameter in the Lasso through the SURE framework.

  4. 微生物酯酶的研究进展%The Research Progress of Microbial Esterases

    Institute of Scientific and Technical Information of China (English)

    张敏文; 刘悦; 李荷

    2012-01-01

    微生物酯酶是一种广泛应用于食品、医药、精细化工等领域的工业化酶,特别是近年来随着手性化合物的深入研究,酯酶作为手性化合物拆分的高效催化剂,而微生物的来源又十分广泛,因此微生物酯酶成为研究热点.从组成及来源、产酯酶微生物的筛选、微生物酯酶的基因克隆以及微生物酯酶的应用等几方面对微生物酯酶进行综述,微生物酯酶已经在食品加工、精细化工、手性化合物拆分、环境治理中有所应用,随着研究的深入,酯酶的工业化生产及其在各领域的应用将会逐步实现.%Microbial Esterases play an important role in food industry, medicine industry, industry of fine chemicals and so on. In recent years, with thorough research of chiral compounds, esterase was a high-effective catalys for chiral resolution, and microbial sources is very extensive, so microbial esterases become research focus. This paper summarized many aspects in research progress of microbial esterases, including the composition and the source, screening of microbial esterase-producing strains, the gene cloning of microbial esterases, and the application of microbial esterases. Microbial esterase had already applied in food processing, fine chemical industry, chiral compounds and environmental treatment. With the development of research, the industrial production of microbial esterase and its application in various fields will gradually realize.

  5. Genome-wide analysis of esterase-like genes in the striped rice stem borer, Chilo suppressalis.

    Science.gov (United States)

    Wang, Baoju; Wang, Ying; Zhang, Yang; Han, Ping; Li, Fei; Han, Zhaojun

    2015-06-01

    The striped rice stem borer, Chilo suppressalis, a destructive pest of rice, has developed high levels of resistance to certain insecticides. Esterases are reported to be involved in insecticide resistance in several insects. Therefore, this study systematically analyzed esterase-like genes in C. suppressalis. Fifty-one esterase-like genes were identified in the draft genomic sequences of the species, and 20 cDNA sequences were derived which encoded full- or nearly full-length proteins. The putative esterase proteins derived from these full-length genes are overall highly diversified. However, key residues that are functionally important including the serine residue in the active site are conserved in 18 out of the 20 proteins. Phylogenetic analysis revealed that most of these genes have homologues in other lepidoptera insects. Genes CsuEst6, CsuEst10, CsuEst11, and CsuEst51 were induced by the insecticide triazophos, and genes CsuEst9, CsuEst11, CsuEst14, and CsuEst51 were induced by the insecticide chlorantraniliprole. Our results provide a foundation for future studies of insecticide resistance in C. suppressalis and for comparative research with esterase genes from other insect species.

  6. 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.

  7. AnL1 smoothing spline algorithm with cross validation

    Science.gov (United States)

    Bosworth, Ken W.; Lall, Upmanu

    1993-08-01

    We propose an algorithm for the computation ofL1 (LAD) smoothing splines in the spacesWM(D), with . We assume one is given data of the formyiD(f(ti) +ɛi, iD1,...,N with {itti}iD1N ⊂D, theɛi are errors withE(ɛi)D0, andf is assumed to be inWM. The LAD smoothing spline, for fixed smoothing parameterλ?;0, is defined as the solution,sλ, of the optimization problem (1/N)∑iD1N yi-g(ti +λJM(g), whereJM(g) is the seminorm consisting of the sum of the squaredL2 norms of theMth partial derivatives ofg. Such an LAD smoothing spline,sλ, would be expected to give robust smoothed estimates off in situations where theɛi are from a distribution with heavy tails. The solution to such a problem is a "thin plate spline" of known form. An algorithm for computingsλ is given which is based on considering a sequence of quadratic programming problems whose structure is guided by the optimality conditions for the above convex minimization problem, and which are solved readily, if a good initial point is available. The "data driven" selection of the smoothing parameter is achieved by minimizing aCV(λ) score of the form .The combined LAD-CV smoothing spline algorithm is a continuation scheme in λ↘0 taken on the above SQPs parametrized inλ, with the optimal smoothing parameter taken to be that value ofλ at which theCV(λ) score first begins to increase. The feasibility of constructing the LAD-CV smoothing spline is illustrated by an application to a problem in environment data interpretation.

  8. 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

  9. Antibody fragments directed against different portions of the human neural cell adhesion molecule L1 act as inhibitors or activators of L1 function.

    Directory of Open Access Journals (Sweden)

    Yan Wang

    Full Text Available The neural cell adhesion molecule L1 plays important roles in neuronal migration and survival, neuritogenesis and synaptogenesis. L1 has also been found in tumors of different origins, with levels of L1 expression correlating positively with the metastatic potential of tumors. To select antibodies targeting the varied functions of L1, we screened the Tomlinson library of recombinant human antibody fragments to identify antibodies binding to recombinant human L1 protein comprising the entire extracellular domain of human L1. We obtained four L1 binding single-chain variable fragment antibodies (scFvs, named I4, I6, I13, and I27 and showed by enzyme-linked immunosorbent assay (ELISA that scFvs I4 and I6 have high affinity to the immunoglobulin-like (Ig domains 1-4 of L1, while scFvs I13 and I27 bind strongly to the fibronectin type III homologous (Fn domains 1-3 of L1. Application of scFvs I4 and I6 to human SK-N-SH neuroblastoma cells reduced proliferation and transmigration of these cells. Treatment of SK-N-SH cells with scFvs I13 and I27 enhanced cell proliferation and migration, neurite outgrowth, and protected against the toxic effects of H(2O(2 by increasing the ratio of Bcl-2/Bax. In addition, scFvs I4 and I6 inhibited and scFvs I13 and I27 promoted phosphorylation of src and Erk. Our findings indicate that scFvs reacting with the immunoglobulin-like domains 1-4 inhibit L1 functions, whereas scFvs interacting with the fibronectin type III domains 1-3 trigger L1 functions of cultured neuroblastoma cells.

  10. Characterization of EST3: a metagenome-derived esterase with suitable properties for biotechnological applications.

    Science.gov (United States)

    Maester, Thaís Carvalho; Pereira, Mariana Rangel; Machado Sierra, E G; Balan, Andrea; de Macedo Lemos, Eliana Gertrudes

    2016-07-01

    Metagenomic libraries from diverse environments have been extensive sources of many lipases and esterases; nevertheless, most of these enzymes remain biochemically uncharacterized. We previously built a metagenomic fosmid library from a microbial consortium specialized for diesel oil degradation and tested it for lipolytic activity. In the present study, we identified the PL14.H10 clone that was subcloned and sequenced, which enabled the identification of the EST3 protein. This enzyme exhibited 74 % amino acid identity with the uncharacterized alpha/beta hydrolase from Parvibaculum lavamentivorans [GenBank: WP012110575.1] and was classified into lipolytic enzyme family IV. Biochemical characterization revealed that EST3 presents high activity in a wide range of temperature with highest activity from 41 to 45 °C. Also, this thermostable esterase acts from mild acidic to alkaline conditions with an optimum pH of 6.0. The enzyme exhibited activity against p-nitrophenyl esters of different chain lengths and highest catalytic efficiency against p-nitrophenyl caprylate. The activity of the protein was increased in the presence of 0.5 mM of Mn(+2), Li(+), EDTA, and 1 % of CTAB and exhibited half of the activity in the presence of 10 % methanol and ethanol. Moreover, the homology model of EST3 was built and compared to other esterases, revealing a substrate channel that should fit a wide range of substrates. Taken together, the data presented in this work reveal the unique and interesting characteristics of EST3 that might be explored for further use in biotechnological applications.

  11. Structural insights into the substrate specificity of two esterases from the thermophilic Rhizomucor miehei.

    Science.gov (United States)

    Yang, Shaoqing; Qin, Zhen; Duan, Xiaojie; Yan, Qiaojuan; Jiang, Zhengqiang

    2015-08-01

    Two hormone-sensitive lipase (HSL) family esterases (RmEstA and RmEstB) from the thermophilic fungus Rhizomucor miehei, exhibiting distinct substrate specificity, have been recently reported to show great potential in industrial applications. In this study, the crystal structures of RmEstA and RmEstB were determined at 2.15 Å and 2.43 Å resolutions, respectively. The structures of RmEstA and RmEstB showed two distinctive domains, a catalytic domain and a cap domain, with the classical α/β-hydrolase fold. Catalytic triads consisting of residues Ser161, Asp262, and His292 in RmEstA, and Ser164, Asp261, and His291 in RmEstB were found in the respective canonical positions. Structural comparison of RmEstA and RmEstB revealed that their distinct substrate specificity might be attributed to their different substrate-binding pockets. The aromatic amino acids Phe222 and Trp92, located in the center of the substrate-binding pocket of RmEstB, blocked this pocket, thus narrowing its catalytic range for substrates (C2-C8). Two mutants (F222A and W92F in RmEstB) showing higher catalytic activity toward long-chain substrates further confirmed the hypothesized interference. This is the first report of HSL family esterase structures from filamentous fungi. The information on structure-function relationships could open important avenues of exploration for further industrial applications of esterases.

  12. Expression of a fungal ferulic acid esterase in alfalfa modifies cell wall digestibility

    Science.gov (United States)

    2014-01-01

    Background Alfalfa (Medicago sativa) is an important forage crop in North America owing to its high biomass production, perennial nature and ability to fix nitrogen. Feruloyl esterase (EC 3.1.1.73) hydrolyzes ester linkages in plant cell walls and has the potential to further improve alfalfa as biomass for biofuel production. Results In this study, faeB [GenBank:AJ309807] was synthesized at GenScript and sub-cloned into a novel pEACH vector containing different signaling peptides to target type B ferulic acid esterase (FAEB) proteins to the apoplast, chloroplast, endoplasmic reticulum and vacuole. Four constructs harboring faeB were transiently expressed in Nicotiana leaves, with FAEB accumulating at high levels in all target sites, except chloroplast. Stable transformed lines of alfalfa were subsequently obtained using Agrobacterium tumefaciens (LBA4404). Out of 136 transgenic plants regenerated, 18 independent lines exhibited FAEB activity. Subsequent in vitro digestibility and Fourier transformed infrared spectroscopy (FTIR) analysis of FAEB-expressing lines showed that they possessed modified cell wall morphology and composition with a reduction in ester linkages and elevated lignin content. Consequently, they were more recalcitrant to digestion by mixed ruminal microorganisms. Interestingly, delignification by alkaline peroxide treatment followed by exposure to a commercial cellulase mixture resulted in higher glucose release from transgenic lines as compared to the control line. Conclusion Modifying cell wall crosslinking has the potential to lower recalcitrance of holocellulose, but also exhibited unintended consequences on alfalfa cell wall digestibility due to elevated lignin content. The combination of efficient delignification treatment (alkaline peroxide) and transgenic esterase activity complement each other towards efficient and effective digestion of transgenic lines. PMID:24650274

  13. NTE and non-NTE esterases in brain membrane: kinetic characterization with organophosphates.

    Science.gov (United States)

    Mangas, Iris; Vilanova, Eugenio; Estévez, Jorge

    2012-07-16

    Some effects of organophosphorus compounds (OPs) esters cannot be explained by action on currently recognized targets. In this work, we evaluate and characterize the interaction (inhibition, reactivation and "ongoing inhibition") of two model compounds: paraoxon (non-neuropathy-inducer) and mipafox (neuropathy-inducer), with esterases of chicken brain membranes, an animal model, tissue and fractions, where neuropathy target esterase (NTE) was first described and isolated. Four enzymatic components were discriminated. The relative sensitivity of time-progressive inhibition differed for paraoxon and mipafox. The most sensitive component for paraoxon was also the most sensitive component for mipafox (EPα: 4.4-8.3% of activity), with I(50) (30 min) of 15-43 nM with paraoxon and 29 nM with mipafox, and it spontaneously reactivated after inhibition with paraoxon. The second most sensitive component to paraoxon (EPβ: 38.3% of activity) had I(50) (30 min) of 1540 nM, and was practically resistant to mipafox. The third component (EPγ: 38.6-47.6% of activity) was paraoxon-resistant and sensitive to micromolar concentrations of mipafox; this component meets the operational criteria of being NTE (target of organophosphorus-induced delayed neuropathy). It had I(50) (30 min) of 5.3-6.6 μM with mipafox. The fourth component (EPδ: 9.8-10.7% of activity) was practically resistant to both inhibitors. Two paraoxon-resistant and mipafox-sensitive esterases were found using the sequential assay removing paraoxon, but only one was paraoxon-resistant and mipafox-sensitive according to the assay without removing paraoxon. We demonstrate that this apparent discrepancy, interpreted as reversible NTE inhibition with paraoxon, is the result of spontaneous reactivation after paraoxon inhibition of a non-NTE component. Some of these esterases' sensitivity to OPs suggests that they may play a role in toxicity in low-level exposure to organophosphate compounds or have a protective effect

  14. A novel alkaliphilic bacillus esterase belongs to the 13(th bacterial lipolytic enzyme family.

    Directory of Open Access Journals (Sweden)

    Lang Rao

    Full Text Available BACKGROUND: Microbial derived lipolytic hydrolysts are an important class of biocatalysts because of their huge abundance and ability to display bioactivities under extreme conditions. In spite of recent advances, our understanding of these enzymes remains rudimentary. The aim of our research is to advance our understanding by seeking for more unusual lipid hydrolysts and revealing their molecular structure and bioactivities. METHODOLOGY/PRINCIPAL FINDINGS: Bacillus. pseudofirmus OF4 is an extreme alkaliphile with tolerance of pH up to 11. In this work we successfully undertook a heterologous expression of a gene estof4 from the alkaliphilic B. pseudofirmus sp OF4. The recombinant protein called EstOF4 was purified into a homologous product by Ni-NTA affinity and gel filtration. The purified EstOF4 was active as dimer with the molecular weight of 64 KDa. It hydrolyzed a wide range of substrates including p-nitrophenyl esters (C2-C12 and triglycerides (C2-C6. Its optimal performance occurred at pH 8.5 and 50°C towards p-nitrophenyl caproate and triacetin. Sequence alignment revealed that EstOF4 shared 71% identity to esterase Est30 from Geobacillus stearothermophilus with a typical lipase pentapeptide motif G91LS93LG95. A structural model developed from homology modeling revealed that EstOF4 possessed a typical esterase 6α/7β hydrolase fold and a cap domain. Site-directed mutagenesis and inhibition studies confirmed the putative catalytic triad Ser93, Asp190 and His220. CONCLUSION: EstOF4 is a new bacterial esterase with a preference to short chain ester substrates. With a high sequence identity towards esterase Est30 and several others, EstOF4 was classified into the same bacterial lipolytic family, Family XIII. All the members in this family originate from the same bacterial genus, bacillus and display optimal activities from neutral pH to alkaline conditions with short and middle chain length substrates. However, with roughly 70% sequence

  15. Production of extracellular ferulic acid esterases by Lactobacillus strains using natural and synthetic carbon sources

    Directory of Open Access Journals (Sweden)

    Dominik Szwajgier

    2011-09-01

    Full Text Available Background. Ferulic acid esterases (FAE, EC 3.1.1.73, also known as feruloyl esterases, cinnamic acid esterases or cinnamoyl esterases, belong to a common group of hydrolases distributed in the plant kingdom. Especially the fungal enzymes were very well characterised in the past whereas the enzyme was rarely found in the lactic acid bacteria (LAB strains. It is well known that strong antioxidants free phenolic acids can be released from the dietary fiber by the action of intestinal microflora composed among others also of Lactobacillus strains. The aim of this study was to examine four Lactobacillus strains (L. acidophilus K1, L. rhamnosus E/N, PEN, OXYfor the ability to produce extracellular FAE on different synthetic and natural carbon sources. Material and methods. The LAB strains were grown in the minimal growth media using German wheat bran, rye bran, brewers’ spent grain, isolated larchwood arabinogalactan, apple pectin, corn pectin, methyl ferulate, methyl p-coumarate, methyl syringate or methyl vanillate as the sole carbon source. FAE activity was determined using the post-cultivation supernatants, methyl ferulate and HPLC with UV detection. Results. The highest FAE activity was obtained with L. acidophilus K1 and methyl ferulate (max. 23.34 ±0.05 activity units and methyl p-coumarate (max. 14.96 ±0.47 activity units as carbon sources. L. rhamnosus E/N, OXY and PEN exhibited the limited ability to produce FAE with cinnamic acids methyl esters. Methyl syringate and methyl vanillate (MS and MV were insufficient carbon sources for FAE production. Brewers’ spent grain was the most suitable substrate for FAE production by L. acidophilus K1 (max. 2.64 ±0.06 activity units and L. rhamnosus E/N, OXY and PEN. FAE was also successfully induced by natural substrates rye bran, corn pectin (L. acidophilus K1, German wheat bran and larchwood arabinogalactan (E/N, PEN or German wheat bran and corn pectin (OXY. Conclusions. This study proved the

  16. 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

    . 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...

  17. Balance of activities of alcohol acetyltransferase and esterase in Saccharomyces cerevisiae is important for production of isoamyl acetate.

    Science.gov (United States)

    Fukuda, K; Yamamoto, N; Kiyokawa, Y; Yanagiuchi, T; Wakai, Y; Kitamoto, K; Inoue, Y; Kimura, A

    1998-10-01

    Isoamyl acetate is synthesized from isoamyl alcohol and acetyl coenzyme A by alcohol acetyltransferase (AATFase) in Saccharomyces cerevisiae and is hydrolyzed by esterases at the same time. We hypothesized that the balance of both enzyme activities was important for optimum production of isoamyl acetate in sake brewing. To test this hypothesis, we constructed yeast strains with different numbers of copies of the AATFase gene (ATF1) and the isoamyl acetate-hydrolyzing esterase gene (IAH1) and used these strains in small-scale sake brewing. Fermentation profiles as well as components of the resulting sake were largely alike; however, the amount of isoamyl acetate in the sake increased with an increasing ratio of AATFase/Iah1p esterase activity. Therefore, we conclude that the balance of these two enzyme activities is important for isoamyl acetate accumulation in sake mash.

  18. The L2 Acquisition of Spanish Rhotics by L1 English Speakers: The Effect of L1 Articulatory Routines and Phonetic Context for Allophonic Variation

    Science.gov (United States)

    Olsen, Michael K.

    2012-01-01

    This article offers a fine-grained investigation of how first-language (L1) phonetics involving English rhotics affect Spanish rhotic production by second-language (L2) learners. Specifically, this study investigates how different L1 English rhotic articulatory routines (retroflex-like and bunched-like) and the phonetic context that produces…

  19. Do L2 Writing Courses Affect the Improvement of L1 Writing Skills via Skills Transfer from L2 to L1?

    Science.gov (United States)

    Gonca, Altmisdort

    2016-01-01

    This study investigates the relationship of second language (L2) writing skills proficiency with the first language (L1) writing skills, in light of the language transfer. The study aims to analyze the positive effects of L2 writing proficiency on L1 writing proficiency. Forty native Turkish-speaking university students participated in the study.…

  20. 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...

  1. Extensive transduction of nonrepetitive DNA mediated by L1 retrotransposition in cancer genomes

    NARCIS (Netherlands)

    J.M.C. Tubio (Jose M.); Y. Li (Yilong); Y.S. Ju (Young Seok); I. Martincorena (Inigo); S.L. Cooke (Susanna); M. Tojo (Marta); G. Gundem (Gunes); C.P. Pipinikas (Christodoulos); J. Zamora (Jorge); J.W. Raine (John); D. Menzies; P. Roman-Garcia (Pablo); A. Fullam (Anthony); M. Gerstung (Moritz); A. Shlien (Adam); P.S. Tarpey (Patrick); E. Papaemmanuil (Elli); S. Knappskog (Stian); P. van Loo (Peter); M. Ramakrishna (Manasa); H. Davies (Helen); J. Marshall (John); D.C. Wedge (David); J. Teague (Jon); A. Butler (Adam); S. Nik-Zainal (Serena); L.B. Alexandrov (Ludmil); S. Behjati (Sam); L.R. Yates (Lucy); N. Bolli (Niccolò); L. Mudie (Laura); C. Hardy (Claire); S. Martin (Sandra); S. McLaren (Stuart); S. O'Meara (Sarah); E. Anderson (Elizabeth); M. Maddison (Mark); S. Gamble (Stephen); C. Foster (Christopher); A.Y. Warren (Anne); H.J. Whitaker (Heather); D. Brewer (Daniel); R. Eeles (Rosalind); C. Cooper (Colin); D. Neal (David); A.G. Lynch (Andy); T. Visakorpi (Tapio); W.B. Isaacs (William); L.J. van 't Veer (Laura); C. Caldas (Carlos); C. Desmedt (Christine); C. Sotiriou (Christos); S. Aparicio (Sam); J.A. Foekens (John); J. Eyfjord; S. Lakhani (Sunil); G. Thomas (Gilles); O. Myklebost (Ola); P.N. Span (Paul); A.L. Børresen-Dale (Anne Lise); A.L. Richardson (Andrea); M.J. Vijver (Marc ); A. Vincent-Salomon (Anne); G.G. van den Eynden (Gert); A.M. Flanagan (Adrienne); P.A. Futreal (Andrew); H. Janes (Holly); G.S. Bova (G. Steven); M.R. Stratton (Michael); U. McDermott (Ultan); P.J. Campbell (Peter)

    2014-01-01

    textabstractLong interspersed nuclear element–1 (L1) retrotransposons are mobile repetitive elements that are abundant in the human genome. L1 elements propagate through RNA intermediates. In the germ line, neighboring, nonrepetitive sequences are occasionally mobilized by the L1 machinery, a proces

  2. English Teachers' Use of Learners' L1 (Arabic) in College Classrooms in Kuwait

    Science.gov (United States)

    Alrabah, Sulaiman; Wu, Shu-hua; Alotaibi, Abdullah M.; Aldaihani, Hussein A.

    2016-01-01

    This study investigated English teachers' use of learners' L1 (Arabic) in college classrooms in Kuwait. The purpose of the study was three-fold: (1) to describe the functions for which L1 was employed by the teachers, (2) to explore the affective, sociolinguistic, and psycholinguistic factors that may have led teachers to use L1 in L2 teaching,…

  3. The Influence of Schema and Cultural Difference on L1 and L2 Reading

    Science.gov (United States)

    Yang, Shi-sheng

    2010-01-01

    Reading in L1 shares numerous basic elements with reading in L2, and the processes also differ greatly. Intriguing questions involve whether there are two parallel cognitive processes at work, or whether there are processing strategies that accommodate both L1 and L2. This paper examines how reading in L1 is different from and similar to reading…

  4. PD-L1 expression in non-small cell lung cancer : Correlations with genetic alterations

    NARCIS (Netherlands)

    Scheel, Andreas H.; Ansen, Sascha; Schultheis, Anne M.; Scheffler, Matthias; Fischer, Rieke N.; Michels, Sebastian; Hellmich, Martin; George, Julie; Zander, Thomas; Brockmann, Michael; Stoelben, Erich; Groen, Harry; Timens, Wim; Perner, Sven; von Bergwelt-Baildon, Michael; Buettner, Reinhard; Wolf, Juergen

    2016-01-01

    Inhibition of the PD-1/PD-L1 pathway may induce anticancer immune responses in non-small cell lung cancer (NSCLC). Two PD-L1 immunohistochemistry (IHC) assays have been approved as companion diagnostic tests for therapeutic anti-PD-1 antibodies. However, many aspects of PD-L1 prevalence and associat

  5. PD-L1 expression by neurons nearby tumors indicates better prognosis in glioblastoma patients

    DEFF Research Database (Denmark)

    Liu, Yawei; Carlsson, Robert; Ambjørn, Malene;

    2013-01-01

    the functional consequences of neuronal Ifnb gene deletion on PD-L1 signaling and function. Ifnb-/- neurons lacked PD-L1 and were defective in inducing glioma cell death; this effect was reversed on PD-L1 gene transfection. Ifnb-/- mice with intracerebral isografts survived poorly. Similar to the observations...

  6. Does Reading in Shallow L1 Orthography Slow Attrition of Language-Specific Morphological Structures?

    Science.gov (United States)

    Zaretsky, Elena; Bar-Shalom, Eva G.

    2010-01-01

    This study looks at the relationship between L1 (Russian) attrition and L1 reading ability in Russian-English-speaking bilingual children. Ten Russian-English bilingual children and 10 adults participated in this study. Nine out of 10 children participants were born in the US and used L1 as their primary language of interaction within the family,…

  7. L1 use during L2 writing: an empirical study of a complex phenomenon

    NARCIS (Netherlands)

    D. van Weijen; H. van den Bergh; G. Rijlaarsdam; T. Sanders

    2009-01-01

    This study examined writers’ use of their first language (L1) while writing in their second language (L2). Twenty students each wrote four short argumentative essays in their L1 (Dutch) and four in their L2 (English) under think-aloud conditions. We analysed whether L1 use varied between writers and

  8. CALL versus Paper: In Which Context Are L1 Glosses More Effective?

    Science.gov (United States)

    Taylor, Alan M.

    2013-01-01

    CALL glossing in first language (L1) or second language (L2) texts has been shown by previous studies to be more effective than traditional, paper-and-pen L1 glossing. Using a pool of studies with much more statistical power and more accurate results, this meta-analysis demonstrates more precisely the degree to which CALL L1 glossing can be more…

  9. A Study of Relationships between L1 Pragmatic Transfer and L2 Proficiency

    Science.gov (United States)

    Bu, Jiemin

    2012-01-01

    Studies in interlanguage pragmatics have shown that L2 learners' proficiency has an influence on the occurrences of L1 pragmatic transfer. However, questions remain whether the relationship between L1 pragmatic transfer and L2 proficiency is positive or negative. This paper is designed to study L1 pragmatic transfer in requests made by Chinese…

  10. Identification of novel esterase-active enzymes from hot environments by use of the host bacterium Thermus thermophilus.

    Science.gov (United States)

    Leis, Benedikt; Angelov, Angel; Mientus, Markus; Li, Haijuan; Pham, Vu T T; Lauinger, Benjamin; Bongen, Patrick; Pietruszka, Jörg; Gonçalves, Luís G; Santos, Helena; Liebl, Wolfgang

    2015-01-01

    Functional metagenomic screening strategies, which are independent of known sequence information, can lead to the identification of truly novel genes and enzymes. Since E. coli has been used exhaustively for this purpose as a host, it is important to establish alternative expression hosts and to use them for functional metagenomic screening for new enzymes. In this study we show that Thermus thermophilus HB27 is an excellent screening host and can be used as an alternative provider of truly novel biocatalysts. In a previous study we constructed mutant strain BL03 with multiple markerless deletions in genes for major extra- and intracellular lipolytic activities. This esterase-diminished strain was no longer able to grow on defined minimal medium supplemented with tributyrin as the sole carbon source and could be used as a host to screen for metagenomic DNA fragments that could complement growth on tributyrin. Several thousand single fosmid clones from thermophilic metagenomic libraries from heated compost and hot spring water samples were subjected to a comparative screening for esterase activity in both T. thermophilus strain BL03 and E. coli EPI300. We scored a greater number of active esterase clones in the thermophilic bacterium than in the mesophilic E. coli. From several thousand functionally screened clones only two thermostable α/β-fold hydrolase enzymes with high amino acid sequence similarity to already characterized enzymes were identifiable in E. coli. In contrast, five further fosmids were found that conferred lipolytic activities in T. thermophilus only. Four open reading frames (ORFs) were found which did not share significant similarity to known esterase enzymes but contained the conserved GXSXG motif regularly found in lipolytic enzymes. Two of the genes were expressed in both hosts and the novel thermophilic esterases, which based on their primary structures could not be assigned to known esterase or lipase families, were purified and

  11. Est10: A Novel Alkaline Esterase Isolated from Bovine Rumen Belonging to the New Family XV of Lipolytic Enzymes

    Science.gov (United States)

    Rodríguez, María Cecilia; Loaces, Inés; Amarelle, Vanesa; Senatore, Daniella; Iriarte, Andrés; Fabiano, Elena; Noya, Francisco

    2015-01-01

    A metagenomic fosmid library from bovine rumen was used to identify clones with lipolytic activity. One positive clone was isolated. The gene responsible for the observed phenotype was identified by in vitro transposon mutagenesis and sequencing and was named est10. The 367 amino acids sequence harbors a signal peptide, the conserved secondary structure arrangement of alpha/beta hydrolases, and a GHSQG pentapeptide which is characteristic of esterases and lipases. Homology based 3D-modelling confirmed the conserved spatial orientation of the serine in a nucleophilic elbow. By sequence comparison, Est10 is related to hydrolases that are grouped into the non-specific Pfam family DUF3089 and to other characterized esterases that were recently classified into the new family XV of lipolytic enzymes. Est10 was heterologously expressed in Escherichia coli as a His-tagged fusion protein, purified and biochemically characterized. Est10 showed maximum activity towards C4 aliphatic chains and undetectable activity towards C10 and longer chains which prompted its classification as an esterase. However, it was able to efficiently catalyze the hydrolysis of aryl esters such as methyl phenylacetate and phenyl acetate. The optimum pH of this enzyme is 9.0, which is uncommon for esterases, and it exhibits an optimal temperature at 40°C. The activity of Est10 was inhibited by metal ions, detergents, chelating agents and additives. We have characterized an alkaline esterase produced by a still unidentified bacterium belonging to a recently proposed new family of esterases. PMID:25973851

  12. An L1-script-transfer-effect fallacy: a rejoinder to Wang et al. (2003).

    Science.gov (United States)

    Yamada, Jun

    2004-09-01

    Do different L1 (first language) writing systems differentially affect word identification in English as a second language (ESL)? Wang, Koda, and Perfetti [Cognition 87 (2003) 129] answered yes by examining Chinese students with a logographic L1 background and Korean students with an alphabetic L1 background for their phonological and orthographic processing skills on English word identification. Such a conclusion is premature, however. We propose that the L1 phonological system (rather than the L1 writing system) of the learner largely accounts for cognitive processes in learning to read a second language (L2).

  13. L^1 stability of conservation laws for a traffic flow model

    Directory of Open Access Journals (Sweden)

    Tong Li

    2001-02-01

    Full Text Available We establish the $L^1$ well-posedness theory for a system of nonlinear hyperbolic conservation laws with relaxation arising in traffic flows. In particular, we obtain the continuous dependence of the solution on its initial data in $L^1$ topology. We construct a functional for two solutions which is equivalent to the $L^1$ distance between the solutions. We prove that the functional decreases in time which yields the $L^1$ well-posedness of the Cauchy problem. We thus obtain the $L^1$-convergence to and the uniqueness of the zero relaxation limit.

  14. A dimeric structure of PD-L1: functional units or evolutionary relics?

    OpenAIRE

    Chen, Yong; Liu, Peipei; Gao, Feng; Cheng, Hao; Qi, Jianxun; Gao, George F

    2010-01-01

    PD-L1 is a member of the B7 protein family, most of whose members so far were identified as dimers in a solution and crystalline state, either complexed or uncomplexed with their ligand(s). The binding of PD-L1 with its receptor PD-1 (CD279) delivers an inhibitory signal regulating the T cell function. Simultaneously with the Garboczi group, we successfully solved another structure of human PD-L1 (hPD-L1). Our protein crystallized in the space group of C2221 with two hPD-L1 molecules per asym...

  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

    (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......Production of feruloyl esterases (FAEs) by Fusarium oxysporum was enhanced by optimization of initial pH of the culture medium, the type and concentration of nitrogen and carbon source. Submerged batch cultivation in a laboratory bioreactor (17 1) produced activity at 82 nkat g(-1) dry substrate....... Production of FAE does not therefore, require FA, however, production is diminished by the removal of esterified FA from the growth substrate. Optimal FAE activity was observed at pH 7 and 50 degreesC with 68 and 55% activity at pH 8 and pH 9, respectively. The esterase was fully stable at pH 5-8 and up...

  16. 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.

  17. Aspartyl proteinase, phospholipase, esterase and hemolysin activities of clinical isolates of the Candida parapsilosis species complex.

    Science.gov (United States)

    Treviño-Rangel, Rogelio de J; González, J Gerardo; González, Gloria M

    2013-04-01

    Candida parapsilosis is considered as an important emerging fungal pathogen and was recently found to be a complex that include three species, i.e., Candida parapsilosis sensu stricto, Candida orthopsilosis and Candida metapsilosis. The aim of this study was to determine the in vitro aspartyl proteinase, phospholipase, esterase and hemolysin activities of 65 clinical isolates of the C. parapsilosis complex, which had been previously identified by RFLP-BanI analysis. Of the enzymes evaluated, aspartyl proteinase was the least produced by the C. parapsilosis species complex. Phospholipase and esterase were strongly expressed by C. orthopsilosis (67% of isolates), while 10% and 13% of C. parapsilosis sensu stricto isolates were strong producers, respectively, of these two enzymes. In contrast, high production of both enzymes was not detected in C. metapsilosis. Hemolysin activity was significantly more abundant in C. orthopsilosis (87%) than C. parapsilosis sensu stricto (67%). Overall, C. orthopsilosis isolates were statistically associated with the production of hemolysins (P= 0.048) and phospholipases (Porthopsilosis and 20% of C. metapsilosis.

  18. Modelling substrate specificity and enantioselectivity for lipases and esterases by substrate-imprinted docking

    Directory of Open Access Journals (Sweden)

    Tyagi Sadhna

    2009-06-01

    Full Text Available Abstract Background Previously, ways to adapt docking programs that were developed for modelling inhibitor-receptor interaction have been explored. Two main issues were discussed. First, when trying to model catalysis a reaction intermediate of the substrate is expected to provide more valid information than the ground state of the substrate. Second, the incorporation of protein flexibility is essential for reliable predictions. Results Here we present a predictive and robust method to model substrate specificity and enantioselectivity of lipases and esterases that uses reaction intermediates and incorporates protein flexibility. Substrate-imprinted docking starts with covalent docking of reaction intermediates, followed by geometry optimisation of the resulting enzyme-substrate complex. After a second round of docking the same substrate into the geometry-optimised structures, productive poses are identified by geometric filter criteria and ranked by their docking scores. Substrate-imprinted docking was applied in order to model (i enantioselectivity of Candida antarctica lipase B and a W104A mutant, (ii enantioselectivity and substrate specificity of Candida rugosa lipase and Burkholderia cepacia lipase, and (iii substrate specificity of an acetyl- and a butyrylcholine esterase toward the substrates acetyl- and butyrylcholine. Conclusion The experimentally observed differences in selectivity and specificity of the enzymes were reproduced with an accuracy of 81%. The method was robust toward small differences in initial structures (different crystallisation conditions or a co-crystallised ligand, although large displacements of catalytic residues often resulted in substrate poses that did not pass the geometric filter criteria.

  19. Molecular cloning and characterization of a juvenile hormone esterase gene from brown planthopper, Nilaparvata lugens.

    Science.gov (United States)

    Liu, Shuhua; Yang, Baojun; Gu, Jianhua; Yao, Xiangmei; Zhang, Yixi; Song, Feng; Liu, Zewen

    2008-12-01

    Juvenile hormone (JH) plays key roles in the regulation of growth, development, diapause and reproduction in insects, and juvenile hormone esterase (JHE) plays an important role in regulating JH titers. We obtained a full-length cDNA encoding JHE in Nilaparvata lugens (NlJHE), the first JHE gene cloned from the hemipteran insects. The deduced protein sequence of Nljhe contains the five conserved motifs identified in JHEs of other insect species, including a consensus GQSAG motif that is required for the enzymatic activity of JHE proteins. Nljhe showed high amino acid similarities with Athalia rosae JHE (40%) and Apis mellifera JHE (39%). Recombinant NlJHE protein expressed in the baculovirus expression system hydrolyzed [3H] JH III at high activity and yielded the specificity constants (kcat/KM=4.28x10(6) M(-1) s(-1)) close to those of the validated JHEs from other insect species, indicating that Nljhe cDNA encodes a functional JH esterase. The Nljhe transcript was expressed mainly in the fat body and the expression level reached a peak at 48 h after ecdysis of the 5th instar nymphs. In the 5th instar, macropterous insects showed significantly higher Nljhe mRNA levels and JHE activities, but much lower JH III levels, than those detected in the brachypterous insects soon after ecdysis and at 48 h after ecdysis. These data suggest that NlJHE might play important roles in regulation of JH levels and wing form differentiation.

  20. Denaturing Effects of Urea and Guanidine Hydrochloride on Hyperthermophilic Esterase from Aeropyrum pernix K1

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The changes in the activity and the conformation of the hyperthermophilic esterase derived from aerobic thermophilic Aeropyrumpernix K1 (APE1547) were studied during denaturation by guanidine hydrochloride (GdnHCl)and urea. The denaturation course of APE1547 was followed by the steady-state and time resolved fluorescence methods. An increase in the denaturant concentration in the denatured system can significantly enhance the inactivation and unfolding of APE1547. The enzyme can be completely inactivated with a urea concentration of 2. 7 mol/L or a GdnHCl concentration of 7.5 mol/L. The fluorescence emission maximum of the enzyme protein red shifts in magnitude to a maximum value(355 nm) when the concentration of GdnHCl is 5.1 mol/L. The experimental results indicate that APE1547 has a high resistance to urea. Unfolding of APE1547 in GdnHCl(4.2-6.0 mol/L) was shown to be an irreversible process. The present results indicate that the ion pairs in this protein may be a key factor for the stability of this esterase.

  1. Diet quality determines lipase gene expression and lipase/esterase activity in Daphnia pulex

    Directory of Open Access Journals (Sweden)

    Apostolos-Manuel Koussoroplis

    2017-02-01

    Full Text Available We studied the short- (12 h and long-term (144 h response of Daphnia pulex lipases to quality shifts in diets consisting of different mixtures of the green alga Scenedesmus with the cyanobacterium Synechococcus, two species with contrasting lipid compositions. The lipase/esterase activity in both the gut and the body tissues had fast responses to the diet shift and increased with higher dietary contributions of Synechococcus. When screening the Daphnia genome for TAG lipases, we discovered a large gene-family expansion of these enzymes. We used a subset of eight genes for mRNA expression analyses and distinguished between influences of time and diet on the observed gene expression patterns. We identified five diet-responsive lipases of which three showed a sophisticated short- and long-term pattern of expression in response to small changes in food-quality. Furthermore, the gene expression of one of the lipases was strongly correlated to lipase/esterase activity in the gut suggesting its potentially major role in digestion. These findings demonstrate that the lipid-related enzymatic machinery of D. pulex is finely tuned to diet and might constitute an important mechanism of physiological adaptation in nutritionally complex environments.

  2. Crystallization and preliminary crystallographic studies of the metalloglycoprotein esterase A4 using a baculovirus expression system

    Energy Technology Data Exchange (ETDEWEB)

    Hiraki, Toshiki [Protein Design Laboratory, Yokohama City University, 1-7-29 Suehiro, Tsurumi, Yokohama 230-0045 (Japan); Shibayama, Naoya [Department of Physiology, Division of Biophysics, Jichi Medical University, 3311-1 Yakushiji, Shimotsuke, Tochigi 329-0498 (Japan); Yoon, Young-Ho [Protein Design Laboratory, Yokohama City University, 1-7-29 Suehiro, Tsurumi, Yokohama 230-0045 (Japan); Yun, Kyung-Mook [Department of Physiology, Division of Biophysics, Jichi Medical University, 3311-1 Yakushiji, Shimotsuke, Tochigi 329-0498 (Japan); Hamamoto, Toshiro [Department of Biochemistry, Jichi Medical University, 3311-1 Yakushiji, Shimotsuke, Tochigi 329-0498 (Japan); Tame, Jeremy R. H.; Park, Sam-Yong, E-mail: park@tsurumi.yokohama-cu.ac.jp [Protein Design Laboratory, Yokohama City University, 1-7-29 Suehiro, Tsurumi, Yokohama 230-0045 (Japan)

    2007-09-01

    Esterase A4 (EA4) is a timer protein found in diapause eggs of the silkworm Bombyx mori. The gene for this metalloglycoprotein was cloned from B. mori eggs and expressed using a baculovirus expression system in silkworm pupae. Crystals of the purified protein have been grown that diffract to beyond 2.1 Å resolution at 100 K using synchrotron radiation. Esterase A4 (EA4) is a timer protein found in diapause eggs of the silkworm Bombyx mori. The gene for this metalloglycoprotein was cloned from B. mori eggs and expressed using a baculovirus expression system in silkworm pupae. Crystals of the purified protein have been grown that diffract to beyond 2.1 Å resolution at 100 K using synchrotron radiation. The protein crystals belong to space group P2{sub 1}, with unit-cell parameters a = 47.1, b = 73.9, c = 47.4 Å, β = 104.1°. With one dimer per asymmetric unit, the crystal volume per unit protein weight (V{sub M}) is 2.3 Å{sup 3} Da{sup −1} and the solvent content is 47%.

  3. 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.

  4. An Esterase from Anaerobic Clostridium hathewayi Can Hydrolyze Aliphatic-Aromatic Polyesters.

    Science.gov (United States)

    Perz, Veronika; Hromic, Altijana; Baumschlager, Armin; Steinkellner, Georg; Pavkov-Keller, Tea; Gruber, Karl; Bleymaier, Klaus; Zitzenbacher, Sabine; Zankel, Armin; Mayrhofer, Claudia; Sinkel, Carsten; Kueper, Ulf; Schlegel, Katharina; Ribitsch, Doris; Guebitz, Georg M

    2016-03-15

    Recently, a variety of biodegradable polymers have been developed as alternatives to recalcitrant materials. Although many studies on polyester biodegradability have focused on aerobic environments, there is much less known on biodegradation of polyesters in natural and artificial anaerobic habitats. Consequently, the potential of anaerobic biogas sludge to hydrolyze the synthetic compostable polyester PBAT (poly(butylene adipate-co-butylene terephthalate) was evaluated in this study. On the basis of reverse-phase high-performance liquid chromatography (RP-HPLC) analysis, accumulation of terephthalic acid (Ta) was observed in all anaerobic batches within the first 14 days. Thereafter, a decline of Ta was observed, which occurred presumably due to consumption by the microbial population. The esterase Chath_Est1 from the anaerobic risk 1 strain Clostridium hathewayi DSM-13479 was found to hydrolyze PBAT. Detailed characterization of this esterase including elucidation of the crystal structure was performed. The crystal structure indicates that Chath_Est1 belongs to the α/β-hydrolases family. This study gives a clear hint that also micro-organisms in anaerobic habitats can degrade manmade PBAT.

  5. [Optimized expression of the L1 protein of human papillomavirus in Hansenula polymorpha].

    Science.gov (United States)

    Li, Weiwei; He, Xiuping; Guo, Xuena; Zhang, Zhenying; Zhang, Borun

    2009-10-01

    The heterologously expressed L1 protein of human papilomavirus 16 can assembly into virus-like particles (VLPs), which has been used as prophylactic vaccine for cervical carcinoma. To express L1 protein in Hansenula polymorpha, we analyzed the codon usage of the native gene of L1 protein and redesigned the encoding sequence according to the codon bias of H. polymorpha. We used assembly PCR to synthesize the native gene HPV16L1-N and the codon optimized gene HPV16L1. The synthesized genes were cloned into pMOXZa-A vector to generate plasmids pMOXZ-HPV16N and pMOXZ-HPV16. The expression cassettes MOXp-HPV16L1(N)-AOXTT were cloned into YEp352 vector and transferred into H. polymorpha. After methanol inducement, the expression of L1 protein in H. polymorpha was detected from the codon optimized gene HPV16L1 rather than the native gene HPVI6L1-N. The parameters for induced cultivation for strain HP-U-16L with HPV16L1 were investigated in shaking flask cultures. After induced cultivation in YPM (pH 7.0) medium supplemented with methanol to a final concentration of 1.0% every 12 h at 37 degrees C for 72 h, the recombinant produced 78.6 mg/L of L1 protein. This work offers the possibility for the production of prophylactic vaccine for cervical carcinoma by H. polymorpha.

  6. Effects of ethanol and NAP on cerebellar expression of the neural cell adhesion molecule L1.

    Directory of Open Access Journals (Sweden)

    Devon M Fitzgerald

    Full Text Available The neural cell adhesion molecule L1 is critical for brain development and plays a role in learning and memory in the adult. Ethanol inhibits L1-mediated cell adhesion and neurite outgrowth in cerebellar granule neurons (CGNs, and these actions might underlie the cerebellar dysmorphology of fetal alcohol spectrum disorders. The peptide NAP potently blocks ethanol inhibition of L1 adhesion and prevents ethanol teratogenesis. We used quantitative RT-PCR and Western blotting of extracts of cerebellar slices, CGNs, and astrocytes from postnatal day 7 (PD7 rats to investigate whether ethanol and NAP act in part by regulating the expression of L1. Treatment of cerebellar slices with 20 mM ethanol, 10(-12 M NAP, or both for 4 hours, 24 hours, and 10 days did not significantly affect L1 mRNA and protein levels. Similar treatment for 4 or 24 hours did not regulate L1 expression in primary cultures of CGNs and astrocytes, the predominant cerebellar cell types. Because ethanol also damages the adult cerebellum, we studied the effects of chronic ethanol exposure in adult rats. One year of binge drinking did not alter L1 gene and protein expression in extracts from whole cerebellum. Thus, ethanol does not alter L1 expression in the developing or adult cerebellum; more likely, ethanol disrupts L1 function by modifying its conformation and signaling. Likewise, NAP antagonizes the actions of ethanol without altering L1 expression.

  7. Expression in Escherichia coli, refolding and crystallization of Aspergillus niger feruloyl esterase A using a serial factorial approach

    NARCIS (Netherlands)

    Benoit, Isabelle; Coutard, Bruno; Oubelaid, Rachid; Asther, Marcel; Bignon, Christophe

    2007-01-01

    Hydrolysis of plant biomass is achieved by the combined action of enzymes secreted by microorganisms and directed against the backbone and the side chains of plant cell wall polysaccharides. Among side chains degrading enzymes, the feruloyl esterase A (FAEA) specifically removes feruloyl residues. T

  8. Homologous expression of the feruloyl esterase B gene from Aspergillus niger and characterization of the recombinant enzyme

    NARCIS (Netherlands)

    Levasseur, Anthony; Benoit, Isabelle; Asther, Michèle; Asther, Marcel; Record, Eric

    2004-01-01

    The faeB gene encoding the feruloyl esterase B (FAEB) was isolated from Aspergillus niger BRFM131 genomic DNA. The faeB gene, with additional sequence coding for a C-terminal histidine tag, was inserted into an expression vector under the control of the gpd promoter and trpC terminator and expressed

  9. Monitoring Lipase/Esterase Activity by Stopped Flow in a Sequential Injection Analysis System Using p-Nitrophenyl Butyrate

    Directory of Open Access Journals (Sweden)

    Jorge Pliego

    2015-01-01

    Full Text Available Lipases and esterases are biocatalysts used at the laboratory and industrial level. To obtain the maximum yield in a bioprocess, it is important to measure key variables, such as enzymatic activity. The conventional method for monitoring hydrolytic activity is to take out a sample from the bioreactor to be analyzed off-line at the laboratory. The disadvantage of this approach is the long time required to recover the information from the process, hindering the possibility to develop control systems. New strategies to monitor lipase/esterase activity are necessary. In this context and in the first approach, we proposed a lab-made sequential injection analysis system to analyze off-line samples from shake flasks. Lipase/esterase activity was determined using p-nitrophenyl butyrate as the substrate. The sequential injection analysis allowed us to measure the hydrolytic activity from a sample without dilution in a linear range from 0.05–1.60 U/mL, with the capability to reach sample dilutions up to 1000 times, a sampling frequency of five samples/h, with a kinetic reaction of 5 min and a relative standard deviation of 8.75%. The results are promising to monitor lipase/esterase activity in real time, in which optimization and control strategies can be designed.

  10. 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.

  11. 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...

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

    NARCIS (Netherlands)

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

    2006-01-01

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

  13. 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...

  14. SSoNΔ and SsoNΔlong: two thermostable esterases from the same ORF in the archaeon Sulfolobus solfataricus?

    Directory of Open Access Journals (Sweden)

    Luigi Mandrich

    2006-01-01

    Full Text Available Previously, we reported from the Sulfolobus solfataricus open reading frame (ORF SSO2517 the cloning, overexpression and characterization of an esterase belonging to the hormone-sensitive lipase (HSL family and apparently having a deletion at the N-terminus, which we named SsoNΔ. Searching the recently reported Sulfolobus acidocaldarius genome by sequence alignment, using SSO2517 as a query, allowed identity of a putative esterase (ORF SAC1105 sharing high sequence similarity (82% with SSO2517. This esterase displays an N-terminus and total length similar to other known esterases of the HSL family. Analysis of the upstream DNA sequence of SS02517 revealed the possibility of expressing a longer version of the protein with an extended N-terminus; however, no clear translation signal consistent with a longer protein version was detected. This new version of SSO2517 was cloned, over-expressed, purified and characterized. The resulting protein, named SsoNΔlong, was 15-fold more active with the substrate p-nitrophenyl hexanoate than SsoNΔ. Furthermore, SsoNΔlong and SsoNΔ displayed different substrate specificities for triacylglycerols. These results and the phylogenetic relationship between S. solfataricus and S. acidocaldarius suggest a common origin of SSO2517 and SAC1105 from an ancestral gene, followed by divergent evolution. Alternatively, a yet-to-be discovered mechanism of translation that directs the expression of SsoNΔlong under specific metabolic conditions could be hypothesized.

  15. Crystallization and Preliminary X-ray Diffraction Analysis of the Glucuronoyl Esterase Catalytic Domain from Hypocrea jecorina

    Science.gov (United States)

    The catalytic domain of the glucuronoyl esterase from Hypocrea jecorina (anamorph Trichoderma reesei) was over-expressed, purified, and crystallized by sitting-drop vapor-diffusion method using 1.4 M sodium/potassium phosphate pH 6.9. Crystals had space group P212121 and X-ray diffraction data were...

  16. Identification and functional characterization of esterases in Euschistus heros (Hemiptera, Pentatomidae) and their relationship with thiamethoxam and lambda-cyhalothrin.

    Science.gov (United States)

    Hegeto, L A; Ronqui, L; Lapenta, A S; Albuquerque, F A

    2015-09-22

    The brown stink bug Euschistus heros is the most abundant species of the soybean-sucking bugs, and causes large economic losses. Applying different chemical groups of organosynthetic insecticides for its control increases the potential for resistance. Esterases are a group of enzymes that play a variety of roles in insects, and some of them are related to the metabolism of xenobiotics. The aim of this study was to analyze the esterase isoenzyme system of this species and investigate its response to Engeo™ Pleno (thiamethoxam and lambda-cyhalothrin), which is the most widely used pesticide in soybean crops. Two strains were analyzed: the EB strain, which had been free of insecticides for several generations; and the MA strain, which was collected in a location exposed to agrochemicals. By analyzing the polyacrylamide gel electrophoresis profile, seven different esterases in adults and nymphs of both strains were found. Eight gene loci were responsible for the synthesis of these enzymes. The differences in esterases between the two strains and enzyme changes in insects exposed to Engeo™ Pleno suggest that EST-2 and EST-4 are related to the metabolism of the agrochemical used and are mechanisms of resistance.

  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 produc

  18. 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 ...

  19. 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 promin...

  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 leuko

  1. 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.

  2. Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm.

    Science.gov (United States)

    Harholt, Jesper; Bach, Inga C; Lind-Bouquin, Solveig; Nunan, Kylie J; Madrid, Susan M; Brinch-Pedersen, Henrik; Holm, Preben B; Scheller, Henrik V

    2010-04-01

    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. 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 threefold relative to wild type. The grains were shrivelled 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 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 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.

  3. Effect of Temperature and High Pressure on the Activity and Mode of Action of Fungal Pectin Methyl Esterase

    NARCIS (Netherlands)

    Duvetter, T.; Fraeye, I.; Sila, D.N.; Verlent, I.; Smout, C.; Clynen, E.; Schoofs, L.; Schols, H.A.; Hendrickx, M.; Loey, van A.

    2006-01-01

    Pectin was de-esterified with purified recombinant Aspergillus aculeatus pectin methyl esterase (PME) during isothermal-isobaric treatments. By measuring the release of methanol as a function of treatment time, the rate of enzymatic pectin conversion was determined. Elevated temperature and pressure

  4. 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

    2013-09-01

    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.

  5. The carboxyl-terminal tail of Noxa protein regulates the stability of Noxa and Mcl-1.

    Science.gov (United States)

    Pang, Xiaming; Zhang, Jingjing; Lopez, Hernando; Wang, Yushu; Li, Wenyang; O'Neill, Katelyn L; Evans, Jacquelynn J D; George, Nicholas M; Long, Jianhong; Chen, Yi; Luo, Xu

    2014-06-20

    The BH3-only protein Noxa is a critical mediator of apoptosis and functions primarily by sequestering/inactivating the antiapoptotic Bcl-2 family protein Mcl-1. Although Noxa is a highly labile protein, recent studies suggested that it is degraded by the proteasome in a ubiquitylation-independent manner. In the present study, we investigated the mechanism of Noxa degradation and its ability to regulate the stability of Mcl-1. We found that the ubiquitylation-independent degradation of Noxa does not require a physical association with Mcl-1. A short stretch of amino acid residues in the C-terminal tail was found to mediate the proteasome-dependent degradation of Noxa. Ectopic placement of this degron was able to render other proteins unstable. Surprisingly, mutation of this sequence not only attenuated the rapid degradation of Noxa, but also stabilized endogenous Mcl-1 through the BH3-mediated direct interaction. Together, these results suggest that the C-terminal tail of Noxa regulates the stability of both Noxa and Mcl-1.

  6. Carboxyl terminal of rhodopsin kinase is required for the phosphorylation of photo—activated rhodopsin

    Institute of Scientific and Technical Information of China (English)

    YUQINGMING; LANMA; 等

    1998-01-01

    Human rhodopsin kinase (RK) and a carboxyl terminus-truncated mutant RK lacking the last 59 amino acids (RKC) were expressed in human embryonic kidney 293 cells to investigate the role of the carboxyl terminus of RK in recognition and phosphorylation of rhodopsin.RKC,like the wild-type RK,was detected in both plasma membranes and cytosolic fractions.The Cterminal truncated rhodopsin kinase was unable to phosphorylate photo-activated rhodopsin,but possesses kinase activity similar to the wild-type RK in phosphorylation of small peptide substrate.It suggests that the truncation did not disturb the gross structures of RK catalytic domain.Our results also show that RKC failed to translocate to photo-activated rod out segments.Taken together,our study demonstrate the carboxyl terminus of RK is required for phosphorylation of photo-activated rhodopsin and strongly indicate that carboxyl-terminus of RK may be involved in interaction with photo-activated rhodopsin.

  7. 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

  8. Esterases activity in the axolotl Ambystoma mexicanum exposed to chlorpyrifos and its implication to motor activity.

    Science.gov (United States)

    Robles-Mendoza, Cecilia; Zúñiga-Lagunes, Sebastian R; Ponce de León-Hill, Claudia A; Hernández-Soto, Jesús; Vanegas-Pérez, Cecilia

    2011-10-01

    The axolotl Ambystoma mexicanum is a neotenic salamander considered a good biological model due to its ability to regenerate limbs, tail, brain and heart cells. Nevertheless, severe reduction of A. mexicanum wild populations in the lacustrine area of Xochimilco, the natural habitat of the axolotl, could be related to several environmental pressures as the presence of organophosphate pesticides (OPPs), intensively applied in agricultural activities in Xochimilco. Thus the aim of this study was to evaluate the effect of environmentally realistic chlorpyrifos (CPF) concentrations, a OPP commonly used in this zone, on esterases activity (acetylcholinesterase and carboxylesterase) and bioconcentration of CPF and to relate them with the motor activity of A. mexicanum juveniles. Axolotls were exposed 48 h to 0.05 and 0.1mg CPF/L, and the responses were evaluated at the end of the CPF exposure. Results suggest that CPF is bioconcentrated into axolotls and that the CPF internal concentrations are related with the observed inhibition activity of AChE (>50%) and CbE (≈ 50%). CPF concentration responsible of the inhibition of the 50% of AChE activity (IC50) was estimated in 0.04 mg CPF/L; however IC50 for CbE activity was not possible to calculate since inhibition levels were lower than 50%, results that suggest a higher resistance of CbE enzymatic activity to CPF. However, motor activity was a more sensitive endpoint to CPF poisoning since time that axolotls spent active and walking, frequency and speed of swimming, frequency of prey attack were reduced >90% of control groups. The motor activity alterations in the axolotl could be related with the registered esterases inhibition. Thus important alterations on axolotls were identified even at short time and low concentrations of CPF exposure. Also, it was possible to link biochemical responses as esterases activity with higher levels of biological organization as behavior. This study provides tools for the regulation of the

  9. Subband Adaptive Filtering with l1-Norm Constraint for Sparse System Identification

    Directory of Open Access Journals (Sweden)

    Young-Seok Choi

    2013-01-01

    Full Text Available This paper presents a new approach of the normalized subband adaptive filter (NSAF which directly exploits the sparsity condition of an underlying system for sparse system identification. The proposed NSAF integrates a weighted l1-norm constraint into the cost function of the NSAF algorithm. To get the optimum solution of the weighted l1-norm regularized cost function, a subgradient calculus is employed, resulting in a stochastic gradient based update recursion of the weighted l1-norm regularized NSAF. The choice of distinct weighted l1-norm regularization leads to two versions of the l1-norm regularized NSAF. Numerical results clearly indicate the superior convergence of the l1-norm regularized NSAFs over the classical NSAF especially when identifying a sparse system.

  10. Molecular characterization of endophytes isolated from Saccharum spp based on esterase and ribosomal DNA (ITS1-5.8S-ITS2) analyses.

    Science.gov (United States)

    Leme, A C; Bevilaqua, M R R; Rhoden, S A; Mangolin, C A; Machado, M F P S; Pamphile, J A

    2013-09-27

    This study used esterases and ribosomal DNA (rDNA) markers to determine endophytic variability in order to better understand endophyte-host interactions. Polyacrylamide gel electrophoresis and esterase isoenzymes (EST; EC 3.1.1.3), with α-naphthyl acetate and β-naphthyl acetate as substrates, were used to assess relationships among endophytes. ITS1-5.8S-ITS2 sequencing data were used as rDNA markers. Thirty-two esterases were obtained from 37 isolates of Saccharum spp, which clustered into five endophyte groups. Esterase EST-06 was observed with the highest frequency, being present in 22 of the 37 isolates analyzed, followed by esterase EST-11, which was present in 20 isolates. The esterases EST-10 and EST-14 were present in 19 isolates and EST-09 was present in 18 isolates. The esterase EST-01 was unique to isolate 33 and can, therefore, be used as a marker for this isolate. None of the esterases identified were common to all isolates tested. Similarly, phylogenetic analysis, based on rDNA sequence data, classified the isolates into 5 genus groups: 1) Curvularia with a 100% bootstrap value (BP), 2) Alternaria with 100% BP, 3) Epicoccum with 60% BP, 4) Phoma with 89% BP, and 5) Saccharicola with 100% BP. This polyphyletic analysis based on several markers, therefore, proved to be a valuable approach in determining the relationship between variation in endophytes and their associated host plants. Furthermore, both the esterase and rDNA analyses obtained similar results and were equally effective in resolving relationships.

  11. In vitro screening for compounds that enhance human L1 mobilization.

    Directory of Open Access Journals (Sweden)

    Natsuko Terasaki

    Full Text Available The Long interspersed element 1 (LINE1 or L1 retrotransposon constitutes 17% of the human genome. There are currently 80-100 human L1 elements that are thought to be active in any diploid human genome. These elements can mobilize into new locations of the genome, resulting in changes in genomic information. Active L1s are thus considered to be a type of endogenous mutagen, and L1 insertions can cause disease. Certain stresses, such as gamma radiation, oxidative stress, and treatment with some agents, can induce transcription and/or mobilization of retrotransposons. In this study, we used a reporter gene assay in HepG2 cells to screen compounds for the potential to enhance the transcription of human L1. We assessed 95 compounds including genotoxic agents, substances that induce cellular stress, and commercially available drugs. Treatment with 15 compounds increased the L1 promoter activity by >1.5-fold (p<0.05 after 6 or 24 hours of treatment. In particular, genotoxic agents (benzo[a]pyrene, camptothecin, cytochalasin D, merbarone, and vinblastine, PPARα agonists (bezafibrate and fenofibrate, and non-steroidal anti-inflammatory drugs (diflunisal, flufenamic acid, salicylamide, and sulindac induced L1 promoter activity. To examine their effects on L1 retrotransposition, we developed a high-throughput real-time retrotransposition assay using a novel secreted Gaussia luciferase reporter cassette. Three compounds (etomoxir, WY-14643, and salicylamide produced a significant enhancement in L1 retrotransposition. This is the first study to report the effects of a wide variety of compounds on L1 transcription and retrotransposition. These results suggest that certain chemical- and drug-induced stresses might have the potential to cause genomic mutations by inducing L1 mobilization. Thus, the risk of induced L1 transcription and retrotransposition should be considered during drug safety evaluation and environmental risk assessments of chemicals.

  12. The Effects of L2 Reading Skills on L1 Reading Skills through Transfer

    Science.gov (United States)

    Altmisdort, Gonca

    2016-01-01

    This study investigated whether transfer from L2 to L1 in reading occurs, and if so, which reading sub-skills are transferred into L1 reading. The aim is to identify the role of second language reading skills in L1 reading skills by means of transfer. In addition, the positive effects of the second language transfer to the first language in the…

  13. UCH-L1 induces podocyte hypertrophy in membranous nephropathy by protein accumulation.

    Science.gov (United States)

    Lohmann, Frithjof; Sachs, Marlies; Meyer, Tobias N; Sievert, Henning; Lindenmeyer, Maja T; Wiech, Thorsten; Cohen, Clemens D; Balabanov, Stefan; Stahl, R A K; Meyer-Schwesinger, Catherine

    2014-07-01

    Podocytes are terminally differentiated cells of the glomerular filtration barrier that react with hypertrophy in the course of injury such as in membranous nephropathy (MGN). The neuronal deubiquitinase ubiquitin C-terminal hydrolase L1 (UCH-L1) is expressed and activated in podocytes of human and rodent MGN. UCH-L1 regulates the mono-ubiquitin pool and induces accumulation of poly-ubiquitinated proteins in affected podocytes. Here, we investigated the role of UCH-L1 in podocyte hypertrophy and in the homeostasis of the hypertrophy associated "model protein" p27(Kip1). A better understanding of the basic mechanisms leading to podocyte hypertrophy is crucial for the development of specific therapies in MGN. In human and rat MGN, hypertrophic podocytes exhibited a simultaneous up-regulation of UCH-L1 and of cytoplasmic p27(Kip1) content. Functionally, inhibition of UCH-L1 activity and knockdown or inhibition of UCH-L1 attenuated podocyte hypertrophy by decreasing the total protein content in isolated glomeruli and in cultured podocytes. In contrast, UCH-L1 levels and activity increased podocyte hypertrophy and total protein content in culture, specifically of cytoplasmic p27(Kip1). UCH-L1 enhanced cytoplasmic p27(Kip1) levels by nuclear export and decreased poly-ubiquitination and proteasomal degradation of p27(Kip1). In parallel, UCH-L1 increased podocyte turnover, migration and cytoskeletal rearrangement, which are associated with known oncogenic functions of cytoplasmic p27(Kip1) in cancer. We propose that UCH-L1 induces podocyte hypertrophy in MGN by increasing the total protein content through altered degradation and accumulation of proteins such as p27(Kip1) in the cytoplasm of podocytes. Modification of both UCH-L1 activity and levels could be a new therapeutic avenue to podocyte hypertrophy in MGN.

  14. Adult ESL Korean Readers' Responses about Their Reading in L1 Korean and L2 English

    Science.gov (United States)

    Kim, Misun

    2010-01-01

    In this research study I explore: (a) beliefs Korean ESL readers have about reading in L1 and L2 prior to the Retrospective Miscue Analysis (RMA) sessions, (b) how their beliefs about reading affect the way they read in L1 and L2 and their evaluation of themselves as readers in L1 and L2 reading, and (c) change of their beliefs about reading and…

  15. A Study of the Differences Between Children's L1 and Adults'L2 in SLA

    Institute of Scientific and Technical Information of China (English)

    孙岩; 马炳军

    2015-01-01

    The differences between children's L1 acquisition and adults'L2 acquisition based on a qualitative analysis are dis-cussed and compared.Through systematic review of the relevant literature in light of the theories of L1 and L2 acquisition between children and adults,and analysis of the factors both influencing children's L1 and adults'L2 acquisition.The findings show that two different acquisitions are distinguished in such aspects as in acquisition age,device,mode,environment and motivation,which conclude that children's L1 acquisition is effortless while adults'L2 acquisition is painful.

  16. An adaptive support driven reweighted L1-regularization algorithm for fluorescence molecular tomography.

    Science.gov (United States)

    Shi, Junwei; Liu, Fei; Pu, Huangsheng; Zuo, Simin; Luo, Jianwen; Bai, Jing

    2014-11-01

    Fluorescence molecular tomography (FMT) is a promising in vivo functional imaging modality in preclinical study. When solving the ill-posed FMT inverse problem, L1 regularization can preserve the details and reduce the noise in the reconstruction results effectively. Moreover, compared with the regular L1 regularization, reweighted L1 regularization is recently reported to improve the performance. In order to realize the reweighted L1 regularization for FMT, an adaptive support driven reweighted L1-regularization (ASDR-L1) algorithm is proposed in this work. This algorithm has two integral parts: an adaptive support estimate and the iteratively updated weights. In the iteratively reweighted L1-minimization sub-problem, different weights are equivalent to different regularization parameters at different locations. Thus, ASDR-L1 can be considered as a kind of spatially variant regularization methods for FMT. Physical phantom and in vivo mouse experiments were performed to validate the proposed algorithm. The results demonstrate that the proposed reweighted L1-reguarization algorithm can significantly improve the performance in terms of relative quantitation and spatial resolution.

  17. Negative Transfer of L1 on English Grammar Learning in SLA

    Institute of Scientific and Technical Information of China (English)

    马秀琳

    2015-01-01

    At present,many scholars pay more attention to the positive transfer of native language on the English learning,while ignoring the negative transfer of L1 on English grammar learning.Therefore native transfer of L1 often appears on English grammar learning.This paper aims to point out that the negative transfer of L1 has a profound and vast influence on the English grammar learning,to find out the countermeasures to reduce the influence of negative transfer of L1 and finally to bring the benefits to the following relative studies.

  18. Negative Transfer of L1 on English Grammar Learning in SLA

    Institute of Scientific and Technical Information of China (English)

    马秀琳

    2015-01-01

    At present,many scholars pay more attention to the positive transfer of native language on the English learning,while ignoring the negative transfer of L1 on English grammar learning. Therefore native transfer of L1 often appears on English grammar learning.This paper aims to point out that the negative transfer of L1 has a profound and vast influence on the English grammar learning,to find out the countermeasures to reduce the influence of negative transfer of L1 and finally to bring the benefits to the following relative studies.

  19. Feruloyl esterases from Schizophyllum commune to treat food industry side-streams.

    Science.gov (United States)

    Nieter, Annabel; Kelle, Sebastian; Linke, Diana; Berger, Ralf G

    2016-11-01

    Agro-industrial side-streams are abundant and renewable resources of hydroxycinnamic acids with potential applications as antioxidants and preservatives in the food, health, cosmetic, and pharmaceutical industries. Feruloyl esterases (FAEs) from Schizophyllum commune were functionally expressed in Pichia pastoris with extracellular activities of 6000UL(-1). The recombinant enzymes, ScFaeD1 and ScFaeD2, released ferulic acid from destarched wheat bran and sugar beet pectin. Overnight incubation of coffee pulp released caffeic (>60%), ferulic (>80%) and p-coumaric acid (100%) indicating applicability for the valorization of food processing wastes and enhanced biomass degradation. Based on substrate specificity profiling and the release of diferulates from destarched wheat bran, the recombinant FAEs were characterized as type D FAEs. ScFaeD1 and ScFaeD2 preferably hydrolyzed feruloylated saccharides with ferulic acid esterified to the O-5 position of arabinose residues and showed an unprecedented ability to hydrolyze benzoic acid esters.

  20. C1-esterase inhibitor blocks T lymphocyte proliferation and cytotoxic T lymphocyte generation in vitro

    DEFF Research Database (Denmark)

    Nissen, Mogens Holst; Bregenholt, S; Nording, J A

    1998-01-01

    We have previously shown that activated C1s complement and activated T cells cleave beta2-microglobulin (beta2m) in vitro leading to the formation of desLys58 beta2m. This process can specifically be inhibited by C1-esterase inhibitor (C1-inh). Furthermore we showed that exogenously added desLys58...... beta2m in nanomolar amounts to a one-way allogenic mixed lymphocyte culture (MLC) increased the endogenous production of IL-2 and the generation of allo-specific cytotoxic T lymphocytes. C1-inh was purified from fresh human plasma and added to human or murine MLC and mitogen-stimulated lymphocyte...... cultures grown in the presence of complement-inactivated serum. Read-outs were cell proliferation, lymphokine production and development of T cell-mediated cytotoxicity. We found that addition of C1-inh to MLC and mitogen-exposed murine and human lymphocyte cultures inhibited proliferation, the development...

  1. Functional-based screening methods for lipases, esterases, and phospholipases in metagenomic libraries.

    Science.gov (United States)

    Reyes-Duarte, Dolores; Ferrer, Manuel; García-Arellano, Humberto

    2012-01-01

    The use of metagenomic techniques for enzyme discovery constitutes a powerful approach. Functional screens, in contrast to sequence homology search, enable us to select enzymes based on their activity. It is noteworthy that they additionally guarantee the identification of genes coding for enzymes that exhibited no sequence similarity to known counterparts from public databases and that even do not match any putative catalytic residues, involved in the selected catalytic function. Therefore, this strategy not only provides new enzymes for new biotechnological applications, but also allows functional assignment of many proteins, found in abundance in the databases, currently designated as "hypothetical" or "conserved hypothetical" proteins. In the past decade, there has been an exponential increase in the design of functional screening programmes, the majority of them established for hydrolases and oxidoreductases. Here, functional screening methods that guarantee the greatest enzyme diversity, for mining esterases and lipases, are described.

  2. Fundamental reaction mechanism and free energy profile for (-)-cocaine hydrolysis catalyzed by cocaine esterase.

    Science.gov (United States)

    Liu, Junjun; Hamza, Adel; Zhan, Chang-Guo

    2009-08-26

    The fundamental reaction mechanism of cocaine esterase (CocE)-catalyzed hydrolysis of (-)-cocaine and the corresponding free energy profile have been studied by performing pseudobond first-principles quantum mechanical/molecular mechanical free energy (QM/MM-FE) calculations. On the basis of the QM/MM-FE results, the entire hydrolysis reaction consists of four reaction steps, including the nucleophilic attack on the carbonyl carbon of (-)-cocaine benzoyl ester by the hydroxyl group of Ser117, dissociation of (-)-cocaine benzoyl ester, nucleophilic attack on the carbonyl carbon of (-)-cocaine benzoyl ester by water, and finally dissociation between the (-)-cocaine benzoyl group and Ser117 of CocE. The third reaction step involving the nucleophilic attack of a water molecule was found to be rate-determining, which is remarkably different from (-)-cocaine hydrolysis catalyzed by wild-type butyrylcholinesterase (BChE; where the formation of the prereactive BChE-(-)-cocaine complex is rate-determining) or its mutants containing Tyr332Gly or Tyr332Ala mutation (where the first chemical reaction step is rate-determining). Besides, the role of Asp259 in the catalytic triad of CocE does not follow the general concept of the "charge-relay system" for all serine esterases. The free energy barrier calculated for the rate-determining step of CocE-catalyzed hydrolysis of (-)-cocaine is 17.9 kcal/mol, which is in good agreement with the experimentally derived activation free energy of 16.2 kcal/mol. In the present study, where many sodium ions are present, the effects of counterions are found to be significant in determining the free energy barrier. The finding of the significant effects of counterions on the free energy barrier may also be valuable in guiding future mechanistic studies on other charged enzymes.

  3. Phenylmethylsulfonyl fluoride, a potentiator of neuropathy, alters the interaction of organophosphorus compounds with soluble brain esterases.

    Science.gov (United States)

    Mangas, Iris; Vilanova, Eugenio; Estévez, Jorge

    2012-11-19

    Phenylmethylsulfonyl fluoride (PMSF) is a protease and esterase inhibitor that causes protection or potentiation/promotion of organophosphorus delayed neuropathy (OPIDN) depending on whether it is dosed before or after an inducer of delayed neuropathy. The molecular target of promotion has not yet been identified. Kinetic data of esterase inhibition were first obtained for PMSF with a soluble chicken brain fraction and then analyzed using a kinetic model with a multienzymatic system in which inhibition occurred with the simultaneous chemical hydrolysis of the inhibitor and ongoing inhibition (inhibition during the substrate reaction). The best fitting model was a model with resistant fraction, Eα (28%), and two sensitive enzymatic entities, Eβ (61%) and Eγ (11%), with I(50) at 20 min of 70 and 447 μM, respectively. The estimated constant of the chemical hydrolysis of PMSF was kh = 0.23 min(-1). Eα, which is sensitive to mipafox and resistant to PMSF, became less sensitive to mipafox when the preparation was preincubated with PMSF. Its Eα I(50) (30 min) of mipafox increased with the PMSF concentration used to preincubate it. Eγ is sensitive to both PMSF and mipafox, and after preincubation with PMSF, Eγ became less sensitive to mipafox and was totally resistant after preincubation with 10 μM PMSF or more. The sensitivity of Eα to paraoxon (I(50) 30 min from 9 to 11 nM) diminished after PMSF preincubation (I(50) 30 min 185 nM) and showed no spontaneous reactivation capacity. The nature of these interactions is unknown but might be due to covalent binding at sites other than the substrate catalytic center. Such interactions should be considered to interpret the potentiation/promotion phenomenon of PMSF and to understand the effects of multiple exposures to chemicals.

  4. Initial clinical experience with remifentanil, a new opioid metabolized by esterases.

    Science.gov (United States)

    Dershwitz, M; Randel, G I; Rosow, C E; Fragen, R J; Connors, P M; Librojo, E S; Shaw, D L; Peng, A W; Jamerson, B D

    1995-09-01

    Remifentanil is a new, esterase-metabolized opioid for anesthesia. Nonspecific esterases terminate the drug effect, with a context-sensitive half-time which plateaus at 3-4 min. This dose-ranging pilot study was designed to estimate the dose requirement of remifentanil for abolition of the responses to skin incision and intraoperative stimuli, and to determine the speed of recovery. Fifty-one unpremedicated patients took part at two centers. Anesthesia was induced with propofol, 67% nitrous oxide, and vecuronium. Remifentanil was then given (1 microgram/kg, plus an infusion of 0.0125-1.0 micrograms.kg-1.min-1). Responses were defined as: > 15% increase in systolic blood pressure or > 20% increase in heart rate, tearing, sweating, movement, or coughing. Responses to incision or surgery were treated with 0.5 micrograms/kg remifentanil boluses and a 50% increase in infusion rate, which could be done twice. Subsequent responses were treated with propofol or isoflurane. Remifentanil and nitrous oxide administration were terminated after the incision was closed. ED50 for response to skin incision varied between the two study sites (0.020 and 0.087 microgram.kg-1.min-1). ED50 for response to all surgical stimuli was 0.52 microgram.kg-1.min-1. At 0.3 microgram.kg-1.min-1 or more, only 3 of 21 patients required isoflurane. Recovery was not longer in patients receiving larger doses to spontaneous ventilation (2.5-4.6 min), tracheal extubation (4.2-7.0 min), and response to verbal command (3.0-4.6 min). Postoperative pain was reported in most patients (92%) at a median time of 21 min. We conclude that remifentanil was effective and well tolerated as a component of nitrous oxide-opioid-relaxant anesthesia.(ABSTRACT TRUNCATED AT 250 WORDS)

  5. C1-esterase inhibitor treatment: preclinical safety aspects on the potential prothrombotic risk.

    Science.gov (United States)

    Schürmann, Daniel; Herzog, Eva; Raquet, Elmar; Nolte, Marc W; May, Frauke; Müller-Cohrs, Jochen; Björkqvist, Jenny; Dickneite, Gerhard; Pragst, Ingo

    2014-11-01

    Human plasma-derived C1-esterase inhibitor (C1-INH) is an efficacious and safe treatment for hereditary angioedema. However, thrombotic events in subjects treated with C1-INH at recommended or off-label, high doses have been reported. In this study, we addressed the potential prothrombotic risk of C1-INH treatment in high doses using a non-clinical rabbit model. Following intravenous infusion of C1-INH to rabbits at doses up to 800 IU/kg, the exposure and the pharmacodynamic efficacy of C1-INH in rabbits were confirmed by activity measurements of C1-esterase, and coagulation factors XIa and XIIa, respectively. Potential prothrombotic effects were assessed following induction of venous and arterial thrombosis using in vivo models of venous and arterial stasis, complemented by various in vitro assays of coagulation markers. Administration of C1-INH at doses up to 800 IU/kg did not potentiate thrombus formation during venous stasis. In contrast, inhibition of arterial occlusion was observed upon C1-INH administration when compared with isotonic saline treatment, indicating antithrombotic rather than prothrombotic activity of high dose C1-INH treatment in vivo. This was further confirmed in vitro by decreased thrombin generation, increased activated partial thromboplastin time, clotting time and clot formation time, and inhibition of platelet aggregation. No relevant changes in fibrinolysis or in the levels of thrombin-antithrombin complexes, and prothrombin fragment 1+2 were observed upon high dose C1-INH treatment. The data suggest that treatment of healthy rabbits with high doses of C1-INH could potentially inhibit coagulation and thrombus formation rather than induce a prothrombotic risk.

  6. Direct imaging of ER calcium with targeted-esterase induced dye loading (TED).

    Science.gov (United States)

    Samtleben, Samira; Jaepel, Juliane; Fecher, Caroline; Andreska, Thomas; Rehberg, Markus; Blum, Robert

    2013-05-07

    Visualization of calcium dynamics is important to understand the role of calcium in cell physiology. To examine calcium dynamics, synthetic fluorescent Ca(2+) indictors have become popular. Here we demonstrate TED (= targeted-esterase induced dye loading), a method to improve the release of Ca(2+) indicator dyes in the ER lumen of different cell types. To date, TED was used in cell lines, glial cells, and neurons in vitro. TED bases on efficient, recombinant targeting of a high carboxylesterase activity to the ER lumen using vector-constructs that express Carboxylesterases (CES). The latest TED vectors contain a core element of CES2 fused to a red fluorescent protein, thus enabling simultaneous two-color imaging. The dynamics of free calcium in the ER are imaged in one color, while the corresponding ER structure appears in red. At the beginning of the procedure, cells are transduced with a lentivirus. Subsequently, the infected cells are seeded on coverslips to finally enable live cell imaging. Then, living cells are incubated with the acetoxymethyl ester (AM-ester) form of low-affinity Ca(2+) indicators, for instance Fluo5N-AM, Mag-Fluo4-AM, or Mag-Fura2-AM. The esterase activity in the ER cleaves off hydrophobic side chains from the AM form of the Ca(2+) indicator and a hydrophilic fluorescent dye/Ca(2+) complex is formed and trapped in the ER lumen. After dye loading, the cells are analyzed at an inverted confocal laser scanning microscope. Cells are continuously perfused with Ringer-like solutions and the ER calcium dynamics are directly visualized by time-lapse imaging. Calcium release from the ER is identified by a decrease in fluorescence intensity in regions of interest, whereas the refilling of the ER calcium store produces an increase in fluorescence intensity. Finally, the change in fluorescent intensity over time is determined by calculation of ΔF/F0.

  7. Propoxur-induced acetylcholine esterase inhibition and impairment of cognitive function: attenuation by Withania somnifera.

    Science.gov (United States)

    Yadav, C S; Kumar, V; Suke, S G; Ahmed, R S; Mediratta, P K; Banerjee, B D

    2010-04-01

    Propoxur (2-isopropoxyphenyl N-methylcarbamate) is widely used as an acaricide in agriculture and public health programs. Studies have shown that sub-chronic exposure to propoxur can cause oxidative stress and immuno-suppression in rats. Carbamates are also known to exhibit inhibitory effect on cholinesterase activity, which is directly related to their cholinergic effects. In the present study, the effect of Withania somnifera (Ashwagandha), a widely used herbal drug possessing anti-stress and immunomodulatory properties was studied on propoxur-induced acetylcholine esterase inhibition and impairment of cognitive function in rats. Male Wistar rats were divided into four groups. Group I was treated with olive oil and served as control. Group II was administered orally with propoxur (10 mg/kg b.wt.) in olive oil, group III received a combination of propoxur (10 mg/kg b.wt.) and W. somnifera (100 mg/kg b.wt.) suspension and group IV W. somnifera (100 mg/kg b.wt.) only. All animals were treated for 30 days. Cognitive behaviour was assessed by transfer latency using elevated plus maze. Blood and brain acetylcholine esterase (AChE) activity was also assessed. Oral administration of propoxur (10 mg/kg b.wt.) resulted in a significant reduction of brain and blood AChE activity. A significant prolongation of the acquisition as well as retention transfer latency was observed in propoxur-treated rats. Oral treatment of W. somnifera exerts protective effect and attenuates AChE inhibition and cognitive impairment caused by sub-chronic exposure to propoxur.

  8. L1/2 regularization: a thresholding representation theory and a fast solver.

    Science.gov (United States)

    Xu, Zongben; Chang, Xiangyu; Xu, Fengmin; Zhang, Hai

    2012-07-01

    The special importance of L1/2 regularization has been recognized in recent studies on sparse modeling (particularly on compressed sensing). The L1/2 regularization, however, leads to a nonconvex, nonsmooth, and non-Lipschitz optimization problem that is difficult to solve fast and efficiently. In this paper, through developing a threshoding representation theory for L1/2 regularization, we propose an iterative half thresholding algorithm for fast solution of L1/2 regularization, corresponding to the well-known iterative soft thresholding algorithm for L1 regularization, and the iterative hard thresholding algorithm for L0 regularization. We prove the existence of the resolvent of gradient of ||x||1/2(1/2), calculate its analytic expression, and establish an alternative feature theorem on solutions of L1/2 regularization, based on which a thresholding representation of solutions of L1/2 regularization is derived and an optimal regularization parameter setting rule is formulated. The developed theory provides a successful practice of extension of the well- known Moreau's proximity forward-backward splitting theory to the L1/2 regularization case. We verify the convergence of the iterative half thresholding algorithm and provide a series of experiments to assess performance of the algorithm. The experiments show that the half algorithm is effective, efficient, and can be accepted as a fast solver for L1/2 regularization. With the new algorithm, we conduct a phase diagram study to further demonstrate the superiority of L1/2 regularization over L1 regularization.

  9. Effects of MAPK and PI3K Pathways on PD-L1 Expression in Melanoma

    Science.gov (United States)

    Atefi, Mohammad; Avramis, Earl; Lassen, Amanda; Wong, Deborah; Robert, Lidia; Foulad, David; Cerniglia, Michael; Titz, Bjoern; Chodon, Thinle; Graeber, Thomas G.; Comin-Anduix, Begonya; Ribas, Antoni

    2014-01-01

    Purpose PD-L1 is the main ligand for the immune inhibitory receptor PD-1. This ligand is frequently expressed by melanoma cells. In this study we investigated whether PD-L1 expression is controlled by melanoma driver mutations and modified by oncogenic signaling inhibition. Experimental Design Expression of PD-L1 was investigated in a panel of 51 melanoma cell lines containing different oncogenic mutations, including cell lines with innate and acquired resistance to BRAF inhibitors. The effects of targeted therapy drugs on expression of PD-L1 by melanoma cells were investigated. Results No association was found between the level of PD-L1 expression and mutations in BRAF, NRAS, PTEN or amplification of AKT. Resistance to vemurafenib due to the activation of alternative signaling pathways was accompanied with the induction of PD-L1 expression, while the resistance due to the reactivation of the MAPK pathway had no effect on PD-L1 expression. In melanoma cell lines the effects of BRAF, MEK and PI3K inhibitors on expression of PD-L1 were variable from reduction to induction, particularly in the presence of INFγ. In PD-L1-exposed lymphocytes, vemurafenib paradoxically restored activity of the MAPK pathway and increased the secretion of cytokines. Conclusions In melanoma cell lines, including BRAF inhibitor-resistant cells, PD-L1 expression is variably regulated by oncogenic signaling pathways. PD-L1-exposed lymphocytes decrease MAPK signaling, which is corrected by exposure to vemurafenib, providing potential benefits of combining this drug with immunotherapies. PMID:24812408

  10. Clinical Neuropathology mini-review 6-2015: PD-L1: emerging biomarker in glioblastoma?

    Science.gov (United States)

    Preusser, Matthias; Berghoff, Anna S; Wick, Wolfgang; Weller, Michael

    2015-01-01

    Programmed death 1 (PD-1, CD279) and programmed death ligand 1 (PD-L1, CD274) are involved in generating tumor-associated immunosuppression by suppression of T-cell proliferation and interleukin 2 (IL-2) production and immune checkpoint inhibitors targeting these molecules are showing compelling activity against a variety of human cancers. PD-L1 expression has shown a positive association with response to PD-1 inhibition in noncentral nervous system (CNS) tumors, e.g., melanoma or non-small cell lung cancer, and is discussed as a potential predictive biomarker for patient selection in these tumor types. This review summarizes current knowledge and potential clinical implications of PD-L1 expression in glioblastoma. At present, the following conclusions are drawn: (a) functional data support a role for PD-1/PD-L1 in tumor-associated immunosuppression in glioblastoma; (b) the incidence of PD-L1-expressing glioblastomas seems to be relatively high in comparison to other tumor types, however, the reported rates of glioblastomas with PD-L1 protein expression vary and range from 61 to 88%; (c) there is considerable variability in the methodology of PD-L1 assessment in glioblastoma across studies with heterogeneity in utilized antibodies, tissue sampling strategies, immunohistochemical staining protocols, cut-off definitions, and evaluated staining patterns; (d) there are conflicting data on the prognostic role and so far no data on the predictive role of PD-L1 gene and protein expression in glioblastoma. In summary, the ongoing clinical studies evaluating the activity of PD-1/PD-L1 inhibitors in glioblastoma need to be complemented with well designed and stringently executed studies to understand the influence of PD-1/PD-L1 expression on therapy response or failure and to develop robust means of PD-L1 assessment for meaningful biomarker development.

  11. The specificity and flexibility of l1 reverse transcription priming at imperfect T-tracts.

    Directory of Open Access Journals (Sweden)

    Clément Monot

    2013-05-01

    Full Text Available L1 retrotransposons have a prominent role in reshaping mammalian genomes. To replicate, the L1 ribonucleoprotein particle (RNP first uses its endonuclease (EN to nick the genomic DNA. The newly generated DNA end is subsequently used as a primer to initiate reverse transcription within the L1 RNA poly(A tail, a process known as target-primed reverse transcription (TPRT. Prior studies demonstrated that most L1 insertions occur into sequences related to the L1 EN consensus sequence (degenerate 5'-TTTT/A-3' sites and frequently preceded by imperfect T-tracts. However, it is currently unclear whether--and to which degree--the liberated 3'-hydroxyl extremity on the genomic DNA needs to be accessible and complementary to the poly(A tail of the L1 RNA for efficient priming of reverse transcription. Here, we employed a direct assay for the initiation of L1 reverse transcription to define the molecular rules that guide this process. First, efficient priming is detected with as few as 4 matching nucleotides at the primer 3' end. Second, L1 RNP can tolerate terminal mismatches if they are compensated within the 10 last bases of the primer by an increased number of matching nucleotides. All terminal mismatches are not equally detrimental to DNA extension, a C being extended at higher levels than an A or a G. Third, efficient priming in the context of duplex DNA requires a 3' overhang. This suggests the possible existence of additional DNA processing steps, which generate a single-stranded 3' end to allow L1 reverse transcription. Based on these data we propose that the specificity of L1 reverse transcription initiation contributes, together with the specificity of the initial EN cleavage, to the distribution of new L1 insertions within the human genome.

  12. Nephrogenic diabetes insipidus in a patient with L1 syndrome: a new report of a contiguous gene deletion syndrome including L1CAM and AVPR2.

    Science.gov (United States)

    Knops, Noël B B; Bos, Krista K; Kerstjens, Mieke; van Dael, Karin; Vos, Yvonne J

    2008-07-15

    We report on an infant boy with congenital hydrocephalus due to L1 syndrome and polyuria due to diabetes insipidus. We initially believed his excessive urine loss was from central diabetes insipidus and that the cerebral malformation caused a secondary insufficient pituitary vasopressin release. However, he failed to respond to treatment with a vasopressin analogue, which pointed to nephrogenic diabetes insipidus (NDI). L1 syndrome and X-linked NDI are distinct clinical disorders caused by mutations in the L1CAM and AVPR2 genes, respectively, located in adjacent positions in Xq28. In this boy we found a deletion of 61,577 basepairs encompassing the entire L1CAM and AVPR2 genes and extending into intron 7 of the ARHGAP4 gene. To our knowledge this is the first description of a patient with a deletion of these three genes. He is the second patient to be described with L1 syndrome and NDI. During follow-up he manifested complications from the hydrocephalus and NDI including global developmental delay and growth failure with low IGF-1 and hypothyroidism.

  13. Bilingual lexical access during L1 sentence reading: The effects of L2 knowledge, semantic constraint, and L1-L2 intermixing.

    Science.gov (United States)

    Titone, Debra; Libben, Maya; Mercier, Julie; Whitford, Veronica; Pivneva, Irina

    2011-11-01

    Libben and Titone (2009) recently observed that cognate facilitation and interlingual homograph interference were attenuated by increased semantic constraint during bilingual second language (L2) reading, using eye movement measures. We now investigate whether cross-language activation also occurs during first language (L1) reading as a function of age of L2 acquisition and task demands (i.e., inclusion of L2 sentences). In Experiment 1, participants read high and low constraint English (L1) sentences containing interlingual homographs, cognates, or control words. In Experiment 2, we included French (L2) filler sentences to increase salience of the L2 during L1 reading. The results suggest that bilinguals reading in their L1 show nonselective activation to the extent that they acquired their L2 early in life. Similar to our previous work on L2 reading, high contextual constraint attenuated cross-language activation for cognates. The inclusion of French filler items promoted greater cross-language activation, especially for late stage reading measures. Thus, L1 bilingual reading is modulated by L2 knowledge, semantic constraint, and task demands.

  14. Ternary Weighted Function and Beurling Ternary Banach Algebra l1ω(S

    Directory of Open Access Journals (Sweden)

    Mehdi Dehghanian

    2011-01-01

    Full Text Available Let S be a ternary semigroup. In this paper, we introduce our notation and prove some elementary properties of a ternary weight function ω on S. Also, we make ternary weighted algebra l1ω(S and show that l1ω(S is a ternary Banach algebra.

  15. L1 and L2 Distance Effects in Learning L3 Dutch

    Science.gov (United States)

    Schepens, Job J.; der Slik, Frans; Hout, Roeland

    2016-01-01

    Many people speak more than two languages. How do languages acquired earlier affect the learnability of additional languages? We show that linguistic distances between speakers' first (L1) and second (L2) languages and their third (L3) language play a role. Larger distances from the L1 to the L3 and from the L2 to the L3 correlate with lower…

  16. Multilingual Acquisition of Vowels in L1 Polish, L2 Danish and L3 English

    Science.gov (United States)

    Sypianska, Jolanta

    2016-01-01

    The aim of this paper is to determine whether all languages in the linguistic repertoire of a multilingual speaker manifest cross-linguistic influence (CLI) and establish the directions of CLI on the basis of chosen vowels from the linguistic repertoire of two groups: the Bilingual group (L1 Polish/L2 Danish) and the Multilingual group (L1

  17. The Impact of L1 Writing System on ESL Knowledge of Vowel and Consonant Spellings

    Science.gov (United States)

    Martin, Katherine I.

    2017-01-01

    Orthographic knowledge, the general ability to learn, store, and use information about the orthographic form of words (Stanovich & West, 1989), is a crucial skill for supporting literacy. Although the development of first language (L1) orthographic awareness is impacted by the characteristics of a learner's L1 writing system, relatively little…

  18. Epigenetic silencing of engineered L1 retrotransposition events in human embryonic carcinoma cells

    Science.gov (United States)

    Garcia-Perez, Jose L.; Morell, Maria; Scheys, Joshua O.; Kulpa, Deanna A.; Morell, Santiago; Carter, Christoph C.; Hammer, Gary D.; Collins, Kathleen L.; O’Shea, K. Sue; Menendez, Pablo; Moran, John V.

    2010-01-01

    Long INterspersed Element-1 (LINE-1 or L1) retrotransposition continues to impact human genome evolution1,2. L1s can retrotranspose in the germline, during early development, and in select somatic cells3,4,5,6,7,8; however, the host response to L1 retrotransposition remains largely unexplored. Here, we show that reporter genes introduced into the genome of various human embryonic carcinoma-derived cell lines (ECs) by L1 retrotransposition are rapidly and efficiently silenced either during or immediately after their integration. Treating ECs with histone deacetylase inhibitors (IHDACs) rapidly reverses this silencing, and chromatin immunoprecipitation (ChIP) experiments revealed that reactivation of the reporter gene was correlated with changes in chromatin status at the L1 integration site. Under our assay conditions, rapid silencing also was observed when reporter genes were delivered into ECs by mouse L1s and a zebrafish LINE-2 element, but not when similar reporter genes were delivered into ECs by Moloney murine leukemia virus (MMLV) or human immunodeficiency virus (HIV), suggesting these integration events are silenced by distinct mechanisms. Finally, we demonstrate that subjecting ECs to culture conditions that promote differentiation attenuates the silencing of reporter genes delivered by L1 retrotransposition, but that differentiation, per se, is not sufficient to reactivate previously silenced reporter genes. Thus, our data suggest that ECs differ from many differentiated cells in their ability to silence reporter genes delivered by L1 retrotransposition. PMID:20686575

  19. Perception of Mandarin Tones: The Effect of L1 Background and Training

    Science.gov (United States)

    Wang, Xinchun

    2013-01-01

    This study investigates whether native Hmong speakers' first language (L1) lexical tone experience facilitates or interferes with their perception of Mandarin tones and whether training is effective for perceptual learning of second (L2) tones. In Experiment 1, 3 groups of beginning level learners of Mandarin with different L1 prosodic background…

  20. Heterologous production of human papillomavirus type-16 L1 protein by a lactic acid bacterium

    Directory of Open Access Journals (Sweden)

    Bermúdez-Humarán Luis G

    2009-08-01

    Full Text Available Abstract Background The expression of vaccine antigens in lactic acid bacteria (LAB is a safe and cost-effective alternative to traditional expression systems. In this study, we investigated i the expression of Human papillomavirus type 16 (HPV-16 L1 major capsid protein in the model LAB Lactococcus lactis and ii the ability of the resulting recombinant strain to produce either capsomer-or virus-like particles (VLPs. Results and conclusion HPV-16 L1 gene was cloned into two vectors, pCYT and pSEC, designed for controlled intra- or extracellular heterologous expression in L. lactis, respectively. The capacity of L. lactis harboring either pCYT:L1 or pSEC:L1 plasmid to accumulate L1 in the cytoplasm and supernatant samples was confirmed by Western blot assays. Electron microscopy analysis suggests that, L1 protein produced by recombinant lactococci can self-assemble into structures morphologically similar to VLPs intracellularly. The presence of conformational epitopes on the L. lactis-derived VLPs was confirmed by ELISA using an anti-HPV16 L1 capsid antigen antibody. Our results support the feasibility of using recombinant food-grade LAB, such as L. lactis, for the production of L1-based VLPs and open the possibility for the development of a new safe mucosal vector for HPV-16 prophylactic vaccination.

  1. Effects of Age of L2 Acquisition on L1 Event Conceptualization Patterns

    Science.gov (United States)

    Bylund, Emanuel

    2009-01-01

    This study explores the effects that the age of onset (AO) of second language (L2) acquisition exerts on the attrition of first language (L1) event conceptualization patterns. The subjects studied are L1 Spanish-L2 Swedish bilinguals living in Sweden. The specific research questions addressed in the study concern the role of AO in endpoint…

  2. The Role of L1 Literacy on L2 Literacy Learning

    Institute of Scientific and Technical Information of China (English)

    张洁

    2014-01-01

    A native language can be learned effortlessly, while a second language is always difficult for people to learn. L1 literacy may impede or promote L2 literacy learning. This paper discusses the role L1 Literacy on L2 Literacy Learning.

  3. Long-Term Crosslinguistic Transfer of Skills from L1 to L2

    Science.gov (United States)

    Sparks, Richard; Patton, Jon; Ganschow, Leonore; Humbach, Nancy

    2009-01-01

    This study investigated the relationship of first language (L1) skills in elementary school and second language (L2) learning in high school. Students classified as high-, average-, and low-proficiency L2 learners were compared on L1 achievement measures of reading, spelling, vocabulary, phonological awareness, and listening comprehension…

  4. Code-Switching: L1-Coded Mediation in a Kindergarten Foreign Language Classroom

    Science.gov (United States)

    Lin, Zheng

    2012-01-01

    This paper is based on a qualitative inquiry that investigated the role of teachers' mediation in three different modes of coding in a kindergarten foreign language classroom in China (i.e. L2-coded intralinguistic mediation, L1-coded cross-lingual mediation, and L2-and-L1-mixed mediation). Through an exploratory examination of the varying effects…

  5. Image reconstruction based on L1 regularization and projection methods for electrical impedance tomography.

    Science.gov (United States)

    Wang, Qi; Wang, Huaxiang; Zhang, Ronghua; Wang, Jinhai; Zheng, Yu; Cui, Ziqiang; Yang, Chengyi

    2012-10-01

    Electrical impedance tomography (EIT) is a technique for reconstructing the conductivity distribution by injecting currents at the boundary of a subject and measuring the resulting changes in voltage. Image reconstruction in EIT is a nonlinear and ill-posed inverse problem. The Tikhonov method with L(2) regularization is always used to solve the EIT problem. However, the L(2) method always smoothes the sharp changes or discontinue areas of the reconstruction. Image reconstruction using the L(1) regularization allows addressing this difficulty. In this paper, a sum of absolute values is substituted for the sum of squares used in the L(2) regularization to form the L(1) regularization, the solution is obtained by the barrier method. However, the L(1) method often involves repeatedly solving large-dimensional matrix equations, which are computationally expensive. In this paper, the projection method is combined with the L(1) regularization method to reduce the computational cost. The L(1) problem is mainly solved in the coarse subspace. This paper also discusses the strategies of choosing parameters. Both simulation and experimental results of the L(1) regularization method were compared with the L(2) regularization method, indicating that the L(1) regularization method can improve the quality of image reconstruction and tolerate a relatively high level of noise in the measured voltages. Furthermore, the projected L(1) method can also effectively reduce the computational time without affecting the quality of reconstructed images.

  6. From weak to strong L1-convergence by an oscillation restriction criterion of BMO type

    NARCIS (Netherlands)

    Balder, E.J.

    2001-01-01

    Recently, Girardi gave acharacterization of relative strong L 1 R-compactness in terms of relative weak L 1 R-compactness and the Bocce criterion [18]. Here this result is generalized and extended by presenting a less stringent oscillation restriction condition (ORC) which enforces the transcendence

  7. Modeling the Development of L1 and EFL Writing Proficiency of Secondary School Students

    NARCIS (Netherlands)

    Schoonen, Rob; van Gelderen, Amos; Stoel, Reinoud D.; Hulstijn, Jan; de Glopper, Cornelis

    2011-01-01

    This longitudinal study investigates the development of writing proficiency in English as a foreign language (EFL), in contrast to the development of first language (L1) writing proficiency in Dutch L1, in a sample of almost 400 secondary school students in the Netherlands. Students performed severa

  8. Modeling the development of L1 and EFL writing proficiency of secondary school students

    NARCIS (Netherlands)

    Schoonen, R.; van Gelderen, A.; Stoel, R.D.; Hulstijn, J.; de Glopper, K.

    2011-01-01

    This longitudinal study investigates the development of writing proficiency in English as a foreign language (EFL), in contrast to the development of first language (L1) writing proficiency in Dutch L1, in a sample of almost 400 secondary school students in the Netherlands. Students performed severa

  9. Assay for the pattern recognition molecule collectin liver 1 (CL-L1)

    DEFF Research Database (Denmark)

    Axelgaard, Esben; Jensenius, Jens Christian; Jensen, Lisbeth;

    Collectin liver 1 (also termed collectin 10 and CL-L1) is a C-type lectin that functions as a pattern recognition molecule (PRM) in the innate immune system1. We have produced antibodies against CL-L1 and have developed a sandwich-type time-resolved immuno-fluorometric assay (TRIFMA...

  10. L1 retrotransposition in neurons is modulated by MeCP2.

    Science.gov (United States)

    Muotri, Alysson R; Marchetto, Maria C N; Coufal, Nicole G; Oefner, Ruth; Yeo, Gene; Nakashima, Kinichi; Gage, Fred H

    2010-11-18

    Long interspersed nuclear elements-1 (LINE-1 or L1s) are abundant retrotransposons that comprise approximately 20% of mammalian genomes. Active L1 retrotransposons can impact the genome in a variety of ways, creating insertions, deletions, new splice sites or gene expression fine-tuning. We have shown previously that L1 retrotransposons are capable of mobilization in neuronal progenitor cells from rodents and humans and evidence of massive L1 insertions was observed in adult brain tissues but not in other somatic tissues. In addition, L1 mobility in the adult hippocampus can be influenced by the environment. The neuronal specificity of somatic L1 retrotransposition in neural progenitors is partially due to the transition of a Sox2/HDAC1 repressor complex to a Wnt-mediated T-cell factor/lymphoid enhancer factor (TCF/LEF) transcriptional activator. The transcriptional switch accompanies chromatin remodelling during neuronal differentiation, allowing a transient stimulation of L1 transcription. The activity of L1 retrotransposons during brain development can have an impact on gene expression and neuronal function, thereby increasing brain-specific genetic mosaicism. Further understanding of the molecular mechanisms that regulate L1 expression should provide new insights into the role of L1 retrotransposition during brain development. Here we show that L1 neuronal transcription and retrotransposition in rodents are increased in the absence of methyl-CpG-binding protein 2 (MeCP2), a protein involved in global DNA methylation and human neurodevelopmental diseases. Using neuronal progenitor cells derived from human induced pluripotent stem cells and human tissues, we revealed that patients with Rett syndrome (RTT), carrying MeCP2 mutations, have increased susceptibility for L1 retrotransposition. Our data demonstrate that L1 retrotransposition can be controlled in a tissue-specific manner and that disease-related genetic mutations can influence the frequency of neuronal L

  11. 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 wa...

  12. SREBP2 mediates the modulation of intestinal NPC1L1 expression by curcumin.

    Science.gov (United States)

    Kumar, Pradeep; Malhotra, Pooja; Ma, Ke; Singla, Amika; Hedroug, Omar; Saksena, Seema; Dudeja, Pradeep K; Gill, Ravinder K; Alrefai, Waddah A

    2011-07-01

    Curcumin, the major phenolic compound in the spice turmeric, exhibits numerous biological effects, including lowering plasma cholesterol and preventing diet-induced hypercholesterolemia. The mechanisms underlying the hypocholesterolemic effect of curcumin are not fully understood. In this regard, intestinal Niemann-Pick C1-like 1 (NPC1L1) cholesterol transporter, the molecular target of intestinal cholesterol absorption inhibitor ezetimibe, plays an essential role in the maintenance of cholesterol homeostasis. The current studies were designed to investigate the effect of curcumin on NPC1L1 function, expression, and promoter activity in intestinal Caco-2 monolayers. NPC1L1 function was evaluated by the measurement of ezetimibe-sensitive [(3)H]cholesterol esterification. Relative abundance of NPC1L1 mRNA and protein was evaluated by real-time PCR and Western blotting, respectively. Luciferase assays were used to measure NPC1L1 promoter activity. Our results showed that curcumin significantly inhibited ezetimibe-sensitive cholesterol esterification in a dose-dependent manner with a maximum decrease (by 52% compared with control) occurring at 50 μM concentration. Curcumin treatment of Caco-2 monolayers also significantly decreased NPC1L1 mRNA and protein expression. Similarly, the promoter activity of the NPC1L1 gene was inhibited significantly (55%) by 50 μM curcumin. The decrease in NPC1L1 promoter activity by curcumin was associated with a reduction in the expression and the DNA-binding activity of the sterol response element-binding protein 2 (SREBP2) transcription factor. Furthermore, the overexpression of active SREBP2 protected NPC1L1 from the inhibitory effect of curcumin. Our studies demonstrate that curcumin directly modulates intestinal NPC1L1 expression via transcriptional regulation and the involvement of SREBP2 transcription factor.

  13. PD-1/PD-L1 expression in chromophobe renal cell carcinoma: An immunological exception?

    Science.gov (United States)

    Erlmeier, Franziska; Hartmann, Arndt; Autenrieth, Michael; Wiedemann, Max; Ivanyi, Philipp; Steffens, Sandra; Weichert, Wilko

    2016-11-01

    Immune checkpoint inhibitors targeting the inhibitory cross talk between tumor and immune cells have been approved for therapy in renal cell carcinoma (RCC). In contrast to clear cell RCC, little is known on PD-1/PD-L1 expression patterns in rarer RCC subtypes. The aim of this study was to evaluate the prevalence, distribution and prognostic impact of PD-1 and PD-L1 expression in chromophobe (ch)RCC. Patients who underwent renal surgery due to chRCC were retrospectively evaluated. Tumor specimen was analyzed for PD-1 and PD-L1 expression by immunohistochemistry. Expression data were correlated with clinic-pathological parameters including patient survival. Eighty-one chRCC patients were eligible for analysis, thereof 25 (30.9 %) and 11 (13.6 %) patients were positive for PD-1(+) tumor-infiltrating mononuclear cells (TIMCs) and tumoral PD-L1(+) expression, respectively. No significant associations were found for PD-1(+) TIMC or tumoral PD-L1(+) expression and clinical attributes. In addition, no differences in 5- and 10-year overall survival for PD-1(-) TIMC compared to PD-1(+) TIMC (90.5 and 72.2 vs. 100 and 75 %; p = 0.41) and for PD-L1(-) tumors compared to PD-L1(+) tumors (91.9 and 76.4 vs. 100 and 50 %; p = 0.48) were observed. In conclusion, to our knowledge this is the first study to evaluate the prognostic impact of PD-1 and PD-L1 in chRCC. PD-L1 does seem to be expressed in a minority of all chRCC, likewise only a minority of chRCC was infiltrated by PD-1-positive inflammatory cells. Neither PD-1(+) TIMC nor tumoral PD-L1(+) expression was associated with parameters of aggressiveness or survival.

  14. HPV L1 and P16 Expression in CIN1 to Predict Future CIN2.

    Science.gov (United States)

    Liu, Chong; Du, Hui; Wang, Chun; Belinson, Jerome L; Yang, Bin; Zhang, Wei; Tang, Jinlong; Wu, Ruifang

    2017-03-08

    To use the biomarkers human papillomavirus (HPV) L1 and p16 to develop an algorithm that could triage the individual patient with CIN1 for the risk of progression. A total of 82 patients initially diagnosed with CIN1 at Peking University Shenzhen Hospital in China had their initial and follow-up paraffin-embedded tissue blocks immune-stained for HPV L1 capsid protein and p16. For CIN1, any staining of abnormal epithelium was considered positive. All patients were followed until they developed CIN2+ or for ≥3 years. About 38 patients regressed (HPV-, Cytology-), 17 persisted (CIN1), and 27 progressed (≥CIN2+). At initial diagnosis, HPV L1 capsid protein was expressed in 42.7% of the CIN1 cases. There was no difference in L1 expression among the 3 groups. However, p16-positive staining in the progression group was significantly higher than in the regression group (Pp16- category was significantly higher than that in the progression group. In the progression group, when CIN1 lesions progressed to CIN2+, the L1-positive rate was significantly decreased from 51.9% to 18.5%, the p16+/HPV L1+ rate decreased from the initial (44.4%) to the final diagnosis (14.8%), and the p16+/HPV L1- rate increased from the initial (25.9%) to the final diagnosis (66.7%). P16 expression is a clear risk factor for the progression of CIN1. The p16-/HPV L1- pattern was significantly associated with the regression of CIN1. Moving from CIN1 to CIN2+ over time, p16+/HPV L1+ decreased, and p16+/HPV L1- increased. Unfortunately, our objective of finding a sensitive and specific triage algorithm for the individual patient with CIN1 was still not achieved.

  15. Identification of CREB3L1 as a Biomarker Predicting Doxorubicin Treatment Outcome.

    Directory of Open Access Journals (Sweden)

    Bray Denard

    Full Text Available Doxorubicin has been shown to inhibit proliferation of cancer cells through proteolytic activation of CREB3L1 (cAMP response element binding protein 3-like 1, a transcription factor synthesized as a membrane-bound precursor. Upon doxorubicin treatment, CREB3L1 is cleaved so that the N-terminal domain of the protein can reach the nucleus where it activates transcription of genes that inhibit cell proliferation. These results suggest that the level of CREB3L1 in cancer cells may determine their sensitivity to doxorubicin.Mice transplanted with 6 lines of renal cell carcinoma (RCC were injected with doxorubicin to observe the effect of the chemotherapy on tumor growth. Immunohistochemistry and bioinformatics analyses were performed to compare CREB3L1 levels in types of cancer known to respond to doxorubicin versus those resistant to doxorubicin.Higher levels of CREB3L1 protein are correlated with increased doxorubicin sensitivity of xenograft RCC tumors (p = 0.017 by Pearson analysis. From patient tumor biopsies we analyzed, CREB3L1 was expressed in 19% of RCC, which is generally resistant to doxorubicin, but in 70% of diffuse large B-cell lymphoma that is sensitive to doxorubicin. Doxorubicin is used as the standard treatment for cancers that express the highest levels of CREB3L1 such as osteosarcoma and malignant fibrous histiocytoma but is not generally used to treat those that express the lowest levels of CREB3L1 such as RCC.Identification of CREB3L1 as the biomarker for doxorubicin sensitivity may markedly improve the doxorubicin response rate by applying doxorubicin only to patients with cancers expressing CREB3L1.

  16. Sekuens Gen Protein Kapsid Mayor L1 Human Papilomavirus 16 dari Isolat Klinik Asal Bandung

    Directory of Open Access Journals (Sweden)

    Anandayu Pradita

    2014-09-01

    Full Text Available Cervical cancer is strongly associated with chronic human papillomavirus (HPV infection. HPV-16 is the most prevalent genotype infecting cervical epithelium. The major coat protein of viral particle (L1 plays a key role in the infection process. Our study aimed to isolate the HPV-16 L1 gene and analyze its sequence. Samples used were samples collected from the Department of Obstetrics and Gynaecology, Dr. Hasan Sadikin General Hospital, Bandung during the period of June to October 2010. In this study, the HPV-16 L1 sequence was analyzed from the viral deoxyribonucleic acid (DNA extracted from biopsy sample of cervical cancer patient biopsy samples.The HPV-16 L1 amplification was performed using the polymerase chain reaction with specific primer. The HPV infection in the cervical tissue was confirmed by commercial HPV genotyping test. The L1 fragment was cloned into plasmid and the insert of the recombinant clone pJET1.2/L1-16 was digested using BamHI and BgIII. The amplicon result showed HPV-16 L1 gene with a length of 1.595 base pairs. The sequence analysis of two samples using software BIOEDIT dan Basic Local Alignment Search Tool revealed a high level of sequence similarity to L1 HPV-16 from Thailand (99% and 97% as registered in GenBank. In conclusion, the L1 HPV-16 gene from Bandung isolates revealed variations from published sequence. Knowledge on L1 gene sequence may give additional information to the development of vaccine. Further study on vaccine development is currently ongoing using this HPV-16 clone that may be specific to Indonesian population.

  17. Construction of a recombinant adenovirus Vector of human papillomavirus type 16 L1_E7c

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Human papillomaviruses are closely associated with human cervical cancer, especially HPV types 16 and 18. At present, HPV can not be produced in large quantity; it also has tumorgenicity and these properties of HPV have seriously hampered the development of HPV vaccine. HPV type 16 L1 proteins can assembled into virus-like particles (VLP), which are morphologically identical to the nature virion. In order to develop the recombinant adenovirus vectors of HPV, we constructed a recombinant adenovirus shuttle plasmid pCA14 L1-E7c. Methods: Human papillomavirus type 16 L1 open reading frame without terminator codon (TAA) (5559- 7152) and E7c (682- 855) were amplified using PCR. The L1 and E7c fragments were inserted into pGEM-T easy vectors by T- A strategy, named pTAL1 and pTAE7c. pTAL1 was cut with Hind III and BglII, the pTAE7c with BamHI and ClaI. The L1 DNA fragment, E7c and pBluesscript SK were ligated together using T4 DNA ligase. pBSL1-E7c and pBSL1-E7c was digested with Hind III and Xhol. The L1-E7c fragment was inserted into adenovirus shuttle plasmids pCAl4, named pCAl4L1-E7c. DNA sequence results indicated that The L1-E7c DNA fragment can encode the HPV16L1-E7 fusion protein correctly. Results: The L1 and E7c DNA fragments were amplified by PCR and recombinant plasmid pTAL1, pTAE7c, pBSL1-E7c and pCA14L1-E7c were constructed correctly. The pCAl4L1-E7c can be used in the further research work, cotransfected the 293 cell with the parent adenovirus pBHG10. Conclusion: Our results indicated that we have constructed a HPV16L1-E7 fusion DNA fragments and the adenovirus shuttle plasmids pCALl-E7c for the further research.

  18. Activity of increased specific and non-specific esterases and glutathione transferases associated with resistance to permethrin in pediculus humanus capitis (phthiraptera: pediculidae) from Argentina.

    Science.gov (United States)

    Barrios, Silvia; Zerba, Eduardo; Picollo, Maria I; Audino, Paola Gonzalez

    2010-01-01

    Enhanced metabolism by oxidative enzymes is a major cause of pyrethroid resistance in insects. In this work, we evaluated the role of specific and non-specific esterases in head louse populations from Buenos Aires with different levels of resistance to permethrin. As esterase activity is substrate-dependent, four different esters were used as unspecific substrates in order to obtain a better characterization of the possible role of these enzymes in the resistance phenomenon. The unspecific substrates were phenylthioacetate, 1- and 2-naphtyl-acetate, and p-nitrophenyl acetate. A 7-coumaryl permethrate was synthesized and used as a specific substrate to measure pyrethroid esterases by a very sensitive microfluorometric method. The results on pyrethroid esterase activity obtained with this substrate showed that these enzymes contribute to the detoxifying activity in resistant populations, although no correlation was found between pyrethroid esterase activity and resistance ratios. In this study, we established that the activity of esterase against specific and non-specific substrates is increased in pyrethroid-resistant populations of head lice from Buenos Aires. Also, dichlorodiphenyltrichloroethane (DDT) resistance values demonstrated that there is a DDT cross-resistance phenomenon in pyrethroid-resistant head louse populations and suggested that an alteration in the receptor of the nervous system (kdr gen) is a key factor of the resistance phenomena in these head louse populations.

  19. Cloning, expression and characterization of a novel cold-active and organic solvent-tolerant esterase from Monascus ruber M7.

    Science.gov (United States)

    Guo, Hailun; Zhang, Yan; Shao, Yanchun; Chen, Wanping; Chen, Fusheng; Li, Mu

    2016-07-01

    Cold active esterases are a class of important biocatalysts that exhibit high activity at low temperatures. In this study, a search for putative cold-active esterase encoding genes from Monascus ruber M7 was performed. A cold-active esterase, named Lip10, was isolated, cloned, purified, and characterized. Amino acid sequence analysis reveals that Lip10 contained a conserved sequence motif Gly(173)-Xaa-Ser(175)-Xaa-Gly(177) that is also present in the majority of esterases and lipases. Phylogenetic analysis indicated that Lip10 was a novel microbial esterase. The lip10 gene was cloned and heterologously expressed in Escherichia coli BL21(DE3), resulting in the expression of an active and soluble protein that constituted 40 % of the total cell protein content. Lip10 maintained almost 50 % of its maximal activity at 4-10 °C, with optimal activity at 40 °C. Furthermore, Lip10 retained 184-216 % of its original activity, after incubation in 50 % (v/v) hydrophobic organic solvents for 24 h. The enzyme also exhibited high activity under alkaline conditions and good tolerance to metal ions in the reaction mixture. These results indicate that Lip10 may have potential uses in chemical synthesis and food processing industrial applications as an esterase.

  20. 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.

  1. Identification of a Marine Bacillus Strain C5 and Parathion-Methyl Degradation Characteristics of the Extracellular Esterase B1

    Directory of Open Access Journals (Sweden)

    Jianhua Hao

    2014-01-01

    Full Text Available A bacterial strain C5 that can produce new type of marine esterase was isolated and screened from marine sludge. According to 16S rRNA sequence analysis and physiological and biochemical experiments, the strain was identified as Bacillus subtilis. A single isozyme with a molecular weight of 86 kDa was observed by SDS-PAGE and native-PAGE. On this basis, the mechanism of esterase B1 secreted by strain C5 degrading parathion-methyl was explored, and the effects of temperature and pH on the degradation rate were investigated. From the results, p-nitrophenol was one of the degradation products of B1 degrading parathion-methyl, and the best degradation effect could be achieved at the temperature of 40°C and the neutral pH value.

  2. Development of Phonological Awareness in English-Mandarin Bilinguals: A Comparison of English-L1 and Mandarin-L1 Kindergarten Children

    Science.gov (United States)

    Yeong, Stephanie H. M.; Rickard Liow, Susan J.

    2012-01-01

    Phoneme awareness is critical for literacy acquisition in English, but relatively little is known about the early development of phonological awareness in ESL (English as a second language) bilinguals when their two languages have different phonological structures. Using parallel tasks in English and Mandarin, we tracked the development of L1

  3. 反思L1在SLA中的迁移现象%Reflection on the Transference of L1 in SLA

    Institute of Scientific and Technical Information of China (English)

    唐雪梅

    2006-01-01

    正确认识和处理L1在SLA中的作用,对英语教学非常重要.L1在SLA中的负迁移作用在我国的外语教学界被广泛地接受,汉语通常被认为是掌握外语的绊脚石.但是,由此产生的教学方法并不能很理想地提高学习者的L2水平.通过对L1在SLA中负迁移观点的反思,发现应该更多地去注意学习者脑海中语际语的本质,进一步更科学地了解L1在SLA中的作用,以便克服教学的盲目性,对症下药,推进教学改革.

  4. Quels changements linguistiques dans l'attrition de la L1 chez le bilingue tardif (What Linguistic Changes in the Attrition of the L1 in Late Bilinguals)?

    Science.gov (United States)

    Kopke, Barbara

    2001-01-01

    This article reports on a psycholinguistic study of first language attrition in German first generation immigrants. On the basis of the individual variation in performance evidenced by the data, the study claims that first language (L1) attrition in late bilinguals is not only the consequence of a lack of use. A comparison of the performance of…

  5. 人乳头瘤病毒衣壳蛋白L1的相关研究%Relative research on human papillomavirus L1 protein

    Institute of Scientific and Technical Information of China (English)

    卜璋于; 俞小虹

    2011-01-01

    人乳头瘤病毒(HPY)是引起尖锐湿疣等上皮乳头瘤样增生性疾病的主要病原。HPV衣壳蛋白L1,由于其结构特征及精确的抗原特性,在黏附宿主细胞、识别病毒受体、协助病毒DNA的人胞转运中均发挥重要作用,在HPV感染的临床评估和HPV疫苗的研究中日益受到关注。该文对HPV衣壳蛋白L1的结构和功能及其相关的应用进行了综述。%Human papillomavirus(HPV)is a main pathogen causing epithelial papillomatosis disease like condyloma acuminatum. HPV L1 protein plays an important role in adhering host cell, recognizing virus receptor and trafficking DNA to cellular nucleus due to its structure and antigenic character, and attack great concern is in the clinical evaluations of HPV infectivity and the researches of HPV vac cines. In the article, the structure and function of HPV L1 protein and its application are reviewed.

  6. Wnt/β-Catenin Signaling Mediated-UCH-L1 Expression in Podocytes of Diabetic Nephropathy

    Science.gov (United States)

    Zhang, Hongxia; Luo, Weili; Sun, Yonghong; Qiao, Yanchun; Zhang, Liying; Zhao, Zhilian; Lv, Shijun

    2016-01-01

    Increasing studies identified podocyte injury as a key early risk factor resulting in diabetic nephropathy (DN). The ubiquitin carboxy-terminal hydrolase 1 (UCH-L1) participates in podocyte differentiation and injury, which is elevated in the podocytes of a variety of nephritis. Whether UCH-L1 expression is positively related to podocyte injury of DN remains unclear. In this study, elevated expression of UCH-L1 and its intrinsic mechanism in high glucose (HG)-stimulated murine podocytes were investigated using western blot and real-time quantitative PCR. Kidney biopsies of DN patients and health individuals were stained by immunofluorescence (IF) method. The morphological and functional changes of podocytes were tested by F-actin staining and cell migration assay. Results demonstrated that HG induced upregulation of UCH-L1 and activation of the Wnt/β-catenin signaling pathway in podocytes. However, blocking of the Wnt pathway by dickkopf related protein 1 (DKK1) eliminated the above changes. Furthermore, IF staining confirmed that, compared with healthy individuals, the expression of UCH-L1 and β-catenin were obviously increased in kidney biopsy of DN patients. Overexpression of UCH-L1 remodeled its actin cytoskeleton, increased its cell migration and impacted its important proteins. All the findings manifested that Wnt/β-catenin/UCH-L1 may be a new potential therapy method in the treatment of DN in future. PMID:27571062

  7. Novel functions for the endocytic regulatory proteins MICAL-L1 and EHD1 in mitosis.

    Science.gov (United States)

    Reinecke, James B; Katafiasz, Dawn; Naslavsky, Naava; Caplan, Steve

    2015-01-01

    During interphase, recycling endosomes mediate the transport of internalized cargo back to the plasma membrane. However, in mitotic cells, recycling endosomes are essential for the completion of cytokinesis, the last phase of mitosis that promotes the physical separation the two daughter cells. Despite recent advances, our understanding of the molecular determinants that regulate recycling endosome dynamics during cytokinesis remains incomplete. We have previously demonstrated that Molecule Interacting with CasL Like-1 (MICAL-L1) and C-terminal Eps15 Homology Domain protein 1 (EHD1) coordinately regulate receptor transport from tubular recycling endosomes during interphase. However, their potential roles in controlling cytokinesis had not been addressed. In this study, we show that MICAL-L1 and EHD1 regulate mitosis. Depletion of either protein resulted in increased numbers of bi-nucleated cells. We provide evidence that bi-nucleation in MICAL-L1- and EHD1-depleted cells is a consequence of impaired recycling endosome transport during late cytokinesis. However, depletion of MICAL-L1, but not EHD1, resulted in aberrant chromosome alignment and lagging chromosomes, suggesting an EHD1-independent function for MICAL-L1 earlier in mitosis. Moreover, we provide evidence that MICAL-L1 and EHD1 differentially influence microtubule dynamics during early and late mitosis. Collectively, our new data suggest several unanticipated roles for MICAL-L1 and EHD1 during the cell cycle.

  8. Increased Oxidative Stress in Cultured 3T3-L1 Cells was Attenuated by Berberine Treatment.

    Science.gov (United States)

    Dong, Shi-Fen; Yasui, Naomi; Negishb, Hiroko; Kishimoto, Aya; Sun, Jian-Ning; Ikeda, Katsumi

    2015-06-01

    The 3T3-L1 cell line is one of the most well-characterized and reliable models for studying adipocytes. Increased oxidative stress in accumulated fat was found in 3T3-L1 cells. Berberine, an isoquinoline alkaloid, could suppress fat deposition in 3T3-L1 cells; however, whether berberine suppresses increased oxidative stress is not well known. In this study, we observed the effect of berberine on increased oxidative stress in 3T3-L1 cells. 3T3-L1 cells were cultured and treated with berberine (5-20 μM) from day 3 to day 8. We confirmed that berberine markedly inhibited fat accumulation and lipid droplets in 3T3-L1 adipocytes and decreased triglyceride content. Berberine inhibited increased oxidative stress in 3T3-L1 cells by suppressing reactive oxygen species (ROS) production, and increased glutathione peroxidase (GPx) gene expression and GPx activity. Berberine also markedly reduced adipokines secreted by adipocytes, including leptin and resistin.

  9. Mutation in the sixth immunoglobulin domain of L1CAM is associated with migrational brain anomalies

    Science.gov (United States)

    Shieh, Christine; Moser, Franklin; Graham, John M.; Watiker, Valerie

    2015-01-01

    Objective: To describe the phenotype of a patient with classical features of X-linked L1 syndrome associated with novel brain malformations. Methods: Diagnostic analysis included physical and dysmorphology examinations, MRI of the brain, and exome sequencing of the family trio. Results: We report a 2.5-year-old boy with developmental delay, dysmorphic facies, and adducted thumbs. MRI of the brain showed a truncated corpus callosum and periventricular heterotopias associated with polymicrogyria (PMG). Variant segregation analysis with exome sequencing discovered a novel maternally derived hemizygous variant in exon 14 of the L1CAM gene (c.1759 G>C; p.G587R). Conclusions: This novel L1CAM mutation was located in the protein's sixth immunoglobin domain and involved glycine-587, a key residue in the structure of L1CAM because of its interactions with lysine-606, which indicates that any mutation at this site would likely affect the secondary structure and function of the protein. The replacement of the small nonpolar glycine residue with a large basic arginine would have an even more dramatic result. The presentation of periventricular nodular heterotopias with overlying PMG is very uncommon, and its association with L1CAM may provide insight into other similar cases. Furthermore, this presentation indicates the important role that L1CAM plays in neuronal migration and brain development and extends the phenotype associated with L1CAM-associated disorders. PMID:27066571

  10. Reaction Mechanism for Cocaine Esterase-Catalyzed Hydrolyses of (+)- and (−)-Cocaine: Unexpected Common Rate-Determining Step

    OpenAIRE

    Liu, Junjun; Zhao, Xinyun; Yang, Wenchao; Zhan, Chang-Guo

    2011-01-01

    First-principles quantum mechanical/molecular mechanical (QM/MM)-free energy (FE) calculations have been performed to examine catalytic mechanism for cocaine esterase (CocE)-catalyzed hydrolysis of (+)-cocaine in comparison with CocE-catalyzed hydrolysis of (−)-cocaine. It has been shown that the acylation of (+)-cocaine consists of nucleophilic attack of hydroxyl group of Ser117 on carbonyl carbon of (+)-cocaine benzoyl ester and the dissociation of (+)-cocaine benzoyl ester. The first react...

  11. Identification of novel esterase-active enzymes from hot environments by use of the host bacterium Thermus thermophilus

    Directory of Open Access Journals (Sweden)

    Benedikt eLeis

    2015-04-01

    Full Text Available Functional metagenomic screening strategies, which are independent of known sequence information, can lead to the identification of truly novel genes and enzymes. Since E. coli has been used exhaustively for this purpose as a host, it is important to establish alternative expression hosts and to use them for functional metagenomic screening for new enzymes. In this study we show that Thermus thermophilus HB27 is an excellent screening host and can be used as an alternative provider of truly novel biocatalysts. In a previous study we constructed the mutant strain BL03 that was no longer able to grow on defined minimal medium supplemented with tributyrin as the sole carbon source and could be used as a host to screen for metagenomic DNA fragments that could complement growth on tributyrin. Several thousand single fosmid clones from thermophilic metagenomic libraries from heated compost and hot spring water samples were subjected to a comparative screening for esterase activity in both T. thermophilus strain BL03 and E. coli EPI300. We scored a greater number of active clones in the thermophilic bacterium than in the mesophilic E. coli. From all clones functionally screened in E. coli, only two thermostable α/β-fold hydrolase enzymes with high amino acid sequence similarity to already characterized enzymes were identifiable. In contrast, five further fosmids were found that conferred lipolytic activities in T. thermophilus. Four open reading frames (ORFs were found which did not share significant similarity to known esterase enzymes. Two of the genes were expressed in both hosts and the novel thermophilic esterases, which based on their primary structures could not be assigned to known esterase or lipase families, were purified and preliminarily characterized. Our work underscores the benefit of using additional screening hosts other than E. coli for the identification of novel biocatalysts with industrial relevance.

  12. Diversity of esterase isozyme in Aegilops tauschii Cosson%节节麦的酯酶同工酶分析

    Institute of Scientific and Technical Information of China (English)

    兰秀锦; 刘登才; 魏育明; 颜泽红; 郑有良

    2001-01-01

    The esterase isozyme of 30 accessions of Aegilops tauschii were studied by means of polyarylamide gel electrophoresis. The results showed significant difference of esterase in all of the four stages i.e. seeding,Shooting, flag leaf and young ear, which patterns can be divided into 15 types. Ten accessions from middle reaches of the Yellow River belonged to two patterns with a similarity coefficient 0. 984. Two accessions of Xinjiang belonged to one pattern and was different from that of middle reaches of Yellow River. No same esterase isozyme has been found in the four stages. It showed that esterase isozyme related to growing and development of plants.%对30份不同来源的节节麦进行4个时期的酯酶同工酶分析。结果表明:不同来源节节麦的酯酶同工酶存在较大差异,共分成15种基本类型。我国黄河流域的10份节节麦被划分为2个基本类型,但二者关系极为相近;新疆节节麦与之有一定差异,但在相似系数≤0.820时可视为一类。所有材料在4个时期之间没有出现一个完全相同的酶带类型,说明酯酶同工酶随发育时期而不断变化。

  13. Expression of Human Papillomavirus Type 16 L1 Protein in Transgenic Tobacco Plants

    Institute of Scientific and Technical Information of China (English)

    Hong-Li LIU; Wen-Sheng LI; Ting LEI; Jing ZHENG; Zheng ZHANG; Xiao-Fei YAN; Zhe-Zhi WANG; Yi-Li WANG; Lü-Sheng SI

    2005-01-01

    To develop a plant expression system for the production of the human papillomavirus type 16(HPV16) vaccine, we investigated whether the HPV16 L1 protein can be expressed in tobacco plants and whether it can be used as the cheapest form of edible vaccine. The HPV16 L1 coding sequence was amplified by PCR using specific primers from the plasmid pGEM-T-HPV16 containing the template sequence, and subcloned into the intermediate vector pUCmT and binary vector pBI121 consecutively to obtain the plant expression plasmid pBI-L1. The T-DNA regions of the pBI-L1 binary vector contained the constitutive Cauliflower mosaic virus (CaMV) 35S promoter and the neomycin phosphotransferase npt Ⅱ gene, which allowed the selection of transformed plants using kanamycin. The tobacco plants were transformed by cocultivating them, using the leaf disc method, with Agrobacterium tumefaciens LBA4404, which harbored the plant expression plasmid. The regenerated transgenic tobacco plants were selected using kanamycin, and confirmed by PCR. The results of the Southern blot assay also showed that the HPV16 L1 gene was integrated stably into the genome of the transformed tobacco plants. The Western blot analysis showed that the transformed tobacco leaves could express the HPV16 L1 protein. Furthermore, it was demonstrated by ELISA assay that the expressed protein accounted for 0.034%-0.076% of the total soluble leaf protein, was able to form 55 nm virus-like particles compatible with HPV virus-like particle (VLP), and induced mouse erythrocyte hemagglutination in vitro. The present results indicate that the HPV16 L1 protein can be expressed in transgenic tobacco plants and the expressed protein possesses the natural features of the HPV 16L1 protein, implying that the HPV16 L1 transgenic plants can be potentially used as an edible vaccine.

  14. Active form Notch4 promotes the proliferation and differentiation of 3T3-L1 preadipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Lai, Peng-Yeh [Institute of Molecular Biology and Department of Life Science, National Chung Cheng University, Chiayi 621, Taiwan, ROC (China); Tsai, Chong-Bin [Institute of Molecular Biology and Department of Life Science, National Chung Cheng University, Chiayi 621, Taiwan, ROC (China); Department of Ophthalmology, Chiayi Christian Hospital, Chiayi 600, Taiwan, ROC (China); Tseng, Min-Jen, E-mail: biomjt@ccu.edu.tw [Institute of Molecular Biology and Department of Life Science, National Chung Cheng University, Chiayi 621, Taiwan, ROC (China)

    2013-01-18

    Highlights: ► Notch4IC modulates the ERK pathway and cell cycle to promote 3T3-L1 proliferation. ► Notch4IC facilitates 3T3-L1 differentiation by up-regulating proadipogenic genes. ► Notch4IC promotes proliferation during the early stage of 3T3-L1 adipogenesis. ► Notch4IC enhances differentiation during subsequent stages of 3T3-L1 adipogenesis. -- Abstract: Adipose tissue is composed of adipocytes, which differentiate from precursor cells in a process called adipogenesis. Many signal molecules are involved in the transcriptional control of adipogenesis, including the Notch pathway. Previous adipogenic studies of Notch have focused on Notch1 and HES1; however, the role of other Notch receptors in adipogenesis remains unclear. Q-RT-PCR analyses showed that the augmentation of Notch4 expression during the differentiation of 3T3-L1 preadipocytes was comparable to that of Notch1. To elucidate the role of Notch4 in adipogenesis, the human active form Notch4 (N4IC) was transiently transfected into 3T3-L1 cells. The expression of HES1, Hey1, C/EBPδ and PPARγ was up-regulated, and the expression of Pref-1, an adipogenic inhibitor, was down-regulated. To further characterize the effect of N4IC in adipogenesis, stable cells expressing human N4IC were established. The expression of N4IC promoted proliferation and enhanced differentiation of 3T3-L1 cells compared with those of control cells. These data suggest that N4IC promoted proliferation through modulating the ERK pathway and the cell cycle during the early stage of 3T3-L1 adipogenesis and facilitated differentiation through up-regulating adipogenic genes such as C/EBPα, PPARγ, aP2, LPL and HSL during the middle and late stages of 3T3-L1 adipogenesis.

  15. Engineering the vaccinia virus L1 protein for increased neutralizing antibody response after DNA immunization

    Directory of Open Access Journals (Sweden)

    Moss Bernard

    2009-03-01

    Full Text Available Abstract Background The licensed smallpox vaccine, comprised of infectious vaccinia virus, has associated adverse effects, particularly for immunocompromised individuals. Therefore, safer DNA and protein vaccines are being investigated. The L1 protein, a component of the mature virion membrane that is conserved in all sequenced poxviruses, is required for vaccinia virus entry into host cells and is a target for neutralizing antibody. When expressed by vaccinia virus, the unglycosylated, myristoylated L1 protein attaches to the viral membrane via a C-terminal transmembrane anchor without traversing the secretory pathway. The purpose of the present study was to investigate modifications of the gene expressing the L1 protein that would increase immunogenicity in mice when delivered by a gene gun. Results The L1 gene was codon modified for optimal expression in mammalian cells and potential N-glycosylation sites removed. Addition of a signal sequence to the N-terminus of L1 increased cell surface expression as shown by confocal microscopy and flow cytometry of transfected cells. Removal of the transmembrane domain led to secretion of L1 into the medium. Induction of binding and neutralizing antibodies in mice was enhanced by gene gun delivery of L1 containing the signal sequence with or without the transmembrane domain. Each L1 construct partially protected mice against weight loss caused by intranasal administration of vaccinia virus. Conclusion Modifications of the vaccinia virus L1 gene including codon optimization and addition of a signal sequence with or without deletion of the transmembrane domain can enhance the neutralizing antibody response of a DNA vaccine.

  16. Identification and Characterization of MEDI4736, an Antagonistic Anti-PD-L1 Monoclonal Antibody.

    Science.gov (United States)

    Stewart, Ross; Morrow, Michelle; Hammond, Scott A; Mulgrew, Kathy; Marcus, Danielle; Poon, Edmund; Watkins, Amanda; Mullins, Stefanie; Chodorge, Matthieu; Andrews, John; Bannister, David; Dick, Emily; Crawford, Nicola; Parmentier, Julie; Alimzhanov, Marat; Babcook, John S; Foltz, Ian N; Buchanan, Andrew; Bedian, Vahe; Wilkinson, Robert W; McCourt, Matthew

    2015-09-01

    Programmed cell-death 1 ligand 1 (PD-L1) is a member of the B7/CD28 family of proteins that control T-cell activation. Many tumors can upregulate expression of PD-L1, inhibiting antitumor T-cell responses and avoiding immune surveillance and elimination. We have identified and characterized MEDI4736, a human IgG1 monoclonal antibody that binds with high affinity and specificity to PD-L1 and is uniquely engineered to prevent antibody-dependent cell-mediated cytotoxicity. In vitro assays demonstrate that MEDI4736 is a potent antagonist of PD-L1 function, blocking interaction with PD-1 and CD80 to overcome inhibition of primary human T-cell activation. In vivo MEDI4736 significantly inhibits the growth of human tumors in a novel xenograft model containing coimplanted human T cells. This activity is entirely dependent on the presence of transplanted T cells, supporting the immunological mechanism of action for MEDI4736. To further determine the utility of PD-L1 blockade, an anti-mouse PD-L1 antibody was investigated in immunocompetent mice. Here, anti-mouse PD-L1 significantly improved survival of mice implanted with CT26 colorectal cancer cells. The antitumor activity of anti-PD-L1 was enhanced by combination with oxaliplatin, which resulted in increased release of HMGB1 within CT26 tumors. Taken together, our results demonstrate that inhibition of PD-L1 function can have potent antitumor activity when used as monotherapy or in combination in preclinical models, and suggest it may be a promising therapeutic approach for the treatment of cancer. MEDI4736 is currently in several clinical trials both alone and in combination with other agents, including anti-CTLA-4, anti-PD-1, and inhibitors of IDO, MEK, BRAF, and EGFR.

  17. Characterization of new cell line stably expressing CHI3L1 oncogene

    Directory of Open Access Journals (Sweden)

    Chekhonin V. P.

    2011-06-01

    Full Text Available Aim. To characterize the immortalized 293 cell line after stable transfection with human oncogene (CHI3L1. Methods. 293 cells, stably transfected with pcDNA3.1_CHI3L1, and 293 cells, stably transfected with pcDNA3.1 as a negative control, were used throughout all experiments. The clones of CHI3L1-expressing 293 cells and 293 cells, transfected with pcDNA3.1, were analyzed by immunofluorescence and confocal microscopy. Cell proliferation was measured using MTT assay; analyses of ERK1/2 and AKT activation and their cellular localization were performed with anti-phospho-ERK and anti-phospho-AKT antibodies. Specific activation of MAP and PI3 kinases was measured by densitometric analysis of Western-blot signals. Results. The obtained results show quite modest ability of CHI3L1 to stimulate cell growth and reflect rather an improved cellular plating efficiency of the 293 cells stably transfected with pcDNA3.1_CHI3L1 as compared to the 293 cells transfected with an «empty» vector. ERK1/2 and AKT are activated in the 293_CHI3L1 cells. In these cells phosphorylated ERK1/2 were localized in both cell cytoplasm and nuclei while AKT only in cytoplasm. The 293_CHI3L1 cells differed from the 293 cells, transfected with an «empty» vector, in their size and ability to adhere to the culture plates. Conclusions. The overexpression of CHI3L1 is likely to have an important role in tumorigenesis via a mechanism which involves activation of PI3K and ERK1/2 pathways. The tumors which can be induced by orthotopic implantation of the transformed human cells with overexpressed human oncogene CHI3L1 into the rat brain can be used as a target for anticancer drug development.

  18. Human Papillomavirus Type16- L1 VLP Production in Insect Cells

    Directory of Open Access Journals (Sweden)

    Asghar Abdoli

    2013-08-01

    Full Text Available   Objective(s:  Infection by high-risk papillomavirus is regarded as the major risk factor in the development of cervical cancer. Recombinant DNA technology allows expression of the L1 major capsid protein of HPV in different expression systems, which has intrinsic capacity to self-assemble into viral-like particles (VLP. VLPS are non-infectious, highly immunogenic and can elicit neutralizing antibodies. VLP-based HPV vaccines can prevent persistent HPV infections and cervical cancer. In this study recombinant HPV-16 L1 protein was produced in Sf9 insect cells and VLP formation was confirmed. Materials and Methods: Complete HPV-16 L1 gene was inserted into pFast HTa plasmid and transformed into DH10BAC Escherichia coli containing bacmid and helper plasmid. The recombinant Bacmid colonies turned to white and non-recombinant colonies harboring L1 gene remained blue in the presence of X-gal and IPTG in colony selection strategy. To confirm the recombinant bacmid production, PCR was applied using specific L1 primers. To produce recombinant baculovirus, the recombinant bacmid DNA was extracted and transfected into Sf9 cells using Cellfectin. The expression of L1 in Sf9 cells was identified through SDS-PAGE and western blot analysis using specific L1 monoclonal antibody. Self-assembled HPV16L-VLPs in Sf9 cells was confirmed by electron microscopy. Results:The recombinant protein L1 was predominantly ~60 KD in SDS-PAGE with distinct immunoreactivity in western blot analysis and formed VLPS as confirmed by electron microscopy. Conclusion:Application of recombinant baculovirus containing HPV-16 L1 gene will certainly prove to be a constructive tool in production of VLPs for prophylactic vaccine development as well as diagnostic tests.

  19. HPV 6b L1 VIRUS-LIKE PARTICLES ELICIT HUMORAL IMMUNITY IN MICE

    Institute of Scientific and Technical Information of China (English)

    Liu Yuehua(刘跃华); Liu Wenjun(刘文军); Liu Xiaosong(刘晓松); Ian H.Frazer

    2003-01-01

    Objective. To test whether intrarnuscular,intranasal, intrarectal and intravaginal administration of HPV 6b L1 virus-like particles (VLPs) could induce immune response in mice and to assess whether intra muscular and mucosal vaccination against HPV is feasible. Methods. HPV6b L1 proteins self-assembled into VLPs in Sf-9 cell in vitro. Mice were immunized on day 0 and 21 with 50 μg HPV 6b L1 VLPs intramuscularly, intranasally, intrarectally and intravagi nally respectively. Sera were collected for testing IgG titer after a further 7 days and 3 months respec tively. Results. After immunizations, all mice developed significant anti-HPV 6b L1 antibody titers in serum by 7 days after the second immunization. The titer of the serum IgG antibody against HPV 6b L1 VLPs in the intramuscularly immunized group was higher than that in the intranasally, intrarectally and intravaginally immunized groups respectively, indicating that both muscular and mucosal administration of HPV 6b L1 VLPs can stimulate a systemic HPV-specific antibody response. Sera of the mice in the in tramuscularly immunized group still maintained a high titer of the serum IgG antibody against HPV 6b L1 VLPs 3 months after the immunization. Conclusion. The results demonstrated that the HPV 6b L1 VLPs maintain strong antigenicity. Immu nization with HPV 6b L1 VLPs via intramuscular and mucosal routes, without adjuvant, can elicit spe cific antibody in sera. These findings suggest that the VLPs are able to induce protective antibodies.

  20. L1+L2正则化逻辑斯蒂模型分类算法%Logistic Model Classification Algorithm via L1+L2 Regularization

    Institute of Scientific and Technical Information of China (English)

    刘建伟; 付捷; 罗雄麟

    2012-01-01

    提出一种L1+L2范数正则化逻辑斯蒂模型分类算法.该算法引入L2范数正则化,解决L1正则化逻辑斯蒂算法迭代过程奇异问题,通过引入样本向量的扩展和新的权值向量完成L1范数非平滑问题,最终使用共轭梯度方法求解经过转化的最优化问题.在各种实际数据集上的实验结果表明,该算法优于L2范数、L1范数和Lp范数正则化逻辑斯蒂模型,具有较好的特征选择和分类性能.%This paper proposes an L1+L2 norm regularized logistic model classification algorithm, and the singularity of iterative process in LI norm regularized logistic classification algorithm is solved by using L2 norm regularization. The non-smooth problem is transformed into smooth one via argumentation of vector of samples and introduction of new weight vector, and classification object function is solved using the conjugate gradient method. Performance of classification and feature selection on real datasets shows that the algorithm is better than L2 norm, LI nrom and Lp norm regularized logistic model.

  1. AFAP-1L1-mediated actin filaments crosslinks hinder Trypanosoma cruzi cell invasion and intracellular multiplication.

    Science.gov (United States)

    de Araújo, Karine Canuto Loureiro; Teixeira, Thaise Lara; Machado, Fabrício Castro; da Silva, Aline Alves; Quintal, Amanda Pifano Neto; da Silva, Claudio Vieira

    2016-10-01

    Host actin cytoskeleton polymerization has been shown to play an important role during Trypanosoma cruzi internalization into mammalian cell. The structure and dynamics of the actin cytoskeleton in cells are regulated by a vast number of actin-binding proteins. Here we aimed to verify the impact of AFAP-1L1, during invasion and multiplication of T. cruzi. Knocking-down AFAP-1L1 increased parasite cell invasion and intracellular multiplication. Thus, we have shown that the integrity of the machinery formed by AFAP-1L1 in actin cytoskeleton polymerization is important to hinder parasite infection.

  2. Ins-4 and daf-28 function redundantly to regulate C. elegans L1 arrest.

    Science.gov (United States)

    Chen, Yutao; Baugh, L Ryan

    2014-10-15

    Caenorhabditis elegans larvae reversibly arrest development in the first larval stage in response to starvation (L1 arrest or L1 diapause). Insulin-like signaling is a critical regulator of L1 arrest. However, the C. elegans genome encodes 40 insulin-like peptides, and it is unknown which peptides participate in nutritional control of L1 development. Work in other contexts has revealed that insulin-like genes can promote development ("agonists") or developmental arrest ("antagonists"), suggesting that such agonists promote L1 development in response to feeding. We measured mRNA expression dynamics with high temporal resolution for all 40 insulin-like genes during entry into and recovery from L1 arrest. Nutrient availability influences expression of the majority of insulin-like genes, with variable dynamics suggesting complex regulation. We identified thirteen candidate agonists and eight candidate antagonists based on expression in response to nutrient availability. We selected ten candidate agonists (daf-28, ins-3, ins-4, ins-5, ins-6, ins-7, ins-9, ins-26, ins-33 and ins-35) for further characterization in L1 stage larvae. We used destabilized reporter genes to determine spatial expression patterns. Expression of candidate agonists is largely overlapping in L1 stage larvae, suggesting a role of the intestine, chemosensory neurons ASI and ASJ, and the interneuron PVT in control of L1 development. Transcriptional regulation of candidate agonists is most significant in the intestine, as if internal nutrient status is a more important influence on transcription than sensory perception. Phenotypic analysis of single and compound deletion mutants did not reveal effects on L1 developmental dynamics, though simultaneous disruption of ins-4 and daf-28 increases survival of L1 arrest. Furthermore, overexpression of ins-4, ins-6 or daf-28 alone decreases survival and promotes cell division during starvation. These results suggest extensive functional overlap among insulin

  3. L1/ℓ1-Gain analysis and synthesis of Markovian jump positive systems with time delay.

    Science.gov (United States)

    Zhang, Junfeng; Zhao, Xudong; Zhu, Fubo; Han, Zhengzhi

    2016-07-01

    This paper is concerned with stability analysis and control synthesis of Markovian jump positive systems with time delay. The notions of stochastic stability with L1- and ℓ1-gain performances are introduced for continuous- and discrete-time contexts, respectively. Using a stochastic copositive Lyapunov function, sufficient conditions for the stability with L1/ℓ1-gain performance of the systems are established. Furthermore, mode-dependent controllers are designed to achieve the stabilization with L1/ℓ1-gain of the resulting closed-loop systems. All proposed conditions are formulated in terms of linear programming. Numerical examples are provided to verify the effectiveness of the findings of theory.

  4. Effects of high hydrostatic pressure and temperature increase on Escherichia coli spp. and pectin methyl esterase inactivation in orange juice.

    Science.gov (United States)

    Torres, E F; González-M, G; Klotz, B; Rodrigo, D

    2016-03-01

    The aim of this study was to evaluate the effect of high hydrostatic pressure treatment combined with moderate processing temperatures (25 ℃-50 ℃) on the inactivation of Escherichia coli O157: H7 (ATCC 700728), E. coli K12 (ATCC 23716), and pectin methyl esterase in orange juice, using pressures of 250 to 500 MPa with times ranging between 1 and 30 min. Loss of viability of E. coli O157:H7 increased significantly as pressure and treatment time increased, achieving a 6.5 log cycle reduction at 400 MPa for 3 min at 25 ℃ of treatment. With regard to the inactivation of pectin methyl esterase, the greatest reduction obtained was 90.05 ± 0.01% at 50 ℃ and 500 MPa of pressure for 15 min; therefore, the pectin methyl esterase enzyme was highly resistant to the treatments by high hydrostatic pressure. The results obtained in this study showed a synergistic effect between the high pressure and moderate temperatures in inactivating E. coli cells.

  5. Identification of a novel carbohydrate esterase from Bjerkandera adusta: structural and function predictions through bioinformatics analysis and molecular modeling.

    Science.gov (United States)

    Cuervo-Soto, Laura I; Valdés-García, Gilberto; Batista-García, Ramón; del Rayo Sánchez-Carbente, María; Balcázar-López, Edgar; Lira-Ruan, Verónica; Pastor, Nina; Folch-Mallol, Jorge Luis

    2015-03-01

    A new gene from Bjerkandera adusta strain UAMH 8258 encoding a carbohydrate esterase (designated as BacesI) was isolated and expressed in Pichia pastoris. The gene had an open reading frame of 1410 bp encoding a polypeptide of 470 amino acid residues, the first 18 serving as a secretion signal peptide. Homology and phylogenetic analyses showed that BaCesI belongs to carbohydrate esterases family 4. Three-dimensional modeling of the protein and normal mode analysis revealed a breathing mode of the active site that could be relevant for esterase activity. Furthermore, the overall negative electrostatic potential of this enzyme suggests that it degrades neutral substrates and will not act on negative substrates such as peptidoglycan or p-nitrophenol derivatives. The enzyme shows a specific activity of 1.118 U mg(-1) protein on 2-naphthyl acetate. No activity was detected on p-nitrophenol derivatives as proposed from the electrostatic potential data. The deacetylation activity of the recombinant BaCesI was confirmed by measuring the release of acetic acid from several substrates, including oat xylan, shrimp shell chitin, N-acetylglucosamine, and natural substrates such as sugar cane bagasse and grass. This makes the protein very interesting for the biofuels production industry from lignocellulosic materials and for the production of chitosan from chitin.

  6. 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.

  7. 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.

  8. 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.

  9. INTERRELATION BETWEEN 3-HYDROXY-1,4-BENZODIAZEPINE-2-ONE ESTERS STRUCTURE ON THEIR HYDROLYSIS BY CARBOXYL ESTERASE

    Directory of Open Access Journals (Sweden)

    E. A. Shesterenko

    2013-04-01

    Full Text Available The synthesis of new series of 7-bromo-5-aryl-1,2-dihydro-3H-1,4-benzdiazepine-2-one derivatives, containing in the tree position phthalimidoacyl and hexylacyl fragments was accomplished. The structure of new compounds was proved by mass-spectrometry and PMRspectroscopy methods. For the first time, hydrolysis of the earlier synthesized 3-hydroxy-7-bromo-5-aryl-1,2-dihydro-3H-1,4-benzdiazepine-2-one esters, potential anxiolytic and hypnotic means, catalyzed by carboxyl esterase in composition of pig liver microsomal fraction was studied. The quantitative inhibition of pig liver microsomal fraction esterase activity in the presence of carboxyl esterase selective inhibitor di-(pnitrophenyl-phosphate was shown. The nonlinear dependence both of hydrolysis degree with acyl moiety length in 3-acyloxy-7-bromo-5-aryl-1,2-dihydro-3H-1,4-benzdiazepine-2-ones and decreased substrate transformation degree after substituent introduction in the first position of molecule was established. For the derivatives with phthalimidoacyl and hexylacyl-mo ie ties in the molecule 3 position it was shown, that increasing of CH2-groups number in this substituents and incorporation of chlorine atom in o-position of phenyl ring bring to increasing of hydrolysis degree.

  10. Screening, purification, and characterization of a novel organic solvent-tolerant esterase, Lip2, from Monascus purpureus strain M7.

    Science.gov (United States)

    Kang, Li-Jing; Meng, Zi-Tong; Hu, Chen; Zhang, Yan; Guo, Hai-Lun; Li, Qing; Li, Mu

    2017-03-01

    Organic solvent-tolerant esterases are proven to be excellent biocatalysts in chemical and pharmaceutical industries. A novel organic solvent-tolerant esterase gene, lip2, was isolated from filamentous fungi Monascus purpureus M7. The sequence analysis suggested that lip2 has a conserved "GDSL" motif near the active center. The multiple-sequence alignment and phylogenetic analysis revealed that Lip2 displayed two unique amino-acid sequence motifs that clearly separate it from any other previously described lipase family. After incubation in 20% methanol and ethanol for 3 h, the Lip2 displayed 190 and 180% residual activities, respectively. It retained 99-110% relative activity in 20% (v/v) hydrophilic organic solvents after incubation for 1 day. This esterase showed optimal activity at 40 °C and retained about 70% maximal activity at 60 °C. The enzyme also displayed more than 50% residual activity over a range of pH 5-11. In the presence of most of metal ions or additives, Lip2 retained most of the activity. These unique properties of Lip2 make it a promising as biocatalyst for industrial processes.

  11. 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

  12. Robust L1 filtering with pole constraint in a disk via parameter-dependent Lyapunov functions

    Institute of Scientific and Technical Information of China (English)

    Li Yanhui; Wen Qiyong; Wang Junling; Wang Changhong; Gao Huijun

    2005-01-01

    The problem of robust L1 filtering with pole constraint in a disk for linear continuous polytopic uncertain systems is discussed. The attention is focused on design a linear asymptotically stable filter such that the filtering error system remains robustly stable, and has a L1 performance constraint and pole constraint in a disk. The new robust L1 performance criteria and regional pole placement condition are obtained via parameter-dependent Lyapunov functions method. Upon the proposed multiobjective performance criteria and by means of LMI technique, both full-order and reducedorder robust L1 filter with suitable dynamic behavior can be obtained from the solution of convex optimization problems.Compared with earlier result in the quadratic framework, this approach turns out to be less conservative. The efficiency of the proposed technique is demonstrated by a numerical example.

  13. Exactly Recovering Low-Rank Matrix in Linear Time via $l_1$ Filter

    CERN Document Server

    Liu, Risheng; Su, Zhixun

    2011-01-01

    Recovering a low rank matrix from corrupted data, which is known as Robust PCA, has attracted considerable interests in recent years. This problem can be exactly solved by a combined nuclear norm and $l_1$ norm minimization. However, due to the computational burden of SVD inherent with the nuclear norm minimization, traditional methods suffer from high computational complexity, especially for large scale datasets. In this paper, inspired by digital filtering idea in image processing, we propose a novel algorithm, named $l_1$ Filter, for solving Robust PCA with linear cost. The $l_1$ Filter is defined by a seed, which is a exactly recovered small submatrix of the underlying low rank matrix. By solving some $l_1$ minimization problems in parallel, the full low rank matrix can be exactly recovered from corrupted observations with linear cost. Both theoretical analysis and experimental results exhibit that our method is an efficient way to exactly recovering low rank matrix in linear time.

  14. A study of the universal threshold in the L1 recovery by statistical mechanics

    CERN Document Server

    Takeda, Koujin

    2012-01-01

    We discuss the universality of the L1 recovery threshold in compressed sensing. Previous studies in the fields of statistical mechanics and random matrix integration have shown that L1 recovery under a random matrix with orthogonal symmetry has a universal threshold. This indicates that the threshold of L1 recovery under a non-orthogonal random matrix differs from the universal one. Taking this into account, we use a simple random matrix without orthogonal symmetry, where the random entries are not independent, and show analytically that the threshold of L1 recovery for such a matrix does not coincide with the universal one. The results of an extensive numerical experiment are in good agreement with the analytical results, which validates our methodology. Though our analysis is based on replica heuristics in statistical mechanics and is not rigorous, the findings nevertheless support the fact that the universality of the threshold is strongly related to the symmetry of the random matrix.

  15. Assay for the pattern recognition molecule collectin liver 1 (CL-L1)

    DEFF Research Database (Denmark)

    Axelgaard, Esben; Jensenius, Jens Christian; Thiel, Steffen

    Collectin liver 1 (also termed collectin 10 and CL-L1) is a C-type lectin that functions as a pattern recognition molecule (PRM) in the innate immune system1. We have produced antibodies against CL-L1 and have developed a sandwich-type time-resolved immuno-fluorometric assay (TRIFMA...... to co-purify with MASPs, possibly rendering it a role in complement. CL-L1 showed binding activity towards mannose-TSK beads in a Ca2+-dependent manner. This binding could be inhibited by mannose and glucose, but not by galactose, indicating that CL-L1 binds via its carbohydrate-recognition domain (CRD)....

  16. EX1504L1 Water Column Summary Report and Profile Data Collection

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — A complete set of water column profile data and CTD Summary Report (if generated) generated by the Okeanos Explorer during EX1504L1: CAPSTONE NWHI & Johnston...

  17. Polymorphic L1 retrotransposons are frequently in strong linkage disequilibrium with neighboring SNPs.

    Science.gov (United States)

    Higashino, Saneyuki; Ohno, Tomoyuki; Ishiguro, Koichi; Aizawa, Yasunori

    2014-05-10

    L1 retrotransposons have been the major driver of structural variation of the human genome. L1 insertion polymorphism (LIP)-mediated genomic variation can alter the transcriptome and contribute to the divergence of human phenotypes. To assess this possibility, a genome-wide association study (GWAS) including LIPs is required. Toward this ultimate goal, the present study examined linkage disequilibrium between six LIPs and their neighboring single nucleotide polymorphisms (SNPs). Genomic PCR and sequencing of L1-plus and -minus alleles from different donors revealed that all six LIPs were in strong linkage disequilibrium with at least one SNP. In addition, comparison of syntenic regions containing the identified SNP nucleotides was performed among modern humans (L1-plus and -minus alleles), archaic humans and non-human primates, revealing two different evolutionary schemes that might have resulted in the observed strong SNP-LIP linkage disequilibria. This study provides an experimental framework and guidance for a future SNP-LIP integrative GWAS.

  18. A Review of Fast l1-Minimization Algorithms for Robust Face Recognition

    CERN Document Server

    Yang, Allen Y; Zhou, Zihan; Sastry, S Shankar; Ma, Yi

    2010-01-01

    l1-minimization refers to finding the minimum l1-norm solution to an underdetermined linear system b=Ax. It has recently received much attention, mainly motivated by the new compressive sensing theory that shows that under quite general conditions the minimum l1-norm solution is also the sparsest solution to the system of linear equations. Although the underlying problem is a linear program, conventional algorithms such as interior-point methods suffer from poor scalability for large-scale real world problems. A number of accelerated algorithms have been recently proposed that take advantage of the special structure of the l1-minimization problem. In this paper, we provide a comprehensive review of five representative approaches, namely, Gradient Projection, Homotopy, Iterative Shrinkage-Thresholding, Proximal Gradient, and Augmented Lagrange Multiplier. The work is intended to fill in a gap in the existing literature to systematically benchmark the performance of these algorithms using a consistent experimen...

  19. EX1402L1 Water Column Summary Report and Profile Data Collection

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — A complete set of water column profile data and CTD Summary Report (if generated) generated by the Okeanos Explorer during EX1402L1: Gulf of Mexico Mapping and...

  20. EX1503L1 Water Column Summary Report and Profile Data Collection

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — A complete set of water column profile data and CTD Summary Report (if generated) generated by the Okeanos Explorer during EX1503L1: Tropical Exploration (Mapping...

  1. EX1202L1 Water Column Summary Report and Profile Data Collection

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — A complete set of water column profile data and CTD Summary Report (if generated) generated by the Okeanos Explorer during EX1202L1: Gulf of Mexico Exploration...

  2. EX1605L1 Water Column Summary Report and Profile Data Collection

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — A complete set of water column profile data and CTD Summary Report (if generated) generated by the Okeanos Explorer during EX1605L1: CAPSTONE CNMI & Mariana...

  3. Batch gradient method with smoothing L1/2 regularization for training of feedforward neural networks.

    Science.gov (United States)

    Wu, Wei; Fan, Qinwei; Zurada, Jacek M; Wang, Jian; Yang, Dakun; Liu, Yan

    2014-02-01

    The aim of this paper is to develop a novel method to prune feedforward neural networks by introducing an L1/2 regularization term into the error function. This procedure forces weights to become smaller during the training and can eventually removed after the training. The usual L1/2 regularization term involves absolute values and is not differentiable at the origin, which typically causes oscillation of the gradient of the error function during the training. A key point of this paper is to modify the usual L1/2 regularization term by smoothing it at the origin. This approach offers the following three advantages: First, it removes the oscillation of the gradient value. Secondly, it gives better pruning, namely the final weights to be removed are smaller than those produced through the usual L1/2 regularization. Thirdly, it makes it possible to prove the convergence of the training. Supporting numerical examples are also provided.

  4. TES/Aura L1B Spectra Special Observation Low Resolution V004

    Data.gov (United States)

    National Aeronautics and Space Administration — The L1B Low Resolution granule consists of radiometrically calibrated spectra & associated NESR, observed at 0.1 cm-1 resolution for a Special Observation &...

  5. TES/Aura L1B Spectra Special Observation Low Resolution V003

    Data.gov (United States)

    National Aeronautics and Space Administration — The L1B Low Resolution granule consists of radiometrically calibrated spectra & associated NESR, observed at 0.1 cm-1 resolution for a Special Observation &...

  6. TES/Aura L1B Spectra Special Observation Low Resolution V005

    Data.gov (United States)

    National Aeronautics and Space Administration — The L1B Low Resolution granule consists of radiometrically calibrated spectra & associated NESR, observed at 0.1 cm-1 resolution for a Special Observation &...

  7. The effects of Ganoderma lucidum herba pharmacopuncture on 3T3-L1 preadipocyte differentiation

    Directory of Open Access Journals (Sweden)

    Chea-woo Lee

    2008-09-01

    Full Text Available Objective : The purpose of this study is to investigate the effects of Ganoderma lucidum herba pharmacopuncture (GHP on the adipogenesis in 3T3-L1 preadipocytes. Methods : 3T3- L1 preadipocytes were differentiated with adipogenic reagents by incubating for 2 days in the absence or presence of GHP ranging from 1 and 2%. The effect of GHP on cell proliferation of 3T3-L1 preadipocytes was investigated using MTT assay. The effect of GHP on adipogenesis was examined by Oil red O staining and measuring glycerol-3-phosphate dehydrogenase (GPDH and intracellular triglyceride (TG content. Results : Following results were obtained from the preadipocyte proliferation and adipocyte differentiation of 3T3-L1. We observed no effect of GHP on preadipocyte proliferation. GHP inhibited adipogenesis, the activity of GPDH and accumulation of intracellular TG content. Conclusions : These results suggest that GHP inhibit differentiation of preadipocyte.

  8. Approximation methods of mixed l 1/H2 optimization problems for MIMO discrete-time systems

    Institute of Scientific and Technical Information of China (English)

    李昇平

    2004-01-01

    The mixed l1/H2 optimization problem for MIMO (multiple input-multiple output) discrete-time systems is eonsidered. This problem is formulated as minimizing the l1-norm of a dosed-loop transfer matrix while maintaining the H2-norm of another closed-loop transfer matrix at prescribed level. The continuity property of the optimal value in respect to changes in the H2-norm constraint is studied. The existence of the optimal solutions of mixed l1/H2 problem is proved. Becatse the solution of the mixed l1/H2 problem is based on the scaled-Q method, it avoids the zero interpolation difficulties. The convergent upper and lower bounds can be obtained by solving a sequence of finite dimensional nonlinear programming for which many efficient numerical optimization algorithms exist.

  9. EX1004L1 Water Column Summary Report and Profile Data Collection

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — A complete set of water column profile data and CTD Summary Report (if generated) generated by the Okeanos Explorer during EX1004L1: Exploration Indonesia - Guam to...

  10. Hydrography - MO 2014 Class L1 Lake Watersheds WQS TableG (SHP)

    Data.gov (United States)

    NSGIC GIS Inventory (aka Ramona) — This feature class contains watersheds for Class L1 lakes listed in Table G - Lake Classifications and Use Designations of the Water Quality Standards rule published...

  11. EX1404L1 Water Column Summary Report and Profile Data Collection

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — A complete set of water column profile data and CTD Summary Report (if generated) generated by the Okeanos Explorer during EX1404L1: Shakedown and Mapping, NE...

  12. TES/Aura L1B Spectra Special Observation High Resolution V004

    Data.gov (United States)

    National Aeronautics and Space Administration — A L1B High Resolution granule consists of radiometrically calibrated spectra & associated NESR, observed at 0.025 cm-1 resolution for a Special Observation &...

  13. Bayesian Network Classifier Based on L1 Regularization%基于L1正则化的贝叶斯网络分类器

    Institute of Scientific and Technical Information of China (English)

    王影; 王浩; 俞奎; 姚宏亮

    2012-01-01

    Variable order-based Bayesian network classifiers ignore the information of the selected variables in their sequence and their class label, which significantly hurts the classification accuracy. To address this problem, we proposed a simple and efficient LI regularized Bayesian network classifier (Ll-BNC). Through adjusting the constraint value of Lasso and fully taking advantage of the regression residuals of the information, Ll-BNC takes the information of the sequence of selected variables and the class label into account,and then generates an excellent variable ordering sequence (LI regularization path) for constructing a good Bayesian network classifier by the K2 algorithm. Experimental results show that Ll-BNC outperforms existing state-of-the-art Bayesian network classifiers. In addition, in comparison with SVM.Knn and J48 classification algorithms,Ll-BNC is also superior to those algorithms on most datasets.%目前基于节点排序的贝叶斯网络分类器忽略了节点序列中已选变量和类标签之间的信息,导致分类器的准确率很难进一步提高.针对这个问题,提出了一种简单高效的贝叶斯网络分类器的学习算法:L1正则化的贝叶斯网络分类器(L1-BNC).通过调整Lasso方法中的约束值,充分利用回归残差的信息,结合点序列中已选变量和类标签的信息,形成一条优秀的有序变量拓扑序列(L1正则化路径);基于该序列,利用K2算法生成优良的贝叶斯网络分类器.实验表明,L1-BNC在分类精度上优于已有的贝叶斯网络分类器.L1-BNC也与SVM,KNN和J48分类算法进行了比较,在大部分数据集上,L1-BNC优于这些算法.

  14. Characterization of an upstream regulatory element of adenovirus L1 poly (A) site.

    Science.gov (United States)

    Liu, Li

    2005-06-20

    The transition from early to late stage infection by adenovirus involves a change in mRNA expression from the adenovirus major late transcription unit (AdMLTU). This early to late switch centers around alternative selection of one of five poly (A) sites (L1-L5) that code for the major structural proteins of Adenovirus. During the early stage of infection, steady state mRNA is primarily derived from the L1 poly (A) site. During the late stage of infection, each of the MLTU poly (A) sites is represented in the steady state mRNA pool (Falck-Pedersen, E., Logan, J., 1989. Regulation of poly(A) site selection in adenovirus. J. Virol. 63 (2), 532-541.). Using transient transfection of a plasmid expressing Chloramphenicol Acetyl Transferase with a tandem poly (A) minigene system (L13) (DeZazzo, J.D., Falck-Pedersen, E., Imperiale, M.J., 1991. Sequences regulating temporal poly(A) site switching in the adenovirus major late transcription unit. Mol. Cell. Biol. 11 (12), 5977-5984; Prescott, J., Falck-Pedersen, E., 1994. Sequence elements upstream of the 3' cleavage site confer substrate strength to the adenovirus L1 and L3 polyadenylation sites. Mol. Cell. Biol. 14 (7), 4682-4693.), it has been demonstrated that the promoter-proximal L1 poly (A) site which is poorly recognized by the 3' end processing machinery, contains an upstream repressor element (URE) that influences steady state levels of mRNA (Prescott, J.C., Liu, L., Falck-Pedersen, E., 1997. Sequence-mediated regulation of adenovirus gene expression by repression of mRNA accumulation. Mol. Cell. Biol. 17 (4), 2207-2216.). In this study, we have further characterized the elements that mediate L1URE function. These studies indicate that the L1 upstream regulatory element (L1 URE) contains a complex RNA architecture that serves to repress gene expression through multiple sub-effectors. The L1URE functions when located upstream of a heterologous poly (A) site, and is able to strongly suppress steady state m

  15. Uncoupling of 3T3-L1 gene expression from lipid accumulation during adipogenesis

    OpenAIRE

    Temple, Karla A.; Basko, Xheni; Allison, Margaret B.; Brady, Matthew J.

    2007-01-01

    Adipocyte differentiation comprises altered gene expression and increased triglyceride storage. To investigate the interdependency of these two events, 3T3-L1 cells were differentiated in the presence of glucose or pyruvate. All adipocytic proteins examined were similarly increased between the two conditions. In contrast, 3T3-L1 adipocytes differentiated with glucose exhibited significant lipid accumulation, which was largely suppressed in the presence of pyruvate. Subsequent addition of gluc...

  16. High-dose Resveratrol Inhibits Insulin Signaling Pathway in 3T3-L1 Adipocytes

    OpenAIRE

    Lee, Haemi; Kim, Jae-Woo

    2013-01-01

    Background Insulin resistance is a major factor in the development of metabolic syndrome and is associated with central obesity and glucose intolerance. Resveratrol, a polyphenol found in fruits, has been shown to improve metabolic conditions. Although it has been widely studied how resveratrol affects metabolism, little is known about how resveratrol regulates lipogenesis with insulin signaling in 3T3-L1 adipocytes. Methods: We treated differentiated 3T3-L1 adipocytes with resveratrol to obs...

  17. L1 Use in EFL Classes with English-only Policy: Insights from Triangulated Data

    Directory of Open Access Journals (Sweden)

    Seyyed Hatam Tamimi Sa’d

    2015-06-01

    Full Text Available This study examines the role of the use of the L1 in EFL classes from the perspective of EFL learners. The triangulated data were collected using class observations, focus group semi-structured interviews and the learners’ written reports of their perceptions and attitudes in a purpose-designed questionnaire. The participants consisted of sixty male Iranian EFL learners who constituted three classes. The results indicated a strong tendency among the participants toward L1 and its positive effects on language learning; while only a minority of the learners favoured an English-only policy, the majority supported the judicious, limited and occasional use of the L1, particularly on the part of the teacher. The participants mentioned the advantages as well as the disadvantages of the use/non-use of the L1. While the major advantage and the main purpose of L1 use was said to be the clarification and intelligibility of instructions, grammatical and lexical items, the main advantages of avoiding it were stated as being the improvement of speaking and listening skills, aximizing learners’ exposure to English and their becoming accustomed to it. The study concludes that, overall and in line with the majority of the previous research studies, a judicious, occasional and limited use of the L1 is a better approach to take in EFL classes than to include or exclude it totally. In conclusion, a re-examination of the English-only policy and a reconsideration of the role of the L1 are recommended. Finally, the commonly held assumption that L1 is a hindrance and an impediment to the learners’ language learning is challenged.

  18. Intonational differences between L1 and L2 english in South Africa.

    Science.gov (United States)

    Swerts, Marc; Zerbian, Sabine

    2010-01-01

    Previous studies have shown that characteristics of a person's first language (L1) may transfer to a second language (L2). The current study looks at the extent to which this holds for aspects of intonation as well. More specifically, we investigate to what extent traces of the L1 can be discerned in the way intonation is used in the L2 for two functions: (1) to highlight certain words by making them sound more prominent and (2) to signal continuation or finality in a list by manipulating the speech melody. To this end, the article presents an explorative study into the way focus and boundaries are marked prosodically in Zulu, and it also compares such prosodic functions in two variants of English in South Africa, i.e., English spoken as an L1, and English spoken as an L2/additional language by speakers who have Zulu as their L1. The latter language is commonly referred to as Black South African English. This comparison is interesting from a typological perspective, as Zulu is intonationally different from English, especially in the way prosody is exploited for signalling informationally important stretches of speech. Using a specific elicitation procedure, we found in a first study that speakers of South African English (as L1) mark focused words and position within a list by intonational means, just as in other L1 varieties of English, whereas Zulu only uses intonation for marking continuity or finality. A second study focused on speakers of Black South African English, and compared the prosody of proficient versus less proficient speakers. We found that the proficient speakers were perceptually equivalent to L1 speakers of English in their use of intonation for marking focus and boundaries. The less proficient speakers marked boundaries in a similar way as L1 speakers of English, but did not use prosody for signalling focus, analogous to what is typical of their native language. Acoustic observations match these perceptual results.

  19. Nebulized C1-Esterase Inhibitor does not Reduce Pulmonary Complement Activation in Rats with Severe Streptococcus Pneumoniae Pneumonia.

    Science.gov (United States)

    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

    2016-12-01

    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 lung injury and lung inflammation. This was investigated in a rat model of severe Streptococcus pneumoniae pneumonia. Rats were intra-tracheally challenged with S. pneumoniae to induce pneumonia. Nebulized C1-esterase inhibitor or saline (control animals) was repeatedly administered to rats, 30 min before induction of pneumonia and every 6 h thereafter. Rats were sacrificed 20 or 40 h after inoculation with bacteria. Brochoalveolar lavage fluid and lung tissue were obtained for measuring levels of complement activation (C4b/c), lung injury and inflammation. Induction of pneumonia was associated with pulmonary complement activation (C4b/c at 20 h 1.24 % [0.56-2.59] and at 40 h 2.08 % [0.98-5.12], compared to 0.50 % [0.07-0.59] and 0.03 % [0.03-0.03] in the healthy control animals). The functional fraction of C1-INH was detectable in BALF, but no effect was found on pulmonary complement activation (C4b/c at 20 h 0.73 % [0.16-1.93] and at 40 h 2.38 % [0.54-4.19]). Twenty hours after inoculation, nebulized C1-esterase inhibitor treatment reduced total histology score, but this effect was no longer seen at 40 h. Nebulized C1-esterase inhibitor did not affect other markers of lung injury or lung inflammation. In this negative experimental animal study, severe S. pneumoniae pneumonia in rats is associated with pulmonary complement activation. Repeated treatment with nebulized C1-esterase inhibitor, although successfully delivered to the lungs, does not affect pulmonary complement activation, lung inflammation or lung injury.

  20. An increase in pectin methyl esterase activity accompanies dormancy breakage and germination of yellow cedar seeds.

    Science.gov (United States)

    Ren, C; Kermode, A R

    2000-09-01

    Pectin methyl esterase (PME) (EC 3.1.1.11) catalyzes the hydrolysis of methylester groups of cell wall pectins. We investigated the role of this enzyme in dormancy termination and germination of yellow cedar (Chamaecyparis nootkatensis [D. Don] Spach) seeds. PME activity was not detected in dormant seeds of yellow cedar but was induced and gradually increased during moist chilling; high activity coincided with dormancy breakage and germination. PME activity was positively correlated to the degree of dormancy breakage of yellow cedar seeds. The enzyme produced in different seed parts and in seeds at different times during moist chilling, germination, and early post-germinative growth consisted of two isoforms, both basic with isoelectric points of 8.7 and 8.9 and the same molecular mass of 62 kD. The pH optimum for the enzyme was between 7.4 and 8.4. In intact yellow cedar seeds, activities of the two basic isoforms of PME that were induced in embryos and in megagametophytes following dormancy breakage were significantly suppressed by abscisic acid. Gibberellic acid had a stimulatory effect on the activities of these isoforms in embryos and megagametophytes of intact seeds at the germinative stage. We hypothesize that PME plays a role in weakening of the megagametophyte, allowing radicle emergence and the completion of germination.

  1. 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.

  2. Diagnosis of spontaneous bacterial peritonitis: An update on leucocyte esterase reagent strips

    Institute of Scientific and Technical Information of China (English)

    Anastasios Koulaouzidis

    2011-01-01

    Ascites remain the commonest complication of decom-pensated cirrhosis. Spontaneous bacterial peritonitis (SBP) is defined as the infection of ascitic fluid (AF) in the ab-sence of a contiguous source of infection and/or an intra-abdominal inflammatory focus. An AF polymorphonuclear (PMN) leucocyte count ≥ 250/mm3 -irrespective of the AF culture result- is universally accepted nowadays as the best surrogate marker for diagnosing SBP. Frequently the results of the manual or automated PMN count do not reach the hands of the responsible medical personnel in a timely manner. However, this is a crucial step in SBP man-agement. Since 2000, 26 studies (most of them published as full papers) have checked the validity of using leukocyte esterase reagent strips (LERS) in SBP diagnosis. LERS appear to have low sensitivity for SBP, some LERS types more than others. On the other hand, though, LERS have consistently given a high negative predictive value (> 95% in the majority of the studies) and this supports the use of LERS as a preliminary screening tool for SBP diagnosis. Finally, an AF-tailored dipstick has been developed. Within the proper setting, it is set to become the mainstream pro-cess for handling AF samples.

  3. Enzymatic hydrolysis of structurally diverse phthalic acid esters by porcine and bovine pancreatic cholesterol esterases.

    Science.gov (United States)

    Saito, Takao; Hong, Peng; Tanabe, Rima; Nagai, Kazuo; Kato, Katsuya

    2010-12-01

    A weak hydrolyzing activity against bis (2-ethylhexyl) phthalate (DEHP) was discovered in a commercial crude lipase (EC 3.1.1.3) preparation from porcine pancreas. DEHP was hydrolyzed to mono (2-ethylhexyl) phthalate (MEHP) not by a pancreatic lipase but by a cholesterol esterase (CEase, EC 3.1.1.13), a trace contaminant in the crude lipase preparation. Enzymatic hydrolysis of phthalic acid esters (PAEs), suspected to be endocrine-disrupting chemicals, was investigated using CEases from two species of mammals and a microorganism. Eight structurally diverse PAEs, namely diethyl phthalate (DEP), di-n-propyl phthalate (DPrP), di-n-butyl phthalate (DBP), di-n-pentyl phthalate (DPeP), di-n-hexyl phthalate (DHP), DEHP, n-butyl benzyl phthalate (BBP), and dicyclohexyl phthalate (DCHP), were hydrolyzed to their corresponding monoesters by both porcine and bovine pancreatic CEases, while a microbial CEase from Pseudomonas sp. had no hydrolyzing activity against these PAEs. The hydrolysis experiments with bovine pancreatic CEase (50 U) indicated complete hydrolysis of every PAE (5 μmole) except for BBP and DCHP within 15 min; BBP and DCHP were hydrolyzed within 30 min and 6h, respectively. The rates of PAE hydrolysis could be affected by the bulkiness of alkyl side chains in the PAEs. This study provides important evidence that mammalian pancreatic CEases, such as those from porcine and bovine sources, are potential enzymes for nonspecific degradation of structurally diverse PAEs.

  4. Glucuronoyl Esterase Screening and Characterization Assays Utilizing Commercially Available Benzyl Glucuronic Acid Ester

    Directory of Open Access Journals (Sweden)

    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.

  5. Construction of an Immobilized Thermophilic Esterase on Epoxy Support for Poly(ε-caprolactone Synthesis

    Directory of Open Access Journals (Sweden)

    Hui Ren

    2016-06-01

    Full Text Available Developing an efficient immobilized enzyme is of great significance for improving the operational stability of enzymes in poly(ε-caprolactone synthesis. In this paper, a thermophilic esterase AFEST from the archaeon Archaeoglobus fulgidus was successfully immobilized on the epoxy support Sepabeads EC-EP via covalent attachment, and the immobilized enzyme was then employed as a biocatalyst for poly(ε-caprolactone synthesis. The enzyme loading and recovered activity of immobilized enzyme was measured to be 72 mg/g and 10.4 U/mg using p-nitrophenyl caprylate as the substrate at 80 °C, respectively. Through the optimization of reaction conditions (enzyme concentration, temperature, reaction time and medium, poly(ε-caprolactone was obtained with 100% monomer conversion and low number-average molecular weight (Mn < 1300 g/mol. Further, the immobilized enzyme exhibited excellent reusability, with monomer conversion values exceeding 75% during 15 batch reactions. Finally, poly(ε-caprolactone was enzymatically synthesized with an isolated yield of 75% and Mn value of 3005 g/mol in a gram-scale reaction.

  6. Expression and characterization of a new esterase with GCSAG motif from a permafrost metagenomic library.

    Science.gov (United States)

    Petrovskaya, Lada E; Novototskaya-Vlasova, Ksenia A; Spirina, Elena V; Durdenko, Ekaterina V; Lomakina, Galina Yu; Zavialova, Maria G; Nikolaev, Evgeny N; Rivkina, Elizaveta M

    2016-05-01

    As a result of construction and screening of a metagenomic library prepared from a permafrost-derived microcosm, we have isolated a novel gene coding for a putative lipolytic enzyme that belongs to the hormone-sensitive lipase family. It encodes a polypeptide of 343 amino acid residues whose amino acid sequence displays maximum likelihood with uncharacterized proteins from Sphingomonas species. A putative catalytic serine residue of PMGL2 resides in a new variant of a recently discovered GTSAG sequence in which a Thr residue is replaced by a Cys residue (GCSAG). The recombinant PMGL2 was produced in Escherichia coli cells and purified by Ni-affinity chromatography. The resulting protein preferably utilizes short-chain p-nitrophenyl esters (C4 and C8) and therefore is an esterase. It possesses maximum activity at 45°C in slightly alkaline conditions and has limited thermostability at higher temperatures. Activity of PMGL2 is stimulated in the presence of 0.25-1.5 M NaCl indicating the good salt tolerance of the new enzyme. Mass spectrometric analysis demonstrated that N-terminal methionine in PMGL2 is processed and cysteine residues do not form a disulfide bond. The results of the study demonstrate the significance of the permafrost environment as a unique genetic reservoir and its potential for metagenomic exploration.

  7. Esterase catalysis of substrate vapour: enzyme activity occurs at very low hydration.

    Science.gov (United States)

    Lind, Penelope A; Daniel, Roy M; Monk, Colin; Dunn, Rachel V

    2004-10-01

    It has been generally accepted that enzyme activity requires a minimal hydration of about 0.2 g H2O g(-1) protein. This fits well with evidence that hydration above this level is associated with the onset of intramolecular motions. The influence of enzyme hydration on the hydrolysis of substrate by Candida rugosa Lipase B and pig liver esterase was investigated. Each enzyme was studied as a powder at various hydration levels, using vapour phase ethyl butyrate as substrate. This procedure allows the separation of those effects that are due to hydration from those arising from diffusional constraints. We found hydrolytic activity in both enzymes at all hydration levels above zero (between 0.054-0.47 and 0.029-0.60 g H2O g(-1) protein, respectively) that were investigated. The lowest hydration level investigated, hydrolysis of ethyl butyrate requires water as a second substrate, the absence of activity at zero hydration does not rule out the possibility of enzyme activity in the absence of water. These results suggest that the properties conferred on proteins by water, at least above 10% surface coverage (in this case corresponding to a hydration level of 0.03 g H2O g(-1) protein), are not a requirement for enzyme catalysis.

  8. 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.)

  9. Acquired C1 esterase inhibitor deficiency in lymphomas: prevalence, symptoms, and response to treatment.

    Science.gov (United States)

    Bekos, Christine; Perkmann, Thomas; Krauth, Maria; Raderer, Markus; Lechner, Klaus; Jaeger, Ulrich

    2016-09-01

    We retrospectively studied the prevalence of C1 esterase inhibitor (C1 INH) deficiency in 131 patients with various lymphomas. We determined C1 INH activity, C1 INH antigen, and C4 concentration at diagnosis and after chemotherapy. In follicular lymphoma (FL), diffuse large B-cell lymphoma (DLBCL) and chronic lymphocytic leukemia (CLL) consecutive patients were studied. In these entities, the prevalence of C1 INH deficiency was 10.2% in DLBCL, 4.1% in CLL, and 0% in FL and Hodgkin lymphoma. In indolent lymphomas, we identified only single cases of C1 INH deficiency, predominantly in splenic marginal zone lymphomas (SMZL) (four cases). Only three patients were symptomatic while the majority (11 cases) was asymptomatic. In DLBCL patients who were successfully treated with chemotherapy, complete normalization of C1 INH activity and C4 was observed. In contrast, C1 INH deficiency remained in SMZL patients after splenectomy. We conclude that C1 INH deficiency in lymphomas is frequently asymptomatic and responsive to immunochemotherapy.

  10. 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.

  11. 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.

  12. Heterologous production of a feruloyl esterase from Pleurotus sapidus synthesizing feruloyl-saccharide esters.

    Science.gov (United States)

    Kelle, Sebastian; Nieter, Annabel; Krings, Ulrich; Zelena, Katerina; Linke, Diana; Berger, Ralf G

    2016-11-01

    The feruloyl esterase (FAE) gene EST1 from the basidiomycete Pleurotus sapidus was heterologously expressed in Escherichia coli and Pichia pastoris. Catalytically active recombinant Est1 was secreted using P. pastoris as a host. For expression in P. pastoris, the expression vector pPIC9K was applied. The EST1 gene was cloned with an N-terminal α-mating factor pre-pro sequence and expressed under the control of a methanol inducible alcohol oxidase 1 promotor. Est1 was purified to homogeneity using ion exchange and hydrophobic interaction chromatography. The recombinant Est1 showed optima at pH 5.0 and 50 °C, and released ferulic acid from saccharide esters and from the natural substrate destarched wheat bran. Substrate specificity profile and descriptor-based analysis demonstrated unique properties, showing that Est1 did not fit into the current FAE classification model. Transferuloylation synthesis of feruloyl-saccharide esters was proven for mono- and disaccharides.

  13. Immobilization and Biochemical Properties of the Enantioselective Recombinant NStcI Esterase of Aspergillus nidulans

    Directory of Open Access Journals (Sweden)

    Carolina Peña-Montes

    2013-01-01

    Full Text Available The recombinant NStcI A. nidulans esterase was adsorbed on Accurel MP1000, where protein yield and immobilization efficiency were 42.48% and 81.94%, respectively. Storage stability test at 4°C and RT showed 100% of residual activity after 40 days at both temperatures. The biocatalyst retains more than 70% of its initial activity after 3 cycles of repeated use. Biochemical properties of this new biocatalyst were obtained. Maximum activity was achieved at pH 11 and 30°C, while the best stability was observed with the pH between 9 and 11 at 40°C. NStcI thermostability was increased after immobilization, as it retained 47.5% of its initial activity after 1 h at 60°C, while the free enzyme under the same conditions displayed no activity. NStcI preserved 70% of its initial activity in 100% hexane after 72 h. Enzymatic kinetic resolution of (R,S-1-phenylethanol was chosen as model reaction, using vinyl acetate as acyl donor. After optimization of reaction parameters, the highest possible conversion (42% was reached at 37°C, aw of 0.07, and 120 h of bioconversion in hexane with an enantiomeric excess of 71.7%. NStcI has selectivity for (R-enantiomer. The obtained E value (31.3 is in the range considered useful to resolve enantiomeric mixtures.

  14. A Lactobacillus plantarum esterase active on a broad range of phenolic esters.

    Science.gov (United States)

    Esteban-Torres, María; Landete, José María; Reverón, Inés; Santamaría, Laura; de las Rivas, Blanca; Muñoz, Rosario

    2015-05-01

    Lactobacillus plantarum is the lactic acid bacterial species most frequently found in the fermentation of food products of plant origin on which phenolic compounds are abundant. L. plantarum strains showed great flexibility in their ability to adapt to different environments and growth substrates. Of 28 L. plantarum strains analyzed, only cultures from 7 strains were able to hydrolyze hydroxycinnamic esters, such as methyl ferulate or methyl caffeate. As revealed by PCR, only these seven strains possessed the est_1092 gene. When the est_1092 gene was introduced into L. plantarum WCFS1 or L. lactis MG1363, their cultures acquired the ability to degrade hydroxycinnamic esters. These results support the suggestion that Est_1092 is the enzyme responsible for the degradation of hydroxycinnamic esters on the L. plantarum strains analyzed. The Est_1092 protein was recombinantly produced and biochemically characterized. Surprisingly, Est_1092 was able to hydrolyze not only hydroxycinnamic esters, since all the phenolic esters assayed were hydrolyzed. Quantitative PCR experiments revealed that the expression of est_1092 was induced in the presence of methyl ferulate, an hydroxycinnamic ester, but was inhibited on methyl gallate, an hydroxybenzoic ester. As Est_1092 is an enzyme active on a broad range of phenolic esters, simultaneously possessing feruloyl esterase and tannase activities, its presence on some L. plantarum strains provides them with additional advantages to survive and grow on plant environments.

  15. CHI3L1 nuclear localization in monocyte derived dendritic cells.

    Science.gov (United States)

    Di Rosa, Michelino; Tibullo, Daniele; Saccone, Salvatore; Distefano, Gisella; Basile, Maria Sofia; Di Raimondo, Francesco; Malaguarnera, Lucia

    2016-02-01

    Chitinase-3-like-1 protein (CHI3L1) is a glycosyl hydrolase (GH) highly expressed in a variety of inflammatory diseases at infectious and non-infectious etiology. CHI3L1 is produced by a wide variety of cells including monocyte-derived macrophages cell lines such as polarized M1 and M2 type macrophages, osteoclasts and Kupffer cells. In this study we have examined the expression of CHI3L1 during the differentiation and maturation of dendritic cells. Magnetically-isolated peripheral blood monocytes were differentiated toward immature DCs (iDC) and mature DCs (mDCs) through a combination of factors and cytokines. Our result showed, for the first time, that CHI3L1 is expressed during the process of differentiation and maturation of dendritic cells in time dependent manner. Furthermore, the CHI3L1 is evenly distributed in cytoplasm and in the nucleus of both the iDCs and mDCs. These results suggest that CHI3L1 may play crucial role in the DCs immunoresponse.

  16. Characterization of an RNA aptamer against HPV-16 L1 virus-like particles.

    Science.gov (United States)

    Leija-Montoya, Ana Gabriela; Benítez-Hess, María Luisa; Toscano-Garibay, Julia Dolores; Alvarez-Salas, Luis Marat

    2014-10-01

    The human papillomavirus (HPV) capsid is mainly composed of the L1 protein that can self-assemble into virus-like particles (VLPs) that are structurally and immunologically similar to the infectious virions. We report here the characterization of RNA aptamers that recognize baculovirus-produced HPV-16 L1 VLPs. Interaction and slot-blot binding assays showed that all isolated aptamers efficiently bound HPV-16 VLPs, although the Sc5-c3 aptamer showed the highest specificity and affinity (Kd=0.05 pM). Sc5-c3 secondary structure consisted of a hairpin with a symmetric bubble and an unstructured 3'end. Biochemical and genetic analyses showed that the Sc5-c3 main loop is directly involved on VLPs binding. In particular, binding specificity appeared mediated by five non-consecutive nucleotide positions. Experiments using bacterial-produced HPV-16 L1 resulted in low Sc5-c3 binding, suggesting that recognition of HPV-16 L1 VLPs relies on quaternary structure features not present in bacteria-produced L1 protein. Sc5-c3 produced specific and stable binding to HPV-16 L1 VLPs even in biofluid protein mixes and thus it may provide a potential diagnostic tool for active HPV infection.

  17. Interaction of monosulfonate tetraphenyl porphyrin (H 2TPPS 1) with plant-esterase: Determination of the binding mechanism by spectroscopic methods

    Science.gov (United States)

    Yang, Limin; Huo, Danqun; Hou, Changjun; Yang, Mei; Fa, Huanbao; Luo, Xiaogang

    2011-05-01

    The interaction of monosulfonate tetraphenyl porphyrin (H 2TPPS 1) with plant-esterase was investigated using fluorescence and UV-vis absorption spectroscopy. Fluorescence quenching, from which the binding parameters were evaluated, revealed that the quenching of the esterase by H 2TPPS 1 resulted from the formation of a dye-esterase complex. According to the modified Stern-Volmer equation, the effective quenching constants ( Ka) between H 2TPPS 1 and plant-esterase at four different temperatures (297 K, 300 K, 303 K, and 306 K) were obtained to be 14.132 × 10 5, 5.734 × 10 5, 2.907 × 10 5, and 2.291 × 10 5 M -1, respectively. The thermodynamic parameters, enthalpy change (Δ H) and entropy change (Δ S) for the reaction were calculated to be -181.67 kJ M -1 and -0.49 kJ M -1 K -1, indicating that van der Waals force and hydrogen bonds were the dominant intermolecular force in stabilizing the complex. Site marker competitive experiments showed that the binding of H 2TPPS 1 to plant-esterase primarily took place in the active site. The binding distance ( r) was obtained to be 5.99 nm according to Förster theory of non-radioactive energy transfer. The conformation of plant-esterase was investigated by synchronous fluorescence and UV-vis absorption spectroscopy, and the results confirmed some micro-environmental and conformational changes of plant-esterase molecules.

  18. 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.

  19. 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

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    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.

  20. Response of GSTase and Liver Esterase in Goldfish (Carrasius auratus) and Topmouth Gudegon (Pseudorasbora parva) after Sublethal Exposure to Cyhalofop-butyl and Profurite-aminium%氰氟草酯、杀虫安亚致死剂量对金鱼Carrasius auratus和麦穗鱼Pseudorasbora parva肝脏酯酶及谷胱甘肽-S-转移酶活性的影响

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    研究了10%千金乳油(有效成分:氰氟草酯)和78%杀虫安可溶性粉剂对金鱼和麦穗鱼肝脏酯酶及谷胱甘肽-S-转移酶(GST)活性的亚致死剂量效应.发现杀虫安(0.234 mg*L-1)对两种鱼的GST活性均具诱导作用,而氰氟草酯(1,2 mg*L-1)仅诱导了麦穗鱼的GST活性;当杀虫安与氰氟草酯混合处理时,对麦穗鱼GST活性的诱导效应最为明显.在一定的浓度范围内,氰氟草酯(1,2 mg*L-1)和杀虫安(0.117,0.234 mg*L-1)均可诱导金鱼肝脏酯酶活性;对麦穗鱼肝脏酯酶而言,杀虫安为诱导作用,氰氟草酯则抑制其活性.研究结果表明:两种酶的活性直接或间接地受供试药剂的影响,两种试鱼对氰氟草酯和杀虫安的生物反应存在差异.%Investigations were carried to assess the sublethal effects of a herbicide clincher (100 EC) containing cyhalofop-butyl [100 g (a.i)*L-1] as the active ingredient and the insecticide profurite-aminium [78% SP, 780 g (a.i)*kg-1] on liver esterases and hepatic glutathione-S-transferase (GST) in goldfish (Carassius auratus) and topmouth gudgeon (Pseudorasbora parva). GST was found to be induced in both goldfish and topmouth gudgeon by the profurite-aminium at concentration of 0.234 mg*L-1. Clincher both at concentration of 1 and 2 mg*L-1 caused GST induction in topmouth gudgeon. The mixture of clincher and profurite-aminium had the highest effect in topmouth gudgeon 722.3 nmol*(min-1*mg-1 protein) with a possible indication of additive toxicity. Liver esterases were induced by both clincher (1 and 2 mg*L-1)and profurite-aminium (0.117 and 0.234 mg*L-1) in goldfish. In topmouth gudgeon liver esterases were induced by profurite-aminium but inhibited by clincher. It is suggested that the two biotransformation enzymes may be directly or indirectly affected by the tested chemicals and that there are differences between them.

  1. FIP1L1-PDGFRA molecular analysis in the differential diagnosis of eosinophilia

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    Matsouka Panagiota

    2009-02-01

    Full Text Available Abstract Background Primary eosinophlia associated with the FIP1L1-PDGFRA rearrangement represents a subset of chronic eosinophilic leukaemia (CEL and affected patients are very sensitive to imatinib treatment. This study was undertaken in order to examine the prevalence and the associated clinicopathologic and genetic features of FIP1L1-PDGFRA rearrangement in a cohort of 15 adult patients presenting with profound eosinophilia (> 1.5 × 109/L. Methods Reverse transcriptase-polymerase chain reaction (RT-PCR was used for the detection of FIP1L1-PDGFRA rearrangement and the results confirmed by direct sequencing. C-KIT-D816V mutation was analysed retrospectively by PCR and restriction-fragment-length-polymorphism (PCR-RFLP, in all cases with primary eosinophilia. Results Two male patients with splenomegaly carried the FIP1L1-PDGFRA rearrangement, whilst 2 others were ultimately classified as suffering from idiopathic hypereosinophlic syndrome (HES and one from systemic mastocytosis. These patients were negative for the C-KIT-D816V mutation and received imatinib (100–400 mg daily. Patients with CEL and HES responded to imatinib and remained in complete haematological, clinical and molecular (for carriers of FIP1L1-PDGFRA rearrangement remission for a median of 28.2 months (range: 11–54, whilst the patient with systemic mastocytosis did not respond. Interestingly, in both patients with FIP1L1-PDGFRA rearrangement, the breakpoints into PDGFRA were located within exon 12 and fused with exons 8 and 8a of FIP1L1, respectively. Conclusion An early diagnosis of FIPIL1-PDGFRA-positive CEL and imatinib treatment offer to the affected patients an excellent clinical therapeutic result, avoiding undesirable morbidity. Moreover, although the molecular mechanisms underlying disease pathogenesis remain to be determined, imatinib can be effective in patients with idiopathic HES.

  2. L1-norm locally linear representation regularization multi-source adaptation learning.

    Science.gov (United States)

    Tao, Jianwen; Wen, Shiting; Hu, Wenjun

    2015-09-01

    In most supervised domain adaptation learning (DAL) tasks, one has access only to a small number of labeled examples from target domain. Therefore the success of supervised DAL in this "small sample" regime needs the effective utilization of the large amounts of unlabeled data to extract information that is useful for generalization. Toward this end, we here use the geometric intuition of manifold assumption to extend the established frameworks in existing model-based DAL methods for function learning by incorporating additional information about the target geometric structure of the marginal distribution. We would like to ensure that the solution is smooth with respect to both the ambient space and the target marginal distribution. In doing this, we propose a novel L1-norm locally linear representation regularization multi-source adaptation learning framework which exploits the geometry of the probability distribution, which has two techniques. Firstly, an L1-norm locally linear representation method is presented for robust graph construction by replacing the L2-norm reconstruction measure in LLE with L1-norm one, which is termed as L1-LLR for short. Secondly, considering the robust graph regularization, we replace traditional graph Laplacian regularization with our new L1-LLR graph Laplacian regularization and therefore construct new graph-based semi-supervised learning framework with multi-source adaptation constraint, which is coined as L1-MSAL method. Moreover, to deal with the nonlinear learning problem, we also generalize the L1-MSAL method by mapping the input data points from the input space to a high-dimensional reproducing kernel Hilbert space (RKHS) via a nonlinear mapping. Promising experimental results have been obtained on several real-world datasets such as face, visual video and object.

  3. Genetically engineered Oenococcus oeni strains to highlight the impact of estA2 and estA7 esterase genes on wine ester profile.

    Science.gov (United States)

    Darsonval, M; Alexandre, H; Grandvalet, C

    2016-12-01

    Besides deacidifying wine, Oenococcus oeni bring significant changes in the chemical composition of wine by releasing esters by the action of their own esterases. The impact of O. oeni esterases remains relatively unexplored. Four esterase genes were identified from O. oeni genome (estA2, estA7, estC, and estB). The dual objective of this study was, first to use a genetic tool enabling the expression of esterase genes in enological conditions and, second, to investigate the impact of O. oeni esterase gene expression during winemaking on wine aromatic profile. Both estA2 and estA7 genes were successfully cloned and expressed in O. oeni and recombinant strains were inoculated in Aligoté wine to initiate malolactic fermentation (MLF). Ester profile of experimental wine was established by SPME-GC-MS. EstA2 caused significant decreases in the concentrations of isoamyl acetate, ethyl hexanoate, isobutyl acetate, and hexyl acetate, by 42.7%, 23.4%, 51.5%, and 28.9%, respectively. EstA2 has preferential hydrolytic activity toward acetate esters from higher alcohols. EstA7 has synthetic activity toward hexyl acetate with a significant 22.7% increase. This study reports the first efficient expression system enabling the production of a functional protein in O. oeni in enological conditions.

  4. Wind Turbine Pitch Control and Load Mitigation Using an L1 Adaptive Approach

    Directory of Open Access Journals (Sweden)

    Danyong Li

    2014-01-01

    Full Text Available We present an application of L1 adaptive output feedback control design to wind turbine collective pitch control and load mitigation. Our main objective is the design of an L1 output feedback controller without wind speed estimation, ensuring that the generator speed tracks the reference trajectory with robustness to uncertain parameters and time-varying disturbances (mainly the uniform wind disturbance across the wind turbine rotor. The wind turbine model CART (controls advanced research turbine developed by the national renewable energy laboratory (NREL is used to validate the performance of the proposed L1 adaptive controller using the FAST (fatigue, aerodynamics, structures, and turbulence code. A comparative study is also conducted between the proposed controller and the most popular methods in practice: gain scheduling PI (GSPI controls and disturbance accommodating control (DAC methods. The results show better performance of L1 output feedback controller over the other two methods. Moreover, based on the FAST software and LQR analysis in the reference model selection of L1 adaptive controller, tradeoff can be achieved between control performance and loads mitigation.

  5. Ubiquitin C-terminal hydrolase-L1 potentiates cancer chemosensitivity by stabilizing NOXA.

    Science.gov (United States)

    Brinkmann, Kerstin; Zigrino, Paola; Witt, Axel; Schell, Michael; Ackermann, Leena; Broxtermann, Pia; Schüll, Stephan; Andree, Maria; Coutelle, Oliver; Yazdanpanah, Benjamin; Seeger, Jens Michael; Klubertz, Daniela; Drebber, Uta; Hacker, Ulrich T; Krönke, Martin; Mauch, Cornelia; Hoppe, Thorsten; Kashkar, Hamid

    2013-03-28

    The BH3-only protein NOXA represents one of the critical mediators of DNA-damage-induced cell death. In particular, its involvement in cellular responses to cancer chemotherapy is increasingly evident. Here, we identify a strategy of cancer cells to escape genotoxic chemotherapy by increasing proteasomal degradation of NOXA. We show that the deubiquitylating enzyme UCH-L1 is a key regulator of NOXA turnover, which protects NOXA from proteasomal degradation by removing Lys(48)-linked polyubiquitin chains. In the majority of tumors from patients with melanoma or colorectal cancer suffering from high rates of chemoresistance, NOXA fails to accumulate because UCH-L1 expression is epigenetically silenced. Whereas UCH-L1/NOXA-positive tumor samples exhibit increased sensitivity to genotoxic chemotherapy, downregulation of UCH-L1 or inhibition of its deubiquitylase activity resulted in reduced NOXA stability and resistance to genotoxic chemotherapy in both human and C. elegans cells. Our data identify the UCH-L1/NOXA interaction as a therapeutic target for overcoming cancer chemoresistance.

  6. L1 literacy affects L2 pronunciation intake and text vocalization

    Science.gov (United States)

    Walton, Martin

    2005-04-01

    For both deaf and hearing learners, L1 acquisition calls on auditive, gestural and visual modes in progressive processes over longer stages imposed in strictly anatomical and social order from the earliest pre-lexical phase [Jusczyk (1993), Kuhl & Meltzoff (1996)] to ultimate literacy. By contrast, L2 learning will call on accelerating procedures but with restricted input, arbitrated by L1 literacy as can be traced in the English of French-speaking learners, whether observed in spontaneous speech or in text vocalization modes. An inventory of their predictable omissions, intrusions and substitutions at suprasegmental and syllabic levels, many of which they can actually hear while unable to vocalize in real-time, suggests that a photogenic segmentation of continuous speech into alphabetical units has eclipsed the indispensable earlier phonogenic module, filtering L2 intake and output. This competing mode analysis hypothesizes a critical effect on L2 pronunciation of L1 graphemic procedures acquired usually before puberty, informing data for any Critical Period Hypothesis or amounts of L1 activation influencing L2 accent [Flege (1997, 1998)] or any psychoacoustic French deafness with regard to English stress-timing [Dupoux (1997)]. A metaphonic model [Howell & Dean (1991)] adapted for French learners may remedially distance L1 from L2 vocalization procedures.

  7. The L(1/2) regularization approach for survival analysis in the accelerated failure time model.

    Science.gov (United States)

    Chai, Hua; Liang, Yong; Liu, Xiao-Ying

    2015-09-01

    The analysis of high-dimensional and low-sample size microarray data for survival analysis of cancer patients is an important problem. It is a huge challenge to select the significantly relevant bio-marks from microarray gene expression datasets, in which the number of genes is far more than the size of samples. In this article, we develop a robust prediction approach for survival time of patient by a L(1/2) regularization estimator with the accelerated failure time (AFT) model. The L(1/2) regularization could be seen as a typical delegate of L(q)(0regularization methods and it has shown many attractive features. In order to optimize the problem of the relevant gene selection in high-dimensional biological data, we implemented the L(1/2) regularized AFT model by the coordinate descent algorithm with a renewed half thresholding operator. The results of the simulation experiment showed that we could obtain more accurate and sparse predictor for survival analysis by the L(1/2) regularized AFT model compared with other L1 type regularization methods. The proposed procedures are applied to five real DNA microarray datasets to efficiently predict the survival time of patient based on a set of clinical prognostic factors and gene signatures.

  8. Polarized targeting of L1-CAM regulates axonal and dendritic bundling in vitro.

    Science.gov (United States)

    Barry, Joshua; Gu, Yuanzheng; Gu, Chen

    2010-11-01

    Proper axonal and dendritic bundling is essential for the establishment of neuronal connections and the synchronization of synaptic inputs, respectively. Cell adhesion molecules of the L1-CAM (L1-cell adhesion molecule) family regulate axon guidance and fasciculation, neuron migration, dendrite morphology, and synaptic plasticity. It remains unclear how these molecules play so many different roles. Here we show that polarized axon-dendrite targeting of an avian L1-CAM protein, NgCAM (neuron-glia cell adhesion molecule), can regulate the switch of bundling of the two major compartments of rat hippocampal neurons. Using a new in-vitro model for studying neurite-neurite interactions, we found that expressed axonal NgCAM induced robust axonal bundling via the trans-homophilic interaction of immunoglobulin domains. Interestingly, dendritic bundling was induced by the dendritic targeting of NgCAM, caused by either deleting its fibronectin repeats or blocking activities of protein kinases. Consistent with the NgCAM results, expression of mouse L1-CAM also induced axonal bundling and blocking kinase activities disrupted its axonal targeting. Furthermore, the trans-homophilic interaction stabilized the bundle formation, probably through recruiting NgCAM proteins to contact sites and promoting guided axon outgrowth. Taken together, our results suggest that precise localization of L1-CAM is important for establishing proper cell-cell contacts in neural circuits.

  9. Amperometric determination of serum total cholesterol with nanoparticles of cholesterol esterase and cholesterol oxidase.

    Science.gov (United States)

    Aggarwal, V; Malik, J; Prashant, A; Jaiwal, P K; Pundir, C S

    2016-05-01

    We describe the preparation of glutaraldehyde cross-linked and functionalized cholesterol esterase nanoparticles (ChENPs) and cholesterol oxidase nanoparticles (ChOxNPs) aggregates and their co-immobilization onto Au electrode for improved amperometric determination of serum total cholesterol. Transmission electron microscope (TEM) images of ChENPs and ChOxNPs showed their spherical shape and average size of 35.40 and 56.97 nm, respectively. Scanning electron microscope (SEM) studies of Au electrode confirmed the co-immobilization of enzyme nanoparticles (ENPs). The biosensor exhibited optimal response at pH 5.5 and 40°C within 5 s when polarized at +0.25 V versus Ag/AgCl. The working/linear range of the biosensor was 10-700 mg/dl for cholesterol. The sensor showed high sensitivity and measured total cholesterol as low as 0.1 mg/dl. The biosensor was evaluated and employed for total cholesterol determination in sera of apparently healthy and diseased persons. The analytical recovery of added cholesterol was 90%, whereas the within-batch and between-batch coefficients of variation (CVs) were less than 2% and less than 3%. There was a good correlation (r = 0.99) between serum cholesterol values as measured by the standard enzymic colorimetric method and the current method. The initial activity of ENPs/working electrode was reduced by 50% during its regular use (200 times) over a period of 60 days when stored dry at 4°C.

  10. 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.

  11. Common and distant structural characteristics of feruloyl esterase families from Aspergillus oryzae.

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    D B R K Gupta Udatha

    Full Text Available BACKGROUND: Feruloyl esterases (FAEs are important biomass degrading accessory enzymes due to their capability of cleaving the ester links between hemicellulose and pectin to aromatic compounds of lignin, thus enhancing the accessibility of plant tissues to cellulolytic and hemicellulolytic enzymes. FAEs have gained increased attention in the area of biocatalytic transformations for the synthesis of value added compounds with medicinal and nutritional applications. Following the increasing attention on these enzymes, a novel descriptor based classification system has been proposed for FAEs resulting into 12 distinct families and pharmacophore models for three FAE sub-families have been developed. METHODOLOGY/PRINCIPAL FINDINGS: The feruloylome of Aspergillus oryzae contains 13 predicted FAEs belonging to six sub-families based on our recently developed descriptor-based classification system. The three-dimensional structures of the 13 FAEs were modeled for structural analysis of the feruloylome. The three genes coding for three enzymes, viz., A.O.2, A.O.8 and A.O.10 from the feruloylome of A. oryzae, representing sub-families with unknown functional features, were heterologously expressed in Pichia pastoris, characterized for substrate specificity and structural characterization through CD spectroscopy. Common feature-based pharamacophore models were developed according to substrate specificity characteristics of the three enzymes. The active site residues were identified for the three expressed FAEs by determining the titration curves of amino acid residues as a function of the pH by applying molecular simulations. CONCLUSIONS/SIGNIFICANCE: Our findings on the structure-function relationships and substrate specificity of the FAEs of A. oryzae will be instrumental for further understanding of the FAE families in the novel classification system. The developed pharmacophore models could be applied for virtual screening of compound databases for

  12. SulE, a sulfonylurea herbicide de-esterification esterase from Hansschlegelia zhihuaiae S113.

    Science.gov (United States)

    Hang, Bao-Jian; Hong, Qing; Xie, Xiang-Ting; Huang, Xing; Wang, Cheng-Hong; He, Jian; Li, Shun-Peng

    2012-03-01

    De-esterification is an important degradation or detoxification mechanism of sulfonylurea herbicide in microbes and plants. However, the biochemical and molecular mechanisms of sulfonylurea herbicide de-esterification are still unknown. In this study, a novel esterase gene, sulE, responsible for sulfonylurea herbicide de-esterification, was cloned from Hansschlegelia zhihuaiae S113. The gene contained an open reading frame of 1,194 bp, and a putative signal peptide at the N terminal was identified with a predicted cleavage site between Ala37 and Glu38, resulting in a 361-residue mature protein. SulE minus the signal peptide was synthesized in Escherichia coli BL21 and purified to homogeneity. SulE catalyzed the de-esterification of a variety of sulfonylurea herbicides that gave rise to the corresponding herbicidally inactive parent acid and exhibited the highest catalytic efficiency toward thifensulfuron-methyl. SulE was a dimer without the requirement of a cofactor. The activity of the enzyme was completely inhibited by Ag(+), Cd(2+), Zn(2+), methamidophos, and sodium dodecyl sulfate. A sulE-disrupted mutant strain, ΔsulE, was constructed by insertion mutation. ΔsulE lost the de-esterification ability and was more sensitive to the herbicides than the wild type of strain S113, suggesting that sulE played a vital role in the sulfonylurea herbicide resistance of the strain. The transfer of sulE into Saccharomyces cerevisiae BY4741 conferred on it the ability to de-esterify sulfonylurea herbicides and increased its resistance to the herbicides. This study has provided an excellent candidate for the mechanistic study of sulfonylurea herbicide metabolism and detoxification through de-esterification, construction of sulfonylurea herbicide-resistant transgenic crops, and bioremediation of sulfonylurea herbicide-contaminated environments.

  13. Carboxylic Esterase and Its Associations With Long-term Effects of Organophosphorus Pesticides

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective To examine a) the effect of organophosphorus pesticide exposure on activity of carboxylic esterases, namely butyrylcholinesterase (BChE), carboxylesterase (CarbE) and paraoxonase (PonE); and b) the association of polymorphisms of BChE and PonE with individual genetic susceptibility to organophosphorus pesticide exposure. Methods A cross-sectional study was conducted in 75 workers exposed to organophosphorus pesticides and 100 non-exposed controls. The serum activity of these enzymes was measured. Variant forms of BCHE-K, PON-192, and PON-55 were detected. A symptom score was developed as a proxy measure of clinical outcomes. Results Activities of both BChE and CarbE were lower in exposed exposed workers with BCHE-K genotype UU (61 cases), genotype UK (12 cases) and genotype KK (2 cases) was 105.05, 84.42 activity in the exposed workers with PON-192 genotype BB (37), genotype AB (27) and genotype AA (11) was 116.8, 91.2, and 9.20. The symptom score was the highest in individuals with abnormal homozygote for each of the three gene loci. Conclusions Long-term exposure to organophosphorus pesticides can inhibit BChE and CarbE activity, but exerts no inhibitory effect on PonE activity. Different genotypes of BCHE-K, PON-192, and PON-55 may be related to the severity of adverse health effects of organophosphorus pesticide exposure. Implications of potentially higher susceptibility of workers with mutant homozygotes should be evaluated to reduce health risks.

  14. 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.

  15. Kinetic imaging of NPC1L1 and sterol trafficking between plasma membrane and recycling endosomes in hepatoma cells

    DEFF Research Database (Denmark)

    Hartwig Petersen, Nicole; Færgeman, Nils J; Yu, Liqing;

    2008-01-01

    Niemann-Pick C1-like 1 (NPC1L1) is a recently identified protein that mediates intestinal cholesterol absorption and regulates biliary cholesterol excretion. The itineraries and kinetics of NPC1L1 trafficking remain uncertain. In this study, we have visualized movement of NPC1L1-enhanced green...... fluorescent protein (NPC1L1-EGFP) and cholesterol analogues in hepatoma cells. At steady state about 42% of NPC1L1 resided in the transferrin (Tf) positive, sterol enriched endocytic recycling compartment (ERC), while time-lapse microscopy demonstrated NPC1L1 traffic between plasma membrane and ERC...... exclusively in the canalicular membrane, where the protein is highly mobile. Our study demonstrates dynamic trafficking of NPC1L1 between cell surface and intracellular compartments and suggests that this transport is involved in NPC1L1 mediated cellular sterol uptake....

  16. Experiences with Sentinel-3 Optical Sensor L1 and L2 Products

    Science.gov (United States)

    Tomažić, Igor; Bonekamp, Hans; Kwiatkowska, Ewa; Montagner, Francois; O'Carroll, Anne; Santacesaria, Vincenzo

    2015-12-01

    The European Organisation for the Exploitation of Meteorological Satellites (EUMETSAT) is contributing to the overall Sentinel-3 Mission Performance Activities in Commissioning (E1) and Routine Operations (E2) phases. During the pre-launch phase we are preparing for upcoming Cal/Val activities and extensively analyzing existing Test Data Sets (TDS) but also preparing scientifically meaningful SLSTR L1 TDS. We are giving an overview of Sea and Land Surface Temperature Radiometer (SLSTR) and Ocean and Land Colour Instrument (OLCI) L1 /L2 products based on the preparatory Cal/Val activities that could be also useful for the scientific community. This includes an in-house developed product reader, extracting data from manifest file, the overview of scientifically meaningful SLSTR L1 TDS, distinction between the instrument and image grid in SLSTR products. Furthermore, we are presenting information related to SLSTR products in commissioning phase that are relevant for the scientific community.

  17. A new approach on designing l1 optimal regulator with minimum order for SISO linear systems

    Institute of Scientific and Technical Information of China (English)

    Xiang LIU

    2006-01-01

    For a SISO linear discrete-time system with a specified input signal, a novel method to realize optimal l1 regulation control is presented. Utilizing the technique of converting a polynomial equation to its corresponding matrix equation, a linear programming problem to get an optimal l1 norm of the system output error map is developed which includes the first term and the last term of the map sequence in the objective function and the right vector of its constraint matrix equation, respectively. The adjustability for the width of the constraint matrix makes the trade-off between the order of the optimal regulator and the value of the minimum objective norm become possible, especially for achieving the optimal regulator with minimum order. By norm scaling rules for the constraint matrix equation, the optimal solution can be scaled directly or be obtained by solving a linear programming problem with l1 norm objective.

  18. The inversion of 2D NMR relaxometry data using L1 regularization

    Science.gov (United States)

    Zhou, Xiaolong; Su, Guanqun; Wang, Lijia; Nie, Shengdong; Ge, Xinmin

    2017-02-01

    NMR relaxometry has been used as a powerful tool to study molecular dynamics. Many algorithms have been developed for the inversion of 2D NMR relaxometry data. Unlike traditional algorithms implementing L2 regularization, high order Tikhonov regularization or iterative regularization, L1 penalty term is involved to constrain the sparsity of resultant spectra in this paper. Then fast iterative shrinkage-thresholding algorithm (FISTA) is proposed to solve the L1 regularization problem. The effectiveness, noise vulnerability and practical utility of the proposed algorithm are analyzed by simulations and experiments. The results demonstrate that the proposed algorithm has a more excellent capability to reveal narrow peaks than traditional inversion algorithms. The L1 regularization implemented by our algorithm can be a useful complementary to the existing algorithms.

  19. L1/2 regularization based numerical method for effective reconstruction of bioluminescence tomography

    Science.gov (United States)

    Chen, Xueli; Yang, Defu; Zhang, Qitan; Liang, Jimin

    2014-05-01

    Even though bioluminescence tomography (BLT) exhibits significant potential and wide applications in macroscopic imaging of small animals in vivo, the inverse reconstruction is still a tough problem that has plagued researchers in a related area. The ill-posedness of inverse reconstruction arises from insufficient measurements and modeling errors, so that the inverse reconstruction cannot be solved directly. In this study, an l1/2 regularization based numerical method was developed for effective reconstruction of BLT. In the method, the inverse reconstruction of BLT was constrained into an l1/2 regularization problem, and then the weighted interior-point algorithm (WIPA) was applied to solve the problem through transforming it into obtaining the solution of a series of l1 regularizers. The feasibility and effectiveness of the proposed method were demonstrated with numerical simulations on a digital mouse. Stability verification experiments further illustrated the robustness of the proposed method for different levels of Gaussian noise.

  20. Critical contrastive rhetoric: The influence of L2 letter writing instruction on L1letter writing

    Directory of Open Access Journals (Sweden)

    Mehrnoosh Fakharzadeh

    2014-08-01

    Full Text Available The present study employed critical contrastive rhetoric to investigate the L2 to L1 transfer of organizational pattern and directness level of speech acts in business complaint letters. By examining the L1 complaint letters of 30 tourism university students in two phases of study, pre and post instruction of English complaint letter, the study revealed that the rhetorical organization of Persian letters are in a state of hybridity. The post instruction comparison of letters, however, showed a tendency towards applying English conventions both in organization and directness level of complaint speech act in the L1 complaint letters. The results also revealed that after instruction the expert in the field of tourism viewed some letters as inappropriate in terms of politeness which is reflected through some lexical items.

  1. An alteration in ATG16L1 stability in Crohn disease.

    Science.gov (United States)

    Lassen, Kara G; Xavier, Ramnik J

    2014-10-01

    Individuals who harbor a common coding polymorphism (Thr300Ala) within a structurally unclassified region of ATG16L1 are at increased risk for the development of Crohn disease. Recently, we reported on the generation and characterization of knockin mice carrying the ATG16L1 T300A variant. We demonstrate that multiple cell types from T300A knock-in mice exhibit reduced selective autophagy, and we mechanistically link this phenotype with an increased susceptibility of ATG16L1 T300A to CASP3- and CASP7-mediated cleavage. These findings demonstrate how a single polymorphism can result in cell type- and pathway-specific disruptions of selective autophagy and alterations in the inflammatory milieu that can contribute to disease.

  2. MODIS L1B数据在水利中的应用%Application of MODIS L1B Data to Water Resources

    Institute of Scientific and Technical Information of China (English)

    蒯晓童; 陈德清; 陈子丹

    2006-01-01

    MODIS是EOS计划中一个重要的传感器,从水利应用的角度,介绍了MODIS L1B数据的特征和处理流程,综合说明了MODIS数据在水旱情监测中的应用前景、处理方法和流程,并在此基础上讨论了深入研究过程中的主要问题.

  3. PD-L1 Expression and Survival among Patients with Advanced Non–Small Cell Lung Cancer Treated with Chemotherapy

    Directory of Open Access Journals (Sweden)

    Steffen Filskov Sorensen

    2016-02-01

    Full Text Available BACKGROUND: Recent clinical trial results have suggested that programmed cell death ligand 1 (PD-L1 expression measured by immunohistochemistry may predict response to anti–programmed cell death 1 (PD-1 therapy. Results on the association between PD-L1 expression and survival among patients with advanced non–small cell lung cancer (NSCLC treated with chemotherapy are inconsistent. MATERIAL AND METHODS: We evaluated the relationship between PD-L1 expression and overall survival (OS among 204 patients with advanced NSCLC treated at Aarhus University Hospital, Aarhus, Denmark, from 2007 to 2012. PD-L1 expression was measured using a prototype immunohistochemistry assay with the anti–PD-L1 22C3 antibody (Merck. PD-L1 strong positivity and weak positivity were defined to be traceable to the clinical trial version of the assay. RESULTS: Twenty-five percent of patients had PD-L1 strong-positive tumors, and 50% had PD-L1 weak-positive tumors. No statistically significant association was found between PD-L1 expression and survival; adjusted hazard ratio of 1.34 (95% confidence interval, 0.88-2.03; median OS, 9.0 months for the PD-L1 strong-positive group and 1.07 (0.74-1.55; median OS, 9.8 months for the PD-L1 weak-positive group compared with the PD-L1–negative group (median OS, 7.5 months. No association was seen between PD-L1 expression and OS when PD-L1 expression levels were stratified by median or tertiles. CONCLUSIONS: In concordance with previous studies, we found PD-L1 measured by immunohistochemistry to be frequently expressed in patients with advanced NSCLC. However, PD-L1 expression is not a strong prognostic marker in patients with advanced NSCLC treated with chemotherapy.

  4. Targeting the PD-1/PD-L1 axis in non-small cell lung cancer.

    Science.gov (United States)

    Kumar, Rajiv; Collins, Dearbhaile; Dolly, Saoirse; McDonald, Fiona; O'Brien, Mary E R; Yap, Timothy A

    2016-12-23

    The last decade has witnessed rapid advances in the discovery and development of immune checkpoint inhibitors in cancer medicine, particularly drugs targeting programmed cell death 1 (PD-1) and programmed cell death ligand 1 (PD-L1) in non-small cell lung cancer (NSCLC). The proven antitumor efficacy coupled with low rates of drug-related toxicities observed, albeit idiosyncratic, with these novel immunotherapeutics have led to the registration of multiple PD-1 and PD-L1 inhibitors, such as nivolumab, pembrolizumab, and atezolizumab, in second-line advanced NSCLC, whereas durvalumab and avelumab are in late-phase clinical testing. Moreover, pembrolizumab has shown a survival advantage in the first-line setting; however, nivolumab failed to show a survival benefit possibly relating to patient selection based on PD-L1 expression. Current patient selection is based on PD-L1 expression, using the relevant companion diagnostic test, where patients with strong PD-L1 expression being more likely to respond to these novel agents. Ongoing clinical research focuses on the development of PD-1 and PD-L1 inhibitor monotherapy in neoadjuvant and adjuvant NSCLC. There is also much interest in using these drugs as a therapeutic backbone for rational combinations with other treatment modalities including cytotoxic chemotherapies in the first-line NSCLC, other immunotherapies such as cytotoxic T-lymphocyte-associated protein 4 antagonists, molecularly targeted agents including EGFR and ALK inhibitors, and radiotherapy. Concurrent treatment with radiotherapy is of particular interest owing to the potential for the abscopal effect, using radiotherapy to facilitate systemic treatment.

  5. A Crohn's disease variant in Atg16l1 enhances its degradation by caspase 3

    Science.gov (United States)

    Murthy, Aditya; Li, Yun; Peng, Ivan; Reichelt, Mike; Katakam, Anand Kumar; Noubade, Rajkumar; Roose-Girma, Merone; Devoss, Jason; Diehl, Lauri; Graham, Robert R.; van Lookeren Campagne, Menno

    2014-02-01

    Crohn's disease is a debilitating inflammatory bowel disease (IBD) that can involve the entire digestive tract. A single-nucleotide polymorphism (SNP) encoding a missense variant in the autophagy gene ATG16L1 (rs2241880, Thr300Ala) is strongly associated with the incidence of Crohn's disease. Numerous studies have demonstrated the effect of ATG16L1 deletion or deficiency; however, the molecular consequences of the Thr300Ala (T300A) variant remains unknown. Here we show that amino acids 296-299 constitute a caspase cleavage motif in ATG16L1 and that the T300A variant (T316A in mice) significantly increases ATG16L1 sensitization to caspase-3-mediated processing. We observed that death-receptor activation or starvation-induced metabolic stress in human and murine macrophages increased degradation of the T300A or T316A variants of ATG16L1, respectively, resulting in diminished autophagy. Knock-in mice harbouring the T316A variant showed defective clearance of the ileal pathogen Yersinia enterocolitica and an elevated inflammatory cytokine response. In turn, deletion of the caspase-3-encoding gene, Casp3, or elimination of the caspase cleavage site by site-directed mutagenesis rescued starvation-induced autophagy and pathogen clearance, respectively. These findings demonstrate that caspase 3 activation in the presence of a common risk allele leads to accelerated degradation of ATG16L1, placing cellular stress, apoptotic stimuli and impaired autophagy in a unified pathway that predisposes to Crohn's disease.

  6. Crystal structure and characterization of esterase Est25 mutants reveal improved enantioselectivity toward (S)-ketoprofen ethyl ester.

    Science.gov (United States)

    Kim, Jinyeong; Seok, Seung-Hyeon; Hong, Eunsoo; Yoo, Tae Hyeon; Seo, Min-Duk; Ryu, Yeonwoo

    2017-03-01

    Esterases comprise a group of enzymes that catalyze the cleavage and synthesis of ester bonds. They are important in biotechnological applications owing to their enantioselectivity, regioselectivity, broad substrate specificity, and the fact that they do not require cofactors. In a previous study, we isolated the esterase Est25 from a metagenomic library. Est25 showed catalytic activity toward the (R,S)-ketoprofen ethyl ester but had low enantioselectivity toward the (S)-ketoprofen ethyl ester. Because (S)-ketoprofen has stronger anti-inflammatory effects and fewer side effects than (R)-ketoprofen, enantioselectivity of this esterase is important. In this study, we generated Est25 mutants with improved enantioselectivity toward the (S)-ketoprofen ethyl ester; improved enantioselectivity of mutants was established by analysis of their crystal structures. The enantioselectivity of mutants was influenced by substitution of Phe72 and Leu255. Substituting these residues changed the size of the binding pocket and the entrance hole that leads to the active site. The enantioselectivity of Est25 (E = 1.1 ± 0.0) was improved in the mutants F72G (E = 1.9 ± 0.2), L255W (E = 16.1 ± 1.1), and F72G/L255W (E = 60.1 ± 0.5). Finally, characterization of Est25 mutants was performed by determining the optimum reaction conditions, thermostability, effect of additives, and substrate specificity after substituting Phe72 and Leu255.

  7. Crystal structure of hyperthermophilic esterase EstE1 and the relationship between its dimerization and thermostability properties

    Directory of Open Access Journals (Sweden)

    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.

  8. Regulation of neutral cholesterol esterase and acyl-CoA : cholesterol acyltransferase in the rat adrenal gland.

    Science.gov (United States)

    Beins, D M; Vining, R; Balasubramaniam, S

    1982-03-15

    The activities of neutral cholesterol esterase and acyl-CoA : cholesterol acyltransferase in rat adrenal gland were measured at various time intervals over 24 h. The activity of cholesterol esterase displayed diurnal rhythm, with a major peak at the onset of darkness coinciding with the peak in the diurnal rhythm of plasma corticosterone concentration. The activity of acyl-CoA : cholesterol acyltransferase also exhibited a characteristic diurnal rhythm, with the minimum activity occurring 3 h after the onset of darkness. The profile of the rhythm exhibited by the activity of the esterifying enzyme was similar to the mirror image of the pattern of diurnal rhythm in the activity of 3-hydroxy-3-methylglutaryl-CoA reductase. Microsomal non-esterified cholesterol showed a gradual decline with a significant decrease in concentration at the onset of darkness, thus suggesting that diurnal removal of cholesterol in the environment of the esterifying enzyme and hydroxymethylglutaryl-CoA reductase leads to such diurnal decrease or increase in the activities of these two enzymes. Acute administration of corticotropin led to a 3-fold increase in the activity of cholesterol esterase, a 50% decrease in the activity of acyl-CoA : cholesterol acyltransferase and a 2-fold increase in the activity of hydroxymethylglutaryl-CoA reductase. Corticotropin administration also resulted in a significant decrease in microsomal non-esterified cholesterol and increase in plasma corticosterone concentration. These observations suggest that corticotropin plays an important part in generating the diurnal rhythm in the activities of the three enzymes.

  9. Characterization of a Novel Alkaline Family VIII Esterase with S-Enantiomer Preference from a Compost Metagenomic Library.

    Science.gov (United States)

    Lee, Hyun Woo; Jung, Won Kyeong; Kim, Yong Ho; Ryu, Bum Han; Kim, T Doohun; Kim, Jungho; Kim, Hoon

    2016-02-01

    A novel esterase gene, est7K, was isolated from a compost metagenomic library. The gene encoded a protein of 411 amino acids and the molecular mass of the Est7K was estimated to be 44,969 Da with no signal peptide. Est7K showed the highest identity of 57% to EstA3, which is an esterase from a drinking water metagenome, when compared with the enzymes with reported properties. Est7K had three motifs, SMTK, YSV, and WGG, which correspond to the typical motifs of family VIII esterases, SxxK, Yxx, and WGG, respectively. Est7K did not have the GxSxG motif in most lipolytic enzymes. Three additional motifs, LxxxPGxxW, PLGMxDTxF, and GGxG, were found to be conserved in family VIII enzymes. The results of the phylogenetic analysis and the alignment study suggest that family VIII enzymes could be classified into two subfamilies, VIII.1 and VIII.2. The purified Est7K was optimally active at 40°C and pH 10.0. It was activated to exhibit a 2.1-fold higher activity by the presence of 30% methanol. It preferred short-length p-nitrophenyl esters, particularly p-nitrophenyl butyrate, and efficiently hydrolyzed glyceryl tributyrate. It did not hydrolyze β-lactamase substrates, tertiary alcohol esters, glyceryl trioleate, fish oil, and olive oil. Est7K preferred an Senantiomer, such as (S)-methyl-3-hydroxy-2-methylpropionate, as the substrate. The tolerance to methanol and the substrate specificity may provide potential advantage in the use of the enzyme in pharmaceutical and other biotechnological processes.

  10. Cyst fluid NB/70K concentration and leukocyte esterase: two new markers for differentiating pancreatic serous tumors from pseudocysts.

    Science.gov (United States)

    Yong, W H; Southern, J F; Pins, M R; Warshaw, A L; Compton, C C; Lewandrowski, K B

    1995-05-01

    Cystic lesions of the pancreas include inflammatory pseudocysts, serous cystadenomas, and mucinous tumors, some of which are malignant. Preoperative clinical and radiological parameters are unreliable and may result in incorrect diagnosis and inappropriate treatment. Cyst fluid analysis for cytology, viscosity, carcino-embryonic antigen, CA 72-4, and CA 15-3 will distinguish mucinous from nonmucinous lesions and usually help in determining malignancy. Currently, there is no reliable method to differentiate inflammatory pseudocysts from serous cystadenomas. This distinction is important because the treatment of these two lesions is different; pseudocysts are either observed or drained, whereas serous tumors are usually resected. The tumor marker NB/70K was measured in aspirated cyst fluid from 13 inflammatory pseudocysts and 11 serous cystadenomas by a commercial immunoassay. Leukocyte esterase was measured using Chemstrip SG urine test strips and amylase and lipase on a routine chemistry analyzer. The cyst fluid NB/70K concentration was significantly higher in pseudocysts (mean, 555 U/ml; range, 42-1,926 U/ml) than in serous cystadenomas (mean, 12 U/ml; range 0-130 U/ml) and this difference was significant (p < 0.0002). Leukocyte esterase was detected in 7 of 11 pseudocysts but was absent in 10 of 10 serous tumors (p = 0.002). Amylase and lipase values were generally higher in pseudocysts but these markers were unreliable due to marked outliers. Cyst fluid NB/70K and leukocyte esterase are promising markers to help differentiate pseudocysts from serous tumors on percutaneous aspirates. When combined with previously reported cyst fluid parameters (amylase, lipase, cytology, and amylase isoenzymes), these two cystic lesions can be reliably distinguished.

  11. Observation and elimination of broken symmetry in L1{sub 0} FePt nanostructures

    Energy Technology Data Exchange (ETDEWEB)

    Quarterman, P.; Wang, Hao; Qiu, Jiao-Ming; Ma, Bin; Liu, Xiaoqi; Wang, Jian-Ping, E-mail: jpwang@umn.edu [Department of Electrical and Computer Engineering and the Center for Micromagnetics and Information Technologies (MINT), University of Minnesota, Minnesota 55455 (United States); Guo, Honghua [Department of Optical Science and Engineering, Fudan University, Shanghai (China)

    2015-12-07

    An unexplained surface anisotropy effect was observed and confirmed in the magnetization reversal process of both L1{sub 0} phase FePt nanoparticles with octahedral shape and (001) textured L1{sub 0} FePt thin films with island nanostructures. We suggest that the nature of the observed surface effect is caused by broken symmetry on the FePt surface, which results in weakened exchange coupling for surface atoms. Furthermore, we propose, and experimentally demonstrate, a method to repair the broken symmetry by capping the FePt islands with a Pt layer, which could prove invaluable in understanding fundamental limitations of magnetic nanostructures.

  12. Uncoupling of 3T3-L1 gene expression from lipid accumulation during adipogenesis.

    Science.gov (United States)

    Temple, Karla A; Basko, Xheni; Allison, Margaret B; Brady, Matthew J

    2007-02-06

    Adipocyte differentiation comprises altered gene expression and increased triglyceride storage. To investigate the interdependency of these two events, 3T3-L1 cells were differentiated in the presence of glucose or pyruvate. All adipocytic proteins examined were similarly increased between the two conditions. In contrast, 3T3-L1 adipocytes differentiated with glucose exhibited significant lipid accumulation, which was largely suppressed in the presence of pyruvate. Subsequent addition of glucose to the latter cells restored lipid accumulation and acute rates of insulin-stimulated lipogenesis. These data indicate that extracellular energy is required for induction of adipocytic proteins, while only glucose sustained the parallel increase in triglyceride storage.

  13. A pruning algorithm with L1/2 regularizer for extreme learning machine

    Institute of Scientific and Technical Information of China (English)

    Ye-tian FAN; Wei WU; Wen-yu YANG; Qin-wei FAN; Jian WANG

    2014-01-01

    Compared with traditional learning methods such as the back propagation (BP) method, extreme learning machine provides much faster learning speed and needs less human intervention, and thus has been widely used. In this paper we combine the L1/2 regularization method with extreme learning machine to prune extreme learning machine. A variable learning coefficient is employed to prevent too large a learning increment. A numerical experiment demonstrates that a network pruned by L1/2 regularization has fewer hidden nodes but provides better performance than both the original network and the network pruned by L2 regularization.

  14. Spiroimidazolidinone NPC1L1 inhibitors. 1: Discovery by 3D-similarity-based virtual screening.

    Science.gov (United States)

    McMasters, Daniel R; Garcia-Calvo, Margarita; Maiorov, Vladimir; McCann, Margaret E; Meurer, Roger D; Bull, Herbert G; Lisnock, Jeanmarie; Howell, Kobporn L; Devita, Robert J

    2009-06-01

    A series of spiroimidazolidinone NPC1L1 inhibitors was discovered by virtual screening of the Merck corporate sample repository using 3D-similarity-based screening. Selection of 330 compounds for testing in an in vitro NPC1L1 binding assay yielded six hits in six distinct chemical series. Follow-up 2D similarity searching yielded several sub- to low-micromolar leads; among these was spiroimidazolidinone 10, with an IC(50) of 2.5 microM. Compound 10 provided a useful scaffold to initiate a medicinal chemistry campaign.

  15. L1{sub 0} phase formation in ternary FePdNi alloys

    Energy Technology Data Exchange (ETDEWEB)

    Montes-Arango, A.M. [Department of Mechanical and Industrial Engineering, Northeastern University, Boston, MA 02115 (United States); Bordeaux, N.C. [Department of Chemical Engineering, Northeastern University, Boston, MA 02115 (United States); Liu, J.; Barmak, K. [Department of Applied Physics and Applied Mathematics, Columbia University, New York, NY 10027 (United States); Lewis, L.H., E-mail: lhlewis@neu.edu [Department of Mechanical and Industrial Engineering, Northeastern University, Boston, MA 02115 (United States); Department of Chemical Engineering, Northeastern University, Boston, MA 02115 (United States)

    2015-11-05

    Metallurgical routes to highly metastable phases are required to access new materials with new functionalities. To this end, the stability of the tetragonal chemically ordered L1{sub 0} phase in the ternary Fe–Pd–Ni system is quantified to provide enabling information concerning synthesis of L1{sub 0}-type FeNi, a highly attractive yet highly elusive advanced permanent magnet candidate. Fe{sub 50}Pd{sub 50−x}Ni{sub x} (x = 0–7 at%) samples were arc-melted and annealed at 773 K (500 °C) for 100 h to induce formation of the chemically ordered L1{sub 0} phase. Coupled calorimetry, structural and magnetic investigations allow determination of an isothermal section of the ternary Fe–Pd–Ni phase diagram featuring a single phase L1{sub 0} region near the FePd boundary for x < 6 at%. It is demonstrated that increased Ni content in Fe{sub 50}Pd{sub 50−x}Ni{sub x} alloys systematically decreases the order-disorder transition temperature, resulting in a lower thermodynamic driving force for the ordering phase transformation. The Fe{sub 50}Pd{sub 50−x}Ni{sub x} L1{sub 0} → fcc disordering transformation is determined to occur via a two-step process, with compositionally-dependent enthalpies and transition temperatures. These results highlight the need to investigate ternary alloys with higher Ni content to determine the stability range of the L1{sub 0} phase near the FeNi boundary, thereby facilitating kinetic access to the important L1{sub 0} FeNi ferromagnetic phase. - Highlights: • Chemical ordering in FePdNi enhances intrinsic and extrinsic magnetic properties. • 773 K annealed FePdNi alloys studied show a stable L1{sub 0} phase for Ni ≤ 5.2 at%. • Chemical disordering in FePdNi occurs by a previously unreported two-step process. • Ni additions to FePd dramatically decrease the chemical order-disorder temperature. • The chemical-ordering transformation kinetics are greatly affected by Ni content.

  16. The clinical spectrum of mutations in L1, a neuronal cell adhesion molecule

    Energy Technology Data Exchange (ETDEWEB)

    Fransen, E.; Vits, L.; Van Camp, G.; Willems, P.J. [Univ. of Antwerp (Belgium)

    1996-07-12

    Mutations in the gene encoding the neuronal cell adhesion molecule L1 are responsible for several syndromes with clinical overlap, including X-linked hydrocephalus (XLH, HSAS), MASA (mental retardation, aphasia, shuffling gait, adducted thumbs) syndrome, complicated X-linked spastic paraplegia (SP 1), X-linked mental retardation-clasped thumb (MR-CT) syndrome, and some forms of X-linked agenesis of the corpus callosum (ACC). We review 34 L1 mutations in patients with these phenotypes. 22 refs., 3 figs., 4 tabs.

  17. Association between PD-1/PD-L1 and T regulate cells in early recurrent miscarriage.

    Science.gov (United States)

    Li, Guiyu; Lu, Caixia; Gao, Jing; Wang, Xietong; Wu, Huanling; Lee, Chao; Xing, Baoxiang; Zhang, Qi

    2015-01-01

    In this study, we try to testify the relationship between the programmed cell death receptor-1 (PD-1)/programmed cell death ligand 1 (PD-L1) passway and Treg cells in maternal-fetal immune regulation through PD-1 blockade on lymphocytes of normal early pregnancy in vitro and investigation of the PD-1 and PD-L1 changes in early recurrent miscarriage patients. CD4+ CD25+ Treg cells and PD-1 (CD279) positive cell were detected in deciduas in early recurrent miscarriage patients by flow cytometry. And the normal early pregnant women were as controls. Meanwhile the mRNA level of PD-1 and molecular expression of PD-L1 in deciduas of early recurrent miscarriage patients were detected by real time RT-PCR test and Immunohistochemical staining respectively. Also through antibody blocking assay to block PD-1 on lymphocytes of normal early pregnancy in vitro further testify the relationship between PD-1/PD-L1 and Treg cells, the results were analyzed by flow cytometry. CD4+ CD25+ Treg cells decreased both in deciduas in RM (P 0.05). PD-L1 mRNA in deciduas decreased in RM (P 0.1). After PD-1 blockade there was no change in CD4+ CD25+ Treg cells percentage, while the CD4+ T cell percentage increased (P < 0.01), as well as the level of IFN-gamma in cells supernatant (P < 0.01). PD-1 blockade has a little influence on the number of Treg cells, and may lead to impaired Treg cells function, the decrease of PD-L1 may closely relates to the occurrence of early recurrent miscarriage and implies that Treg cells may through PD-1/PD-L1 pathway play a role of immunosuppression regulation, and the impairment of Treg cells function in recurrent early abortion cases may be due to PD-L1 decrease in deciduas or trophoblast cells rather than PD-1 change.

  18. L1(-∞,∞)上的Fourier变换

    Institute of Scientific and Technical Information of China (English)

    王海华; 张学俊

    2008-01-01

    我们讨论L1(-∞,∞)上的Fourier变换,给出Fourier变换的若干重要性质,还讨论Fourier变换的部分反演性质,这些都是Fourier系数在L1(-∞,∞)中的推广,但具有更广泛的应用,本文的证明比[1]中详细,证明的方法也有所不同.

  19. File list: Oth.ALL.05.Whsc1l1.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.05.Whsc1l1.AllCell mm9 TFs and others Whsc1l1 All cell types SRX1114542,SRX...1114547 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.ALL.05.Whsc1l1.AllCell.bed ...

  20. File list: Oth.ALL.20.Whsc1l1.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.20.Whsc1l1.AllCell mm9 TFs and others Whsc1l1 All cell types SRX1114542,SRX...1114547 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.ALL.20.Whsc1l1.AllCell.bed ...

  1. HUMAN PAPILLOMAVIRUS TYPE 16 L1 PROTEIN CAN BE EXPRESSED IN LIVE ATTENUATED SHIGELLA FLEXNERI 5A STRAIN SH42

    Institute of Scientific and Technical Information of China (English)

    Qu Xinzhong; Yang Xiaofeng; Zheng Jin; Wang Kai; Si Lüsheng; Wang Yili

    2005-01-01

    Objective Attenuated strains of Shigella are attractive live vaccine candidates for eliciting mucosal immune responses which is a suitable carrier for the prophylactic human papillomaviruses (HPV) vaccine development, To examine the potential of a live Shigella based prophylactic HPV vaccine, HPV16L1should be expressed in attenuated shigella strain. Methods A Shigella large invasive plasmid (icsA/virG) based prokaryotic expression plasmid pHS3199 was constructed. HPV16L1 gene was inserted into plasmid pHS3199 to form pHS3199-HPV16 L1 construct, and pHS3199-hpv16L1 was electroporated into a live attenuated shigella strain sh42. The expression of HPV16L1 protein was demonstrated by Western blotting with monoclonal antibody to HPV16L1, The genetic stability of recombinant strain sh42-HPV16 L1 was monitored by consecutive passage culture. Invasive ability of sh42-HPV16L1 was evaluated by Hela cell infection assay. Results HPV16 L1 protein can be expressed in recombinant strain sh42-HPV16 L1, and the protein stably expressed over 140 generations. The invasive ability of sh42-HPV16L1 was diminished dramatically compared to its parent strain, but not abolished completely. Conclusion HPV16L1 protein was constitutively expressed in the attenuated strain of shigella flexneri sh42, and maintained partial invasive ability. Our strategy may represent a promising vaccine candidate against genital HPV16 infection.

  2. 26 CFR 36.3121(l)(1)-1 - Agreements entered into by domestic corporations with respect to foreign subsidiaries.

    Science.gov (United States)

    2010-04-01

    ... corporations with respect to foreign subsidiaries. 36.3121(l)(1)-1 Section 36.3121(l)(1)-1 Internal Revenue... TAX AT SOURCE CONTRACT COVERAGE OF EMPLOYEES OF FOREIGN SUBSIDIARIES § 36.3121(l)(1)-1 Agreements entered into by domestic corporations with respect to foreign subsidiaries. (a) In general. (1)...

  3. L2 Acquisition of Prosodic Properties of Speech Rhythm: Evidence from L1 Mandarin and German Learners of English

    Science.gov (United States)

    Li, Aike; Post, Brechtje

    2014-01-01

    This study examines the development of speech rhythm in second language (L2) learners of typologically different first languages (L1s) at different levels of proficiency. An empirical investigation of durational variation in L2 English productions by L1 Mandarin learners and L1 German learners compared to native control values in English and the…

  4. Predicting pauses in L1 and L2 speech: the effects of utterance boundaries and word frequency

    NARCIS (Netherlands)

    de Jong, N.H.

    2016-01-01

    This paper compares the distribution of silent and filled pauses in first (L1) and second language (L2) speech. The occurrence of pauses of 52 L2 and 18 L1 Dutch speakers was evaluated with respect to utterance boundaries and word frequency. We found that L2 speakers paused more often than L1 speake

  5. 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

  6. Estrogenic and esterase-inhibiting potency in rainwater in relation to pesticide concentrations, sampling season and location

    Energy Technology Data Exchange (ETDEWEB)

    Hamers, T.; Brink, P.J. van den; Mos, L.; Linden, S.C. van der; Legler, J.; Koeman, J.H.; Murk, A.J

    2003-05-01

    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

  7. 发酵性丝孢酵母酯酶的稳定性研究%Stabilities of Esterase from Fermentable Filamentous Spore Yeast(Trichosporon fermentans)

    Institute of Scientific and Technical Information of China (English)

    陈洋洋; 张晓锋; 侯英敏; 孙玉梅

    2015-01-01

    Thestabilitiesofesterasefromcellwalloffilamentoussporeyeast(FSY)(Trichosporonfermentans)were studied in different physical and chemical conditions. The results showed that the esterase remained 81. 5% of its ini-tial activity after being stored at 0 ℃ for 24 h and became totally inactive at 40 ℃. K+,Mn2+ and Fe2+ at 5 mmol/L increased esterase activity by fewer than 6. 9%. While Ca2+ and Cu2+ at 5 mmol/L reduced esterase activities by respectively 90. 7% and 73. 3%. The residual activity of esterase was kept at more than 77% of original activity in 1%(w/w)Tween-40 and TritonX-100,and increased by 23. 3% in 1%(w/w)Tween-80. While in 1%(w/w) SDS( sodium dodecyl sulfate)and CTAB( cetyl trimethyl ammonium bromide)totally inhibited the esterase activity. 5%( w/w)glycerol,Arabic gum,and dextrin could enhance esterase activity,and enhanced the esterase activity by respectively 57%,29. 6%,and 2%. 5% soluble starch and BSA(albumin from bovine serum)decreased esterase activity by respectively 20. 43% and 71. 5%. The FSY esterase should be stored below 10 ℃ due to its poor thermal stability. Glycerol had good protection effect on the esterase,while BSA had strong inhibition against the esterase. The esterase had potential application values to treat oleo-waste-water and to enhance washing effects of detergent.%以发酵性丝孢酵母胞壁酯酶为研究对象,研究了其在不同理化条件下的稳定性。结果表明:0℃保存24 h,酯酶仍保持81.5%原酶活;在40℃完全失活;5 mmol/L K+、Mn2+和Fe2+提高酶活力6.9%以下,5 mmol/L Ca2+、Cu2+分别降低酶活力90.7%和73.3%;1%(质量分数,下同)Tween-40和TritonX-100保持77%以上原酶活,1%Tween-80提高酶活力23.3%,1%十二烷基磺酸钠( Sodium dodecyl sulfate,SDS)和十六烷基三甲基溴化铵( Cetyl trimethyl ammonium bromide,CTAB)完全抑制酶活;5%甘油、阿拉伯胶和糊精分别提高酶活力57%、29.6%和2%,5%可溶

  8. Interaction of Plant Epicuticular Waxes and Extracellular Esterases of Curvularia eragrostidis during Infection of Digitaria sanguinalis and Festuca arundinacea by the Fungus

    Directory of Open Access Journals (Sweden)

    Lang-Lai Xu

    2006-09-01

    Full Text Available Curvularia eragrostidis, a causal agent of head blight on the weed (Digitariasanguinalis, did not cause disease on the turfgrass Festuca arundinacea. Differentextracellular esterase isoenzymes were detected in saprophytic and parasitic phases duringthe fungal germination. The epicuticular waxes of D. sanguinalis were more efficient toinduce the secretion of esterases from the fungus than that of F. arundinacea, but were morerapidly degraded by the fungal enzymes. Component analysis indicated that the epicuticularwaxes from D. sanguinalis were mostly composed of alcohols, with 54.3% being 9,12-Octadecadien-1-ol. The main component of F arundinacea waxes was alkyl compounds,with 49.8% being olefin, 9-Tricosence. More long-chained esters were found in D.sanguinalis waxes, which were easier to be digested than those in F. arundinacea waxes byextreacellular esterases of the fungus. Epicuticular waxes play a role in varyingpathogenicity of C. eragrostidis on D. sanguinalis and F arundinacea.

  9. The effect of C1-esterase inhibitor on systemic inflammation in trauma patients with a femur fracture - The CAESAR study: study protocol for a randomized controlled trial

    Directory of Open Access Journals (Sweden)

    Strengers Paul FW

    2011-10-01

    Full Text Available Abstract Background Systemic inflammation in response to a femur fracture and the additional fixation is associated with inflammatory complications, such as acute respiratory distress syndrome and multiple organ dysfunction syndrome. The injury itself, but also the additional procedure of femoral fixation induces a release of pro-inflammatory cytokines such as interleukin-6. This results in an aggravation of the initial systemic inflammatory response, and can cause an increased risk for the development of inflammatory complications. Recent studies have shown that administration of the serum protein C1-esterase inhibitor can significantly reduce the release of circulating pro-inflammatory cytokines in response to acute systemic inflammation. Objective Attenuation of the surgery-induced additional systemic inflammatory response by perioperative treatment with C1-esterase inhibitor of trauma patients with a femur fracture. Methods The study is designed as a double-blind randomized placebo-controlled trial. Trauma patients with a femur fracture, Injury Severity Score ≥ 18 and age 18-80 years are included after obtaining informed consent. They are randomized for administration of 200 U/kg C1-esterase inhibitor intravenously or placebo (saline 0.9% just before the start of the procedure of femoral fixation. The primary endpoint of the study is Δ interleukin-6, measured at t = 0, just before start of the femur fixation surgery and administration of C1-esterase inhibitor, and t = 6, 6 hours after administration of C1-esterase inhibitor and the femur fixation. Conclusion This study intents to identify C1-esterase inhibitor as a safe and potent anti-inflammatory agent, that is capable of suppressing systemic inflammation in trauma patients. This might facilitate early total care procedures by lowering the risk of inflammation in response to the surgical intervention. This could result in increased functional outcomes and reduced health care related

  10. KSHV LANA and EBV LMP1 induce the expression of UCH-L1 following viral transformation

    Energy Technology Data Exchange (ETDEWEB)

    Bentz, Gretchen L.; Bheda-Malge, Anjali; Wang, Ling [Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill (United States); Shackelford, Julia [Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill (United States); Department of Cell and Developmental Biology, University of North Carolina, Chapel Hill (United States); Damania, Blossom [Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill (United States); Departments of Medicine and of Microbiology and Immunology, University of North Carolina, Chapel Hill, NC (United States); Pagano, Joseph S., E-mail: joseph_pagano@med.unc.edu [Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill (United States); Departments of Medicine and of Microbiology and Immunology, University of North Carolina, Chapel Hill, NC (United States)

    2014-01-05

    Ubiquitin C-terminal Hydrolase L1 (UCH-L1) has oncogenic properties and is highly expressed during malignancies. We recently documented that Epstein-Barr virus (EBV) infection induces uch-l1 expression. Here we show that Kaposi's Sarcoma-associated herpesvirus (KSHV) infection induced UCH-L1 expression, via cooperation of KSHV Latency-Associated Nuclear Antigen (LANA) and RBP-Jκ and activation of the uch-l1 promoter. UCH-L1 expression was also increased in Primary Effusion Lymphoma (PEL) cells co-infected with KSHV and EBV compared with PEL cells infected only with KSHV, suggesting EBV augments the effect of LANA on uch-l1. EBV latent membrane protein 1 (LMP1) is one of the few EBV products expressed in PEL cells. Results showed that LMP1 was sufficient to induce uch-l1 expression, and co-expression of LMP1 and LANA had an additive effect on uch-l1 expression. These results indicate that viral latency products of both human γ-herpesviruses contribute to uch-l1 expression, which may contribute to the progression of lymphoid malignancies. - Highlights: • Infection of endothelial cells with KSHV induced UCH-L1 expression. • KSHV LANA is sufficient for the induction of uch-l1. • Co-infection with KSHV and EBV (observed in some PELs) results in the additive induction of uch-l1. • EBV LMP1 also induced UCH-L1 expression. • LANA- and LMP1-mediated activation of the uch-l1 promoter is in part through RBP-Jκ.

  11. 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

  12. Increase of cells expressing PD-L1 in bovine leukemia virus infection and enhancement of anti-viral immune responses in vitro via PD-L1 blockade

    Directory of Open Access Journals (Sweden)

    Ikebuchi Ryoyo

    2011-09-01

    Full Text Available Abstract The inhibitory receptor programmed death-1 (PD-1 and its ligand, programmed death-ligand 1 (PD-L1 are involved in immune evasion mechanisms for several pathogens causing chronic infections. Blockade of the PD-1/PD-L1 pathway restores anti-virus immune responses, with concomitant reduction in viral load. In a previous report, we showed that, in bovine leukemia virus (BLV infection, the expression of bovine PD-1 is closely associated with disease progression. However, the functions of bovine PD-L1 are still unknown. To investigate the role of PD-L1 in BLV infection, we identified the bovine PD-L1 gene, and examined PD-L1 expression in BLV-infected cattle in comparison with uninfected cattle. The deduced amino acid sequence of bovine PD-L1 shows high homology to the human and mouse PD-L1. The proportion of PD-L1 positive cells, especially among B cells, was upregulated in cattle with the late stage of the disease compared to cattle at the aleukemic infection stage or uninfected cattle. The proportion of PD-L1 positive cells correlated positively with prediction markers for the progression of the disease such as leukocyte number, virus load and virus titer whilst on the contrary, it inversely correlated with the degree of interferon-gamma expression. Blockade of the PD-1/PD-L1 pathway in vitro by PD-L1-specific antibody upregulated the production of interleukin-2 and interferon-gamma, and correspondingly, downregulated the BLV provirus load and the proportion of BLV-gp51 expressing cells. These data suggest that PD-L1 induces immunoinhibition in disease progressed cattle during chronic BLV infection. Therefore, PD-L1 would be a potential target for developing immunotherapies against BLV infection.

  13. 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

    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......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...

  14. Translation assistance by translation of L1 fragments in an L2 context

    NARCIS (Netherlands)

    Gompel, M. van; Bosch, A.P.J. van den

    2014-01-01

    In this paper we present new research in translation assistance. We describe a system capable of translating native language (L1) fragments to foreign language (L2) fragments in an L2 context. Practical applications of this research can be framed in the context of second language learning. The type

  15. Data of evolutionary structure change: 1DS8L-1AIJM [Confc[Archive

    Lifescience Database Archive (English)

    Full Text Available 1DS8L-1AIJM 1DS8 1AIJ L M --------------ALLSFERKYRVPGGTL-------VG...A 245 SER CA 310 ALA CA 263 VAL CA 196 1AIJ... M 1AIJMLEU CA 264 ALA CA 260 LYS CA 280 1AIJ... M 1AIJM YGLSFAAPLKE

  16. Spanish as a Second Language when L1 Is Quechua: Endangered Languages and the SLA Researcher

    Science.gov (United States)

    Kalt, Susan E.

    2012-01-01

    Spanish is one of the most widely spoken languages in the world. Quechua is the largest indigenous language family to constitute the first language (L1) of second language (L2) Spanish speakers. Despite sheer number of speakers and typologically interesting contrasts, Quechua-Spanish second language acquisition is a nearly untapped research area,…

  17. PD-L1 biomarker testing for non-small cell lung cancer: truth or fiction?

    Science.gov (United States)

    Grigg, Claud; Rizvi, Naiyer A

    2016-01-01

    Research in cancer immunology is currently accelerating following a series of cancer immunotherapy breakthroughs during the last 5 years. Various monoclonal antibodies which block the interaction between checkpoint molecules PD-1 on immune cells and PD-L1 on cancer cells have been used to successfully treat non-small cell lung cancer (NSCLC), including some durable responses lasting years. Two drugs, nivolumab and pembrolizumab, are now FDA approved for use in certain patients who have failed or progressed on platinum-based or targeted therapies while agents targeting PD-L1, atezolizumab and durvalumab, are approaching the final stages of clinical testing. Despite impressive treatment outcomes in a subset of patients who receive these immune therapies, many patients with NSCLC fail to respond to anti-PD-1/PD-L1 and the identification of a biomarker to select these patients remains highly sought after. In this review, we discuss the recent clinical trial results of pembrolizumab, nivolumab, and atezolizumab for NSCLC, and the significance of companion diagnostic testing for tumor PD-L1 expression.

  18. Modified Sparseness Controlled IPNLMS Algorithm Based on l_1, l_2 and l_∞ Norms

    Directory of Open Access Journals (Sweden)

    Krishna Samalla

    2013-04-01

    Full Text Available In the context of Acoustic Echo Cancellation (AEC, sparseness level of acoustic impulse response (AIR varies greatly in mobile environments. The modified sparseness-controlled Improved PNLMS (MSC-IPNLMS algorithm proposed in this paper, exploits the sparseness measure of AIR using l1, l2 and l∞ norms. The MSC-IPNLMS is the modified version of SC-IPNLMS which uses sparseness measure based on l1 and l2 norms. Sparseness measure using l1, l2 and l∞ norms is the good representation of both sparse and dense impulse response, where as the measure which uses l1 and l2 norms is the good representation of sparse impulse response only. The MSC-IPNLMS is based on IPNLMS which allocates adaptation step size gain in proportion to the magnitude of estimated filter weights. By estimating the sparseness of the AIR, the proposed MSC-IPNLMS algorithm assigns the gains for each step size such that the proportionate term of the IPNLMS will be given higher weighting for sparse impulse responses. For a less sparse impulse response, a higher weighting will be given to the NLMS term. Simulation results, with input as white Gaussian noise (WGN, show the improved performance over the SC-IPNLMS algorithm in both sparse and dense AIR.

  19. Task-Modality and L1 Use in EFL Oral Interaction

    Science.gov (United States)

    Azkarai, Agurtzane; del Pilar García Mayo, María

    2015-01-01

    This study examines whether task-modality (speaking vs. speaking+writing) influences first language (L1) use in task-based English as a foreign language (EFL) learner-learner interaction. Research on the topic has shown that different task-modality triggers different learning opportunities with collaborative speaking tasks drawing learners'…

  20. Perceptual Processing of Mandarin Nasals by L1 and L2 Mandarin Speakers

    Science.gov (United States)

    Lai, Yi-hsiu

    2012-01-01

    Nasals are cross-linguistically susceptible to change, especially in the syllable final position. Acoustic reports on Mandarin nasal production have recently shown that the syllable-final distinction is frequently dropped. Few studies, however, have addressed the issue of perceptual processing in Mandarin nasals for L1 and L2 speakers of Mandarin…

  1. Germline Chromothripsis Driven by L1-Mediated Retrotransposition and Alu/Alu Homologous Recombination

    DEFF Research Database (Denmark)

    Nazaryan-Petersen, Lusine; Bertelsen, Birgitte; Bak, Mads;

    2016-01-01

    L1-endonuclease potential target sites in other breakpoints. In addition, we found four Alu elements flanking the 110-kb deletion and associated with an inversion. We suggest that chromatin looping mediated by homologous Alu elements may have brought distal DNA regions into close proximity...

  2. Adult Learners' Perceptions of the Incorporation of Their L1 in Foreign Language Teaching and Learning

    Science.gov (United States)

    Brooks-Lewis, Kimberly Anne

    2009-01-01

    This article challenges the theory and practice of the exclusion of the adult learner's first language (L1) by reporting learners' overwhelmingly positive perceptions of its incorporation in foreign language teaching and learning. Classroom-based research was undertaken with university students in an English as a foreign language course which…

  3. Fault Tolerance Analysis of L1 Adaptive Control System for Unmanned Aerial Vehicles

    Science.gov (United States)

    Krishnamoorthy, Kiruthika

    Trajectory tracking is a critical element for the better functionality of autonomous vehicles. The main objective of this research study was to implement and analyze L1 adaptive control laws for autonomous flight under normal and upset flight conditions. The West Virginia University (WVU) Unmanned Aerial Vehicle flight simulation environment was used for this purpose. A comparison study between the L1 adaptive controller and a baseline conventional controller, which relies on position, proportional, and integral compensation, has been performed for a reduced size jet aircraft, the WVU YF-22. Special attention was given to the performance of the proposed control laws in the presence of abnormal conditions. The abnormal conditions considered are locked actuators (stabilator, aileron, and rudder) and excessive turbulence. Several levels of abnormal condition severity have been considered. The performance of the control laws was assessed over different-shape commanded trajectories. A set of comprehensive evaluation metrics was defined and used to analyze the performance of autonomous flight control laws in terms of control activity and trajectory tracking errors. The developed L1 adaptive control laws are supported by theoretical stability guarantees. The simulation results show that L1 adaptive output feedback controller achieves better trajectory tracking with lower level of control actuation as compared to the baseline linear controller under nominal and abnormal conditions.

  4. Structural basis for small molecule targeting of the programmed death ligand 1 (PD-L1)

    NARCIS (Netherlands)

    Zak, Krzysztof M.; Grudnik, Przemyslaw; Guzik, Katarzyna; Zieba, Bartosz J.; Musielak, Bogdan; Dömling, Alexander; Dubin, Grzegorz; Holak, Tad A.

    2016-01-01

    Targeting the PD-1/PD-L1 immunologic checkpoint with monoclonal antibodies has provided unprecedented results in cancer treatment in the recent years. Development of chemical inhibitors for this pathway lags the antibody development because of insufficient structural information. The first nonpeptid

  5. Assessment of Serum UCH-L1 and GFAP in Acute Stroke Patients

    Science.gov (United States)

    Ren, Changhong; Kobeissy, Firas; Alawieh, Ali; Li, Na; Li, Ning; Zibara, Kazem; Zoltewicz, Susie; Guingab-Cagmat, Joy; Larner, Stephen F.; Ding, Yuchuan; Hayes, Ronald L.; Ji, Xunming; Mondello, Stefania

    2016-01-01

    A rapid and reliable diagnostic test to distinguish ischemic from hemorrhagic stroke in patients presenting with stroke-like symptoms is essential to optimize management and triage for thrombolytic therapy. The present study measured serum concentrations of ubiquitin C-terminal hydrolase (UCH-L1) and glial fibrillary astrocytic protein (GFAP) in acute stroke patients and healthy controls and investigated their relation to stroke severity and patient characteristics. We also assessed the diagnostic performance of these markers for the differentiation of intracerebral hemorrhage (ICH) from ischemic stroke (IS). Both UCH-L1 and GFAP concentrations were significantly greater in ICH patients than in controls (p < 0.0001). However, exclusively GFAP differed in ICH compared with IS (p < 0.0001). GFAP yielded an AUC of 0.86 for differentiating between ICH and IS within 4.5hrs of symptom onset with a sensitivity of 61% and a specificity of 96% using a cut-off of 0.34ng/ml. Higher GFAP levels were associated with stroke severity and history of prior stroke. Our results demonstrate that blood UCH-L1 and GFAP are increased early after stroke and distinct biomarker-specific release profiles are associated with stroke characteristics and type. We also confirmed the potential of GFAP as a tool for early rule-in of ICH, while UCH-L1 was not clinically useful. PMID:27074724

  6. Isoproterenol Increases Uncoupling, Glycolysis, and Markers of Beiging in Mature 3T3-L1 Adipocytes.

    Science.gov (United States)

    Miller, Colette N; Yang, Jeong-Yeh; England, Emily; Yin, Amelia; Baile, Clifton A; Rayalam, Srujana

    2015-01-01

    Beta-adrenergic activation stimulates uncoupling protein 1 (UCP1), enhancing metabolic rate. In vitro, most work has studied brown adipocytes, however, few have investigated more established adipocyte lines such as the murine 3T3-L1 line. To assess the effect of beta-adrenergic activation, mature 3T3-L1s were treated for 6 or 48 hours with or without isoproterenol (10 and 100 μM) following standard differentiation supplemented with thyroid hormone (T3; 1 nM). The highest dose of isoproterenol increased lipid content following 48 hours of treatment. This concentration enhanced UCP1 mRNA and protein expression. The increase in UCP1 following 48 hours of isoproterenol increased oxygen consumption rate. Further, coupling efficiency of the electron transport chain was disturbed and an enhancement of glycolytic rate was measured alongside this, indicating an attempt to meet the energy demands of the cell. Lastly, markers of beige adipocytes (protein content of CD137 and gene transcript of CITED1) were also found to be upregulated at 48 hours of isoproterenol treatment. This data indicates that mature 3T3-L1 adipocytes are responsive to isoproterenol and induce UCP1 expression and activity. Further, this finding provides a model for further pharmaceutical and nutraceutical investigation of UCP1 in 3T3-L1s.

  7. Isoproterenol Increases Uncoupling, Glycolysis, and Markers of Beiging in Mature 3T3-L1 Adipocytes.

    Directory of Open Access Journals (Sweden)

    Colette N Miller

    Full Text Available Beta-adrenergic activation stimulates uncoupling protein 1 (UCP1, enhancing metabolic rate. In vitro, most work has studied brown adipocytes, however, few have investigated more established adipocyte lines such as the murine 3T3-L1 line. To assess the effect of beta-adrenergic activation, mature 3T3-L1s were treated for 6 or 48 hours with or without isoproterenol (10 and 100 μM following standard differentiation supplemented with thyroid hormone (T3; 1 nM. The highest dose of isoproterenol increased lipid content following 48 hours of treatment. This concentration enhanced UCP1 mRNA and protein expression. The increase in UCP1 following 48 hours of isoproterenol increased oxygen consumption rate. Further, coupling efficiency of the electron transport chain was disturbed and an enhancement of glycolytic rate was measured alongside this, indicating an attempt to meet the energy demands of the cell. Lastly, markers of beige adipocytes (protein content of CD137 and gene transcript of CITED1 were also found to be upregulated at 48 hours of isoproterenol treatment. This data indicates that mature 3T3-L1 adipocytes are responsive to isoproterenol and induce UCP1 expression and activity. Further, this finding provides a model for further pharmaceutical and nutraceutical investigation of UCP1 in 3T3-L1s.

  8. Octanoate and decanoate induce apoptosis in 3T3-L1 adipocytes.

    Science.gov (United States)

    Yang, Jeong-Yeh; Rayalam, Srujana; Della-Fera, Mary Anne; Ambati, Suresh; Baile, Clifton A

    2009-10-01

    The effect of octanoate and decanoate, respectively, eight- and 10-carbon medium-chain fatty acids (MCFAs), on apoptotic signaling in 3T3-L1 adipocytes was investigated. 3T3-L1 adipocytes were treated with various concentrations of octanoate or decanoate. Cell viability, apoptosis, and expression of apoptosis-related proteins were investigated. Results indicated that both octanoate and decanoate decreased viability, increased apoptosis, and increased reactive oxygen species production. Immunoblotting analysis showed an increase in the levels of cytoplasmic cytochrome c and cleaved poly(ADP-ribose) polymerase by octanoate and decanoate. Concomitantly, we observed that pro-caspase-3 was decreased, resulting in the induced accumulation of the cleaved form of caspase-3 by both octanoate and decanoate. In addition, both octanoate and decanoate increased the expression of pro-apoptotic Bax with an accompanied decrease of anti-apoptotic Bcl-2. These results show that octanoate and decanoate mediate adipocyte apoptosis via a caspase-dependent mitochondrial pathway in 3T3-L1 adipocytes. MCFAs thus decrease adipocyte number by initiating the apoptotic process in 3T3-L1 adipocytes.

  9. Selectivity in L1 Attrition: Differential Object Marking in Spanish Near-Native Speakers of English

    Science.gov (United States)

    Chamorro, Gloria; Sturt, Patrick; Sorace, Antonella

    2016-01-01

    Previous research has shown L1 attrition to be restricted to structures at the interfaces between syntax and pragmatics, but not to occur with syntactic properties that do not involve such interfaces ("Interface Hypothesis", Sorace and Filiaci in "Anaphora resolution in near-native speakers of Italian." "Second Lang…

  10. NOD2 and ATG16L1 polymorphisms affect monocyte responses in Crohn's disease

    Institute of Scientific and Technical Information of China (English)

    Dylan M Glubb; Richard B Gearry; Murray L Barclay; Rebecca L Roberts; John Pearson; Jacqui I Keenan; Judy McKenzie; Robert W Bentley

    2011-01-01

    AIM: To assess whether polymorphisms in NOD2 and ATG16L1 affect cytokine responses and mycobacterium avium subspecies paratuberculosis (MAP) survival in monocytes from Crohn's disease (CD) patients.METHODS: Monocytes were isolated from peripheral blood of CD patients of known genotype for common single nucleotide polymorphisms of NOD2 and ATG16L1 .Monocytes were challenged with MAP and bacterial persistence assessed at subsequent time-points. Cytokine responses were assayed using a Milliplex multi-analyte profiling assay for 13 cytokines.RESULTS: Monocytes heterozygous for a NOD2 polymorphism (R702W, P268S, or 1007fs) were more permissive for growth of MAP (P = 0.045) than those without. There was no effect of NOD2 genotype on subsequent cytokine expression. The T300A polymorphism of ATG16L1 did not affect growth of MAP in our model (P= 0.175), but did increase expression of cytokines interleukin (IL)-10 (P = 0.047) and IL-6 (P = 0.019).CONCLUSION: CD-associated polymorphisms affected the elimination of MAP from ex vivo monocytes (NOD2 ), or expression of certain cytokines (ATG16L1 ), implying independent but contributory roles in the pathogenesis of CD.

  11. Processing Focus Structure in L1 and L2 French: L2 Proficiency Effects on ERPs

    Science.gov (United States)

    Reichle, Robert V.; Birdsong, David

    2014-01-01

    This study examined the event-related potentials (ERPs) elicited by focus processing among first language (L1) speakers and second language (L2) learners of French. Participants read wh-questions containing explicit focus marking, followed by responses instantiating contrastive and informational focus. We hypothesized that L2 proficiency would…

  12. The Effect of L1 Transfer on Pronunciation of English Interdentals

    Institute of Scientific and Technical Information of China (English)

    张景京

    2011-01-01

    In this paper,the author makes a hypothesis that negative transfer from L1(Mandarin Chinese) has an effect on EFL learners' pronunciation of the two English interdentals /θ/ and //.To analyze this hypothesis the author applies OT(the Optimality Theory) to illustrate the way Mandarin Chinese influences the interdental production.

  13. Using L1 to Enhance the Grammar Learning and Having Only English Policy in EFL Classes

    Science.gov (United States)

    Uyar, Yusuf

    2012-01-01

    The purpose of this study is to determine the differences of grammar learning, if any, between the EFL classes in which native language (L1) is sometimes used and only target language (L2) is used. Participants were 42 prep year students from one of the universities in Turkey. They have been studying English for 9 months, and now they are in level…

  14. Cdk5 disruption attenuates tumor PD-L1 expression and promotes antitumor immunity

    Science.gov (United States)

    Dorand, R. Dixon; Nthale, Joseph; Myers, Jay T.; Barkauskas, Deborah S.; Avril, Stefanie; Chirieleison, Steven M.; Pareek, Tej K.; Abbott, Derek W.; Stearns, Duncan S.; Letterio, John J.

    2016-01-01

    Cancers often evade immune surveillance by adopting peripheral tissue–tolerance mechanisms, such as the expression of programmed cell death ligand 1 (PD-L1), the inhibition of which results in potent antitumor immunity. Here, we show that cyclin-dependent kinase 5 (Cdk5), a serine-threonine kinase that is highly active in postmitotic neurons and in many cancers, allows medulloblastoma (MB) to evade immune elimination. Interferon-γ (IFN-γ)-induced PD-L1 up-regulation on MB requires Cdk5, and disruption of Cdk5 expression in a mouse model of MB results in potent CD4+ T cell–mediated tumor rejection. Loss of Cdk5 results in persistent expression of the PD-L1 transcriptional repressors, the interferon regulatory factors IRF2 and IRF2BP2, which likely leads to reduced PD-L1 expression on tumors. Our finding highlights a central role for Cdk5 in immune checkpoint regulation by tumor cells. PMID:27463676

  15. L1/L2/L3 Writing Development: Longitudinal Case Study of a Japanese Multicompetent Writer

    Science.gov (United States)

    Kobayashi, Hiroe; Rinnert, Carol

    2013-01-01

    This longitudinal case study, supplemented by cross-sectional comparisons among five groups of writers with differing backgrounds, investigates how Natsu, a Japanese multilingual writer, developed her L1, L2 (English), and L3 (Chinese) writing competence over two and a half years. To create a comprehensive picture of this multilingual writer, the…

  16. Data of evolutionary structure change: 1CF8L-1MCEA [Confc[Archive

    Lifescience Database Archive (English)

    Full Text Available 1CF8L-1MCEA 1CF8 1MCE L A DIVLTQSPTIMSVSPGEKVTLTCSASSSV--SSNYVYWY...DINVKWKIDGSERQNGVLNSWTDQDSKDSTYSMSSTLTLTKDEYERHNSYTCEATHKTSTSPIVKSFNRNEC- PSALTQ-PPSASGSLGQSVTI...ntryChain> 1MCE A 1MCEA A 1MCEA KVTVLGQPKAN 1MCE A 1MCEA KPSKQ-SNNKY

  17. L1 Shapes L2 Auditory Representation Elicited Imitation of Arabic-Speaking Learners of English

    Directory of Open Access Journals (Sweden)

    Rajaa Aquil

    2012-05-01

    Full Text Available L1 interference plays a major role in second language acquisition, as evidenced by empirical studies (Kellerman & Sharwood Smith, 1986. The interference could result from a learner's conscious or unconscious judgment that some linguistic features in L1 and L2 are similar (Odlin, 1989, particularly in phonology (MacKain, Best, & Strange, 1981. This paper reports on two experiments using Elicited Imitation and Reading Tasks to investigate whether L1, Cairene Arabic prosodic strategy of epenthesis to break up consonant clusters is transferred to the participants' English output. Results of Experiment A showed that epenthesis took place more in reading than in repetition, as tested by the Elicited Imitation and Reading Tasks. Mimicking was suspected to be behind the results. To control for mimicking, a second experiment (Experiment B was conducted following the same design, but with the addition of a familiarity task to ensure that the participants knew and understood the words of an utterance and did not just mimic them. Results of Experiment B showed that epenthesis instances were the same in repetition as in reading. Epenthesis of a vowel to break consonant clusters suggests that participants of the study reconstructed the utterances based not only on how English words are stored in their mental representation, but also on Cairene Arabic syllable structure rules. This study, through the usage of Elicited Imitation Task, is able to tap into L2 Arabic speaking learners’ auditory mental representation of L2 input and demonstrate the influence of L1 transfer.

  18. Aspectual perspective taking in event construal in L1 and L2 Dutch

    NARCIS (Netherlands)

    Beek, G.; Flecken, M.E.P.; Starren, M.

    2013-01-01

    The present study focuses on the role of grammatical aspect in event construal and its function in encoding the specificity of an event. We investigate whether advanced L2 learners (L1 German) acquire target-like patterns of use of progressive aspect in Dutch, a language in which use of aspect depen

  19. Predicting the Nuclear Localization Signals of 107 Types of HPV L1 Proteins by Bioinformatic Analysis

    Institute of Scientific and Technical Information of China (English)

    Jun Yang; Yi-Li Wang; Lü-Sheng Si

    2006-01-01

    In this study, 107 types of human papillomavirus (HPV) L1 protein sequences were obtained from available databases, and the nuclear localization signals (NLSs) of these HPV L1 proteins were analyzed and predicted by bioinformatic analysis.Out of the 107 types, the NLSs of 39 types were predicted by PredictNLS software (35 types of bipartite NLSs and 4 types of monopartite NLSs). The NLSs of the remaining HPV types were predicted according to the characteristics and the homology of the already predicted NLSs as well as the general rule of NLSs.According to the result, the NLSs of 107 types of HPV L1 proteins were classified into 15 categories. The different types of HPV L1 proteins in the same NLS category could share the similar or the same nucleocytoplasmic transport pathway.They might be used as the same target to prevent and treat different types of HPV infection. The results also showed that bioinformatic technology could be used to analyze and predict NLSs of proteins.

  20. L1-L2 Convergence in Clausal Packaging in Japanese and English

    Science.gov (United States)

    Brown, Amanda; Gullberg, Marianne

    2013-01-01

    This study investigates L1-L2 convergence among bilinguals at an intermediate (CEFR-B2) level of L2 proficiency, focusing on the clausal packaging of Manner and Path of motion. Previous research has shown cross-linguistic differences between English and Japanese in this domain (Allen et al., 2003; Kita & Ozyurek, 2003, though note Brown &…

  1. mdm-2 gene amplification in 3T3-L1 preadipocytes.

    Science.gov (United States)

    Berberich, S J; Litteral, V; Mayo, L D; Tabesh, D; Morris, D

    1999-05-01

    In this study the regulation of the murine double minute-2 (mdm-2) gene was examined in NIH 3T3-L1 preadipocytes. The 3T3-L1 cell line, under proper conditions, has the capacity to differentiate from fibroblasts into adipocytes [15]. A recent report demonstrated that mdm-2 overexpression could block myogenesis [12]. While examining the regulation of the mdm-2 gene during adipogenesis, it was discovered that 3T3-L1 cells possess a 36-fold elevation of mdm-2 mRNA relative to A31 cells, another immortalized Balb/c 3T3 fibroblast cell line that lacks the capacity to differentiate. Based on Southern blot analysis, the increase in mdm-2 mRNA was the result of a mdm-2 gene amplification. The level of Mdm-2 protein in undifferentiated 3T3-L1 cells was elevated relative to A31 fibroblasts and resulted from translation of mRNA transcripts initiating from the p53-independent P1 promoter. We also examined how mdm-2 and p53 levels changed as undifferentiated fibroblasts converted to adipocytes. While mdm-2 mRNA levels remained elevated, p53 mRNA, protein, and DNA-binding activity decreased. These results suggest that adipogenesis is unaffected by elevated Mdm-2 levels and that the overexpression of mdm-2 mRNA is predominantly p53 independent.

  2. The future of cancer therapy: Selecting patients who respond to PD-1/L1 blockade

    Science.gov (United States)

    Ribas, Antoni; Tumeh, Paul C.

    2014-01-01

    Summary It is conceivable that, in the near future, an assay that defines the likelihood of a patient with advanced cancer to respond to immunotherapy based on PD-1/L1 blockade will be the initial decision point to select the treatment of patients with any cancer type. PMID:24970841

  3. Conceptualisations of "Grammar Teaching": L1 English Teachers' Beliefs about Teaching Grammar for Writing

    Science.gov (United States)

    Watson, Annabel Mary

    2015-01-01

    This paper reports on an investigation of L1 English teachers' conceptual and evaluative beliefs about teaching grammar, one strand of a larger Economic and Social Research Council (ESRC)-funded investigation into the impact of contextualised grammar teaching [RES-062-23-0775]. Thirty-one teachers in English secondary schools were interviewed…

  4. 26 CFR 36.3121(l)(1)-2 - Amendment of agreement.

    Science.gov (United States)

    2010-04-01

    ... under the same conditions, with respect to any one or more of the foreign subsidiaries of the domestic...) EMPLOYMENT TAXES AND COLLECTION OF INCOME TAX AT SOURCE CONTRACT COVERAGE OF EMPLOYEES OF FOREIGN SUBSIDIARIES § 36.3121(l)(1)-2 Amendment of agreement. (a) An agreement entered into by a domestic...

  5. Developmental Trends and L1 Effects in Early L2 Learners' Onset Cluster Production

    Science.gov (United States)

    Tessier, Anne-Michelle; Duncan, Tamara Sorenson; Paradis, Johanne

    2013-01-01

    This study focuses on English onset cluster production in spontaneous speech samples of 10 children aged 5;04-6;09 from Chinese and Hindi/Punjabi first language (L1) backgrounds, each with less than a year of exposure to English. The results suggest commonalities between early second language (L2) learners and both monolingual and adult L2…

  6. Reading Skills and Strategies: Assessing Primary School Students’ Awareness in L1 and EFL Strategy Use

    Directory of Open Access Journals (Sweden)

    Evdokimos Aivazoglou

    2014-09-01

    Full Text Available The present study was designed and conducted with the purpose to assess primary school students’ awareness in GL1 (Greek as first language and EFL (English as a foreign language strategy use and investigate the relations between the reported reading strategies use in first (L1 and foreign language (FL.  The sample (455 students attending the fifth and sixth grades of primary schools in Northern Greece was first categorized into skilled and less skilled L1 and EFL readers through screening reading comprehension tests, one in L1 and one in FL, before filling in the reading strategy questionnaires. The findings revealed participants’ preference for “problem solving” strategies, while “global strategies” coming next. Girls were proved to be more aware of their reading strategies use with the boys reporting a more frequent use in both languages. Also, skilled readers were found to use reading strategies more effectively, and appeared to be more flexible in transferring strategies from L1 to FL compared to less-skilled readers. Keywords: reading awareness, reading strategies, reading difficulties, primary school students

  7. Similarities of L1 (Mother Tongue) in Terms of Grammar and Language Structure in Translation

    Science.gov (United States)

    Subramaniam, Vijayaletchumy

    2011-01-01

    Malaysian philosophy, purpose and objective of the education system are rooted and based on policies stated in the National Education Policy 1956 and Education Act 1961. Whereas the Education Ordinance 1952 urged all Chinese and Tamil schools to be given an equal opportunity to learn English and Malay language together with their L1 (mother…

  8. Identification of dynamical hinge points of the L1 ligase molecular switch.

    Science.gov (United States)

    Giambasu, George M; Lee, Tai-Sung; Sosa, Carlos P; Robertson, Michael P; Scott, William G; York, Darrin M

    2010-04-01

    The L1 ligase is an in vitro selected ribozyme that uses a noncanonically base-paired ligation site to catalyze regioselectively and regiospecifically the 5' to 3' phosphodiester bond ligation, a reaction relevant to origin of life hypotheses that invoke an RNA world scenario. The L1 ligase crystal structure revealed two different conformational states that were proposed to represent the active and inactive forms. It remains an open question as to what degree these two conformers persist as stable conformational intermediates in solution, and along what pathway are they able to interconvert. To explore these questions, we have performed a series of molecular dynamics simulations in explicit solvent of the inactive-active conformational switch in L1 ligase. Four simulations were performed departing from both conformers in both the reactant and product states, in addition to a simulation where local unfolding in the active state was induced. From these simulations, along with crystallographic data, a set of four virtual torsion angles that span two evolutionarily conserved and restricted regions were identified as dynamical hinge points in the conformational switch transition. The ligation site visits three distinct states characterized by hydrogen bond patterns that are correlated with the formation of specific contacts that may promote catalysis. The insights gained from these simulations contribute to a more detailed understanding of the coupled catalytic/conformational switch mechanism of L1 ligase that may facilitate the design and engineering of new catalytic riboswitches.

  9. EXPLORING L1 INTERFERENCE IN THE WRITINGS OF KADAZANDUSUN ESL STUDENTS

    Directory of Open Access Journals (Sweden)

    Chelster Sherralyn Jeoffrey Pudin

    2015-07-01

    Full Text Available For many ethnic KadazanDusuns from Sabah, North Borneo, English is a third language after their mother tongue and Malay. The burden of having to contend with an additional language frequently leads to errors, particularly those caused by interference from the first language (L1. This study set out to identify the types and frequency of English language errors and their correlations in the writing of KadazanDusun ESL students at Universiti Malaysia Sabah. A further aim of the study was to establish which of these errors could be attributed to L1 interference. A total of 54 students with lower Malaysian University Entrance Test (MUET band scores were asked to complete a questionnaire and write a short essay on a designated topic. The language errors were categorized and analysed via statistical analysis. Errors considered to be related to L1 interference were then identified after consultation with an experienced KadazanDusun language lecturer. The most common errors were those involving singular /plural nouns and unusual sentence structures. The results show that approximately 25% of the errors were attributable to L1 interference, i.e. mode (normal/involuntary, voice (actor (-ing form /undergoer (-ed form, overuse of article, linker (when linker is used, no article is needed, auxiliary verb and direct translation. The findings of this study give ESL practitioners a better insight into student errors and should lead to improved writing performance in the classroom.

  10. A Prerequisite to L1 Homophone Effects in L2 Spoken-Word Recognition

    Science.gov (United States)

    Nakai, Satsuki; Lindsay, Shane; Ota, Mitsuhiko

    2015-01-01

    When both members of a phonemic contrast in L2 (second language) are perceptually mapped to a single phoneme in one's L1 (first language), L2 words containing a member of that contrast can spuriously activate L2 words in spoken-word recognition. For example, upon hearing cattle, Dutch speakers of English are reported to experience activation…

  11. THE ROLE OF L1 IN SECOND LANGUAGE ACQUISITION (SLA)

    Institute of Scientific and Technical Information of China (English)

    LiuTianyun

    2004-01-01

    This paper based on the related SLA theories, attempts to present the development of the roles of the first language on language learning before focusing on what have been found in this area. The purpose of the writing is to call the public attention on the rediscovery of the role L1 plays in SLA.

  12. MASA syndrome is caused by mutations in the neural cell adhesion gene, L1CAM

    Energy Technology Data Exchange (ETDEWEB)

    Schwartz, C.E.; Wang, Y.; Schroer, R.J.; Stevenson, R.E. [Greenwood Genetic Center, SC (United States)

    1994-09-01

    The MASA syndrome is a recessive X-linked disorder characterized by Mental retardation, Adducted thumbs, Shuffling gait and Aphasia. Recently we found that MASA in one family was likely caused by a point mutation in exon 6 of the L1CAM gene. This gene has also been shown to be involved in X-linked hydrocephalus (HSAS). We have screened 60 patients with either sporadic HSAS or MASA as well as two additional families with MASA. For the screening, we initially utilized 3 cDNA probes for the L1CAM gene. In one of the MASA families, K8310, two affected males were found to have an altered BglII band. The band was present in their carrier mother but not in their normal brothers. This band was detected by the entire cDNA probe as well as the cDNA probe for 3{prime} end of the gene. Analysis of the L1CAM sequence indicated the altered BglII site is distal to the exon 28 but proximal to the punative poly A signal site. It is hypothesized that this point mutation alters the stability of the L1CAM mRNA. This is being tested using cell lines established from the two affected males.

  13. An optimal L1-minimization algorithm for stationary Hamilton-Jacobi equations

    KAUST Repository

    Guermond, Jean-Luc

    2009-01-01

    We describe an algorithm for solving steady one-dimensional convex-like Hamilton-Jacobi equations using a L1-minimization technique on piecewise linear approximations. For a large class of convex Hamiltonians, the algorithm is proven to be convergent and of optimal complexity whenever the viscosity solution is q-semiconcave. Numerical results are presented to illustrate the performance of the method.

  14. L1 Influence on the Acquisition Order of English Grammatical Morphemes

    Science.gov (United States)

    Murakami, Akira; Alexopoulou, Theodora

    2016-01-01

    We revisit morpheme studies to evaluate the long-standing claim for a universal order of acquisition. We investigate the L2 acquisition order of six English grammatical morphemes by learners from seven L1 groups across five proficiency levels. Data are drawn from approximately 10,000 written exam scripts from the Cambridge Learner Corpus. The…

  15. L1-determined ideals in group algebras of exponential Lie groups

    CERN Document Server

    Ungermann, Oliver

    2012-01-01

    A locally compact group $G$ is said to be $\\ast$-regular if the natural map $\\Psi:\\Prim C^\\ast(G)\\to\\Prim_{\\ast} L^1(G)$ is a homeomorphism with respect to the Jacobson topologies on the primitive ideal spaces $\\Prim C^\\ast(G)$ and $\\Prim_{\\ast} L^1(G)$. In 1980 J. Boidol characterized the $\\ast$-regular ones among all exponential Lie groups by a purely algebraic condition. In this article we introduce the notion of $L^1$-determined ideals in order to discuss the weaker property of primitive $\\ast$-regularity. We give two sufficient criteria for closed ideals $I$ of $C^\\ast(G)$ to be $L^1$-determined. Herefrom we deduce a strategy to prove that a given exponential Lie group is primitive $\\ast$-regular. The author proved in his thesis that all exponential Lie groups of dimension $\\le 7$ have this property. So far no counter-example is known. Here we discuss the example $G=B_5$, the only critical one in dimension $\\le 5$.

  16. High ethanol tolerance of the thermophilic anaerobic ethanol producer Thermoanaerobacter BG1L1

    DEFF Research Database (Denmark)

    Georgieva, Tania I.; Mikkelsen, Marie Just; Ahring, Birgitte Kiær

    2007-01-01

    The low ethanol tolerance of thermophilic anaerobic bacteria, generally less than 2% (v/v) ethanol, is one of the main limiting factors for their potential use for second generation fuel ethanol production. In this work, the tolerance of thermophilic anaerobic bacterium Thermoanaerobacter BG 1L1 ...

  17. Reprogramming triggers endogenous L1 and Alu retrotransposition in human induced pluripotent stem cells.

    Science.gov (United States)

    Klawitter, Sabine; Fuchs, Nina V; Upton, Kyle R; Muñoz-Lopez, Martin; Shukla, Ruchi; Wang, Jichang; Garcia-Cañadas, Marta; Lopez-Ruiz, Cesar; Gerhardt, Daniel J; Sebe, Attila; Grabundzija, Ivana; Merkert, Sylvia; Gerdes, Patricia; Pulgarin, J Andres; Bock, Anja; Held, Ulrike; Witthuhn, Anett; Haase, Alexandra; Sarkadi, Balázs; Löwer, Johannes; Wolvetang, Ernst J; Martin, Ulrich; Ivics, Zoltán; Izsvák, Zsuzsanna; Garcia-Perez, Jose L; Faulkner, Geoffrey J; Schumann, Gerald G

    2016-01-08

    Human induced pluripotent stem cells (hiPSCs) are capable of unlimited proliferation and can differentiate in vitro to generate derivatives of the three primary germ layers. Genetic and epigenetic abnormalities have been reported by Wissing and colleagues to occur during hiPSC derivation, including mobilization of engineered LINE-1 (L1) retrotransposons. However, incidence and functional impact of endogenous retrotransposition in hiPSCs are yet to be established. Here we apply retrotransposon capture sequencing to eight hiPSC lines and three human embryonic stem cell (hESC) lines, revealing endogenous L1, Alu and SINE-VNTR-Alu (SVA) mobilization during reprogramming and pluripotent stem cell cultivation. Surprisingly, 4/7 de novo L1 insertions are full length and 6/11 retrotransposition events occurred in protein-coding genes expressed in pluripotent stem cells. We further demonstrate that an intronic L1 insertion in the CADPS2 gene is acquired during hiPSC cultivation and disrupts CADPS2 expression. These experiments elucidate endogenous retrotransposition, and its potential consequences, in hiPSCs and hESCs.

  18. Comparing Hypertext Reading in L1 and L2: The Case of Filipino Adults

    Science.gov (United States)

    Gruspe, Michael Angelo M.; Marinas, Christian Joshua L.; Villasin, Marren Nicole F.; Villanueva, Ariel Josephe Therese R.; Vizconde, Camilla J.

    2015-01-01

    This research probed into the reading experiences of adult readers in their first language (L1) and second language (L2). Qualitative in nature, the investigation focused on twelve (12) adult readers , six (6) males and six (6) females, whose first language is Filipino. Data were gathered through interviews and focus-group discussions. Based on…

  19. Reshaping Teacher Cognition about L1 Use through Critical ELT Teacher Education

    Science.gov (United States)

    Miri, Mowla; Alibakhshi, Goudarz; Mostafaei-Alaei, Mahnaz

    2017-01-01

    The authors report on a study aimed at exploring the role of a teacher education program (TEP) rested upon the tenets of critical pedagogy in influencing teachers' cognitions and practices concerning first-language (L1) use in second-language (L2) classrooms. Participants were 10 Iranian English as a foreign language teachers, whose cognitions…

  20. Fluorescence lifetime imaging of lipids during 3T3-L1 cell differentiation

    Science.gov (United States)

    Song, Young Sik; Won, Young Jae; Lee, Sang-Hak; Kim, Dug Young

    2014-03-01

    Obesity is becoming a big health problem in these days. Since increased body weight is due to increased number and size of the triglyceride-storing adipocytes, many researchers are working on differentiation conditions and processes of adipocytes. Adipocytes also work as regulators of whole-body energy homeostasis by secreting several proteins that regulate processes as diverse as haemostasis, blood pressure, immune function, angiogenesis and energy balance. 3T3-L1 cells are widely used cell line for studying adipogenesis because it can differentiate into an adipocyte-like phenotype under appropriate conditions. In this paper, we propose an effective fluorescence lifetime imaging technique which can easily distinguish lipids in membrane and those in lipid droplets. Nile red dyes are attached to lipids in 3T3-L1 cells. Fluorescence lifetime images were taken for 2 week during differentiation procedure of 3T3-L1 cells into adipocytes. We used 488 nm pulsed laser with 5MHz repetition rate and emission wavelength is 520 nm of Nile Red fluorescent dye. Results clearly show that the lifetime of Nile red in lipid droplets are smaller than those in cell membrane. Our results suggest that fluorescence lifetime imaging can be a very powerful tool to monitor lipid droplet formation in adipocytes from 3T3-L1 cells.