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Sample records for acetate esterase anae

  1. Determination of acid alpha-naphthyl acetate esterase enzyme activity in peripheral blood leukocytes of gazelles (Gazella subgutturosa).

    PubMed

    Altunay, H; Harem, I S; Harem, M K; Asti, R N; Kurtdede, N

    2008-12-01

    We examined gazelle peripheral blood leucocytes using the alpha-Naphthyl acetate esterase (ANAE) staining technique (pH 5.8). Our purpose was to determine the percentage of ANAE positive lymphocytes. The proportion of ANAE positive T-lymphocytes was 72%. T-lymphocytes showed an ANAE positive reaction, but eosinophilic granulocytes and monocytes also showed a positive reaction. By contrast, no reaction was detected in B-lymphocytes, neutrophil granulocytes or platelets. The reaction observed in T-lymphocytes was a red-brown coloration, usually 1-2 granules, but enough granules to fill the cytoplasm were detected rarely. As a result of ANAE enzyme staining, we concluded that the staining technique can be used as a cytochemical marker for gazelle T-lymphocytes. PMID:19085516

  2. Determination of organophosphorus pesticide residues in vegetables by an enzyme inhibition method using α-naphthyl acetate esterase extracted from wheat flour*

    PubMed Central

    Wang, Jun-liang; Xia, Qing; Zhang, An-ping; Hu, Xiao-yan; Lin, Chun-mian

    2012-01-01

    The widespread use of organophosphorus pesticides (OPs) poses a great threat to human health and has made the detection of OP residues in food an important task, especially in view of the fact that easy and rapid detection methods are needed. Because OPs have inhibitory effects on the activity of α-naphthyl acetate esterase (ANAE) in plants, in this work we evaluated the possibility of detecting OPs in vegetables with ANAE extracted from commercial flour. The limits of detection (LODs) obtained for methamidophos, dichlorvos, phoxim, dimethoate, and malathion in lettuce samples with crude ANAE were 0.17, 0.11, 0.11, 0.96, and 1.70 mg/kg, respectively. Based on the maximum residue limits (MRLs) for OPs in food stipulated by Chinese laws which are 0.05, 0.20, 0.05, 1.00, and 8.00 mg/kg for methamidophos, dichlorvos, phoxim, dimethoate, and malathion, respectively, the esterase inhibition method with crude ANAE had sufficient sensitivity to detect the residues of dichlorvos, dimethoate, and malathion in lettuce, but it could not be used to guarantee the safety of the same samples if methamidophos or phoxim residue was present. The sensitivity of the method was improved by the use of esterase purified by ammonium sulfate salting-out. The LODs obtained for methamidophos and phoxim with purified esterase were lower than the MRLs for these OPs in food. This is a very promising method for the detection of OP residues in vegetables using crude or purified esterase because of its cheapness, sensitivity, and convenience. PMID:22467368

  3. Differences in Esterase Activity to Aspirin and p-Nitrophenyl Acetate among Human Serum Albumin Preparations.

    PubMed

    Tatsumi, Akitoshi; Okada, Masaya; Inagaki, Yoshihiro; Inoue, Sachiyo; Hamaguchi, Tsuneo; Iwakawa, Seigo

    2016-01-01

    Human serum albumin (HSA) has two major ligand-binding sites, sites I and II, and also hydrolyzes some compounds at both sites. In the present study, we investigated differences in esterase activity among HSA preparations, and also the effects of warfarin, indomethacin, and naproxen on the hydrolytic activities of HSA to aspirin and p-nitrophenyl acetate. The esterase activities of HSA to aspirin or p-nitrophenyl acetate were measured from the pseudo-first-order formation rate constant (kobs) of salicylic acid or p-nitrophenol by HSA. Inter-lot variations were observed in the esterase activities of HSA to aspirin and p-nitrophenyl acetate; however, the esterase activity of HSA to aspirin did not correlate with that to p-nitrophenyl acetate. The inhibitory effects of warfarin and indomethacin on the esterase activity of HSA to aspirin were stronger than that of naproxen. In contrast, the inhibitory effect of naproxen on the esterase activity of HSA to p-nitrophenyl acetate was stronger than those of warfarin and indomethacin. These results suggest that the administration of different commercial HSA preparations and the co-administration with site I or II high-affinity binding drugs may change the pharmacokinetic profiles of drugs that are hydrolyzed by HSA. PMID:27476944

  4. Purification and characterization of an esterase involved in cellulose acetate degradation by Neisseria sicca SB.

    PubMed

    Moriyoshi, K; Ohmoto, T; Ohe, T; Sakai, K

    1999-10-01

    An esterase catalyzing the hydrolysis of acetyl ester moieties in cellulose acetate was purified 1,110-fold to electrophoretic homogeneity from the culture supernatant of Neisseria sicca SB, which can assimilate cellulose acetate as the sole carbon and energy source. The purified enzyme was a monomeric protein with a molecular mass of 40 kDa and the isoelectric point was 5.3. The pH and temperature optima of the enzyme were 8.0-8.5 and 45 degrees C. The enzyme catalyzed the hydrolysis of acetyl saccharides, p-nitrophenyl esters of short-chain fatty acids, and was slightly active toward aliphatic and aromatic esters. The K(m) and Vmax for cellulose acetate (degree of substitution, 0.88) and p-nitrophenyl acetate were 0.0162% (716 microM as acetyl content in the polymer) and 36.0 microM, and 66.8 and 39.1 mumol/min/mg, respectively. The enzyme was strongly inhibited by phenylmethylsulfonyl fluoride and diisopropyl fluorophosphate, which indicated that the enzyme was a serine esterase.

  5. Enantioselective Resolution of (±)-1-Phenylethanol and (±)-1-Phenylethyl Acetate by a Novel Esterase from Bacillus sp. SCSIO 15121.

    PubMed

    Liang, Jiayuan; Zhang, Yun; Sun, Aijun; Deng, Dun; Hu, Yunfeng

    2016-02-01

    A novel microbial esterase BSE01281 identified from the Indian Ocean was cloned, expressed, and functionally characterized. Esterase BSE01281 could enanoselectively resolve (±)-1-phenylethanol and (±)-1-phenylethyl acetate through two types of enzymatic reactions. After the optimization of enzymatic reactions, BSE01281 could efficiently generate (R)-1-phenylethyl acetate with high enantiomeric excess (>99%) and high conversion (42%) after 96 h trans-esterification reactions. Additionally, BSE01281 could also produce (R)-1-phenylethanol (e.e. > 99%) and (S)-1-phenylethyl acetate (e.e. > 95%) at a conversion of 49% through direct hydrolysis of inexpensive racemic 1-phenylethyl acetate for 8 h. Optically pure (R)-1-phenylethanol generated from direct enzymatic hydrolysis of racemic 1-phenylethyl acetate by BSE01281 is not easily prepared by dehydrogenases, which generally follow the "Prelog's rule" and give (S)-1-phenylethanol instead.

  6. Enhancement of acetyl xylan esterase activity on cellulose acetate through fusion to a family 3 cellulose binding module.

    PubMed

    Mai-Gisondi, Galina; Turunen, Ossi; Pastinen, Ossi; Pahimanolis, Nikolaos; Master, Emma R

    2015-11-01

    The current study investigates the potential to increase the activity of a family 1 carbohydrate esterase on cellulose acetate through fusion to a family 3 carbohydrate binding module (CBM). Specifically, CtCBM3 from Clostridium thermocellum was fused to the carboxyl terminus of the acetyl xylan esterase (AnAXE) from Aspergillus nidulans, and active forms of both AnAXE and AnAXE-CtCBM3 were produced in Pichia pastoris. CtCBM3 fusion had negligible impact on the thermostability or regioselectivity of AnAXE; activities towards acetylated corncob xylan, 4-methylumbelliferyl acetate, p-nitrophenyl acetate, and cellobiose octaacetate were also unchanged. By contrast, the activity of AnAXE-CtCBM3 on cellulose acetate increased by two to four times over 24 h, with greater differences observed at earlier time points. Binding studies using microcrystalline cellulose (Avicel) and a commercial source of cellulose acetate confirmed functional production of the CtCBM3 domain; affinity gel electrophoresis using acetylated xylan also verified the selectivity of CtCBM3 binding to cellulose. Notably, gains in enzyme activity on cellulose acetate appeared to exceed gains in substrate binding, suggesting that fusion to CtCBM3 increases functional associations between the enzyme and insoluble, high molecular weight cellulosic substrates.

  7. Cloning, Purification and Characterization of Acetyl Xylane Esterase from Anoxybacillus flavithermus DSM 2641(T) with Activity on Low Molecular-Weight Acetates.

    PubMed

    Eminoğlu, Ayşenur; Ülker, Serdar; Sandallı, Cemal

    2015-08-01

    Family 4 carbohydrate esterases (CE-4) have deacetylate different forms of acetylated poly/oligosaccharides in nature. This family is recognized with a specific polysaccharide deacetylase domain assigned as NodB homology domain in their secondary structure. Most family 4 carbohydrate esterases have been structurally and biochemically characterized. However, this is the first study about the enzymological function of pdaB-like CE4s from thermophilic bacterium Anoxybacillus flavithermus DSM 2641(T). A. flavithermus WK1 genome harbors five putative CE4 family genes. One of them is 762 bp long and encodes a protein of 253 amino acids in length and it was used as reference sequence in this study. It was described as acetyl xylane esterase (AXE) in genome project and this AfAXE gene was amplified without signal sequence and cloned. The recombinant protein was expressed in E. coli BL21 (DE3), purified by nickel affinity chromatography and its purity was visualized on SDS-PAGE. The activity of the recombinant enzyme was shown by zymogram analysis with α-naphtyl acetate as a substrate. The enzyme was characterized spectrophotometrically using chromogenic p-nitrophenyl acetate. Optimum temperature and pH were determined as 50 °C and 7.5, respectively. Km and Vmax were determined as 0.43 mM and 3333.33 U/mg, respectively under optimum conditions. To our knowledge this is the first enzymological characterization of a pdaB-like family 4 carbohydrate esterase from the members of Anoxybacillus genus.

  8. Geranyl acetate esterase controls and regulates the level of geraniol in lemongrass (Cymbopogon flexuosus Nees ex Steud.) mutant cv. GRL-1 leaves.

    PubMed

    Ganjewala, Deepak; Luthra, Rajesh

    2009-01-01

    Essential oil isolated from lemongrass (Cymbopogon flexuosus) mutant cv. GRL-1 leaves is mainly composed of geraniol (G) and geranyl acetate (GA). The proportion of G and GA markedly fluctuates during leaf development. The proportions of GA and G in the essential oil recorded at day 10 after leaf emergence were approximately 59% and approximately 33% respectively. However, the level of GA went down from approximately 59 to approximately 3% whereas the level of G rose from approximately 33 to approximately 91% during the leaf growth period from day 10 to day 50. However, the decline in the level of GA was most pronounced in the early (day 10 to day 30) stage of leaf growth. The trend of changes in the proportion of GA and G has clearly indicated the role of an esterase that must be involved in the conversion of GA to G during leaf development. We isolated an esterase from leaves of different ages that converts GA into G and has been given the name geranyl acetate esterase (GAE). The GAE activity markedly varied during the leaf development cycle; it was closely correlated with the monoterpene (GA and G) composition throughout leaf development. GAE appeared as several isoenzymes but only three (GAE-I, GAE-II, and GAE-III) of them had significant GA cleaving activity. The GAE isoenzymes pattern was greatly influenced by the leaf developmental stages and so their GA cleaving activities. Like the GAE activity, GAE isoenzyme patterns were also found to be consistent with the monoterpene (GA and G) composition. GAE had an optimum pH at 8.5 and temperature at 30 degrees C. Besides GAE, a compound with phosphatase activity capable of hydrolyzing geranyl diphosphate (GPP) to produce geraniol has also been isolated.

  9. Use of 'small but smart' libraries to enhance the enantioselectivity of an esterase from Bacillus stearothermophilus towards tetrahydrofuran-3-yl acetate.

    PubMed

    Nobili, Alberto; Gall, Markus G; Pavlidis, Ioannis V; Thompson, Mark L; Schmidt, Marlen; Bornscheuer, Uwe T

    2013-07-01

    Two libraries of simultaneous double mutations in the active site region of an esterase from Bacillus stearothermophilus were constructed to improve the enantioselectivity in the hydrolysis of tetrahydrofuran-3-yl acetate. As screening of large mutant libraries is hampered by the necessity for GC/MS analysis, mutant libraries were designed according to a 'small but smart' concept. The design of focused libraries was based on data derived from a structural alignment of 3317 amino acid sequences of α/β-hydrolase fold enzymes with the bioinformatic tool 3DM. In this way, the number of mutants to be screened was substantially reduced as compared with a standard site-saturation mutagenesis approach. Whereas the wild-type esterase showed only poor enantioselectivity (E = 4.3) in the hydrolysis of (S)-tetrahydrofuran-3-yl acetate, the best variants obtained with this approach showed increased E-values of up to 10.4. Furthermore, some variants with inverted enantiopreference were found. PMID:23331978

  10. Esterase patterns of species in the Drosophila buzzatii cluster.

    PubMed

    Lapenta, A S; de Campos Bicudo, H E; Ceron, C R; Cordeiro, J A

    1995-01-01

    A comparative analysis was made of the esterase isoenzyme patterns of eight iso-female lines, four of Drosophila serido (B31 D1, A55, B59, Q1, B50Q3), two of D. koepferae (B20D2 and B25D7), one of D. seriema (A95) and one of D. buzzatii (Buz). In all, 43 bands in the spectrum of esterase isoenzymes were detected by electrophoresis in polyacrylamide gels. They showed variations in specific reactions with alpha and beta-naphthyl acetate, number of patterns yielded in their intra-isofemale line combinations, frequencies of such combinations and the thickness and staining degree of some bands, in different individuals, lines and species. Among bands detected exclusively in males, seven may be considered sex-specific (5 alpha-esterases and 2 beta-esterases). These male-specific alpha-esterases have in common the inability to cleave beta-naphthyl acetate in the absence of alpha-naphthyl, denoting a possible common function. The similarity index (SI) and analysis of dependence were calculated in an attempt to quantify the differentiation of the iso-female lines studied, on the basis of esterase bands. SI mean value allowed the separation of the isofemale lines into five classes. Each species had its own pattern of esterase bands, but some bands were shared. A divergence hypothesis for the isofemale lines and the species is discussed.

  11. Esterases and putative lipases from tropical isolates of Aureobasidium pullulans.

    PubMed

    Kudanga, Tukayi; Mwenje, Eddie; Mandivenga, Faith; Read, John S

    2007-04-01

    Esterases and lipases have been studied in a number of fungi, though very little is known about esterases from Aureobasidium pullulans especially from the African tropics. In this study, forty-two Zimbabwean isolates were screened for lipase activity on tributyrin agar. Extracellular esterase activities of seven selected isolates were studied under varying conditions using para-nitrophenol acetate as substrate. Twenty isolates (48%) showed lipolytic activity; sixteen showed negative results for lipase activity while the rest showed weak activities. Esterase activities in broth cultures ranged from 0.011-0.223 mmol/microg protein/min while activities ranged from 1.5-12.8 U/ml under solid state fermentation. The esterases were optimally active at pH 7.6-8.0, showed a temperature optimum of 35 degrees C and retained more than 50% activity at temperatures up to 60 degrees C and at pH 4.0-7.0 after 150 min. Enzyme production was optimal after 5-6 days with diammonium hydrogen phosphate as nitrogen source. Isolates showed variations in preference for carbon source for esterase production. The A. pullulans esterases differed from most fungal esterases in that they are optimally active in alkaline conditions and are active over a broad pH range. PMID:17440916

  12. Antinuclear Antibodies (ANA)

    MedlinePlus

    ... drugs you take. ANA testing can produce a “false positive.” This typically signals the presence of antinuclear ... keep looking. In fact, you may have a “false positive” ANA, which means that the evidence is ...

  13. Esterase Activity and Intracellular Localization in Reconstructed Human Epidermal Cultured Skin Models

    PubMed Central

    Katayanagi, Mishina; Hashimoto, Fumie

    2015-01-01

    Background Reconstructed human epidermal culture skin models have been developed for cosmetic and pharmaceutical research. Objective This study evaluated the total and carboxyl esterase activities (i.e., Km and Vmax, respectively) and localization in two reconstructed human epidermal culture skin models (LabCyte EPI-MODEL [Japan Tissue Engineering] and EpiDerm [MatTek/Kurabo]). The usefulness of the reconstruction cultured epidermis was also verified by comparison with human and rat epidermis. Methods Homogenized epidermal samples were fractioned by centrifugation. p-nitrophenyl acetate and 4-methylumbelliferyl acetate were used as substrates of total esterase and carboxyl esterase, respectively. Results Total and carboxyl esterase activities were present in the reconstructed human epidermal culture skin models and were localized in the cytosol. Moreover, the activities and localization were the same as those in human and rat epidermis. Conclusion LabCyte EPI-MODEL and EpiDerm are potentially useful for esterase activity prediction in human epidermis. PMID:26082583

  14. Esterase patterns and phylogenetic relationships of Drosophila species in the saltans subgroup (saltans group).

    PubMed

    Nascimento, A P; de Campos, Bicudo H E M

    2002-01-01

    The esterase patterns of sixteen strains from four species in the saltans subgroup were analyzed using polyacrylamide gel electrophoresis. Thirty-four esterase bands were detected. By using alpha and beta naphthyl acetates as substrates, they were classified in 18 alpha-esterases (they hydrolyse the alpha-naphtyl substrate), 15 beta-esterases (they hydrolyse the beta-naphtyl substrate) and 1 alpha/beta-esterase (it hydrolyses the alpha and beta-naphtyl substrates). Among the alpha-esterases, three were detected exclusively in males. Malathion, Eserine and pCMB were used as inhibitors in order to characterize biochemically the esterases. The results indicated the presence of cholinesterases, carboxylesterases and acetylesterases. The degree of mobility of the bands in the gels, their specificity to alpha and beta naphthyl acetates and the results of the inhibition tests allowed us to recognize tentatively nine genetic loci. Phylogenetic relationships among species inferred on the basis of the esterase patterns by PAUP 4.0b8, with neighbor-joining search and a bootstrap analysis showed that, although the four species are closely related, D. septentriosaltans, D. saltans and D. austrosaltans are closer to each other than to D. prosaltans. These results showed to be consistent with phylogenetic relationships previously inferred from inversion polymorphism.

  15. A New Family of Carbohydrate Esterases Is Represented by a GDSL Hydrolase/Acetylxylan Esterase from Geobacillus stearothermophilus*

    PubMed Central

    Alalouf, Onit; Balazs, Yael; Volkinshtein, Margarita; Grimpel, Yael; Shoham, Gil; Shoham, Yuval

    2011-01-01

    Acetylxylan esterases hydrolyze the ester linkages of acetyl groups at positions 2 and/or 3 of the xylose moieties in xylan and play an important role in enhancing the accessibility of xylanases to the xylan backbone. The hemicellulolytic system of the thermophilic bacterium Geobacillus stearothermophilus T-6 comprises a putative acetylxylan esterase gene, axe2. The gene product belongs to the GDSL hydrolase family and does not share sequence homology with any of the carbohydrate esterases in the CAZy Database. The axe2 gene is induced by xylose, and the purified gene product completely deacetylates xylobiose peracetate (fully acetylated) and hydrolyzes the synthetic substrates 2-naphthyl acetate, 4-nitrophenyl acetate, 4-methylumbelliferyl acetate, and phenyl acetate. The pH profiles for kcat and kcat/Km suggest the existence of two ionizable groups affecting the binding of the substrate to the enzyme. Using NMR spectroscopy, the regioselectivity of Axe2 was directly determined with the aid of one-dimensional selective total correlation spectroscopy. Methyl 2,3,4-tri-O-acetyl-β-d-xylopyranoside was rapidly deacetylated at position 2 or at positions 3 and 4 to give either diacetyl or monoacetyl intermediates, respectively; methyl 2,3,4,6-tetra-O-acetyl-β-d-glucopyranoside was initially deacetylated at position 6. In both cases, the complete hydrolysis of the intermediates occurred at a much slower rate, suggesting that the preferred substrate is the peracetate sugar form. Site-directed mutagenesis of Ser-15, His-194, and Asp-191 resulted in complete inactivation of the enzyme, consistent with their role as the catalytic triad. Overall, our results show that Axe2 is a serine acetylxylan esterase representing a new carbohydrate esterase family. PMID:21994937

  16. 1. 'SANTA ANA RIVER IN SANTA ANA CANYON. ORANGE COUNTY.' ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    1. 'SANTA ANA RIVER IN SANTA ANA CANYON. ORANGE COUNTY.' This is an oblique aerial view to the northeast taken from the northeast extremity of the canyon, showing, in the middle distance, the confluence of Chino Creek and the Santa Ana River, site of the future Prado Dam. File number written on negative: R & H 80 026. - Prado Dam, Santa Ana River near junction of State Highways 71 & 91, Corona, Riverside County, CA

  17. Production, Purification, and Properties of Extracellular Carboxyl Esterases from Bacillus subtilis NRRL 365

    PubMed Central

    Meghji, K.; Ward, O. P.; Araujo, A.

    1990-01-01

    Bacillus subtilis NRRL 365 produced high extracellular carboxyl esterase activity in submerged culture media containing wheat bran, corn steep liquor, and salts. Supplementation of this medium with glucose reduced esterase activity to 37% of that in the unsupplemented control. Esterase activity was purified by ammonium sulfate fractionation, DEAE-Sephadex A-50 ion-exchange chromatography with sodium chloride gradient elution, and preparative polyacrylamide gel electrophoresis. The resultant purified components, esterases I and II, manifested single bands following silver staining of polyacrylamide gel electrophoresis gels and had final specific activities of 80 and 520 U/mg, respectively. Molecular weights for components I and II were 36,000 and 105,000 to 110,000, respectively. Esterases I and II both had a pH optimum of 8.0, with relative activities of 10 and 85%, respectively, at pH 9.0. Kms with p-nitrophenylacetate were 0.91 mM for esterase I and 0.67 mM for esterase II. In general, patterns of enzyme inhibition were similar for both components. Differences were observed in the relative activities of esterases I and II towards p-nitrophenyl esters of acetate, propionate, and butyrate; Activity ratios for components I and II were 100:94:48 and 100:36:23, respectively. The purified components did not hydrolyze long-chain triglycerides and did not manifest proteolytic activity. Images PMID:16348375

  18. Production and purification of a solvent-resistant esterase from Bacillus licheniformis S-86.

    PubMed

    Torres, Sebastián; Baigorí, Mario D; Pandey, Ashok; Castro, Guillermo R

    2008-12-01

    New thermophilic and organic-solvent-tolerant Bacillus licheniformis S-86 strain is able to produce two active and solvent-stable esterases. Production of type I and II esterases was substantially enhanced when oils and surfactants were supplied as carbon sources. Grape oil (0.1% v/v) and Tween 20 to 60 (0.1% v/v) had enhanced enzyme production between 1.6- and 2.2-folds. Type II esterase was purified to homogeneity in a five-step procedure. This esterase was purified 76.7-fold with a specific activity of 135 U mg(-1). Molecular mass of the enzyme was estimated to be 38.4 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Type II esterase was active mostly on esters with short acyl chains, which allowed to classify the enzyme as a carboxylesterase with a K (m) of 80.2 mmol l(-1) and a V (max) of 256.4 micromol min(-1) mg(-1) for p-nitrophenyl acetate. Also, B. licheniformis S-86 type II esterase displayed activity in presence of water-miscible organic solvents at 50% concentration and stability after 1-h incubation. PMID:18543118

  19. Esterases immobilized on aminosilane modified magnetic nanoparticles as a catalyst for biotransformation reactions.

    PubMed

    Alex, Deepthy; Mathew, Abraham; Sukumaran, Rajeev K

    2014-09-01

    Magnetite nanoparticles were prepared by reacting ferrous and ferric salts in presence of aqueous ammonia. The magnetic nanoparticles (MNPs) were amino functionalized by treating with 3-aminopropyl triethoxy silane (APTES) and was coupled with glutaraldehyde. A novel solvent tolerant esterase from Pseudozyma sp. NII 08165 was immobilized on the MNPs through covalent bonding to the glutaraldehyde. The magnetite nanoparticles had a size range of 10-100 nm, confirmed by DLS. Lipases immobilized on MNPs were evaluated for biotransformation reactions including synthesis of ethyl acetate and transesterification of vegetable oil for producing biodiesel. The MNP immobilized esterase had prolonged shelf life and there was no loss in enzyme activity. PMID:24968816

  20. Esterases immobilized on aminosilane modified magnetic nanoparticles as a catalyst for biotransformation reactions.

    PubMed

    Alex, Deepthy; Mathew, Abraham; Sukumaran, Rajeev K

    2014-09-01

    Magnetite nanoparticles were prepared by reacting ferrous and ferric salts in presence of aqueous ammonia. The magnetic nanoparticles (MNPs) were amino functionalized by treating with 3-aminopropyl triethoxy silane (APTES) and was coupled with glutaraldehyde. A novel solvent tolerant esterase from Pseudozyma sp. NII 08165 was immobilized on the MNPs through covalent bonding to the glutaraldehyde. The magnetite nanoparticles had a size range of 10-100 nm, confirmed by DLS. Lipases immobilized on MNPs were evaluated for biotransformation reactions including synthesis of ethyl acetate and transesterification of vegetable oil for producing biodiesel. The MNP immobilized esterase had prolonged shelf life and there was no loss in enzyme activity.

  1. Immunoelectron microscopic demonstration of an esterase on the outer membrane of Xanthomonas maltophilia.

    PubMed Central

    Debette, J; Prensier, G

    1989-01-01

    Xanthomonas maltophilia (later synonym of Pseudomonas maltophilia), an ubiquitous species, is known to show proteolytic and lipolytic activities. A cell-bound esterase which hydrolyzes beta-naphthyl acetate during growth has been extracted from a strain isolated from soil. Because of its strongly hydrophobic character, the enzyme could be efficiently solubilized only by Triton X-100. This nonionic detergent must be added in polyacrylamide gels to permit migration. Polyclonal rabbit antibodies raised against the Triton-soluble esterase complex were used to localize the enzyme at the ultrastructural level. Electron microscopy of cell sections of this organism and immunogold labeling demonstrated that the enzyme was located on the outer membrane. Such an envelope-bound esterase may produce assimilable substrates for X. maltophilia which can grow in various environments. Images PMID:2495761

  2. Santa Ana Forecasting and Classification

    NASA Astrophysics Data System (ADS)

    Rolinski, T.; Eichhorn, D.; D'Agostino, B. J.; Vanderburg, S.; Means, J. D.

    2011-12-01

    Southern California experiences wildfires every year, but under certain circumstances these fires grow into extremely large and destructive fires, such as the Cedar Fire of 2003 and the Witch Fire of 2007. The Cedar Fire burned over 1100 km2 , destroyed more than 2200 homes and killed 15 people; the Witch fire burned more than 800 km2, destroyed more than 1000 homes and killed 2 people. Fires can quickly become too large and dangerous to fight if they are accompanied by a very strong "Santa Ana" condition, which is a foehn-like wind that may bring strong winds and very low humidities. However there is an entire range of specific weather conditions that fall into the broad category of Santa Anas, from cold and blustery to hot with very little wind. All types are characterized by clear skies and low humidity. Since the potential for destructive fire is dependent on the characteristics of Santa Anas, as well as the level of fuel moisture, there exists a need for further classification, such as is done with tropical cyclones and after-the-fact with tornadoes. We use surface data and fuel moisture combined with reanalysis to diagnose those conditions that result in Santa Anas with the greatest potential for destructive fires. We use this data to produce a new classification system for Santa Anas. This classification system should be useful for informing the relevant agencies for mitigation and response planning. In the future this same classification may be made available to the general public.

  3. A thermoactive uropygial esterase from chicken: purification, characterisation and synthesis of flavour esters.

    PubMed

    Fendri, Ahmed; Louati, Hanen; Sellami, Mohamed; Gargouri, Héla; Smichi, Nabil; Zarai, Zied; Aissa, Imen; Miled, Nabil; Gargouri, Youssef

    2012-06-01

    A lipolytic activity was located in the chicken uropygial glands, from which a carboxylesterase (CUE) was purified. Pure CUE has an apparent molecular mass of 50 kDa. The purified esterase displayed its maximal activity (200 U/mg) on short-chain triacylglycerols (tributyrin) at a temperature of 50°C. No significant lipolytic activity was found when medium chain (trioctanoin) or long chain (olive oil) triacylglycerols were used as substrates. The enzyme retained 75% of its maximal activity when incubated during 2h at 50°C. The NH(2)-terminal amino acid sequence showed similarities with the esterase purified recently from turkey pharyngeal tissue. Esterase activity remains stable after its incubation during 30 min in presence of organic solvents such as hexane or butanol. CUE is a serine enzyme since it was inactivated by phenylmethanesulphonyl fluoride (PMSF), a serine-specific inhibitor. The purified enzyme, which tolerates the presence of some organic solvent and a high temperature, can be used in non-aqueous synthesis reactions. Hence, the uropygial esterase immobilised onto CaCO(3) was tested to produce the isoamyl and the butyl acetate (flavour esters). Reactions were performed at 50°C in presence of hexane. High synthesis yields of 91 and 67.8% were obtained for isoamyl and butyl acetate, respectively. PMID:22531158

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

    PubMed

    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. PMID:27554142

  5. Comparison of fungal carbohydrate esterases of family CE16 on artificial and natural substrates.

    PubMed

    Puchart, Vladimír; Agger, Jane W; Berrin, Jean-Guy; Várnai, Anikó; Westereng, Bjørge; Biely, Peter

    2016-09-10

    The enzymatic conversion of acetylated hardwood glucuronoxylan to functional food oligomers, biochemicals or fermentable monomers requires besides glycoside hydrolases enzymes liberating acetic acid esterifying position 2 and/or 3 in xylopyranosyl (Xylp) residues. The 3-O-acetyl group at internal Xylp residues substituted by MeGlcA is the only acetyl group of hardwood acetylglucuronoxylan and its fragments not attacked by acetylxylan esterases of carbohydrate esterase (CE) families 1, 4, 5 and 6 and by hemicellulolytic acetyl esterases classified in CE family 16. Monoacetylated aldotetraouronic acid 3″-Ac(3)MeGlcA(3)Xyl3, generated from the polysaccharide by GH10 endoxylanases, appears to be one of the most resistant fragments. The presence of the two substituents on the non-reducing-end Xylp residue prevents liberation of MeGlcA by α-glucuronidase of family GH67 and blocks the action of acetylxylan esterases. The Ac(3)MeGlcA(3)Xyl3 was isolated from an enzymatic hydrolysate of birchwood acetylglucuronoxylan and characterized by (1)H NMR spectroscopy as a mixture of two positional isomers, 3″-Ac(3)MeGlcA(3)Xyl3 and 4″-Ac(3)MeGlcA(3)Xyl3, the latter being the result of acetyl group migration. The mixture was used as a substrate for three members of CE16 family of fungal origin. Trichoderma reesei CE16 esterase, inactive on polymeric substrate, deacetylated both isomers. Podospora anserina and Aspergillus niger esterases, active on acetylglucuronoxylan, deesterified effectively only the 4″-isomer. The results indicate catalytic diversity among CE16 enzymes, but also their common and unifying catalytic ability to exo-deacetylate positions 3 and 4 on non-reducing-end Xylp residues, which is an important step in plant hemicellulose saccharification. PMID:27439201

  6. Purification and Characterization of Anacardium occidentale (Cashew) Allergens Ana o 1, Ana o 2, and Ana o 3.

    PubMed

    Reitsma, Marit; Bastiaan-Net, Shanna; Sforza, Stefano; van der Valk, Johanna P M; van Gerth van Wijk, Roy; Savelkoul, Huub F J; de Jong, Nicolette W; Wichers, Harry J

    2016-02-10

    In this study a fast and simple purification procedure for the three known allergens from cashew (7S globulin Ana o 1, 11S globulin Ana o 2, and 2S albumin Ana o 3) is described. The purified allergens are characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), Western blot, glycoprotein stain, and protein identification. The purified proteins still bind IgE, and this IgE binding varied between different pools of patient serum. Ana o 1 was found to be a glycoprotein. Ana o 3 has been studied more in detail to identify both the small and large subunits, both displaying microheterogeneity, and epitope mapping of Ana o 3 has been performed. PMID:26769082

  7. Purification and Characterization of Anacardium occidentale (Cashew) Allergens Ana o 1, Ana o 2, and Ana o 3.

    PubMed

    Reitsma, Marit; Bastiaan-Net, Shanna; Sforza, Stefano; van der Valk, Johanna P M; van Gerth van Wijk, Roy; Savelkoul, Huub F J; de Jong, Nicolette W; Wichers, Harry J

    2016-02-10

    In this study a fast and simple purification procedure for the three known allergens from cashew (7S globulin Ana o 1, 11S globulin Ana o 2, and 2S albumin Ana o 3) is described. The purified allergens are characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), Western blot, glycoprotein stain, and protein identification. The purified proteins still bind IgE, and this IgE binding varied between different pools of patient serum. Ana o 1 was found to be a glycoprotein. Ana o 3 has been studied more in detail to identify both the small and large subunits, both displaying microheterogeneity, and epitope mapping of Ana o 3 has been performed.

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

    PubMed Central

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

    2015-01-01

    Background: In the earlier studies, methanolic extract of Mangifera indica L leaf was exhibited hypocholesterol activity. However, the bioactive compounds responsible for the same are not reported so far. Objective: To isolate the bioactive compounds with hypocholesterol activity from the leaf extract using cholesterol esterase inhibition assay which can be used for the standardization of extract. Materials and Methods: The leaf methanolic extract of M. indica (Sindoora variety) was partitioned with ethyl acetate and chromatographed on silica gel to yield twelve fractions and the activity was monitored by using cholesterol esterase inhibition assay. Active fractions were re-chromatographed to yield individual compounds. Results and Discussion: A major compound mangiferin present in the extract was screened along with other varieties of mango leaves for cholesterol esterase inhibition assay. However, the result indicates that compounds other than mangiferin may be active in the extract. Invitro pancreatic cholesterol esterase inhibition assay was used for bioactivity guided fractionation (BAGF) to yield bioactive compound for standardization of extract. Bioactivity guided fractionation afford the active fraction containing 3b-taraxerol with an IC50 value of 0.86μg/ml. Conclusion: This study demonstrates that M. indica methanol extract of leaf have significant hypocholesterol activity which is standardized with 3b-taraxerol, a standardized extract for hypocholesterol activity resulted in development of dietary supplement from leaves of Mangifera indica. PMID:26692750

  9. Isolation of acetyl esterase mutants of Bacillus subtilis 168.

    PubMed Central

    Higerd, T B

    1977-01-01

    Five mutants of Bacillus subtilis 168 defective in an intracellular esterase activity were identified. By polyacrylamide gel electrophoresis, four of the mutants were shown to lack esterase B activity, and the fifth lacked esterase A activity. All of the back-crossed esterase mutants were able to sporulate at wild-type frequency and produce exoprotease(s) and antibiotic(s). No difference in motility could be attributed to the esterase mutation. PBS1 transduction analysis showed all the esterase B mutations to be linked to the hisA marker. Images PMID:402361

  10. Novel Redox-Dependent Esterase Activity (EC 3.1.1.2) for DJ-1: Implications for Parkinson's Disease.

    PubMed

    Vázquez-Mayorga, Emmanuel; Díaz-Sánchez, Ángel G; Dagda, Ruben K; Domínguez-Solís, Carlos A; Dagda, Raul Y; Coronado-Ramírez, Cynthia K; Martínez-Martínez, Alejandro

    2016-01-01

    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. PMID:27556455

  11. Association of esterases with insecticide resistance in Culex quinquefasciatus (Diptera: Culicidae).

    PubMed

    Gordon, Jennifer R; Ottea, James

    2012-06-01

    The southern house mosquito, Culex quinquefasciatus Say, is a competent vector of human disease and an important target of mosquito abatement programs. However, these management programs have been compromised by development of insecticide resistance. In the current study, susceptibilities to naled and resmethrin, two adulticides used in mosquito abatement, were monitored using a topical and contact bioassay, respectively, in five field- collected populations of C. quinquefasciatus (MARC, HOOD1, HOOD2, MINLOVE, and THIB). Frequencies of resistance, measured as survival after treatment with discriminating concentrations (i.e., sufficient to kill > 90% of a reference susceptible strain) were high (88.0-96.8%) in all field collections treated with naled, but were variable (3.3-94.2%) with resmethrin. In addition, esterase activities in mosquitoes from these collections were quantified using alpha-naphthyl acetate and ranged from 1.08 to 3.39 micromol alpha-naphthol produced min(-1) mg prot(-1). Heightened activities were associated with decreased insecticide susceptibility in HOOD1, THIB, and MINLOVE but not HOOD2. Esterases were visualized using native polyacrylamide gel electrophoresis, and intra- and interstrain differences in banding patterns were detected. In addition, esterases from MINLOVE mosquitoes were more numerous and intensely staining when compared with those from a laboratory-susceptible strain. Finally, naled synergized the toxicity of resmethrin in populations with decreased insecticide susceptibility and increased esterase activity by 2.5-(MINLOVE) to three-fold (THIB). Results from this study will allow management strategies for populations of C. quinquefasciatus to be optimized, and provide a foundation for further studies exploring use of esterase inhibitors as synergists of pyrethroid toxicity. PMID:22812138

  12. Phenolic acid esterases, coding sequences and methods

    DOEpatents

    Blum, David L.; Kataeva, Irina; Li, Xin-Liang; Ljungdahl, Lars G.

    2002-01-01

    Described herein are four phenolic acid esterases, three of which correspond to domains of previously unknown function within bacterial xylanases, from XynY and XynZ of Clostridium thermocellum and from a xylanase of Ruminococcus. The fourth specifically exemplified xylanase is a protein encoded within the genome of Orpinomyces PC-2. The amino acids of these polypeptides and nucleotide sequences encoding them are provided. Recombinant host cells, expression vectors and methods for the recombinant production of phenolic acid esterases are also provided.

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

    SciTech Connect

    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

  14. Mesoxalaldehyde acetals

    SciTech Connect

    Gordeeva, G.N.; Kalashnikov, S.M.; Popov, Yu.N.; Kruglov, E.A.; Imashev, U.B.

    1987-11-10

    The treatment of methylglyoxal acetals by alkyl nitrites in the presence of the corresponding aliphatic alcohols and hydrochloric acid leads to the formation of linear mesoxalaldehyde acetals, whose structure was established by NMR spectroscopy and mass spectrometry. The major pathways for the decomposition of these molecules upon electron impact were established.

  15. Uncovering divergent evolution of α/β-hydrolases: a surprising residue substitution needed to convert Hevea brasiliensis hydroxynitrile lyase into an esterase.

    PubMed

    Nedrud, David M; Lin, Hui; Lopez, Gilsinia; Padhi, Santosh K; Legatt, Graig A; Kaz-Lauskas, Romas J

    2014-11-01

    Hevea brasiliensis hydroxynitrile lyase (HbHNL) and salicylic acid binding protein 2 (SABP2, an esterase) share 45% amino acid sequence identity, the same protein fold, and even the same catalytic triad of Ser-His-Asp. However, they catalyze different reactions: cleavage of hydroxynitriles and hydrolysis of esters, respectively. To understand how other active site differences in the two enzymes enable the same catalytic triad to catalyze different reactions, we substituted amino acid residues in HbHNL with the corresponding residues from SABP2, expecting hydroxynitrile lyase activity to decrease and esterase activity to increase. Previous mechanistic studies and x-ray crystallography suggested that esterase activity requires removal of an active site lysine and threonine from the hydroxynitrile lyase. The Thr11Gly Lys236Gly substitutions in HbHNL reduced hydroxynitrile lyase activity for cleavage of mandelonitrile 100-fold, but increased esterase activity only threefold to kcat ~ 0.1 min(-1) for hydrolysis of p-nitrophenyl acetate. Adding a third substitution - Glu79His - increased esterase activity more than tenfold to kcat ~ 1.6 min(-1). The specificity constant (kcat/KM) for this triple substitution variant versus wild type HbHNL shifted more than one million-fold from hydroxynitrile lyase activity (acetone cyanohydrin substrate) to esterase activity (p-nitrophenyl acetate substrate). The contribution of Glu79His to esterase activity was surprising since esterases and lipases contain many different amino acids at this position, including glutamate. Saturation mutagenesis at position 79 showed that 13 of 19 possible amino acid substitutions increased esterase activity, suggesting that removal of glutamate, not addition of histidine, increased esterase activity. Molecular modeling indicates that Glu79 disrupts esterase activity in HbHNL when its negatively charged side chain distorts the orientation of the catalytic histidine. Naturally occurring glutamate at the

  16. Uncovering divergent evolution of α/β-hydrolases: a surprising residue substitution needed to convert Hevea brasiliensis hydroxynitrile lyase into an esterase

    PubMed Central

    Nedrud, David M.; Lin, Hui; Lopez, Gilsinia; Padhi, Santosh K.; Legatt, Graig A.

    2014-01-01

    Hevea brasiliensis hydroxynitrile lyase (HbHNL) and salicylic acid binding protein 2 (SABP2, an esterase) share 45% amino acid sequence identity, the same protein fold, and even the same catalytic triad of Ser-His-Asp. However, they catalyze different reactions: cleavage of hydroxynitriles and hydrolysis of esters, respectively. To understand how other active site differences in the two enzymes enable the same catalytic triad to catalyze different reactions, we substituted amino acid residues in HbHNL with the corresponding residues from SABP2, expecting hydroxynitrile lyase activity to decrease and esterase activity to increase. Previous mechanistic studies and x-ray crystallography suggested that esterase activity requires removal of an active site lysine and threonine from the hydroxynitrile lyase. The Thr11Gly Lys236Gly substitutions in HbHNL reduced hydroxynitrile lyase activity for cleavage of mandelonitrile 100-fold, but increased esterase activity only threefold to kcat ~ 0.1 min−1 for hydrolysis of p-nitrophenyl acetate. Adding a third substitution – Glu79His – increased esterase activity more than tenfold to kcat ~ 1.6 min−1. The specificity constant (kcat/KM) for this triple substitution variant versus wild type HbHNL shifted more than one million-fold from hydroxynitrile lyase activity (acetone cyanohydrin substrate) to esterase activity (p-nitrophenyl acetate substrate). The contribution of Glu79His to esterase activity was surprising since esterases and lipases contain many different amino acids at this position, including glutamate. Saturation mutagenesis at position 79 showed that 13 of 19 possible amino acid substitutions increased esterase activity, suggesting that removal of glutamate, not addition of histidine, increased esterase activity. Molecular modeling indicates that Glu79 disrupts esterase activity in HbHNL when its negatively charged side chain distorts the orientation of the catalytic histidine. Naturally occurring glutamate at

  17. Uncovering divergent evolution of α/β-hydrolases: a surprising residue substitution needed to convert Hevea brasiliensis hydroxynitrile lyase into an esterase.

    PubMed

    Nedrud, David M; Lin, Hui; Lopez, Gilsinia; Padhi, Santosh K; Legatt, Graig A; Kaz-Lauskas, Romas J

    2014-11-01

    Hevea brasiliensis hydroxynitrile lyase (HbHNL) and salicylic acid binding protein 2 (SABP2, an esterase) share 45% amino acid sequence identity, the same protein fold, and even the same catalytic triad of Ser-His-Asp. However, they catalyze different reactions: cleavage of hydroxynitriles and hydrolysis of esters, respectively. To understand how other active site differences in the two enzymes enable the same catalytic triad to catalyze different reactions, we substituted amino acid residues in HbHNL with the corresponding residues from SABP2, expecting hydroxynitrile lyase activity to decrease and esterase activity to increase. Previous mechanistic studies and x-ray crystallography suggested that esterase activity requires removal of an active site lysine and threonine from the hydroxynitrile lyase. The Thr11Gly Lys236Gly substitutions in HbHNL reduced hydroxynitrile lyase activity for cleavage of mandelonitrile 100-fold, but increased esterase activity only threefold to kcat ~ 0.1 min(-1) for hydrolysis of p-nitrophenyl acetate. Adding a third substitution - Glu79His - increased esterase activity more than tenfold to kcat ~ 1.6 min(-1). The specificity constant (kcat/KM) for this triple substitution variant versus wild type HbHNL shifted more than one million-fold from hydroxynitrile lyase activity (acetone cyanohydrin substrate) to esterase activity (p-nitrophenyl acetate substrate). The contribution of Glu79His to esterase activity was surprising since esterases and lipases contain many different amino acids at this position, including glutamate. Saturation mutagenesis at position 79 showed that 13 of 19 possible amino acid substitutions increased esterase activity, suggesting that removal of glutamate, not addition of histidine, increased esterase activity. Molecular modeling indicates that Glu79 disrupts esterase activity in HbHNL when its negatively charged side chain distorts the orientation of the catalytic histidine. Naturally occurring glutamate at the

  18. VvMJE1 of the grapevine (Vitis vinifera) VvMES methylesterase family encodes for methyl jasmonate esterase and has a role in stress response

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The known members of the plant methyl esterase (MES) family catalyze hydrolysis of a C-O ester linkage of methyl esters of several phytohormones including indole-3-acetic acid, salicylic acid, and jasmonic acid. The genome of grapevine (Vitis vinifera) was found to contain 15 MES genes, designated V...

  19. Phenol esterase activity of porcine skin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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 ...

  20. New Extremophilic Lipases and Esterases from Metagenomics

    PubMed Central

    López-López, Olalla; Cerdán, Maria E; González Siso, Maria I

    2014-01-01

    Lipolytic enzymes catalyze the hydrolysis of ester bonds in the presence of water. In media with low water content or in organic solvents, they can catalyze synthetic reactions such as esterification and transesterification. Lipases and esterases, in particular those from extremophilic origin, are robust enzymes, functional under the harsh conditions of industrial processes owing to their inherent thermostability and resistance towards organic solvents, which combined with their high chemo-, regio- and enantioselectivity make them very attractive biocatalysts for a variety of industrial applications. Likewise, enzymes from extremophile sources can provide additional features such as activity at extreme temperatures, extreme pH values or high salinity levels, which could be interesting for certain purposes. New lipases and esterases have traditionally been discovered by the isolation of microbial strains producing lipolytic activity. The Genome Projects Era allowed genome mining, exploiting homology with known lipases and esterases, to be used in the search for new enzymes. The Metagenomic Era meant a step forward in this field with the study of the metagenome, the pool of genomes in an environmental microbial community. Current molecular biology techniques make it possible to construct total environmental DNA libraries, including the genomes of unculturable organisms, opening a new window to a vast field of unknown enzymes with new and unique properties. Here, we review the latest advances and findings from research into new extremophilic lipases and esterases, using metagenomic approaches, and their potential industrial and biotechnological applications. PMID:24588890

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

    PubMed Central

    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

  2. Structure and activity of the cold-active and anion-activated carboxyl esterase OLEI01171 from the oil-degrading marine bacterium Oleispira antarctica

    PubMed Central

    Lemak, Sofia; Tchigvintsev, Anatoli; Petit, Pierre; Flick, Robert; Singer, Alexander U.; Brown, Greg; Evdokimova, Elena; Egorova, Olga; Gonzalez, Claudio F.; Chernikova, Tatyana N.; Yakimov, Michail M.; Kube, Michael; Reinhardt, Richard; Golyshin, Peter N.; Savchenko, Alexei; Yakunin, Alexander F.

    2014-01-01

    The uncharacterized α/β-hydrolase protein OLEI01171 from the psychrophilic marine bacterium Oleispira antarctica belongs to the PF00756 family of putative esterases, which also includes human esterase D. In the present paper we show that purified recombinant OLEI01171 exhibits high esterase activity against the model esterase substrate α-naphthyl acetate at 5 – 30°C with maximal activity at 15–20°C. The esterase activity of OLEI01171 was stimulated 3–8-fold by the addition of chloride or several other anions (0.1–1.0 M). Compared with mesophilic PF00756 esterases, OLEI01171 exhibited a lower overall protein thermostability. Two crystal structures ofOLEI01171 were solved at 1.75 and 2.1 Å resolution and revealed a classical serine hydrolase catalytic triad and the presence of a chloride or bromide ion bound in the active site close to the catalytic Ser148.Both anions were found to co-ordinate a potential catalytic water molecule located in the vicinity of the catalytic triad His257. The results of the present study suggest that the bound anion perhaps contributes to the polarization of the catalytic water molecule and increases the rate of the hydrolysis of an acyl-enzyme intermediate. Alanine replacement mutagenesis of OLEI01171 identified ten amino acid residues important for esterase activity. The replacement of Asn225 by lysine had no significant effect on the activity or thermostability of OLEI01171, but resulted in a detectable increase of activity at 35–45°C. The present study has provided insight into the molecular mechanisms of activity of a cold-active and anion-activated carboxyl esterase. PMID:22519667

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

    PubMed

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

    2008-01-01

    Two of the six esterases identified in Cucurbita pepo cv. "Eskandrani" were purified to homogeneity using two chromatography steps: anion exchange and gel filtration. The molecular weights of C. pepo esterases EIc and EII were 50,000 +/- 1500 and 68,000 +/- 1900 Da from gel filtration and 47,000 and 66,000 Da from SDS/PAGE, respectively, suggesting a monomeric structure for both enzymes. Esterases EIc and EII had K(m) values of 1.22 and 1.56 mM and pH optima at 9.0 and 8.0, respectively. The substrate specificity of C. pepo esterases EIc and EII were determined for a number of p-nitrophenyl esters, where their affinity toward these substrates were decreased as carbon atom number increased. Esterases EIc and EII had the same temperature optima, 40 degrees C. Thermal stability studies of esterases EIc and EII indicated that half maximal activities of EIc and EII esterases were reached at 55 degrees C and 50 degrees C, while they lost 45%, 51% and 70%, 77% of their activities after 30 and 90 min of incubation at 40 degrees C, respectively. The effect of different metal cations and inhibitors were examined. The inhibition studies revealed that the active sites of the two esterases contain serine and cysteine residues. The characteristics of C. pepo esterases are closely similar to those of microbial esterases used in food processing and food industry. PMID:17321740

  4. Characterization of two metagenome-derived esterases that reactivate chloramphenicol by counteracting chloramphenicol acetyltransferase.

    PubMed

    Tao, Weixin; Lee, Myung Hwan; Yoon, Mi-Young; Kim, Jin-Cheol; Malhotra, Shweta; Wu, Jing; Hwang, Eul Chul; Lee, Seon-Woo

    2011-12-01

    Function-driven metagenomic analysis is a powerful approach to screening for novel biocatalysts. In this study, we investigated lipolytic enzymes selected from an alluvial soil metagenomic library, and identified two novel esterases, EstDL26 and EstDL136. EstDL26 and EstDL136 reactivated chloramphenicol from its acetyl derivates by counteracting the chloramphenicol acetyltransferase (CAT) activity in Escherichia coli. These two enzymes showed only 27% identity in amino acid sequence to each other; however both preferentially hydrolyzed short-chain p-nitrophenyl esters (< or =C5) and showed mesophilic properties. In vitro, EstDL136 catalyzed the deacetylation of 1- and 3- acetyl and 1,3-diacetyl derivates; in contrast, EstDL26 was not capable of the deacetylation at C1, indicating a potential regioselectivity. EstDL26 and EstDL136 were similar to microbial hormone-sensitive lipase (HSL), and since chloramphenicol acetate esterase (CAE) activity was detected from two other soil esterases in the HSL family, this suggests a distribution of CAE among the soil microorganisms. The isolation and characterization of EstDL26 and EstDL136 in this study may be helpful in understanding the diversity of CAE enzymes and their potential role in releasing active chloramphenicol in the producing bacteria. PMID:22210605

  5. Plasma B-esterase activities in European raptors.

    PubMed

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

    2005-01-01

    Strigidae families were characterized by a BChE contribution that dominated the total ChE activity, while in the species of the Falconidae family, AChE activity dominated. With the exception of the barn owl, CbE activity (eserine-insensitive alpha-naphthyl acetate esterase [alpha-NAE] activity) in all species was almost absent or very low. The values obtained in this study for ChE, AChE, and BChE activities and the AChE:BChE ratios for buzzard, kestrel, barn owl, and tawny owl provide a good estimate of the normal values in free-living individuals of these European species. They can be used as a baseline to evaluate the effect of anticholinesterase insecticides in the field. PMID:15827224

  6. Xylella fastidiosa esterase rather than hydroxynitrile lyase.

    PubMed

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

    2015-03-01

    In 2009, we reported that the product of the gene SCJ21.16 (XFa0032) from Xylella fastidiosa, a xylem-restricted plant pathogen that causes a range of diseases in several important crops, encodes a protein (XfHNL) with putative hydroxynitrile lyase activity. Sequence analysis and activity tests indicated that XfHNL exhibits an α/β-hydrolase fold and could be classified as a member of the family of FAD-independent HNLs. Here we provide a more detailed sequence analysis and new experimental data. Using pure heterologously expressed XfHNL we show that this enzyme cannot catalyse the cleavage/synthesis of mandelonitrile and that this protein is in fact a non-enantioselective esterase. Homology modelling and ligand docking simulations were used to study the active site and support these results. This finding could help elucidate the common ancestor of esterases and hydroxynitrile lyases with an α/β -hydrolase fold. PMID:25684099

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

    PubMed Central

    De Santi, Concetta; Willassen, Nils Peder

    2016-01-01

    Background 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. Results 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. PMID:27433797

  8. Novel choline esterase based sensor for monitoring of organophosphorus pollutants

    SciTech Connect

    Wilkins, E.S.; Ghindilis, A.L.; Atanasov, P.

    1996-12-31

    Organophosphorus compounds are significant major environmental pollutants due to their intensive use as pesticides. The modern techniques based on inhibition of choline esterase enzyme activity are discussed. Potentiometric electrodes based on detection of choline esterase inhibition by analytes has been developed. The detection of choline esterase activity is based on the novel principle of molecular transduction. Immobilized peroxidase acting as the molecular transducer, catalyzes the electroreduction of hydrogen peroxide by direct (mediatorless) electron transfer. The sensing element consists of a carbon based electrode containing an assembly of co-immobilized enzymes: choline esterase, choline oxidase and peroxidase.

  9. Ethyl acetate

    Integrated Risk Information System (IRIS)

    Ethyl acetate ; CASRN 141 - 78 - 6 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinogenic Eff

  10. Phenylmercuric acetate

    Integrated Risk Information System (IRIS)

    Phenylmercuric acetate ; CASRN 62 - 38 - 4 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinog

  11. Vinyl acetate

    Integrated Risk Information System (IRIS)

    Vinyl acetate ; CASRN 108 - 05 - 4 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinogenic Eff

  12. Ammonium acetate

    Integrated Risk Information System (IRIS)

    Ammonium acetate ; CASRN 631 - 61 - 8 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinogenic

  13. Thallium acetate

    Integrated Risk Information System (IRIS)

    Jump to main content . Integrated Risk Information System Recent Additions | Contact Us Search : All EPA IRIS • You are here : EPA Home • Research • Environmental Assessment • IRIS • IRIS Summaries Redirect Page As of September 30 , 2009 , the assessment summary for Thallium acetate is included in t

  14. Adsorption of microbial esterases on Bacillus subtilis-templated cobalt oxide nanoparticles.

    PubMed

    Jang, Eunjin; Ryu, Bum Han; Shim, Hyun-Woo; Ju, Hansol; Kim, Dong-Wan; Kim, T Doohun

    2014-04-01

    Due to low diffusion rates and large surface areas, nanomaterials have received great interest as supporting materials for enzyme immobilization. Here, the preparation of a cobalt oxide nanoparticle using Bacillus subtilis as a biological template and use of the nanostructure for microbial esterase immobilization is described. Morphological features and size distributions were investigated using electron microscopy (EM) and dynamic light scattering (DLS). Catalytic properties of enzyme-coated nanostructures were investigated using 4-methylumbelliferyl acetate and p-nitrophenyl (PNP) acetate as model substrates. Enzyme-coated nanostructures were observed to retain ∼85% of the initial activity after 15 successive reaction cycles, and enzyme immobilization processes could be repeated four times without a loss of immobilization potential. The present work demonstrates that B. subtilis-templated cobalt oxide nanoparticles have the potential to be used as biocompatible immobilization materials, and are promising candidates for the preparation of effective nanobiocatalysts.

  15. Biochemical studies on a versatile esterase that is most catalytically active with polyaromatic esters

    PubMed Central

    Martínez-Martínez, Mónica; Lores, Iván; Peña-García, Carlina; Bargiela, Rafael; Reyes-Duarte, Dolores; Guazzaroni, María-Eugenia; Peláez, Ana Isabel; Sánchez, Jesús; Ferrer, Manuel

    2014-01-01

    Herein, we applied a community genomic approach using a naphthalene-enriched community (CN1) to isolate a versatile esterase (CN1E1) from the α/β-hydrolase family. The protein shares low-to-medium identity (≤ 57%) with known esterase/lipase-like proteins. The enzyme is most active at 25–30°C and pH 8.5; it retains approximately 55% of its activity at 4°C and less than 8% at ≥ 55°C, which indicates that it is a cold-adapted enzyme. CN1E1 has a distinct substrate preference compared with other α/β-hydrolases because it is catalytically most active for hydrolysing polyaromatic hydrocarbon (phenanthrene, anthracene, naphthalene, benzoyl, protocatechuate and phthalate) esters (7200–21 000 units g−1 protein at 40°C and pH 8.0). The enzyme also accepts 44 structurally different common esters with different levels of enantio-selectivity (1.0–55 000 units g−1 protein), including (±)-menthyl-acetate, (±)-neomenthyl acetate, (±)-pantolactone, (±)-methyl-mandelate, (±)-methyl-lactate and (±)-glycidyl 4-nitrobenzoate (in that order). The results provide the first biochemical evidence suggesting that such broad-spectrum esterases may be an ecological advantage for bacteria that mineralize recalcitrant pollutants (including oil refinery products, plasticizers and pesticides) as carbon sources under pollution pressure. They also offer a new tool for the stereo-assembly (i.e. through ester bonds) of multi-aromatic molecules with benzene rings that are useful for biology, chemistry and materials sciences for cases in which enzyme methods are not yet available. PMID:24418210

  16. Biochemical studies on a versatile esterase that is most catalytically active with polyaromatic esters.

    PubMed

    Martínez-Martínez, Mónica; Lores, Iván; Peña-García, Carlina; Bargiela, Rafael; Reyes-Duarte, Dolores; Guazzaroni, María-Eugenia; Peláez, Ana Isabel; Sánchez, Jesús; Ferrer, Manuel

    2014-03-01

    Herein, we applied a community genomic approach using a naphthalene-enriched community (CN1) to isolate a versatile esterase (CN1E1) from the α/β-hydrolase family. The protein shares low-to-medium identity (≤ 57%) with known esterase/lipase-like proteins. The enzyme is most active at 25-30°C and pH 8.5; it retains approximately 55% of its activity at 4°C and less than 8% at ≥ 55°C, which indicates that it is a cold-adapted enzyme. CN1E1 has a distinct substrate preference compared with other α/β-hydrolases because it is catalytically most active for hydrolysing polyaromatic hydrocarbon (phenanthrene, anthracene, naphthalene, benzoyl, protocatechuate and phthalate) esters (7200-21 000 units g(-1) protein at 40°C and pH 8.0). The enzyme also accepts 44 structurally different common esters with different levels of enantio-selectivity (1.0-55 000 units g(-1) protein), including (±)-menthyl-acetate, (±)-neomenthyl acetate, (±)-pantolactone, (±)-methyl-mandelate, (±)-methyl-lactate and (±)-glycidyl 4-nitrobenzoate (in that order). The results provide the first biochemical evidence suggesting that such broad-spectrum esterases may be an ecological advantage for bacteria that mineralize recalcitrant pollutants (including oil refinery products, plasticizers and pesticides) as carbon sources under pollution pressure. They also offer a new tool for the stereo-assembly (i.e. through ester bonds) of multi-aromatic molecules with benzene rings that are useful for biology, chemistry and materials sciences for cases in which enzyme methods are not yet available.

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

    PubMed

    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.

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

    PubMed

    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. PMID:25586442

  19. Decreasing the Level of Ethyl Acetate in Ethanolic Fermentation Broths of Escherichia coli KO11 by Expression of Pseudomonas putida estZ Esterase†

    PubMed Central

    Hasona, Adnan; York, S. W.; Yomano, L. P.; Ingram, L. O.; Shanmugam, K. T.

    2002-01-01

    During the fermentation of sugars to ethanol relatively high levels of an undesirable coproduct, ethyl acetate, are also produced. With ethanologenic Escherichia coli strain KO11 as the biocatalyst, the level of ethyl acetate in beer containing 4.8% ethanol was 192 mg liter−1. Although the E. coli genome encodes several proteins with esterase activity, neither wild-type strains nor KO11 contained significant ethyl acetate esterase activity. A simple method was developed to rapidly screen bacterial colonies for the presence of esterases which hydrolyze ethyl acetate based on pH change. This method allowed identification of Pseudomonas putida NRRL B-18435 as a source of this activity and the cloning of a new esterase gene, estZ. Recombinant EstZ esterase was purified to near homogeneity and characterized. It belongs to family IV of lipolytic enzymes and contains the conserved catalytic triad of serine, aspartic acid, and histidine. As expected, this serine esterase was inhibited by phenylmethylsulfonyl fluoride and the histidine reagent diethylpyrocarbonate. The native and subunit molecular weights of the recombinant protein were 36,000, indicating that the enzyme exists as a monomer. By using α-naphthyl acetate as a model substrate, optimal activity was observed at pH 7.5 and 40°C. The Km and Vmax for α-naphthyl acetate were 18 μM and 48.1 μmol · min−1 · mg of protein−1, respectively. Among the aliphatic esters tested, the highest activity was obtained with propyl acetate (96 μmol · min−1 · mg of protein−1), followed by ethyl acetate (66 μmol · min−1 · mg of protein−1). Expression of estZ in E. coli KO11 reduced the concentration of ethyl acetate in fermentation broth (4.8% ethanol) to less than 20 mg liter−1. PMID:12039716

  20. Novel Redox-Dependent Esterase Activity (EC 3.1.1.2) for DJ-1: Implications for Parkinson’s Disease

    PubMed Central

    Vázquez-Mayorga, Emmanuel; Díaz-Sánchez, Ángel G.; Dagda, Ruben K.; Domínguez-Solís, Carlos A.; Dagda, Raul Y.; Coronado-Ramírez, Cynthia K.; Martínez-Martínez, Alejandro

    2016-01-01

    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. PMID:27556455

  1. 53. SIPHON NO. 1, SANTA ANA RIVER NO. 2 PROJECT, ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    53. SIPHON NO. 1, SANTA ANA RIVER NO. 2 PROJECT, EXHIBIT L, PROJECT 1933, MAY 1973. SCE drawing no. 5110869 (sheet no. 11; for filing with Federal Power Commission). - Santa Ana River Hydroelectric System, Redlands, San Bernardino County, CA

  2. 4. INTERIOR OF ABANDONED SANTA ANA CANAL TUNNEL, SHOWING CEMENT ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    4. INTERIOR OF ABANDONED SANTA ANA CANAL TUNNEL, SHOWING CEMENT TROUGH FLOOR AND UNFINISHED GRANITE ROOF. VIEW TO SOUTHWEST. - Santa Ana River Hydroelectric System, Abandoned Tunnel, Redlands, San Bernardino County, CA

  3. 42. FOUNDATIONS TAIL RACE, ETC., POWER HOUSE SANTA ANA ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    42. FOUNDATIONS - TAIL RACE, ETC., POWER HOUSE SANTA ANA RIVER NO. 2, EDISON ELECTRIC CO., NOV. 3, 1904. SCE drawing no. 5393. - Santa Ana River Hydroelectric System, SAR-2 Powerhouse, Redlands, San Bernardino County, CA

  4. 34. ELEVATION OF RELAY AND CONTROL SWITCHBOARD, SANTA ANA RIVER ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    34. ELEVATION OF RELAY AND CONTROL SWITCHBOARD, SANTA ANA RIVER P.H. #3, JUNE 23, 1943. SCE drawing no. 413187-1. - Santa Ana River Hydroelectric System, SAR-3 Powerhouse, San Bernardino National Forest, Redlands, San Bernardino County, CA

  5. 57. ELECTRICAL EQUIPMENT PLOT PLAN, SANTA ANA NO. 1 HYDRO ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    57. ELECTRICAL EQUIPMENT PLOT PLAN, SANTA ANA NO. 1 HYDRO PLANT, OCTOBER 10, 1958. SCE drawing no. 428615-0. - Santa Ana River Hydroelectric System, SAR-1 Powerhouse, Redlands, San Bernardino County, CA

  6. 53. NEW BCB AND LIGHTNING ARRESTER ARRANGEMENT, SANTA ANA RIVER ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    53. NEW BCB AND LIGHTNING ARRESTER ARRANGEMENT, SANTA ANA RIVER NO. 2, JAN. 24, 1977. SCE drawing no. 455670-0. - Santa Ana River Hydroelectric System, SAR-2 Powerhouse, Redlands, San Bernardino County, CA

  7. 60. NEEDLE AND NOZZLE TIP, SANTA ANA NO. 1, SOUTHERN ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    60. NEEDLE AND NOZZLE TIP, SANTA ANA NO. 1, SOUTHERN CALIFORNIA EDISON CO., APR. 28, 1910, REVISED MAY 12, 1910. SCE drawing no. 4500. - Santa Ana River Hydroelectric System, SAR-1 Powerhouse, Redlands, San Bernardino County, CA

  8. Structural features determining thermal adaptation of esterases.

    PubMed

    Kovacic, Filip; Mandrysch, Agathe; Poojari, Chetan; Strodel, Birgit; Jaeger, Karl-Erich

    2016-02-01

    The adaptation of microorganisms to extreme living temperatures requires the evolution of enzymes with a high catalytic efficiency under these conditions. Such extremophilic enzymes represent valuable tools to study the relationship between protein stability, dynamics and function. Nevertheless, the multiple effects of temperature on the structure and function of enzymes are still poorly understood at the molecular level. Our analysis of four homologous esterases isolated from bacteria living at temperatures ranging from 10°C to 70°C suggested an adaptation route for the modulation of protein thermal properties through the optimization of local flexibility at the protein surface. While the biochemical properties of the recombinant esterases are conserved, their thermal properties have evolved to resemble those of the respective bacterial habitats. Molecular dynamics simulations at temperatures around the optimal temperatures for enzyme catalysis revealed temperature-dependent flexibility of four surface-exposed loops. While the flexibility of some loops increased with raising the temperature and decreased with lowering the temperature, as expected for those loops contributing to the protein stability, other loops showed an increment of flexibility upon lowering and raising the temperature. Preserved flexibility in these regions seems to be important for proper enzyme function. The structural differences of these four loops, distant from the active site, are substantially larger than for the overall protein structure, indicating that amino acid exchanges within these loops occurred more frequently thereby allowing the bacteria to tune atomic interactions for different temperature requirements without interfering with the overall enzyme function.

  9. 2. 'SANTA ANA RIVER AT CHINO CREEK, RIVERSIDE COUNTY.' This ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    2. 'SANTA ANA RIVER AT CHINO CREEK, RIVERSIDE COUNTY.' This is an oblique aerial view to the north, looking over the flooded fields between Chino Creek and the Santa Ana River, just upstream of the Prado Dam site. File number written on negative: R & H 80 024. - Prado Dam, Santa Ana River near junction of State Highways 71 & 91, Corona, Riverside County, CA

  10. 51. INTAKE AND POWER HOUSE AREAS, SANTA ANA NO. 1; ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    51. INTAKE AND POWER HOUSE AREAS, SANTA ANA NO. 1; DETAIL MAP OF SANTA ANA NO. 1 AND NO. 2 HYDROELECTRIC PROJECT, EXHIBIT K, APR. 30, 1945. SCE drawing no. 523690 (sheet no. 5; for filing with the Federal Power Commission). - Santa Ana River Hydroelectric System, Redlands, San Bernardino County, CA

  11. Degradation of vinyl acetate by soil, sewage, sludge, and the newly isolated aerobic bacterium V2.

    PubMed Central

    Nieder, M; Sunarko, B; Meyer, O

    1990-01-01

    Vinyl acetate is subject to microbial degradation in the environment and by pure cultures. It was hydrolyzed by samples of soil, sludge, and sewage at rates of up to 6.38 and 1 mmol/h per g (dry weight) under aerobic and anaerobic conditions, respectively. Four yeasts and thirteen bacteria that feed aerobically on vinyl acetate were isolated. The pathway of vinyl acetate degradation was studied in bacterium V2. Vinyl acetate was degraded to acetate as follows: vinyl acetate + NAD(P)+----2 acetate + NAD(P)H + H+. The acetate was then converted to acetyl coenzyme A and oxidized through the tricarboxylic acid cycle and the glyoxylate bypass. The key enzyme of the pathway is vinyl acetate esterase, which hydrolyzed the ester to acetate and vinyl alcohol. The latter isomerized spontaneously to acetaldehyde and was then converted to acetate. The acetaldehyde was disproportionated into ethanol and acetate. The enzymes involved in the metabolism of vinyl acetate were studied in extracts. Vinyl acetate esterase (Km = 6.13 mM) was also active with indoxyl acetate (Km = 0.98 mM), providing the basis for a convenient spectrophotometric test. Substrates of aldehyde dehydrogenase were formaldehyde, acetaldehyde, propionaldehyde, and butyraldehyde. The enzyme was equally active with NAD+ or NADP+. Alcohol dehydrogenase was active with ethanol (Km = 0.24 mM), 1-propanol (Km = 0.34 mM), and 1-butanol (Km = 0.16 mM) and was linked to NAD+. The molecular sizes of aldehyde dehydrogenase and alcohol dehydrogenase were 145 and 215 kilodaltons, respectively. PMID:2285314

  12. [Role of Human Orphan Esterases in Drug-induced Toxicity].

    PubMed

    Fukami, Tatsuki

    2015-01-01

    Esterases hydrolyze compounds containing ester, amide, and thioester bonds, causing prodrug activation or detoxification. Among esterases, carboxylesterases have been studied in depth due to their ability to hydrolyze a variety of drugs. However, there are several drugs for which the involved esterase(s) is unknown. We found that flutamide, phenacetin, rifamycins (rifampicin, rifabutin, and rifapentine), and indiplon are hydrolyzed by arylacetamide deacetylase (AADAC), which is highly expressed in human liver and gastrointestinal tissues. Flutamide hydrolysis is considered associated with hepatotoxicity. Phenacetin, a prodrug of acetaminophen, was withdrawn due to side effects such as methemoglobinemia and renal failure. It was demonstrated in vitro and in vivo using mice that AADAC is responsible for phenacetin hydrolysis, which leads to methemoglobinemia. In addition, it was shown that AADAC-mediated hydrolysis attenuates the cytotoxicity of rifamycins. Thus AADAC plays critical roles in drug-induced toxicity. Another orphan esterase, α/β hydrolase domain containing 10 (ABHD10), was found responsible for deglucuronidation of acyl-glucuronides including mycophenolic acid acyl-glucuronide and probenecid acyl-glucuronide. Because acyl-glucuronides appear associated with toxicity, ABHD10 would function as a detoxification enzyme. The roles of orphan esterases are becoming increasingly understood. Further studies will facilitate our knowledge of the pharmacologic and toxicological significance of orphan esterases in drug therapy. PMID:26521872

  13. Novel organic solvent-tolerant esterase isolated by metagenomics: insights into the lipase/esterase classification.

    PubMed

    Berlemont, Renaud; Spee, Olivier; Delsaute, Maud; Lara, Yannick; Schuldes, Jörg; Simon, Carola; Power, Pablo; Daniel, Rolf; Galleni, Moreno

    2013-01-01

    in order to isolate novel organic solvent-tolerant (OST) lipases, a metagenomic library was built using DNA derived from a temperate forest soil sample. A two-step activity-based screening allowed the isolation of a lipolytic clone active in the presence of organic solvents. Sequencing of the plasmid pRBest recovered from the positive clone revealed the presence of a putative lipase/esterase encoding gene. The deduced amino acid sequence (RBest1) contains the conserved lipolytic enzyme signature and is related to the previously described OST lipase from Lysinibacillus sphaericus 205y, which is the sole studied prokaryotic enzyme belonging to the 4.4 α/β hydrolase subgroup (abH04.04). Both in vivo and in vitro studies of the substrate specificity of RBest1, using triacylglycerols or nitrophenyl-esters, respectively, revealed that the enzyme is highly specific for butyrate (C4) compounds, behaving as an esterase rather than a lipase. The RBest1 esterase was purified and biochemically characterized. The optimal esterase activity was observed at pH 6.5 and at temperatures ranging from 38 to 45 °C. Enzymatic activity, determined by hydrolysis of p-nitrophenyl esters, was found to be affected by the presence of different miscible and non-miscible organic solvents, and salts. Noteworthy, RBest1 remains significantly active at high ionic strength. These findings suggest that RBest1 possesses the ability of OST enzymes to molecular adaptation in the presence of organic compounds and resistance of halophilic proteins.

  14. Mechanism of action of Neisseria gonorrhoeae O-acetylpeptidoglycan esterase, an SGNH serine esterase.

    PubMed

    Pfeffer, John M; Weadge, Joel T; Clarke, Anthony J

    2013-01-25

    O-Acetylpeptidoglycan esterase from Neisseria gonorrhoeae functions to release O-acetyl groups from the C-6 position of muramoyl residues in O-acetylated peptidoglycan, thereby permitting the continued metabolism of this essential cell wall heteropolymer. It has been demonstrated to be a serine esterase with sequence similarity to the family CE-3 carbohydrate esterases of the CAZy classification system. In the absence of a three-dimensional structure for any Ape, further knowledge of its structure and function relationship is dependent on modeling and kinetic studies. In this study, we predicted Neisseria gonorrhoeae Ape1a to be an SGNH hydrolase with an adopted α/β-hydrolase fold containing a central twisted four-stranded parallel β-sheet flanked by six α-helices with the putative catalytic triad, Asp-366, His-369, and Ser-80 appropriately aligned within a pocket. The role of eight invariant and highly conserved residues localized to the active site was investigated by site-directed replacements coupled with kinetic characterization and binding studies of the resultant engineered enzymes. Based on these data and theoretical considerations, Gly-236 and Asn-268 were identified as participating at the oxyanion hole to stabilize the tetrahedral species in the reaction mechanism, whereas Gly-78, Asp-79, His-81, Asn-235, Thr-267, and Val-368 are proposed to position appropriately the catalytic residues and participate in substrate binding. PMID:23209280

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

    PubMed Central

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

    2011-01-01

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

  16. Purification and characterization of an esterase involved in poly(vinyl alcohol) degradation by Pseudomonas vesicularis PD.

    PubMed

    Sakai, K; Fukuba, M; Hasui, Y; Moriyoshi, K; Ohmoto, T; Fujita, T; Ohe, T

    1998-10-01

    An esterase catalyzing the hydrolysis of acetyl ester moieties in poly(vinyl alcohol) was purified 400-fold to electrophoretic homogeneity from the cytoplasmic fraction of Pseudomonas vesicularis PD, which was capable of assimilating poly(vinyl alcohol) as the sole carbon and energy source. The purified enzyme was a homodimeric protein with a molecular mass of 80 kDa and the isoelectric point was 6.8. The pH and temperature optima of the enzyme were 8.0 and 45 degrees C. The enzyme catalyzed the hydrolysis of side chains of poly(vinyl alcohol), short-chain p-nitrophenyl esters, 2-naphthyl acetate, and phenyl acetate, and was slightly active toward aliphatic esters. The enzyme was also active toward the enzymatic degradation products, acetoxy hydroxy fatty acids, of poly(vinyl alcohol). The K(m) and Vmax of poly(vinyl alcohol) (degree of polymerization, 500; saponification degree, 86.5-89.0 mol%) and p-nitrophenyl acetate were 0.381% (10.6 mM as acetyl content in the polymer) and 2.56 microM, and 6.52 and 12.6 mumol/min/mg, respectively. The enzyme was strongly inhibited by phenylmethylsulfonyl fluoride and diisopropyl fluorophosphate at a concentration of 5 mM, which indicated that the enzyme was a serine esterase. The pathway for the metabolism of poly(vinyl alcohol) is also discussed.

  17. Recruit an ANA for RCS tests

    NASA Astrophysics Data System (ADS)

    Boyles, J.

    1985-03-01

    For the conduction of valid radar cross-section (RCS) measurements, it is necessary to remove unwanted signals from the test data. However, the absolute accuracy of the measurements is limited because even the most carefully designed anechoic chamber allows some residual energy to reflect from the walls, floor, and ceiling. The present investigation is concerned with an automatic network analyzer (ANA) which makes it possible to eliminate the unwanted signals from the final results on the basis of a careful calibration. The ANA system consists of a network analyzer, a swept signal source, and a test set. Attention is given to hardware choices for RCS measurements, an extension of the dynamic range, an error model, error-correction procedures for RCS measurements, and the conduction of the measurements.

  18. Enzymes involved in vinyl acetate decomposition by Pseudomonas fluorescens PCM 2123 strain.

    PubMed

    Szczyrba, Elżbieta; Greń, Izabela; Bartelmus, Grażyna

    2014-03-01

    Esterases are widely used in food processing industry, but there is little information concerning enzymes involved in decompositions of esters contributing to pollution of environment. Vinyl acetate (an ester of vinyl alcohol and acetic acid) is a representative of volatile organic compounds (VOCs) in decomposition, of which hydrolyses and oxidoreductases are mainly involved. Their activities under periodically changing conditions of environment are essential for the removal of dangerous VOCs. Esterase and alcohol/aldehyde dehydrogenase activities were determined in crude cell extract from Pseudomonas fluorescens PMC 2123 after vinyl acetate induction. All examined enzymes exhibit their highest activity at 30-35 °C and pH 7.0-7.5. Esterase preferably hydrolyzed ester bonds with short fatty chains without plain differences for C2 or C4. Comparison of Km values for alcohol and aldehyde dehydrogenases for acetaldehyde suggested that this metabolite was preferentially oxidized than reduced. Activity of alcohol dehydrogenase reducing acetaldehyde to ethanol suggested that one mechanism of defense against the elevated concentration of toxic acetaldehyde could be its temporary reduction to ethanol. Esterase activity was inhibited by phenylmethanesulfonyl fluoride, while β-mercaptoethanol, dithiothreitol, and ethylenediaminetetraacetic acid had no inhibitor effect. From among metal ions, only Mg(2+) and Fe(2+) stimulated the cleavage of ester bond.

  19. Functional Analysis of Esterase TCE2 Gene from Tetranychus cinnabarinus (Boisduval) involved in Acaricide Resistance

    PubMed Central

    Shi, Li; Wei, Peng; Wang, Xiangzun; Shen, Guangmao; Zhang, Jiao; Xiao, Wei; Xu, Zhifeng; Xu, Qiang; He, Lin

    2016-01-01

    The carmine spider mite, Tetranychus cinnabarinus is an important pest of crops and vegetables worldwide, and it has the ability to develop resistance against acaricides rapidly. Our previous study identified an esterase gene (designated TCE2) over-expressed in resistant mites. To investigate this gene’s function in resistance, the expression levels of TCE2 in susceptible, abamectin-, fenpropathrin-, and cyflumetofen-resistant strains were knocked down (65.02%, 63.14%, 57.82%, and 63.99%, respectively) via RNA interference. The bioassay data showed that the resistant levels to three acaricides were significantly decreased after the down-regulation of TCE2, indicating a correlation between the expression of TCE2 and the acaricide-resistance in T. cinnabarinus. TCE2 gene was then re-engineered for heterologous expression in Escherichia coli. The recombinant TCE2 exhibited α-naphthyl acetate activity (483.3 ± 71.8 nmol/mg pro. min−1), and the activity of this enzyme could be inhibited by abamectin, fenpropathrin, and cyflumetofen, respectively. HPLC and GC results showed that 10 μg of the recombinant TCE2 could effectively decompose 21.23% fenpropathrin and 49.70% cyflumetofen within 2 hours. This is the first report of a successful heterologous expression of an esterase gene from mites. This study provides direct evidence that TCE2 is a functional gene involved in acaricide resistance in T. cinnabarinus. PMID:26725309

  20. Characterization of a Feruloyl Esterase from Lactobacillus plantarum

    PubMed Central

    Esteban-Torres, María; Reverón, Inés; Mancheño, José Miguel; de las Rivas, Blanca

    2013-01-01

    Lactobacillus plantarum is frequently found in the fermentation of plant-derived food products, where hydroxycinnamoyl esters are abundant. L. plantarum WCFS1 cultures were unable to hydrolyze hydroxycinnamoyl esters; however, cell extracts from the strain partially hydrolyze methyl ferulate and methyl p-coumarate. In order to discover whether the protein Lp_0796 is the enzyme responsible for this hydrolytic activity, it was recombinantly overproduced and enzymatically characterized. Lp_0796 is an esterase that, among other substrates, is able to efficiently hydrolyze the four model substrates for feruloyl esterases (methyl ferulate, methyl caffeate, methyl p-coumarate, and methyl sinapinate). A screening test for the detection of the gene encoding feruloyl esterase Lp_0796 revealed that it is generally present among L. plantarum strains. The present study constitutes the description of feruloyl esterase activity in L. plantarum and provides new insights into the metabolism of hydroxycinnamic compounds in this bacterial species. PMID:23793626

  1. Endophytic fungi producing of esterases: Evaluation in vitro of the enzymatic activity using pH indicator

    PubMed Central

    Lisboa, Helen Cristina Fávero; Biasetto, Carolina Rabal; de Medeiros, João Batista; Araújo, Ângela 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. PMID:24516461

  2. International consensus on ANA patterns (ICAP): the bumpy road towards a consensus on reporting ANA results.

    PubMed

    Damoiseaux, Jan; von Mühlen, Carlos A; Garcia-De La Torre, Ignacio; Carballo, Orlando Gabriel; de Melo Cruvinel, Wilson; Francescantonio, Paulo Luiz Carvalho; Fritzler, Marvin J; Herold, Manfred; Mimori, Tsuneyo; Satoh, Minoru; Andrade, Luis E C; Chan, Edward K L; Conrad, Karsten

    2016-12-01

    The International Consensus on ANA Patterns (ICAP) was initiated as a workshop aiming to thoroughly discuss and achieve consensus regarding the morphological patterns observed in the indirect immunofluorescence assay on HEp-2 cells. One of the topics discussed at the second ICAP workshop, and addressed in this paper, was the harmonization of reporting ANA test results. This discussion centered on the issue if cytoplasmic and mitotic patterns should be reported as positive or negative. This report outlines the issues that impact on two major different reporting methods. Although it was appreciated by all participants that cytoplasmic and mitotic patterns are clinically relevant, implications for existing diagnostic/classification criteria for ANA-associated diseases in particular hampered a final consensus on this topic. Evidently, a more concerted action of all relevant stakeholders is required. Future ICAP workshops may help to facilitate this action.

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

    PubMed

    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.

  4. Structural studies of a thermophilic esterase from a new Planctomycetes species, Thermogutta terrifontis.

    PubMed

    Sayer, Christopher; Isupov, Michail N; Bonch-Osmolovskaya, Elizaveta; Littlechild, Jennifer A

    2015-08-01

    Thermogutta terrifontis esterase (TtEst), a carboxyl esterase identified in the novel thermophilic bacterium T. terrifontis from the phylum Planctomycetes, has been cloned and over-expressed in Escherichia coli. The enzyme has been characterized biochemically and shown to have activity towards small p-nitrophenyl (pNP) carboxylic esters, with optimal activity for pNP-propionate. The enzyme retained 95% activity after incubation for 1 h at 80 °C. The crystal structures of the native TtEst and its complexes with the substrate analogue D-malate and the product acetate have been determined to high resolution. The bound ligands have allowed the identification of the carboxyl and alcohol binding pockets in the enzyme active site. Comparison of TtEst with structurally related enzymes provides insight into how differences in their catalytic activity can be rationalized based upon the properties of the amino acid residues in their active site pockets. The mutant enzymes L37A and L251A have been constructed to extend the substrate range of TtEst towards the larger butyrate and valerate pNP-esters. These mutant enzymes have also shown a significant increase in activity towards acetate and propionate pNP esters. A crystal structure of the L37A mutant was determined with the butyrate product bound in the carboxyl pocket of the active site. The mutant structure shows an expansion of the pocket that binds the substrate carboxyl group, which is consistent with the observed increase in activity towards pNP-butyrate. PMID:26011036

  5. Characterization of Esterases Produced by a Ruminal Bacterium Identified as Butyrivibrio fibrisolvens1

    PubMed Central

    Lanz, Wayne W.; Williams, Phletus P.

    1973-01-01

    An obligately anaerobic ruminal bacterial isolate was selected from 18 tributyrin-degrading isolates and identified as Butyrivibrio fibrisolvens strain 53. The culture in late exponential phase contained enzymes which could be released by sonic disruption. These enzymes degraded substrates at a rate in the order 1-naphthyl acetate (NA) > 1-naphthyl butyrate > 1-naphthyl propionate but did not degrade 1-naphthyl palmitate or 1-naphthyl phosphate. The enzymes on NA were neither stimulated nor inhibited by CoCl2, MgCl2, and MnCl (each varied from 10−6 to 10−4 M). CaCl at 10−3 M stimulated esterase activity by 16%. Aliphatic substrates were hydrolyzed at a rate in the order triacetin > tributyrin > tripropionin, and ethyl acetate > ethyl formate. Similarly, aromatic fluorescein diesters were degraded at a rate in the order acetyl > propionyl > caproyl > butyryl > capryl > lauryl. Polyacrylamide gel electrophoretic zymograms indicated that the enzyme composite contained cathodally migrating bands. By column chromatography, these enzymes were separated into six NA-degrading fractions. Fraction V contained an esterase which had an optimal temperature of 39 C, a Km of 7.6 × 10−4 on NA, and a molecular weight of about 66,000. This enzyme was inhibited by paraoxon (41%, 10−4 M), eserine (17%, 10−2 M), NaF (17%, 10−2 M), and diisopropyl fluorophosphate (62%, 10−4 M) but not by 1-naphthyl N-methyl carbamate at 8.4 × 10−4 M. PMID:4734862

  6. Tissue distribution, characterization and in vitro inhibition of B-esterases in the earwig Forficula auricularia.

    PubMed

    Malagnoux, Laure; Capowiez, Yvan; Rault, Magali

    2014-10-01

    Earwigs are important natural enemies of numerous pests in pome fruit orchards worldwide. Studying the effects of agricultural practices on these biological control agents is important for understanding its vulnerability in the field. The aim of this study was to characterize the B-esterase activities in the European earwig Forficula auricularia and to evaluate in vitro its sensitivity to organophosphate and carbamate pesticides. Acetylcholinesterase (AChE) activity was mainly measured with 1.5 mM acetylthiocholine as the substrate in the microsomal fraction of earwig heads (70% of total AChE activity). Carboxylesterase (CbE) activities were measured with three substrates [5 mM 4-nitrophenyl acetate (4-NPA), 1mM 4-nitrophenyl valerate (4-NPV), and 2 mM α-naphtyl acetate (α-NA)] to examine different isoenzymes, which were present mainly in the cytosolic fraction (about 70-88% of total activities) of all earwig tissues. CbE activity was higher than AChE activity, especially with α-NA, then 4-NPA and lastly 4-NPV. Chlorpyrifos-oxon an organophosphate, and carbaryl a carbamate pesticide, inhibited AChE and CbE activities in a concentration-dependent manner. Earwig CbE activities showed a stronger sensitivity to organophosphate than AChE, with the strongest effect for chlorpyrifos-oxon on male carboxylesterase activities. CbE and AChE showed about the same sensitivity to carbamate pesticides regardless of sex. These results suggest that B-type esterases in the European earwig F.auricularia are suitable biomarkers of pesticide exposure.

  7. Characterization of a novel highly thermostable esterase from the Gram-positive soil bacterium Streptomyces lividans TK64.

    PubMed

    Wang, Baojuan; Wang, Ao; Cao, Zhengyu; Zhu, Guoping

    2016-05-01

    A novel esterase gene (estW) from soil bacterium Streptomyces lividans TK64 was successfully cloned using a pair of homologous primers. The estW gene encoded a protein (EstW) of 289 amino acid residues with a predicted molecular weight of 31.43 kDa. Sequence alignment revealed that EstW show relatively high levels of homology to other lipolytic enzymes characterized from Streptomyces and phylogenetic analysis suggested EstW belongs to the bacterial lipase/esterase family I. The estW gene was expressed at a high level in Escherichia coli and the recombinant enzyme was purified to homogeneity. The purified EstW was characterized via hydrolysis of various p-nitrophenyl esters and the best substrate was found to be p-nitrophenyl acetate (pNPA). Maximal activity of the recombinant protein was observed at pH 8.0 and 50 °C with pNPA as the substrate. The calculated activation energy (Ea ) of the esterase reaction was 9.12 kcal/mol. Half-life of EstW at 95 °C was approximately 12.5 H, making it the most thermostable esterase among all of the known lipolytic enzymes from Streptomyces, and the thermostability of EstW was similar to those of some enzymes characterized from the thermophilic bacteria. EstW exhibited relatively high tolerance to several detergents and required no cations for its maximal activity. The unique properties of EstW, namely its high thermostability and stability in the presence of organic solvents, may render it a potential candidate for industrial applications.

  8. Preliminary X-ray analysis of twinned crystals of the Q88Y25_Lacpl esterase from Lactobacillus plantarum WCFS1

    PubMed Central

    Álvarez, Yanaisis; Esteban-Torres, María; Acebrón, Iván; de las Rivas, Blanca; Muñoz, Rosario; Martínez-Ripoll, Martín; Mancheño, José M.

    2011-01-01

    Q88Y25_Lacpl is an esterase produced by the lactic acid bacterium Lactobacillus plantarum WCFS1 that shows amino-acid sequence similarity to carboxyl­esterases from the hormone-sensitive lipase family, in particular the AFEST esterase from the archaeon Archaeoglobus fulgidus and the hyperthermophilic esterase EstEI isolated from a metagenomic library. N-­terminally His6-tagged Q88Y25_Lacpl has been overexpressed in Escherichia coli BL21 (DE3) cells, purified and crystallized at 291 K using the hanging-drop vapour-diffusion method. Mass spectrometry was used to determine the purity and homogeneity of the enzyme. Crystals of His6-tagged Q88Y25_Lacpl were prepared in a solution containing 2.8 M sodium acetate trihydrate pH 7.0. X-ray diffraction data were collected to 2.24 Å resolution on beamline ID29 at the ESRF. The apparent crystal point group was 422; however, initial global analysis of the intensity statistics (data processed with high symmetry in space group I422) and subsequent tests on data processed with low symmetry (space group I4) showed that the crystals were almost perfectly merohedrally twinned. Most probably, the true space group is I4, with unit-cell parameters a = 169.05, b = 169.05, c = 183.62 Å. PMID:22102251

  9. Communicating stigma: the pro-ana paradox.

    PubMed

    Yeshua-Katz, Daphna; Martins, Nicole

    2013-01-01

    This study explores the personal experience of pro-ana bloggers, members of an online community for people with eating disorders. Using Erving Goffman's work on stigma, this study explores the motivations, benefits, and drawbacks of blogging about a stigmatized mental illness, as taken from the bloggers' own perceptive. We conducted 33 interviews with bloggers from seven different countries via phone, Skype, and e-mail. Participants were motivated to blog because they found social support, a way to cope with a stigmatized illness, and means of self-expression. Participants described blogging as a cathartic experience and perceived the social support they received from other members of the pro-ana community as a benefit. The fear that the eating disorder will be revealed if the blog is exposed and the concern that the blog encourages disordered eating were the perceived negative consequences of maintaining such a blog. Thus, blogging about anorexia serves to both alleviate and trigger anxiety about living with this stigmatized illness. Recommendations for future research are made.

  10. 29. ELECTRICAL EQUIPMENT. PLAN FOR POWER HOUSE, SANTA ANA RIVER ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    29. ELECTRICAL EQUIPMENT. PLAN FOR POWER HOUSE, SANTA ANA RIVER P. H. NO. 3, JUNE 23, 1943; REVISIONS, MAR. 14, 1945 AND MAY 17, 1954. SCE drawing no. 523219-2. - Santa Ana River Hydroelectric System, SAR-3 Powerhouse, San Bernardino National Forest, Redlands, San Bernardino County, CA

  11. Free inside: The Music Class at Santa Ana Jail

    ERIC Educational Resources Information Center

    Fierro, Joe

    2010-01-01

    This article examines the workings of the music class at the Santa Ana Jail in Santa Ana, California. It gives us insight into a jail system and a music class focused on helping inmates position themselves to become productive members of society. In this article I examine how the facility encourages inmates' good behaviour and why the music class…

  12. 48. MAP OF SANTA ANA RIVER POWER PLANT NO. 2 ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    48. MAP OF SANTA ANA RIVER POWER PLANT NO. 2 OF THE EDISON ELECTRIC CO. THROUGH UNSURVEYED LAND IN THE SAN BERNARDINO FOREST RESERVE, APPROVED MAY 26, 1904, F. C. FINKLE, CHIEF HYDRAULIC ENGINEER. SCE drawing no. 53988. - Santa Ana River Hydroelectric System, Redlands, San Bernardino County, CA

  13. Comparative toxicity of lead shot in black ducks (Anas rubripes) and mallards (Anas platyrhynchos).

    PubMed

    Rattner, B A; Fleming, W J; Bunck, C M

    1989-04-01

    In winter, pen-reared and wild black ducks (Anas rubripes), and game farm and wild mallards (Anas platyrhynchos), maintained on pelleted feed, were sham-dosed or given one number 4 lead shot. After 14 days, dosed birds were redosed with two or four additional lead shot. This dosing regimen also was repeated in summer using pen-reared black ducks and game farm mallards. Based upon mortality, overt intoxication, weight change, delta-aminolevulinic acid dehydratase activity and protoporphyrin concentration, black ducks and mallards were found to be equally tolerant to lead shot. However, captive wild ducks were more sensitive than their domesticated counterparts, as evidenced by greater mortality and weight loss following lead shot administration. This difference may be related to stress associated with captivity and unnatural diet. PMID:2716097

  14. Comparative toxicity of lead shot in black ducks (Anas rubripes) and mallards (Anas platyrhynchos)

    USGS Publications Warehouse

    Rattner, B.A.; Fleming, W.J.; Bunck, C.M.

    1989-01-01

    In winter, pen-reared and wild black ducks (Anas rubripes), and game farm and wild mallards (Anas platyrhynchos), maintained on pelleted feed, were sham-dosed or given one number 4 lead shot. After 14 days, dosed birds were redosed with two or four additional lead shot. This dosing regimen also was repeated in summer using pen-reared black ducks and game farm mallards. Based upon mortality, overt intoxication, weight change, delta-aminolevulinic acid dehydratase activity and protoporphyrin concentration, black ducks and mallards were found to be equally tolerant to lead shot. However, captive wild ducks were more sensitive than their domesticated counterparts, as evidenced by greater mortality and weight loss following lead shot administration. This difference may be related to stress associated with captivity and unnatural diet.

  15. Bacteriophage 933W encodes a functional esterase downstream of the Shiga toxin 2a operon.

    PubMed

    Nübling, Simone; Eisele, Thomas; Stöber, Helen; Funk, Joschua; Polzin, Sabrina; Fischer, Lutz; Schmidt, Herbert

    2014-05-01

    In this study, the 1938bp open reading frame z1466, which is encoded directly downstream the Shiga toxin 2a (Stx2a) operon in E. coli O157:H7 phage 933W was cloned and expressed recombinantly. Purification with Ni-NTA agarose beads with subsequent SDS-PAGE revealed a 68kDa protein, designated 933Wp42-His. Analysis of 933Wp42-His demonstrated an esterase activity by activity staining of native gels using triacetin as a substrate. Purified 933Wp42-His demonstrated a Km value of about 10mM and a Vmax value of 1.667nkat/ml for 4-methylumbelliferyl-acetate (4-MUF-Ac) as a substrate. The enzyme was most active in the pH-range of 7.0-8.0, and at 50°C. Furthermore, 933Wp42-His was able to hydrolyze acetic acid from mucin, and 5-N-acetyl-9-O-acetyl neuraminic acid (Neu5,9Ac2). This is the first description of an enzymatic activity of the Stx-phage-encoded protein 933Wp42. Its role in substrate utilization during colonization and human infection is discussed.

  16. Exploring the aryl esterase catalysis of paraoxonase-1 through solvent kinetic isotope effects and phosphonate-based isosteric analogues of the tetrahedral reaction intermediate.

    PubMed

    Bavec, Aljoša; Knez, Damijan; Makovec, Tomaž; Stojan, Jure; Gobec, Stanislav; Goličnik, Marko

    2014-11-01

    Although a recent study of Debord et al. in Biochimie (2014; 97:72-77) described the thermodynamics of the catalysed hydrolysis of phenyl acetate by human paraoxonase-1, the mechanistic details along the reaction route of this enzyme remain unclear. Therefore, we briefly present the solvent kinetic isotope effects on the phenyl acetate esterase activity of paraoxonase-1 and its inhibition with the phenyl methylphosphonate anion, which is a stable isosteric analogue that mimics the high-energy tetrahedral intermediate on the hydroxide-promoted hydrolysis pathway. The data show normal isotope effects, while proton inventory analysis indicates that two protons contribute to the kinetic isotope effect. Coherently, moderate competitive inhibition with the phenyl methylphosphonate anion reveals that the rate-limiting transition state suboptimally resembles the tetrahedral intermediate. The implications of these findings can be attributed to two possible reaction mechanisms that might occur during the paraoxonase-1-catalysed hydrolysis of phenyl acetate.

  17. Esterase detoxication of acetylcholinesterase inhibitors using human liver samples in vitro.

    PubMed

    Moser, Virginia C; Padilla, Stephanie

    2016-04-15

    Organophosphorus (OP) and N-methylcarbamate pesticides inhibit acetylcholinesterase (AChE), but differences in metabolism and detoxication can influence potency of these pesticides across and within species. Carboxylesterase (CaE) and A-esterase (paraoxonase, PON1) are considered factors underlying age-related sensitivity differences. We used an in vitro system to measure detoxication of AChE-inhibiting pesticides mediated via these esterases. Recombinant human AChE was used as a bioassay of inhibitor concentration following incubation with detoxifying tissue: liver plus Ca(+2) (to stimulate PON1s, measuring activity of both esterases) or EGTA (to inhibit PON1s, thereby measuring CaE activity). AChE inhibitory concentrations of aldicarb, chlorpyrifos oxon, malaoxon, methamidophos, oxamyl, paraoxon, and methylparaoxon were incubated with liver homogenates from adult male rat or one of 20 commercially provided human (11-83 years of age) liver samples. Detoxication was defined as the difference in inhibition produced by the pesticide alone and inhibition measured in combination with liver plus Ca(+2) or liver plus EGTA. Generally, rat liver produced more detoxication than did the human samples. There were large detoxication differences across human samples for some pesticides (especially malaoxon, chlorpyrifos oxon) but not for others (e.g., aldicarb, methamidophos); for the most part these differences did not correlate with age or sex. Chlorpyrifos oxon was fully detoxified only in the presence of Ca(+2) in both rat and human livers. Detoxication of paraoxon and methylparaoxon in rat liver was greater with Ca(+2), but humans showed less differentiation than rats between Ca(+2) and EGTA conditions. This suggests the importance of PON1 detoxication for these three OPs in the rat, but mostly only for chlorpyrifos oxon in human samples. Malaoxon was detoxified similarly with Ca(+2) or EGTA, and the differences across humans correlated with metabolism of p

  18. Regiospecific Ester Hydrolysis by Orange Peel Esterase - An Undergraduate Experiment.

    NASA Astrophysics Data System (ADS)

    Bugg, Timothy D. H.; Lewin, Andrew M.; Catlin, Eric R.

    1997-01-01

    A simple but effective experiment has been developed to demonstrate the regiospecificity of enzyme catalysis using an esterase activity easily isolated from orange peel. The experiment involves the preparation of diester derivatives of para-, meta- and ortho-hydroxybenzoic acid (e.g. methyl 4-acetoxy-benzoic acid). The derivatives are incubated with orange peel esterase, as a crude extract, and with commercially available pig liver esterase and porcine pancreatic lipase. The enzymatic hydrolysis reactions are monitored by thin layer chromatography, revealing which of the two ester groups is hydrolysed, and the rate of the enzyme-catalysed reaction. The results of a group experiment revealed that in all cases hydrolysis was observed with at least one enzyme, and in most cases the enzymatic hydrolysis was specific for production of either the hydroxy-ester or acyl-acid product. Specificity towards the ortho-substituted series was markedly different to that of the para-substituted series, which could be rationalised in the case of pig liver esterase by a published active site model.

  19. Glucuronoyl esterases are active on polymeric substrate, methyl esterified glucuronoxylan

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Alkali extracted beechwood glucuronoxylan methyl ester prepared by esterification of 4-O-methyl-D-glucuronic acid side residues by methanol was found to serve as substrate of microbial glucuronoyl esterases from Ruminococcus flavefaciens, Schizophyllum commune and Trichoderma reesei. The enzymatic d...

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

    SciTech Connect

    Novak, R.; Padilla, S.

    1986-04-01

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

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

    PubMed

    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.

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

    PubMed

    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. PMID:22426713

  3. Improving enantioselectivity towards tertiary alcohols using mutants of Bacillus sp. BP-7 esterase EstBP7 holding a rare GGG(X)-oxyanion hole.

    PubMed

    Fillat, Amanda; Romea, Pedro; Urpí, Fèlix; Pastor, F I Javier; Diaz, Pilar

    2014-05-01

    Lipases and esterases are important biocatalysts for synthetic organic fine chemistry. An esterase from Bacillus sp. BP-7 (EstBP7) bears in its amino acid sequence a rare GGG(A)X oxyanion hole motif, where an uncommon threonine (T) is found at the third position. Detection of this pattern motivated evaluation of the ability of EstBP7 for conversion of tertiary alcohols. The enzyme was engineered in order to optimize its performance to provide important chiral building blocks: five variants with mutations in the oxyanion hole motif were created to investigate the influence on activity and enantioselectivity in the kinetic resolution of eight acetates of tertiary alcohols. Wild-type enzyme converted all esters of tertiary alcohols assayed with low enantioselectivity, whereas some of the mutants displayed significantly increased E-values. One of the mutants (EstBP7-AGA; Mut 5) showed an E >100 towards a complex tertiary alcohol acetate (2-(4-pyridyl)but-3-yn-2-yl acetate) at low reaction temperature (4 °C). Therefore, the catalytic toolbox was expanded for biocatalysis of optically pure tertiary alcohols valuable for the pharmaceutical industry. PMID:24407449

  4. Naval Air Station, Santa Ana, Calif. Lighterthanairhangar roof truss details. ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Naval Air Station, Santa Ana, Calif. Lighter-than-air-hangar roof truss details. Drawing no. 212817. - Marine Corps Air Station Tustin, East of Red Hill Avenue between Edinger Avenue & Barranca Parkway, Tustin, Orange County, CA

  5. Naval Air Station, Santa Ana, Calif. Lighterthanairhangar roof truss details. ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Naval Air Station, Santa Ana, Calif. Lighter-than-air-hangar roof truss details. Drawing no. 212817. - Marine Corps Air Station Tustin, Northern Lighter Than Air Ship Hangar, Meffett Avenue & Maxfield Street, Tustin, Orange County, CA

  6. The solution structure of double helical arabino nucleic acids (ANA and 2'F-ANA): effect of arabinoses in duplex-hairpin interconversion.

    PubMed

    Martín-Pintado, Nerea; Yahyaee-Anzahaee, Maryam; Campos-Olivas, Ramón; Noronha, Anne M; Wilds, Christopher J; Damha, Masad J; González, Carlos

    2012-10-01

    We report here the first structure of double helical arabino nucleic acid (ANA), the C2'-stereoisomer of RNA, and the 2'-fluoro-ANA analogue (2'F-ANA). A chimeric dodecamer based on the Dickerson sequence, containing a contiguous central segment of arabino nucleotides, flanked by two 2'-deoxy-2'F-ANA wings was studied. Our data show that this chimeric oligonucleotide can adopt two different structures of comparable thermal stabilities. One structure is a monomeric hairpin in which the stem is formed by base paired 2'F-ANA nucleotides and the loop by unpaired ANA nucleotides. The second structure is a bimolecular duplex, with all the nucleotides (2'F-ANA and ANA) forming Watson-Crick base pairs. The duplex structure is canonical B-form, with all arabinoses adopting a pure C2'-endo conformation. In the ANA:ANA segment, steric interactions involving the 2'-OH substituent provoke slight changes in the glycosidic angles and, therefore, in the ANA:ANA base pair geometry. These distortions are not present in the 2'F-ANA:2'F-ANA regions of the duplex, where the -OH substituent is replaced by a smaller fluorine atom. 2'F-ANA nucleotides adopt the C2'-endo sugar pucker and fit very well into the geometry of B-form duplex, allowing for favourable 2'F···H8 interactions. This interaction shares many features of pseudo-hydrogen bonds previously observed in 2'F-ANA:RNA hybrids and in single 2'F-ANA nucleotides.

  7. Electrophoretic and densitometric analysis of esterase activity as an indicator of mercury toxicity

    SciTech Connect

    Benton, M.J.; Guttman, S.I.

    1995-12-31

    In an earlier experiment, esterase activity as determined by starch gel electrophoresis was absent in larval caddisflies (Nectopsyche albida) that succumbed to mercury exposure, but was present in control larvae. To test the effects of mercury exposure duration on esterase activity, additional larval N. albida were exposed under conditions identical to those in the earlier experiment, and esterase activity was determined by electrophoresis of several live individuals every 12 hours. To test the effects of mercury concentration on esterase activity, homogenates of unexposed N. albida were electrophoresed, and esterase activity was determined using esterase-specific stains spiked with various concentrations of mercury. Following both experiments, esterase activity was quantified by laser densitometry of stained electrophoresis gels, Results indicate that: (1) inorganic mercury inhibited esterase activity, (2) inhibition increased with exposure time, and (3) inhibition increased with mercury concentration. Esterase inhibition may be a causal factor in mortality related to mercury exposure. Quantification of esterase activity by densitometry of electrophoretic gels may be an alternative method of rapid toxicity assessment.

  8. Identification of a bacterial pectin acetyl esterase in Erwinia chrysanthemi 3937.

    PubMed

    Shevchik, V E; Hugouvieux-Cotte-Pattat, N

    1997-06-01

    Erwinia chrysanthemi causes soft-rot diseases of various plants by enzymatic degradation of the pectin in plant cell walls. The structural complexity of pectin requires the combined action of several pectinases for its efficient breakdown. Three types of pectinases have so far been identified in E. chrysanthemi: two pectin methyl esterases (PemA, PemB), a polygalacturonase (PehX), and eight pectate lyases (PelA, PelB, PelC, PelD, PelE, PelL, PelZ, PelX). We report in this paper the analysis of a novel enzyme, the pectin acetyl esterase encoded by the paeY gene. No bacterial form of pectin acetyl esterases has been described previously, while plant tissues and some pectinolytic fungi were found to produce similar enzymes. The paeY gene is present in a cluster of five pectinase-encoding genes, pelA-pelE-pelD-paeY-pemA. The paeY open reading frame is 1650 bases long and encodes a 551-residue precursor protein of 60704Da, including a 25-amino-acid signal peptide. PaeY shares one region of homology with a rhamnogalacturonan acetyl esterase of Aspergillus aculeatus. To characterize the enzyme, the paeY gene was overexpressed and its protein product was purified. PaeY releases acetate from sugar-beet pectin and from various synthetic substrates. Moreover, the enzyme was shown to act in synergy with other pectinases. The de-esterification rate by PaeY increased after previous demethylation of the pectins by PemA and after depolymerization of the pectin by pectate lyases. In addition, the degradation of sugar-beet pectin by pectate lyases is favoured after the removal of methyl and acetyl groups by PemA and PaeY, respectively. The paeY gene was first identified on the basis of its regulation, which shares several characteristics with that of other pectinases. Analysis of the paeY transcription, using gene fusions, revealed that it is induced by pectic catabolic products and is affected by growth phase, oxygen limitation and catabolite repression. Regulation of pae

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

    SciTech Connect

    Loughna, S.; Moore, G. ); Gau, G.; Blunt, S. ); Nicolaides, K. )

    1993-10-01

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

  10. The Serratia marcescens bioH gene encodes an esterase.

    PubMed

    Akatsuka, Hiroyuki; Kawai, Eri; Sakurai, Naoki; Omori, Kenji

    2003-01-01

    The 3.9 kb chromosomal DNA was cloned from Serratia marcescens Sr41, which confers on Escherichia coli cells a phenotype of clear halo formation on tributyrin agar plates. Three complete open reading frames (ORFs) were identified in the inserted DNA, and one ORF was demonstrated to encode a 28 kDa protein of 255 amino acids related to esterase activity. Interestingly, the ORF was 70% identical to a product of the E. coli bioH gene, which lies at a locus separated from the bioABFCD operon and acts in the early steps of the biotin synthetic pathway before pimeloyl-CoA synthesis. This gene complemented a bioH-deficient mutation of E. coli. From the sequence analysis, BioH is presumed to be a serine hydrolase, which belongs to the alpha/beta hydrolase-fold family comprising a wide variety of hydrolases including esterases. A catalytic triad composed of a nucleophilic residue (Ser80), an acidic residue (Asp206), and histidine (His234) was conserved in BioH, and the nucleophilic residue Ser, a catalytic center, was situated in the consensus sequence of G-X-S-X-G-G, a nucleophile elbow. Although the enzymatic function of BioH is not yet elucidated, the bioH gene products from S. marcescens and E. coli show esterase activity, which may imply the hydrolysis of a precursor leading to pimeloyl-CoA ester. The esterase activity of BioH and its CoA binding activity recently reported agree with a current hypothesis of pimeloyl-CoA ester synthesis from CoA and acylester derivatives including an acyl-carrier protein.

  11. The conversion of C'IS to C'1 esterase by plasmin and trypsin.

    PubMed

    Ratnoff, O D; Naff, G B

    1967-02-01

    The formation of C'1 esterase from C'1, the first component of complement, may be brought about by the action of plasmin or trypsin upon C'1s, a subcomponent of C'1. These enzymes also decrease the esterolytic activity of C'1 esterase. The formation of C'1 esterase was demonstrated by measuring the appearance of an agent or agents with esterolytic properties and the capacity to inactivate C'2 and C'4, attributes of C'1 esterase. The activity of the agent which evolved was blocked by serum inhibitor of C'1 esterase. The implications of these observations, that the formation of C'1 esterase during complement fixation is mediated by proteolytic processes, are under study. The possible inhibition of C'1q by soybean trypsin inhibitor is in agreement with this hypothesis.

  12. 3 Benzyl-6-chloropyrone: a suicide inhibitor of cholesterol esterase

    SciTech Connect

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

    1986-05-01

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

  13. Preparation and Properties of Novel Dentin Adhesives with Esterase Resistance

    PubMed Central

    Park, Jong-Gu; Ye, Qiang; Topp, Elizabeth M.; Kostoryz, Elisabet L.; Wang, Yong; Kieweg, Sarah L.; Spencer, Paulette

    2012-01-01

    A new methacrylate monomer, trimethylolpropane mono allyl ether dimethacrylate (TMPEDMA), was synthesized and evaluated. This branched methacrylate was designed to increase esterase-resistance when incorporated into conventional HEMA (2-hydroxyethyl methacrylate)/BisGMA (2,2-bis[4(2-hydroxy-3-methacryloyloxy-propyloxy)-phenyl] propane) dental adhesives. The new adhesives, HEMA/BisGMA/TMPEDMA in a 45/30/25 (w/w) ratio were formulated with H2O at 0 (A0T) and 8 wt % water (A8T) and compared with control adhesives (HEMA/BisGMA, 45/55 (w/w), at 0 (A0) and 8 wt % (A8) water). Camphoroquinone (CQ), 2-(dimethylamino) ethyl methacrylate and diphenyliodonium hexafluorophosphate were used as photoinitiators. The new adhesives showed a degree of conversion comparable with the control and improved modulus and glass transition temperature (Tg). Exposure of photopolymerized discs to porcine liver esterase for up to eight days showed that the net cumulative methacrylic acid (MAA) release in adhesives formulated with the new monomer and 8% water (A8T: 182 μg/mL) was dramatically (P < 0.05) decreased in comparison to the control (A8: 361.6 μg/mL). The results demonstrate that adhesives made with the new monomer and cured in water to simulate wet bonding are more resistant to esterase than conventional HEMA/BisGMA adhesive. PMID:22919119

  14. Tumor promoter 12-O-tetradecanoyl phorbol 13-acetate and regulatory diacylglycerols are substrates for the same carboxylesterase

    SciTech Connect

    Mentlein, R.

    1986-06-15

    Rat liver homogenate or cell fractions deacylate 12-O-tetradecanoyl phorbol 13-acetate (TPA) in vitro mainly by conversion to phorbol 13-acetate. The highest specific activity is located in the microsomal fraction. The deacylation is inhibited by bis-(4-nitrophenyl) phosphate, a selective inhibitor of nonspecific carboxylesterases. Only two of five purified esterases from rat liver endoplasmic reticulum deacylate TPA. These two esterases have formerly been characterized as acylcarnitine hydrolases and the more active one is also a potent diacylglycerol lipase. Its TPA-hydrolyzing activity is inhibited by other substrates like 1-naphthylacetate, lauroylcarnitine, or dioleoyl glycerol. The results support the view that phorbol esters act like structural analogs of diacylglycerols, not only with respect to their activating effect on protein kinase C, but also as substrates for the same lipases.

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

    PubMed Central

    Peña-Montes, Carolina; Mondragón-Tintor, María Elena; Castro-Rodríguez, José Augusto; Bustos-Jaimes, Ismael; Navarro-Ocaña, Arturo; Farrés, Amelia

    2013-01-01

    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. PMID:23781330

  16. Preparation of vinyl acetate

    DOEpatents

    Tustin, Gerald Charles; Zoeller, Joseph Robert; Depew, Leslie Sharon

    1998-01-01

    This invention pertains to the preparation of vinyl acetate by contacting a mixture of hydrogen and ketene with a heterogeneous catalyst containing a transition metal to produce acetaldehyde, which is then reacted with ketene in the presence of an acid catalyst to produce vinyl acetate.

  17. Preparation of vinyl acetate

    DOEpatents

    Tustin, G.C.; Zoeller, J.R.; Depew, L.S.

    1998-03-24

    This invention pertains to the preparation of vinyl acetate by contacting a mixture of hydrogen and ketene with a heterogeneous catalyst containing a transition metal to produce acetaldehyde, which is then reacted with ketene in the presence of an acid catalyst to produce vinyl acetate.

  18. Effect of UVB on hydrolysis of alpha-tocopherol acetate to alpha-tocopherol in mouse skin.

    PubMed

    Kramer-Stickland, K; Liebler, D C

    1998-08-01

    We have assessed the hydrolysis of alpha-tocopherol acetate (alpha-TAc) to the active antioxidant alpha-tocopherol (alpha-TH) in mouse epidermis and in supernatant from epidermal homogenates. Topically administered alpha-TH prevents UVB photocarcinogenesis in C3H mice, whereas alpha-TAc does not. Hydrolysis in skin was monitored in mice treated topically with deuterium labeled alpha-TAc (d3-alpha-TAc). Epidermal samples were isolated from mice and analyzed for endogenous (d0-alpha-TAc) and d3-alpha-TH by gas chromatography-mass spectrometry. Within 24 h, the levels of d3-alpha-TH increased up to 10-fold over endogenous d0-alpha-TH levels; however, in mice irradiated with UVB prior to the application of d3-alpha-TAc, levels of d3-alpha-TH increased up to 30-40-fold over endogenous d0-alpha-TH. This enhancement of alpha-TAc hydrolysis increased with increasing UVB dose. Prior UVB exposure may increase hydrolysis of alpha-TAc by increasing epidermal esterase activity. Nonspecific esterase activity was measured in the 2000 x g supernatant from epidermis of unirradiated and irradiated mice. Alpha-napthyl acetate, a nonspecific esterase substrate, was converted to alpha-napthol in supernatants from unirradiated mice. Hydrolysis to alpha-napthol increased approximately 3-fold in supernatants from irradiated mice. Hydrolysis of alpha-TAc to alpha-TH also occurred in supernatant from unirradiated mice, and this hydrolysis increased approximately 3-fold in supernatant from irradiated animals. These data indicate that nonspecific esterase activity was increased by UVB in the skin, that alpha-TAc is converted to alpha-TH in the homogenate fraction containing nonspecific esterase, and that UVB exposure modulates the metabolism of alpha-TAc to alpha-TH in vivo.

  19. Interaction between human serum esterases and environmental metal compounds.

    PubMed

    Hernández, Antonio F; Gil, Fernando; Leno, Esther; López, Olga; Rodrigo, Lourdes; Pla, Antonio

    2009-07-01

    Paraoxonase-1 (PON1) and cholinesterase (BChE) are two of the major human serum esterases. Although most of variation in PON1 activity results from genetic factors, there is growing evidence that environmental chemicals also modulate its activity. The aim of this study was to investigate whether environmental exposure to metal compounds has any influence on those esterases. A cross-sectional study was conducted in a representative sample of the general population of Andalusia, South of Spain. PON1 activity against different substrates (paraoxon, phenylacetate, diazoxon and dihydrocoumarin) and BChE were measured in serum from 536 healthy subjects. Potential associations of these esterases with metal compounds, age, sex and body mass index as well as life-style habits (smoking, alcohol drinking and food habits) were explored. Multiple linear regression analysis showed that blood lead levels were significantly associated with increased PON1 in serum regardless of the substrate used for the assay. Mercury also showed a significant and direct association with PON1 towards paraoxon and phenylacetate. In turn, cadmium and zinc levels were significantly associated with a decreased PON1 activity (zinc was associated with all PON1 activities and cadmium with PON1 towards paraoxon and diazoxon). Arsenic, nickel and manganese failed to be significantly associated with any of the PON1 activities assayed. PON1 192R alloform predicted significantly higher levels of arsenic and lead. BChE, however, was inversely associated with serum levels of manganese and zinc. These results suggest that PON1 and BChE activities are modulated by background exposure to metal compounds, which may have implications in public health given the defensive role played by both enzyme proteins against environmental toxicants. The potential underlying mechanisms merit further investigation.

  20. 31. CRANE RUNWAY FOR 5TON PUSH TYPE CRANE, SANTA ANA ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    31. CRANE RUNWAY FOR 5-TON PUSH TYPE CRANE, SANTA ANA RIVER NO. 3, SEPT. 4, 1945. SCE drawing no. 523856-2. - Santa Ana River Hydroelectric System, SAR-3 Powerhouse, San Bernardino National Forest, Redlands, San Bernardino County, CA

  1. Structure of the catalytic domain of glucuronoyl esterase Cip2 from Hypocrea jecorina

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The structure of the catalytic domain of glucuronoyl esterase Cip2 from the fungus Hypocrea jecorina was determined at a resolution of 1.9 Angstroms. This is the first structure of the newly established carbohydrate esterase family 15. The structure has revealed the residues Ser278–His411–Glu301 pre...

  2. 21 CFR 173.140 - Esterase-lipase derived from Mucor miehei.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... (CONTINUED) SECONDARY DIRECT FOOD ADDITIVES PERMITTED IN FOOD FOR HUMAN CONSUMPTION Enzyme Preparations and Microorganisms § 173.140 Esterase-lipase derived from Mucor miehei. Esterase-lipase enzyme, consisting of enzyme... animals. (c) The enzyme is produced by a process which completely removes the organism Mucor miehei...

  3. Para-nitrobenzyl esterases with enhanced activity in aqueous and nonaqueous media

    DOEpatents

    Arnold, Frances H.; Moore, Jeffrey C.

    1998-01-01

    A method for isolating and identifying modified para-nitrobenzyl esterases which exhibit improved stability and/or esterase hydrolysis activity toward selected substrates and under selected reaction conditions relative to the unmodified para-nitrobenzyl esterase. The method involves preparing a library of modified para-nitrobenzyl esterase nucleic acid segments (genes) which have nucleotide sequences that differ from the nucleic acid segment which encodes for unmodified para-nitrobenzyl esterase. The library of modified para-nitrobenzyl nucleic acid segments is expressed to provide a plurality of modified enzymes. The clones expressing modified enzymes are then screened to identify which enzymes have improved esterase activity by measuring the ability of the enzymes to hydrolyze the selected substrate under the selected reaction conditions. Specific modified para-nitrobenzyl esterases are disclosed which have improved stability and/or ester hydrolysis activity in aqueous or aqueous-organic media relative to the stability and/or ester hydrolysis activity of unmodified naturally occurring para-nitrobenzyl esterase.

  4. Para-nitrobenzyl esterases with enhanced activity in aqueous and nonaqueous media

    DOEpatents

    Arnold, Frances H.; Moore, Jeffrey C.

    1999-01-01

    A method for isolating and identifying modified para-nitrobenzyl esterases which exhibit improved stability and/or esterase hydrolysis activity toward selected substrates and under selected reaction conditions relative to the unmodified para-nitrobenzyl esterase. The method involves preparing a library of modified para-nitrobenzyl esterase nucleic acid segments (genes) which have nucleotide sequences that differ from the nucleic acid segment which encodes for unmodified para-nitrobenzyl esterase. The library of modified para-nitrobenzyl nucleic acid segments is expressed to provide a plurality of modified enzymes. The clones expressing modified enzymes are then screened to identify which enzymes have improved esterase activity by measuring the ability of the enzymes to hydrolyze the selected substrate under the selected reaction conditions. Specific modified para-nitrobenzyl esterases are disclosed which have improved stability and/or ester hydrolysis activity in aqueous or aqueous-organic media relative to the stability and/or ester hydrolysis activity of unmodified naturally occurring para-nitrobenzyl esterase.

  5. Para-nitrobenzyl esterases with enhanced activity in aqueous and nonaqueous media

    DOEpatents

    Arnold, F.H.; Moore, J.C.

    1998-04-21

    A method is disclosed for isolating and identifying modified para-nitrobenzyl esterases. These enzymes exhibit improved stability and/or esterase hydrolysis activity toward selected substrates and under selected reaction conditions relative to the unmodified para-nitrobenzyl esterase. The method involves preparing a library of modified para-nitrobenzyl esterase nucleic acid segments (genes) which have nucleotide sequences that differ from the nucleic acid segment which encodes for unmodified para-nitrobenzyl esterase. The library of modified para-nitrobenzyl nucleic acid segments is expressed to provide a plurality of modified enzymes. The clones expressing modified enzymes are then screened to identify which enzymes have improved esterase activity by measuring the ability of the enzymes to hydrolyze the selected substrate under the selected reaction conditions. Specific modified para-nitrobenzyl esterases are disclosed which have improved stability and/or ester hydrolysis activity in aqueous or aqueous-organic media relative to the stability and/or ester hydrolysis activity of unmodified naturally occurring para-nitrobenzyl esterase. 43 figs.

  6. 21 CFR 173.140 - Esterase-lipase derived from Mucor miehei.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... HUMAN CONSUMPTION Enzyme Preparations and Microorganisms § 173.140 Esterase-lipase derived from Mucor miehei. Esterase-lipase enzyme, consisting of enzyme derived from Mucor miehei var. Cooney et Emerson by... Emerson is nonpathogenic and nontoxic in man or other animals. (c) The enzyme is produced by a...

  7. 21 CFR 173.140 - Esterase-lipase derived from Mucor miehei.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... HUMAN CONSUMPTION Enzyme Preparations and Microorganisms § 173.140 Esterase-lipase derived from Mucor miehei. Esterase-lipase enzyme, consisting of enzyme derived from Mucor miehei var. Cooney et Emerson by... Emerson is nonpathogenic and nontoxic in man or other animals. (c) The enzyme is produced by a...

  8. 21 CFR 173.140 - Esterase-lipase derived from Mucor miehei.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... HUMAN CONSUMPTION Enzyme Preparations and Microorganisms § 173.140 Esterase-lipase derived from Mucor miehei. Esterase-lipase enzyme, consisting of enzyme derived from Mucor miehei var. Cooney et Emerson by... Emerson is nonpathogenic and nontoxic in man or other animals. (c) The enzyme is produced by a...

  9. 21 CFR 173.140 - Esterase-lipase derived from Mucor miehei.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... HUMAN CONSUMPTION Enzyme Preparations and Microorganisms § 173.140 Esterase-lipase derived from Mucor miehei. Esterase-lipase enzyme, consisting of enzyme derived from Mucor miehei var. Cooney et Emerson by... Emerson is nonpathogenic and nontoxic in man or other animals. (c) The enzyme is produced by a...

  10. Para-nitrobenzyl esterases with enhanced activity in aqueous and nonaqueous media

    DOEpatents

    Arnold, F.H.; Moore, J.C.

    1999-05-25

    A method is disclosed for isolating and identifying modified para-nitrobenzyl esterases which exhibit improved stability and/or esterase hydrolysis activity toward selected substrates and under selected reaction conditions relative to the unmodified para-nitrobenzyl esterase. The method involves preparing a library of modified para-nitrobenzyl esterase nucleic acid segments (genes) which have nucleotide sequences that differ from the nucleic acid segment which encodes for unmodified para-nitrobenzyl esterase. The library of modified para-nitrobenzyl nucleic acid segments is expressed to provide a plurality of modified enzymes. The clones expressing modified enzymes are then screened to identify which enzymes have improved esterase activity by measuring the ability of the enzymes to hydrolyze the selected substrate under the selected reaction conditions. Specific modified para-nitrobenzyl esterases are disclosed which have improved stability and/or ester hydrolysis activity in aqueous or aqueous-organic media relative to the stability and/or ester hydrolysis activity of unmodified naturally occurring para-nitrobenzyl esterase. 43 figs.

  11. Cloning, overexpression in Escherichia coli, and characterization of a thermostable fungal acetylxylan esterase from Talaromyces emersonii.

    PubMed

    Waters, Deborah M; Murray, Patrick G; Miki, Yuta; Martínez, Angel T; Tuohy, Maria G; Faulds, Craig B

    2012-05-01

    The gene encoding an acetylxylan esterase (AXE1) from the thermophilic ascomycete Talaromyces emersonii was cloned, expressed in Escherichia coli, and characterized. This form of AXE1, rTeAXE1, exhibits increased thermostability and activity at a higher temperature than other known fungal acetyl esterases, thus having huge potential application in biomass bioconversion to high value chemicals or biofuels. PMID:22407679

  12. Lipases or esterases: does it really matter? Toward a new bio-physico-chemical classification.

    PubMed

    Ali, Yassine Ben; Verger, Robert; Abousalham, Abdelkarim

    2012-01-01

    Carboxylester hydrolases, commonly named esterases, consist of a large spectrum of enzymes defined by their ability to catalyze the hydrolysis of carboxylic ester bonds and are widely distributed among animals, plants, and microorganisms. Lipases are lipolytic enzymes which constitute a special class of carboxylic esterases capable of releasing long-chain fatty acids from natural water-insoluble carboxylic esters. However, up to now, several unsuccessful attempts aimed at differentiating "lipases" from "esterases" by using various criteria. These criteria were based on the first substrate used chronologically, primary sequence comparisons, some kinetic parameters, or some structural features.Lipids are biological compounds which, by definition, are insoluble in water. Taking into account this basic physico-chemical criterion, we primarily distinguish lipolytic esterases (L, acting on lipids) from nonlipolytic esterases (NL, not acting on lipids). In view of the biochemical data accumulated up to now, we proposed a new classification of esterases based on various criteria of physico-chemical, chemical, anatomical, or cellular nature. We believe that the present attempt matters scientifically for several reasons: (1) to help newcomers in the field, performing a few key experiments to figure out if a newly isolated esterase is lipolytic or not; (2) to clarify a debate between scientists in the field; and (3) to formulate questions which are relevant to the still unsolved problem of the structure-function relationships of esterases. PMID:22426710

  13. Correlation of leukocyte esterase activity and bacterial isolation from body fluids.

    PubMed Central

    Smalley, D L; Bradley, M E

    1984-01-01

    We evaluated 230 body fluid samples, of which 131 were peritoneal effluents and 99 were other body fluids. Of these, 63 dialysates were culture positive, and 54 (85.7%) of these 63 were leukocyte esterase positive. Of 99 other body fluids, 8 were both culture positive and leukocyte esterase positive. PMID:6520224

  14. Lipases or esterases: does it really matter? Toward a new bio-physico-chemical classification.

    PubMed

    Ali, Yassine Ben; Verger, Robert; Abousalham, Abdelkarim

    2012-01-01

    Carboxylester hydrolases, commonly named esterases, consist of a large spectrum of enzymes defined by their ability to catalyze the hydrolysis of carboxylic ester bonds and are widely distributed among animals, plants, and microorganisms. Lipases are lipolytic enzymes which constitute a special class of carboxylic esterases capable of releasing long-chain fatty acids from natural water-insoluble carboxylic esters. However, up to now, several unsuccessful attempts aimed at differentiating "lipases" from "esterases" by using various criteria. These criteria were based on the first substrate used chronologically, primary sequence comparisons, some kinetic parameters, or some structural features.Lipids are biological compounds which, by definition, are insoluble in water. Taking into account this basic physico-chemical criterion, we primarily distinguish lipolytic esterases (L, acting on lipids) from nonlipolytic esterases (NL, not acting on lipids). In view of the biochemical data accumulated up to now, we proposed a new classification of esterases based on various criteria of physico-chemical, chemical, anatomical, or cellular nature. We believe that the present attempt matters scientifically for several reasons: (1) to help newcomers in the field, performing a few key experiments to figure out if a newly isolated esterase is lipolytic or not; (2) to clarify a debate between scientists in the field; and (3) to formulate questions which are relevant to the still unsolved problem of the structure-function relationships of esterases.

  15. Dona Ana Branch Community College Annual Report, 1990-1991.

    ERIC Educational Resources Information Center

    New Mexico State Univ., Las Cruces. Dona Ana Branch Community Coll.

    During 1990-91, New Mexico State University's (NMSU's) Dona Ana Branch Community College (DABCC) continued to feel the effects of its fourth year of rapidly increasing enrollments. The defeat of bond issues that would have funded facility expansions resulted in critical space shortages. The 27% increase in headcount enrollments between spring 1990…

  16. 21. ORIGINAL COMPANY HOUSE AT CORNER OF SANTA ANA AND ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    21. ORIGINAL COMPANY HOUSE AT CORNER OF SANTA ANA AND ANAHEIM BLVDS. (BEHIND HOUSE IN CA-242-20), WHICH IS BEING PREPARED FOR DEMOLITION. - Gene Pump Plant, South of Gene Wash Reservoir, 2 miles west of Whitsett Pump Plant, Parker Dam, San Bernardino County, CA

  17. A comparison of multiple esterases as biomarkers of organophosphate exposure and effect in two earthworm species.

    PubMed

    Henson-Ramsey, Heather; Schneider, Ashley; Stoskopf, Michael K

    2011-04-01

    Two different earthworm species, Eisenia fetida and Lumbricus terrestris, were exposed to 5 μg/cm(2) of malathion to evaluate their usefulness as sentinels of organophosphate exposure and to assess three different esterases, as biomarkers of malathion exposure and effect. Tissue xenobiotic burdens and esterase activity were determined for each species and each esterase in order to assess variability. E. fetida exhibited 4-fold less variability in tissue burdens than did L. terrestris and had less variable basal esterase activities. An attempt was made to correlate malathion and malaoxon tissue burdens with esterase activity post-exposure. There was no malaoxon present in the earthworm tissues. No significant correlations were determined by comparing acetylcholinesterase, butyrylcholinesterase, nor carboxylesterase activities with malathion burdens. PMID:21404045

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

    PubMed

    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.

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

    PubMed Central

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

    2011-01-01

    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. PMID:21887044

  20. Developing and Validating a Santa Ana Wildfire Threat Index

    NASA Astrophysics Data System (ADS)

    Capps, S. B.; Rolinski, T.; DAgostino, B.; Vanderburg, S.; Fovell, R. G.; Cao, Y.

    2014-12-01

    Santa Ana winds, common to southern California during the fall through spring, are a type of katabatic wind that originates from a direction generally ranging from 360°/0° to 100° and is usually accompanied by very low humidity. Since fuel conditions tend to be driest from late September through the middle of November, Santa Ana winds occurring during this period have the greatest potential to produce large, devastating fires when an ignition occurs. Such catastrophic fires occurred in 1993, 2003, 2007, and 2008. Because of the destructive nature of these fires, there has been a growing desire to categorize Santa Ana wind events in much the same way that tropical cyclones have been categorized. The Santa Ana Wildfire Threat index (SAWT) is an attempt to categorize such events with respect to fire activity, based on surface wind velocity, dew point depression, and forecasted fuel conditions. The index, a USDA Forest Service product, was developed by the Forest Service in collaboration with San Diego Gas and Electric Utility (SDG&E), the Department of Atmospheric and Oceanic Sciences at UCLA, The Desert Research Institute (DRI), and Vertum Partners. The methodology behind the SAWT index, along with the index itself will be presented in detail. Also, there will be a discussion on the construction of a 30-year climatology of the index, which includes various meteorological and fuel parameters. We will demonstrate the usefulness of the index as another decision support tool for fire agencies and first responders, and how it could assist the general public and private industry in the preparation of critical Santa Ana wind events.

  1. The Impact of Santa Ana Winds on Wildland Fire Regimes

    NASA Astrophysics Data System (ADS)

    Billmire, M.; Loboda, T. V.; French, N. H.; Tyner, M.

    2011-12-01

    Santa Ana winds have been implicated as major drivers of extensive wildfires that occur annually in southern California. While numerous anecdotal reports dictate an obvious relationship, there is little quantitative analysis in current literature on how this loosely-defined weather phenomenon impacts fire progression regimes. A new satellite-derived fire progression dataset developed using MODIS and Landsat was linked to a network of 82 Remote Automated Weather Stations (RAWS) to evaluate three differing a priori classifications of Santa Ana events defined using three weather parameters: wind speed, wind direction, and relative humidity. The fire progression dataset comprised 528 burn area polygons representing 151 distinct fire events from 2001-2009 in southern California. Each burn area polygon was assigned weather data from one of the RAWS units by shortest straight-line distance. These data and methods show quantitatively that burn area is dramatically larger under Santa Ana conditions than under non-Santa Ana conditions (see table). Outliers of both types (large burn areas under non-SA conditions; small burn areas under SA conditions) were identified and closely examined to identify potentially confounding variables. Time-lag effects (particularly with respect to cumulative precipitation preceding day of burn) and effects with regard to local vs. regional measurements were examined as well. We tested 17 variables (3 relative humidity variables, 5 wind speed variables, 3 temperature variables, 3 moisture variables, previous day burn area, census-derived population density, and the number of hours meeting one Santa Ana classification) individually and in combination for correlation with both burn area and burn area change from the previous day to test their predictive power. Mean daily relative humidity was found to have the strongest correlation (Pearson's r = -0.451) with burn area. All variables except moisture variables were more strongly correlated with the

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

    PubMed Central

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

    2012-01-01

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

  3. Characterization of Heterologously Expressed Acetyl Xylan Esterase1 Isolated from the Anaerobic Rumen Fungus Neocallimastix frontalis PMA02

    PubMed Central

    Kwon, Mi; Song, Jaeyong; Park, Hong-Seog; Park, Hyunjin; Chang, Jongsoo

    2016-01-01

    Acetyl xylan esterase (AXE), which hydrolyzes the ester linkages of the naturally acetylated xylan and thus known to have an important role for hemicellulose degradation, was isolated from the anaerobic rumen fungus Neocallimastix frontatlis PMA02, heterologously expressed in Escherichi coli (E.coli) and characterized. The full-length cDNA encoding NfAXE1 was 1,494 bp, of which 978 bp constituted an open reading frame. The estimated molecular weight of NfAXE1 was 36.5 kDa with 326 amino acid residues, and the calculated isoelectric point was 4.54. The secondary protein structure was predicted to consist of nine α-helixes and 12 β-strands. The enzyme expressed in E.coli had the highest activity at 40°C and pH 8. The purified recombinant NfAXE1 had a specific activity of 100.1 U/mg when p-nitrophenyl acetate (p-NA) was used as a substrate at 40°C, optimum temperature. The amount of liberated acetic acids were the highest and the lowest when p-NA and acetylated birchwood xylan were used as substrates, respectively. The amount of xylose released from acetylated birchwod xylan was increased by 1.4 fold when NfAXE1 was mixed with xylanase in a reaction cocktail, implying a synergistic effect of NfAXE1 with xylanase on hemicellulose degradation. PMID:27383808

  4. Branched nanotrees with immobilized acetylcholine esterase for nanobiosensor applications

    NASA Astrophysics Data System (ADS)

    Risveden, Klas; Dick, Kimberly A.; Bhand, Sunil; Rydberg, Patrik; Samuelson, Lars; Danielsson, Bengt

    2010-02-01

    A novel lab-on-a-chip nanotree enzyme reactor is demonstrated for the detection of acetylcholine. The reactors are intended for use in the RISFET (regional ion sensitive field effect transistor) nanosensor, and are constructed from gold-tipped branched nanorod structures grown on SiNx-covered wafers. Two different reactors are shown: one with simple, one-dimensional nanorods and one with branched nanorod structures (nanotrees). Significantly higher enzymatic activity is found for the nanotree reactors than for the nanorod reactors, most likely due to the increased gold surface area and thereby higher enzyme binding capacity. A theoretical calculation is included to show how the enzyme kinetics and hence the sensitivity can be influenced and increased by the control of electrical fields in relation to the active sites of enzymes in an electronic biosensor. The possible effects of electrical fields employed in the RISFET on the function of acetylcholine esterase is investigated using quantum chemical methods, which show that the small electric field strengths used are unlikely to affect enzyme kinetics. Acetylcholine esterase activity is determined using choline oxidase and peroxidase by measuring the amount of choline formed using the chemiluminescent luminol reaction.

  5. A halotolerant type A feruloyl esterase from Pleurotus eryngii.

    PubMed

    Nieter, Annabel; Haase-Aschoff, Paul; Linke, Diana; Nimtz, Manfred; Berger, Ralf G

    2014-03-01

    An extracellular feruloyl esterase (PeFaeA) from the culture supernatant of Pleurotus eryngii was purified to homogeneity using cation exchange, hydrophobic interaction, and size exclusion chromatography. The length of the complete coding sequence of PeFaeA was determined to 1668 bp corresponding to a protein of 555 amino acids. The catalytic triad of Ser-Glu-His demonstrated the uniqueness of the enzyme compared to previously published FAEs. The purified PeFaeA was a monomer with an estimated molecular mass of 67 kDa. Maximum feruloyl esterase (FAE) activity was observed at pH 5.0 and 50 °C, respectively. Metal ions (5 mM), except Hg(2+), had no significant influence on the enzyme activity. Substrate specificity profiling characterized the enzyme as a type A FAE preferring bulky natural substrates, such as feruloylated saccharides, rather than small synthetic ones. Km and kcat of the purified enzyme for methyl ferulate were 0.15 mM and 0.85 s(-1). In the presence of 3 M NaCl activity of the enzyme increased by 28 %. PeFaeA alone released only little ferulic acid from destarched wheat bran (DSWB), whereas after addition of Trichoderma viride xylanase the concentration increased more than 20 fold. PMID:24607359

  6. Food induced esterase phenocopies in the snail Cepaea nemoralis.

    PubMed

    Oxford, G S

    1975-12-01

    Hepatopancreatic extracts from the snail Cepaea nemoralis, assayed straight from the field, often contain three or four heavily staining esterase zones which migrate to the cathodal end of polyacrylamide disc gels during electrophoresis. Previous breeding results showed that the heavily straining zones appeared allelic but to incorporate these multibanded phenotypes, a super gene of five closely linked loci was tentatively proposed. Further breeding work again failed to demonstrate multiple zones in parents or offspring and so experiments were conducted to see whether the multi-zoned phenotypes in the wild were produced by secondary modification of single primary products. Wild snails yielding extracts containing more than two heavily staining zones were shown to possess only two such zones after three months under laboratory conditions. Also, the ingestion of nettle (Urtica dioica L.) has been demonstrated to induce extra esterase zones in laboratory-reared animals. Some of the secondarily induced zones appear identical in physical, biochemical and electrophoretic properties to the primary products of other alleles, and thus appear to be electrophoretic phenocopies. A model is suggested which could account for this phenomenon. PMID:1061709

  7. Methane from acetate.

    PubMed

    Ferry, J G

    1992-09-01

    The general features are known for the pathway by which most methane is produced in nature. All acetate-utilizing methanogenic microorganisms contain CODH which catalyzes the cleavage of acetyl-CoA; however, the pathway differs from all other acetate-utilizing anaerobes in that the methyl group is reduced to methane with electrons derived from oxidation of the carbonyl group of acetyl-CoA to CO2. The current understanding of the methanogenic fermentation of acetate provides impressions of nature's novel solutions to problems of methyl transfer, electron transport, and energy conservation. The pathway is now at a level of understanding that will permit productive investigations of these and other interesting questions in the near future. PMID:1512186

  8. An organic-solvent-tolerant esterase from thermophilic Bacillus licheniformis S-86.

    PubMed

    Torres, Sebastián; Martínez, M Alejandra; Pandey, Ashok; Castro, Guillermo R

    2009-01-01

    A thermophile, halotolerant and organic-solvent-tolerant esterase producer Bacillus sp. S-86 strain previously isolated was found to belong to Bacillus licheniformis species through morphological, biochemical, 16S rRNA gene sequence analyses and rDNA intergenic spacers amplification (ITS-PCR). The strain can grow at 55 degrees C in presence of C2-C7 alkanols (log P=-0.86 to 2.39), and NaCl concentrations up to 15% (w/v). This bacterium showed optimal growth and esterase production at 50 degrees C. Two different molecular weight esterase activities were detected in zymographic assays. PMSF inhibited type I esterase activity, showing no inhibitory effect on type II esterase activity. B. licheniformis S-86 was able to grow in presence of hydroxylic organic-solvents like propan-2-ol, butan-1-ol and 3-methylbutan-1-ol. At a sub-lethal concentration of these solvents (392 mmoll(-1) propan-2-ol; 99 mmol l(-1) butan-1-ol, 37 mmol l(-1) 3-methylbutan-1-ol), adequate to produce 50% cell growth inhibition at 50 degrees C, an increment between 1.9 and 2.3 times was observed in type I esterase production, and between 2.2 and 3.1 times in type II esterase production. PMID:18723341

  9. Esterase and lipase in camel tick Hyalomma dromedarii (Acari: Ixodidae) during embryogenesis.

    PubMed

    Fahmy, Afaf S; Abdel-Gany, Somia S; Mohamed, Tarek M; Mohamed, Saleh A

    2004-02-01

    Esterase and lipase activity showed significant changes during embryogenesis of camel tick Hyalomma dromedarii. From the elution profile of chromatography on DEAE-cellulose, six forms of H. dromedarii esterase (El to EVI) can be distinguished. Esterase EIII was purified to homogeneity after chromatography on Sepharose 6B. The molecular mass of esterase EIII was 45 kDa for the native enzyme and represented a monomer of 45 kDa by SDS-PAGE. Esterase EIII had an acidic pI at 5.3. Lipase activity was detected in the same DEAE-cellulose peaks (LI to LVI) of H. dromedarii esterases. The highest lipase activity was exhibited by lipase LIII. Esterase EIII and lipase LIII were compared with respect to Michaelis constant, substrate specificity, temperature optimum, heat stability, pH optimum, effect of metal ions and inhibitors. This study suggests that H. dromedarii lipolytic enzymes may play a central role in the interconversion of lipovitellins during embryogenesis. PMID:14990212

  10. Comparison of mesophilic and thermophilic feruloyl esterases: characterization of their substrate specificity for methyl phenylalkanoates.

    PubMed

    Topakas, Evangelos; Christakopoulos, Paul; Faulds, Craig B

    2005-02-23

    The active sites of feruloyl esterases from mesophilic and thermophilic sources were probed using methyl esters of phenylalkanoic acids. Only 13 out of 26 substrates tested were significant substrates for all the enzymes. Lengthening or shortening the aliphatic side chain while maintaining the same aromatic substitutions completely abolished activity for both enzymes, which demonstrates the importance of the correct distance between the aromatic group and the ester bond. Maintaining the phenylpropanoate structure but altering the substitutions of the aromatic ring demonstrated that the type-A esterase from the mesophilic fungus Fusarium oxysporum (FoFaeA) showed a preference for methoxylated substrates, in contrast to the type-B esterase from the same source (FoFaeB) and the thermophilic type-B (StFaeB) and type-C (StFaeC) from Sporotrichum thermophile, which preferred hydroxylated substrates. All four esterases hydrolyzed short chain aliphatic acid (C2-C4) esters of p-nitrophenol, but not the C12 ester of laurate. All the feruloyl esterases were able to release ferulic acid from the plant cell wall material in conjunction with a xylanase, but only the type-A esterase FoFaeA was effective in releasing the 5,5' form of diferulic acid. The thermophilic type-B esterase had a lower catalytic efficiency than its mesophilic counterpart, but released more ferulic acid from plant cell walls.

  11. Purification and properties of an esterase from organophosphate-resistant strain of the mosquito Culex quinquefasciatus.

    PubMed Central

    Merryweather, A T; Crampton, J M; Townson, H

    1990-01-01

    Organophosphate-resistant and -susceptible strains of Culex quinquefasciatus (mosquito) have been compared on the basis of their esterase activities. The homozygous resistant strain (Dar) shows two highly active esterases after starch-gel electrophoresis, of Rm 0.2 and 0.4, which are absent from susceptible strains (Apo, Mon), and which previous selection studies have shown to be inseparable from organophosphate resistance. After SDS/polyacrylamide-gel electrophoresis and silver staining of total C. quinquefasciatus proteins, a 62 kDa band is observed in strain Dar at high concentrations, and in susceptible strains in trace amounts. After Western blotting, this 62 kDa protein is recognized by antisera raised against the two esterases eluted from starch gels. After chromatofocusing of Dar proteins, the 62 kDa protein is seen to be associated with esterase activity, and of a similar pI to that observed for esterases after isoelectric focusing. Post-translational modification is not required for recognition of the 62 kDa putative esterase, since the protein is immunoprecipitated by the anti-esterase serum from products of translation of Dar mRNA in vitro. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 6. Fig. 7. PMID:2178604

  12. Distribution of esterase activity in porcine ear skin, and the effects of freezing and heat separation.

    PubMed

    Lau, Wing Man; Ng, Keng Wooi; Sakenyte, Kristina; Heard, Charles M

    2012-08-20

    Porcine ear skin is widely used to study skin permeation and absorption of ester compounds, whose permeation and absorption profiles may be directly influenced by in situ skin esterase activity. Importantly, esterase distribution and activity in porcine ear skin following common protocols of skin handling and storage have not been characterised. Thus, we have compared the distribution and hydrolytic activity of esterases in freshly excised, frozen, heated and explanted porcine ear skin. Using an esterase staining kit, esterase activity was found to be localised in the stratum corneum and viable epidermis. Under frozen storage and a common heating protocol of epidermal sheet separation, esterase staining in the skin visibly diminished. This was confirmed by a quantitative assay using HPLC to monitor the hydrolysis of aspirin, in freshly excised, frozen or heated porcine ear skin. Compared to vehicle-only control, the rate of aspirin hydrolysis was approximately three-fold higher in the presence of freshly excised skin, but no different in the presence of frozen or heated skin. Therefore, frozen and heat-separated porcine ear skin should not be used to study the permeation of ester-containing permeants, in particular co-drugs and pro-drugs, whose hydrolysis or degradation can be modulated by skin esterases.

  13. An antennal carboxylesterase from Drosophila melanogaster, esterase 6, is a candidate odorant-degrading enzyme toward food odorants.

    PubMed

    Chertemps, Thomas; Younus, Faisal; Steiner, Claudia; Durand, Nicolas; Coppin, Chris W; Pandey, Gunjan; Oakeshott, John G; Maïbèche, Martine

    2015-01-01

    Reception of odorant molecules within insect olfactory organs involves several sequential steps, including their transport through the sensillar lymph, interaction with the respective sensory receptors, and subsequent inactivation. Odorant-degrading enzymes (ODEs) putatively play a role in signal dynamics by rapid degradation of odorants in the vicinity of the receptors, but this hypothesis is mainly supported by in vitro results. We have recently shown that an extracellular carboxylesterase, esterase-6 (EST-6), is involved in the physiological and behavioral dynamics of the response of Drosophila melanogaster to its volatile pheromone ester, cis-vaccenyl acetate. However, as the expression pattern of the Est-6 gene in the antennae is not restricted to the pheromone responding sensilla, we tested here if EST-6 could play a broader function in the antennae. We found that recombinant EST-6 is able to efficiently hydrolyse several volatile esters that would be emitted by its natural food in vitro. Electrophysiological comparisons of mutant Est-6 null flies and a control strain (on the same genetic background) showed that the dynamics of the antennal response to these compounds is influenced by EST-6, with the antennae of the null mutants showing prolonged activity in response to them. Antennal responses to the strongest odorant, pentyl acetate, were then studied in more detail, showing that the repolarization dynamics were modified even at low doses but without modification of the detection threshold. Behavioral choice experiments with pentyl acetate also showed differences between genotypes; attraction to this compound was observed at a lower dose among the null than control flies. As EST-6 is able to degrade various bioactive odorants emitted by food and plays a role in the response to these compounds, we hypothesize a role as an ODE for this enzyme toward food volatiles. PMID:26594178

  14. An antennal carboxylesterase from Drosophila melanogaster, esterase 6, is a candidate odorant-degrading enzyme toward food odorants

    PubMed Central

    Chertemps, Thomas; Younus, Faisal; Steiner, Claudia; Durand, Nicolas; Coppin, Chris W.; Pandey, Gunjan; Oakeshott, John G.; Maïbèche, Martine

    2015-01-01

    Reception of odorant molecules within insect olfactory organs involves several sequential steps, including their transport through the sensillar lymph, interaction with the respective sensory receptors, and subsequent inactivation. Odorant-degrading enzymes (ODEs) putatively play a role in signal dynamics by rapid degradation of odorants in the vicinity of the receptors, but this hypothesis is mainly supported by in vitro results. We have recently shown that an extracellular carboxylesterase, esterase-6 (EST-6), is involved in the physiological and behavioral dynamics of the response of Drosophila melanogaster to its volatile pheromone ester, cis-vaccenyl acetate. However, as the expression pattern of the Est-6 gene in the antennae is not restricted to the pheromone responding sensilla, we tested here if EST-6 could play a broader function in the antennae. We found that recombinant EST-6 is able to efficiently hydrolyse several volatile esters that would be emitted by its natural food in vitro. Electrophysiological comparisons of mutant Est-6 null flies and a control strain (on the same genetic background) showed that the dynamics of the antennal response to these compounds is influenced by EST-6, with the antennae of the null mutants showing prolonged activity in response to them. Antennal responses to the strongest odorant, pentyl acetate, were then studied in more detail, showing that the repolarization dynamics were modified even at low doses but without modification of the detection threshold. Behavioral choice experiments with pentyl acetate also showed differences between genotypes; attraction to this compound was observed at a lower dose among the null than control flies. As EST-6 is able to degrade various bioactive odorants emitted by food and plays a role in the response to these compounds, we hypothesize a role as an ODE for this enzyme toward food volatiles. PMID:26594178

  15. Identification of a Secreted Lipolytic Esterase in Propionibacterium freudenreichii, a Ripening Process Bacterium Involved in Emmental Cheese Lipolysis▿ †

    PubMed Central

    Dherbécourt, J.; Falentin, H.; Jardin, J.; Maillard, M.-B.; Baglinière, F.; Barloy-Hubler, F.; Thierry, A.

    2010-01-01

    Lipolysis plays an important role in the formation of cheese flavor. In Emmental cheese, the main part of lipolysis has been associated with the presence of Propionibacterium freudenreichii, a species used as a ripening culture. Our aim was to identify the most probable lipolytic esterase(s) involved in cheese lipolysis by P. freudenreichii. Since cheese lipolysis mainly occurs during P. freudenreichii growth, we hypothesized that P. freudenreichii possesses secreted lipolytic esterase(s). For 12 putative esterase genes previously identified from the genome of P. freudenreichii CIRM1, the level of expression was quantified by real-time reverse transcriptase (RT)-PCR, and the subcellular localization of esterases was predicted in silico. The esterase activity in extracellular and intracellular extracts of P. freudenreichii was characterized by zymography, and the extracellular esterases were identified by mass spectrometry. Finally, the best candidate was overexpressed in the same strain. All of the 12 genes encoding putative esterases were expressed. Esterase PF#279 was predicted to be secreted in the medium, PF#774 to be surface exposed, and the 10 remaining putative esterases to be intracellular. Zymography revealed that esterase activities in culture supernatant differed from the ones detected in intracellular extracts. PF#279 was identified as the sole esterase present in culture supernatant. Transformed P. freudenreichii CIRM1 clones overexpressing PF#279 showed 5 to 8 times more lipolytic activity on milk fat than the wild-type strain. Combining in silico, biochemical, and genetic approaches, we showed that PF#279 is the sole secreted esterase in P. freudenreichii and is active on milk fat. Therefore, it is likely a key component in cheese lipolysis by P. freudenreichii. PMID:20038704

  16. DOM in recharge waters of the Santa Ana River Basin

    USGS Publications Warehouse

    Leenheer, J.A.; Aiken, G.R.; Woodside, G.; O'Connor-Patel, K.

    2007-01-01

    The urban Santa Ana River in California is the primary source of recharge water for Orange County's groundwater basin, which provides water to more than two million residents. This study was undertaken to determine the unidentified portion of dissolved organic matter (DOM) in various natural surface and reclaimed waters of the Santa Ana River Basin and to assess the potential health risk of this material. The most abundant organic contaminants were anionic detergent degradation products (constituting about 12% of the DOM), which have no known adverse health effects. In addition, high percentages of dissolved colloids from bacterial cell walls were found during storm flows; these colloids foul membranes used in water treatment. Although no significant health risks were ascribed to the newly characterized DOM, the authors note that even the small amounts of humic substances deposited during storm flow periods were responsible for significant increases in disinfection by_product formation potential in these waters.

  17. Online Stigma Resistance in the Pro-Ana Community.

    PubMed

    Yeshua-Katz, Daphna

    2015-10-01

    Media scholars often use concepts from Goffman's dramaturgical approach to study online communities of stigmatized individuals as "backstages," spaces where members take refuge from social disapproval. In this study, I extend this view through an examination of in-depth interviews with bloggers from the "pro-ana" community, an online community for people with eating disorders. To explore how this community uses an online environment that is both anonymous and public, I fuse Goffman's ideas about identity performance and stigma with more recent theories about boundary maintenance. In-depth interviews with "pro-ana" bloggers reveal that to protect this virtual group and resist stigmas associated both with their illness and with their online presence, they construct their own norms and rules in the online realm, and discipline and eject members deemed to be out-group.

  18. Online Stigma Resistance in the Pro-Ana Community.

    PubMed

    Yeshua-Katz, Daphna

    2015-10-01

    Media scholars often use concepts from Goffman's dramaturgical approach to study online communities of stigmatized individuals as "backstages," spaces where members take refuge from social disapproval. In this study, I extend this view through an examination of in-depth interviews with bloggers from the "pro-ana" community, an online community for people with eating disorders. To explore how this community uses an online environment that is both anonymous and public, I fuse Goffman's ideas about identity performance and stigma with more recent theories about boundary maintenance. In-depth interviews with "pro-ana" bloggers reveal that to protect this virtual group and resist stigmas associated both with their illness and with their online presence, they construct their own norms and rules in the online realm, and discipline and eject members deemed to be out-group. PMID:25667161

  19. Evolutionary routes from a prebiotic ANA-world.

    PubMed

    Braun, Sebastian; Humphreys, Christine; Dale, Trevor C

    2012-03-01

    Recent experimental support has been generated for a model of prebiotic development that postulates a role for Amyloid-Nucleic Acid (ANA)-fibers as the earliest replicating entities capable of undergoing Darwinian evolution. Here, this new model is compared with existing RNA-world models with a particular focus on trajectories that lead to evolutionary-beneficial interactions between nucleic acid, protein and lipid components. This analysis suggests a number of new areas for fruitful experimental studies. PMID:22808333

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

    NASA Astrophysics Data System (ADS)

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

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

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

    SciTech Connect

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

    2008-01-01

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

  2. Esterase variation at three loci in meat ants.

    PubMed

    Halliday, R B

    1979-01-01

    The meat ant (Iridomyrmex purpureus) occurs in a number of color forms, with uncertain taxonomic status. Gel electrophresis of meat ant extracts, followed by nonspecific esterase staining, reveals several zones of activity. Allelic variation at three loci is proposed to account for variation in some of these zones. Two of the loci (Es-1, Es-2) appear to have recessive null alleles, whose frequencies have been estimated by the method of maximum likelihood. Geographic variation in allele frequency is attributed to behavioral and geographic subdivision of the population. Apparent disturbances in segregation ratios and deviations from Hardy-Weinberg equilibrium can be accounted for if it is argued that some nests contain more than one queen. Differences in gene frequency between sympatric populations of the red and blue forms of I. purpureus are observed, confirming their reproductive is isolation and sibling species status.

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

    PubMed

    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. PMID:27452816

  4. Contrasting controls on wildland fires in Southern California during periods with and without Santa Ana winds

    NASA Astrophysics Data System (ADS)

    Jin, Yufang; Randerson, James T.; Faivre, Nicolas; Capps, Scott; Hall, Alex; Goulden, Michael L.

    2014-03-01

    Wildland fires in Southern California can be divided into two categories: fall fires, which are typically driven by strong offshore Santa Ana winds, and summer fires, which occur with comparatively weak onshore winds and hot and dry weather. Both types of fire contribute significantly to annual burned area and economic loss. An improved understanding of the relationship between Southern California's meteorology and fire is needed to improve predictions of how fire will change in the future and to anticipate management needs. We used output from a regional climate model constrained by reanalysis observations to identify Santa Ana events and partition fires into those occurring during periods with and without Santa Ana conditions during 1959-2009. We then developed separate empirical regression models for Santa Ana and non-Santa Ana fires to quantify the effects of meteorology on fire number and size. These models explained approximately 58% of the seasonal and interannual variation in the number of Santa Ana fires and 36% of the variation in non-Santa Ana fires. The number of Santa Ana fires increased during years when relative humidity during Santa Ana events and fall precipitation were below average, indicating that fuel moisture is a key controller of ignition. Relative humidity strongly affected Santa Ana fire size. Cumulative precipitation during the previous three winters was significantly correlated with the number of non-Santa Ana fires, presumably through increased fine fuel density and connectivity between infrastructure and nearby vegetation. Both relative humidity and the preceding wet season precipitation influenced non-Santa Ana fire size. Regression models driven by meteorology explained 57% of the temporal variation in Santa Ana burned area and 22% of the variation in non-Santa Ana burned area. The area burned by non-Santa Ana fires has increased steadily by 1.7% year-1 since 1959 (p < 0.006) the occurrence of extremely large Santa Ana fires has

  5. Drosophila Ana1 is required for centrosome assembly and centriole elongation

    PubMed Central

    Saurya, Saroj; Roque, Hélio; Novak, Zsofia A.; Wainman, Alan; Aydogan, Mustafa G.; Volanakis, Adam; Sieber, Boris; Pinto, David Miguel Susano

    2016-01-01

    ABSTRACT Centrioles organise centrosomes and cilia, and these organelles have an important role in many cell processes. In flies, the centriole protein Ana1 is required for the assembly of functional centrosomes and cilia. It has recently been shown that Cep135 (also known as Bld10) initially recruits Ana1 to newly formed centrioles, and that Ana1 then recruits Asl (known as Cep152 in mammals) to promote the conversion of these centrioles into centrosomes. Here, we show that ana1 mutants lack detectable centrosomes in vivo, that Ana1 is irreversibly incorporated into centrioles during their assembly and appears to play a more important role in maintaining Asl at centrioles than in initially recruiting Asl to centrioles. Unexpectedly, we also find that Ana1 promotes centriole elongation in a dose-dependent manner: centrioles are shorter when Ana1 dosage is reduced and are longer when Ana1 is overexpressed. This latter function of Ana1 appears to be distinct from its role in centrosome and cilium function, as a GFP–Ana1 fusion lacking the N-terminal 639 amino acids of the protein can support centrosome assembly and cilium function but cannot promote centriole over-elongation when overexpressed. PMID:27206860

  6. Drosophila Ana1 is required for centrosome assembly and centriole elongation.

    PubMed

    Saurya, Saroj; Roque, Hélio; Novak, Zsofia A; Wainman, Alan; Aydogan, Mustafa G; Volanakis, Adam; Sieber, Boris; Pinto, David Miguel Susano; Raff, Jordan W

    2016-07-01

    Centrioles organise centrosomes and cilia, and these organelles have an important role in many cell processes. In flies, the centriole protein Ana1 is required for the assembly of functional centrosomes and cilia. It has recently been shown that Cep135 (also known as Bld10) initially recruits Ana1 to newly formed centrioles, and that Ana1 then recruits Asl (known as Cep152 in mammals) to promote the conversion of these centrioles into centrosomes. Here, we show that ana1 mutants lack detectable centrosomes in vivo, that Ana1 is irreversibly incorporated into centrioles during their assembly and appears to play a more important role in maintaining Asl at centrioles than in initially recruiting Asl to centrioles. Unexpectedly, we also find that Ana1 promotes centriole elongation in a dose-dependent manner: centrioles are shorter when Ana1 dosage is reduced and are longer when Ana1 is overexpressed. This latter function of Ana1 appears to be distinct from its role in centrosome and cilium function, as a GFP-Ana1 fusion lacking the N-terminal 639 amino acids of the protein can support centrosome assembly and cilium function but cannot promote centriole over-elongation when overexpressed.

  7. A novel cold-adapted and highly salt-tolerant esterase from Alkalibacterium sp. SL3 from the sediment of a soda lake.

    PubMed

    Wang, Guozeng; Wang, Qiaohuang; Lin, Xianju; Ng, Tzi Bun; Yan, Renxiang; Lin, Juan; Ye, Xiuyun

    2016-01-01

    A novel esterase gene (estSL3) was cloned from the Alkalibacterium sp. SL3, which was isolated from the sediment of soda lake Dabusu. The 636-bp full-length gene encodes a polypeptide of 211 amino acid residues that is closely related with putative GDSL family lipases from Alkalibacterium and Enterococcus. The gene was successfully expressed in E. coli, and the recombinant protein (rEstSL3) was purified to electrophoretic homogeneity and characterized. rEstSL3 exhibited the highest activity towards pNP-acetate and had no activity towards pNP-esters with acyl chains longer than C8. The enzyme was highly cold-adapted, showing an apparent temperature optimum of 30 °C and remaining approximately 70% of the activity at 0 °C. It was active and stable over the pH range from 7 to 10, and highly salt-tolerant up to 5 M NaCl. Moreover, rEstSL3 was strongly resistant to most tested metal ions, chemical reagents, detergents and organic solvents. Amino acid composition analysis indicated that EstSL3 had fewer proline residues, hydrogen bonds and salt bridges than mesophilic and thermophilic counterparts, but more acidic amino acids and less hydrophobic amino acids when compared with other salt-tolerant esterases. The cold active, salt-tolerant and chemical-resistant properties make it a promising enzyme for basic research and industrial applications.

  8. A novel cold-adapted and highly salt-tolerant esterase from Alkalibacterium sp. SL3 from the sediment of a soda lake

    PubMed Central

    Wang, Guozeng; Wang, Qiaohuang; Lin, Xianju; Bun Ng, Tzi; Yan, Renxiang; Lin, Juan; Ye, Xiuyun

    2016-01-01

    A novel esterase gene (estSL3) was cloned from the Alkalibacterium sp. SL3, which was isolated from the sediment of soda lake Dabusu. The 636-bp full-length gene encodes a polypeptide of 211 amino acid residues that is closely related with putative GDSL family lipases from Alkalibacterium and Enterococcus. The gene was successfully expressed in E. coli, and the recombinant protein (rEstSL3) was purified to electrophoretic homogeneity and characterized. rEstSL3 exhibited the highest activity towards pNP-acetate and had no activity towards pNP-esters with acyl chains longer than C8. The enzyme was highly cold-adapted, showing an apparent temperature optimum of 30 °C and remaining approximately 70% of the activity at 0 °C. It was active and stable over the pH range from 7 to 10, and highly salt-tolerant up to 5 M NaCl. Moreover, rEstSL3 was strongly resistant to most tested metal ions, chemical reagents, detergents and organic solvents. Amino acid composition analysis indicated that EstSL3 had fewer proline residues, hydrogen bonds and salt bridges than mesophilic and thermophilic counterparts, but more acidic amino acids and less hydrophobic amino acids when compared with other salt-tolerant esterases. The cold active, salt-tolerant and chemical-resistant properties make it a promising enzyme for basic research and industrial applications. PMID:26915906

  9. The Chlamydia trachomatis CT149 protein exhibits esterase activity in vitro and catalyzes cholesteryl ester hydrolysis when expressed in HeLa cells

    PubMed Central

    Peters, Jan; Onguri, Vijaya; Nishimoto, Satoru K.; Marion, Tony N.; Byrne, Gerald I.

    2012-01-01

    Chlamydia, like other intracellular bacteria, are auxotrophic for a variety of essential metabolites and obtain cholesterol and fatty acids from their eukaryotic host cell, however not many Chlamydia-specific enzymes have been identified that are involved in lipid metabolism. In silico analysis of one candidate C. trachomatis enzyme, annotated as a conserved putative hydrolase (CT149), identified two lipase/esterase GXSXG motifs, and a potential cholesterol recognition/interaction amino acid consensus (CRAC) sequence. His-tag purified recombinant CT149 exhibited ester hydrolysis activity in a nitrophenyl acetate-based cell-free assay system. When cholesteryl linoleate was used as substrate, ester hydrolysis occurred and production of cholesterol was detected by high performance liquid chromatography. Exogenous expression of transfected CT149 in HeLa cells resulted in a significant decrease of cytoplasmic cholesteryl esters within 48 hrs. These results demonstrate that CT149 has cholesterol esterase activity and is likely to contribute to the hydrolysis of eukaryotic cholesteryl esters during intracellular chlamydial growth. PMID:22940277

  10. Influenza C virus esterase: analysis of catalytic site, inhibition, and possible function.

    PubMed Central

    Vlasak, R; Muster, T; Lauro, A M; Powers, J C; Palese, P

    1989-01-01

    The active site serine of the acetylesterase of influenza C virus was localized to amino acid 71 of the hemagglutinin-esterase protein by affinity labeling with 3H-labeled diisopropylfluorophosphate. This serine and the adjacent amino acids (Phe-Gly-Asp-Ser) are part of a consensus sequence motif found in serine hydrolases. Since comparative analysis failed to reveal esterase sequence similarities with other serine hydrolases, we suggest that this viral enzyme is a serine hydrolase constituting a new family of serine esterases. Furthermore, we found that the influenza C virus esterase was inhibited by isocoumarin derivatives, with 3,4-dichloroisocoumarin being the most potent inhibitor. Addition of this compound prevented elution of influenza C virus from erythrocytes and inhibited virus infectivity, possibly through inhibition of virus entry into cells. Images PMID:2495370

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

    EPA Science Inventory

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

  12. Exposure to a Cutinase-like Serine Esterase Triggers Rapid Lysis of Multiple Mycobacterial Species*

    PubMed Central

    Yang, Yong; Bhatti, Alexandra; Ke, Danxia; Gonzalez-Juarrero, Mercedes; Lenaerts, Anne; Kremer, Laurent; Guerardel, Yann; Zhang, Peijun; Ojha, Anil K.

    2013-01-01

    Mycobacteria are shaped by a thick envelope made of an array of uniquely structured lipids and polysaccharides. However, the spatial organization of these molecules remains unclear. Here, we show that exposure to an esterase from Mycobacterium smegmatis (Msmeg_1529), hydrolyzing the ester linkage of trehalose dimycolate in vitro, triggers rapid and efficient lysis of Mycobacterium tuberculosis, Mycobacterium bovis BCG, and Mycobacterium marinum. Exposure to the esterase immediately releases free mycolic acids, while concomitantly depleting trehalose mycolates. Moreover, lysis could be competitively inhibited by an excess of purified trehalose dimycolate and was abolished by a S124A mutation affecting the catalytic activity of the esterase. These findings are consistent with an indispensable structural role of trehalose mycolates in the architectural design of the exposed surface of the mycobacterial envelope. Importantly, we also demonstrate that the esterase-mediated rapid lysis of M. tuberculosis significantly improves its detection in paucibacillary samples. PMID:23155047

  13. Esterase and Malate Dehydrogenase Phenotypes in Portuguese Populations of Meloidogyne Species

    PubMed Central

    Pais, Célia S.; de O. Abrantes, Isabel M.

    1989-01-01

    Nonspecific esterases and malate dehydrogenases of 1-5 females from 40 root-knot nematode populations from Portugal were analyzed by electrophoresis in 0.4-mm-thick polyacrylamide gels. Fourteen major bands of esterase activity were detected, corresponding to 10 distinct phenotypes, Meloidogyne javanica and M. hapla had distinct species-specific phenotypes. Two phenotypes occurred in M. arenaria. The most variability was found among M. incognita populations. Of the remaining two phenotypes, one was associated with M. hispanica and the other belonged to a new species. Three malate dehydrogenase phenotypes were discerned on the basis of particular combinations of the eight main bands of activity found. As previously found, esterases were more useful than malate dehydrogenases in identification of the major Meloidogyne species. The host plant had no effect on the nematode esterase or malate dehydrogenase phenotypes. PMID:19287618

  14. Influenza C virus esterase: analysis of catalytic site, inhibition, and possible function

    SciTech Connect

    Vlasak, R.; Muster, T.; Lauro, A.M.; Powers, J.C.; Palese, P.

    1989-05-01

    The active site serine of the acetylesterase of influenza C virus was localized to amino acid 71 of the hemagglutinin-esterase protein by affinity labeling with /sup 3/H-labeled diisopropylfluorophosphate. This serine and the adjacent amino acids (Phe-Gly-Asp-Ser) are part of a consensus sequence motif found in serine hydrolases. Since comparative analysis failed to reveal esterase sequence similarities with other serine hydrolases, the authors suggest that this viral enzyme is a serine hydrolase constituting a new family of serine esterases. Furthermore, they found that the influenza C virus esterase was inhibited by isocoumarin derivatives, with 3,4-dichloroisocoumarin being the most potent inhibitor. Addition of this compound prevented elution of influenza C virus from erythrocytes and inhibited virus infectivity, possibly through inhibition of virus entry into cells.

  15. Acetate Production by Methanogenic Bacteria

    PubMed Central

    Westermann, Peter; Ahring, Birgitte K.; Mah, Robert A.

    1989-01-01

    Methanosarcina barkeri MS and 227 and Methanosarcina mazei S-6 produced acetate when grown on H2-CO2, methanol, or trimethylamine. Marked differences in acetate production by the two bacterial species were found, even though methane and cell yields were nearly the same. M. barkeri produced 30 to 75 μmol of acetate per mmol of CH4 formed, but M. mazei produced only 8 to 9 μmol of acetate per mmol of CH4. PMID:16348006

  16. The search of the target of promotion: Phenylbenzoate esterase activities in hen peripheral nerve

    SciTech Connect

    Moretto, A. . E-mail: angelo.moretto@icps.it; Nicolli, A.; Lotti, M.

    2007-03-15

    Certain esterase inhibitors, such as carbamates, phosphinates and sulfonyl halides, do not cause neuropathy as some organophosphates, but they may exacerbate chemical or traumatic insults to axons. This phenomenon is called promotion of axonopathies. Given the biochemical and toxicological characteristics of these compounds, the hypothesis was made that the target of promotion is a phenyl valerate (PV) esterase similar to neuropathy target esterase (NTE), the target of organophosphate induced delayed polyneuropathy. However, attempts to identify a PV esterase in hen peripheral nerve have been, so far, unsuccessful. We tested several esters, other than PV, as substrates of esterases from crude homogenate of the hen peripheral nerve. The ideal substrate should be poorly hydrolysed by NTE but extensively by enzyme(s) that are insensitive to non-promoters, such as mipafox, and sensitive to promoters, such as phenyl methane sulfonyl fluoride (PMSF). When phenyl benzoate (PB) was used as substrate, about 65% of total activity was resistant to the non-promoter mipafox (up to 0.5 mM, 20 min, pH 8.0), that inhibits NTE and other esterases. More than 90% of this resistant activity was sensitive to the classical promoter PMSF (1 mM, 20 min, pH 8.0) with an IC{sub 50} of about 0.08 mM (20 min, pH 8.0). On the contrary, the non-promoter p-toluene sulfonyl fluoride caused only about 10% inhibition at 0.5 mM. Several esterase inhibitors including, paraoxon, phenyl benzyl carbamate, di-n-butyl dichlorovinyl phosphate and di-isopropyl fluorophosphate, were tested both in vitro and in vivo for inhibition of this PB activity. Mipafox-resistant PMSF-sensitive PB esterase activity(ies) was inhibited by promoters but not by non promoters and neuropathic compounds.

  17. Acetate dependence of tumors.

    PubMed

    Comerford, Sarah A; Huang, Zhiguang; Du, Xinlin; Wang, Yun; Cai, Ling; Witkiewicz, Agnes K; Walters, Holly; Tantawy, Mohammed N; Fu, Allie; Manning, H Charles; Horton, Jay D; Hammer, Robert E; McKnight, Steven L; Tu, Benjamin P

    2014-12-18

    Acetyl-CoA represents a central node of carbon metabolism that plays a key role in bioenergetics, cell proliferation, and the regulation of gene expression. Highly glycolytic or hypoxic tumors must produce sufficient quantities of this metabolite to support cell growth and survival under nutrient-limiting conditions. Here, we show that the nucleocytosolic acetyl-CoA synthetase enzyme, ACSS2, supplies a key source of acetyl-CoA for tumors by capturing acetate as a carbon source. Despite exhibiting no gross deficits in growth or development, adult mice lacking ACSS2 exhibit a significant reduction in tumor burden in two different models of hepatocellular carcinoma. ACSS2 is expressed in a large proportion of human tumors, and its activity is responsible for the majority of cellular acetate uptake into both lipids and histones. These observations may qualify ACSS2 as a targetable metabolic vulnerability of a wide spectrum of tumors.

  18. Characterization and structural modeling of a new type of thermostable esterase from Thermotoga maritima.

    PubMed

    Levisson, Mark; van der Oost, John; Kengen, Servé W M

    2007-06-01

    A bioinformatic screening of the genome of the hyperthermophilic bacterium Thermotoga maritima for ester-hydrolyzing enzymes revealed a protein with typical esterase motifs, though annotated as a hypothetical protein. To confirm its putative esterase function the gene (estD) was cloned, functionally expressed in Escherichia coli and purified to homogeneity. Recombinant EstD was found to exhibit significant esterase activity with a preference for short acyl chain esters (C4-C8). The monomeric enzyme has a molecular mass of 44.5 kDa and optimal activity around 95 degrees C and at pH 7. Its thermostability is relatively high with a half-life of 1 h at 100 degrees C, but less stable compared to some other hyperthermophilic esterases. A structural model was constructed with the carboxylesterase Est30 from Geobacillus stearothermophilus as a template. The model covered most of the C-terminal part of EstD. The structure showed an alpha/beta-hydrolase fold and indicated the presence of a typical catalytic triad consisting of a serine, aspartate and histidine, which was verified by site-directed mutagenesis and inhibition studies. Phylogenetic analysis showed that EstD is only distantly related to other esterases. A comparison of the active site pentapeptide motifs revealed that EstD should be grouped into a new family of esterases (Family 10). EstD is the first characterized member of this family. PMID:17466017

  19. VvMJE1 of the grapevine (Vitis vinifera) VvMES methylesterase family encodes for methyl jasmonate esterase and has a role in stress response.

    PubMed

    Zhao, Nan; Lin, Hong; Lan, Suque; Jia, Qidong; Chen, Xinlu; Guo, Hong; Chen, Feng

    2016-05-01

    The known members of plant methyl esterase (MES) family catalyze the hydrolysis of a C-O ester linkage of methyl esters of several phytohormones including indole-3-acetic acid, salicylic acid and jasmonic acid. The genome of grapevine (Vitis vinifera) was found to contain 15 MES genes, designated VvMES1-15. In this report, VvMES5 was selected for molecular, biochemical and structural studies. VvMES5 is most similar to tomato methyl jasmonate esterase. E. coli-expressed recombinant VvMES5 displayed methyl jasmonate (MeJA) esterase activity, it was renamed VvMJE1. Under steady-state conditions, VvMJE1 exhibited an apparent Km value of 92.9 μM with MeJA. VvMJE1 was also shown to have lower activity with methyl salicylate (MeSA), another known substrate of the MES family, and only at high concentrations of the substrate. To understand the structural basis of VvMJE1 in discriminating MeJA and MeSA, a homolog model of VvMJE1 was made using the X-ray structure of tobacco SABP2, which encodes for methyl salicylate esterase, as a template. Interestingly, two bulky residues at the binding site and near the surface of tobacco SABP2 are replaced by relatively small residues in VvMJE1. Such a change enables the accommodation of a larger substrate MeJA in VvMJE1. The expression of VvMJE1 was compared in control grape plants and grape plants treated with one of the three stresses: heat, cold and UV-B. While the expression of VvMJE1 was not affected by heat treatment, its expression was significantly up-regulated by cold treatment and UV-B treatment. This result suggests that VvMJE1 has a role in response of grape plants to these two abiotic stresses. PMID:26934101

  20. Functional Characterization of a Novel Dactylosporangium Esterase and Its Utilization in the Asymmetric Synthesis of (R)-Methyl Mandelate.

    PubMed

    Deng, Dun; Zhang, Yun; Sun, Aijun; Hu, Yunfeng

    2016-09-01

    One novel esterase DAEst6 was identified from the genome of Dactylosporangium aurantiacum subsp. Hamdenensis NRRL 18085. DAEst6 was further characterized to be an esterase which exhibited high resistance to high pH values. Esterase DAEst6 could resolve racemic methyl mandelate and generate (R)-methyl mandelate, one key drug intermediate, with an enantiomeric excess and a conversion of 99 and 49 %, respectively, after process optimization. The optimal working condition for the preparation of (R)-methyl mandelate through DAEst6 was found to be 10-mM racemic methyl mandelate, no organic co-solvents, pH 7.5, and 40 °C, for 5 h. Our work was the first report about the functional characterization of one novel Dactylosporangium esterase and the utilization of one Dactylosporangium esterase in kinetic resolution. Dactylosporangium esterases represented by DAEst6 possess great potential in the generation of valuable chiral drug intermediates and chemicals.

  1. Contribution of Doñana Wetlands to Carbon Sequestration

    PubMed Central

    Morris, Edward P.; Flecha, Susana; Figuerola, Jordi; Costas, Eduardo; Navarro, Gabriel; Ruiz, Javier; Rodriguez, Pablo; Huertas, Emma

    2013-01-01

    Inland and transitional aquatic systems play an important role in global carbon (C) cycling. Yet, the C dynamics of wetlands and floodplains are poorly defined and field data is scarce. Air-water fluxes in the wetlands of Doñana Natural Area (SW Spain) were examined by measuring alkalinity, pH and other physiochemical parameters in a range of water bodies during 2010–2011. Areal fluxes were calculated and, using remote sensing, an estimate of the contribution of aquatic habitats to gaseous transport was derived. Semi-permanent ponds adjacent to the large Guadalquivir estuary acted as mild sinks, whilst temporal wetlands were strong sources of (−0.8 and 36.3 ). Fluxes in semi-permanent streams and ponds changed seasonally; acting as sources in spring-winter and mild sinks in autumn (16.7 and −1.2 ). Overall, Doñana's water bodies were a net annual source of (5.2 ). Up–scaling clarified the overwhelming contribution of seasonal flooding and allochthonous organic matter inputs in determining regional air-water gaseous transport (13.1 ). Nevertheless, this estimate is about 6 times < local marsh net primary production, suggesting the system acts as an annual net sink. Initial indications suggest longer hydroperiods may favour autochthonous C capture by phytoplankton. Direct anthropogenic impacts have reduced the hydroperiod in Doñana and this maybe exacerbated by climate change (less rainfall and more evaporation), suggesting potential for the modification of C sequestration. PMID:23977044

  2. Esterase SeE of Streptococcus equi ssp. equi is a Novel Non-specific Carboxylic Ester Hydrolase

    PubMed Central

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

    2009-01-01

    Extracellular carboxylic ester hydrolases are produced by many bacterial pathogens and have been shown recently to be important for virulence of some pathogens. However, these hydrolases are poorly characterized in enzymatic activity. This study prepared and characterized the secreted ester hydrolase of Streptococcus equi ssp. equi (designated SeE for S. equi esterase). SeE hydrolyzes ethyl acetate, acetylsalicylic acid, and tributyrin but not ethyl butyrate. This substrate specificity pattern does not match those of the three conventional types of non-specific carboxylic ester hydrolases (carboxylesterases, arylesterases, and acetylesterases). To determine whether SeE has lipase activity, a number of triglycerides and vinyl esters were tested in SeE-catalyzed hydrolysis. SeE does not hydrolyze triglycerides and vinyl esters of long chain carboxylic acids nor display interfacial activation, indicating that SeE is not a lipase. Like the conventional carboxylesterases, SeE is inhibited by diisopropylfluorophosphate. These findings indicate that SeE is a novel non-specific carboxylic ester hydrolase that has broader substrate specificity than the conventional carboxylesterases. PMID:19054107

  3. Comparison of chaparral regrowth patterns between Santa Ana wind-driven and non-Santa Ana fire areas

    NASA Astrophysics Data System (ADS)

    Rachels, Diane Helen

    Wildfires are a common occurrence in California shrublands and island forests. Fire has a fundamental role in maintaining the ecosystem functions in chaparral where fire intensity and severity play important roles in the regeneration of species. In San Diego, the Cedar Fire that occurred in the fall of 2003 was unique in that one side was burned with wildfire fueled by dry, strong easterly Santa Ana winds that later died down, burning the remainder of the area under a mild westerly wind, allowing fuel-fed conditions. The objective of this study was to understand the connection between vegetation type and structure and environmental response to extreme fire events by analyzing life form regrowth in chaparral communities from the Santa Ana wind driven, Santa Ana backing, and non-Santa Ana fire types. Environmental factors of slope angle, aspect, elevation and soils were investigated in an effort to isolate shrub regrowth patterns. Fire burn characteristics, anthropogenic disturbance, fire history, and moisture availability were also analyzed to identify additional factors that may have influenced shrub regrowth. Shrub extents before the fire and six year after the fire were examined per slope aspect, slope angle, elevation, and fire characteristic categories. The closed canopy and natural features of the chaparral environment make ground based mapping very difficult. Remote sensing data and methods can be very helpful to evaluate the health of the vegetation and condition of the watershed for flood, erosion, and fire control. This study used high spatial resolution aerial imagery and a machine learning algorithm with a spatial contextual classifier to map three different areas from within the Cedar Fire perimeter. Geographic information science (GIS), field mapping, and image interpretation methods were used to identify vegetation samples for the classification and accuracy assessment of the vegetation maps. Object-based image samples were selected for the classifier

  4. Structural analysis of thermostabilizing mutations of cocaine esterase

    SciTech Connect

    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.

    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.

  5. Inhibition of monocyte esterase activity by organophosphate insecticides.

    PubMed

    Lee, M J; Waters, H C

    1977-11-01

    Organophosphate insecticides, such as Vapona, Naled, and Rabon, are highly potent inhibitors of an enzyme found in human monocytes. The enzyme, a specific monocyte esterase, could be inhibited by Vapona in blood samples via airborne contamination at levels easily achieved from commercial slow-release insecticide strips. Fifty percent inhibition (I50)--as measured on the Hemalog D (Technicon Corp.)--occurred at solution concentrations of 0.22, 1.5, and 2.6 X 10(-6) g/liter for Vapona, Rabon, and Naled, respectively. Parathion (a thiophosphate) and Baygon (a carbamate) were less potent, with I50 values of 3.7 X 10(-5) and 1.5 X 10(-4) g/liter, respectively. Dursban (another thiophosphate) and Carbaryl (a carbamate) showed only marginal inhibition. Eserine, malathion, nicotine and pyrethrum had no inhibitory effect up to 0.5 g/liter. The occurrence of this effect in vivo has not yet been shown, nor is it clear what the implications of such an effect would be. The inhibition of this enzyme by airborne contaminants, however, may interfere with the proper functioning of the Hemalog D. PMID:907842

  6. Identification of petrogenic produced water components as acetylcholine esterase inhibitors.

    PubMed

    Froment, Jean; Langford, Katherine; Tollefsen, Knut Erik; Bråte, Inger Lise N; Brooks, Steven J; Thomas, Kevin V

    2016-08-01

    Effect-directed analysis (EDA) was applied to identify acetylcholine esterase (AChE) inhibitors in produced water. Common produced water components from oil production activities, such as polycyclic aromatic hydrocarbons (PAHs), alkylphenols, and naphthenic acids were tested for AChE inhibition using a simple mixture of PAHs and naphthenic acids. Produced water samples collected from two offshore platforms in the Norwegian sector of the North Sea were extracted by solid phase extraction and fractionated by open-column liquid solid chromatography and high-performance liquid chromatography (HPLC) before being tested using a high-throughput and automated AChE assay. The HPLC fractions causing the strongest AChE inhibition were analysed by gas chromatography coupled to a high-resolution time-of-flight mass spectrometry (GC-HR-ToF-MS). Butylated hydroxytoluene and 4-phenyl-1,2-dihydronaphthalene were identified as two produced water components capable of inhibiting AChE at low concentrations. In order to assess the potential presence of such compounds discharged into aquatic ecosystems, AChE activity in fish tissues was measured. Saithe (Pollachius virens) caught near two offshore platforms showed lower enzymatic activity than those collected from a reference location. Target analysis of saithe did not detected the presence of these two putative AChE inhibitors and suggest that additional compounds such as PAHs, naphthenic acids and yet un-identified compounds may also contribute to the purported AChE inhibition observed in saithe. PMID:27176761

  7. Mycobacteriophage Lysin B is a novel mycolylarabinogalactan esterase

    SciTech Connect

    Payne, K.; Sun, Q.; Sacchettini, J.; Hatfull, G.F.

    2010-08-27

    Mycobacteriophages encounter a unique problem among phages of Gram-positive bacteria, in that lysis must not only degrade the peptidoglycan layer but also circumvent a mycolic acid-rich outer membrane covalently attached to the arabinogalactan-peptidoglycan complex. Mycobacteriophages accomplish this by producing two lysis enzymes, Lysin A (LysA) that hydrolyses peptidoglycan, and Lysin B (LysB), a novel mycolylarabinogalactan esterase, that cleaves the mycolylarabinogalactan bond to release free mycolic acids. The D29 LysB structure shows an {alpha}/{beta} hydrolase organization with a catalytic triad common to cutinases, but which contains an additional four-helix domain implicated in the binding of lipid substrates. Whereas LysA is essential for mycobacterial lysis, a Giles {Delta}lysB mutant mycobacteriophage is viable, but defective in the normal timing, progression and completion of host cell lysis. We propose that LysB facilitates lysis by compromising the integrity of the mycobacterial outer membrane linkage to the arabinogalactan-peptidoglycan layer.

  8. Lipases and esterases from extremophiles: overview and case example of the production and purification of an esterase from Thermus thermophilus HB27.

    PubMed

    Fuciños, Pablo; González, Roberto; Atanes, Estrella; Sestelo, Ana Belén Fernández; Pérez-Guerra, Nelson; Pastrana, Lorenzo; Rúa, María Luisa

    2012-01-01

    Extremophiles are organisms that have evolved to exist in a variety of extreme environments. They fall into a number of different classes that include thermophiles, halophiles, acidophiles, alkalophiles, psychrophiles, and barophiles (piezophiles). Extremophiles have the potential to produce uniquely valuable biocatalysts that function under conditions in which usually the enzymes of their nonextremophilic counterparts could not. Among novel enzymes isolated from extremophilic microorganisms, hydrolases, and particularly lipases and esterases are experiencing a growing demand. Lipases (EC 3.1.1.3) and esterases (EC 3.1.1.1) catalyze the cleavage of ester bounds in aqueous media and the reverse reaction in organic solvents. Both lipolytic enzymes have relevant applications in food, dairy, detergent, biofuel, and pharmaceutical industries. Here, we summarize the properties of lipases and esterases from the main extremophile groups: thermophiles and hyperthermophiles, psychrophiles, halophiles, alkalophiles/acidophiles, and solvent-resistant microorganisms.We report the biomass and lipolytic activity production by Thermus thermophilus HB27 in 5-L stirred-tank bioreactor at 70°C. Suitability of thermal spring water for culture media formulation is shown. In addition, a protocol to isolate and purify a cell-bound esterase from this microorganism is described.

  9. Lipases and esterases from extremophiles: overview and case example of the production and purification of an esterase from Thermus thermophilus HB27.

    PubMed

    Fuciños, Pablo; González, Roberto; Atanes, Estrella; Sestelo, Ana Belén Fernández; Pérez-Guerra, Nelson; Pastrana, Lorenzo; Rúa, María Luisa

    2012-01-01

    Extremophiles are organisms that have evolved to exist in a variety of extreme environments. They fall into a number of different classes that include thermophiles, halophiles, acidophiles, alkalophiles, psychrophiles, and barophiles (piezophiles). Extremophiles have the potential to produce uniquely valuable biocatalysts that function under conditions in which usually the enzymes of their nonextremophilic counterparts could not. Among novel enzymes isolated from extremophilic microorganisms, hydrolases, and particularly lipases and esterases are experiencing a growing demand. Lipases (EC 3.1.1.3) and esterases (EC 3.1.1.1) catalyze the cleavage of ester bounds in aqueous media and the reverse reaction in organic solvents. Both lipolytic enzymes have relevant applications in food, dairy, detergent, biofuel, and pharmaceutical industries. Here, we summarize the properties of lipases and esterases from the main extremophile groups: thermophiles and hyperthermophiles, psychrophiles, halophiles, alkalophiles/acidophiles, and solvent-resistant microorganisms.We report the biomass and lipolytic activity production by Thermus thermophilus HB27 in 5-L stirred-tank bioreactor at 70°C. Suitability of thermal spring water for culture media formulation is shown. In addition, a protocol to isolate and purify a cell-bound esterase from this microorganism is described. PMID:22426723

  10. The Secreted Esterase of Propionibacterium freudenreichii Has a Major Role in Cheese Lipolysis

    PubMed Central

    Abeijón Mukdsi, María Claudia; Falentin, Hélène; Maillard, Marie-Bernadette; Chuat, Victoria; Medina, Roxana Beatriz; Parayre, Sandrine

    2014-01-01

    Free fatty acids are important flavor compounds in cheese. Propionibacterium freudenreichii is the main agent of their release through lipolysis in Swiss cheese. Our aim was to identify the esterase(s) involved in lipolysis by P. freudenreichii. We targeted two previously identified esterases: one secreted esterase, PF#279, and one putative cell wall-anchored esterase, PF#774. To evaluate their role in lipolysis, we constructed overexpression and knockout mutants of P. freudenreichii CIRM-BIA1T for each corresponding gene. The sequences of both genes were also compared in 21 wild-type strains. All strains were assessed for their lipolytic activity on milk fat. The lipolytic activity observed matched data previously reported in cheese, thus validating the relevance of the method used. The mutants overexpressing PF#279 or PF#774 released four times more fatty acids than the wild-type strain, demonstrating that both enzymes are lipolytic esterases. However, inactivation of the pf279 gene induced a 75% reduction in the lipolytic activity compared to that of the wild-type strain, whereas inactivation of the pf774 gene did not modify the phenotype. Two of the 21 wild-type strains tested did not display any detectable lipolytic activity. Interestingly, these two strains exhibited the same single-nucleotide deletion at the beginning of the pf279 gene sequence, leading to a premature stop codon, whereas they harbored a pf774 gene highly similar to that of the other strains. Taken together, these results clearly demonstrate that PF#279 is the main lipolytic esterase in P. freudenreichii and a key agent of Swiss cheese lipolysis. PMID:24242250

  11. A carboxylesterase, Esterase-6, modulates sensory physiological and behavioral response dynamics to pheromone in Drosophila

    PubMed Central

    2012-01-01

    Background Insects respond to the spatial and temporal dynamics of a pheromone plume, which implies not only a strong response to 'odor on', but also to 'odor off'. This requires mechanisms geared toward a fast signal termination. Several mechanisms may contribute to signal termination, among which odorant-degrading enzymes. These enzymes putatively play a role in signal dynamics by a rapid inactivation of odorants in the vicinity of the sensory receptors, although direct in vivo experimental evidences are lacking. Here we verified the role of an extracellular carboxylesterase, esterase-6 (Est-6), in the sensory physiological and behavioral dynamics of Drosophila melanogaster response to its pheromone, cis-vaccenyl acetate (cVA). Est-6 was previously linked to post-mating effects in the reproductive system of females. As Est-6 is also known to hydrolyze cVA in vitro and is expressed in the main olfactory organ, the antenna, we tested here its role in olfaction as a putative odorant-degrading enzyme. Results We first confirm that Est-6 is highly expressed in olfactory sensilla, including cVA-sensitive sensilla, and we show that expression is likely associated with non-neuronal cells. Our electrophysiological approaches show that the dynamics of olfactory receptor neuron (ORN) responses is strongly influenced by Est-6, as in Est-6° null mutants (lacking the Est-6 gene) cVA-sensitive ORN showed increased firing rate and prolonged activity in response to cVA. Est-6° mutant males had a lower threshold of behavioral response to cVA, as revealed by the analysis of two cVA-induced behaviors. In particular, mutant males exhibited a strong decrease of male-male courtship, in association with a delay in courtship initiation. Conclusions Our study presents evidence that Est-6 plays a role in the physiological and behavioral dynamics of sex pheromone response in Drosophila males and supports a role of Est-6 as an odorant-degrading enzyme (ODE) in male antennae. Our results

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

    PubMed

    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.

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

    PubMed

    Alex, Deepthy; Shainu, Anju; Pandey, Ashok; Sukumaran, Rajeev K

    2014-01-01

    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 K m and V max of 0.01 and 1.12, respectively. The purified esterase lost its thermo and halo tolerance but interestingly, retained 97% activity in methanol. PMID:24800063

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

    PubMed Central

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

    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 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. PMID:24204998

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

    PubMed

    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. PMID:25575887

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

    PubMed Central

    Shainu, Anju; Pandey, Ashok; Sukumaran, Rajeev K.

    2014-01-01

    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. PMID:24800063

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

    PubMed

    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.

  18. Selective induction of xenobiotic metabolizing esterases/amidases of liver by methaqualone consumption.

    PubMed

    Kaur, S; Ali, B

    1983-08-01

    The present investigation reports the influence of po and ip methaqualone administration on the hydrolytic metabolism of acetylsalicylic acid, procaine, p-nitrophenylacetate, acetanilid, and butyrylcholine in the liver, kidney, and brain of male rats. Oral administration of methaqualone (60 mg/kg/day) to rats for 20 days caused 41.0, 46.5, and 55.0% stimulation of acetylsalicyclic acid esterase I, acetylsalicyclic acid esterase II, and acetanilid N-deacetylase, respectively, in the liver. Under such conditions, the activities of other esterases remained unaffected. The responses of tissue esterases to ip methaqualone treatment (40 mg/kg/day for 6 days) were similar to those observed after po methaqualone administration. Since a single po dose of methaqualone failed to produce any alteration in the rate of metabolism of acetylsalicylic acid, procaine, p-nitrophenylacetate, acetanilid, and butyrylcholine within 20 hr, it may be interpreted that the stimulation of acetylsalicylic acid and acetanilid metabolism is possibly due to selective enhanced de novo synthesis of the enzymes/isozymes necessary for the hydrolysis of the two drugs. The ability of the kidney and brain to metabolize the esters/amides was not modified by po or ip methaqualone pretreatment suggesting the possibility of noninducible forms of renal and neuronal esterases/amidases.

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

    PubMed

    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.

  20. The classification of esterases: an important gene family involved in insecticide resistance--a review.

    PubMed

    Montella, Isabela Reis; Schama, Renata; Valle, Denise

    2012-06-01

    The use of chemical insecticides continues to play a major role in the control of disease vector populations, which is leading to the global dissemination of insecticide resistance. A greater capacity to detoxify insecticides, due to an increase in the expression or activity of three major enzyme families, also known as metabolic resistance, is one major resistance mechanisms. The esterase family of enzymes hydrolyse ester bonds, which are present in a wide range of insecticides; therefore, these enzymes may be involved in resistance to the main chemicals employed in control programs. Historically, insecticide resistance has driven research on insect esterases and schemes for their classification. Currently, several different nomenclatures are used to describe the esterases of distinct species and a universal standard classification does not exist. The esterase gene family appears to be rapidly evolving and each insect species has a unique complement of detoxification genes with only a few orthologues across species. The examples listed in this review cover different aspects of their biochemical nature. However, they do not appear to contribute to reliably distinguish among the different resistance mechanisms. Presently, the phylogenetic criterion appears to be the best one for esterase classification. Joint genomic, biochemical and microarray studies will help unravel the classification of this complex gene family.

  1. Insecticidal properties of genetically engineered baculoviruses expressing an insect juvenile hormone esterase gene.

    PubMed Central

    Eldridge, R; O'Reilly, D R; Hammock, B D; Miller, L K

    1992-01-01

    Exploring the possibility of enhancing the properties of baculoviruses as biological control agents of insect pests, we tested the effect of expressing an insect gene (jhe) encoding juvenile hormone esterase. Juvenile hormone esterase inactivates juvenile hormone, which regulates the outcome of an insect molt. A cDNA encoding the juvenile hormone esterase of Heliothis virescens was inserted into the genome of Autographa californica nuclear polyhedrosis virus such that the gene was expressed under the control of a strong, modified viral promoter. This virus, however, naturally encodes an ecdysteroid UDP-glucosyltransferase which inactivates ecdysone, the hormone which initiates molting. Since ecdysteroid UDP-glucosyltransferase could mask the effects of jhe expression by blocking molting entirely, jhe-expressing viruses in which the ecdysteroid UDP-glucosyltransferase gene was deleted or disrupted were constructed. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of proteins from infected cells revealed several intracellular proteins and two major secreted proteins which reacted with antibodies to authentic juvenile hormone esterase. Western blot analysis coupled with tunicamycin treatment indicated that differential glycosylation was responsible for the multiple products. Hemolymph of recombinant virus-infected fourth-instar Trichoplusia ni larvae contained levels of juvenile hormone esterase activity 40-fold higher than maximal levels found in uninfected larvae. However, little or no difference in developmental characteristics, weight gain, or time of mortality was observed between insects infected with the jhe-expressing viruses and control viruses. Images PMID:1622228

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

    PubMed

    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. PMID:26369647

  3. A novel esterase from a marine metagenomic library exhibiting salt tolerance ability.

    PubMed

    Fang, Zemin; Li, Jingjing; Wang, Quan; Fang, Wei; Peng, Hui; Zhang, Xuecheng; Xiao, Yazhong

    2014-06-28

    A putative lipolytic enzyme gene, named as est9x, was obtained from a marine microbial metagenome of the South China Sea. Sequence analysis showed that Est9X shares lower than 27% sequence identities with the characterized lipolytic enzymes, but possesses a catalytic triad highly conserved in lipolytic enzymes of the α/β hydrolase superfamily. By phylogenetic tree construction, Est9X was grouped into a new lipase/esterase family. To understand Est9X protein in depth, it was recombinantly expressed, purified, and biochemically characterized. Within potential hydrolytic activities, only lipase/esterase activity was detected for Est9X, confirming its identity as a lipolytic enzyme. When using p-nitrophenol esters with varying lengths of fatty acid as substrates, Est9X exhibited the highest activity to the C2 substrate, indicating it is an esterase. The optimal activity of Est9X occurred at a temperature of 65°C, and Est9X was pretty stable below the optimum temperature. Distinguished from other salttolerant esterases, Est9X's activity was tolerant to and even promoted by as high as 4 M NaCl. Our results imply that Est9X is a unique esterase and could be a potential candidate for industrial application under extreme conditions.

  4. The loss rates of web tags applied to day-old Anas and Aythya ducklings

    USGS Publications Warehouse

    Blums, P.; Mednis, A.; Bauga, I.; Nichols, J.D.; Hines, J.E.

    1997-01-01

    Researchers studied the loss rate of web tags on Anas and Aythya ducklings by double marking day-old ducklings of five species with web tags and plasticine-filled rings. Tag loss was examined over three-month, one-year, and three-year periods. Web tag loss was greatest for Anas and occurred mostly in the first three months following tagging.

  5. Cytotoxicity and DNA-protein crosslink formation in rat nasal tissues exposed to vinyl acetate are carboxylesterase-mediated.

    PubMed

    Kuykendall, J R; Taylor, M L; Bogdanffy, M S

    1993-12-01

    Vinyl acetate is used in the paint, adhesive, and paper board industries. Vinyl acetate is a nasal carcinogen in rats exposed by inhalation for 2 years to 200 and 600 ppm, but not 50 ppm. Previous studies from our laboratory suggest that rat liver microsome-activated vinyl acetate induces plasmid DNA-histone crosslinks, in vitro, through esterase-mediated metabolism. Since nasal tissues contain high levels of carboxylesterase, tumorigenesis may be related to in situ production of the hydrolysis products acetaldehyde and acetic acid. Vinyl acetate was cytotoxic to both respiratory and olfactory tissues in vitro at 50-200 mM, but not 25 mM, after 2 hr exposure. Pretreatment of rats with the carboxylesterase inhibitor, bis-(p-nitrophenyl) phosphate (BNPP), attenuated the cytotoxic effects and metabolism of vinyl acetate in both tissue types. Semicarbazide, an aldehyde scavenger, was unable to protect the tissues from vinyl acetate-induced cytotoxicity. When the metabolites were tested, acetic acid, but not acetaldehyde, was cytotoxic to both tissues. The induction of DNA-protein crosslink (DPXL) formation by acetaldehyde and vinyl acetate in rat nasal epithelial tissues was detected using a sodium dodecyl sulfate/KCl precipitation technique. Endogenous crosslink levels ranged from 0.5 to 2.0% of total DNA and were considered background. Epithelial cells isolated from both olfactory and respiratory turbinates exhibited dose- and time-dependent increases in DPXL formation when exposed to 10-150 mM acetaldehyde for 1-2 hr at 37 degrees C. Similarly, respiratory and olfactory epithelial cells exposed to 5-75 mM vinyl acetate for 1-2 hr accumulated up to 12- and 15-fold higher crosslink levels than untreated cells, respectively. However, vinyl acetate appears to induce much higher levels of DPXLs at equimolar doses than acetaldehyde. This is thought to be related to stimulation of acetaldehyde-induced DPXL formation by the pH lowering effect of acetic acid production (via

  6. Cellular function of neuropathy target esterase in lysophosphatidylcholine action

    SciTech Connect

    Vose, Sarah C.; Fujioka, Kazutoshi; Gulevich, Alex G.; Lin, Amy Y.; Holland, Nina T.; Casida, John E.

    2008-11-01

    Neuropathy target esterase (NTE) plays critical roles in embryonic development and maintenance of peripheral axons. It is a secondary target of some organophosphorus toxicants including analogs of insecticides and chemical warfare agents. Although the mechanistic role of NTE in vivo is poorly defined, it is known to hydrolyze lysophosphatidylcholine (LPC) in vitro and may protect cell membranes from cytotoxic accumulation of LPC. To determine the cellular function of NTE, Neuro-2a and COS-7 cells were transfected with a full-length human NTE-containing plasmid yielding recombinant NTE (rNTE). We find the same inhibitor sensitivity and specificity profiles for rNTE assayed with LPC or phenyl valerate (a standard NTE substrate) and that this correlation extends to the LPC hydrolases of human brain, lymphocytes and erythrocytes. All of these LPC hydrolases are therefore very similar to each other in respect to a conserved inhibitor binding site conformation. NTE is expressed in brain and lymphocytes and contributes to LPC hydrolase activities in these tissues. The enzyme or enzymes responsible for erythrocyte LPC hydrolase activity remain to be identified. We also show that rNTE protects Neuro-2a and COS-7 cells from exogenous LPC cytotoxicity. Expression of rNTE in Neuro-2a cells alters their phospholipid balance (analyzed by liquid chromatography-mass spectrometry with single ion monitoring) by lowering LPC-16:0 and LPC-18:0 and elevating glycerophosphocholine without a change in phosphatidylcholine-16:0/18:1 or 16:0/18:2. NTE therefore serves an important function in LPC homeostasis and action.

  7. Erythrodiplax ana sp. nov. (Odonata: Libellulidae) from Brazilian palm swamps.

    PubMed

    Guillermo-Ferreira, Rhainer; Vilela, Diogo S; Del-Claro, Kleber; Bispo, Pitágoras C

    2016-01-01

    Erythrodiplax ana sp. nov. (male holotype, six male and three female paratypes), collected in Vereda wetlands (a unique Neotropical savanna environment) in Uberlândia (Minas Gerais) and Chapada dos Guimarães (Mato Grosso), Brazil, is described and illustrated. The new species fits in Borror's Basalis Group, and can be distinguished from other species by the combination of the following traits: blue pruinosity dorsally on thorax and third to eighth abdominal segments; sides of the thorax olive-green; face ivory or olive-green; wings hyaline with a small apical brown spot on all four wings, well defined in females; male genitalia with sclerotized erectile posterior lobe and inflatable sac-like median process. Last instar larvae were reared in the laboratory, resulting in the description of the larva. We also followed this population for 13 months and present resulting biological notes and comments on ontogenetic color change in males, as well as longevity. PMID:27615887

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

    NASA Astrophysics Data System (ADS)

    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.

  9. Isoenzyme status and genetic variability of serum esterases in the lesser snow goose, Anser caerulescens caerulescens.

    PubMed

    Bargiello, T A; Grossfield, J; Steele, R W; Cooke, F

    1977-08-01

    A maximum of 22 bands comprising four esterase subgroups--acetylesterase, carboxylesterase, cholinesterase, and acetylcholinesterase--were detected following electrophoresis of lesser snow goose sera on polyacrylamide gels. A minimum of seven structural genes was surmised to be involved in the biosynthesis of these enzymes following physiochemical characterizations. The genetic variability of these loci was calculated to be 1.25% average heterozygosity, while 14.3% of the loci were polymorphic. These estimates of genetic variability were substantially lower than those reported for other vertebrate species. The low degree of genetic variability found in snow goose serum esterases coupled with the extensive protein multiplicity observed may possibly reflect an adaptive strategy based on "biochemical plasticity" rather than genic heterozygosity for this species. The nature of evolutionary forces acting upon multiple enzyme systems such as esterases is discussed. The concept of "conditional neutrality" is introduced and defined within this context. PMID:921742

  10. Extraction and purification of wheat-esterase using aqueous two-phase systems of ionic liquid and salt.

    PubMed

    Jiang, Bin; Feng, Zhibiao; Liu, Chunhong; Xu, Yingcao; Li, Dongmei; Ji, Guo

    2015-05-01

    To explore a new and simple rapid extraction and purification technique for wheat-esterase, an ionic liquids (ILs)-based aqueous two-phase system (ATPS) was developed for the purification of wheat-esterase from wheat extracts. Effects of various process parameters such as the concentrations of [Bmim]BF4, the types and concentrations of phase-forming salt, the system pH and the temperature on partitioning of wheat-esterase were evaluated. The obtained data indicated that wheat-esterase was preferentially partitioned into the ILs-rich phase and the ATPS composed of 20 % [Bmim]BF4 (w/w) and 25 % (w/w) NaH2PO4(pH = 4.8) showed good selectivity on wheat-esterase. Under the optimum conditions, wheat-esterase was purified with an acceptable yield (88.93 %), but produced wheat-esterase was 4.23 times as pure. It was obvious that temperature shows little influence on the purification between 10 and 50 °C. Sephadex G-150FF revealed that the band intensity of contaminating proteins in ATPS fraction almost disappeared. Therefore, ILs-based ATPS was an effective method for partitioning and recovery of wheat-esterase from wheat crude extracts. PMID:25892786

  11. The ars detoxification system is advantageous but not required for As(V) respiration by the genetically tractable Shewanella species strain ANA-3.

    PubMed

    Saltikov, Chad W; Cifuentes, Ana; Venkateswaran, Kasthuri; Newman, Dianne K

    2003-05-01

    Arsenate [As(V); HAsO(4)(2-)] respiration by bacteria is poorly understood at the molecular level largely due to a paucity of genetically tractable organisms with this metabolic capability. We report here the isolation of a new As(V)-respiring strain (ANA-3) that is phylogenetically related to members of the genus Shewanella and that also provides a useful model system with which to explore the molecular basis of As(V) respiration. This gram-negative strain stoichiometrically couples the oxidation of lactate to acetate with the reduction of As(V) to arsenite [As(III); HAsO(2)]. The generation time and lactate molar growth yield (Y(lactate)) are 2.8 h and 10.0 g of cells mol of lactate(-1), respectively, when it is grown anaerobically on lactate and As(V). ANA-3 uses a wide variety of terminal electron acceptors, including oxygen, soluble ferric iron, oxides of iron and manganese, nitrate, fumarate, the humic acid functional analog 2,6-anthraquinone disulfonate, and thiosulfate. ANA-3 also reduces As(V) to As(III) in the presence of oxygen and resists high concentrations of As(III) (up to 10 mM) when grown under either aerobic or anaerobic conditions. ANA-3 possesses an ars operon (arsDABC) that allows it to resist high levels of As(III); this operon also confers resistance to the As-sensitive strains Shewanella oneidensis MR-1 and Escherichia coli AW3110. When the gene encoding the As(III) efflux pump, arsB, is inactivated in ANA-3 by a polar mutation that also eliminates the expression of arsC, which encodes an As(V) reductase, the resulting As(III)-sensitive strain still respires As(V); however, the generation time and the Y(lactate) value are two- and threefold lower, respectively, than those of the wild type. These results suggest that ArsB and ArsC may be useful for As(V)-respiring bacteria in environments where As concentrations are high, but that neither is required for respiration.

  12. Quantum magnetic deflagration in acetate.

    PubMed

    Hernández-Mínguez, A; Hernandez, J M; Macià, F; García-Santiago, A; Tejada, J; Santos, P V

    2005-11-18

    We report controlled ignition of magnetization reversal avalanches by surface acoustic waves in a single crystal of acetate. Our data show that the speed of the avalanche exhibits maxima on the magnetic field at the tunneling resonances of Mn(12). Combined with the evidence of magnetic deflagration in Mn(12) acetate, this suggests a novel physical phenomenon: deflagration assisted by quantum tunneling. PMID:16384178

  13. Quantum magnetic deflagration in acetate.

    PubMed

    Hernández-Mínguez, A; Hernandez, J M; Macià, F; García-Santiago, A; Tejada, J; Santos, P V

    2005-11-18

    We report controlled ignition of magnetization reversal avalanches by surface acoustic waves in a single crystal of acetate. Our data show that the speed of the avalanche exhibits maxima on the magnetic field at the tunneling resonances of Mn(12). Combined with the evidence of magnetic deflagration in Mn(12) acetate, this suggests a novel physical phenomenon: deflagration assisted by quantum tunneling.

  14. Enzymatic production of ethanol from cellulose using soluble cellulose acetate as an intermediate.

    PubMed

    Downing, K M; Ho, C S; Zabriskie, D W

    1987-06-01

    A two-stage process for the enzymatic conversion of cellulose to ethanol is proposed as an alternative to currently incomplete and relatively slow enzymatic conversion processes employing natural insoluble cellulose. This alternative approach is designed to promote faster and more complete conversion of cellulose to fermentable sugars through the use of a homogeneous enzymatic hydrolysis reaction. Cellulose is chemically dissolved in the first stage to form water-soluble cellulose acetate (WSCA). The WSCA is then converted to ethanol in a simultaneous saccharification-fermentation with Pestal-otiopsis westerdijkii enzymes (containing cellulolytic and acetyl esterase components) and yeast.Water-soluble cellulose acetate was successfully prepared from purified wood cellulose (Solka Floe) and chemical reagents. Enzyme pretreatment of WSCAto form metabolizable sugars was a necessary step in achieving practical conversion of WSCA to ethanol using yeast. The results showed that WSCA has a low enzyme requirement and a high convertibility to reducing sugars with enzymes from P. westerdijkii fungus. Pestalotiopsis westerdijkii enzymes were found to be superior to enzymes from Trichoderma viride in producing metabolizable glucose from WSCA. The yeast utilized 55-70% of the hydrolyzate sugars that were produced by P. westerrlijkii enzymes on WSCA and produced ethanol. The acetate that was liberated into solution by the action of acetyl esterase enzymes on WSCA was found to have a stimulatory effect on ethanol production in yeast. This is an important feature that can be used to advantage in manipulating the conversion to maximize the production of ethanol. Hence, the simultaneous saccharification-fermentation of WSCA to ethanol using P. westerdijkii enzymes and yeast has features that are highly desirable for developing an economical cellulose conversion process.

  15. [Erythropoietin-forming and esterase activity of rat kidney subcellular fractions during stimulation of erythropoiesis].

    PubMed

    Novikov, N M; Voronkov, S F; Voloshchenko, L G; Mikhaĭlova, S N

    1977-01-01

    Stimulation of erythropoiesis in rats (hemolytic-phenylhydrazine and acute posthemorrhagic anemia, effect of hypoxic hypoxia) was accompanied by an increased erythropoietine-formating activity in kidney microsomes and light mitochondria. The phenomenon correlated with an increased esterase activity in hypotonic supernatant of kidney homogenate mainly due to the enzymatic fraction, corresponding to alpha2-globulin by its mobility. Histochemical examination of kidney showed that the most distinct alterations in the esterase activity were observed in epithelial cells of nephron proximal part and in capillary endothelium.

  16. The ANA-reflex test as a model for improving clinical appropriateness in autoimmune diagnostics.

    PubMed

    Tonutti, Elio; Bizzaro, Nicola; Morozzi, Gabriella; Radice, Antonella; Cinquanta, Luigi; Villalta, Danilo; Tozzoli, Renato; Tampoia, Marilina; Porcelli, Brunetta; Fabris, Martina; Brusca, Ignazio; Alessio, Maria Grazia; Barberio, Giuseppina; Sorrentino, Maria Concetta; Antico, Antonio; Bassetti, Danila; Fontana, Desré Ethel; Imbastaro, Tiziana; Visentini, Daniela; Pesce, Giampaola; Bagnasco, Marcello

    2016-12-01

    Reflex tests are widely used in clinical laboratories, for example, to diagnose thyroid disorders or in the follow-up of prostate cancer. Reflex tests for antinuclear antibodies (ANA) have recently gained attention as a way to improve appropriateness in the immunological diagnosis of autoimmune rheumatic diseases and avoid waste of resources. However, the ANA-reflex test is not as simple as other consolidated reflex tests (the TSH-reflex tests or the PSA-reflex tests) because of the intrinsic complexity of the ANA test performed by the indirect immunofluorescence method on cellular substrates. The wide heterogeneity of the ANA patterns, which need correct interpretation, and the subsequent choice of the most appropriate confirmatory test (ANA subserology), which depend on the pattern feature and on clinical information, hinder any informatics automation, and require the pathologist's intervention. In this review, the Study Group on Autoimmune Diseases of the Italian Society of Clinical Pathology and Laboratory Medicine provides some indications on the configuration of the ANA-reflex test, using two different approaches depending on whether clinical information is available or not. We further give some suggestions on how to report results of the ANA-reflex test. PMID:27423928

  17. The concordance of serial ANA tests in an Australian tertiary hospital pathology laboratory.

    PubMed

    Lee, Adrian Y S; Hudspeth, Andrew R; Adelstein, Stephen

    2016-10-01

    The antinuclear antibody (ANA) tests are some of the more frequently requested tests for the diagnosis of autoimmunity. Although they are used primarily as diagnostic blood tests, multiple requests on the same patient continue to be encountered in the laboratory. This retrospective analysis of serial ANA testing at one pathology laboratory in Australia is the first study that examines the statistical concordance and possible implications of this on clinical practice. High-titred ANA have quite good repeatability for titre and pattern, and low-titred ANA, which can be non-specific, have poor repeatability. Staining patterns are, in general, almost random in nature on serial tests when compared to the first-obtained ANA pattern for each patient. This study confirms that there is little benefit in serial ANA testing, and only if there is a clear change in the patient's clinical picture would repeat of an initial low-titred ANA be useful. The findings reinforce the need for pathology stewardship to minimise costs, wasted resources and unnecessary referrals. PMID:27600602

  18. The ANA-reflex test as a model for improving clinical appropriateness in autoimmune diagnostics.

    PubMed

    Tonutti, Elio; Bizzaro, Nicola; Morozzi, Gabriella; Radice, Antonella; Cinquanta, Luigi; Villalta, Danilo; Tozzoli, Renato; Tampoia, Marilina; Porcelli, Brunetta; Fabris, Martina; Brusca, Ignazio; Alessio, Maria Grazia; Barberio, Giuseppina; Sorrentino, Maria Concetta; Antico, Antonio; Bassetti, Danila; Fontana, Desré Ethel; Imbastaro, Tiziana; Visentini, Daniela; Pesce, Giampaola; Bagnasco, Marcello

    2016-12-01

    Reflex tests are widely used in clinical laboratories, for example, to diagnose thyroid disorders or in the follow-up of prostate cancer. Reflex tests for antinuclear antibodies (ANA) have recently gained attention as a way to improve appropriateness in the immunological diagnosis of autoimmune rheumatic diseases and avoid waste of resources. However, the ANA-reflex test is not as simple as other consolidated reflex tests (the TSH-reflex tests or the PSA-reflex tests) because of the intrinsic complexity of the ANA test performed by the indirect immunofluorescence method on cellular substrates. The wide heterogeneity of the ANA patterns, which need correct interpretation, and the subsequent choice of the most appropriate confirmatory test (ANA subserology), which depend on the pattern feature and on clinical information, hinder any informatics automation, and require the pathologist's intervention. In this review, the Study Group on Autoimmune Diseases of the Italian Society of Clinical Pathology and Laboratory Medicine provides some indications on the configuration of the ANA-reflex test, using two different approaches depending on whether clinical information is available or not. We further give some suggestions on how to report results of the ANA-reflex test.

  19. Life history and ecological characteristics of the Santa Ana sucker, Catostomus santaanae

    USGS Publications Warehouse

    Saiki, Michael K.; Martin, Barbara A.; Knowles, Glen W.; Tennant, Patrick W.

    2007-01-01

    This study was conducted to document the life history and ecological characteristics of the Santa Ana sucker, Catostomus santaanae, within its native range in southern California. Electrofishing surveys were conducted at 3-month intervals from December 1998 to December 1999 at one site on the San Gabriel River and two sites on the Santa Ana River. Suckers were captured in the San Gabriel River (average, 6.6 fish/10-minutes electrofishing) and at an upstream Santa Ana River site (average, 2.3 fish/10-minutes electrofishing) but not at a downstream Santa Ana River site. Length frequency distributions indicated that at least three year classes (modal groups) of suckers were present in the San Gabriel River, whereas one or two year classes were present in the Santa Ana River. Collection of 21-30 mm standard length (SL) juveniles in June in the Santa Ana River and in September in the San Gabriel River indicated that reproduction occurred over several months. In December, Age-0 suckers averaged 36-48 mm SL in the San Gabriel River and 63-65 mm SL in the Santa Ana River, whereas Age-1 suckers averaged 86 mm SL in the San Gabriel River and 115 mm SL in the Santa Ana River. On average, suckers were in better body condition in the San Gabriel River than in the Santa Ana River. Highest abundance of suckers was associated with relativelypristine environmental conditions (especially low specific conductance) where other native fishes were also common or abundant.

  20. A cluster of at least three esterase genes in Lucilia cuprina includes malathion carboxylesterase and two other esterases implicated in resistance to organophosphates

    SciTech Connect

    Smyth, K.A. |; Russell, R.J.; Oakeshott, J.G.

    1994-12-01

    Three distinct malathion carboxylesterase (MCE) phenotypes have been identified among strains of Lucilia cuprina. The high-activity phenotype shows 1.6- and 3.3-fold more MCE specific activity than the intermediate- and low-activity phenotypes, respectively. Flies with high MCE activity are 1000-fold more resistant to malathion than flies with either low or intermediate MCE phenotypes, which are equally susceptible. High and low MCE specific activity are allelic and encoded by the Rmal gene on chromosome 4. Rmal is clustered within one map unit of two other esterase genes, Rop1 and E9, which are implicated in resistance to other organophosphate insecticides. Intermediate MCE specific activity is also inherited within the cluster, although its allelism to Rmal, Rop1, or E9 is unclear. The cluster does not contain the gene for the hemolymph esterase E4, which maps 6.1 map units from Rop1, on the other side of the bubbled wing marker. The cluster appears to be homologous to part of a tandem array of 11 esterase genes on chromosome 3R of Drosophila melanogaster. 41 refs., 4 figs., 2 tabs.

  1. The use of pro-ana blogs for online social support.

    PubMed

    Tong, Stephanie Tom; Heinemann-Lafave, Daria; Jeon, Jehoon; Kolodziej-Smith, Renata; Warshay, Nathaniel

    2013-01-01

    The aim of this research was to document the content on pro-ana blogs and to uncover how the unique socio-technical features of blogs (interactivity, self-disclosure, masspersonal communication) facilitate social support among members. A final sample of 48 pro-ana blogs provided 624 individual units for coding. Results indicate that prevalent forms of social support were emotional support, esteem support, and informational support. A new category, reciprocal self-disclosure, was also revealed to be quite frequent. Blogs are spaces where social support is sought and communicated among members of the pro-ana network. Interpretation of blog communication and implications for treatment and research are discussed.

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

    EPA Science Inventory

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

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

    PubMed

    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.

  4. Tissue-specific inhibition and recovery of esterase activities in Lumbricus terrestris experimentally exposed to chlorpyrifos.

    PubMed

    Vejares, Sandra González; Sabat, Pablo; Sanchez-Hernandez, Juan C

    2010-04-01

    Exposure and effect assessment of organophosphate (OP) pesticides generally involves the use of cholinesterase (ChE) inhibition. In earthworm, this enzyme activity is often measured in homogenates from the whole organism. Here we examine the tissue-specific response of ChE and carboxylesterase (CE) activities in Lumbricus terrestris experimentally exposed to chlorpyrifos-spiked field soils. Esterases were measured in different gut segments and in the seminal vesicles of earthworms following acute exposure (2 d) to the OP and during 35d of a recovery period. We found that inhibition of both esterase activities was dependent on the tissue. Cholinesterase activity decreased in the pharynx, crop, foregut and seminal vesicles in a concentration-dependent way, whereas CE activity (4-nitrophenyl valerate) was strongly inhibited in these tissues. Gizzard CE activity was not inhibited by the OP, even an increase of enzyme activity was evident during the recovery period. These results suggest that both esterases should be determined jointly in selected tissues of earthworms. Moreover, the high levels of gut CE activity and its inhibition and recovery dynamic following OP exposure suggest that this esterase could play an important role as an enzymatic barrier against OP uptake from the ingested contaminated soil. PMID:20045489

  5. Phylogenetic classification of Aureobasidium pullulans strains for production of feruloyl esterase

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective was to phylogenetically classify diverse strains of A. pullulans and determine their production of feruloyl esterase. Seventeen strains from the A. pullulans literature were phylogenetically classified. Phenotypic traits of color variation and endo-ß-1,4-xylanase overproduction were as...

  6. Evolution and homologous recombination of the hemagglutinin-esterase gene sequences from porcine torovirus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of the present study was to gain new insights into the evolution, homologous recombination and selection pressures imposed on the porcine torovirus (PToV), by examining changes in the hemagglutinin-esterase (HE) gene. The most recent common ancestor of PToV was estimated to have emerge...

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

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

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

    PubMed Central

    2015-01-01

    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. PMID:27335975

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

    PubMed

    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.

  10. Chaperone-like activities of {alpha}-synuclein: {alpha}-Synuclein assists enzyme activities of esterases

    SciTech Connect

    Ahn, Misun; Kim, SeungBum; Kang, Mira; Ryu, Yeonwoo . E-mail: ywryu@ajou.ac.kr; Doohun Kim, T. . E-mail: doohunkim@ajou.ac.kr

    2006-08-11

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

  11. Absence of "A"-esterase activity in the serum of a patient with Tangier disease.

    PubMed

    Mackness, M I; Peuchant, E; Dumon, M F; Walker, C H; Clerc, M

    1989-12-01

    The levels of apolipoprotein A-I, A-II and B in subjects who are homozygous or heterozygous for Tangier disease are reported and compared with the amount of "A"-esterase in the serum. The "A"-esterases hydrolyse toxic organophosphate pesticides and are currently classified by the nomenclature committee of the International Union of Biochemistry as arylesterases (EC 3.1.1.2) although recent evidence has cast doubt on this classification. The apolipoprotein data are consistent with previous data reported for a number of Tangier patients. The homozygote has a marked reduction in apo A-I and A-II levels and a 30% reduction in apo B. The heterozygotes have about a 50% reduction of apo A-I, a slight reduction in apo A-II and no change in apo B. These apolipoprotein values correspond to a marked reduction in HDL cholesterol for the homozygote and substantial reductions in the heterozygotes. The "A"-esterase activity is zero in one homozygote while heterozygotes have about 5% of the levels in control subjects. Arylesterase activity appears to be essentially normal. The data thus support previous observations that the HDL "A"-esterase activity is greatly reduced in those conditions where HDL apo A-I is markedly reduced, e.g., in "Fish-eye" Disease.

  12. Cloning, sequencing, and regulation of expression of an extracellular esterase gene from the plant pathogen Streptomyces scabies.

    PubMed Central

    Raymer, G; Willard, J M; Schottel, J L

    1990-01-01

    The gene that encodes the extracellular esterase produced by Streptomyces scabies has been cloned and sequenced. The gene was identified by hybridization to a synthetic oligonucleotide that corresponds to the amino-terminal amino acid sequence determined for the secreted form of the esterase. Nucleotide sequence analysis predicted a 345-amino-acid open reading frame, a putative ribosome-binding site, and 39 amino acids at the amino terminus of the sequence that is not found in the secreted protein. This 39-amino-acid sequence has many of the characteristics common to known signal peptides. End mapping the esterase transcript revealed a single 5' end of the mRNA located 51 nucleotides upstream from the start point for translation. Northern (RNA) hybridization analysis of the esterase message by using the cloned esterase gene as a probe indicated that the esterase mRNA is about 1,440 nucleotides in length and was detected only when the cells were grown in the presence of zinc. These results suggest that the level of esterase mRNA detected in the cells is regulated by zinc. Images PMID:2254271

  13. Role of endo-1,4-beta-glucanases from neisseria sicca SB in synergistic degradation of cellulose acetate.

    PubMed

    Moriyoshi, Kunihiko; Ohmoto, Takashi; Ohe, Tatsuhiko; Sakai, Kiyofumi

    2003-02-01

    An enzyme hydrolyzing beta-1,4 bonds in cellulose acetate was purified 10.5-fold to electrophoretic homogeneity from a culture supernatant of Neisseria sicca SB, which assimilate cellulose acetate as the sole carbon and energy source. The enzyme was an endo-1,4-beta-glucanase, to judge from the substrate specificity and hydrolysis products of cellooligosaccharides, we named it endo-1,4-beta-glucanase I (EG I). Its molecular mass was 50 kDa, 9 kDa larger than EG II from this strain, and its isoelectric point was 5.0. Results of N-terminal and inner-peptide sequences of both enzymes, and a similarity search, suggested that EG I contained a carbohydrate-binding module at the N-terminus and that EG II lacked this module. The pH and temperature optima of EG I were 5.0-6.0 and 45 degrees C. It hydrolyzed water-soluble cellulose acetate (degree of substitution, 0.88) and carboxymethyl cellulose. The Km and Vmax for these compounds were 0.296% and 1.29 micromol min(-1) mg(-1), and 0.448% and 13.6 micromol min(-1) mg(-1), respectively. Both glucanases and cellulose acetate esterase from this strain degraded water-insoluble cellulose acetate synergistically.

  14. Gel-electrophoretic identification of hen brain neurotoxic esterase, labelled with tritiated di-isopropyl phosphorofluoridate.

    PubMed Central

    Williams, D G; Johnson, M K

    1981-01-01

    The particulate fraction from hen brain was labelled with [3H]di-isopropyl phosphorofluoridate (DiPF) and separated by polyacrylamide-gel electrophoresis. Four radioactive protein bands (1--4) of molecular weights 155000, 92000, 60000, and 30000 were resolved. Most of the labelling of bands 2, 3 and 4 was inhibited by preincubation with Paraoxon. The residue in band 4 was sensitive to pH 5.2. Successive treatments with Paraoxon and pH 5.2 resulted in the abolition of bands 3 and 4. Bands 1 and 2 contained one and two polypeptides respectively, whose labelling was sensitive to Mipafox, but one, in band 2, was sensitive to higher concentrations of Paraoxon. The concentrations of the other two polypeptides were 6.7 and 1.95 pmol of DiPF bound/g of brain in bands 1 and 2 respectively. Both were as sensitive to Mipafox as neurotoxic esterase and were also sensitive to phenyl benzylcarbamate. 4-Nitrophenyl di-n-pentylphosphinate given in vivo inhibited neurotoxic esterase and the labelling of the band-1 polypeptide by 82% and 84% respectively, but inhibited the labelling of the band 2 polypeptide by 51%. The phosphinate in vitro produced 98% inhibition of the labelling of the band-1 polypeptide, with only 26% inhibition of the band-2 polypeptide, under conditions sufficient to inhibit neurotoxic esterase totally. Both neurotoxic esterase and the band-1 polypeptide were found in the forebrain at 1.74-fold their concentration in the rest of the brain, whereas the band-2 polypeptide was uniformly distributed. The evidence indicates that the Mipafox-sensitive polypeptide in band 1 is the [3H]DiPF-labelled active-site subunit of neurotoxic esterase. The catalytic-centre activity of the enzyme for phenyl valerate hydrolysis was found to be 2.6 x 10(5) min-1. PMID:7340807

  15. Microbial degradation of poly-b-esters: A mechanistic study, cellulose acetate biodegradability. Final report, 1 May 1990-31 July 1993

    SciTech Connect

    Gross, R.A.

    1993-08-30

    In this Final Report, work carried out under ARO grant C-DAAL03-G-0111 is described. The investigations performed include the following: (1) isolation, purification and characterization of a poly(3-hydroxybutyrate) depolymerase enzyme from Penicillium funiculosum, (2) determination that the depolymerase is a serine esterase, (3) study of the effect of polymer stereochemistry and crystalline order in a semi-crystalline polymer film substrate on enzyme specificity and activity, (3) isolation, purification and characterization of cellulose acetate degrading microorganisms and (4) determination of the biodegradability of cellulose acetate with degrees of substitution up to 2.5 under aerobic thermophilic conditions. Poly(3-hydroxybutyrate) biodegradation, Poly(3-hydroxybutyrate) depolymerase enzyme, Depolymerase from Penicillium funiculosum, Cellulose acetate degrading microorganisms, Composting polymer biodegradable.

  16. Updating movement estimates for American black ducks (Anas rubripes)

    PubMed Central

    McGowan, Conor P.; Devers, Patrick K.

    2016-01-01

    Understanding migratory connectivity for species of concern is of great importance if we are to implement management aimed at conserving them. New methods are improving our understanding of migration; however, banding (ringing) data is by far the most widely available and accessible movement data for researchers. Here, we use band recovery data for American black ducks (Anas rubripes) from 1951–2011 and analyze their movement among seven management regions using a hierarchical Bayesian framework. We showed that black ducks generally exhibit flyway fidelity, and that many black ducks, regardless of breeding region, stopover or overwinter on the Atlantic coast of the United States. We also show that a non-trivial portion of the continental black duck population either does not move at all or moves to the north during the fall migration (they typically move to the south). The results of this analysis will be used in a projection modeling context to evaluate how habitat or harvest management actions in one region would propagate throughout the continental population of black ducks. This analysis may provide a guide for future research and help inform management efforts for black ducks as well as other migratory species. PMID:26989624

  17. Migration strategy affects avian influenza dynamics in mallards (Anas platyrhynchos).

    PubMed

    Hill, Nichola J; Takekawa, John Y; Ackerman, Joshua T; Hobson, Keith A; Herring, Garth; Cardona, Carol J; Runstadler, Jonathan A; Boyce, Walter M

    2012-12-01

    Studies of pathogen transmission typically overlook that wildlife hosts can include both migrant and resident populations when attempting to model circulation. Through the application of stable isotopes in flight feathers, we estimated the migration strategy of mallards (Anas platyrhynchos) occurring on California wintering grounds. Our study demonstrates that mallards- a principal host of avian influenza virus (AIV) in nature, contribute differently to virus gene flow depending on migration strategy. No difference in AIV prevalence was detected between resident (9.6%), intermediate-distance (9.6%) and long-distance migrants (7.4%). Viral diversity among the three groups was also comparable, possibly owing to viral pool mixing when birds converge at wetlands during winter. However, migrants and residents contributed differently to the virus gene pool at wintering wetlands. Migrants introduced virus from northern breeding grounds (Alaska and the NW Pacific Rim) into the wintering population, facilitating gene flow at continental scales, but circulation of imported virus appeared to be limited. In contrast, resident mallards acted as AIV reservoirs facilitating year-round circulation of limited subtypes (i.e. H5N2) at lower latitudes. This study supports a model of virus exchange in temperate regions driven by the convergence of wild birds with separate geographic origins and exposure histories. PMID:22971007

  18. Updating movement estimates for American black ducks (Anas rubripes).

    PubMed

    Robinson, Orin J; McGowan, Conor P; Devers, Patrick K

    2016-01-01

    Understanding migratory connectivity for species of concern is of great importance if we are to implement management aimed at conserving them. New methods are improving our understanding of migration; however, banding (ringing) data is by far the most widely available and accessible movement data for researchers. Here, we use band recovery data for American black ducks (Anas rubripes) from 1951-2011 and analyze their movement among seven management regions using a hierarchical Bayesian framework. We showed that black ducks generally exhibit flyway fidelity, and that many black ducks, regardless of breeding region, stopover or overwinter on the Atlantic coast of the United States. We also show that a non-trivial portion of the continental black duck population either does not move at all or moves to the north during the fall migration (they typically move to the south). The results of this analysis will be used in a projection modeling context to evaluate how habitat or harvest management actions in one region would propagate throughout the continental population of black ducks. This analysis may provide a guide for future research and help inform management efforts for black ducks as well as other migratory species. PMID:26989624

  19. Migration strategy affects avian influenza dynamics in mallards (Anas platyrhynchos).

    USGS Publications Warehouse

    Takekawa, John Y.; Hill, Nichola J.; Ackerman, Joshua T.; Herring, Garth; Hobson, Keith; Cardona, Carol J.; Runstadler, Jonathan; Boyce, Walter M.

    2012-01-01

    Studies of pathogen transmission typically overlook that wildlife hosts can include both migrant and resident populations when attempting to model circulation. Through the application of stable isotopes in flight feathers, we estimated the migration strategy of mallards (Anas platyrhynchos) occurring on California wintering grounds. Our study demonstrates that mallards- a principal host of avian influenza virus (AIV) in nature, contribute differently to virus gene flow depending on migration strategy. No difference in AIV prevalence was detected between resident (9.6%), intermediate-distance (9.6%) and long-distance migrants (7.4%). Viral diversity among the three groups was also comparable, possibly owing to viral pool mixing when birds converge at wetlands during winter. However, migrants and residents contributed differently to the virus gene pool at wintering wetlands. Migrants introduced virus from northern breeding grounds (Alaska and the NW Pacific Rim) into the wintering population, facilitating gene flow at continental scales, but circulation of imported virus appeared to be limited. In contrast, resident mallards acted as AIV reservoirs facilitating year-round circulation of limited subtypes (i.e. H5N2) at lower latitudes. This study supports a model of virus exchange in temperate regions driven by the convergence of wild birds with separate geographic origins and exposure histories.

  20. Migration strategy affects avian influenza dynamics in mallards (Anas platyrhynchos).

    PubMed

    Hill, Nichola J; Takekawa, John Y; Ackerman, Joshua T; Hobson, Keith A; Herring, Garth; Cardona, Carol J; Runstadler, Jonathan A; Boyce, Walter M

    2012-12-01

    Studies of pathogen transmission typically overlook that wildlife hosts can include both migrant and resident populations when attempting to model circulation. Through the application of stable isotopes in flight feathers, we estimated the migration strategy of mallards (Anas platyrhynchos) occurring on California wintering grounds. Our study demonstrates that mallards- a principal host of avian influenza virus (AIV) in nature, contribute differently to virus gene flow depending on migration strategy. No difference in AIV prevalence was detected between resident (9.6%), intermediate-distance (9.6%) and long-distance migrants (7.4%). Viral diversity among the three groups was also comparable, possibly owing to viral pool mixing when birds converge at wetlands during winter. However, migrants and residents contributed differently to the virus gene pool at wintering wetlands. Migrants introduced virus from northern breeding grounds (Alaska and the NW Pacific Rim) into the wintering population, facilitating gene flow at continental scales, but circulation of imported virus appeared to be limited. In contrast, resident mallards acted as AIV reservoirs facilitating year-round circulation of limited subtypes (i.e. H5N2) at lower latitudes. This study supports a model of virus exchange in temperate regions driven by the convergence of wild birds with separate geographic origins and exposure histories.

  1. Belgian recommendations on ANA, anti-dsDNA and anti-ENA antibody testing.

    PubMed

    Van Blerk, M; Bossuyt, X; Humbel, R; Mewis, A; Servais, G; Tomasi, J P; Van Campenhout, C; Van Hoovels, L; Vercammen, M; Damoiseaux, J; Coucke, W; Van de Walle, P

    2014-04-01

    Autoantibodies to nuclear antigens, i.e. antinuclear antibodies (ANA), antibodies to double-stranded DNA (dsDNA) and extractable nuclear antigens (ENA), are useful as diagnostic markers for a variety of autoimmune diseases. In March 2010, the Belgian national External Quality Assessment Scheme sent a questionnaire on ANA, anti-dsDNA and anti-ENA antibody testing designed by the Dutch EASI (European Autoimmunity Standardization Initiative) team, to all clinical laboratories performing ANA testing. Virtually all laboratories completed the questionnaire (97·7%, 127/130). This paper discusses the results of this questionnaire and provides valuable information on the state-of-the-art of ANA, anti-dsDNA and anti-ENA antibody testing as practiced in the Belgian laboratories. In addition, this work presents practical recommendations developed by the members of the advisory board of the scheme as a result of the outcome of this study.

  2. DEVELOPMENT OF A PHYSIOLOGICALLY BASED PHARMACOKINETIC MODEL FOR PROPYLENE GLYCOL MONOMETHYL ETHER AND ITS ACETATE IN RATS AND HUMANS

    SciTech Connect

    Corley, Rick A.; Gies, Richard A.; Wu, Hong; Weitz, Karl K.

    2005-03-05

    Propylene glycol monomethyl ether (PM), along with its acetate, is the most widely used of the propylene glycol ether family of solvents. The most common toxic effects of PM observed in animal studies include sedation, very slight alpha2u globulin-mediated nephropathy (male rats only) and hepatomegally at high exposures (typically >1000 ppm). Sedation in animal studies usually resolves within a few exposures to 3000 ppm (the highest concentration used in subchronic and chronic inhalation studies) due to the induction of metabolizing enzymes. Data from a variety of pharmacokinetic and mechanistic studies have been incorporated into a PBPK model for PM and its acetate in rats and mice. Published controlled exposure and workplace biomonitoring studies have also been included for comparisons of the internal dosimetry of PM and its acetate between laboratory animals and humans. PM acetate is rapidly hydrolyzed to PM, which is further metabolized to either glucuronide or sulphate conjugates (minor pathways) or propylene glycol (major pathway). In vitro half-lives for PM acetate range from 14-36 min depending upon the tissue and species. In vivo half-lives are considerably faster, reflecting the total contributions of esterases in the blood and tissues of the body, and are on the order of just a few minutes. Thus, very little PM acetate is found in vivo and, other than potential portal of entry irritation, the toxicity of PM acetate is related to PM. Regardless of the source for PM (either PM or its acetate), rats were predicted to have a higher Cmax and AUC for PM in blood than humans, especially at concentrations greater than the current ACGIH TLV of 100 ppm. This would indicate that the major systemic effects of PM would be expected to be less severe in humans than rats at comparable inhalation exposures.

  3. Biochemical identification of the mallard, Anas platyrhynchos, and black duck, A. rubripes

    USGS Publications Warehouse

    Morgan, R.P.; Noe, L.A.; Henny, C.J.

    1976-01-01

    1. 1. Eleven tissue systems from mallards and black ducks were examined for soluble proteins, lactate dehydrogenases and non-specific esterases through discontinuous polyacrylamide techniques. 2. 2. Biochemical relationships between the black duck and mallard are extremely similar. 3. 3. Hemoglobins and lactate dehydrogenase appear to be common in electrophoretic mobility between the two species. 4. 4. Approximately 89% of the soluble proteins and 58% of the non-specific esterases are common among the two species, indicating both biochemical similarity at the genus level and species-specificity.

  4. Chemotactic activity from rabbit peritoneal neutrophils. Lack of identity with N-acetyl-DL-phenylalanine beta-napthyl esterase.

    PubMed

    Tsung, P K; Showell, H J; Kegeles, S W; Becker, E L

    1976-08-12

    The chemotactic and N-acetyl-DL-phenylalanine beta-naphthyl esterase activities of rabbit peritoneal neutrophils are separable from each other by both DEAE cellulose and Sephadex G-100 column chromatography. Partially purified esterase obtained from DEAE-cellulose chromatography had molecular weight of 70 000. However, the partially purified fraction contained chemotactic activities with major activity in molecular weight of 28000 and minor activities in the molecular weights of 45000, 21900, 14500 and 10500. Esterase activity is inhibited by 10(-7) M p-nitrophenylethyl-5-chloropentylphosphonate but chemotactic activity is not.

  5. Use of ammonium nitrate-alcohol (ANA) for ballistic range propellant

    NASA Astrophysics Data System (ADS)

    Sasoh, A.; Ogawa, T.; Takayama, K.

    Mixtures of ammonium-nitrate and alcohol (ANA) are usable as range propellants in ballistics. The molecular mass and specific energy are controlled by altering the type of alcohol used and by varying their molar ratio. This is confirmed by experiments carried out in a 25-mm-bore single-stage gun with ANA using smokeless powder as igniter. It is found that an important parameter determining propellant performance is the quantity of the smokeless powder used for igniting the mixture.

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

    PubMed Central

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

    2012-01-01

    Soft rot (type II) fungi belonging to the family Xylariaceae are known to substantially degrade hardwood by means of their poorly understood lignocellulolytic system, which comprises various hydrolases, including feruloyl esterases and laccase. In the present study, several members of the Xylariaceae were found to exhibit high feruloyl esterase activity during growth on lignocellulosic materials such as wheat straw (up to 1,675 mU g−1) or beech wood (up to 80 mU g−1). Following the ester-cleaving activity toward methyl ferulate, a hydrolase of Xylaria polymorpha was produced in solid-state culture on wheat straw and purified by different steps of anion-exchange and size-exclusion chromatography to apparent homogeneity (specific activity, 2.2 U mg−1). The peptide sequence of the purified protein deduced from the gene sequence and verified by de novo peptide sequencing shows high similarity to putative α-l-rhamnosidase sequences belonging to the glycoside hydrolase family 78 (GH78; classified under EC 3.2.1.40). The purified enzyme (98 kDa by SDS-PAGE, 103 kDa by size-exclusion chromatography; pI 3.7) converted diverse glycosides (e.g., α-l-rhamnopyranoside and α-l-arabinofuranoside) but also natural and synthetic esters (e.g., chlorogenic acid, hydroxycinnamic acid glycoside esters, veratric acid esters, or p-nitrophenyl acetate) and released free hydroxycinnamic acids (ferulic and coumaric acid) from arabinoxylan and milled wheat straw. These catalytic properties strongly suggest that X. polymorpha GH78 is a multifunctional enzyme. It is the first fungal enzyme that combines glycosyl hydrolase with esterase activities and may help this soft rot fungus to degrade lignocelluloses. PMID:22544251

  7. Report on the second International Consensus on ANA Pattern (ICAP) workshop in Dresden 2015.

    PubMed

    Chan, E K L; Damoiseaux, J; de Melo Cruvinel, W; Carballo, O G; Conrad, K; Francescantonio, P L C; Fritzler, M J; Garcia-De La Torre, I; Herold, M; Mimori, T; Satoh, M; von Mühlen, C A; Andrade, L E C

    2016-07-01

    The second meeting for the International Consensus on Antinuclear antibody (ANA) Pattern (ICAP) was held on 22 September 2015, one day prior to the opening of the 12th Dresden Symposium on Autoantibodies in Dresden, Germany. The ultimate goal of ICAP is to promote harmonization and understanding of autoantibody nomenclature, and thereby optimizing ANA usage in patient care. The newly developed ICAP website www.ANApatterns.org was introduced to the more than 50 participants. This was followed by several presentations and discussions focusing on key issues including the two-tier classification of ANA patterns into competent-level versus expert-level, the consideration of how to report composite versus mixed ANA patterns, and the necessity for developing a consensus on how ANA results should be reported. The need to establish on-line training modules to help users gain competency in identifying ANA patterns was discussed as a future addition to the website. To advance the ICAP goal of promoting wider international participation, it was agreed that there should be a consolidated plan to translate consensus documents into other languages by recruiting help from members of the respective communities. PMID:27252255

  8. 21 CFR 173.228 - Ethyl acetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... the specifications of the Food Chemicals Codex, 1 (Ethyl Acetate; p. 372, 3d Ed., 1981), which are... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Ethyl acetate. 173.228 Section 173.228 Food and..., Lubricants, Release Agents and Related Substances § 173.228 Ethyl acetate. Ethyl acetate (CAS Reg. No....

  9. 21 CFR 173.228 - Ethyl acetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... the specifications of the Food Chemicals Codex, 1 (Ethyl Acetate; p. 372, 3d Ed., 1981), which are... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Ethyl acetate. 173.228 Section 173.228 Food and..., Lubricants, Release Agents and Related Substances § 173.228 Ethyl acetate. Ethyl acetate (CAS Reg. No....

  10. 21 CFR 73.2396 - Lead acetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 1 2014-04-01 2014-04-01 false Lead acetate. 73.2396 Section 73.2396 Food and... ADDITIVES EXEMPT FROM CERTIFICATION Cosmetics § 73.2396 Lead acetate. (a) Identity. The color additive lead acetate is the trihydrate of lead (2+) salt of acetic acid. The color additive has the chemical formula...

  11. 21 CFR 73.2396 - Lead acetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 1 2012-04-01 2012-04-01 false Lead acetate. 73.2396 Section 73.2396 Food and... ADDITIVES EXEMPT FROM CERTIFICATION Cosmetics § 73.2396 Lead acetate. (a) Identity. The color additive lead acetate is the trihydrate of lead (2+) salt of acetic acid. The color additive has the chemical formula...

  12. 21 CFR 73.2396 - Lead acetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 1 2011-04-01 2011-04-01 false Lead acetate. 73.2396 Section 73.2396 Food and... ADDITIVES EXEMPT FROM CERTIFICATION Cosmetics § 73.2396 Lead acetate. (a) Identity. The color additive lead acetate is the trihydrate of lead (2+) salt of acetic acid. The color additive has the chemical formula...

  13. 21 CFR 73.2396 - Lead acetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 1 2013-04-01 2013-04-01 false Lead acetate. 73.2396 Section 73.2396 Food and... ADDITIVES EXEMPT FROM CERTIFICATION Cosmetics § 73.2396 Lead acetate. (a) Identity. The color additive lead acetate is the trihydrate of lead (2+) salt of acetic acid. The color additive has the chemical formula...

  14. 21 CFR 73.2396 - Lead acetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 1 2010-04-01 2010-04-01 false Lead acetate. 73.2396 Section 73.2396 Food and... ADDITIVES EXEMPT FROM CERTIFICATION Cosmetics § 73.2396 Lead acetate. (a) Identity. The color additive lead acetate is the trihydrate of lead (2+) salt of acetic acid. The color additive has the chemical formula...

  15. Mortality in the endangered Laysan teal, Anas laysanensis: conservation implications

    USGS Publications Warehouse

    Reynolds, M.H.; Work, T.M.

    2005-01-01

    The Laysan Teal Anas laysanensis is an endangered anatid of the Hawaiian Islands, currently restricted to an emergent atoll, Laysan Island. Laysan Island lacks terrestrial mammalian predators, which permits the examination of mortality rates and causes without the anthropogenic effects of introduced predators. Mass and morophometrics were measured during the colour-marking of 297 Laysan Teal between 1998 and 2001. Intensive mark-resighting and recovery methods were used to estimate adult and juvenile mortality. One hundred and nineteen carcasses were collected on Laysan between 1998 and 2003, and systematic gross and microscopic examinations were undertaken on 63 of these. Causes of mortality were categorised as trauma, emaciation, miscellaneous or undetermined. Annual adult mortality rates were low, 0.05-0.10 (s.e. <0.01), but duckling mortality was much higher, varying from approximately 0.7-0.9 during 1998-2000 and 2003. Body condition of both sexes deteriorates during the breeding season, and most adult mortality (88%) occurred during or post-breeding (May-October). Cause of mortality was determined via necropsy in 22 ducks. Of three adults, one died from bacterial infections, one was egg bound, and one died from botulism concomitant with nematode infestation. Fourteen ducklings died from acute trauma, four from emaciation sometimes associated with nematode infection, and one from bacterial pneumonia Trauma is a significant factor in Laysan duckling mortalities, and elucidating the cause of and preventing such trauma may allow for management measures to enhance duckling survivability. High duckling mortality rates and emaciation also indicate that habitat on Laysan Island may have limited capacity to support broods.

  16. Mortality in the endangered Laysan Teal Anas laysanensis: Conservation implications

    USGS Publications Warehouse

    Reynolds, M.H.; Work, T.M.

    2005-01-01

    The Laysan Teal Anas laysanensis is an endangered anatid of the Hawaiian Islands, currently restricted to an emergent atoll, Laysan Island. Laysan Island lacks terrestrial mammalian predators, which permits the examination of mortality rates and causes without the anthropogenic effects of introduced predators. Mass and morphometrics were measured during the colour-marking of 297 Laysan Teal between 1998 and 2001. Intensive mark-resighting and recovery methods were used to estimate adult and juvenile mortality. One hundred and nineteen carcasses were collected on Laysan between 1998 and 2003, and systematic gross and microscopic examinations were undertaken on 63 of these. Causes of mortality were categorised as trauma, emaciation, miscellaneous or undetermined. Annual adult mortality rates were low, 0.05-0.10 (s.e. < 0.01), but duckling mortality was much higher, varying from approximately 0.7-0.9 during 1998-2000 and 2003. Body condition of both sexes deteriorates during the breeding season, and most adult mortality (88%) occurred during or post-breeding (May-October). Cause of mortality was determined via necropsy in 22 ducks. Of three adults, one died from bacterial infections, one was egg bound, and one died from botulism concomitant with nematode infestation. Fourteen ducklings died from acute trauma, four from emaciation sometimes associated with nematode infection, and one from bacterial pneumonia. Trauma is a significant factor in Laysan duckling mortalities, and elucidating the cause of and preventing such trauma may allow for management measures to enhance duckling survivability. High duckling mortality rates and emaciation also indicate that habitat on Laysan Island may have limited capacity to support broods. ?? Wildfowl & Wetlands Trust.

  17. Historical Climate and Streamflow Trends in Santa Ana River Basin

    NASA Astrophysics Data System (ADS)

    Mejia, D.; Sultana, R.; Tang, V.

    2015-12-01

    Santa Ana River watershed, located in Southern California, is the home of more than 5 million people. Population is projected to double within the next 50 years in the 2,650 square miles watershed. With prolonged drought conditions, and projected climate change, a strong concern exists about sustainable water supply of the area. In this study, historic climate and streamflow trend from water year 1965 to 2014 is analyzed using the nonparametric Mann-Kendall test. Climate trends are studied using annual rainfall, and annual average maximum and minimum temperature at 5 and 4 weather stations, respectively. Three of the precipitation stations show precipitation is decreasing in the watershed while minimum and maximum temperature has an increasing trend at three stations (p < 0.05). To assess whether streamflow and stream-channel characteristics are tended to increase or decrease monotonically with time, four variables - (1) annual maximum peak, (2) annual mean, (3) low to moderate and (4) moderate to high maximum peak streamflow were tested at 20 stream gauge sites. Only at 5 stream gage stations, significant streamflow trend is observed. At two stream gages, annual peak and annual average streamflow is increasing and at two stations, annual average streamflow has a decreasing trend. Low to moderate peak streamflow is increasing at two gage locations but there is no monotonic trend in moderate to high flows. As precipitation is decreasing in some part of the watershed, the effect of increasing urbanization in the area can be attributed for the localized increase in mean and peak streamflow. The trend analysis in weather and stream gage data will be presented in detail.

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

    PubMed

    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.

  19. 21 CFR 184.1185 - Calcium acetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Calcium acetate. 184.1185 Section 184.1185 Food... GRAS § 184.1185 Calcium acetate. (a) Calcium acetate (Ca (C2H3O2)2, CAS Reg. No. 62-54-4), also known as acetate of lime or vinegar salts, is the calcium salt of acetic acid. It may be produced by...

  20. 21 CFR 184.1185 - Calcium acetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Calcium acetate. 184.1185 Section 184.1185 Food and... Substances Affirmed as GRAS § 184.1185 Calcium acetate. (a) Calcium acetate (Ca (C2H3O2)2, CAS Reg. No. 62-54-4), also known as acetate of lime or vinegar salts, is the calcium salt of acetic acid. It may...

  1. 21 CFR 184.1185 - Calcium acetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Calcium acetate. 184.1185 Section 184.1185 Food... Specific Substances Affirmed as GRAS § 184.1185 Calcium acetate. (a) Calcium acetate (Ca (C2H3O2)2, CAS Reg. No. 62-54-4), also known as acetate of lime or vinegar salts, is the calcium salt of acetic acid....

  2. 21 CFR 184.1185 - Calcium acetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Calcium acetate. 184.1185 Section 184.1185 Food... Specific Substances Affirmed as GRAS § 184.1185 Calcium acetate. (a) Calcium acetate (Ca (C2H3O2)2, CAS Reg. No. 62-54-4), also known as acetate of lime or vinegar salts, is the calcium salt of acetic acid....

  3. 21 CFR 184.1185 - Calcium acetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Calcium acetate. 184.1185 Section 184.1185 Food... Specific Substances Affirmed as GRAS § 184.1185 Calcium acetate. (a) Calcium acetate (Ca (C2H3O2)2, CAS Reg. No. 62-54-4), also known as acetate of lime or vinegar salts, is the calcium salt of acetic acid....

  4. Analytical performance of the AtheNA MultiLyte ANA II assay in sera from lupus patients with multiple positive ANAs.

    PubMed

    Biagini, Raymond E; Parks, Christine G; Smith, Jerome P; Sammons, Deborah L; Robertson, Shirley A

    2007-06-01

    The purpose of this study was to evaluate the precision and accuracy of a commercial multiplexed kit for the measurement of 9 anti-nuclear antibodies (ANAs; anti-SS/A, anti-SS/B, anti-Sm, anti-RNP, anti-Jo-1, anti-Scl-70, anti-dsDNA, anti-Centromere B, and anti-Histone), and to compare these results to a subset of ANAs measured by enzyme-linked immunosorbent assays (ELISA) and immunodiffusion (ID). Sera were obtained from 22 systemic lupus erythematosus (SLE) patients, twelve controls and five others (commercial source) with various autoimmune diseases. ANA results from the AtheNA MultiLyte ANA II Assay (AtheNA) were compared to ELISA results (controls) and patients (ID). The AtheNA interassay coefficients of variation (CVs, N = 39, performed in duplicate; replicated 3x) ranged from 6.2% to 16.7% (mean = 9.8%), while the intra-assay CVs ranged from 5.8% to 14.3% (mean = 10.8%). Compared to results for SLE cases and controls, the sensitivity of AtheNA ranged from 85.7% to 100% (mean = 97.1%), while diagnostic specificity ranged from 16.7% to 100% (mean = 71.6%). There was significant agreement (P values ranging from 0.0001 to 0.03) when analytes coanalyzed by AtheNA and ELISA/ID were evaluated using Cohen's kappa (kappa values ranging from 0.376 to 1.000). No false positive ANA results were observed for either the control or commercial source autoimmune disease sera. These results indicate that the AtheNA assay is a precise and accurate alternative for performing multiple ELISAs or IDs in the diagnosis of autoimmune diseases, especially when the number of sera to be tested is large, such as in clinical screening or epidemiologic studies. It also appears that the AtheNA assay identifies positive ANA specificities which are missed by ID techniques, suggesting that it may have greater analytical sensitivity for some ANAs.

  5. 40 CFR 721.10001 - 2-Ethoxyethanol, 2-ethoxyethanol acetate, 2-methoxyethanol, and 2-methoxyethanol acetate.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... acetate, 2-methoxyethanol, and 2-methoxyethanol acetate. 721.10001 Section 721.10001 Protection of...-ethoxyethanol acetate, 2-methoxyethanol, and 2-methoxyethanol acetate. (a) Chemical substances and significant...-80-5), 2-ethoxyethanol acetate (CAS No. 111-15-9), 2-methoxyethanol (CAS No. 109-86-4), and...

  6. An esterase on the outer membrane of Pseudomonas aeruginosa for the hydrolysis of long chain acyl esters.

    PubMed

    Ohkawa, I; Shiga, S; Kageyama, M

    1979-09-01

    A new esterase activity which hydrolyzes palmitoyl-CoA was found in the membrane fraction of Pseudomonas aeruginosa. All the 11 strains of P. aeruginosa tested possessed this esterase activity. The esterase was constitutive and was fully active on the intact cell bodies toward substrates in the medium. It was located on the outer membrane of the cell envelope, and was not released into the culture medium. This activity was designated as OM (outer membrane) esterase. OM esterase was solubilized from the cell envelope with EDTA-Triton X-100 and purified 690-fold. It was a minor component of the outer membrane. Its molecular weight was approximately 55,000. The activity was rather stable to heat, a wide range of pH, and treatment with detergents and organic solvents. No cofactors were required. The pH optimum of the reaction was 8.5. Among various acyl-CoAs, only long chain (C12--C18) thioesters were hydrolyzed. OM esterase also hydrolyzed some kinds of oxy-esters such as p-nitrophenyl acyl esters, monoacyl esters of sucrose and Tween 80 (polyoxyethylene sorbitan monooleate). On the other hand, triglycerides, phospholipids, or hydrophobic monoesters were not hydrolyzed at all. Thus, this enzyme seems to have specificity for long chain acyl esters with hydrophilic groups, whether thio- or oxy-ester. Mutants deficient in this esterase activity were isolated. These mutants were unable to grow on Tween 80 as a sole carbon source. This suggests a possible role of OM esterase in the utilization of acyl esters as carbon sources.

  7. Analysis of proteins responsive to acetic acid in Acetobacter: molecular mechanisms conferring acetic acid resistance in acetic acid bacteria.

    PubMed

    Nakano, Shigeru; Fukaya, Masahiro

    2008-06-30

    Acetic acid bacteria are used for industrial vinegar production because of their remarkable ability to oxidize ethanol and high resistance to acetic acid. Although several molecular machineries responsible for acetic acid resistance in acetic acid bacteria have been reported, the entire mechanism that confers acetic acid resistance has not been completely understood. One of the promising methods to elucidate the entire mechanism is global analysis of proteins responsive to acetic acid by two-dimensional gel electrophoresis. Recently, two proteins whose production was greatly enhanced by acetic acid in Acetobacter aceti were identified to be aconitase and a putative ABC-transporter, respectively; furthermore, overexpression or disruption of the genes encoding these proteins affected acetic acid resistance in A. aceti, indicating that these proteins are involved in acetic acid resistance. Overexpression of each gene increased acetic acid resistance in Acetobacter, which resulted in an improvement in the productivity of acetic acid fermentation. Taken together, the results of the proteomic analysis and those of previous studies indicate that acetic acid resistance in acetic acid bacteria is conferred by several mechanisms. These findings also provide a clue to breed a strain having high resistance to acetic acid for vinegar fermentation.

  8. ANA-Negative Lupus Presenting with Heart Failure and Severe Valvular Dysfunction: Case Report and Review of the Literature.

    PubMed

    Hoang, Vu; Addison, Daniel; Lakkis, Nasser; Tabbaa, Rashed

    2015-01-01

    Antinuclear antibody (ANA) negative lupus is an important subset of the systemic lupus erythematosus (SLE) disease spectrum. Since the introduction of human cell line for ANA assay, the occurrence of true ANA-negative SLE has been a rare clinical phenomenon. The nature of cardiac involvement in ANA-negative SLE is not well understood, although any cardiac involvement, including valvular dysfunction, should be considered as a presenting manifestation of SLE irrespective of serology status. Early recognition and intervention appears to be associated with decreased morbidity. The following report describes our first case of ANA-negative SLE with an initial presentation of severe cardiac valvular dysfunction and heart failure. It also characterizes the spectrum of disease severity in ANA-negative SLE and demonstrates how aggressive SLE therapy can improve cardiac disease.

  9. The role of calcium in the hydrolysis of the organophosphate paraoxon by human serum A-esterase.

    PubMed

    Vitarius, J A; Sultatos, L G

    1995-01-01

    Human serum A-esterase is a calcium-dependent enzyme that hydrolyzes the organophosphate paraoxon by an Ordered Uni Bi kinetic mechanism. Incubation of various concentrations of calcium chloride with human serum A-esterase resulted in corresponding changes in appk3 and appE for the reaction, while appk2 was unaffected. Carboxyglutamic acid (CAG) prevented calcium chloride from altering appk3, but not appE. Similarly CAG reduced the calcium-stimulated nonenzymatic hydrolysis of paraoxon, as well as the calcium-stimulated de-phosphorylation of chymotrypsin phosphorylated by paraoxon. These results suggest that calcium plays two roles in the hydrolysis of paraoxon by A-esterase. Firstly, calcium is required in order to maintain an active site. In this capacity calcium might participate directly in the catalytic reaction, or it might be required in order to maintain the appropriate confirmation of the active site. And secondly, free calcium (or calcium weakly associated with A-esterase) facilitates the removal of diethyl phosphate from A-esterase, probably by polarizing the P = O bond of the diethyl phosphate-A-esterase intermediate, thereby rendering phosphorus more susceptible to nucleophilic attack by hydroxide ions. PMID:7823759

  10. The ferulic acid esterases of Chrysosporium lucknowense C1: purification, characterization and their potential application in biorefinery.

    PubMed

    Kühnel, S; Pouvreau, L; Appeldoorn, M M; Hinz, S W A; Schols, H A; Gruppen, H

    2012-01-01

    Three ferulic acid esterases from the filamentous fungus Chrysosporium lucknowense C1 were purified and characterized. The enzymes were most active at neutral pH and temperatures up to 45 °C. All enzymes released ferulic acid and p-coumaric acid from a soluble corn fibre fraction. Ferulic acid esterases FaeA1 and FaeA2 could also release complex dehydrodiferulic acids and dehydrotriferulic acids from corn fibre oligomers, but released only 20% of all ferulic acid present in sugar beet pectin oligomers. Ferulic acid esterase FaeB2 released almost no complex ferulic acid oligomers from corn fibre oligomers, but 60% of all ferulic acid from sugar beet pectin oligomers. The ferulic acid esterases were classified based on both, sequence similarity and their activities toward synthetic substrates. The type A ferulic acid esterases FaeA1 and FaeA2 are the first members of the phylogenetic subfamily 5 to be biochemically characterized. Type B ferulic acid esterase FaeB2 is a member of subfamily 6.

  11. Gene Cloning and Nucleotide Sequencing and Properties of a Cocaine Esterase from Rhodococcus sp. Strain MB1

    PubMed Central

    Bresler, Matthew M.; Rosser, Susan J.; Basran, Amrik; Bruce, Neil C.

    2000-01-01

    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 Mr of approximately 65,000. The apparent Km 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. PMID:10698749

  12. Cocaine metabolism: cocaine and norcocaine hydrolysis by liver and serum esterases.

    PubMed

    Stewart, D J; Inaba, T; Lucassen, M; Kalow, W

    1979-04-01

    The hydrolysis of cocaine and its N-demethylated product, norcocaine, by esterases was examined in liver and serum. Both liver and serum enzymatically formed ecgonine methyl ester from cocaine. The liver enzyme had a much lower affinity for cocaine than that of serum, indicating that a different form of esterase was present in liver. The liver enzyme had a similar affinity for both norcocaine and cocaine. Likewise, the serum enzyme showed similar affinities for both substrates. The Vmax estimates, however, were consistently higher for norcocaine than cocaine in both liver and serum. Benzoyl ecgonine, a major metabolite of cocaine formed by hydrolysis, was not produced enzymatically in either serum or liver; the rate of spontaneous formation at physiological pH suggests that this metabolite may arise nonenzymatically in the body.

  13. A New Strategy for Fluorogenic Esterase Probes Displaying Low Levels of Non-specific Hydrolysis.

    PubMed

    Kim, Sungwoo; Kim, Hyunjin; Choi, Yongdoo; Kim, Youngmi

    2015-06-26

    A new design for fluorescence probes of esterase activity that features a carboxylate-side pro-fluorophore is demonstrated with boron dipyrromethene (BODIPY)-based probes 1 a and 1 b. Because the design relies on the enzyme-catalyzed hydrolysis of an ester group that is not electronically activated, these probes exhibit a stability to background hydrolysis that is far superior to classical alcohol-side profluorophore-based probes, large signal-to-noise ratios, reduced sensitivity to pH variations, and high enzymatic reactivity. The utility of probe 1 a was established with a real-time fluorescence imaging experiment of endogenous esterase activity that does not require washing of the extracellular medium. PMID:26033618

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

    PubMed

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

    2016-02-10

    Lignin-carbohydrate complexes (LCCs) are in part responsible for the recalcitrance of lignocellulosics in relation to industrial utilization of biomass for biofuels. Glucuronoyl esterases (GEs) belonging to the carbohydrate esterase family 15 have been proposed to be able to degrade ester LCCs between glucuronic acids in xylans and lignin alcohols. By means of synthesized complex LCC model substrates we provide kinetic data suggesting a preference of fungal GEs for esters of bulky arylalkyl alcohols such as ester LCCs. Furthermore, using natural corn fiber substrate we report the first examples of improved degradation of lignocellulosic biomass by the use of GEs. Improved C5 sugar, glucose and glucuronic acid release was observed when heat pretreated corn fiber was incubated in the presence of GEs from Cerrena unicolor and Trichoderma reesei on top of different commercial cellulase/hemicellulase preparations. These results emphasize the potential of GEs for delignification of biomass thereby improving the overall yield of fermentable sugars for biofuel production. PMID:26712478

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

    PubMed

    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.

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

    PubMed

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

    2015-10-01

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

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

    PubMed

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

    2015-10-01

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

  18. Application of glutaraldehyde for the staining of esterase-active cells with carboxyfluorescein diacetate.

    PubMed

    Morono, Yuki; Takano, Suguru; Miyanaga, Kazuhiko; Tanji, Yasunori; Unno, Hajime; Hori, Katsutoshi

    2004-03-01

    Staining of esterase-active bacteria with carboxyfluorescein diacetate (CFDA) has been used to evaluate the viability of various types of cell. However, the outer membrane of Gram-negative bacteria prevents CFDA from permeating into the cell. Although EDTA can increase the permeability of the outer membrane allowing CFDA to enter the cells, it was experimentally confirmed that there is still considerable difficulty in visualizing viable cells due to passive diffusion of carboxyfluorescein (CF), a hydrolyzed product of CFDA, out of the cells. We found that glutaraldehyde enhances the discriminative recognition of esterase-active Gram-negative bacteria under microscopic observation by improving the efficacy of staining. We believe the successful staining in the presence of glutaraldehyde is due to two separate effects: an increase in the permeability of CFDA into the cell and prevention of leakage of CF out of the cell.

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

    PubMed

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

    2016-02-10

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

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

    PubMed

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

    2013-04-12

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

  1. A Chlorogenic Acid Esterase with a Unique Substrate Specificity from Ustilago maydis

    PubMed Central

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

    2014-01-01

    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. PMID:25548041

  2. Environmental Factors Modulating the Stability and Enzymatic Activity of the Petrotoga mobilis Esterase (PmEst).

    PubMed

    Lopes, Jose L S; Yoneda, Juliana S; Martins, Julia M; DeMarco, Ricardo; Jameson, David M; Castro, Aline M; Bossolan, Nelma R S; Wallace, B A; Araujo, Ana P U

    2016-01-01

    Enzymes isolated from thermophilic organisms found in oil reservoirs can find applications in many fields, including the oleochemical, pharmaceutical, bioenergy, and food/dairy industries. In this study, in silico identification and recombinant production of an esterase from the extremophile bacteria Petrotoga mobilis (designated PmEst) were performed. Then biochemical, bioinformatics and structural characterizations were undertaken using a combination of synchrotron radiation circular dichroism (SRCD) and fluorescence spectroscopies to correlate PmEst stability and hydrolytic activity on different substrates. The enzyme presented a high Michaelis-Menten constant (KM 0.16 mM) and optimum activity at ~55°C for p-nitrophenyl butyrate. The secondary structure of PmEst was preserved at acid pH, but not under alkaline conditions. PmEst was unfolded at high concentrations of urea or guanidine through apparently different mechanisms. The esterase activity of PmEst was preserved in the presence of ethanol or propanol and its melting temperature increased ~8°C in the presence of these organic solvents. PmEst is a mesophilic esterase with substrate preference towards short-to medium-length acyl chains. The SRCD data of PmEst is in agreement with the prediction of an α/β protein, which leads us to assume that it displays a typical fold of esterases from this family. The increased enzyme stability in organic solvents may enable novel applications for its use in synthetic biology. Taken together, our results demonstrate features of the PmEst enzyme that indicate it may be suitable for applications in industrial processes, particularly, when the use of polar organic solvents is required. PMID:27351338

  3. Environmental Factors Modulating the Stability and Enzymatic Activity of the Petrotoga mobilis Esterase (PmEst)

    PubMed Central

    Martins, Julia M.; DeMarco, Ricardo; Jameson, David M.; Castro, Aline M.; Bossolan, Nelma R. S.; Wallace, B. A.; Araujo, Ana P. U.

    2016-01-01

    Enzymes isolated from thermophilic organisms found in oil reservoirs can find applications in many fields, including the oleochemical, pharmaceutical, bioenergy, and food/dairy industries. In this study, in silico identification and recombinant production of an esterase from the extremophile bacteria Petrotoga mobilis (designated PmEst) were performed. Then biochemical, bioinformatics and structural characterizations were undertaken using a combination of synchrotron radiation circular dichroism (SRCD) and fluorescence spectroscopies to correlate PmEst stability and hydrolytic activity on different substrates. The enzyme presented a high Michaelis-Menten constant (KM 0.16 mM) and optimum activity at ~55°C for p-nitrophenyl butyrate. The secondary structure of PmEst was preserved at acid pH, but not under alkaline conditions. PmEst was unfolded at high concentrations of urea or guanidine through apparently different mechanisms. The esterase activity of PmEst was preserved in the presence of ethanol or propanol and its melting temperature increased ~8°C in the presence of these organic solvents. PmEst is a mesophilic esterase with substrate preference towards short-to medium-length acyl chains. The SRCD data of PmEst is in agreement with the prediction of an α/β protein, which leads us to assume that it displays a typical fold of esterases from this family. The increased enzyme stability in organic solvents may enable novel applications for its use in synthetic biology. Taken together, our results demonstrate features of the PmEst enzyme that indicate it may be suitable for applications in industrial processes, particularly, when the use of polar organic solvents is required. PMID:27351338

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

    PubMed

    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.

  5. Heterologous Expression and Biochemical Characterisation of Fourteen Esterases from Helicoverpa armigera

    PubMed Central

    Li, Yongqiang; Coppin, Chris W.; Devonshire, Alan L.; Scott, Colin; East, Peter; Russell, Robyn J.; Oakeshott, John G.

    2013-01-01

    Esterases have recurrently been implicated in insecticide resistance in Helicoverpa armigera but little is known about the underlying molecular mechanisms. We used a baculovirus system to express 14 of 30 full-length esterase genes so far identified from midgut cDNA libraries of this species. All 14 produced esterase isozymes after native PAGE and the isozymes for seven of them migrated to two regions of the gel previously associated with both organophosphate and pyrethroid resistance in various strains. Thirteen of the enzymes obtained in sufficient yield for further analysis all showed tight binding to organophosphates and low but measurable organophosphate hydrolase activity. However there was no clear difference in activity between the isozymes from regions associated with resistance and those from elsewhere in the zymogram, or between eight of the isozymes from a phylogenetic clade previously associated with resistance in proteomic and quantitative rtPCR experiments and five others not so associated. By contrast, the enzymes differed markedly in their activities against nine pyrethroid isomers and the enzymes with highest activity for the most insecticidal isomers were from regions of the gel and, in some cases, the phylogeny that had previously been associated with pyrethroid resistance. Phospholipase treatment confirmed predictions from sequence analysis that three of the isozymes were GPI anchored. This unusual feature among carboxylesterases has previously been suggested to underpin an association that some authors have noted between esterases and resistance to the Cry1Ac toxin from Bacillus thuringiensis. However these three isozymes did not migrate to the zymogram region previously associated with Cry1Ac resistance. PMID:23799064

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

    PubMed

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

    2013-01-01

    Genetic diversity and structure were analyzed in 10 accessions belonging to Banco Ativo de Germoplasma de Capsicum located at Federal University of Piauí in northwestern Brazil that receives pepper samples grown in community gardens in various regions and Brazilian states. Selections were made from seeds of C. chinense (4 accessions), C. annuum (5 accessions), and C. baccatum (1 accession). Samples consisting of leaves were collected from 4-10 plants of each accession (a total of 85 plants). Native polyacrylamide gel electrophoresis was used to identify α- and β-esterase polymorphisms. Polymorphism was clearly detected in 5 loci. Sixteen alleles were found at 5 α/β-esterase loci of the three Capsicum species. In the C. chinense samples, the highest HO and HE values were 0.3625 and 0.4395, respectively, whereas in C. annuum samples, HO and HE values were 0.2980 and 0.3310, respectively; the estimated HO and HE values in C. chinense samples were higher than those detected in C. annuum samples. A deficit of homozygous individuals was found in C. chinense (FIS = -0.6978) and C. annuum (FIS = 0.7750). Genetic differentiation between C. chinense and C. annuum at these loci was high (FST = 0.1867) indicating that C. chinense and C. annuum are genetically structured species for α/β- esterase isozymes. The esterase analysis showed high genetic diversity among the C. chinense and C. annuum samples and very high genetic differentiation (FST = 0.6321) among the C. chinense and C. annuum samples and the C. baccatum accession. PMID:23661440

  7. Purification and characterization of an endo-1,4-beta-glucanase from Neisseria sicca SB that hydrolyzes beta-1,4 linkages in cellulose acetate.

    PubMed

    Moriyoshi, Kunihiko; Ohmoto, Takashi; Ohe, Tatsuhiko; Sakai, Kiyofumi

    2002-03-01

    An enzyme catalyzing hydrolysis of beta-1,4 bonds in cellulose acetate was purified 18.3-fold to electrophoretic homogeneity from a culture supernatant of Neisseria sicca SB, which can assimilate cellulose acetate as the sole carbon and energy source. The molecular mass of the enzyme was 41 kDa and the isoelectric point was 4.8. The pH and temperature optima of the enzyme were 6.0-7.0 and 60 degrees C. The enzyme catalyzed hydrolysis of water-soluble cellulose acetate (degree of substitution, 0.88) and carboxymethyl cellulose. The Km and Vmax for water-soluble cellulose acetate and carboxymethyl cellulose were 0.242% and 2.24 micromol/min/mg, and 2.28% and 12.8 micromol/min/mg, respectively. It is estimated that the enzyme is a kind of endo-1,4-beta-glucanase (EC 3.2.1.4) from the substrate specificity and hydrolysis products of cellooligosaccharides. The enzyme and cellulose acetate esterase from Neisseria sicca SB degraded water-insoluble cellulose acetate by synergistic action.

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

    NASA Astrophysics Data System (ADS)

    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.

  9. Hormone-sensitive lipase is a cholesterol esterase of the intestinal mucosa.

    PubMed

    Grober, Jacques; Lucas, Stéphanie; Sörhede-Winzell, Maria; Zaghini, Isabelle; Mairal, Aline; Contreras, Juan-Antonio; Besnard, Philippe; Holm, Cecilia; Langin, Dominique

    2003-02-21

    The identity of the enzymes responsible for lipase and cholesterol esterase activities in the small intestinal mucosa is not known. Because hormone-sensitive lipase (HSL) catalyzes the hydrolysis of acylglycerols and cholesteryl esters, we sought to determine whether HSL could be involved. HSL mRNA and protein were detected in all segments of the small intestine by Northern and Western blot analyses, respectively. Immunocytochemistry experiments revealed that HSL was expressed in the differentiated enterocytes of the villi and was absent in the undifferentiated cells of the crypt. Diacylglycerol lipase and cholesterol esterase activities were found in the different segments. Analysis of gut from HSL-null mice showed that diacylglycerol lipase activity was unchanged in the duodenum and reduced in jejunum. Neutral cholesterol esterase activity was totally abolished in duodenum, jejunum, and ileum of HSL-null mice. Analysis of HSL mRNA structure showed two types of transcripts expressed in equal amounts with alternative 5'-ends transcribed from two exons. This work demonstrates that HSL is expressed in the mucosa of the small intestine. The results also reveal that the enzyme participates in acylglycerol hydrolysis in jejunal enterocytes and cholesteryl ester hydrolysis throughout the small intestine. PMID:12482847

  10. Newly Identified Thermostable Esterase from Sulfobacillus acidophilus: Properties and Performance in Phthalate Ester Degradation

    PubMed Central

    Zhang, Xiao-Yan; Fan, Xiang; Qiu, Yong-Jun; Li, Cheng-Yuan; Xing, Shuai; Zheng, Yi-Tao

    2014-01-01

    EstS1, a newly identified thermostable esterase from Sulfobacillus acidophilus DSM10332, was heterologously expressed in Escherichia coli and shown to enzymatically degrade phthalate esters (PAEs) to their corresponding monoalkyl PAEs. The optimal pH and temperature of the esterase were found to be 8.0 and 70°C, respectively. The half-life of EstS1 at 60°C was 15 h, indicating that the enzyme had good thermostability. The specificity constant (kcat/Km) of the enzyme for p-nitrophenyl butyrate was as high as 6,770 mM−1 s−1. The potential value of EstS1 was demonstrated by its ability to effectively hydrolyze 35 to 82% of PAEs (10 mM) within 2 min at 37°C, with all substrates being completely degraded within 24 h. At 60°C, the time required for complete hydrolysis of most PAEs was reduced by half. To our knowledge, this enzyme is a new esterase identified from thermophiles that is able to degrade various PAEs at high temperatures. PMID:25149523

  11. Identification and characterization of a novel salt-tolerant esterase from a Tibetan glacier metagenomic library.

    PubMed

    De Santi, Concetta; Ambrosino, Luca; Tedesco, Pietro; Zhai, Lei; Zhou, Cheng; Xue, Yanfen; Ma, Yanhe; de Pascale, Donatella

    2015-01-01

    A salt-tolerant esterase, designated H9Est, was identified from a metagenomic library of the Karuola glacier. H9Est gene comprised 1071 bp and encoded a polypeptide of 357 amino acids with a molecular mass of 40 kDa. Sequence analysis revealed that H9Est belonged to the family IV of bacterial lypolitic enzyme. H9Est was overexpressed in Escherichia coli and the purified enzyme showed hydrolytic activity towards p-nitrophenyl esters with carbon chain from 2 to 8. The optimal esterase activity was at 40°C and pH 8.0 and the enzyme retained its activity towards some miscible organic solvents such as polyethylene glycol. A three-dimensional model of H9Est revealed that S200, D294, and H324 formed the H9Est catalytic triad. Circular Dichroism spectra and molecular dynamic simulation indicated that the esterase had a wide denaturation temperature range and flexible loops that would be beneficial for H9Est performance at low temperatures while retaining heat-resistant features. PMID:25920073

  12. A software tool for material data analysis and property prediction: CASAC-ANA

    SciTech Connect

    Zhou, J.; Xie, Q.; Feng, J.; Li, S.; Xu, Z.; Chen, L.; Gui, Z.

    1995-12-31

    In this paper, a user-friendly software, CASAC-ANA, for material data analysis and property prediction is presented. In CASAC-ANA, there are seven methods: Nonlinear Mapping (NLM), Principal Component Analysis (PCA), Stepwise Discriminant Analysis (SDA), Discriminant Analysis with Constellation Graph (DACG), Hierarchical Clustering Analysis (HCA), Stepwise Multiple Linear Regression (SMLR), and Artificial Neural Networks (ANN). The software has some noteworthy features: (1) only one input file is needed and multipath output is produced; (2) both quantitative and qualitative data of dependent variables are accepted; and (3) it is easy to link with materials property databases. As a generalized modeling tool, CASAC-ANA can be used to treat material data concerning composition, technological processes, properties, and to predict properties of materials. The validity of the CASAC-ANA software has been tested successfully with three typical case studies concerning structural alloy steels, nickel-base superalloys, and continuously cast copper alloys. These CASAC-ANA methods have been compared and discussed.

  13. Reductive opening of carbohydrate phenylsulfonylethylidene (PSE) acetals.

    PubMed

    Chéry, Florence; Cabianca, Elena; Tatibouët, Arnaud; De Lucchi, Ottorino; Lindhorst, Thisbe K; Rollin, Patrick

    2015-11-19

    The phenylsulfonylethylidene (PSE) acetal is a relatively new protecting group in carbohydrate chemistry. However, carbohydrate-derived phenylsulfonylethylidene (PSE) acetals show a different behavior in reductive desulfonylation than simple symmetrical acetals. Here we have investigated various SET-type reaction conditions in order to open PSE acetals regioselectively and to produce chiral ω-hydroxyethenyl ethers. Whereas sodium amalgam leads to a mixture of regioisomeric vinyl ethers besides the ethylidene acetal, samarium iodide is suited for regioselective ring opening. This is shown with seven different carbohydrate PSE acetals, both of the 1,3-dioxane and the 1,3-dioxolane type. PMID:26469209

  14. Sediment Dynamics Affecting the Threatened Santa Ana Sucker in the Highly-modified Santa Ana River and Inset Channel, Southern California, USA

    NASA Astrophysics Data System (ADS)

    Minear, J. T.; Wright, S. A.

    2015-12-01

    In this study, we investigate the sediment dynamics of the low-flow channel of the Santa Ana River that is formed by wastewater discharges and contains some of the last remaining habitat of the Santa Ana Sucker (Catostomus santaanae). The Santa Ana River is a highly-modified river draining the San Bernardino Mountains and Inland Empire metropolitan area east of Los Angeles. Home to over 4 million people, the watershed provides habitat for the federally-threatened Santa Ana Sucker, which presently reside within the mainstem Santa Ana River in a reach supported by year-round constant discharges from water treatment plants. The nearly constant low-flow wastewater discharges and infrequent runoff events create a small, approximately 8 m wide, inset channel within the approximately 300 m wide mainstem channel that is typically dry except for large flood flows. The sediment dynamics within the inset channel are characterized by constantly evolving bed substrate and sediment transport rates, and occasional channel avulsions. The sediment dynamics have large influence on the Sucker, which rely on coarse-substrate (gravel and cobble) for their food production. In WY 2013 through the present, we investigated the sediment dynamics of the inset channel using repeat bathymetric and substrate surveys, bedload sampling, and discharge measurements. We found two distinct phases of the inset channel behavior: 1. 'Reset' flows, where sediment-laden mainstem discharges from upstream runoff events result in sand deposition in the inset channel or avulse the inset channel onto previously dry riverbed; and 2. 'Winnowing' flows, whereby the sand within the inset channel is removed by clear-water low flows from the wastewater treatment plant discharges. Thus, in contrast to many regulated rivers where high flows are required to flush fine sediments from the bed (for example, downstream from dams), in the Santa Ana River the low flows from wastewater treatment plants serve as the flushing

  15. Biochemical characterization and structural analysis of a new cold-active and salt-tolerant esterase from the marine bacterium Thalassospira sp.

    PubMed

    De Santi, Concetta; Leiros, Hanna-Kirsti S; Di Scala, Alessia; de Pascale, Donatella; Altermark, Bjørn; Willassen, Nils-Peder

    2016-05-01

    A gene encoding an esterase, ThaEst2349, was identified in the marine psychrophilic bacterium Thalassospira sp. GB04J01. The gene was cloned and overexpressed in E. coli as a His-tagged fusion protein. The recombinant enzyme showed optimal activity at 45 °C and the thermal stability displayed a retention of 75 % relative activity at 40 °C after 2 h. The optimal pH was 8.5 but the enzyme kept more than 75 % of its maximal activity between pH 8.0 and 9.5. ThaEst2349 also showed remarkable tolerance towards high concentrations of salt and it was active against short-chain p-nitrophenyl esters, displaying optimal activity with the acetate. The enzyme was tested for tolerance of organic solvents and the results are suggesting that it could function as an interesting candidate for biotechnological applications. The crystal structure of ThaEst2349 was determined to 1.69 Å revealing an asymmetric unit containing two chains, which also is the biological unit. The structure has a characteristic cap domain and a catalytic triad comprising Ser158, His285 and Asp255. To explain the cold-active nature of the enzyme, we compared it against thermophilic counterparts. Our hypothesis is that a high methionine content, less hydrogen bonds and less ion pairs render the enzyme more flexible at low temperatures. PMID:27016194

  16. The Structure of a Novel Thermophilic Esterase from the Planctomycetes Species, Thermogutta terrifontis Reveals an Open Active Site Due to a Minimal ‘Cap’ Domain

    PubMed Central

    Sayer, Christopher; Szabo, Zalan; Isupov, Michail N.; Ingham, Colin; Littlechild, Jennifer A.

    2015-01-01

    A carboxyl esterase (TtEst2) has been identified in a novel thermophilic bacterium, Thermogutta terrifontis from the phylum Planctomycetes and has been cloned and over-expressed in Escherichia coli. The enzyme has been characterized biochemically and shown to have activity toward small p-nitrophenyl (pNP) carboxylic esters with optimal activity for pNP-acetate. The enzyme shows moderate thermostability retaining 75% activity after incubation for 30 min at 70°C. The crystal structures have been determined for the native TtEst2 and its complexes with the carboxylic acid products propionate, butyrate, and valerate. TtEst2 differs from most enzymes of the α/β-hydrolase family 3 as it lacks the majority of the ‘cap’ domain and its active site cavity is exposed to the solvent. The bound ligands have allowed the identification of the carboxyl pocket in the enzyme active site. Comparison of TtEst2 with structurally related enzymes has given insight into how differences in their substrate preference can be rationalized based upon the properties of their active site pockets. PMID:26635762

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

    PubMed

    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

  18. Organophosphorus compound esterase profiles as predictors of therapeutic and toxic effects.

    PubMed

    Makhaeva, Galina F; Radchenko, Eugene V; Palyulin, Vladimir A; Rudakova, Elena V; Aksinenko, Alexey Yu; Sokolov, Vladimir B; Zefirov, Nikolay S; Richardson, Rudy J

    2013-03-25

    Certain organophosphorus compounds (OPCs) inhibit various serine esterases (EOHs) via phosphorylation of their active site serines. We focused on 4 EOHs of particular toxicological interest: acetylcholinesterase (AChE: acute neurotoxicity; cognition enhancement), butyrylcholinesterase (BChE: inhibition of drug metabolism and/or stoichiometric scavenging of EOH inhibitors; cognition enhancement), carboxylesterase (CaE: inhibition of drug metabolism and/or stoichiometric scavenging of EOH inhibitors), and neuropathy target esterase (NTE: delayed neurotoxicity, OPIDN). The relative degree of inhibition of these EOHs constitutes the "esterase profile" of an OPC and serves as a major determinant of its net physiological effects. Thus, understanding and controlling the esterase profile of OPC activity and selectivity toward these 4 target enzymes is a significant undertaking. In the present study, we analyzed the inhibitor properties of 52 OPCs against the 4 EOHs, along with pairwise and multitarget selectivities between them, using 2 QSAR approaches: Hansch modeling and Molecular Field Topology Analysis (MFTA). The general formula of the OPCs was (RO)(2)P(O)X, where R = alkyl, X = - SCH(Hal)COOEt (Hal = Cl, Br), -SCHCl(2), -SCH(2)Br, -OCH(CF(3))R(1) (R(1) = C(6)H(5), CF(3), COOEt, COOMe). The Hansch model showed that increasing neuropathic potential correlated with rising R hydrophobicity; moreover, OPC binding to scavenger EOHs (BChE and CaE) had different effects on potential acute and delayed neurotoxicity. Predicted protective roles of BChE and CaE against acute toxicity were enhanced with increasing hydrophobicity, but projected protection against OPIDN was decreased. Next, Molecular Field Topology Analysis (MFTA) models were built, considering atomic descriptors, e.g., effective charge, van der Waals radius of environment, and group lipophilicity. Activity/selectivity maps confirmed predictions from Hansch models and revealed other structural factors affecting

  19. Automated tests of ANA immunofluorescence as throughput autoantibody detection technology: strengths and limitations

    PubMed Central

    2014-01-01

    Anti-nuclear antibody (ANA) assay is a screening test used for almost all autoimmune rheumatic diseases, and in a number of these cases, it is a diagnostic/classification parameter. In addition, ANA is also a useful test for additional autoimmune disorders. The indirect immunofluorescence technique on monolayers of cultured epithelial cells is the current recommended method because it has higher sensitivity than solid phase assays. However, the technique is time-consuming and requires skilled operators. Automated ANA reading systems have recently been developed, which offer the advantage of faster and much easier performance as well as better harmonization in the interpretation of the results. Preliminary validation studies of these systems have given promising results in terms of analytical specificity and reproducibility. However, these techniques require further validation in clinical studies and need improvement in their recognition of mixed or less common staining patterns. PMID:24589329

  20. Inside the "pro-ana" community: a covert online participant observation.

    PubMed

    Brotsky, Sarah R; Giles, David

    2007-01-01

    A covert participant observation was conducted into the meanings of interaction in the "pro-ana" online community. Specifically, the researchers were interested in the kind of psychological support offered by such websites and by the beliefs of community members towards eating disorders and the processes of treatment and recovery. One of the authors joined a number of pro-ana sites in the guise of a plausible persona and experienced a variety of responses from community members, some extremely hostile, others very supportive, yet without ever being exposed as an interloper. These starkly different responses challenge the notion of a broad "pro-ana" philosophy, suggesting that the sites are best understood as local cliques offering temporary relief from offline hostility, but it is doubtful whether they can be said to possess any therapeutic value beyond the immediate online context.

  1. Santa Ana Winds and Fire Regimes of Southern California National Forests

    NASA Astrophysics Data System (ADS)

    Bendix, J.

    2015-12-01

    In Southern California, it has long been understood that foehn-type Santa Ana winds are an important factor in the occurrence of large wildfires. Although a variety of anecdotal observations and statistical analyses have confirmed the importance of these winds to wildfire, particularly in the Fall months when Santa Ana winds overlap with dry fuels from summer drought, many of the details of those winds' impacts on fire remain obscure. This paper uses data regarding individual fires from California's Fire and Resource Assessment Program database and a compilation of Santa Ana Wind days (SAW days) published by Abatzoglou et al. in 2013 to assess the relationship of Santa Ana winds to fire occurrence and size in Southern California. The analysis included 474 fires larger than 20 ha (~50 acres).that burned on the four Southern California national forests (Angeles, Cleveland, Los Padres and San Bernardino) between 1948 and 2010. Overall, just 10.3% of the fires started on SAW days, and 14.4% experienced at least one SAW day between start and containment dates. The impact of Santa Ana winds is greater, however, with increasing fire size. For fires > 4000 ha, 18.4% began on SAW days, with 30.4% experiencing at least one SAW day before containment. And 20% of fires > 20000 ha started on SAW days, with 50% including one or more SAW days. Fires beginning on SAW days were larger, with a mean of 6239 ha compared to 2150 ha for fires that began on non-SAW days. Only 2% of the fires that began on SAW days were started by lightning, suggesting that the impact of Santa Ana winds on Southern California fire regimes may be enhanced by humans' role in ignitions.

  2. Carbon-isotopic analysis of dissolved acetate

    NASA Technical Reports Server (NTRS)

    Gelwicks, J. T.; Hayes, J. M.

    1990-01-01

    Heating of dried, acetate-containing solids together with oxalic acid dihydrate conveniently releases acetic acid for purification by gas chromatography. For determination of the carbon-isotopic composition of total acetate, the acetate-containing zone of the chromatographic effluent can be routed directly to a combustion furnace coupled to a vacuum system allowing recovery, purification, and packaging of CO2 for mass-spectrometric analysis. For analysis of methyl carbon, acetic acid can be cryogenically trapped from the chromatographic effluent, then transferred to a tube containing excess NaOH. The tube is evacuated, sealed, and heated to 500 degrees C to produce methane by pyrolysis of sodium acetate. Subsequent combustion of the methane allows determination of the 13C content at the methyl position in the parent acetate. With typical blanks, the standard deviation of single analyses is less than 0.4% for acetate samples larger than 5 micromoles. A full treatment of uncertainties is outlined.

  3. Ozone decomposition in aqueous acetate solutions

    SciTech Connect

    Sehested, K.; Holcman, J.; Bjergbakke, E.; Hart, E.J.

    1987-01-01

    The acetate radical ion reacts with ozone with a rate constant of k = (1.5 +/- 0.5) x 10Z dmT mol s . The products from this reaction are CO2, HCHO, and O2 . By subsequent reaction of the peroxy radical with ozone the acetate radical ion is regenerated through the OH radical. A chain decomposition of ozone takes place. It terminates when the acetate radical ion reacts with oxygen forming the unreactive peroxy acetate radical. The chain is rather short as oxygen is developed, as a result of the ozone consumption. The inhibiting effect of acetate on the ozone decay is rationalized by OH scavenging by acetate and successive reaction of the acetate radical ion with oxygen. Some products from the bimolecular disappearance of the peroxy acetate radicals, however, react further with ozone, reducing the effectiveness of the stabilization.

  4. Project W-314 updated acceptance test report HNF-4649 for HNF-4648 241-AN-A pit leak detection ANA-WT-LDSTA-331 for project W-314

    SciTech Connect

    HAMMERS, J.S.

    1999-09-30

    The purpose of the test was to verify that the AN Tank Farm AN-A Pit Leak Detector components are functionally integrated and operate in accordance with engineering design specifications. The Acceptance Test Procedure HNF-4648,24l-AN-A-Pit Leak Detection ANA-WT-LDSTA-331 was conducted between 23 June and 01 July 1999 at the 200E AN Tank Farm. The test has been completed with no open test exceptions. The test was conducted prior to final engineering ''as built'' activities being completed this had no impact on the procedure or test results. All components, identified in the procedure were found to be labeled and identified as written in the procedure.

  5. 241-AN-A valve pit manifold valves and position indication acceptance test procedure

    SciTech Connect

    VANDYKE, D.W.

    1999-08-25

    This document describes the method used to test design criteria for gear actuated ball valves installed in 241-AN-A Valve Pit located at 200E Tank Farms. The purpose of this procedure is to demonstrate the following: Equipment is properly installed, labeled, and documented on As-Built drawings; New Manifold Valves in the 241-AN-A Valve Pit are fully operable using the handwheel of the valve operators; New valve position indicators on the valve operators will show correct valve positions; New valve position switches will function properly; and New valve locking devices function properly.

  6. 21 CFR 184.1721 - Sodium acetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Sodium acetate. 184.1721 Section 184.1721 Food and....1721 Sodium acetate. (a) Sodium acetate (C2H3O2Na, CAS Reg. No. 127-09-3 or C2H3O2Na·3H2O, CAS Reg. No. 6131-90-4) is the sodium salt of acetic acid and occurs naturally in plant and animal tissues....

  7. Direct evidence for bioconversion of vitamin E acetate into vitamin E: an ex vivo study in viable human skin.

    PubMed

    Baschong, W; Artmann, C; Hueglin, D; Roeding, J

    2001-01-01

    For better stability, vitamin E is commonly used as the non-active esterified pro-drug. Such esters are postulated to be hydrolyzed to the free active form by skin-related esterases. So far, successful conversion of esterified vitamin E to free vitamin E (tocopherol) has been mainly delineated from observed biological effects. Quantitative evidence in human skin is poor. In vitro and in vivo studies on human and animal skin have proved ambiguous. Formulation-based effects may have added to this controversy. In the present study, comparable amounts of vitamin E acetate (i) in oil (Mygliol-812N), (ii) surfactant-solubilized in water, (iii) encapsulated in liposomes, or (iv) encapsulated in Nanotopes were applied to human skin mounted in modified Franz-perfusion chambers that permit emulation of both open or occlusive conditions. The distribution of vitamin E(total) (vitamin E acetate + vitamin E) was assessed on the skin surface, in the horny layers, and in the underlying skin by high-pressure liquid chromatography (HPLC), with a recovery higher than 90%. Vitamin E acetate in Mygliol deposited exclusively on the surface and in the stratum corneum. In contrast, solubilized or encapsulated vitamin E acetate deposited also in the underlying skin. Nanotopes performed best, followed by liposomes and solubilized vitamin E acetate. Non-occlusive application favored deposition in the skin relative to occlusive application. Conversion of vitamin E acetate to vitamin E was not observed on the skin surface or in the horny layers, while in the underlying skin up to 50% of the vitamin E(total) was deacetylated.

  8. 21 CFR 582.1721 - Sodium acetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Sodium acetate. 582.1721 Section 582.1721 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1721 Sodium acetate. (a) Product. Sodium acetate. (b) Conditions of use. This substance is...

  9. 21 CFR 582.1721 - Sodium acetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Sodium acetate. 582.1721 Section 582.1721 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1721 Sodium acetate. (a) Product. Sodium acetate. (b) Conditions of use. This substance is...

  10. 21 CFR 582.1721 - Sodium acetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Sodium acetate. 582.1721 Section 582.1721 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1721 Sodium acetate. (a) Product. Sodium acetate. (b) Conditions of use. This substance is...

  11. 21 CFR 582.1721 - Sodium acetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Sodium acetate. 582.1721 Section 582.1721 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1721 Sodium acetate. (a) Product. Sodium acetate. (b) Conditions of use. This substance is...

  12. 21 CFR 582.1721 - Sodium acetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Sodium acetate. 582.1721 Section 582.1721 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1721 Sodium acetate. (a) Product. Sodium acetate. (b) Conditions of use. This substance is...

  13. 21 CFR 556.380 - Melengestrol acetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Melengestrol acetate. 556.380 Section 556.380 Food... Tolerances for Residues of New Animal Drugs § 556.380 Melengestrol acetate. A tolerance of 25 parts per billion is established for residues of the parent compound, melengestrol acetate, in fat of cattle....

  14. 21 CFR 582.6185 - Calcium acetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Calcium acetate. 582.6185 Section 582.6185 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium acetate. (a) Product. Calcium acetate. (b) Conditions of use. This substance is...

  15. 21 CFR 582.6185 - Calcium acetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Calcium acetate. 582.6185 Section 582.6185 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium acetate. (a) Product. Calcium acetate. (b) Conditions of use. This substance is...

  16. 21 CFR 582.6185 - Calcium acetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Calcium acetate. 582.6185 Section 582.6185 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium acetate. (a) Product. Calcium acetate. (b) Conditions of use. This substance is...

  17. 21 CFR 582.6185 - Calcium acetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Calcium acetate. 582.6185 Section 582.6185 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium acetate. (a) Product. Calcium acetate. (b) Conditions of use. This substance is...

  18. 21 CFR 582.6185 - Calcium acetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Calcium acetate. 582.6185 Section 582.6185 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium acetate. (a) Product. Calcium acetate. (b) Conditions of use. This substance is...

  19. 21 CFR 582.1005 - Acetic acid.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Acetic acid. 582.1005 Section 582.1005 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1005 Acetic acid. (a) Product. Acetic acid. (b) Conditions of use. This substance is...

  20. 21 CFR 582.1005 - Acetic acid.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Acetic acid. 582.1005 Section 582.1005 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1005 Acetic acid. (a) Product. Acetic acid. (b) Conditions of use. This substance is...

  1. 21 CFR 582.1005 - Acetic acid.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Acetic acid. 582.1005 Section 582.1005 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1005 Acetic acid. (a) Product. Acetic acid. (b) Conditions of use. This substance is...

  2. 21 CFR 582.1005 - Acetic acid.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Acetic acid. 582.1005 Section 582.1005 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1005 Acetic acid. (a) Product. Acetic acid. (b) Conditions of use. This substance is...

  3. 21 CFR 582.1005 - Acetic acid.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Acetic acid. 582.1005 Section 582.1005 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1005 Acetic acid. (a) Product. Acetic acid. (b) Conditions of use. This substance is...

  4. Photochemistry of 2-nitrobenzylidene acetals.

    PubMed

    Sebej, Peter; Solomek, Tomás; Hroudná, L'ubica; Brancová, Pavla; Klán, Petr

    2009-11-20

    Photolysis of dihydroxy compounds (diols) protected as 2-nitrobenzylidene acetals (ONBA) and subsequent acid- or base-catalyzed hydrolysis of the 2-nitrosobenzoic acid ester intermediates result in an efficient and high-yielding release of the substrates. We investigated the scope and limitations of ONBA photochemistry and expanded upon earlier described two-step procedures to show that the protected diols of many structural varieties can also be liberated in a one-pot procedure. In view of the fact that the acetals of nonsymmetrically substituted diols are converted into one of the corresponding 2-nitrosobenzoic acid ester isomers with moderate to high regioselectivity, the mechanism of their formation was studied using various experimental techniques. The experimental data were found to be in agreement with DFT-based quantum chemical calculations that showed the preferential cleavage occurs on the acetal C-O bond in the vicinity of more electron-withdrawing (or less electron-donating) groups. The study also revealed considerable complexity in the cleavage mechanism and that the structural variations in the substrate can significantly alter the reaction pathway. This deprotection strategy was found to be also applicable for 2-thioethanol when released from the corresponding monothioacetal in the presence of a reducing agent, such as ascorbic acid.

  5. A coordinated set of ecosystem research platforms open to international research in ecotoxicology, AnaEE-France.

    PubMed

    Mougin, Christian; Azam, Didier; Caquet, Thierry; Cheviron, Nathalie; Dequiedt, Samuel; Le Galliard, Jean-François; Guillaume, Olivier; Houot, Sabine; Lacroix, Gérard; Lafolie, François; Maron, Pierre-Alain; Michniewicz, Radika; Pichot, Christian; Ranjard, Lionel; Roy, Jacques; Zeller, Bernd; Clobert, Jean; Chanzy, André

    2015-10-01

    The infrastructure for Analysis and Experimentation on Ecosystems (AnaEE-France) is an integrated network of the major French experimental, analytical, and modeling platforms dedicated to the biological study of continental ecosystems (aquatic and terrestrial). This infrastructure aims at understanding and predicting ecosystem dynamics under global change. AnaEE-France comprises complementary nodes offering access to the best experimental facilities and associated biological resources and data: Ecotrons, seminatural experimental platforms to manipulate terrestrial and aquatic ecosystems, in natura sites equipped for large-scale and long-term experiments. AnaEE-France also provides shared instruments and analytical platforms dedicated to environmental (micro) biology. Finally, AnaEE-France provides users with data bases and modeling tools designed to represent ecosystem dynamics and to go further in coupling ecological, agronomical, and evolutionary approaches. In particular, AnaEE-France offers adequate services to tackle the new challenges of research in ecotoxicology, positioning its various types of platforms in an ecologically advanced ecotoxicology approach. AnaEE-France is a leading international infrastructure, and it is pioneering the construction of AnaEE (Europe) infrastructure in the field of ecosystem research. AnaEE-France infrastructure is already open to the international community of scientists in the field of continental ecotoxicology.

  6. A coordinated set of ecosystem research platforms open to international research in ecotoxicology, AnaEE-France.

    PubMed

    Mougin, Christian; Azam, Didier; Caquet, Thierry; Cheviron, Nathalie; Dequiedt, Samuel; Le Galliard, Jean-François; Guillaume, Olivier; Houot, Sabine; Lacroix, Gérard; Lafolie, François; Maron, Pierre-Alain; Michniewicz, Radika; Pichot, Christian; Ranjard, Lionel; Roy, Jacques; Zeller, Bernd; Clobert, Jean; Chanzy, André

    2015-10-01

    The infrastructure for Analysis and Experimentation on Ecosystems (AnaEE-France) is an integrated network of the major French experimental, analytical, and modeling platforms dedicated to the biological study of continental ecosystems (aquatic and terrestrial). This infrastructure aims at understanding and predicting ecosystem dynamics under global change. AnaEE-France comprises complementary nodes offering access to the best experimental facilities and associated biological resources and data: Ecotrons, seminatural experimental platforms to manipulate terrestrial and aquatic ecosystems, in natura sites equipped for large-scale and long-term experiments. AnaEE-France also provides shared instruments and analytical platforms dedicated to environmental (micro) biology. Finally, AnaEE-France provides users with data bases and modeling tools designed to represent ecosystem dynamics and to go further in coupling ecological, agronomical, and evolutionary approaches. In particular, AnaEE-France offers adequate services to tackle the new challenges of research in ecotoxicology, positioning its various types of platforms in an ecologically advanced ecotoxicology approach. AnaEE-France is a leading international infrastructure, and it is pioneering the construction of AnaEE (Europe) infrastructure in the field of ecosystem research. AnaEE-France infrastructure is already open to the international community of scientists in the field of continental ecotoxicology. PMID:26315587

  7. A non-modular type B feruloyl esterase from Neurospora crassa exhibits concentration-dependent substrate inhibition.

    PubMed Central

    Crepin, Valerie F; Faulds, Craig B; Connerton, Ian F

    2003-01-01

    Feruloyl esterases, a subclass of the carboxylic acid esterases (EC 3.1.1.1), are able to hydrolyse the ester bond between the hydroxycinnamic acids and sugars present in the plant cell wall. The enzymes have been classified as type A or type B, based on their substrate specificity for aromatic moieties. We show that Neurospora crassa has the ability to produce multiple ferulic acid esterase activities depending upon the length of fermentation with either sugar beet pulp or wheat bran substrates. A gene identified on the basis of its expression on sugar beet pulp has been cloned and overexpressed in Pichia pastoris. The gene encodes a single-domain ferulic acid esterase, which represents the first report of a non-modular type B enzyme (fae-1 gene; GenBank accession no. AJ293029). The purified recombinant protein has been shown to exhibit concentration-dependent substrate inhibition (K(m) 0.048 mM, K (i) 2.5 mM and V(max) 8.2 units/mg against methyl 3,4-dihydroxycinnamate). The kinetic behaviour of the non-modular enzyme is discussed in terms of the diversity in the roles of the feruloyl esterases in the mobilization of plant cell wall materials and their respective modes of action. PMID:12435269

  8. Conserved tyrosine 182 residue in hyperthermophilic esterase EstE1 plays a critical role in stabilizing the active site.

    PubMed

    Truongvan, Ngoc; Chung, Hye-Shin; Jang, Sei-Heon; Lee, ChangWoo

    2016-03-01

    An aromatic amino acid, Tyr or Trp, located in the esterase active site wall, is highly conserved, with hyperthermophilic esterases showing preference for Tyr and lower temperature esterases showing preference for Trp. In this study, we investigated the role of Tyr(182) in the active site wall of hyperthermophilic esterase EstE1. Mutation of Tyr to Phe or Ala had a moderate effect on EstE1 thermal stability. However, a small-to-large mutation such as Tyr to His or Trp had a devastating effect on thermal stability. All mutant EstE1 enzymes showed reduced catalytic rates and enhanced substrate affinities as compared with wild-type EstE1. Hydrogen bond formation involving Tyr(182) was unimportant for maintaining EstE1 thermal stability, as the EstE1 structure is already adapted to high temperatures via increased intramolecular interactions. However, removal of hydrogen bond from Tyr(182) significantly decreased EstE1 catalytic activity, suggesting its role in stabilization of the active site. These results suggest that Tyr is preferred over a similarly sized Phe residue or bulky His or Trp residue in the active site walls of hyperthermophilic esterases for stabilizing the active site and regulating catalytic activity at high temperatures. PMID:26838013

  9. Biodegradation of cellulose acetate by Neisseria sicca.

    PubMed

    Sakai, K; Yamauchi, T; Nakasu, F; Ohe, T

    1996-10-01

    Bacteria capable of assimilating cellulose acetate, strains SB and SC, were isolated from soil on a medium containing cellulose acetate as a carbon source, and identified as Neisseria sicca. Both strains degraded cellulose acetate membrane filters (degree of substitution, DS, mixture of 2.8 and 2.0) and textiles (DS, 2.34) in a medium containing cellulose acetate (DS, 2.34) or its oligomer, but were not able to degrade these materials in a medium containing cellobiose octaacetate. Biodegradation of cellulose acetate (DS, 1.81 and 2.34) on the basis of biochemical oxygen demand reached 51 and 40% in the culture of N. sicca SB and 60 and 45% in the culture of N. sicca SC within 20 days. A decrease in the acetyl content of degraded cellulose acetate films and powder was confirmed by infrared and nuclear magnetic resonance analyses. After 10-day cultivation of N. sicca SB and SC, the number-average molecular weight of residual cellulose acetate decreased by 9 and 5%, respectively. Activities of enzymes that released acetic acid and produced reducing sugars from cellulose acetate were mainly present in the culture supernatant. Reactivity of enzymes for cellulose acetate (DS, 1.81) was higher than that for cellulose acetate (DS, 2.34).

  10. Presence of pharmaceutically active compounds in Doñana Park (Spain) main watersheds.

    PubMed

    Camacho-Muñoz, M D; Santos, J L; Aparicio, I; Alonso, E

    2010-05-15

    Among the emerging environmental contaminants, pharmaceutically active compounds have become a growing public concern because of their potential to cause undesirable ecological and human health effects. Doñana Park (South of Spain) includes a mosaic of unique ecosystems known around the world which is particularly affected by the quality of the incoming flowing water. This study reports the presence of a number of priority pharmaceuticals in wastewater and surface water samples from Doñana watersheds. In general, ibuprofen, naproxen, salicylic acid, propranolol, caffeine and gemfibrozil were the compounds most frequently found in all locations, in the range of ng/L to microg/L. Carbamazepine, with high potential risk to the environment, was also detected, although only in a few water samples. The main results are: (i) pharmaceuticals, as water pollutants, are continually discharged into Doñana water bodies and, owing to their biological activity, could lead to adverse effects in this outstanding aquatic ecosystem; (ii) wastewater treatments implemented in the area are insufficient to remove pharmaceuticals; and (iii) therefore, there is a requirement for better wastewater treatments in this natural area to reduce or avoid the presence of organic pollutants in general and pharmaceutical active compounds in particular. To the best of our knowledge, these data constitute the first measurements of pharmaceutical compounds in water not only from the protected area of Doñana Park but also from other Natural or National Parks in the world.

  11. 75 FR 77961 - Endangered and Threatened Wildlife and Plants; Revised Critical Habitat for Santa Ana Sucker

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-12-14

    ... Santa Ana sucker in this final rule. In the proposed rule (74 FR 65056; December 9, 2009) and the document that made available the draft economic analysis (DEA) (75 FR 38441; July 2, 2010), we stated that... River that we did not discuss in the 2005 final critical habitat designation for this species (70 FR...

  12. Map of Naval Air Station (L.T.A.), Santa Ana, Calif. Showing ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Map of Naval Air Station (L.T.A.), Santa Ana, Calif. Showing conditions on June 30, 1949. Drawing no. NA 91/A9-1(1) 1949 - Marine Corps Air Station Tustin, Northern Lighter Than Air Ship Hangar, Meffett Avenue & Maxfield Street, Tustin, Orange County, CA

  13. U.S. Marine Corps Air Facility, Santa Ana, California hangers no. ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    U.S. Marine Corps Air Facility, Santa Ana, California hangers no. 1&2-building no. 28 &29. Drawing no. PW-66-044 - Marine Corps Air Station Tustin, Northern Lighter Than Air Ship Hangar, Meffett Avenue & Maxfield Street, Tustin, Orange County, CA

  14. The Chicana Subject in Ana Castillo's Fiction and the Discursive Zone of Chicana/o Theory

    ERIC Educational Resources Information Center

    Carson, Benjamin D.

    2007-01-01

    In the world of Chicana fiction, Ana Castillo has achieved the kind of status Maxine Hong Kingston has attained within Asian American discourse. Castillo's work is popular not only with the general reading public but in many academic circles as well. What sets Castillo apart from so many other Chicana fiction writers is that she is also a…

  15. Determinants of alcohol preference in the AA and ANA rat lines selected for differential ethanol intake.

    PubMed

    Kiianmaa, K; Stenius, K; Sinclair, J D

    1991-01-01

    A selective breeding program conducted in this laboratory has resulted in the establishment of the alcohol-preferring AA (Alko Alcohol) and alcohol-avoiding ANA (Alko Nonalcohol) rat lines. These lines have been used as a tool for attempting to identify the behavioral, neurochemical, and biochemical correlates of differential voluntary ethanol consumption. Some of the differences that have been found between the lines involve differential reinforcement: AA rats, but not ANA rats, rapidly acquire an ethanol-reinforced operant response. The AA's greater development of tolerance to the depressant effects of ethanol and their faster ethanol metabolism would also allow them to drink more. Neurochemical studies have suggested differential functioning of brain monoaminergic mechanisms. The activity of tyrosine hydroxylase and dopa decarboxylase, and the brain dopamine concentrations are higher in the AA rats than in the ANA rats, and the maximal number of dopamine D2 receptors is lower in the AA rats. The concentration of noradrenaline is higher in the brain of ANA rats than in that of AA rats, while the 5-hydroxytryptamine levels do not seem to differ greatly. The importance of these differences to the line difference in ethanol intake is not, however, clear, since there appears to be no difference in the sensitivity of monoamine systems of the two lines to ethanol. PMID:1726981

  16. Map of Naval Air Station (L.T.A.), Santa Ana, Calif. Showing ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Map of Naval Air Station (L.T.A.), Santa Ana, Calif. Showing conditions on June 30, 1949. Drawing no. NA 91/A9-1(1) 1949 - Marine Corps Air Station Tustin, East of Red Hill Avenue between Edinger Avenue & Barranca Parkway, Tustin, Orange County, CA

  17. Synthesis and catalytic properties of metal clusters encapsulated within small-pore (SOD, GIS, ANA) zeolites.

    PubMed

    Goel, Sarika; Wu, Zhijie; Zones, Stacey I; Iglesia, Enrique

    2012-10-24

    The synthesis protocols for encapsulation of metal clusters reported here expand the diversity in catalytic chemistries made possible by the ability of microporous solids to select reactants, transition states, and products on the basis of their molecular size. We report a synthesis strategy for the encapsulation of noble metals and their oxides within SOD (Sodalite, 0.28 nm × 0.28 nm), GIS (Gismondine, 0.45 nm × 0.31 nm), and ANA (Analcime, 0.42 nm × 0.16 nm) zeolites. Encapsulation was achieved via direct hydrothermal synthesis for SOD and GIS using metal precursors stabilized by ammonia or organic amine ligands, which prevent their decomposition or precipitation as colloidal hydroxides at the conditions of hydrothermal synthesis (<380 K) and favor interactions between metal precursors and incipient aluminosilicate nuclei during self-assembly of microporous frameworks. The synthesis of ANA requires higher crystallization temperatures (~415 K) and high pH (>12), thereby causing precipitation of even ligand-stabilized metal precursors as hydroxides. As a result, encapsulation was achieved by the recrystallization of metal clusters containing GIS into ANA, which retained these metal clusters within voids throughout the GIS-ANA transformation.

  18. A Study of Non-Native English Speakers' Academic Performance at Santa Ana College.

    ERIC Educational Resources Information Center

    Slark, Julie; Bateman, Harold

    A study was conducted in 1980-81 at Santa Ana College (SAC) to collect data on the English communication skills of non-native English speakers and to determine if a relationship existed between these skills and student's educational success. A sample of 22 classes, with an enrollment of at least 50% non-native English speakers and representing a…

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

    PubMed

    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.

  20. Feruloyl esterase activity is influenced by bile, probiotic intestinal adhesion and milk fat.

    PubMed

    Mukdsi, M C Abeijón; Argañaraz Martínez, E; Chaia, A Perez; Medina, R B

    2016-09-01

    Cinnamoyl esterases (CE) are microbial and mammalian intestinal enzymes able to release antioxidant hydroxycinnamic acids from their non-digestible ester-linked forms naturally present in vegetable foods. Previous findings showed that oral administration of Lactobacillus fermentum CRL1446 increased intestinal CE activity and improved oxidative status in mice. The aim of this work was to evaluate the in vitro CE activity of L. fermentum CRL1446 and the effect of bile on this activity, as well as strain resistance to simulated gastrointestinal tract (GIT) conditions and its ability to adhere to intestinal epithelium and influence its basal CE activity. L. fermentum CRL1446 and L. fermentum ATCC14932 (positive control for CE activity) were able to hydrolyse different synthetic hydroxycinnamates, with higher specificity toward methyl ferulate (3,853.73 and 899.19 U/g, respectively). Feruloyl esterase (FE) activity was mainly intracellular in L. fermentum CRL1446 and cell-surface associated in L. fermentum ATCC14932. Both strains tolerated simulated GIT conditions and were able to adhere ex vivo to intestinal epithelium. Pre-incubation of L. fermentum strains with bile increased FE activity in both whole cells and supernatants (~2-fold), compared to controls, suggesting that cells were permeabilised by bile, allowing more substrate to enter the cell and/or leakage of FE enzymes. Three-fold higher FE activities were detected in intestinal tissue fragments with adhered L. fermentum CRL1446 cells compared to control fragments (without bacteria), indicating that this strain provides exogenous FE activity and could stimulate esterase activity in the intestinal mucosa. Finally, we found that milk fat had a negative effect on FE activity of intestinal tissue, in absence or presence of adhered L. fermentum. These results help explaining the increase in intestinal FE activity previously observed in mice fed with L. fermentum CRL1446, and support the potential use of this strain

  1. Crystallization and preliminary X-ray diffraction analysis of the glucuronoyl esterase catalytic domain from Hypocrea jecorina

    SciTech Connect

    Wood, S. J.; Li, X.-L.; Cotta, M. A.; Biely, P.; Duke, N. E. C.; Schiffer, M.; Pokkuluri, P. R.

    2008-04-01

    The catalytic domain of the glucuronoyl esterase from H. jecorina was overexpresssed, purified and crystallized in space group P2{sub 1}2{sub 1}2{sub 1}. X-ray diffraction data were collected to 1.9 Å resolution. The catalytic domain of the glucuronoyl esterase from Hypocrea jecorina (anamorph Trichoderma reesei) was overexpresssed, purified and crystallized by the sitting-drop vapor-diffusion method using 1.4 M sodium/potassium phosphate pH 6.9. The crystals belonged to space group P2{sub 1}2{sub 1}2{sub 1} and X-ray diffraction data were collected to 1.9 Å resolution. This is the first enzyme with glucoronoyl esterase activity to be crystallized; its structure will be valuable in lignocellulose-degradation research.

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  3. A glucuronoyl esterase from Acremonium alcalophilum cleaves native lignin-carbohydrate ester bonds.

    PubMed

    Arnling Bååth, Jenny; Giummarella, Nicola; Klaubauf, Sylvia; Lawoko, Martin; Olsson, Lisbeth

    2016-08-01

    The Glucuronoyl esterases (GE) have been proposed to target lignin-carbohydrate (LC) ester bonds between lignin moieties and glucuronic acid side groups of xylan, but to date, no direct observations of enzymatic cleavage on native LC ester bonds have been demonstrated. In the present investigation, LCC fractions from spruce and birch were treated with a recombinantly produced GE originating from Acremonium alcalophilum (AaGE1). A combination of size exclusion chromatography and (31) P NMR analyses of phosphitylated LCC samples, before and after AaGE1 treatment provided the first evidence for cleavage of the LC ester linkages existing in wood. PMID:27397104

  4. p-Nitrophenylacetate hydrolysis by honey bee esterases: kinetics and inhibition.

    PubMed

    Spoonamore, J E; Frohlich, D R; Wells, M A

    1993-03-01

    1. The kinetics and inhibition of p-nitrophenylacetate hydrolysis by cytosolic esterases of 1-day old female honey bees, Apis mellifera L., were studied. 2. The calculated values obtained were Km = 2.27 x 10(-5)M and Vmax = 2.48 x 10(-8) mol/s per mg protein. 3. The inhibition mechanisms examined for four organophosphorus insecticides were highly competitive in nature and based on competitive inhibition coefficients the order of toxicity was naled > dichlorvos > cis-mevinphos = trans-mevinphos. 4. Comparisons are made with the alfalfa leafcutting bee, Megachile rotundata (Fab). PMID:8498090

  5. Characterization of lymphoid cells in the blood of healthy adults: sequential immunological, cytochemical and cytokinetic studies

    SciTech Connect

    Hirt, A.; Wagner, H.P.

    1980-01-01

    With a new method, sequential immunological, cytochemical and cytokinetic studies were done on lymphoid cells in the peripheral blood of 12 healthy adults. Every single lymphoid cell could therefore be characterized by the following markers: surface immunoglobulins (sIg); rosetting with sheep red blood cells (E); unspecific acid alpha-naphthyl acetate esterase (ANAE); and 3HdT incorporation. Significantly more E+sIg-ANAE-cells (51% and 22% of all lymphoid cells, respectively). Of all ANAE+ cells 90% were E+, but 64% of all ANAE- cells were also E+. In all individuals a subpopulation of E+sIg+ cells was found. The esterase pattern of these cells was similar to that of E-sIg+ cells. The overall labeling index of the lymphoid cells examined was less than or equal to 0.2%.

  6. AnaBench: a Web/CORBA-based workbench for biomolecular sequence analysis

    PubMed Central

    Badidi, Elarbi; De Sousa, Cristina; Lang, B Franz; Burger, Gertraud

    2003-01-01

    Background Sequence data analyses such as gene identification, structure modeling or phylogenetic tree inference involve a variety of bioinformatics software tools. Due to the heterogeneity of bioinformatics tools in usage and data requirements, scientists spend much effort on technical issues including data format, storage and management of input and output, and memorization of numerous parameters and multi-step analysis procedures. Results In this paper, we present the design and implementation of AnaBench, an interactive, Web-based bioinformatics Analysis workBench allowing streamlined data analysis. Our philosophy was to minimize the technical effort not only for the scientist who uses this environment to analyze data, but also for the administrator who manages and maintains the workbench. With new bioinformatics tools published daily, AnaBench permits easy incorporation of additional tools. This flexibility is achieved by employing a three-tier distributed architecture and recent technologies including CORBA middleware, Java, JDBC, and JSP. A CORBA server permits transparent access to a workbench management database, which stores information about the users, their data, as well as the description of all bioinformatics applications that can be launched from the workbench. Conclusion AnaBench is an efficient and intuitive interactive bioinformatics environment, which offers scientists application-driven, data-driven and protocol-driven analysis approaches. The prototype of AnaBench, managed by a team at the Université de Montréal, is accessible on-line at: . Please contact the authors for details about setting up a local-network AnaBench site elsewhere. PMID:14678565

  7. New dinosaur fossils from ANA locality, Arcillas de Morella Formation (Aptian, Lower Cretaceous, Cinctorres, Spain)

    NASA Astrophysics Data System (ADS)

    Santos-Cubedo, A.; de Santisteban, C.; Suñer, M.; Galobart, A.

    2009-04-01

    Ana is one of the several dinosaur bone sites located in the Arcillas de Morella Formation (Aptian, Lower Cretaceous; eastern Iberian Chain, Spain). This site was discovered in 1998, but it remained unexcavated until 2002, when a palaeontologist team formed by members of the Institut Paleontología Miquel Crusafont from Sabadell and the Grup Guix from Vila-real unearthed the first fossil from the locality. Nowadays there are five hundred fossils collected, including vertebrate and invertebrate species. Dinosaur bones (Theropoda and Ornithopoda) are abundant in this assemblage and in the last field season bones determined as Sauropoda were found. Taxonomically, Ana is dominated by disarticulated remains of Ornithopoda, which are usually fragmentary and abraded. Many of the elements may have been reworked (spatial averaging and/or time averaging), and the fossil concentration constitutes an autochthonous to parautochthonous association, in a spatial sense. The remains found in the Ana fossils site are placed in sandstones and limes containing marine autochthonous fauna. These deposits were formed during the transgressive infilling of an incised valley. Sedimentological features indicate that fossils were finally deposited in starved shallow estuarine environment. Mineralogically, the sediment including the fossils contains grains of quartz, illite/mica, kaolinite/clorite, K-feldspar and plagioclase, distributed in two mainly grain populations, a silty-clay and a coarse sand size grain, indicating that the sediments were bedded in a low-medium energy depositional environment. Nowadays we identified in Ana, teeth of Theropoda indet. and Baryonychinae indet., and bones of Iguanodon sp. Herein, we report new fossil findings from Ana site. These materials have been determined as Iguanodontia, Titanosauriformes and Theropoda. These new findings will help to understand the dinosaur fauna present in the Lower Cretaceous of Els Ports (Castellón, Spain). Acknowledgments This

  8. Purification and characterization of an extracellular esterase with organic solvent tolerance from a halotolerant isolate, Salimicrobium sp. LY19

    PubMed Central

    2013-01-01

    Background Halotolerant bacteria are excellent sources for selecting novel enzymes. Being intrinsically stable and active under high salinities, enzymes from these prokaryotes have evolved to function optimally under extreme conditions, making them robust biocatalysts with potential applications in harsh industrial processes. Results A halotolerant strain LY19 showing lipolytic activity was isolated from saline soil of Yuncheng Salt Lake, China. It was identified as belonging to the genus of Salimicrobium by 16S rRNA gene sequence analysis. The extracellular enzyme was purified to homogeneity with molecular mass of 57 kDa by SDS-PAGE. Substrate specificity test revealed that the enzyme preferred short-chain p-nitrophenyl esters and exhibited maximum activity towards p-nitrophenyl butyrate (p-NPB), indicating an esterase activity. The esterase was highly active and stable over broad temperature (20°C-70°C), pH (7.0-10.0) and NaCl concentration (2.5%-25%) ranges, with an optimum at 50°C, pH 7.0 and 5% NaCl. Significant inhibition of the esterase was shown by ethylenediaminetetraacetic acid (EDTA), phenylmethylsulfonyl fluoride (PMSF) and phenylarsine oxide (PAO), which indicated that it was a metalloenzyme with serine and cysteine residues essential for enzyme activity. Moreover, the esterase displayed high activity and stability in the presence of hydrophobic organic solvents with log Pow ≥ 0.88 than in the absence of an organic solvent or in the presence of hydrophilic solvents. Conclusions Results from the present study indicated the novel extracellular esterase from Salimicrobium sp. LY19 exhibited thermostable, alkali-stable, halotolerant and organic solvent-tolerant properties. These features led us to conclude that the esterase may have considerable potential for industrial applications in organic synthesis reactions. PMID:24325447

  9. Functional characterization of a novel microbial esterase identified from the Indian Ocean and its use in the stereoselective preparation of ( R)-methyl mandelate

    NASA Astrophysics Data System (ADS)

    Liang, Jiayuan; Sun, Aijun; Zhang, Yun; Deng, Dun; Wang, Yongfei; Ma, Sanmei; Hu, Yunfeng

    2016-11-01

    Genomic mining has identified a novel microbial alkaline esterase from the Indian Ocean. This esterase was overexpressed in E. coli BL21 (DE3) and further functionally characterized. Under optimal conditions (10 mmol/L substrate, pH 6.0, 2 h at 40 °C), this esterase can hydrolyze racemic methyl mandelate to ( R)-methyl mandelate with very high optical purity ( e.e. >99%) and yield (nearly 90%). Interestingly, the stereoselectivity of this esterase is opposite to that of two previously reported lipases that can generate ( S)-methyl mandelate through the hydrolysis of racemic methyl mandelate. No organic solvents or other additives were required to optimize the optical purity and production of the final chiral product ( R)-methyl mandelate, which can potentially simplify the production procedure of ( R)-methyl mandelate catalyzed by esterase.

  10. Crystallization and preliminary X-ray diffraction analysis of the glucuronoyl esterase catalytic domain from Hypocrea jecorina.

    SciTech Connect

    Wood, S. J.; Li, X. -L.; Cotta, M. A.; Biely, P.; Duke, N. E. C.; Schiffer, M.; Pokkuluri, P. R.; Biosciences Division; National Center for Agricultural Utilization Research; Slovak Academy of Sciences

    2008-01-01

    The catalytic domain of the glucuronoyl esterase from Hypocrea jecorina (anamorph Trichoderma reesei) was overexpresssed, purified and crystallized by the sitting-drop vapor-diffusion method using 1.4 M sodium/potassium phosphate pH 6.9. The crystals belonged to space group P2{sub 1}2{sub 1}2{sub 1} and X-ray diffraction data were collected to 1.9 {angstrom} resolution. This is the first enzyme with glucoronoyl esterase activity to be crystallized; its structure will be valuable in lignocellulose-degradation research.

  11. Medroxyprogesterone acetate exacerbates glutamate excitotoxicity.

    PubMed

    Nilsen, Jon; Morales, Alison; Brinton, Roberta Diaz

    2006-07-01

    We previously demonstrated that progesterone functions as a neuroprotective agent whereas medroxyprogesterone acetate (MPA; Provera) does not. Moreover, MPA antagonized the neuroprotective and neurotrophic outcomes induced by 17beta-estradiol (E2). Towards developing effective hormone therapies for protection against neurodegeneration, we sought to determine whether formulation, chemical features or prevention versus treatment mode of exposure affected the outcome of MPA treatment in survival of primary hippocampal neurons. Results of these analyses indicated that both crystalline MPA and a pharmaceutical formulation (Depo-Provera) lacked neuroprotective efficacy, indicating that the effects were not dependent upon MPA formulation. Likewise, MPA in the prevention and treatment paradigms were equally ineffective at promoting neuronal survival, indicating that timing of MPA administration was not a factor. Further, the detrimental effects of MPA were not due to the presence of the acetate group, as medroxyprogesterone was as ineffective as MPA in promoting neuronal survival. Moreover, MPA pretreatment exacerbated neuron death induced by glutamate excitotoxicity as indicated by a 40% increase in neuron death determined by direct live/dead cell count and a commensurate increase in the number of positive cells by terminal deoxynucleotidyl transferase-mediated nick end-labeling. Collectively these results predict that the progestin formulation of hormone therapy will affect the vulnerability of the central nervous system to degenerative insults.

  12. Reproductive and hormonal risk factors for antinuclear antibodies (ANA) in a representative sample of U.S. women

    PubMed Central

    Parks, Christine G.; Miller, Frederick W.; Satoh, Minoru; Chan, Edward K.L.; Andrushchenko, Zhanna; Birnbaum, Linda S.; Jusko, Todd A.; Kissling, Grace E.; Patel, Mehul D.; Rose, Kathryn M.; Weinberg, Clarice; Zeldin, Darryl C.; Sandler, Dale P.

    2014-01-01

    Background Autoantibodies are of growing interest in cancer research as potential biomarkers; yet the determinants of autoimmunity are not well understood. Antinuclear antibodies (ANA) are common in the general population, and are more prevalent in women and older adults. Here we examined the relationship of ANA with reproductive and hormonal factors in a representative sample of U.S. women. Methods We analyzed data on reproductive history and exogenous hormone use in relation to serum ANA in 2,037 females ages 12 and older from the National Health and Nutrition Examination Survey (NHANES; 1999–2004). Estimated ANA prevalences were adjusted for sampling weights. Prevalence odds ratios (POR) and 95% confidence intervals (CI) were adjusted for age, race and poverty-income-ratio, and models were stratified by menopause status. Results In premenopausal women ages 20 and older, ANA prevalence was associated with parity (p<0.001; parous versus nulliparous POR=2.0; 95%CI 1.2, 3.4), but in parous women ANA did not vary by number of births, age at first birth, years since last birth or breastfeeding. In postmenopausal women, ANA prevalence was associated with an older age at menarche (p=0.019; age 16–20 versus 10–12 years POR=3.0, 95%CI 1.6, 5.9), but not with parity. Oral contraceptives and estrogen therapy were not associated with a higher ANA prevalence. Conclusions Childbearing (having had one or more births) may explain age-associated elevations in ANA prevalence seen in premenopausal women. Impact These findings highlight the importance of considering reproductive history in studies of autoimmunity and cancer in women. PMID:25086100

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

    SciTech Connect

    Hiraki, Toshiki; Shibayama, Naoya; Yoon, Young-Ho; Yun, Kyung-Mook; Hamamoto, Toshiro; Tame, Jeremy R. H.; Park, Sam-Yong

    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%.

  14. Entamoeba histolytica: soluble and membrane-associated neutral sphingomyelinase-C and other unidentified esterase activity.

    PubMed

    Vargas-Villarreal, Javier; Palacios-Corona, Rebeca; Hernández-Luna, Carlos; Mata-Cárdenas, Benito David; Torres de la Cruz, Victor M; Cortés-Gutiérrez, Elva I; González-Salazar, Francisco; Garza-González, Jesús Norberto; Escobedo-Guajardo, Brenda Leticia; Said-Fernández, Salvador

    2010-08-01

    Sphingomyelinase (SMase) activity was measured in Entamoeba histolytica particulate and soluble subcellular fractions. The effects on SMase of incubation time, total protein concentration, pH, and several divalent cations were determined. SMase-C and other unidentified esterase activity were detected in soluble and particulate fractions. SMase-C was 94.5-96.0% higher than the unidentified esterase activity. Soluble and insoluble SMase-C specific activities increased with protein dose and incubation time. Soluble and insoluble SMase-C activities were maximum at pH 7.5 and were dependent on Mg(2+), Mn(2+), or Co(2+), and inhibited by Zn(2+), Hg(2+), Ca(2+), and EDTA. SMase-C was active in the pH range of 3-10 and its maximum activity was at pH 7.5. The soluble and insoluble SMases have remarkably similar physicochemical properties, strongly suggesting that E. histolytica has just one isoform of neutral SMase-C that had not been described before and might be essential for E. histolytica metabolism or virulence. PMID:20350542

  15. Esterase activity of carbonic anhydrases serves as surrogate for selecting antibodies blocking hydratase activity.

    PubMed

    Uda, Narasimha Rao; Seibert, Volker; Stenner-Liewen, Frank; Müller, Philipp; Herzig, Petra; Gondi, Gabor; Zeidler, Reinhard; van Dijk, Marc; Zippelius, Alfred; Renner, Christoph

    2015-12-01

    Carbonic anhydrase 9 (CA9) and carbonic anhydrase 12 (CA12) were proposed as potential targets for cancer therapy more than 20 years ago. However, to date, there are only very few antibodies that have been described to specifically target CA9 and CA12 and also block the enzymatic activity of their targets. One of the early stage bottlenecks in identifying CA9- and CA12-inhibiting antibodies has been the lack of a high-throughput screening system that would allow for rapid assessment of inhibition of the targeted carbon dioxide hydratase activity of carbonic anhydrases. In this study, we show that measuring the esterase activity of carbonic anhydrase offers a robust and inexpensive screening method for identifying antibody candidates that block both hydratase and esterase activities of carbonic anhydrase's. To our knowledge, this is the first implementation of a facile surrogate-screening assay to identify potential therapeutic antibodies that block the clinically relevant hydratase activity of carbonic anhydrases. PMID:25775095

  16. A feruloyl esterase (FAE) characterized by relatively high thermostability from the edible mushroom Russula virescens.

    PubMed

    Wang, Li; Zhang, Rui; Ma, Zengqiang; Wang, Hexiang; Ng, Tzibun

    2014-01-01

    A monomeric feruloyl esterase (FAE) with a molecular mass of 62 kDa was acquired from fresh fruiting bodies of the edible mushroom Russula virescens. The isolation procedure involved ion exchange chromatography on CM-cellulose, Q-Sepharose, and SP-Sepharose and finally fast protein liquid chromatography-gel filtration on Superdex 75. Two amino acid sequences were obtained after tryptic digestion, and they both showed some homology with the esterase of some fungi. Maximal activity was observed at pH 5.0 and at 50 °C. The enzyme displayed relatively high thermostability as evidenced by over 70 % residual activity at 70 °C and about 34 % residual activity at 80 °C. The K m and V max for this enzyme on methyl ferulate were 0.19 mM and 1.65 U/mg proteins, respectively. The purified FAE prefers methyl ferulate over methyl caffeate and is least active on methyl p-coumarate. The FAE activity was not significantly affected by the presence of cations such as Mn(2+), Ca(2+), Cd(2+), Zn(2+), Mg(2+), Cu(2+), and K(+) ions but inhibited by Al(3+), Hg(2+), Fe(2+), and Pb(2+) ions at a tested concentration of 2. 5 mM.

  17. Leukocyte esterase urine strips for the screening of men with urethritis--use in developing countries.

    PubMed Central

    Tyndall, M W; Nasio, J; Maitha, G; Ndinya-Achola, J O; Plummer, F A; Sellors, J W; Luinstra, K E; Jang, D; Mahony, J B; Chernesky, M A

    1994-01-01

    BACKGROUND AND OBJECTIVES--The leukocyte esterase (LE) strip is a useful tool for the screening of men with urethritis. In developing countries, where laboratory facilities are limited, and sexually transmitted diseases endemic, simple and inexpensive diagnostic tests which perform well, would be of great value. METHODS--Men presenting with urethritis to a referral clinic for sexually transmitted diseases in Nairobi, Kenya participated in this cohort analytical study. First-void urine was collected for LE dipstick testing as part of the diagnostic work-up. The results of the dipstick measurement were compared with the laboratory detection of Chlamydia trachomatis and Neisseria gonorrhoeae. RESULTS--Of 200 men with symptoms of urethritis, 33 (17%) had a pathogen detected from the urethra or the urine. Chlamydia was detected in urine by PCR in 22 (11%), and gonorrhoea was cultured from the urethra in 11 (6%). Esterase activity (trace or greater) had a sensitivity of 76%, a specificity of 80%, a positive predictive value of 42% and a negative predictive value of 94% for the presence of chlamydia or gonorrhoea. CONCLUSIONS--The use of the LE dipstick for the screening of men with symptomatic urethritis can improve diagnostic accuracy and reduce the amount of empiric antimicrobial therapy. The low detection rate of chlamydia in these men with a clinical diagnosis of nongonococcal urethritis needs further study. PMID:8300096

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

    SciTech Connect

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

    2007-11-01

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

  19. Crystal structure of human esterase D: a potential genetic marker of retinoblastoma

    SciTech Connect

    Wu, Dong; Li, Yang; Song, Gaojie; Zhang, David; Shaw, Neil; Liu, Zhi-Jie

    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.

  20. Crystal structures of Ophiostoma piceae sterol esterase: structural insights into activation mechanism and product release.

    PubMed

    Gutiérrez-Fernández, Javier; Vaquero, María Eugenia; Prieto, Alicia; Barriuso, Jorge; Martínez, María Jesús; Hermoso, Juan A

    2014-09-01

    Sterol esterases are able to efficiently hydrolyze both sterol esters and triglycerides and to carry out synthesis reactions in the presence of organic solvents. Their high versatility makes them excellent candidates for biotechnological purposes. Sterol esterase from fungus Ophiostoma piceae (OPE) belongs to the family abH03.01 of the Candida rugosa lipase-like proteins. Crystal structures of OPE were solved in this study for the closed and open conformations. Enzyme activation involves a large displacement of the conserved lid, structural rearrangements of loop α16-α17, and formation of a dimer with a large opening. Three PEG molecules are placed in the active site, mimicking chains of the triglyceride substrate, demonstrating the position of the oxyanion hole and the three pockets that accommodate the sn-1, sn-2 and sn-3 fatty acids chains. One of them is an internal tunnel, connecting the active center with the outer surface of the enzyme 30 Å far from the catalytic Ser220. Based on our structural and biochemical results we propose a mechanism by which a great variety of different substrates can be hydrolyzed in OPE paving the way for the construction of new variants to improve the catalytic properties of these enzymes and their biotechnological applications. PMID:25108239

  1. [Characters of two gravity-related esterases in carrot callus cells].

    PubMed

    Guan, P Z; Fei, C K; Yin, J; Liu, M; Zhao, Q; Cai, W M

    1999-01-01

    On the basis of identification of two gravity-related esterases (grEST1 and grEST2) in carrot callus cells (Cai et al. 1998), we continued the study of the characteristics of these two esterases. They have the very special characteristic of SDS resistance. Their activities could be inhibited partially by deoxycholate. beta-Phenylpropionic acid, AgNO3 and CuSO4 had no inhibitory effect on their activities. The activities of grEST1 and grEST2 could be decreased by ascorbic acid and cysteine, and the influence by cysteine was particularly obvious. The molecular weights of grEST1 and grEST2 were shown to be near the ranges of 49-66 kD and 43-59 kD respectively by non-denaturing electrophoresis containing deoxycholate, Triton X-100 and SDS respectively, and the isoelectric points were approximately pH 5.4 and 4.9 respectively. Besides, grEST1 and grEST2 were found in the fraction precipitating at value between 30% and 40% saturation with (NH4)2SO4.

  2. 3-d structure-based amino acid sequence alignment of esterases, lipases and related proteins

    SciTech Connect

    Gentry, M.K.; Doctor, B.P.; Cygler, M.; Schrag, J.D.; Sussman, J.L.

    1993-05-13

    Acetylcholinesterase and butyrylcholinesterase, enzymes with potential as pretreatment drugs for organophosphate toxicity, are members of a larger family of homologous proteins that includes carboxylesterases, cholesterol esterases, lipases, and several nonhydrolytic proteins. A computer-generated alignment of 18 of the proteins, the acetylcholinesases, butyrylcholinesterases, carboxylesterases, some esterases, and the nonenzymatic proteins has been previously presented. More recently, the three-dimensional structures of two enzymes enzymes in this group, acetylcholinesterase from Torpedo californica and lipase from Geotrichum candidum, have been determined. Based on the x-ray structures and the superposition of these two enzymes, it was possible to obtain an improved amino acid sequence alignment of 32 members of this family of proteins. Examination of this alignment reveals that 24 amino acids are invariant in all of the hydrolytic proteins, and an additional 49 are well conserved. Conserved amino acids include those of the active site, the disulfide bridges, the salt bridges, in the core of the proteins, and at the edges of secondary structural elements. Comparison of the three-dimensional structures makes it possible to find a well-defined structural basis for the conservation of many of these amino acids.

  3. Molecular cloning, overexpression and characterization of a novel feruloyl esterase from a soil metagenomic library.

    PubMed

    Sang, Shu Li; Li, Gang; Hu, Xiao Peng; Liu, Yu Huan

    2011-01-01

    The gene estF27, encoding a protein with feruloyl esterase activity, was cloned through functional screening from a soil metagenomic library and expressed in Escherichiacoli BL21 (DE3) with high solubility. Sequence analysis showed that estF27 encoded a protein of 291 amino acids with a predicted molecular mass of 31.16 kDa. According to the substrate specificity, EstF27 was classified as a type A feruloyl esterase. EstF27 displayed optimal activity at 40°C and pH 6.8. This enzyme was stable in a broad pH range of 5.0-10.0 over 24 h, and retained more than 50% of its activity after 96 or 120 h incubation in the presence of 3 M KCl or 5 M NaCl. The enzyme activity was slightly enhanced by the addition of Mg(2+) and Fe(3+) at a low concentration, and completely inhibited by Cu(2+). In the enzymatic hydrolysis of destarched wheat bran, EstF27 could release ferulic acid from it in the presence of xylanase from Thermomyces lanuginosus. Given its alkalitolerance, halotolerance and highly soluble expression, EstF27 is a promising candidate for industrial applications.

  4. Crystal structures of Ophiostoma piceae sterol esterase: structural insights into activation mechanism and product release.

    PubMed

    Gutiérrez-Fernández, Javier; Vaquero, María Eugenia; Prieto, Alicia; Barriuso, Jorge; Martínez, María Jesús; Hermoso, Juan A

    2014-09-01

    Sterol esterases are able to efficiently hydrolyze both sterol esters and triglycerides and to carry out synthesis reactions in the presence of organic solvents. Their high versatility makes them excellent candidates for biotechnological purposes. Sterol esterase from fungus Ophiostoma piceae (OPE) belongs to the family abH03.01 of the Candida rugosa lipase-like proteins. Crystal structures of OPE were solved in this study for the closed and open conformations. Enzyme activation involves a large displacement of the conserved lid, structural rearrangements of loop α16-α17, and formation of a dimer with a large opening. Three PEG molecules are placed in the active site, mimicking chains of the triglyceride substrate, demonstrating the position of the oxyanion hole and the three pockets that accommodate the sn-1, sn-2 and sn-3 fatty acids chains. One of them is an internal tunnel, connecting the active center with the outer surface of the enzyme 30 Å far from the catalytic Ser220. Based on our structural and biochemical results we propose a mechanism by which a great variety of different substrates can be hydrolyzed in OPE paving the way for the construction of new variants to improve the catalytic properties of these enzymes and their biotechnological applications.

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

    PubMed Central

    Juhl, P Benjamin; Trodler, Peter; Tyagi, Sadhna; Pleiss, Jürgen

    2009-01-01

    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. PMID:19493341

  6. Identification of two novel esterases from a marine metagenomic library derived from South China Sea.

    PubMed

    Chu, Xinmin; He, Haoze; Guo, Changquan; Sun, Baolin

    2008-09-01

    The demand for novel biocatalysts is increasing in modern biotechnology, which greatly stimulates the development of powerful tools to explore the genetic resources in the environment. Metagenomics, a culture independent strategy, provides an access to valuable genetic resources of the uncultured microbes. In this study, two novel esterase genes designated as estA and estB, which encoded 277- and 328-amino-acid peptides, respectively, were isolated from a marine microbial metagenomic library by functional screening, and the corresponding esterases EstA and EstB were biochemically characterized. Amino acid sequence comparison and phylogenetic analysis indicated that EstA together with other putative lipolytic enzymes was closely related to family III, and EstB with its relatives formed a subfamily of family IV. Site-directed mutagenesis showed that EstA contained classical catalytic triad made up of S146-D222-H255, whereas EstB contained an unusual catalytic triad which consisted of S-E-H, an important feature of the subfamily. EstA exhibited habitat-specific characteristics such as its high level of stability in the presence of various divalent cations and at high concentrations of NaCl. EstB displayed remarkable activity against p-nitrophenyl esters and was highly stable in 30% methanol, ethanol, dimethylformamide, and dimethyl sulfoxide, making EstB a potential candidate for industrial applications.

  7. Understanding Palladium Acetate from a User Perspective.

    PubMed

    Carole, William A; Colacot, Thomas J

    2016-06-01

    The behavior of palladium acetate is reviewed with respect to its synthesis, characterization, structure (in both solution and solid state), and activation pathways. In addition, comparisons of catalytic activities between pure palladium acetate and two common byproducts, Pd3 (OAc)5 (NO2 ) and polymeric [Pd(OAc)2 ]n , typically present in commercially available material are reviewed. Hence, this minireview serves as a concise guide for the users of palladium acetate from both academia and industry. PMID:27125630

  8. 21 CFR 184.1721 - Sodium acetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... sodium sulfate and sodium bicarbonate. (b) The ingredient meets the specifications of the Food Chemicals... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Sodium acetate. 184.1721 Section 184.1721 Food and... Substances Affirmed as GRAS § 184.1721 Sodium acetate. (a) Sodium acetate (C2H3O2Na, CAS Reg. No. 127-09-3...

  9. 21 CFR 184.1721 - Sodium acetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... sodium sulfate and sodium bicarbonate. (b) The ingredient meets the specifications of the Food Chemicals... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Sodium acetate. 184.1721 Section 184.1721 Food and... Substances Affirmed as GRAS § 184.1721 Sodium acetate. (a) Sodium acetate (C2H3O2Na, CAS Reg. No. 127-09-3...

  10. 21 CFR 184.1721 - Sodium acetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... sodium sulfate and sodium bicarbonate. (b) The ingredient meets the specifications of the Food Chemicals... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Sodium acetate. 184.1721 Section 184.1721 Food and... Substances Affirmed as GRAS § 184.1721 Sodium acetate. (a) Sodium acetate (C2H3O2Na, CAS Reg. No. 127-09-3...

  11. 21 CFR 184.1721 - Sodium acetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... sodium sulfate and sodium bicarbonate. (b) The ingredient meets the specifications of the Food Chemicals... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Sodium acetate. 184.1721 Section 184.1721 Food and... Substances Affirmed as GRAS § 184.1721 Sodium acetate. (a) Sodium acetate (C2H3O2Na, CAS Reg. No. 127-09-3...

  12. Positron scattering from vinyl acetate

    NASA Astrophysics Data System (ADS)

    Chiari, L.; Zecca, A.; Blanco, F.; García, G.; Brunger, M. J.

    2014-09-01

    Using a Beer-Lambert attenuation approach, we report measured total cross sections (TCSs) for positron scattering from vinyl acetate (C4H6O2) in the incident positron energy range 0.15-50 eV. In addition, we also report an independent atom model with screening corrected additivity rule computation results for the TCSs, differential and integral elastic cross sections, the positronium formation cross section and inelastic integral cross sections. The energy range of these calculations is 1-1000 eV. While there is a reasonable qualitative correspondence between measurement and calculation for the TCSs, in terms of the energy dependence of those cross sections, the theory was found to be a factor of ˜2 larger in magnitude at the lower energies, even after the measured data were corrected for the forward angle scattering effect.

  13. Extractive fermentation of acetic acid

    SciTech Connect

    Busche, R.M.

    1991-12-31

    In this technoeconomic evaluation of the manufacture of acetic acid by fermentation, the use of the bacterium: Acetobacter suboxydans from the old vinegar process was compared with expected performance of the newer Clostridium thermoaceticum bacterium. Both systems were projected to operate as immobilized cells in a continuous, fluidized bed bioreactor, using solvent extraction to recover the product. Acetobacter metabolizes ethanol aerobically to produce acid at 100 g/L in a low pH medium. This ensures that the product is in the form of a concentrated extractable free acid, rather than as an unextractable salt. Unfortunately, yields from glucose by way of the ethanol fermentation are poor, but near the biological limits of the organisms involved. Conversely, C. thermoaceticum is a thermophilic anaerobe that operates at high fermentation rates on glucose at neutral pH to produce acetate salts directly in substantially quantitative yields. However, it is severely inhibited by product, which restricts concentration to a dilute 20 g/L. An improved Acetobacter system operating with recycled cells at 50 g/L appears capable of producing acid at $0.38/lb, as compared with a $0.29/lb price for synthetic acid. However, this system has only a limited margin for process improvement. The present Clostridium system cannot compete, since the required selling price would be $0.42/lb. However, if the organism could be adapted to tolerate higher product concentrations at acid pH, selling price could be reduced to $0.22/lb, or about 80% of the price of synthetic acid.

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  15. Structure and properties of the esterase from non-LTR retrotransposons suggest a role for lipids in retrotransposition.

    PubMed

    Schneider, Anna M; Schmidt, Steffen; Jonas, Stefanie; Vollmer, Benjamin; Khazina, Elena; Weichenrieder, Oliver

    2013-12-01

    Non-LTR retrotransposons are mobile genetic elements and play a major role in eukaryotic genome evolution and disease. Similar to retroviruses they encode a reverse transcriptase, but their genomic integration mechanism is fundamentally different, and they lack homologs of the retroviral nucleocapsid-forming protein Gag. Instead, their first open reading frames encode distinct multi-domain proteins (ORF1ps) presumed to package the retrotransposon-encoded RNA into ribonucleoprotein particles (RNPs). The mechanistic roles of ORF1ps are poorly understood, particularly of ORF1ps that appear to harbor an enzymatic function in the form of an SGNH-type lipolytic acetylesterase. We determined the crystal structures of the coiled coil and esterase domains of the ORF1p from the Danio rerio ZfL2-1 element. We demonstrate a dimerization of the coiled coil and a hydrolytic activity of the esterase. Furthermore, the esterase binds negatively charged phospholipids and liposomes, but not oligo-(A) RNA. Unexpectedly, the esterase can split into two dynamic half-domains, suited to engulf long fatty acid substrates extending from the active site. These properties indicate a role for lipids and membranes in non-LTR retrotransposition. We speculate that Gag-like membrane targeting properties of ORF1ps could play a role in RNP assembly and in membrane-dependent transport or localization processes. PMID:24003030

  16. ISOLATION OF JUVENILE HORMONES ESTERASE AND ITS PARTIAL CDNA CLONE FROM THE BEETLE, TENEBRIO MOLITOR. (R825433)

    EPA Science Inventory

    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...

  17. Statistical optimization of medium components and physicochemical parameters to simultaneously enhance bacterial growth and esterase production by Bacillus thuringiensis.

    PubMed

    Mazzucotelli, Cintia Anabela; Moreira, María del Rosario; Ansorena, María Roberta

    2016-01-01

    Bacillus thuringiensis is a genus extensively studied because of its high potential for biotechnological application, principally in biocontrol techniques. However, the optimization of esterase production by this strain has been scarcely studied. The aim of this work was to select and optimize the physicochemical and nutritional parameters that significantly influence the growth and esterase production of B. thuringiensis. To this purpose, 6 nutritional factors and 2 physicochemical parameters were evaluated using a Plackett-Burman design. Significant variables were optimized using a Box-Behnken design and through the desirability function to select the levels of the variables that simultaneously maximize microbial growth and esterase production. The optimum conditions resulting from simultaneous optimization of the responses under study were found to be 1 g/L glucose, 15 g/L peptone, and 3.25 g/L NaCl. Under these optimal conditions, it was possible to achieve a 2.5 log CFU/mL increase in bacterial growth and a 113-fold increase in esterase productivity, compared with minimal medium without agitation.

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

    SciTech Connect

    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.

  19. Novel ferulate esterase from Gram-positive lactic acid bacteria and analyses of the recombinant enzyme produced in E. coli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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 ...

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

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

    PubMed Central

    Pliego, Jorge; Mateos, Juan Carlos; Rodriguez, Jorge; Valero, Francisco; Baeza, Mireia; Femat, Ricardo; Camacho, Rosa; Sandoval, Georgina; Herrera-López, Enrique J.

    2015-01-01

    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. PMID:25633600

  2. Monitoring lipase/esterase activity by stopped flow in a sequential injection analysis system using p-nitrophenyl butyrate.

    PubMed

    Pliego, Jorge; Mateos, Juan Carlos; Rodriguez, Jorge; Valero, Francisco; Baeza, Mireia; Femat, Ricardo; Camacho, Rosa; Sandoval, Georgina; Herrera-López, Enrique J

    2015-01-01

    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. PMID:25633600

  3. Climate change projected fire weather sensitivity: California Santa Ana wind occurrence

    NASA Astrophysics Data System (ADS)

    Miller, Norman L.; Schlegel, Nicole J.

    2006-08-01

    A new method based on global climate model pressure gradients was developed for identifying coastal high-wind fire weather conditions, such as the Santa Ana Occurrence (SAO). Application of this method for determining southern California Santa Ana wind occurrence resulted in a good correlation between derived large-scale SAOs and observed offshore winds during periods of low humidity. The projected change in the number of SAOs was analyzed using two global climate models, one a low temperature sensitivity and the other a middle-temperature sensitivity, both forced with low and high emission scenarios, for three future time periods. This initial analysis shows consistent shifts in SAO events from earlier (September-October) to later (November-December) in the season, suggesting that SAOs may significantly increase the extent of California coastal areas burned by wildfires, loss of life, and property.

  4. Project W-314 241-AN-A valve pit upgrade acceptance for beneficial use

    SciTech Connect

    HAMMERS, J.S.

    1999-07-21

    This report identifies the responsibilities and requirements, applicable to the 241-AN-A Valve Pit Upgrades portion of Project W-314, for Acceptance for Beneficial Use in accordance with HNF-IP-0842, Vol IV, Sec 3.12. At project turnover, the end user accepts the affected Structures, Systems, and Components (SSCs) for beneficial use. This checklist is used to help the end user ensure that all documentation, training, and testing requirements are met prior to turnover. This checklist specifically identifies those items related to the upgrading of the 241-AN-A valve pit. The upgrades include: the installation of jumper/valve manifolds with position sensors, replacement pit leak detection systems, construction of replacement cover blocks, and electrical upgrades to support the instrumentation upgrades.

  5. Climate change projected fire weather sensitivity: CaliforniaSanta Ana wind occurrence

    SciTech Connect

    Miller, Norman L.; Schlegel, Nicole J.

    2006-01-01

    A new methodbased on global climate model pressuregradients was developed for identifying coastal high-wind fire weatherconditions, such as the Santa Ana Occurrence (SAO). Application of thismethod for determining southern California Santa Ana wind occurrenceresulted in a good correlation between derived large-scale SAOs andobserved offshore winds during periods of low humidity. The projectedchange in the number of SAOs was analyzed using two global climatemodels, one a low temperature sensitivity and the other amiddle-temperature sensitivity, both forced with low and high emissionscenarios, for three future time periods. This initial analysis showsconsistent shifts in SAO events from earlier (September-October) to later(November-December) in the season, suggesting that SAOs may significantlyincrease the extent of California coastal areas burned by wildfires, lossof life, and property.

  6. Differences in distribution of esterase between cell fractions of rat liver homogenates prepared in various media. Relevance to the lysosomal location of the enzyme in the intact cell

    PubMed Central

    Barrow, Patience C.; Holt, S. J.

    1971-01-01

    The distribution of esterase in subcellular fractions of rat liver homogenates was compared with that of the lysosomal enzyme acid phosphatase and the microsomal enzyme glucose 6-phosphatase. Most of the esterase from sucrose homogenate sediments with glucose 6-phosphatase and about 8% is recovered in the supernatant. However, up to 53% of the esterase can be washed from microtome sections of unfixed liver, in which less cellular damage would be expected than that caused by homogenization. About 40% of both esterase and acid phosphatase are recovered in the soluble fraction after homogenization in aqueous glycerol or in a two-phase system (Arcton 113–0.25m-sucrose), although glucose 6-phosphatase is still recovered in the microsomal fraction of such homogenates. The esterase of the microsomal fraction prepared from a sucrose homogenate is much more readily released by treatment with 0.26% deoxycholate than are other constituents of this fraction. The release of esterase from the microsomal fraction by the detergent and its concomitant release with acid phosphatase after homogenization in glycerol or the two-phase system suggests that a greater proportion of esterase may be present in lysosomes of the intact cell than is indicated by the results of standard fractionation procedures. PMID:4335692

  7. Project W-314 Specific Test and Evaluation Plan 241-AN-A Valve Pit

    SciTech Connect

    HAMMERS, J.S.

    1999-08-25

    The purpose of this Specific Test and Evaluation Plan (STEP) is to provide a detailed written plan for the systematic testing of modifications made to the 241-AN-A Valve Pit by the W-314 Project. The STEP develops the outline for test procedures that verify the system's performance to the established Project design criteria. The STEP is a lower tier document based on the W-314 Test and Evaluation Plan (TEP).

  8. Metal contamination in interstitial waters of Doñana Park.

    PubMed

    Tovar-Sanchez, Antonio; Huerta-Diaz, Miguel A; Negro, Juan J; Bravo, Miguel A; Sañudo-Wilhelmy, Sergio A

    2006-02-01

    The composition of interstitial waters in Spain's Doñana National Park was assessed 4 years after a major pyrite slurry spill occurred from the Aznalcollar Mine. Metal and nutrient concentrations in pore waters from two of the most important watercourses traversing Doñana Park were measured: Guadiamar River (affected by the accident) and Partido Stream (unimpacted by the accident). Concentrations of dissolved constituents in interstitial waters varied according to land use in the two watersheds and to the effects of the mine spill. Levels of dissolved Co, Cu, Mo, Ti, and Zn were higher in pore waters from the Guadiamar River than in the Partido Stream, suggesting that concentrations of trace elements are still influenced by the spill. In contrast, concentrations of dissolved nutrients (NH4+, NO2-, NO3-, PO4(-3)) and some trace metals used in fertilizers (e.g. Al and Cr) were higher in the Partido Stream. Levels of dissolved As, Cs, DOC, Ge, Hg, Rb and V in the interstitial waters were equal in both watercourses. Metal concentrations in interstitial waters of the Guadiamar River floodplain were between 0.3 (As) and 16,000 (Zn) times lower than those previously reported in the river and groundwater a few weeks after the mine spill. Although metals in pore water appear to have reached levels characteristic of the area before the accident, concentrations are 60-150 times higher than those in pore waters from other regions. Metal:Al ratios in Doñana's pore waters suggest a transport of contaminants from the Iberian Pyrite Belt into Doñana Park.

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

    PubMed

    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

  10. The Reliability of a Novel Automated System for ANA Immunofluorescence Analysis in Daily Clinical Practice

    PubMed Central

    Alsuwaidi, Mohammed; Dollinger, Margit; Fleck, Martin; Ehrenstein, Boris

    2016-01-01

    Automated interpretation (AI) systems for antinuclear antibody (ANA) analysis have been introduced based on assessment of indirect immunofluorescence (IIF) patterns. The diagnostic performance of a novel automated IIF reading system was compared with visual interpretation (VI) of IIF in daily clinical practice to evaluate the reduction of workload. ANA-IIF tests of consecutive serum samples from patients with suspected connective tissue disease were carried out using HEp-2 cells according to routine clinical care. AI was performed using a visual analyser (Zenit G-Sight, Menarini, Germany). Agreement rates between ANA results by AI and VI were calculated. Of the 336 samples investigated, VI yielded 205 (61%) negative, 42 (13%) ambiguous, and 89 (26%) positive results, whereas 82 (24%) were determined to be negative, 176 (52%) ambiguous, and 78 (24%) positive by AI. AI displayed a diagnostic accuracy of 175/336 samples (52%) with a kappa coefficient of 0.34 compared to VI being the gold standard. Solely relying on AI, with VI only performed for all ambiguous samples by AI, would have missed 1 of 89 (1%) positive results by VI and misclassified 2 of 205 (1%) negative results by VI as positive. The use of AI in daily clinical practice resulted only in a moderate reduction of the VI workload (82 of 336 samples: 24%). PMID:27247573

  11. Aesthetic satisfaction scoring - introducing an aesthetic numeric analogue scale (ANA-scale).

    PubMed

    Funk, Wolfgang; Podmelle, Fred; Guiol, Claudia; Metelmann, Hans Robert

    2012-07-01

    To objectively and reproducibly assess the outcome of aesthetic procedures remains one of the major, unmet challenges in maxillo-facial and plastic surgery. Frequently employed scoring systems for the evaluation of aesthetic procedures are confounded by observer bias, be it that of the patient or of the surgeon. A new approach of pragmatic and simple scoring is the ANA [Aesthetic Numeric Analogue] scale, which facilitates the objective, reproducible, standardized and internationally uniform evaluation of aesthetic procedure outcome by converting all ratings for any kind of aesthetic procedures from a subjective value to an objective figure. The intention of the ANA-scale is to relate aesthetic satisfaction from wording to figures and by this create a rating system. The study is arranging matching pairs of verbal description and figures to finally queue up generating a scale. The clinical feasibility of this rating system is demonstrated in a surgical case. As a detail of the results the influence of the viewer's age to the aesthetic benefit assessment is obvious. In summary the ANA-scale looks to be a tool useful in individual treatment protocols as well as analysis of different techniques of aesthetic surgery for rating of the pure aesthetic satisfaction of the patients.

  12. Aesthetic satisfaction scoring - introducing an aesthetic numeric analogue scale (ANA-scale).

    PubMed

    Funk, Wolfgang; Podmelle, Fred; Guiol, Claudia; Metelmann, Hans Robert

    2012-07-01

    To objectively and reproducibly assess the outcome of aesthetic procedures remains one of the major, unmet challenges in maxillo-facial and plastic surgery. Frequently employed scoring systems for the evaluation of aesthetic procedures are confounded by observer bias, be it that of the patient or of the surgeon. A new approach of pragmatic and simple scoring is the ANA [Aesthetic Numeric Analogue] scale, which facilitates the objective, reproducible, standardized and internationally uniform evaluation of aesthetic procedure outcome by converting all ratings for any kind of aesthetic procedures from a subjective value to an objective figure. The intention of the ANA-scale is to relate aesthetic satisfaction from wording to figures and by this create a rating system. The study is arranging matching pairs of verbal description and figures to finally queue up generating a scale. The clinical feasibility of this rating system is demonstrated in a surgical case. As a detail of the results the influence of the viewer's age to the aesthetic benefit assessment is obvious. In summary the ANA-scale looks to be a tool useful in individual treatment protocols as well as analysis of different techniques of aesthetic surgery for rating of the pure aesthetic satisfaction of the patients. PMID:21872486

  13. Esterase mediated resistance against synthetic pyrethroids in field populations of Rhipicephalus (Boophilus) microplus (Acari: Ixodidae) in Punjab districts of India.

    PubMed

    Singh, Nirbhay Kumar; Rath, Shitanshu S

    2014-08-29

    Detection of resistance levels against cypermethrin and deltamethrin, the most commonly used synthetic pyrethroids (SP), in Rhipicephalus (Boophilus) microplus collected from thirteen districts of Punjab (India) was carried out using adult immersion test. The regression graphs of probit mortality of ticks plotted against log values of concentrations of drugs were utilized for the determination of slope of mortality, lethal concentration for 50% (LC50), 95% (LC95) and resistance factor (RF). On the basis of the data generated on variables (mortality, egg mass weight, reproductive index and percentage inhibition of oviposition) the resistance levels were categorized. Against cypermethrin RFs of 1.48-11.22 were recorded in 12 isolates whereas, one isolate was susceptible. Resistance factors against deltamethrin were 2.4-38.54 and all 13 isolates were found to be resistant. Quantitative analysis of general esterase activity (measured by the production of the metabolite naphthol) revealed a range of 3.34 ± 0.30-13.75 ± 1.33 and 1.31 ± 0.15-8.09 ± 0.68 μmol/min/mg protein for α and β-esterase activity, respectively in different field isolates. Further, multiple pairwise comparisons of the mean values with susceptible strain (Tukey, P = 0.05) revealed significant elevated levels of both α-esterase and β-esterase in nine tick isolates resistant to both deltamethrin and cypermethrin. The data generated on acaricide resistant status and esterase mediated mechanism in ticks will help in formulating tick control strategy for the region.

  14. A cold-adapted esterase of a novel marine isolate, Pseudoalteromonas arctica: gene cloning, enzyme purification and characterization.

    PubMed

    Al Khudary, Rami; Venkatachalam, Ramprasath; Katzer, Moritz; Elleuche, Skander; Antranikian, Garabed

    2010-05-01

    A gene encoding an esterase (estO) was identified and sequenced from a gene library screen of the psychrotolerant bacterium Pseudoalteromonas arctica. Analysis of the 1,203 bp coding region revealed that the deduced peptide sequence is composed of 400 amino acids with a predicted molecular mass of 44.1 kDa. EstO contains a N-terminal esterase domain and an additional OsmC domain at the C-terminus (osmotically induced family of proteins). The highly conserved five-residue motif typical for all alpha/beta hydrolases (G x S x G) was detected from position 104 to 108 together with a putative catalytic triad consisting of Ser(106), Asp(196), and His(225). Sequence comparison showed that EstO exhibits 90% amino acid identity with hypothetical proteins containing similar esterase and OsmC domains but only around 10% identity to the amino acid sequences of known esterases. EstO variants with and without the OsmC domain were produced and purified as His-tag fusion proteins in E. coli. EstO displayed an optimum pH of 7.5 and optimum temperature of 25 degrees C with more than 50% retained activity at the freezing point of water. The thermostability of EstO (50% activity after 5 h at 40 degrees C) dramatically increased in the truncated variant (50% activity after 2.5 h at 90 degrees C). Furthermore, the esterase displays broad substrate specificity for esters of short-chain fatty acids (C(2)-C(8)).

  15. Leucocyte esterase, glucose and C-reactive protein in the diagnosis of prosthetic joint infections: a prospective study.

    PubMed

    De Vecchi, E; Villa, F; Bortolin, M; Toscano, M; Tacchini, L; Romanò, C L; Drago, L

    2016-06-01

    Analysis of joint fluid is of paramount importance for the diagnosis of prosthetic joint infections. Different markers of inflammation and/or infection in joint fluid have been proposed for diagnosis of these infections. In this study we evaluated the performance of leucocyte esterase, C-reactive protein (CRP) and glucose assays in synovial fluids from 129 patients with septic (n = 27) or aseptic (n = 102) prosthetic joint failure. Samples were collected in serum tubes and centrifuged to limit the presence of corpuscle interfering with the assays. Determinations of leucocyte esterase and glucose were carried out by means of enzymatic colorimetric reactions performed on strips for urine analysis. Tests were considered positive when graded + or ++ whereas traces or absence of colour were considered negative. CRP was measured using an automated turbidimetric method and considered suggestive for infections when >10 mg/L. Leucocyte esterase was positive in 25/27 infected patients and negative in 99/102 not infected patients (sensitivity 92.6%, specificity 97.0%). CRP was higher than the threshold in 22/27 infected patients and in 6/102 not infected patients (sensitivity: 81.5%; specificity: 94.1%) whereas glucose showed the lowest sensitivity (77.8%) and specificity (81.4%), being negative in 21/27 and 19/102 infected and not infected patients, respectively. CRP led to a correct diagnosis in 19 of 22 patients with discordant esterase and glucose results. In conclusion, evaluation of leucocyte esterase, glucose and CRP may represent a useful tool for rapid diagnosis of prosthetic joint infections. PMID:27040804

  16. Analysing deltamethrin susceptibility and pyrethroid esterase activity variations in sylvatic and domestic Triatoma infestans at the embryonic stage

    PubMed Central

    Santo-Orihuela, Pablo Luis; Carvajal, Guillermo; Picollo, María Inés; Vassena, Claudia Viviana

    2013-01-01

    The aim of the present work was to study the deltamethrin susceptibility of eggs from Triatoma infestans populations and the contribution of pyrethroid esterases to deltamethrin degradation. Insects were collected from sylvatic areas, including Veinte de Octubre and Kirus-Mayu (Bolivia) and from domiciliary areas, including El Palmar (Bolivia) and La Pista (Argentina). Deltamethrin susceptibility was determined by dose-response bioassays. Serial dilutions of deltamethrin (0.0005-1 mg/mL) were topically applied to 12-day-old eggs. Samples from El Palmar had the highest lethal dose ratio (LDR) value (44.90) compared to the susceptible reference strain (NFS), whereas the Veinte de Octubre samples had the lowest value (0.50). Pyrethroid esterases were evaluated using 7-coumaryl permethrate (7-CP) on individually homogenised eggs from each population and from NFS. The El Palmar and La Pista samples contained 40.11 and 36.64 pmol/min/mg protein, respectively, and these values were statistically similar to NFS (34.92 pmol/min/mg protein) and different from Kirus-Mayu and Veinte de Octubre (27.49 and 22.69 pmol/min/mg protein, respectively). The toxicological data indicate that the domestic populations were resistant to deltamethrin, but no statistical contribution of 7-CP esterases was observed. The sylvatic populations had similar LDR values to NFS, but lower 7-CP esterase activities. Moreover, this is the first study of the pyrethroid esterases on T. infestans eggs employing a specific substrate (7-CP). PMID:24402155

  17. 2'-F-ANA-guanosine and 2'-F-guanosine as powerful tools for structural manipulation of G-quadruplexes.

    PubMed

    Lech, Christopher Jacques; Li, Zhe; Heddi, Brahim; Phan, Anh Tuân

    2012-12-01

    Here we demonstrate the applicability of 2'-F-ANA-guanosine and 2'-F-guanosine as powerful tools for manipulating G-quadruplex folding by anti-position-favoring substitutions. A single guanine to 2'-F-ANA-guanine substitution can favor a single (3+1) hybrid conformation from a mixture of conformers. Rational substitutions of either type of 2'-F-modified nucleotide enable conformational switching from a (3+1) hybrid to a parallel folding topology.

  18. Manufacturing Ethyl Acetate From Fermentation Ethanol

    NASA Technical Reports Server (NTRS)

    Rohatgi, Naresh K.; Ingham, John D.

    1991-01-01

    Conceptual process uses dilute product of fermentation instead of concentrated ethanol. Low-concentration ethanol, extracted by vacuum from fermentation tank, and acetic acid constitutes feedstock for catalytic reaction. Product of reaction goes through steps that increases ethyl acetate content to 93 percent by weight. To conserve energy, heat exchangers recycle waste heat to preheat process streams at various points.

  19. The homo-oligomerisation of both Sas-6 and Ana2 is required for efficient centriole assembly in flies.

    PubMed

    Cottee, Matthew A; Muschalik, Nadine; Johnson, Steven; Leveson, Joanna; Raff, Jordan W; Lea, Susan M

    2015-05-23

    Sas-6 and Ana2/STIL proteins are required for centriole duplication and the homo-oligomerisation properties of Sas-6 help establish the ninefold symmetry of the central cartwheel that initiates centriole assembly. Ana2/STIL proteins are poorly conserved, but they all contain a predicted Central Coiled-Coil Domain (CCCD). Here we show that the Drosophila Ana2 CCCD forms a tetramer, and we solve its structure to 0.8 Å, revealing that it adopts an unusual parallel-coil topology. We also solve the structure of the Drosophila Sas-6 N-terminal domain to 2.9 Å revealing that it forms higher-order oligomers through canonical interactions. Point mutations that perturb Sas-6 or Ana2 homo-oligomerisation in vitro strongly perturb centriole assembly in vivo. Thus, efficient centriole duplication in flies requires the homo-oligomerisation of both Sas-6 and Ana2, and the Ana2 CCCD tetramer structure provides important information on how these proteins might cooperate to form a cartwheel structure.

  20. The homo-oligomerisation of both Sas-6 and Ana2 is required for efficient centriole assembly in flies

    PubMed Central

    Cottee, Matthew A; Muschalik, Nadine; Johnson, Steven; Leveson, Joanna; Raff, Jordan W; Lea, Susan M

    2015-01-01

    Sas-6 and Ana2/STIL proteins are required for centriole duplication and the homo-oligomerisation properties of Sas-6 help establish the ninefold symmetry of the central cartwheel that initiates centriole assembly. Ana2/STIL proteins are poorly conserved, but they all contain a predicted Central Coiled-Coil Domain (CCCD). Here we show that the Drosophila Ana2 CCCD forms a tetramer, and we solve its structure to 0.8 Å, revealing that it adopts an unusual parallel-coil topology. We also solve the structure of the Drosophila Sas-6 N-terminal domain to 2.9 Å revealing that it forms higher-order oligomers through canonical interactions. Point mutations that perturb Sas-6 or Ana2 homo-oligomerisation in vitro strongly perturb centriole assembly in vivo. Thus, efficient centriole duplication in flies requires the homo-oligomerisation of both Sas-6 and Ana2, and the Ana2 CCCD tetramer structure provides important information on how these proteins might cooperate to form a cartwheel structure. DOI: http://dx.doi.org/10.7554/eLife.07236.001 PMID:26002084

  1. Non-specific esterase isoenzymes of adult schistosomes from the hippopotamus (Hippopotamus amphibius).

    PubMed

    Fripp, P J

    1981-12-01

    Extracts of adult schistosomes collected from Hippopotamus amphibius in the Kruger National Park gave alpha naphthyl acetate isoenzyme patterns after polyacrylamide gel electrophoresis that did not correspond with those of either Schistosoma mansoni or S. rodhaini.

  2. Biomass-to-bio-products application of feruloyl esterase from Aspergillus clavatus.

    PubMed

    Damásio, André R L; Braga, Cleiton Márcio Pinto; Brenelli, Lívia B; Citadini, Ana Paula; Mandelli, Fernanda; Cota, Junio; de Almeida, Rodrigo Ferreira; Salvador, Victor Hugo; Paixao, Douglas Antonio Alvaredo; Segato, Fernando; Mercadante, Adriana Zerlotti; de Oliveira Neto, Mario; do Santos, Wanderley Dantas; Squina, Fabio M

    2013-08-01

    The structural polysaccharides contained in plant cell walls have been pointed to as a promising renewable alternative to petroleum and natural gas. Ferulic acid is a ubiquitous component of plant polysaccharides, which is found in either monomeric or dimeric forms and is covalently linked to arabinosyl residues. Ferulic acid has several commercial applications in food and pharmaceutical industries. The study herein introduces a novel feruloyl esterase from Aspergillus clavatus (AcFAE). Along with a comprehensive functional and biophysical characterization, the low-resolution structure of this enzyme was also determined by small-angle X-ray scattering. In addition, we described the production of phenolic compounds with antioxidant capacity from wheat arabinoxylan and sugarcane bagasse using AcFAE. The ability to specifically cleave ester linkages in hemicellulose is useful in several biotechnological applications, including improved accessibility to lignocellulosic enzymes for biofuel production.

  3. Structure of EstA esterase from psychrotrophic Pseudoalteromonas sp. 643A covalently inhibited by monoethylphosphonate

    SciTech Connect

    Brzuszkiewicz, Anna; Nowak, Elzbieta; Dauter, Zbigniew; Dauter, Miroslawa; Cieslinski, Hubert; Dlugolecka, Anna; Kur, Józef

    2010-10-28

    The crystal structure of the esterase EstA from the cold-adapted bacterium Pseudoalteromonas sp. 643A was determined in a covalently inhibited form at a resolution of 1.35 {angstrom}. The enzyme has a typical SGNH hydrolase structure consisting of a single domain containing a five-stranded {beta}-sheet, with three helices at the convex side and two helices at the concave side of the sheet, and is ornamented with a couple of very short helices at the domain edges. The active site is located in a groove and contains the classic catalytic triad of Ser, His and Asp. In the structure of the crystal soaked in diethyl p-nitrophenyl phosphate (DNP), the catalytic serine is covalently connected to a phosphonate moiety that clearly has only one ethyl group. This is the only example in the Protein Data Bank of a DNP-inhibited enzyme with covalently bound monoethylphosphate.

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

    PubMed

    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.

  5. Potential of Ophiostoma piceae sterol esterase for biotechnologically relevant hydrolysis reactions

    PubMed Central

    Barba Cedillo, Víctor; Prieto, Alicia; Martínez, María Jesús

    2013-01-01

    The ascomycete Ophiostoma piceae produces a sterol esterase (OPE) with high affinity toward p-nitrophenol, glycerol, and sterol esters. Recently, this enzyme has been heterologously expressed in the methylotrophic yeast Pichia pastoris under the AOX1 methanol-inducible promoter (PAOX1) using sorbitol as co-susbtrate, and the hydrolytic activity of the recombinant protein (OPE*) turned out to be improved from a kinetic point of view. In this study, we analyze the effects of sorbitol during the expression of OPE*, at first added as an additional carbon source, and methanol as inducer. The O. piceae enzyme was successfully used for PVAc hydrolysis, suggesting its potential applicability in recycled paper production to decrease stickies problems. PMID:23138020

  6. Bioassay technique using nonspecific esterase activities of Tetrahymena pyriformis for screening and assessing cytotoxicity of xenobiotics

    SciTech Connect

    Bogaerts, P.; Senaud, J.; Bohatier, J. |

    1998-08-01

    A simple and rapid test for screening and assessing the cytotoxicity of xenobiotics was developed with Tetrahymena pyriformis. The method estimates the activities of nonspecific esterases of a cell by concentrating within it a specific amount of fluorescence associated with fluorescein dye. The 2-h median effective concentration (EC50) values of 10 inorganic and eight organic substances are presented and compared to those of three other bioassays: the conventional T. pyriformis proliferation rate 9-h median inhibitory concentrations, the Microtox 30-min EC50s, and the Daphnia magna 4-methylumbelliferyl {beta}-D galactoside 1-h EC50s. A highly significant correlation was found between the results obtained with the fluorescein diacetate test and those obtained with the growth inhibition and Microtox tests. This in vivo enzymatic test showed high sensitivity to all compounds tested except Cr{sup 6+} and sodium dodecyl sulfate.

  7. Addition of feruloyl esterase and xylanase produced on-site improves sugarcane bagasse hydrolysis.

    PubMed

    Braga, Cleiton Márcio Pinto; Delabona, Priscila da Silva; Lima, Deise Juliana da Silva; Paixão, Douglas Antônio Alvaredo; Pradella, José Geraldo da Cruz; Farinas, Cristiane Sanchez

    2014-10-01

    Accessory enzymes that assist biomass degradation could be used to improve the recovery of fermentable sugar for use in biorefineries. In this study, different fungal strains isolated from the Amazon rainforest were evaluated in terms of their ability to produce feruloyl esterase (FAE) and xylanase enzymes, and an assessment was made of the contributions of the enzymes in the hydrolysis of pretreated sugarcane bagasse. In the selection step, screening using plate assays was followed by shake flask submerged cultivations. After carbon source selection and cultivation in a stirred-tank bioreactor, Aspergillusoryzae P21C3 proved to be a promising strain for production of the enzymes. Supplementation of a commercial enzyme preparation with 30% (v/v) crude enzymatic complex from A. oryzae P21C3 increased the conversion of cellulose derived from pretreated sugarcane bagasse by 36%. Supplementation with FAE and xylanase enzymes produced on-site can therefore be used to improve the hydrolysis of sugarcane bagasse.

  8. Enzyme induced biodegradation of polycarbonate-polyurethanes: dose dependence effect of cholesterol esterase.

    PubMed

    Tang, Y W; Labow, R S; Santerre, J P

    2003-05-01

    The current study has investigated the influence of esterase activity (80-400units/ml) on the biodegradation of polycarbonate-urethanes (PCNUs) by cholesterol esterase (CE), with a particular interest in studying the influence of different hard segment structures and their contribution to sensitizing the polymer towards enzyme catalyzed hydrolysis. Polycarbonate based polyurethanes were synthesized with varying hard segment content as well as hard segment chemistry based on three different diisocyanates, 1,6-hexane diisocyanate (HDI), 4,4'-methylene bisphenyl diisocyanate (MDI) and 4,4-methylene biscyclohexyl diisocyanate (HMDI). The effect of different chemistry on surface contact angle was measured in order to define the relative chemical nature of the surfaces. The enzyme dose response was found to be lower when hard segment content in the polymer was high. There was a very strong dependence on enzyme concentration for polyurethanes with different hard segment chemistry, despite the fact that the nature of the hydrolysable polycarbonate segment remained the same. The PCNU which showed the most dramatic dependence on enzyme concentration was synthesized with HMDI. At low enzyme concentration (80units/ml) this material was the most stable of the polymers while at elevated CE concentration (400units/ml) the polymer underwent a catastrophic breakdown. The findings suggested that protein binding on the surfaces was saturated even though enzyme degradation did not achieve saturation on any of the surfaces. The role of protein binding in modulating the hydrolytic action of the enzymes at different activity levels highlights a need for further study in this area.

  9. Positive and negative selection in the beta-esterase gene cluster of the Drosophila melanogaster subgroup.

    PubMed

    Balakirev, Evgeniy S; Anisimova, Maria; Ayala, Francisco J

    2006-04-01

    We examine the pattern of molecular evolution of the beta-esterase gene cluster, including the Est-6 and psiEst-6 genes, in eight species of the Drosophila melanogaster subgroup. Using maximum likelihood estimates of nonsynonymous/synonymous rate ratios, we show that the majority of Est-6 sites evolves under strong (48% of sites) or moderate (50% of sites) negative selection and a minority of sites (1.5%) is under significant positive selection. Est-6 sites likely to be under positive selection are associated with increased intraspecific variability. One positively selected site is responsible for the EST-6 F/S allozyme polymorphism; the same site is responsible for the EST-6 functional divergence between species of the melanogaster subgroup. For psiEst-6 83.7% sites evolve under negative selection, 16% sites evolve neutrally, and 0.3% sites are under positive selection. The positively selected sites of psiEst-6 are located at the beginning and at the end of the gene, where there is reduced divergence between D. melanogaster and D. simulans; these regions of psiEst-6 could be involved in regulation or some other function. Branch-site-specific analysis shows that the evolution of the melanogaster subgroup underwent episodic positive selection. Collating the present data with previous results for the beta-esterase genes, we propose that positive and negative selection are involved in a complex relationship that may be typical of the divergence of duplicate genes as one or both duplicates evolve a new function.

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

    PubMed

    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.

  11. Functional characterization of an α-esterase gene involving malathion detoxification in Bactrocera dorsalis (Hendel).

    PubMed

    Wang, Luo-Luo; Lu, Xue-Ping; Meng, Li-Wei; Huang, Yong; Wei, Dong; Jiang, Hong-Bo; Smagghe, Guy; Wang, Jin-Jun

    2016-06-01

    Extensive use of insecticides in many orchards has prompted resistance development in the oriental fruit fly, Bactrocera dorsalis (Hendel). In this study, a laboratory selected strain of B. dorsalis (MR) with a 21-fold higher resistance to malathion was used to examine the resistance mechanisms to this organophosphate insecticide. Carboxylesterase (CarE) was found to be involved in malathion resistance in B. dorsalis from the synergism bioassay by CarE-specific inhibitor triphenylphosphate (TPP). Molecular studies further identified a previously uncharacterized α-esterase gene, BdCarE2, that may function in the development of malathion resistance in B. dorsalis via gene upregulation. This gene is predominantly expressed in the Malpighian tubules, a key insect tissue for detoxification. The transcript levels of BdCarE2 were also compared between the MR and a malathion-susceptible (MS) strain of B. dorsalis, and it was significantly more abundant in the MR strain. No sequence mutation or gene copy changes were detected between the two strains. Functional studies using RNA interference (RNAi)-mediated knockdown of BdCarE2 significantly increased the malathion susceptibility in the adult files. Furthermore, heterologous expression of BdCarE2 combined with cytotoxicity assay in Sf9 cells demonstrated that BdCarE2 could probably detoxify malathion. Taken together, the current study bring new molecular evidence supporting the involvement of CarE-mediated metabolism in resistance development against malathion in B. dorsalis and also provide bases on functional analysis of insect α-esterase associated with insecticide resistance. PMID:27155483

  12. Direct Imaging of ER Calcium with Targeted-Esterase Induced Dye Loading (TED)

    PubMed Central

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

    2013-01-01

    Visualization of calcium dynamics is important to understand the role of calcium in cell physiology. To examine calcium dynamics, synthetic fluorescent Ca2+ indictors have become popular. Here we demonstrate TED (= targeted-esterase induced dye loading), a method to improve the release of Ca2+ 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 Ca2+ 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 Ca2+ indicator and a hydrophilic fluorescent dye/Ca2+ 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. PMID:23685703

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

    PubMed

    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.

  14. GDSL esterase/lipase genes in Brassica rapa L.: genome-wide identification and expression analysis.

    PubMed

    Dong, Xiangshu; Yi, Hankuil; Han, Ching-Tack; Nou, Ill-Sup; Hur, Yoonkang

    2016-04-01

    GDSL esterase/lipase proteins (GELPs), a very large subfamily of lipolytic enzymes, have been identified in microbes and many plants, but only a few have been characterized with respect to their roles in growth, development, and stress responses. In Brassica crops, as in many other species, genome-wide systematic analysis and functional studies of these genes are still lacking. As a first step to study their function in B. rapa ssp. pekinensis (Chinese cabbage), we comprehensively identified all GELP genes in the genome. We found a total of 121 Brassica rapa GDSL esterase/lipase protein genes (BrGELPs), forming three clades in the phylogenetic analysis (two major and one minor), with an asymmetrical chromosomal distribution. Most BrGELPs possess four strictly conserved residues (Ser-Gly-Asn-His) in four separate conserved regions, along with short conserved and clade-specific blocks, suggesting functional diversification of these proteins. Detailed expression profiling revealed that BrGELPs were expressed in various tissues, including floral organs, implying that BrGELPs play diverse roles in various tissues and during development. Ten percent of BrGELPs were specifically expressed in fertile buds, rather than male-sterile buds, implying their involvement in pollen development. Analyses of EXL6 (extracellular lipase 6) expression and its co-expressed genes in both B. rapa and Arabidopsis, as well as knockdown of this gene in Arabidopsis, revealed that this gene plays an important role in pollen development in both species. The data described in this study will facilitate future investigations of other BrGELP functions.

  15. GDSL esterase/lipase genes in Brassica rapa L.: genome-wide identification and expression analysis.

    PubMed

    Dong, Xiangshu; Yi, Hankuil; Han, Ching-Tack; Nou, Ill-Sup; Hur, Yoonkang

    2016-04-01

    GDSL esterase/lipase proteins (GELPs), a very large subfamily of lipolytic enzymes, have been identified in microbes and many plants, but only a few have been characterized with respect to their roles in growth, development, and stress responses. In Brassica crops, as in many other species, genome-wide systematic analysis and functional studies of these genes are still lacking. As a first step to study their function in B. rapa ssp. pekinensis (Chinese cabbage), we comprehensively identified all GELP genes in the genome. We found a total of 121 Brassica rapa GDSL esterase/lipase protein genes (BrGELPs), forming three clades in the phylogenetic analysis (two major and one minor), with an asymmetrical chromosomal distribution. Most BrGELPs possess four strictly conserved residues (Ser-Gly-Asn-His) in four separate conserved regions, along with short conserved and clade-specific blocks, suggesting functional diversification of these proteins. Detailed expression profiling revealed that BrGELPs were expressed in various tissues, including floral organs, implying that BrGELPs play diverse roles in various tissues and during development. Ten percent of BrGELPs were specifically expressed in fertile buds, rather than male-sterile buds, implying their involvement in pollen development. Analyses of EXL6 (extracellular lipase 6) expression and its co-expressed genes in both B. rapa and Arabidopsis, as well as knockdown of this gene in Arabidopsis, revealed that this gene plays an important role in pollen development in both species. The data described in this study will facilitate future investigations of other BrGELP functions. PMID:26423069

  16. Experimental West Nile virus infection in aigamo ducks, a cross between wild ducks (Anas platyrhynchos) and domestic ducks (Anas platyrhynchos var. domesticus).

    PubMed

    Shirafuji, Hiroaki; Kanehira, Katsushi; Kubo, Masanori; Shibahara, Tomoyuki; Kamio, Tsugihiko

    2009-06-01

    Four 2-wk-old and four 4-wk-old aigamo ducks, a cross between wild and domestic ducks (Anas platyrhynchos and Anas platyrhynchos var. domesticus, respectively), were infected with the NY99 strain of West Nile virus (WNV) to investigate WNV's pathogenicity in aigamo ducks and the possibility that they could transmit WNV. In the group of infected 2-wk-old aigamo ducks (2w-infection group), all of the ducks ate and drank less and showed decreased activity, some showed ataxia, and one died. Meanwhile, the group of infected 4 wk olds (4w-infection group) showed no clinical signs during the experimental period. Viremia was observed in all of the ducks in both age groups. Peak viral titers in the three surviving members of the 2w-infection group were 10(3.7)-10(5.3) plaque-forming units (PFU)/ml serum; the peak was 10(7.1) PFU/ml serum in the 2w duck that died from the infection. Peak viral titers in the 4w-infection group were 10(4.1)-10(4.9) PFU/ml serum. Viral shedding in the oral and/or cloacal cavity was observed in all four members of the 2w-infection group and in three of the four members of the 4w-infection group. These results suggest that WNV-infected aigamo ducks can transmit WNV. Although aigamo ducks are reared in East Asia, where WNV is an exotic pathogen, the virus could be introduced and spread there in the future; thus it is important to take precautions against an introduction, and measures to prevent infection to aigamo duck operations should be prepared.

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

    PubMed Central

    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

  18. Histochemical studies on genetical control of hormonal enzyme inducibility in the mouse. I. Non-specific esterase activity and regional histology of the epididymis.

    PubMed Central

    Blecher, S R; Kirkeby, S

    1978-01-01

    As a base line for future cell genetical studies the authors record the distribution of non-specific esterase reaction in the various histologically distinguishable cell types of the mouse epididymis. The findings are correlated with previous descriptions of the lobar structure of the organ. Assuming the sequence of lobes of the head to be as implied in these classical descriptions, the esterase activity of the epithelial cells gradates between strong to weak several times along the length of the epididymal duct. The relationship of the lobes to each other, as seen in transverse sections, is described. Methodological studies using different fixatives indicate that apparent similarity of esterase reaction at different sites may camouflage an underlying difference in the nature of the esterases at these sites. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 PMID:564339

  19. Conversion to eslicarbazepine acetate monotherapy

    PubMed Central

    French, Jacqueline; Jacobson, Mercedes P.; Pazdera, Ladislav; Gough, Mallory; Cheng, Hailong; Grinnell, Todd; Blum, David

    2016-01-01

    Objective: To assess the efficacy and safety of eslicarbazepine acetate (ESL) monotherapy. Methods: This post hoc pooled analysis of 2 randomized double-blind studies (093-045 and -046) included adults with partial-onset seizures medically uncontrolled by 1 or 2 antiepileptic drugs (AEDs). Following the baseline period (8 weeks), eligible patients were randomized 2:1 to receive ESL 1,600 mg or 1,200 mg once daily for 18 weeks; the primary endpoint was study exit by meeting predefined exit criteria (signifying worsening seizure control). In each study, treatment was considered effective if the upper 95% confidence limit for exit rate was lower than the historical control threshold (65.3%). Results: Pooled exit rates were as follows: ESL 1,600 mg = 20.6% (95% confidence interval: 15.6%–26.8%); ESL 1,200 mg = 30.8% (23.0%–40.5%). Use of 2 baseline AEDs or rescue medication, US location, epilepsy duration ≥20 years, and higher maximum baseline seizure frequency were associated with higher exit risks. Median percent reductions in standardized seizure frequency between baseline and the 18-week double-blind period were as follows: ESL 1,600 mg = 43.2%; ESL 1,200 mg = 35.7%; baseline carbamazepine use was associated with smaller reductions. Safety profiles were similar between ESL doses. Conclusions: Exit rates for ESL monotherapy (1,600 mg and 1,200 mg once daily) were lower than the historical control threshold, irrespective of baseline AED use and region, with no additional safety concerns identified. Clinical factors and location clearly influence treatment responses in conversion-to-monotherapy trials. Classification of evidence: This pooled analysis provides Class IV evidence that for adults with medically uncontrolled partial-onset seizures, ESL monotherapy is well tolerated and effective. PMID:26911639

  20. Intravitreal injection of octreotide acetate.

    PubMed

    Robertson, J E; Westra, I; Woltering, E A; Winthrop, K L; Barrie, R; O'Dorisio, T M; Holmes, D

    1997-04-01

    This study was conducted to determine the feasibility of injecting the somatostatin analogue, octreotide acetate (OA), into the vitreous cavity. Previous work suggests that octreotide effectively inhibits angiogenesis in vitro, thus its use in vivo may slow the progression of proliferative eye disease. Fifty micrograms of aqueous OA in 50 microliters aqueous solution was injected into the mid-vitreous of kitten eyes (n = 6), and OA levels were monitored over 4 days. A long-acting release form of octreotide (OA-LAR) was also injected into the mid-vitreous of rabbit eyes at doses of 0.36 (n = 16), 1.1 (n = 1), 2.1 (n = 1), 4.05 (n = 1), 8.2 (n = 1), and 36 mg (n = 3) in solution; and octreotide concentrations were measured at various time points over 42 days. OA concentrations were determined by a highly specific radioimmunoassay. Aqueous octreotide was eliminated rapidly (t1/2 = 16 hours) from the vitreous of the kitten eye, with only negligible amounts recoverable 4 days post-injection. In the long-acting form, OA in the rabbit eye reached peak levels at 28 days. By 42 days, OA levels had declined to the 14-day level. Doses of OA-LAR of 1.1 mg or less produced no gross evidence of clinical toxicity and elicited no grossly visible ocular side effects. Doses greater than 1.1 mg produced significant toxicity, including cataracts and rubeosis. The 28-day peak release for long-acting OA implies that monthly intravitreal injections could provide continual high levels of OA. Intravitreal injection of long-acting OA provides sustained, high concentrations of drug, and deserves further study as a potential treatment of proliferative eye diseases.

  1. Autoantibodies Profile in the Sera of Patients with Sjogren]s Syndrome: The ANA Evaluation—A Homogeneous, Multiplexed System

    PubMed Central

    Gilburd, Boris; Abu-Shakra, Mahmoud; Giordano, Andrea; Bocci, Elena Bartoloni; delle Monache, Francesco; Gerli, Roberto

    2004-01-01

    Background: Flow-based, multiplex bead arrays (MBA) have been developed for a variety of applications including the detection of antibodies to extractable nuclear antigens (ENA). It offers a rapid and sensitive method to assess multiple analyses in a single tube/well. Purpose: To evaluate the Athena Multi-Lyte ANA Test System utilizes Luminex Corporation's MBA technology for the detection of antinuclear antibodies (ANA) and ENA antibodies in the sera of patients with Sjogren's syndrome (SS). Methods: MBA assay was used to detect ANA and ENA antibodies in the sera of 37 patients with SS and 96 sera from healthy subjects. Results: All patients were women. Their mean age was 48.7 years and the mean disease duration was 7.27 years. ANA was found in 3 (3%) sera of healthy subjects by the AtheNA system and in 2 (2%) sera by the ELISA kit. A 99% concordance between the 2 assays was found. A 94.6% concordance between the 2 assays was found by testing the sera of patients with SS for ANA. By the AtheNA system, none of the sera of 37 patients with SS had autoantibodies reacting with Sm, Jo-1, dsDNA or histones. Anti-RNP antibody was found in 5.4% of the sera and 2.7% of the sera reacted with Scl-70 and histones. Anti-SS/A and anti-SS/B were identified in 84 and 76% of the sera, respectively. Conclusion: The AtheNa Multi-Lyte ANA Test System offers a sensitive and specific result for the detection of ANA and ENA antibodies in the sera of patients with SS. PMID:15154612

  2. Apoptosis induction and mitochondria alteration in human HeLa tumour cells by photoproducts of Rose Bengal acetate.

    PubMed

    Panzarini, Elisa; Tenuzzo, Bernadette; Palazzo, Fabio; Chionna, Alfonsina; Dini, Luciana

    2006-04-01

    The aim of this work was to investigate the apoptosis induction and mitochondria alteration after photodamage exerted by incubation of HeLa cells with Rose Bengal acetate-derivative (RBAc) followed by irradiation for a total dose of 1.6 J/cm2. This treatment was previously demonstrated to reduce cell viability under mild treatment conditions, suggesting the restoration of the photoactive molecule in particularly sensitive cell sites. Indeed, Rose Bengal (RB) is a very efficient photosensitizer, whose photophysical properties are inactivated by addition of the quencher group acetate. The RBAc behaves as a fluorogenic substrate by entering easily the cells where the original, photoactive molecule is restored by specific esterases. Different intracellular sites of photodamage of RB are present. In particular, fluorescence imaging of Rodamine 123 and JC-1 labelled cells showed altered morphology and loss of potential membrane of mitochondria. MTT and NR assays gave indications of alteration of mitochondrial and lysosomal enzyme activities. These damaged sites were likely responsible for triggering apoptosis. Significant amount of apoptotic cell death (about 40%) was induced after light irradiation followed RBAc incubation as revealed by morphological (modification of cell shape and blebs formation), cytochemical (FITC-Annexin-V positive cells) and nuclear fragmentation assays.

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

    PubMed

    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. PMID:27209523

  4. Fragrance material review on 3-phenylpropyl acetate.

    PubMed

    McGinty, D; Letizia, C S; Api, A M

    2012-09-01

    A toxicologic and dermatologic review of 3-phenylpropyl acetate when used as a fragrance ingredient is presented. 3-Phenylpropyl acetate is a member of the fragrance structural group Aryl Alkyl Alcohol Simple Acid Esters (AAASAE). The AAASAE fragrance ingredients are prepared by reacting an aryl alkyl alcohol with a simple carboxylic acid (a chain of 1-4 carbons) to generate formate, acetate, propionate, butyrate, isobutyrate and carbonate esters. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for 3-phenylpropyl acetate were evaluated, then summarized, and includes: physical properties, acute toxicity, skin irritation, skin sensitization, and toxicokinetics data. A safety assessment of the entire AAASAE will be published simultaneously with this document. Please refer to Belsito et al., 2012 for an overall assessment of the safe use of this material and all AAASAE in fragrances.

  5. Fragrance material review on anisyl acetate.

    PubMed

    McGinty, D; Letizia, C S; Api, A M

    2012-09-01

    A toxicologic and dermatologic review of anisyl acetate when used as a fragrance ingredient is presented. Anisyl acetate is a member of the fragrance structural group Aryl Alkyl Alcohol Simple Acid Esters (AAASAE). The AAASAE fragrance ingredients are prepared by reacting an aryl alkyl alcohol with a simple carboxylic acid (a chain of 1-4 carbons) to generate formate, acetate, propionate, butyrate, isobutyrate and carbonate esters. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for anisyl acetate were evaluated, then summarized, and includes: physical properties, skin irritation, skin sensitization, elicitation, and phototoxicity data. A safety assessment of the entire AAASAE will be published simultaneously with this document. Please refer to Belsito et al., 2012 for an overall assessment of the safe use of this material and all AAASAE in fragrances.

  6. Fragrance material review on piperonyl acetate.

    PubMed

    McGinty, D; Letizia, C S; Api, A M

    2012-09-01

    A toxicologic and dermatologic review of piperonyl acetate when used as a fragrance ingredient is presented. Piperonyl acetate is a member of the fragrance structural group aryl alkyl alcohol simple acid esters (AAASAE). The AAASAE fragrance ingredients are prepared by reacting an aryl alkyl alcohol with a simple carboxylic acid (a chain of 1-4 carbons) to generate formate, acetate, propionate, butyrate, isobutyrate and carbonate esters. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for piperonyl acetate were evaluated, then summarized, and includes: physical properties, acute toxicity, skin irritation, mucous membrane (eye) irritation, skin sensitization, toxicokinetics, and genotoxicity data. A safety assessment of the entire AAASAE will be published simultaneously with this document. Please refer to Belsito et al. (2012) for an overall assessment of the safe use of this material and all AAASAE in fragrances.

  7. Fragrance material review on 2-phenylpropyl acetate.

    PubMed

    McGinty, D; Letizia, C S; Api, A M

    2012-09-01

    A toxicologic and dermatologic review of 2-phenylpropyl acetate when used as a fragrance ingredient is presented. 2-Phenylpropyl acetate is a member of the fragrance structural group Aryl Alkyl Alcohol Simple Acid Esters (AAASAE). The AAASAE fragrance ingredients are prepared by reacting an aryl alkyl alcohol with a simple carboxylic acid (a chain of 1-4 carbons) to generate formate, acetate, propionate, butyrate, isobutyrate and carbonate esters. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for 2-phenylpropyl acetate were evaluated, then summarized, and includes: physical properties, acute toxicity, skin irritation, mucous membrane (eye) irritation, and skin sensitization data. A safety assessment of the entire AAASAE will be published simultaneously with this document. Please refer to Belsito et al. (2012) for an overall assessment of the safe use of this material and all AAASAE in fragrances.

  8. Fragrance material review on 4-methylbenzyl acetate.

    PubMed

    McGinty, D; Letizia, C S; Api, A M

    2012-09-01

    A toxicologic and dermatologic review of 4-methylbenzyl acetate when used as a fragrance ingredient is presented. 4-Methylbenzyl acetate is a member of the fragrance structural group Aryl Alkyl Alcohol Simple Acid Esters (AAASAE). The AAASAE fragrance ingredients are prepared by reacting an aryl alkyl alcohol with a simple carboxylic acid (a chain of 1-4 carbons) to generate formate, acetate, propionate, butyrate, isobutyrate and carbonate esters. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for 4-methylbenzyl acetate were evaluated, then summarized, and includes: physical properties, skin irritation, skin sensitization, and elicitation data. A safety assessment of the entire AAASAE will be published simultaneously with this document. Please refer to Belsito et al. (2012) for an overall assessment of the safe use of this material and all AAASAE in fragrances.

  9. Titan2D Based Pyroclastic Flows Hazard Maps for Santa Ana Volcano, El Salvador

    NASA Astrophysics Data System (ADS)

    Bajo, J. V.; Martinez-Hackert, B.; Escobar, C. D.; Gutierrez, R. E.

    2009-05-01

    Santa Ana Volcano is located in the Apaneca Volcanic Field located to the west of El Salvador, Central America. It is one the six active volcanoes monitor by the Servicios Nacionales de Estudios Territoriales (SNET) in El Salvador, out of twenty that are considered active in this small country by Smithsonian definition. The Santa Ana Volcano is surrounded by rural communities in its proximal areas and in its close distal areas by the second largest city of the country. On October 1st 2005, after a few months of increased fumarolic and seismic activity, it erupted generating a 10 km high steam and ash plume, reportedly seen by some aircraft and estimated using photography by SNET members. Ash was deposited to the west, north-west part of the country, following typical wind pattern for the region, as well as small pyroclastic flows and major lahars in its eastern part. Coffee plantations were lost, as was some crop of coffee in the following season. However, to the west the ash fertilized the land and resulted in an enhanced harvest of coffee beans. Only 2 people were killed from the Blast, thanks to the auto evacuation of proximal communities. Whilst the last eruption had a relatively low human life toll, a stronger eruption spells havoc almost certainly for the region. At this moment no exhaustive study and understanding exists of the pyroclastic flows generated by the Santa Ana Volcano nor a map for this particular hazard. This study proposes the use of Titan2D for those two purposes, using a DEM generated by the SNET using topographic maps as well as DEMs generated using Advanced Spaceborne Thermal Emission and Reflection Radiometer Images (ASTER).

  10. Aquatic assemblages of the highly urbanized Santa Ana River Basin, California

    USGS Publications Warehouse

    Brown, L.R.; Burton, C.A.; Belitz, K.

    2005-01-01

    We assessed the structure of periphyton, benthic macroinvertebrate, and fish assemblages and their associations with environmental variables at 17 sites on streams of the highly urbanized Santa Ana River basin in Southern California. All assemblages exhibited strong differences between highly urbanized sites in the valley and the least-impacted sites at the transition between the valley and undeveloped mountains. Results within the urbanized area differed among taxa. Periphyton assemblages were dominated by diatoms (>75% of total taxa). Periphyton assemblages within the urbanized area were not associated with any of the measured environmental variables, suggesting that structure of urban periphyton assemblages might be highly dependent on colonization dynamics. The number of Ephemeroptera, Trichoptera, and Plecoptera (EPT) taxa included in macroinvertebrate assemblages ranged from 0 to 6 at urbanized sites. Benthic macroinvertebrate assemblages had significant correlations with several environmental variables within the urban area, suggesting that stream size and permanence were important determinants of distribution among the species able to survive conditions in urban streams. Only 4 of 16 fish species collected were native to the drainage. Fish assemblages of urbanized sites included two native species, arroyo chub Gila orcuttii and Santa Ana sucker Catostomus santaanae, at sites that were intermediate in coefficient of variation of bank-full width, depth, bed substrate, and water temperature. Alien species dominated urbanized sites with lesser or greater values for these variables. These results suggest that urban streams can be structured to enhance populations of native fishes. Continued study of urban streams in the Santa Ana River basin and elsewhere will contribute to the basic understanding of ecological principles and help preserve the maximum ecological value of streams in highly urbanized areas.

  11. Audiomagnetotelluric exploration across the Waíanae Range, Óahu, Hawaíi

    NASA Astrophysics Data System (ADS)

    Sigurdardottir, T. D.; Thomas, D. M.; Wallin, E.; Winchester, C.; Sinton, J. M.

    2015-12-01

    The audiomagnetotelluric (AMT) method is capable of providing direct evidence of a geothermal resource within the extinct Waíanae volcano, Óahu, Hawaíi. Geothermal systems are becoming an increasingly important energy source worldwide. With electric energy costs in Hawaíi the most expensive in the US (30.54 cents/kWh), it is important to investigate the potential of local geothermal resources. Slightly elevated temperature and chloride concentrations, measured in the 1970's at wells in the upper Lualualei Valley indicate the possibility of a geothermal resource. Previous geophysical investigations: self-potential, rotating quadripole resistivity, and shallow soil temperature surveys in the caldera measured low resistivity values. Resistivity is related to rock characteristics (e.g., porosity, saturation, salinity, temperature, chemistry, and the presence of weathered minerals). We are investigating the area further using the AMT method. We have collected profiles of AMT measurements across the Lualualei Valley and the Waíanae caldera boundary. Anthropogenic noise and access in this area is problematic. Electrical noise, originating from power lines along roads and very low frequency radio towers in the vicinity, add noise to the data. Limited access to sites on military lands inhibit data collection. However, preliminary results show that we have successfully imaged the expected higher resistivity values as our profiles cross the mountains bounding the caldera. As data continue to be collected across the Waíanae Caldera and Range and we begin modeling our data in two dimensions, we expect to be able to identify water table elevations, detect lateral variability between salt and fresh water saturation, estimate thickness of the freshwater lens and depth to the transition zone, image fault structures at the caldera boundary, and with enough sensitivity to conductivity, we can identify regions of elevated temperature.

  12. Purification and Properties of an Esterase from the Yeast Saccharomyces cerevisiae and Identification of the Encoding Gene

    PubMed Central

    Degrassi, Giuliano; Uotila, Lasse; Klima, Raffaella; Venturi, Vittorio

    1999-01-01

    We purified an intracellular esterase that can function as an S-formylglutathione hydrolase from the yeast Saccharomyces cerevisiae. Its molecular mass was 40 kDa, as determined by gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isoelectric point was 5.0 by isoelectric focusing. The enzyme activity was optimal at 50°C and pH 7.0. The corresponding gene, YJLO68C, was identified by its N-terminal amino acid sequence and is not essential for cell viability. Null mutants have reduced esterase activities and grow slowly in the presence of formaldehyde. This enzyme may be involved in the detoxification of formaldehyde, which can be metabolized to S-formylglutathione by S. cerevisiae. PMID:10427036

  13. Glutaraldehyde cross-linking of immobilized thermophilic esterase on hydrophobic macroporous resin for application in poly(ε-caprolactone) synthesis.

    PubMed

    Wang, Min; Shi, Hui; Wu, Di; Han, Haobo; Zhang, Jianxu; Xing, Zhen; Wang, Shuang; Li, Quanshun

    2014-01-01

    The immobilized thermophilic esterase from Archaeoglobus fulgidus was successfully constructed through the glutaraldehyde-mediated covalent coupling after its physical adsorption on a hydrophobic macroporous resin, Sepabeads EC-OD. Through 0.05% glutaraldehyde treatment, the prevention of enzyme leaching and the maintenance of catalytic activity could be simultaneously realized. Using the enzymatic ring-opening polymerization of ε-caprolactone as a model, effects of organic solvents and reaction temperature on the monomer conversion and product molecular weight were systematically investigated. After the optimization of reaction conditions, products were obtained with 100% monomer conversion and Mn values lower than 1010 g/mol. Furthermore, the cross‑linked immobilized thermophilic esterase exhibited an excellent operational stability, with monomer conversion values exceeding 90% over the course of 12 batch reactions, still more than 80% after 16 batch reactions. PMID:25006789

  14. Directed evolution of an esterase: screening of enzyme libraries based on pH-indicators and a growth assay.

    PubMed

    Bornscheuer, U T; Altenbuchner, J; Meyer, H H

    1999-10-01

    In order to resolve a sterically hindered 3-hydroxy ethyl ester, which was not accepted as substrate by 20 wild-type hydrolases, a directed evolution of an esterase from Pseudomonas fluorescens (PFE) was performed. Mutations were introduced using the mutator strain Epicurian coli XL1-Red. Enzyme libraries derived from seven mutation cycles were assayed on minimal media agar plates supplemented with pH indicators (neutral red and crystal violet), thus allowing the identification of active esterase variants by the formation of a red color caused by a pH decrease due to the released acid. A further selection criteria was introduced by using the corresponding glycerol estar, because release of the carbon source glycerol facilitates growth on minimal media. By this strategy, one double mutant (A209D and L181V) of PFE was identified, which hydrolyzed the 3-hydroxy ethyl ester in a stereoselective manner (25% ee for the remaining ester, E approximate to 5).

  15. The Santa AnaWinds of Southern California in the context of Fire Weather

    NASA Astrophysics Data System (ADS)

    Cao, Yang

    The Santa Ana winds represent a high-impact weather event owing to the intimate relationship between the extremely dry, fast winds and the wildfire threat. The winds can be locally gusty, particularly in the complex terrain of San Diego county, where the airflow has characteristics of downslope windstorms. These winds can cause and/or rapidly spread wildfires, the threat of which is particularly acute during the autumn season before the onset of winter rains. It remains a day-to-day challenge to accurately predict wind gust speed, especially in the mountainous regions. Our study employs large physics ensembles composed of high-resolution simulations of severe downslope windstorms that involve an exhaustive examination of available model physical parameterizations. Model results are calibrated and validated against the San Diego Gas and Electric (SDG&E) mesonet observations, a dense, homogenous, and well-positioned network with uniform high quality. Results demonstrate model horizontal resolution, model physics, random perturbations and landuse database can have a material effect on the strength, location and timing of Santa Ana winds in real-data simulations. A large model physics ensemble reveals the land surface model to be most crucial in skillful wind predictions, which are particularly sensitive to the surface roughness length. A surprisingly simple gust parameterization is proposed for the San Diego network, based on the discovery that this homogeneous mesonet has a nearly invariant network-averaged gust factor. The gust forecast technique is of special interest in the context of routine weather combined with atmospheric humidity and fuel moisture information. A real-time wildfire threat warning system, the Santa Ana Wildfire Threat Index (SAWTI), has been developed to effectively communicate the upcoming Santa Ana wind strength with respect to the anticipated fire danger to first responders and the public. In addition to the wind and gust forecast techniques

  16. Mate preference in wild and domesticated (game-farm) mallards (Anas platyrhynchos): I. Initial preference

    USGS Publications Warehouse

    Cheng, K.M.; Shoffner, R.N.; Phillips, R.E.; Lee, F.B.

    1978-01-01

    Wild and game-farm mallards (Anas platyrhynchos) raised in pure strain and mixed groups were tested for initial mate preference in a choice test. Female mallards showed no significant preference but males of either strain raised with females of their own strain significantly preferred female models of their own strain during the test. Males raised with females of the other strain merely showed attenuation of their preference for female models of their own strain and did not show preference for female models of the other strain. Game-farm mallards approached models significantly sooner than wild mallards and there was a significant sex X mate interaction.

  17. Survival of female black ducks, Anas rubripes, during the breeding season

    USGS Publications Warehouse

    Ringelman, J.K.; Longcore, J.R.

    1983-01-01

    The Mayfield method was used to estimate the survival rate of 19 radio-marked, female Black Ducks (Anas rubripes) in southcentral Maine during 1977-80. An overall survival rate of 0.74 was estimated for the 121-day monitoring period that included the pre-laying and laying, incubation, brood rearing, and post-rearing stages. No differences in survival rates were detected among these stages. Two instrumented hens were killed by Red-shouldered Hawks (Buteo lineatus) and a third was killed by an unknown predator. We found no evidence that the attachment of radio transmitters affected hen survival.

  18. Dynamics of a thermo-Mediterranean coastal environment the Coto Doñana National Park

    NASA Astrophysics Data System (ADS)

    Finlayson, Geraldine; Finlayson, Clive; Espejo, J. M. Recio

    2008-11-01

    Using data collected from existing habitats found in the southwestern Iberian Peninsula, and including data collected in the Biological Reserve of the Parque Nacional de Doñana, Spain, this study considers the seasonal and inter-annual variability of a thermo-Mediterranean, subhumid, environment and the significance of the presence of surface water in the system. This extant environment is then used together with the fossil record from Gorham's Cave in Gibraltar, where Neanderthals lived, as a proxy for the ecology of the emerged landscape outside the cave.

  19. Early imprinting in wild and game-farm mallards (Anas platyrhynchos): genotype and arousal

    USGS Publications Warehouse

    Cheng, K.M.; Shoffner, R.N.; Phillips, R.E.; Shapiro, L.J.

    1979-01-01

    Early imprinting was studied under laboratory conditions in five lines of mallards (Anas platyrhynchos) with different degrees of wildness obtained through pedigreed breeding. Data were analyzed by the least squares method. Wild ducklings imprinted better than game-farm (domesticated) ducklings, and heterosis was demonstrated to exist in imprinting traits. Nonadditive genetic variations and genotype-environmental interactions are discussed as possible causes for the heterosis observed. Differences in imprinting between genetic lines are attributed, at least partly, to differences in arousal level during the ducklings' first exposure to the imprinting stimulus.

  20. Is the ANA guilty of paternalism in its guidelines on withdrawing or withholding food and fluid?

    PubMed

    Light, K; Connelly, R

    1989-01-01

    The ANA Committee on Ethics's Guidelines on Withdrawing or Withholding Food and Fluid (1988) delineates those circumstances under which withholding is normally permissible. The Guidelines direct nurses to respect the wishes of competent adults. What of adults who are considered incompetent? Can they make decisions to refuse food and fluids? The Guidelines ask nurses to judge patient competence and the merit of the reasons behind the decisions. This suggests paternalism. A more appropriate role of the nurse might be to focus on helping the patient to make a decision, rather than on the merit of the decision itself.

  1. NURSING EMERGING. ANA Nursing: Scope and Standards of Practice, (2015) 3rd Edition.

    PubMed

    Mariano, Carla

    2016-04-01

    AHNA Past-President Carla Mariano recently had the privilege of serving on the American Nurses Association's (ANA) Nursing Scope and Standards Revision Workgroup. Representing the specialty practice of holistic nursing, Carla's presence within this workgroup contributed greatly to the inclusion of holistic principles and values throughout the new 2015 Nursing: Scope and Standards of Practice, 3rd edition, the foundational document that informs and guides professional nursing practice within the United States. This is a significant step forward for holistic nursing and an indicator of our growing influence as specialty practice. PMID:27305802

  2. Acetate limitation and nitrite accumulation during denitrification

    SciTech Connect

    Oh, J.; Silverstein, J.

    1999-03-01

    Nitrite accumulated in denitrifying activated sludge mixed liquor when the carbon and electron source, acetate, was limited. If acetate was added to obtain a carbon-to-nitrogen (C:N) ratio in the range of 2:1 to 3:1, nitrate was completely consumed at the same rate with no nitrite accumulation, indicating that nitrate concentration controlled the respiration rate as long as sufficient substrate was present. However, when acetate was reduced to a C:N ratio of 1:1, while nitrate continued to be consumed, > 50% of the initial nitrate-nitrogen accumulated as nitrite and 29% persisted as nitrite throughout an endogenous denitrification period of 8--9 h. While nitrite accumulated during acetate-limited denitrification, the specific nitrate reduction rate increased significantly compared with the rate when excess acetate was provided as follows: 0.034 mg-NO{sub 3}-N/mg-mixed liquid volatile suspended solids/h versus 0.023 mg-NO{sub 3}-N/mg-mixed liquid volatile suspended solids/h, respective. This may be explained by nitrate respiration out-competing nitrite respiration for limited acetate electrons. Complete restoration of balanced denitrification and elimination of nitrite accumulation during denitrification required several weeks after the C:N ratio was increased back to 2:1.

  3. Dehydrogenases, Acid and Alkaline Phosphatases, and Esterases for Chemotaxonomy of Selected Meloidogyne, Ditylenchus, Heterodera and Aphelenchus spp.

    PubMed Central

    Dickson, D. W.; Huisingh, D.; Sasser, J. N.

    1971-01-01

    Various taxonomically useful profiles of four dehydrogenases (lactate, malate, glucose-6-phosphate, and a-glycerophosphate) and three hydrolases (acid and alkaline phosphatase and esterase) were detected in whole nematode homogenates of Meloidogyne javanica, M. hapla, M. incognita, M. arenaria, Ditylenchus dipsaci, D. triformis, Heterodera glycines, and Aphelenchus avenae. The enzyme profiles were stable in populations cultured on several different hosts. A tentative enzymically-determined phylogeny of Meloidogyne is given. PMID:19322334

  4. Identification and characterization of an esterase involved in malathion resistance in the head louse Pediculus humanus capitis.

    PubMed

    Kwon, Deok Ho; Kim, Ju Hyeon; Kim, Young Ho; Yoon, Kyong Sup; Clark, J Marshall; Lee, Si Hyeock

    2014-06-01

    Enhanced malathion carboxylesterase (MCE) activity was previously reported to be involved in malathion resistance in the head louse Pediculus humanus capitis (Gao et al., 2006 [8]). To identify MCE, the transcriptional profiles of all five esterases that had been annotated to be catalytically active were determined and compared between the malathion-resistant (BR-HL) and malathion-susceptible (KR-HL) strains of head lice. An esterase gene, designated HLCbE3, exhibited approximately 5.4-fold higher transcription levels, whereas remaining four esterases did not exhibit a significant increase in their transcription in BR-HL, indicating that HLCbE3 may be the putative MCE. Comparison of the entire cDNA sequences of HLCbE3 revealed no sequence differences between the BR-HL and KR-HL strains and suggested that no single nucleotide polymorphism is associated with enhanced MCE activity. Two copies of the HLCbE3 gene were observed in BR-HL, implying that the over-transcription of HLCbE3 is due to the combination of a gene duplication and up-regulated transcription. Knockdown of HLCbE3 expression by RNA interference in the BR-HL strain led to increases in malathion susceptibility, confirming the identity of HLCbE3 as a MCE responsible for malathion resistance in the head louse. Phylogenetic analysis suggested that HLCbE3 is a typical dietary esterase and belongs to a clade containing various MCEs involved in malathion resistance. PMID:24974112

  5. Exploitation of heterosis for carotenoid content in African marigold (Tagetes erecta L.) and its correlation with esterase polymorphism.

    PubMed

    Sreekala, C; Raghava, S P S

    2003-02-01

    African marigold (Tagetes erecta L.), a major source of carotenoids, is also grown as a cut flower and a garden flower in addition to being grown for its medicinal values. We studied gene action, combining ability and heterosis, aiming at genetic improvement of T. erecta for enhanced carotenoid content in petals, and report for the first time that heterosis can be exploited for total carotenoids and its commercially important fractions. Total content of carotenoids and lutein appears to be governed by dominance (or non-additive) gene action, while content of xanthophyll esters is governed by both additive and dominance (or non-additive) gene actions. Specific combining ability variance was predominant for all the three traits. General and specific combining abilities and heterosis were highly significant. Heterobeltiosis was also positive. General combining ability (GCA) variances were not significantly correlated to performance per se. There was also no correlation between performance per se of normal petalled pollen parents and the performance of crosses made between male-sterile (female) and male-fertile (pollen) parents. These findings suggest that carotenoid content should not be the only criterion considered in the selection of parental lines. Studies on esterase in seeds and peroxidase in seedlings revealed a relatively high level of polymorphism in esterase with a total of 14 isoforms, whereas peroxidase showed low polymorphism. Similarity indices between different parental combinations, calculated based on seed esterase polymorphism, showed a significant negative correlation (r = -0.479, P = 0.05) with heterosis for carotenoid content. This indicates that the selection of parents with wider variation in their esterase profiles may possibly be exploited for genetic enhancement of carotenoids in T. erecta.

  6. Identification and characterization of a periplasmic trilactone esterase, Cee, revealed unique features of ferric enterobactin acquisition in Campylobacter

    PubMed Central

    Zeng, Ximin; Mo, Yiming; Xu, Fuzhou; Lin, Jun

    2012-01-01

    Summary Ferric enterobactin (FeEnt) acquisition is a highly efficient and conserved iron scavenging system in Gram-negative bacteria. Recently, we have characterized two FeEnt receptors (CfrA and CfrB) in Campylobacter jejuni and C. coli, the enteric human pathogens that do not produce any siderophores. In this study, whole genome sequencing and comparative genomic analysis identified a unique Ent trilactone esterase Cee (Cj1376) in C. jejuni. Genomic analysis and biochemical assay strongly suggested that Cee is the sole trilactone esterase in C. jejuni. Thin layer chromatography and HPLC analyses showed high efficiency of the purified Cee to hydrolyze Ent. Three Cee homologs previously characterized from other bacteria (IroE, IroD, and Fes) were also purified and analyzed together with Cee, indicating that Cee, Fes, and IroD displayed similar hydrolysis dynamics for both apo and ferric forms of Ent while IroE catalyzed Ent inefficiently. Unlike cytoplasmic Fes and IroD, Cee is localized in the periplasm as demonstrated by immunoblotting using Cee-specific antibodies. Genetic manipulation of diverse Campylobacter strains demonstrated that Cee is not only essential for CfrB-dependent FeEnt acquisition but also involved in CfrA-dependent pathway. Together, this study identified and characterized a novel periplasmic trilactone esterase and suggested a new model of FeEnt acquisition in Campylobacter. PMID:23278903

  7. Est16, a New Esterase Isolated from a Metagenomic Library of a Microbial Consortium Specializing in Diesel Oil Degradation

    PubMed Central

    Pereira, Mariana Rangel; Mercaldi, Gustavo Fernando; Maester, Thaís Carvalho; Balan, Andrea; de Macedo Lemos, Eliana Gertrudes

    2015-01-01

    Lipolytic enzymes have attracted attention from a global market because they show enormous biotechnological potential for applications such as detergent production, leather processing, cosmetics production, and use in perfumes and biodiesel. Due to the intense demand for biocatalysts, a metagenomic approach provides methods of identifying new enzymes. In this study, an esterase designated as Est16 was selected from 4224 clones of a fosmid metagenomic library, revealing an 87% amino acid identity with an esterase/lipase (accession number ADM63076.1) from an uncultured bacterium. Phylogenetic studies showed that the enzyme belongs to family V of bacterial lipolytic enzymes and has sequence and structural similarities with an aryl-esterase from Pseudomonas fluorescens and a patented Anti-Kazlauskas lipase (patent number US20050153404). The protein was expressed and purified as a highly soluble, thermally stable enzyme that showed a preference for basic pH. Est16 exhibited activity toward a wide range of substrates and the highest catalytic efficiency against p-nitrophenyl butyrate and p-nitrophenyl valerate. Est16 also showed tolerance to the presence of organic solvents, detergents and metals. Based on molecular modeling, we showed that the large alpha-beta domain is conserved in the patented enzymes but not the substrate pocket. Here, it was demonstrated that a metagenomic approach is suitable for discovering the lipolytic enzyme diversity and that Est16 has the biotechnological potential for use in industrial processes. PMID:26214846

  8. Est16, a New Esterase Isolated from a Metagenomic Library of a Microbial Consortium Specializing in Diesel Oil Degradation.

    PubMed

    Pereira, Mariana Rangel; Mercaldi, Gustavo Fernando; Maester, Thaís Carvalho; Balan, Andrea; Lemos, Eliana Gertrudes de Macedo

    2015-01-01

    Lipolytic enzymes have attracted attention from a global market because they show enormous biotechnological potential for applications such as detergent production, leather processing, cosmetics production, and use in perfumes and biodiesel. Due to the intense demand for biocatalysts, a metagenomic approach provides methods of identifying new enzymes. In this study, an esterase designated as Est16 was selected from 4224 clones of a fosmid metagenomic library, revealing an 87% amino acid identity with an esterase/lipase (accession number ADM63076.1) from an uncultured bacterium. Phylogenetic studies showed that the enzyme belongs to family V of bacterial lipolytic enzymes and has sequence and structural similarities with an aryl-esterase from Pseudomonas fluorescens and a patented Anti-Kazlauskas lipase (patent number US20050153404). The protein was expressed and purified as a highly soluble, thermally stable enzyme that showed a preference for basic pH. Est16 exhibited activity toward a wide range of substrates and the highest catalytic efficiency against p-nitrophenyl butyrate and p-nitrophenyl valerate. Est16 also showed tolerance to the presence of organic solvents, detergents and metals. Based on molecular modeling, we showed that the large alpha-beta domain is conserved in the patented enzymes but not the substrate pocket. Here, it was demonstrated that a metagenomic approach is suitable for discovering the lipolytic enzyme diversity and that Est16 has the biotechnological potential for use in industrial processes.

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

    PubMed

    Panagiotou, Gianni; Olavarria, Reyes; Olsson, Lisbeth

    2007-06-30

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

  10. Immobilization of thermoalkalophilic recombinant esterase enzyme by entrapment in silicate coated Ca-alginate beads and its hydrolytic properties.

    PubMed

    Gülay, Seçkin; Şanlı-Mohamed, Gülşah

    2012-04-01

    Thermoalkalophilic esterase enzyme from Balçova (Agamemnon) geothermal site were aimed to be immobilized effectively via a simple and cost-effective protocol in silicate coated Calcium alginate (Ca-alginate) beads by entrapment. The optimal immobilization conditions of enzyme in Ca-alginate beads were investigated and obtained with 2% alginate using 0.5mg/ml enzyme and 0.7 M CaCl(2) solution. In order to prevent enzyme from leaking out of the gel beads, Ca-alginate beads were then coated with silicate. Enzyme loading efficiency and immobilization yield for silicate coated beads was determined as 98.1% and 71.27%, respectively and compared with non-coated ones which were 68.5% and 45.80%, respectively. Surface morphologies, structure and elemental analysis of both silicate coated and non-coated alginate beads were also compared using Fourier Transform Infrared Spectroscopy (FT-IR) and Scanning Electron Microscope (SEM) equipped with Energy-dispersive X-ray spectroscopy (EDX). Moreover, silicate coated alginate beads enhanced reusability of esterase in continuous processes compared to non-coated beads. The hydrolytic properties of free and immobilized enzyme in terms of storage and thermal stability as well as the effects of the temperature and pH were determined. It was observed that operational, thermal and storage stabilities of the esterase were increased with immobilization. PMID:22309712

  11. Immobilization of thermoalkalophilic recombinant esterase enzyme by entrapment in silicate coated Ca-alginate beads and its hydrolytic properties.

    PubMed

    Gülay, Seçkin; Şanlı-Mohamed, Gülşah

    2012-04-01

    Thermoalkalophilic esterase enzyme from Balçova (Agamemnon) geothermal site were aimed to be immobilized effectively via a simple and cost-effective protocol in silicate coated Calcium alginate (Ca-alginate) beads by entrapment. The optimal immobilization conditions of enzyme in Ca-alginate beads were investigated and obtained with 2% alginate using 0.5mg/ml enzyme and 0.7 M CaCl(2) solution. In order to prevent enzyme from leaking out of the gel beads, Ca-alginate beads were then coated with silicate. Enzyme loading efficiency and immobilization yield for silicate coated beads was determined as 98.1% and 71.27%, respectively and compared with non-coated ones which were 68.5% and 45.80%, respectively. Surface morphologies, structure and elemental analysis of both silicate coated and non-coated alginate beads were also compared using Fourier Transform Infrared Spectroscopy (FT-IR) and Scanning Electron Microscope (SEM) equipped with Energy-dispersive X-ray spectroscopy (EDX). Moreover, silicate coated alginate beads enhanced reusability of esterase in continuous processes compared to non-coated beads. The hydrolytic properties of free and immobilized enzyme in terms of storage and thermal stability as well as the effects of the temperature and pH were determined. It was observed that operational, thermal and storage stabilities of the esterase were increased with immobilization.

  12. Recombinant sterol esterase from Ophiostoma piceae: an improved biocatalyst expressed in Pichia pastoris

    PubMed Central

    2012-01-01

    Background The ascomycete Ophiostoma piceae produces a sterol esterase (OPE) with high affinity towards p-nitrophenol, glycerol and sterol esters. Its hydrolytic activity on natural mixtures of triglycerides and sterol esters has been proposed for pitch biocontrol in paper industry since these compounds produce important economic losses during paper pulp manufacture. Results Recently, this enzyme has been heterologously expressed in the methylotrophic yeast Pichia pastoris, and the hydrolytic activity of the recombinant protein (OPE*) studied. After the initial screening of different clones expressing the enzyme, only one was selected for showing the highest production rate. Different culture conditions were tested to improve the expression of the recombinant enzyme. Complex media were better than minimal media for production, but in any case the levels of enzymatic activity were higher (7-fold in the best case) than those obtained from O. piceae. The purified enzyme had a molecular mass of 76 kDa, higher than that reported for the native enzyme under SDS-PAGE (60 kDa). Steady-state kinetic characterization of the recombinant protein showed improved catalytic efficiency for this enzyme as compared to the native one, for all the assayed substrates (p-nitrophenol, glycerol, and cholesterol esters). Different causes for this were studied, as the increased glycosylation degree of the recombinant enzyme, their secondary structures or the oxidation of methionine residues. However, none of these could explain the improvements found in the recombinant protein. N-terminal sequencing of OPE* showed that two populations of this enzyme were expressed, having either 6 or 8 amino acid residues more than the native one. This fact affected the aggregation behaviour of the recombinant protein, as was corroborated by analytical ultracentrifugation, thus improving the catalytic efficiency of this enzyme. Conclusion P. pastoris resulted to be an optimum biofactory for the

  13. Clinical evaluation of the RapID-ANA II panel for identification of anaerobic bacteria.

    PubMed

    Celig, D M; Schreckenberger, P C

    1991-03-01

    The accuracy of the RapID-ANA II system (Innovative Diagnostic Systems, Inc., Atlanta, Ga.) was evaluated by comparing the results obtained with that system with results obtained by the methods described by the Virginia Polytechnic Institute and State University. Three hundred anaerobic bacteria were tested, including 259 clinical isolates and 41 stock strains of anaerobic microorganisms representing 16 genera and 48 species. When identifications to the genus level only were included, 96% of the anaerobic gram-negative bacilli, 94% of the Clostridium species, 83% of the anaerobic, nonsporeforming, gram-positive bacilli, and 97% of the anaerobic cocci were correctly identified. When correct identifications to the genus and species levels were compared, 86% of 152 anaerobic gram-negative bacilli, 76% of 34 Clostridium species, 81% of 41 anaerobic, nonsporeforming, gram-positive bacilli, and 97% of 73 anaerobic cocci were correctly identified. Eight isolates (3%) produced inadequate identification in which the correct identification was listed with one or two other possible choices and extra tests were required for separation. A total of 9 isolates (3%) were misidentified by the RapID-ANA II panel. Overall, the system was able to correctly identify 94% of all the isolates to the genus level and 87% of the isolates to the species level in 4 h by using aerobic incubation.

  14. Cytochrome c Biogenesis Genes Involved in Arsenate Respiration by Shewanella trabarsenatis ANA-3

    NASA Astrophysics Data System (ADS)

    Newman, D. K.

    2002-12-01

    Arsenate can be used as a terminal electron acceptor in anaerobic respiration by diverse bacteria. The detection of these bacteria in numerous contaminated environments suggests that they are widespread and metabolically active in nature. Arsenate-respiring bacteria have been implicated in the mobilization of arsenic from arsenic-contaminated sediments. However, the enzymatic mechanisms supporting arsenate respiration are largely unknown. Here, we describe c-type cytochromes that are involved in arsenate respiration by the bacterium Shewanella trabarsenatis strain ANA-3, a facultative anaerobe that is able to use a variety of electron acceptors for growth. We performed transposon mutagenesis to study the electron transport pathway in ANA-3 during arsenate respiration. 10 arsenate-respiration deficient mutants were found after screening up to 7,000 mutants, and 4 were shown to have unique transposon insertions through Southern Blot analysis. The physiological properties of these mutants were determined, including characterization of their growth on different electron acceptors. The genes flanking the transposon insertions were sequenced for each mutant, and several were found to encode c-type cytochrome biogenesis genes. UV/VIS spectra and SDS/PAGE were used to confirm the absence of c-type cytochromes in the mutants. Based on these findings, we proposed a model for respiratory electron transport to arsenate.

  15. Ground-water geology of the coastal zone, Long Beach-Santa Ana area, California

    USGS Publications Warehouse

    Poland, J.F.; Piper, A.M.

    1956-01-01

    This paper is the first chapter of a comprehensive report on the ground-water features in the southern part of the coastal plain in Los Angeles and Orange Counties, Calif., with special reference to the effectiveness of the so-called coastal barrier--the Newport-Inglewood structural zone--in restraining landwar,-1 movement of saline water. The coastal plain in Los Angeles and Orange Counties, which covers some 775 square miles, sustains a large urban and rural population, diverse industries, and intensive agricultural developments. The aggregate ground-water withdrawal in 1945 was about 400,000 acre-feet a year, an average of about 360 million gallons a day. The dominant land-form elements are a central lowland plain with tongues extending to the coast, bordering highlands and foothills, and a succession of low hills and mesas aligned northwestward along the coastal edge of the central low- land plain. These low hills and mesas are the land-surface expression of geologic structure in the Newport-Inglewood zone. The highland areas that border the inland edge of the coastal plain are of moderate altitude and relief; most of the ridge crests range from 1,400 to 2,500 feet in altitude, but Santiago Peak in the Santa Ana Mountains attains a height of 5,680 feet above sea level. From these highlands the land surface descends across foothills and aggraded alluvial aprons to the central lowland, Downey Plain, here defined as the surface formed by alluvial aggradation during the post-Pleistocene time of rising base level. The Newport-Inglewood belt of hills and plains (mesas) has a maximum relief of some 500 feet but is widely underlain at a depth of about 30 feet by a surface of marine plantation. As initially formed in late Pleistocene time that surface was largely a featureless plain. Thus the present land-surface forms within the Newport-Inglewood belt measure the earth deformation that has occurred there since late Pleistocene time and so are pertinent with respect to

  16. D-penicillamine-induced ANA (+) ANCA (+) vasculitis in pediatric patients with Wilson's disease.

    PubMed

    Lee, Yeonhee; Lee, Sang Taek; Cho, Heeyeon

    2016-05-01

    Anti-neutrophil cytoplasmic antibodies (ANCA) are associated with systemic vasculitis. The pathophysiology of ANCA-associated vasculitis (AAV) has not been clearly proven, and drug-induced ANCA-associated vasculitis has been reported. Wilson's disease is an inborn error of copper metabolism caused by a mutation in the copper transporting gene ATP7B, and traditional treatment is based on copper chelation with agents such as D-penicillamine. There have been rare reports that prolonged D-penicillamine therapy might cause adverse renal events such as membranous nephropathy and minimal change disease, but it is questionable if D-penicillamine induces ANCA-associated vasculitis. We describe 2 patients with Wilson's disease treated with D-penicillamine who presented with ANCA (+) vasculitis and renal involvement. The 2 patients also showed positive results for antinuclear antibody (ANA). Their kidney biopsy findings were compatible with crescentic/necrotizing glomerulonephritis, pauci-immune type. After diagnosis of AAV, D-penicillamine was stopped. Patients were then treated with plasmapheresis and immunosuppressants, including methylprednisolone pulse therapy and intravenous cyclophosphamide. One patient progressed to end-stage renal disease and the other showed persistent proteinuria. These cases suggest that D-penicillamine may induce ANA (+) ANCA (+) vasculitis with severe renal involvement in pediatric patients, and plasmapheresis combined with immunosuppressant should be considered.

  17. D-penicillamine-induced ANA (+) ANCA (+) vasculitis in pediatric patients with Wilson's disease.

    PubMed

    Lee, Yeonhee; Lee, Sang Taek; Cho, Heeyeon

    2016-05-01

    Anti-neutrophil cytoplasmic antibodies (ANCA) are associated with systemic vasculitis. The pathophysiology of ANCA-associated vasculitis (AAV) has not been clearly proven, and drug-induced ANCA-associated vasculitis has been reported. Wilson's disease is an inborn error of copper metabolism caused by a mutation in the copper transporting gene ATP7B, and traditional treatment is based on copper chelation with agents such as D-penicillamine. There have been rare reports that prolonged D-penicillamine therapy might cause adverse renal events such as membranous nephropathy and minimal change disease, but it is questionable if D-penicillamine induces ANCA-associated vasculitis. We describe 2 patients with Wilson's disease treated with D-penicillamine who presented with ANCA (+) vasculitis and renal involvement. The 2 patients also showed positive results for antinuclear antibody (ANA). Their kidney biopsy findings were compatible with crescentic/necrotizing glomerulonephritis, pauci-immune type. After diagnosis of AAV, D-penicillamine was stopped. Patients were then treated with plasmapheresis and immunosuppressants, including methylprednisolone pulse therapy and intravenous cyclophosphamide. One patient progressed to end-stage renal disease and the other showed persistent proteinuria. These cases suggest that D-penicillamine may induce ANA (+) ANCA (+) vasculitis with severe renal involvement in pediatric patients, and plasmapheresis combined with immunosuppressant should be considered. PMID:26784915

  18. Separating esterase targets of organophosphorus compounds in the brain by preparative chromatography.

    PubMed

    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.

  19. Evolutionary endocrinology of juvenile hormone esterase in Gryllus assimilis: direct and correlated responses to selection.

    PubMed

    Zera, A J; Zhang, C

    1995-11-01

    Hemolymph juvenile hormone esterase (JHE) activity on the third day of the last stadium in the cricket, Gryllus assimilis, exhibited a significant response to selection in each of six replicate lines. Mean realized heritability was 0.26 +/- 0.04. The response was due to changes in whole-organism enzyme activity as well as to changes in the proportion of enzyme allocated to the hemolymph compartment. In vivo juvenile hormone metabolism differed between some lines selected for high vs. low enzyme activity. Only minimal differences were observed between lines with respect to hemolymph protein concentration or whole-cricket activity of juvenile hormone epoxide hydrolase, the other major JH-degrading enzyme. Dramatic correlated responses to selection, equal in magnitude to the direct response, were observed for JHE activity on each of three other days of the last juvenile stadium. In contrast, no correlated responses in JHE activity were observed in adults. This indicates that JHE activities throughout the last stadium will evolve as a highly correlated unit independent of adult activities and the evolution of endocrine mechanisms regulating juvenile development can be decoupled from those controlling adult reproduction. This study represents the first quantitative-genetic analysis of naturally occurring endocrine variation in an insect species.

  20. Subunit Stabilization and Polyethylene Glycolation of Cocaine Esterase Improves In Vivo Residence TimeS⃞

    PubMed Central

    Narasimhan, Diwahar; Collins, Gregory T.; Nance, Mark R.; Nichols, Joseph; Edwald, Elin; Chan, Jimmy; Ko, Mei-Chuan; Woods, James H.; Tesmer, John J. G.

    2011-01-01

    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. PMID:21890748

  1. Bacterial cocaine esterase: a protein-based therapy for cocaine overdose and addiction

    PubMed Central

    Narasimhan, Diwahar; Woods, James H; Sunahara, Roger K

    2012-01-01

    Cocaine is highly addictive and there are no pharmacotherapeutic drugs available to treat acute cocaine toxicity or chronic abuse. Antagonizing an inhibitor such as cocaine using a small molecule has proven difficult. The alternative approach is to modify cocaine’s pharmacokinetic properties by sequestering or hydrolyzing it in serum and limiting access to its sites of action. We took advantage of a bacterial esterase (CocE) that has evolved to hydrolyze cocaine and have developed it as a therapeutic that rapidly and specifically clears cocaine from the subject. Native enzyme was unstable at 37°C, thus limiting CocE’s potential. Innovative computational methods based on the protein’s structure helped elucidate its mechanism of destabilization. Novel protein engineering methodologies were applied to substantially improve its stability in vitro and in vivo. These improvements rendered CocE as a powerful and efficacious therapeutic to treat cocaine intoxication and lead the way towards developing a therapy for addiction. PMID:22300094

  2. Discovery of potent carbonic anhydrase and acetylcholine esterase inhibitors: novel sulfamoylcarbamates and sulfamides derived from acetophenones.

    PubMed

    Akıncıoğlu, Akın; Akıncıoğlu, Hülya; Gülçin, İlhami; Durdagi, Serdar; Supuran, Claudiu T; Göksu, Süleyman

    2015-07-01

    In this study, several novel sulfamides were synthesized and evaluated for their acetylcholine esterase (AChE) and human carbonic anhydrase I, and II isoenzymes (hCA I and II) inhibition profiles. Reductive amination of methoxyacetophenones was used for the synthesis of amines. Amines were converted to sulfamoylcarbamates with chlorosulfonyl isocyanate (CSI) in the presence of BnOH. Pd-C catalyzed hydrogenolysis of sulfamoylcarbamates afforded sulfamides. These novel compounds were good inhibitors of the cytosolic hCA I, and hCA II with Ki values in the range of 45.9±8.9-687.5±84.3 pM for hCA I, and 48.80±8.2-672.2±71.9pM for hCA II. The inhibitory effects of the synthesized novel compounds on AChE were also investigated. The Ki values of these compounds were in the range of 4.52±0.61-38.28±6.84pM for AChE. These results show that hCA I, II, and AChE were effectively inhibited by the novel sulfamoylcarbamates 17-21 and sulfamide derivatives 22-26. All investigated compounds were docked within the active sites of the corresponding enzymes revealing the reasons of the effective inhibitory activity. PMID:25921269

  3. Are PECTIN ESTERASE INHIBITOR Genes Involved in Mediating Resistance to Rhynchosporium commune in Barley?

    PubMed Central

    Marzin, Stephan; Hanemann, Anja; Sharma, Shailendra; Hensel, Götz; Kumlehn, Jochen; Schweizer, Günther; Röder, Marion S.

    2016-01-01

    A family of putative PECTIN ESTERASE INHIBITOR (PEI) genes, which were detected in the genomic region co-segregating with the resistance gene Rrs2 against scald caused by Rhynchosporium commune in barley, were characterized and tested for their possible involvement in mediating resistance to the pathogen by complementation and overexpression analysis. The sequences of the respective genes were derived from two BAC contigs originating from the susceptible cultivar ‘Morex’. For the genes HvPEI2, HvPEI3, HvPEI4 and HvPEI6, specific haplotypes for 18 resistant and 23 susceptible cultivars were detected after PCR-amplification and haplotype-specific CAPS-markers were developed. None of the tested candidate genes HvPEI2, HvPEI3 and HvPEI4 alone conferred a high resistance level in transgenic over-expression plants, though an improvement of the resistance level was observed especially with OE-lines for gene HvPEI4. These results do not confirm but also do not exclude an involvement of the PEI gene family in the response to the pathogen. A candidate for the resistance gene Rrs2 could not be identified yet. It is possible that Rrs2 is a PEI gene or another type of gene which has not been detected in the susceptible cultivar ‘Morex’ or the full resistance reaction requires the presence of several PEI genes. PMID:26937960

  4. Isolation and characterization of an enzyme with esterase activity from Micropolyspora faeni.

    PubMed Central

    Bannerman, E N; Nicolet, J

    1976-01-01

    The isolation and the characterization of one of the enzymes of Micropolyspora faeni that hydrolyzes the substrate N-benzoyl-DL-phenylalanine-beta-naphthyl ester and that seems to be of medical importance are described. This enzyme (enzyme 1) was isolated with an 86-fold purification by using the following seven steps: ammonium sulfate precipitation, gel filtration through Sephadex G-150, heat treatment, chromatography on diethylaminoethyl-cellulose, rechromatography on diethylaminoethyl-Sephadex, gel filtration through Sephadex G-200, and affinity chromatography. Enzyme 1 has a molecular weight of approximately 500,000 and maximum activity at pH 7.8 to 8.0 and at 20 degrees C. The enzyme is stable between pH 7.5 and 10.5 and at temperatures up to 60 degrees C. Its activity is not inhibited by ethylenediaminetetraacetic acid. It is, however, sensitive to diisopropyl phosphofluoride and phenylmethyl sulfonyl fluoride. These properties and the ability to hydrolyze the esters of phenylalanine, tyrosine, and tryptophan without endopeptidasic activity and no marked proteolytic activity suggest that the enzyme is an esterase. Images PMID:9899

  5. Characterization of a chimeric enzyme comprising feruloyl esterase and family 42 carbohydrate-binding module.

    PubMed

    Koseki, Takuya; Mochizuki, Keiji; Kisara, Hiroe; Miyanaga, Akimasa; Fushinobu, Shinya; Murayama, Tetsuya; Shiono, Yoshihito

    2010-03-01

    We engineered a chimeric enzyme (AwFaeA-CBM42) comprising of type-A feruloyl esterase from Aspergillus awamori (AwFaeA) and family 42 carbohydrate-binding module (AkCBM42) from glycoside hydrolase family 54 alpha-L-arabinofuranosidase of Aspergillus kawachii. The chimeric enzyme was successfully produced in Pichia pastoris and accumulated in the culture broth. The purified chimeric enzyme had an apparent relative molecular mass (M(r)) of 53,000. The chimeric enzyme binds to arabinoxylan; this indicates that the AkCBM42 in AwFaeA-CBM42 binds to arabinofuranose side chain moiety of arabinoxylan. The thermostability of the chimeric enzyme was greater than that of AwFaeA. No significant difference of the specific activity toward methyl ferulate was observed between the AwFaeA and chimeric enzyme, but the release of ferulic acid from insoluble arabinoxylan by the chimeric enzyme was approximately 4-fold higher than that achieved by AwFaeA alone. In addition, the chimeric enzyme and xylanase acted synergistically for the degradation of arabinoxylan. In conclusion, the findings of our study demonstrated that the components of the AwFaeA-CBM42 chimeric enzyme act synergistically to bring about the degradation of complex substrates and that the family 42 carbohydrate-binding module has potential for application in the degradation of polysaccharides.

  6. Potential human cholesterol esterase inhibitor design: benefits from the molecular dynamics simulations and pharmacophore modeling studies.

    PubMed

    John, Shalini; Thangapandian, Sundarapandian; Lee, Keun Woo

    2012-01-01

    Human pancreatic cholesterol esterase (hCEase) is one of the lipases found to involve in the digestion of large and broad spectrum of substrates including triglycerides, phospholipids, cholesteryl esters, etc. The presence of bile salts is found to be very important for the activation of hCEase. Molecular dynamic simulations were performed for the apoform and bile salt complexed form of hCEase using the co-ordinates of two bile salts from bovine CEase. The stability of the systems throughout the simulation time was checked and two representative structures from the highly populated regions were selected using cluster analysis. These two representative structures were used in pharmacophore model generation. The generated pharmacophore models were validated and used in database screening. The screened hits were refined for their drug-like properties based on Lipinski's rule of five and ADMET properties. The drug-like compounds were further refined by molecular docking simulation using GOLD program based on the GOLD fitness score, mode of binding, and molecular interactions with the active site amino acids. Finally, three hits of novel scaffolds were selected as potential leads to be used in novel and potent hCEase inhibitor design. The stability of binding modes and molecular interactions of these final hits were re-assured by molecular dynamics simulations. PMID:22292952

  7. A cold-adapted, solvent and salt tolerant esterase from marine bacterium Psychrobacter pacificensis.

    PubMed

    Wu, Gaobing; Zhang, Xiangnan; Wei, Lu; Wu, Guojie; Kumar, Ashok; Mao, Tao; Liu, Ziduo

    2015-11-01

    Lipolytic enzymes with unique physico-chemical characteristics are gaining more attention for their immense industrial importance. In this study, a novel lipolytic enzyme (Est11) was cloned from the genomic library of a marine bacterium Psychrobacter pacificensis. The enzyme was expressed in Escherichia coli and purified to homogeneity with molecular mass of 32.9kDa. The recombinant Est11 was able to hydrolyze short chain esters (C2-C8) and displayed an optimum activity against butyrate ester (C4). The optimal temperature and pH were 25°C and 7.5, respectively. Est11 retained more than 70% of its original activity at 10°C, suggesting that it was a cold-active esterase. The enzyme was highly active and stable at high concentration of NaCl (5M). Further, incubation with ethanol, isopropanol, propanediol, DMSO, acetonitrile, and glycerol rendered remarkable positive effects on Est11 activity. Typically, even at the concentration of 30% (v/v), ethanol, DMSO, and propanediol increased Est11 activity by 1.3, 2.0, and 2.4-folds, respectively. This new robust enzyme with remarkable properties like cold-adaptability, exceptional tolerance to salt and organic solvents provides us a promising candidate to meet the needs of some harsh industrial processes. PMID:26231332

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

    PubMed

    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.

  9. A Lactobacillus plantarum Esterase Active on a Broad Range of Phenolic Esters

    PubMed Central

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

    2015-01-01

    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. PMID:25746986

  10. Subunit Stabilization and Polyethylene Glycolation of Cocaine Esterase Improves In Vivo Residence Time

    SciTech Connect

    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.

    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. A thermostable bacterial cocaine esterase rapidly eliminates cocaine from brain in nonhuman primates

    PubMed Central

    Howell, L L; Nye, J A; Stehouwer, J S; Voll, R J; Mun, J; Narasimhan, D; Nichols, J; Sunahara, R; Goodman, M M; Carroll, F I; Woods, J H

    2014-01-01

    A long-acting, thermostable bacterial cocaine esterase (CocE) has been identified that rapidly degrades cocaine with a KM of 1.33+0.085 μM. In vivo evaluation of CocE has shown protection against convulsant and lethal effects of cocaine in rodents, confirming the therapeutic potential of CocE against cocaine overdose. However, the current study is the first to evaluate the effects of CocE on cocaine brain levels. Positron emission tomogrpahy neuroimaging of [11C]cocaine was used to evaluate the time course of cocaine elimination from brain in the presence and absence of CocE in nonhuman primates. Systemic administration of CocE eliminated cocaine from the rhesus-monkey brain approximately three times faster than control conditions via peripheral actions through attenuating the input function from blood plasma. The efficiency of this process is sufficient to alleviate or prevent adverse central nervous system effects induced by cocaine. Although the present study used tracer doses of cocaine to access brain clearance, these findings further support the development of CocE for the treatment of acute cocaine toxicity. PMID:24984194

  12. HIGHLY METHYL ESTERIFIED SEEDS Is a Pectin Methyl Esterase Involved in Embryo Development1[OPEN

    PubMed Central

    Levesque-Tremblay, Gabriel; Müller, Kerstin; Mansfield, Shawn D.; Haughn, George W.

    2015-01-01

    Homogalacturonan pectin domains are synthesized in a highly methyl-esterified form that later can be differentially demethyl esterified by pectin methyl esterase (PME) to strengthen or loosen plant cell walls that contain pectin, including seed coat mucilage, a specialized secondary cell wall of seed coat epidermal cells. As a means to identify the active PMEs in seed coat mucilage, we identified seven PMEs expressed during seed coat development. One of these, HIGHLY METHYL ESTERIFIED SEEDS (HMS), is abundant during mucilage secretion, peaking at 7 d postanthesis in both the seed coat and the embryo. We have determined that this gene is required for normal levels of PME activity and homogalacturonan methyl esterification in the seed. The hms-1 mutant displays altered embryo morphology and mucilage extrusion, both of which are a consequence of defects in embryo development. A significant decrease in the size of cells in the embryo suggests that the changes in embryo morphology are a consequence of lack of cell expansion. Progeny from a cross between hms-1 and the previously characterized PME inhibitor5 overexpression line suggest that HMS acts independently from other cell wall-modifying enzymes in the embryo. We propose that HMS is required for cell wall loosening in the embryo to facilitate cell expansion during the accumulation of storage reserves and that its role in the seed coat is masked by redundancy. PMID:25572606

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

    PubMed

    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. PMID:27566510

  14. Identification and characterization of barley mutants lacking glycine decarboxylase and carboxyl esterase activities

    SciTech Connect

    Blackwell, R.; Lewis, K.; Lea, P. )

    1990-05-01

    A barley mutant has been isolated, from a selection of fifty air-sensitive seed-lines, using a standard gel stain technique which lacks carboxyl esterase activity, but has normal levels of carbonic anhydrase. In addition, two barley mutants lacking the ability to convert glycine to serine in the mitochondria, have been characterized. Both plants accumulate glycine in air and are unable to metabolize ({sup 14}C)glycine in the short-term. When ({sup 14}C)glycine was supplied over 2h LaPr 85/55 metabolized 90%, whereas the second mutant (LaPr 87/30) metabolized 10%. Results indicate that the mutation in LaPr 85/55 is almost certainly in the glycine transporter into the mitochondrion. The mutation in LaPr 87/30 has been shown, using western blotting, to be in both the P and H proteins, two of four proteins which comprise glycine decarboxylase (P, H, T and L).

  15. Discovery a novel organic solvent tolerant esterase from Salinispora arenicola CNP193 through genome mining.

    PubMed

    Fang, Yaowei; Wang, Shujun; Liu, Shu; Jiao, Yuliang

    2015-09-01

    An esterase gene, encoding a 325-amino-acid protein (SAestA), was mined form obligate marine actinomycete strain Salinispora arenicola CNP193 genome sequence. Phylogenetic analysis of the deduced amino acid sequence showed that the enzyme belonged to the family IV of lipolytic enzymes. The gene was cloned, expressed in Escherichia coli as a His-tagged protein, purified and characterized. The molecular weight of His-tagged SAestA is ∼38 kDa. SAestA-His6 was active in a temperature (5-40 °C) and pH range (7.0-11.0), and maximal activity was determined at pH 9.0 and 30 °C. The activity was severely inhibited by Hg(2+), Cu(2+), and Zn(2+). In particular, this enzyme showed remarkable stability in presence of organic solvents (25%, v/v) with log P>2.0 even after incubation for 7 days. All these characteristics suggested that SAestA may be a potential candidate for application in industrial processes in aqueous/organic media. PMID:26118483

  16. Nature versus nurture in two highly enantioselective esterases from Bacillus cereus and Thermoanaerobacter tengcongensis.

    PubMed

    Grosse, Stephan; Bergeron, Hélène; Imura, Akihiro; Boyd, Jason; Wang, Shaozhao; Kubota, Kazuo; Miyadera, Akihiko; Sulea, Traian; Lau, Peter C K

    2010-01-01

    There is an increasing need for the use of biocatalysis to obtain enantiopure compounds as chiral building blocks for drug synthesis such as antibiotics. The principal findings of this study are: (i) the complete sequenced genomes of Bacillus cereus ATCC 14579 and Thermoanaerobacter tengcongensis MB4 contain a hitherto undescribed enantioselective and alkaliphilic esterase (BcEST and TtEST respectively) that is specific for the production of (R)-2-benzyloxy-propionic acid ethyl ester, a key intermediate in the synthesis of levofloxacin, a potent antibiotic; and (ii) directed evolution targeted for increased thermostability of BcEST produced two improved variants, but in either case the 3-5 °C increase in the apparent melting temperature (T(m)) of the mutants over the native BcEST that has a T(m) of 50 °C was outperformed by TtEST, a naturally occurring homologue with a T(m) of 65 °C. Protein modelling of BcEST mapped the S148C and K272R mutations at protein surface and the I88T and Q110L mutations at more buried locations. This work expands the repertoire of characterized members of the α/β-fold hydrolase superfamily. Further, it shows that genome mining is an economical option for new biocatalyst discovery and we provide a rare example of a naturally occurring thermostable biocatalyst that outperforms experimentally evolved homologues that carry out the same hydrolysis. PMID:21255307

  17. Combined activation of methyl paraben by light irradiation and esterase metabolism toward oxidative DNA damage.

    PubMed

    Okamoto, Yoshinori; Hayashi, Tomohiro; Matsunami, Shinpei; Ueda, Koji; Kojima, Nakao

    2008-08-01

    Methyl paraben (MP) is often used as a preservative in foods, drugs, and cosmetics because of its high reliability in safety based on the rapid excretion and nonaccumulation following administration. Light irradiation sometimes produces unexpected activity from chemicals such as MP; furthermore, there is ample opportunity for MP to be exposed to sunlight. Here, we investigated whether MP shows DNA damage after sunlight irradiation. Two major photoproducts, p-hydroxybenzoic acid (PHBA) and 3-hydroxy methyl paraben (MP-3OH), were detected after sunlight irradiation to an aqueous MP solution. Both photoproducts were inactive in the in vitro DNA damage assay that measures oxidized guanine formed in calf thymus DNA in the presence of divalent copper ion, a known mediator of oxidative DNA damage. Simulated MP metabolism using dermal tissues after light irradiation produced these two photoproducts, which reacted with a microsomal fraction (S9) of the skin. A metabolite from MP-3OH, not PHBA, caused distinct DNA damage in the in vitro assay. This active metabolite was identified as protocatechuic acid, a hydrolyzed MP-3OH product. In addition, NADH, a cellular reductant, enhanced DNA damage by approximately five times. These results suggest that reactive oxygen species generated by the redox cycle via metal ion and catechol autoxidation are participating in oxidative DNA damage. This study reveals that MP might cause skin damage involving carcinogenesis through the combined activation of sunlight irradiation and skin esterases.

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

    PubMed

    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.

  19. Activity and dynamics of an enzyme, pig liver esterase, in near-anhydrous conditions

    SciTech Connect

    Lopez, Murielle; Kurkal-Siebert, V; Dunn, Rachel V.; Tehei, M; Finney, J.L.; Smith, Jeremy C; Daniel, R. M.

    2010-10-01

    Water is widely assumed to be essential for life, although the exact molecular basis of this requirement is unclear. Water facilitates protein motions, and although enzyme activity has been demonstrated at low hydrations in organic solvents, such nonaqueous solvents may allow the necessary motions for catalysis. To examine enzyme function in the absence of solvation and bypass diffusional constraints we have tested the ability of an enzyme, pig liver esterase, to catalyze alcoholysis as an anhydrous powder, in a reaction system of defined water content and where the substrates and products are gaseous. At hydrations of 3 ( 2) molecules of water per molecule of enzyme, activity is several orders-of-magnitude greater than nonenzymatic catalysis. Neutron spectroscopy indicates that the fast ( nanosecond) global anharmonic dynamics of the anhydrous functional enzyme are suppressed. This indicates that neither hydration water nor fast anharmonic dynamics are required for catalysis by this enzyme, implying that one of the biological requirements of water may lie with its role as a diffusion medium rather than any of its more specific properties.

  20. Modeling the tertiary structure of the patatin domain of neuropathy target esterase.

    PubMed

    Wijeyesakere, Sanjeeva J; Richardson, Rudy J; Stuckey, Jeanne A

    2007-04-01

    Neuropathy target esterase (NTE) is a transmembrane protein of unknown function whose specific chemical modification by certain organophosphorus (OP) compounds leads to distal axonopathy. Therefore, solving the 3D structure of NTE would advance the understanding of its pathogenic and physiologic roles. In this study, the tertiary structures of the patatin (catalytic) domain and the N-terminal transmembrane domain of NTE were modeled using the crystal structures of patatin (PDB ID 1oxw) and moricin (PDB ID 1kv4) as templates. Sequence alignments and secondary structure predictions were obtained from the INUB server (Buffalo, NY). O and PyMol were used to build the PNTE and NTE TMD chains from these sequence alignments. The PNTE model was refined in the presence of water using the crystallography and NMR system, while the NTE TMD model was refined in vacuo using the GROMOS implementation in the Swiss PDB viewer. The modeled active site of NTE was found to consist of a Ser966-Asp1086 catalytic dyad, which is characteristic of phospholipase A2 enzymes. The Ser966 Ogamma was located 2.93 A from the Odelta2 of Asp1086. In addition, our NTE model was found to contain a single N-terminal transmembrane domain. This modeling effort provided structural and mechanistic predictions about the catalytic domain of NTE that are being verified via experimental techniques.

  1. Subunit stabilization and polyethylene glycolation of cocaine esterase improves in vivo residence time.

    PubMed

    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

    2011-12-01

    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. PMID:21890748

  2. A thermostable bacterial cocaine esterase rapidly eliminates cocaine from brain in nonhuman primates.

    PubMed

    Howell, L L; Nye, J A; Stehouwer, J S; Voll, R J; Mun, J; Narasimhan, D; Nichols, J; Sunahara, R; Goodman, M M; Carroll, F I; Woods, J H

    2014-01-01

    A long-acting, thermostable bacterial cocaine esterase (CocE) has been identified that rapidly degrades cocaine with a K(M) of 1.33+0.085 μM. In vivo evaluation of CocE has shown protection against convulsant and lethal effects of cocaine in rodents, confirming the therapeutic potential of CocE against cocaine overdose. However, the current study is the first to evaluate the effects of CocE on cocaine brain levels. Positron emission tomogrpahy neuroimaging of [(11)C]cocaine was used to evaluate the time course of cocaine elimination from brain in the presence and absence of CocE in nonhuman primates. Systemic administration of CocE eliminated cocaine from the rhesus-monkey brain approximately three times faster than control conditions via peripheral actions through attenuating the input function from blood plasma. The efficiency of this process is sufficient to alleviate or prevent adverse central nervous system effects induced by cocaine. Although the present study used tracer doses of cocaine to access brain clearance, these findings further support the development of CocE for the treatment of acute cocaine toxicity. PMID:24984194

  3. Identification and characterization of novel esterases from a deep-sea sediment metagenome.

    PubMed

    Jiang, Xiawei; Xu, Xuewei; Huo, Yingyi; Wu, Yuehong; Zhu, Xufen; Zhang, Xinqi; Wu, Min

    2012-03-01

    A deep-sea sediment metagenomic library was constructed and screened for lipolytic enzymes by activity-based approach. Nine novel lipolytic enzymes were identified, and the amino acid sequences shared 56% to 84% identity to other lipolytic enzymes in the database. Phylogenetic analysis showed that these enzymes belonged to family IV lipolytic enzymes. One of the lipolytic enzymes, Est6, was successfully cloned and expressed in Escherichia coli Rosetta in a soluble form. The recombinant protein was purified by Ni-nitrilotriacetic affinity chromatography column and characterized using p-nitrophenyl esters with various chain lengths. The est6 gene consisted of 909 bp that encoded 302 amino acid residues. Est6 was most similar to a lipolytic enzyme from uncultured bacterium (ACL67845, 61% identity) isolated from the South China Sea marine sediment metagenome. The characterization of Est6 revealed that it was a cold-active esterase and exhibited the highest activity toward p-nitrophenyl butyrate (C4) at 20°C and pH 7.5.

  4. Feruloyl esterase production by Aspergillus terreus CECT 2808 and subsequent application to enzymatic hydrolysis.

    PubMed

    Pérez-Rodríguez, N; Moreira, C D; Torrado Agrasar, A; Domínguez, J M

    2016-09-01

    Ferulic acid esterases (FAE) were produced by Aspergillus terreus CECT 2808 from vine trimming shoots (VTS) and corn cob. Later, the fungal extracts thus obtained were used to enzymatically release ferulic acid (FA) from both substrates. Our findings showed a higher FAE activity in the enzymatic extracts produced on corn cob (0.070±0.004U/mL). Nevertheless, the enzymatic extracts produced on VTS demonstrated a better performance for FA release from both corn cob (2.05±0.01mg/g) and VTS (0.19±0.003mg/g). This result was probably because of the higher xylanase/FAE ratio determined in VTS extract. Therefore, an additional assay was carried out by supplementing corn cob extract with a commercial xylanase to test the influence of FAE/xylanase ratio in FA release. The results revealed the relevance of the FAE/xylanase ratio for an optimal FA release. PMID:27444329

  5. Acetic acid production from food wastes using yeast and acetic acid bacteria micro-aerobic fermentation.

    PubMed

    Li, Yang; He, Dongwei; Niu, Dongjie; Zhao, Youcai

    2015-05-01

    In this study, yeast and acetic acid bacteria strains were adopted to enhance the ethanol-type fermentation resulting to a volatile fatty acids yield of 30.22 g/L, and improve acetic acid production to 25.88 g/L, with food wastes as substrate. In contrast, only 12.81 g/L acetic acid can be obtained in the absence of strains. The parameters such as pH, oxidation reduction potential and volatile fatty acids were tested and the microbial diversity of different strains and activity of hydrolytic ferment were investigated to reveal the mechanism. The optimum pH and oxidation reduction potential for the acetic acid production were determined to be at 3.0-3.5 and -500 mV, respectively. Yeast can convert organic matters into ethanol, which is used by acetic acid bacteria to convert the organic wastes into acetic acid. The acetic acid thus obtained from food wastes micro-aerobic fermentation liquid could be extracted by distillation to get high-pure acetic acid.

  6. Acetylation of Starch with Vinyl Acetate in Imidazolium Ionic Liquids and Characterization of Acetate Distribution

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Starch was acetylated with vinyl acetate in different 1-butyl-3-methylimidazolium (BMIM) salts as solvent in effort to produce starches with different acetylation patterns. Overall degree of substitution was much higher for basic anions such as acetate and dicyanimide (dca) than for neutral anions ...

  7. 40 CFR 721.10001 - 2-Ethoxyethanol, 2-ethoxyethanol acetate, 2-methoxyethanol, and 2-methoxyethanol acetate.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... new uses subject to reporting. (1) The chemical substances identified as 2-ethoxyethanol (CAS No. 110-80-5), 2-ethoxyethanol acetate (CAS No. 111-15-9), 2-methoxyethanol (CAS No. 109-86-4), and 2-methoxyethanol acetate (CAS No. 110-49-6) are subject to reporting under this section for the significant new...

  8. 40 CFR 721.10001 - 2-Ethoxyethanol, 2-ethoxyethanol acetate, 2-methoxyethanol, and 2-methoxyethanol acetate.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... new uses subject to reporting. (1) The chemical substances identified as 2-ethoxyethanol (CAS No. 110-80-5), 2-ethoxyethanol acetate (CAS No. 111-15-9), 2-methoxyethanol (CAS No. 109-86-4), and 2-methoxyethanol acetate (CAS No. 110-49-6) are subject to reporting under this section for the significant new...

  9. Tested Demonstrations: Buffer Capacity of Various Acetic Acid-Sodium Acetate Systems: A Lecture Experiment.

    ERIC Educational Resources Information Center

    Donahue, Craig J.; Panek, Mary G.

    1985-01-01

    Background information and procedures are provided for a lecture experiment which uses indicators to illustrate the concept of differing buffer capacities by titrating acetic acid/sodium acetate buffers with 1.0 molar hydrochloric acid and 1.0 molar sodium hydroxide. A table with data used to plot the titration curve is included. (JN)

  10. 78 FR 16650 - In the Matter of: Dan Tran Dang, 1010 W. Moore Street, Santa Ana, CA 92707; Order Denying Export...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-18

    ... by successive Presidential Notices, the most recent being that of August 15, 2012 (77 FR 49699... Bureau of Industry and Security In the Matter of: Dan Tran Dang, 1010 W. Moore Street, Santa Ana, CA... address at: 1010 W. Moore Street, Santa Ana, CA 92707, and when acting for or on behalf of Dang,...

  11. "Houses and Fields and Vineyards Shall Yet Again Be Bought in This Land": The Story of Ana, a Public Kindergarten Teacher in Portugal.

    ERIC Educational Resources Information Center

    Vasconcelos, Teresa Maria Sena

    This study examined the teaching style and methods of Ana, a kindergarten teacher in Portugal, chosen because she is considered a master teacher by colleagues and parents and because she grew up in Portugal before democracy. The study attempted to answer the questions: (1) What are the commitments and competencies that distinguish Ana as a master…

  12. Dynamic Protonation Equilibrium of Solvated Acetic Acid

    SciTech Connect

    Gu, Wei; Frigato, Tomaso; Straatsma, TP; Helms, Volkhard H.

    2007-04-13

    For the first time, the dynamic protonation equilibrium between an amino acid side chain analogue and bulk water as well as the diffusion properties of the excess proton were successfully reproduced through unbiased computer simulations. During a 50 ns Q-HOP MD simulation, two different regimes of proton transfer were observed. Extended phases of frequent proton swapping between acetic acid and nearby water were separated by phases where the proton freely diffuses in the simulation box until it is captured again by acetic acid. The pKa of acetic acid was calculated around 3.0 based on the relative population of protonated and deprotonated states and the diffusion coefficient of excess proton was computed from the average mean squared displacement in the simulation. Both calculated values agree well with the experimental measurements.

  13. Leuprolide acetate-induced generalized papular eruption.

    PubMed

    Burris, Katy; Ding, Catherine Y; Lim, Geoffrey F S

    2014-06-01

    Leuprolide acetate, a gonadotropin-releasing hormone agonist, is used in the treatment of prostate cancer. We report a unique case of a disseminated papular rash following leuprolide acetate injections in a 65-year-old man that shares clinical and histopathological features of papuloerythroderma of Ofuji. Leuprolide-induced papuloerythroderma, as well as a limited number of other disseminated cutaneous eruptions caused by this drug, is extremely rare, with only one case previously reported. Our case calls attention to this uncommon side effect in a commonly used hormonal therapy.

  14. Insecticide resistance status, esterase activity, and electromorphs from mosquito populations of Culex quinquefasciatus Say (Diptera: Culicidae), in Houston (Harris County), Texas.

    PubMed

    Pietrantonio, P V; Gibson, G; Nawrocki, S; Carrier, F; Knight, W P

    2000-06-01

    Culex quinquefasciatus Say is a vector of St. Louis encephalitis (SLE) in Texas. This disease is endemic and prevalent in the Houston area. Disease prevention through mosquito control is mainly targeted against adults by application of a resmethrin-piperonyl butoxide formulation (Scourge). Immature mosquitoes were collected from eight areas in Harris County during 1998. The susceptibility status of these populations to Scourge, malathion, and resmethrin, the latter alone or with an esterase inhibitor as a synergist, was determined using a bottle assay with females. The population structure was investigated by electrophoretic analysis of esterases and their activity. Individual females were also analyzed for esterase activity by plate assay and for isoenzyme pattern by native PAGE. Bioassays indicated high levels of resistance to malathion in all areas. In addition, the effectiveness of Scourge in mosquitoes from area 51 deteriorated throughout the season. A localized, distinctive esterase pattern and activity level was observed in mosquitoes from different areas. Overall, the frequency of esterases Est alpha 2 (A2)/Est beta 2 (B2) was higher than that of Est beta 1 (B1). Altogether, these results indicate the onset of a fragile situation for mosquito control that should be further analyzed to effectively maintain the SLE prevention program for Harris County.

  15. Trypanosomatidae produce acetate via a mitochondrial acetate:succinate CoA transferase.

    PubMed

    Van Hellemond, J J; Opperdoes, F R; Tielens, A G

    1998-03-17

    Hydrogenosome-containing anaerobic protists, such as the trichomonads, produce large amounts of acetate by an acetate:succinate CoA transferase (ASCT)/succinyl CoA synthetase cycle. The notion that mitochondria and hydrogenosomes may have originated from the same alpha-proteobacterial endosymbiont has led us to look for the presence of a similar metabolic pathway in trypanosomatids because these are the earliest-branching mitochondriate eukaryotes and because they also are known to produce acetate. The mechanism of acetate production in these organisms, however, has remained unknown. Four different members of the trypanosomatid family: promastigotes of Leishmania mexicana mexicana, L. infantum and Phytomonas sp., and procyclics of Trypanosoma brucei were analyzed as well as the parasitic helminth Fasciola hepatica. They all use a mitochondrial ASCT for the production of acetate from acetyl CoA. The succinyl CoA that is produced during acetate formation by ASCT is recycled presumably to succinate by a mitochondrial succinyl CoA synthetase, concomitantly producing ATP from ADP. The ASCT of L. mexicana mexicana promastigotes was further characterized after partial purification of the enzyme. It has a high affinity for acetyl CoA (Km 0.26 mM) and a low affinity for succinate (Km 6.9 mM), which shows that significant acetate production can occur only when high mitochondrial succinate concentrations prevail. This study identifies a metabolic pathway common to mitochondria and hydrogenosomes, which strongly supports a common origin for these two organelles.

  16. Batrachochytrium dendrobatidis infection of amphibians in the Doñana National Park, Spain.

    PubMed

    Hidalgo-Vila, Judit; Díaz-Paniagua, Carmen; Marchand, Marc A; Cunningham, Andrew A

    2012-03-20

    Amphibian chytridiomycosis, caused by infection with the non-hyphal, zoosporic chytrid fungus Batrachochytrium dendrobatidis (Bd), is an emerging infectious disease recognised as a cause of recent amphibian population declines and extinctions worldwide. The Doñana National Park (DNP) is located in southwestern Spain, a country with widespread Bd infection. This protected area has a great diversity of aquatic habitats that constitute important breeding habitats for 11 native amphibian species. We sampled 625 amphibians in December 2007 and February to March 2008, months that correspond to the early and intermediate breeding seasons for amphibians, respectively. We found 7 of 9 sampled species to be infected with Bd and found differences in prevalence between sampling periods. Although some amphibians tested had higher intensities of infection than others, all animals sampled were apparently healthy and, so far, there has been no evidence of either unusually high rates of mortality or amphibian population declines in the DNP.

  17. Translocation of wild Laysan Teal Anas laysanensis from Laysan Island to Midway Atoll: Project update

    USGS Publications Warehouse

    Reynolds, M.H.; Breeden, J.H.; Klavitter, J.L.

    2007-01-01

    The Laysan Teal Anas laysanensis is endemic to the Hawaiian Islands, where it has been restricted to Laysan Island over the last 150 years. Individuals of this endangered species have recently been translocated to the two largest islands that comprise Midway Atoll National Wildlife Refuge, to reduce the risk of the Laysan Teal becoming extinct. Post-release monitoring with the aid of radio-telemetry was conducted to determine the success of the reintroduction attempt during October 2004-2007. The population was found to have increased after three breeding seasons, from forty-two founders sourced directly from Laysan, to a population of ??? 192 post-fledglings juveniles and adults. ?? Wildfowl & Wetlands Trust.

  18. Novel duck parvovirus identified in Cherry Valley ducks (Anas platyrhynchos domesticus), China.

    PubMed

    Li, Chuanfeng; Li, Qi; Chen, Zongyan; Liu, Guangqing

    2016-10-01

    An unknown infectious disease in Cherry Valley ducks (Anas platyrhynchos domesticus) characterized by short beak and strong growth retardation occurred in China during 2015. The causative agent of this disease, tentatively named duck short beak and dwarfism syndrome (DSBDS), as well as the evolutionary relationships between this causative agent and all currently known avian-origin parvoviruses were clarified by virus isolation, transmission electron microscope (TEM) observation, analysis of nuclear acid type, (RT-)PCR identification, whole genome sequencing, and NS1 protein sequences-based phylogenetic analyses. The results indicated that the causative agent of DSBDS is closely related with the goose parvovirus-like virus, which is divergent from all currently known avian-origin parvoviruses and should be a novel duck parvovirus (NDPV). PMID:27449955

  19. Meteorological Controls on Biomass Burning During Santa Ana Events in Southern California

    NASA Technical Reports Server (NTRS)

    Veraverbeke, Sander; Capps, Scott; Hook, Simon J.; Randerson, James T.; Jin, Yufang; Hall, Alex

    2013-01-01

    Fires occurring during Santa Ana (SA) events in southern California are driven by extreme fire weather characterized by high temperatures, low humidities, and high wind speeds. We studied the controls on burned area and carbon emissions during two intensive SA burning periods in 2003 and 2007. We therefore used remote sensing data in parallel with fire weather simulations of the Weather and Regional Forecast model. Total carbon emissions were approximately 1800 gigagrams in 2003 and 900 gigagrams in 2007, based on a daily burned area and a fire emission model that accounted for spatial variability in fuel loads and combustion completeness. On a regional scale, relatively strong positive correlations were found between the daily Fosberg fire weather index and burned area/emissions (probability is less than 0.01). Our analysis provides a quantitative assessment of relationships between fire activity and weather during severe SA fires in southern California.

  20. Biomimicry of multifunctional nanostructures in the neck feathers of mallard (Anas platyrhynchos L.) drakes

    NASA Astrophysics Data System (ADS)

    Khudiyev, Tural; Dogan, Tamer; Bayindir, Mehmet

    2014-04-01

    Biological systems serve as fundamental sources of inspiration for the development of artificially colored devices, and their investigation provides a great number of photonic design opportunities. While several successful biomimetic designs have been detailed in the literature, conventional fabrication techniques nonetheless remain inferior to their natural counterparts in complexity, ease of production and material economy. Here, we investigate the iridescent neck feathers of Anas platyrhynchos drakes, show that they feature an unusual arrangement of two-dimensional (2D) photonic crystals and further exhibit a superhydrophobic surface, and mimic this multifunctional structure using a nanostructure composite fabricated by a recently developed top-down iterative size reduction method, which avoids the above-mentioned fabrication challenges, provides macroscale control and enhances hydrophobicity through the surface structure. Our 2D solid core photonic crystal fibres strongly resemble drake neck plumage in structure and fully polymeric material composition, and can be produced in wide array of colors by minor alterations during the size reduction process.