The Experts below are selected from a list of 51 Experts worldwide ranked by ideXlab platform
L.-c. Comeau - One of the best experts on this subject based on the ideXlab platform.
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Maximizing production of Penicillium cyclopium partial Acylglycerol Lipase
Applied Microbiology and Biotechnology, 2002Co-Authors: G. Vanot, D. Valérie, M.-c. Guilhem, R. Phan Tan Luu, L.-c. ComeauAbstract:Penicillium cyclopium partial Acylglycerol Lipase production was maximized in shaken batch culture. The effect of inoculum size and substrate concentration on the Lipase activity released in the culture medium was visualized using a surface response methodology based on a Doehlert experimental design. The main advantage of this approach is the low number of experiments required to construct a predictive model of the experimental domain. Substrate percentage (corn steep, w/v) ranged from 0.1% to 1.9% and inoculum from 100 spores/ml to 3,200 spores/ml. We determined that an optimal set of experimental conditions for high Lipase production was 1.0% substrate and 3,200 spores/ml, with initial pH 5.0, temperature 25 °C and shaking speed 120 rpm. Between the conditions giving the minimum and the maximum Lipase production, we observed a three-fold increase in both the predicted and the measured values.
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Lipase-catalysed hydrolysis of short-chain substrates in solution and in emulsion: a kinetic study
Biochimica et biophysica acta, 2001Co-Authors: Lylia Nini, L.-c. Comeau, Louis Sarda, Elisabeth Boitard, J.p. Dubes, Henri ChahinianAbstract:We have studied the enzymatic hydrolysis of solutions and emulsions of vinyl propionate, vinyl butyrate and tripropionin by Lipases of various origin and specificity. Kinetic studies of the hydrolysis of short-chain substrates by microbial triAcylglycerol Lipases from Rhizopus oryzae, Mucor miehei, Candida rugosa, Candida antarctica A and by (phospho)Lipase from guinea-pig pancreas show that these lipolytic enzymes follow the Michaelis-Menten model. Surprisingly, the activity against solutions of tripropionin and vinyl esters ranges from 70% to 90% of that determined against emulsions. In contrast, a non-hyperbolic (sigmoidal) dependence of enzyme activity on ester concentration is found with human pancreatic Lipase, triAcylglycerol Lipase from Humicola lanuginosa (Thermomyces lanuginosa) and partial Acylglycerol Lipase from Penicillium camembertii and the same substrates. In all cases, no abrupt jump in activity (interfacial activation) is observed at substrate concentration corresponding to the solubility limit of the esters. Maximal lipolytic activity is always obtained in the presence of emulsified ester. Despite progress in the understanding of structure-function of Lipases, interpretation of the mode of action of Lipases active against solutions of short-chain substrates remains difficult. Actually, it is not known whether these enzymes, which possess a lid structure, are in open or/and closed conformation in the bulk phase and whether the opening of the lid that gives access to the catalytic triad is triggered by interaction of the enzyme molecule with monomeric substrates or/and multimolecular aggregates (micelles) both present in the bulk phase. From the comparison of the behaviour of Lipases used in this study which, in some cases, follow the Michaelis-Menten model and, in others, deviate from classical kinetics, it appears that the activity of classical Lipases against soluble short-chain vinyl esters and tripropionin depends not only on specific interaction with single substrate molecules at the catalytic site of the enzyme but also on physico-chemical parameters related to the state of association of the substrate dispersed in the aqueous phase. It is assumed that the interaction of Lipase with soluble multimolecular aggregates of tripropionin or short-chain vinyl esters or the formation of enzyme-substrate mixed micelles with ester bound to Lipase, might represent a crucial step that triggers the structural transition to the open enzyme conformation by displacement of the lid.
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Production of extracellular Lipases by Penicillium cyclopium purification and characterization of a partial Acylglycerol Lipase.
Bioscience biotechnology and biochemistry, 2000Co-Authors: Henri Chahinian, G. Vanot, Aida Ibrik, Nathalie Rugani, Louis Sarda, L.-c. ComeauAbstract:Penicillium cyclopium, grown in stationary culture, produces a type I Lipase specific for triAcylglycerols while, in shaken culture, it produces a type II Lipase only active on partial Acylglycerols. Lipase II has been purified by ammonium sulfate precipitation and chromatographies on Sephadex G-75 and DEAE-Sephadex. The enzyme exists in several glycosylated forms of 40-43 kDa, which can be converted to a single protein of 37 kDa by enzymatic deglycosylation. Activity of Lipase II is maximal at pH 7.0 and 40°C. The enzyme is stable from pH 4.5 to 7.0. Activity is rapidly lost at temperatures above 50°C. The enzyme specifically hydrolyzes monoAcylglycerols and diAcylglycerols, especially of medium chain fatty acids. The sequence of the 20 first amino acid residues is similar to the N-terminal region of P. camembertii Lipase and partially similar to Lipases from Humicola lanuginosa and Aspergillus oryzae, but is different from Penicillium cyclopium Lipase I. However, it can be observed that residues of valine and serine at positions 2 and 5 in Penicillium cyclopium Lipase II are conserved in Penicillium expansum Lipase, of which 16 out of the 20 first amino acid residues are similar to Penicillium cyclopium Lipase I.
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production of extracellular Lipases by penicillium cyclopium purification and characterization of a partial Acylglycerol Lipase
Bioscience Biotechnology and Biochemistry, 2000Co-Authors: Henri Chahinian, G. Vanot, Aida Ibrik, Nathalie Rugani, Louis Sarda, L.-c. ComeauAbstract:Penicillium cyclopium, grown in stationary culture, produces a type I Lipase specific for triAcylglycerols while, in shaken culture, it produces a type II Lipase only active on partial Acylglycerols. Lipase II has been purified by ammonium sulfate precipitation and chromatographies on Sephadex G-75 and DEAE-Sephadex. The enzyme exists in several glycosylated forms of 40-43 kDa, which can be converted to a single protein of 37 kDa by enzymatic deglycosylation. Activity of Lipase II is maximal at pH 7.0 and 40 degrees C. The enzyme is stable from pH 4.5 to 7.0. Activity is rapidly lost at temperatures above 50 degrees C. The enzyme specifically hydrolyzes monoAcylglycerols and diAcylglycerols, especially of medium chain fatty acids. The sequence of the 20 first amino acid residues is similar to the N-terminal region of P. camembertii Lipase and partially similar to Lipases from Humicola lanuginosa and Aspergillus oryzae, but is different from Penicillium cyclopium Lipase I. However, it can be observed that residues of valine and serine at positions 2 and 5 in Penicillium cyclopium Lipase II are conserved in Penicillium expansum Lipase, of which 16 out of the 20 first amino acid residues are similar to Penicillium cyclopium Lipase I.
Yonghua Wang - One of the best experts on this subject based on the ideXlab platform.
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A novel strategy to improve the thermostability of Penicillium camembertii mono- and di-Acylglycerol Lipase.
Biochemical and biophysical research communications, 2018Co-Authors: Yanhua Liu, Bo Yang, Dongjuan Yuan, Zexin Zhao, Dongming Lan, Yonghua WangAbstract:Abstract Penicillium camembertii (PCL), a mono- and di-Acylglycerol Lipase (DGL), has the vital potential in the oil chemistry for food industry. However, known DGLs are mesophilic enzymes which restricts its application in the industry. To improve thermostability of PCL, we used amino acid substitution by comparison of amino acids compositions of PCL and protein sequences from typical thermophilic bacteria. Then, some conservative residues around active center were avoided to mutate according to homologous alignment analyses. Furthermore, the list was narrowed down to 28 candidate mutational sites of PCL by analyzing the hydrophobic interaction of amino acids in the structure. And among them only the mutant PCL-D25R had formed an additional salt bridge between R25-D32 and increased more hydrogen bonds interaction. Therefore, mutant PCL-D25R were constructed and expressed. Thermal inactivation assay showed that the half-life of mutant PCL-D25R at 45 °C increased 4-fold compared to that of PCL-WT. Melting temperature of mutant PCL-D25R increased to 49.5 °C from 46.5 °C by fluorescence-based thermal stability assay. This study provides a valuable strategy for engineering DGL thermostability.
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A mechanistic study into the epoxidation of carboxylic acid and alkene in a mono, di-Acylglycerol Lipase
Biochemical and biophysical research communications, 2015Co-Authors: Xuping Wang, Grzegorz M Popowicz, Bo Yang, Qingyun Tang, Yonghua WangAbstract:Abstract More and more industrial chemistry reactions rely on green technologies. Enzymes are finding increasing use in diverse chemical processes. Epoxidized vegetable oils have recently found applications as plasticizers and additives for PVC production. We report here an unusual activity of the Malassezia globosa Lipase (SMG1) that is able to catalyze epoxidation of alkenes. SMG1 catalyzes formation of peroxides from long chain carboxylic acids that subsequently react with double bonds of alkenes to produce epoxides. The SMG1 is selective towards carboxylic acids and active also as a mutant lacking hydrolase activity. Moreover we present previously unobserved mechanism of catalysis that does not rely on acyl–substrate complex nor tetrahedral intermediate. Since SMG1 Lipase is activated by allosteric change upon binding to the lipophilic–hydrophilic phase interface we reason that it can be used to drive the epoxidation in the lipophilic phase exclusively.
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biochemical properties of a new cold active mono and diAcylglycerol Lipase from marine member janibacter sp strain htcc2649
International Journal of Molecular Sciences, 2014Co-Authors: Dongjuan Yuan, Bo Yang, Yonghua WangAbstract:Mono- and di-Acylglycerol Lipase has been applied to industrial usage in oil modification for its special substrate selectivity. Until now, the reported mono- and di-Acylglycerol Lipases from microorganism are limited, and there is no report on the mono- and di-Acylglycerol Lipase from bacteria. A predicted Lipase (named MAJ1) from marine Janibacter sp. strain HTCC2649 was purified and biochemical characterized. MAJ1 was clustered in the family I.7 of esterase/Lipase. The optimum activity of the purified MAJ1 occurred at pH 7.0 and 30 °C. The enzyme retained 50% of the optimum activity at 5 °C, indicating that MAJ1 is a cold-active Lipase. The enzyme activity was stable in the presence of various metal ions, and inhibited in EDTA. MAJ1 was resistant to detergents. MAJ1 preferentially hydrolyzed mono- and di-Acylglycerols, but did not show activity to triAcylglycerols of camellia oil substrates. Further, MAJ1 is low homologous to that of the reported fungal diAcylglycerol Lipases, including Malassezia globosa Lipase 1 (SMG1), Penicillium camembertii Lipase U-150 (PCL), and Aspergillus oryzae Lipase (AOL). Thus, we identified a novel cold-active bacterial Lipase with a sn-1/3 preference towards mono- and di-acylglycerides for the first time. Moreover, it has the potential, in oil modification, for special substrate selectivity.
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Enzymatic synthesis of extremely pure triAcylglycerols enriched in conjugated linoleic acids.
Molecules (Basel Switzerland), 2013Co-Authors: Yu Cao, Bo Yang, Weifei Wang, Yonghua WangAbstract:This work was objectively targeted to synthesize extremely pure triAcylglycerols (TAG) enriched in conjugated linoleic acids (CLAs) for medical and dietetic purposes. Extremely pure CLA-enriched TAG was successfully synthesized by using the multi-step process: TAG was primarily synthesized by Lipase-catalyzed esterification of CLA and glycerol and then the lower glycerides [monoAcylglycerol (MAG) and diAcylglycerol (DAG)] in the esterification mixtures was hydrolyzed to free fatty acids (FFAs) by a mono- and di-Acylglycerol Lipase (Lipase SMG1), finally, the FFAs were further separated from TAG by low temperature (150 °C) molecular distillation. The operation parameters for the Lipase SMG1-catalyzed hydrolysis were optimized using response surface methodology based on the central composite rotatable design (CCRD). The operation parameters included water content, pH and reaction temperature and all of these three parameters showed significant effects on the hydrolysis of lower glycerides. The optimal conditions were obtained with a water content of 66.4% (w/w, with respect to oil mass), pH at 5.7 and 1 h of reaction time at 19.6 °C. Under these conditions, the content of lower glycerides in the reaction mixture decreased from 45.2% to 0.3% and the purity of CLA-enriched TAG reached 99.7%. Further purification of TAG was accomplished by molecular distillation and the final CLA-enriched TAG product yielded 99.8% of TAG. These extremely pure CLA-enriched TAG would be used for in vivo studies in animals and humans in order to get specific information concerning CLA metabolism.
Henri Chahinian - One of the best experts on this subject based on the ideXlab platform.
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Lipase-catalysed hydrolysis of short-chain substrates in solution and in emulsion: a kinetic study
Biochimica et biophysica acta, 2001Co-Authors: Lylia Nini, L.-c. Comeau, Louis Sarda, Elisabeth Boitard, J.p. Dubes, Henri ChahinianAbstract:We have studied the enzymatic hydrolysis of solutions and emulsions of vinyl propionate, vinyl butyrate and tripropionin by Lipases of various origin and specificity. Kinetic studies of the hydrolysis of short-chain substrates by microbial triAcylglycerol Lipases from Rhizopus oryzae, Mucor miehei, Candida rugosa, Candida antarctica A and by (phospho)Lipase from guinea-pig pancreas show that these lipolytic enzymes follow the Michaelis-Menten model. Surprisingly, the activity against solutions of tripropionin and vinyl esters ranges from 70% to 90% of that determined against emulsions. In contrast, a non-hyperbolic (sigmoidal) dependence of enzyme activity on ester concentration is found with human pancreatic Lipase, triAcylglycerol Lipase from Humicola lanuginosa (Thermomyces lanuginosa) and partial Acylglycerol Lipase from Penicillium camembertii and the same substrates. In all cases, no abrupt jump in activity (interfacial activation) is observed at substrate concentration corresponding to the solubility limit of the esters. Maximal lipolytic activity is always obtained in the presence of emulsified ester. Despite progress in the understanding of structure-function of Lipases, interpretation of the mode of action of Lipases active against solutions of short-chain substrates remains difficult. Actually, it is not known whether these enzymes, which possess a lid structure, are in open or/and closed conformation in the bulk phase and whether the opening of the lid that gives access to the catalytic triad is triggered by interaction of the enzyme molecule with monomeric substrates or/and multimolecular aggregates (micelles) both present in the bulk phase. From the comparison of the behaviour of Lipases used in this study which, in some cases, follow the Michaelis-Menten model and, in others, deviate from classical kinetics, it appears that the activity of classical Lipases against soluble short-chain vinyl esters and tripropionin depends not only on specific interaction with single substrate molecules at the catalytic site of the enzyme but also on physico-chemical parameters related to the state of association of the substrate dispersed in the aqueous phase. It is assumed that the interaction of Lipase with soluble multimolecular aggregates of tripropionin or short-chain vinyl esters or the formation of enzyme-substrate mixed micelles with ester bound to Lipase, might represent a crucial step that triggers the structural transition to the open enzyme conformation by displacement of the lid.
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Production of extracellular Lipases by Penicillium cyclopium purification and characterization of a partial Acylglycerol Lipase.
Bioscience biotechnology and biochemistry, 2000Co-Authors: Henri Chahinian, G. Vanot, Aida Ibrik, Nathalie Rugani, Louis Sarda, L.-c. ComeauAbstract:Penicillium cyclopium, grown in stationary culture, produces a type I Lipase specific for triAcylglycerols while, in shaken culture, it produces a type II Lipase only active on partial Acylglycerols. Lipase II has been purified by ammonium sulfate precipitation and chromatographies on Sephadex G-75 and DEAE-Sephadex. The enzyme exists in several glycosylated forms of 40-43 kDa, which can be converted to a single protein of 37 kDa by enzymatic deglycosylation. Activity of Lipase II is maximal at pH 7.0 and 40°C. The enzyme is stable from pH 4.5 to 7.0. Activity is rapidly lost at temperatures above 50°C. The enzyme specifically hydrolyzes monoAcylglycerols and diAcylglycerols, especially of medium chain fatty acids. The sequence of the 20 first amino acid residues is similar to the N-terminal region of P. camembertii Lipase and partially similar to Lipases from Humicola lanuginosa and Aspergillus oryzae, but is different from Penicillium cyclopium Lipase I. However, it can be observed that residues of valine and serine at positions 2 and 5 in Penicillium cyclopium Lipase II are conserved in Penicillium expansum Lipase, of which 16 out of the 20 first amino acid residues are similar to Penicillium cyclopium Lipase I.
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production of extracellular Lipases by penicillium cyclopium purification and characterization of a partial Acylglycerol Lipase
Bioscience Biotechnology and Biochemistry, 2000Co-Authors: Henri Chahinian, G. Vanot, Aida Ibrik, Nathalie Rugani, Louis Sarda, L.-c. ComeauAbstract:Penicillium cyclopium, grown in stationary culture, produces a type I Lipase specific for triAcylglycerols while, in shaken culture, it produces a type II Lipase only active on partial Acylglycerols. Lipase II has been purified by ammonium sulfate precipitation and chromatographies on Sephadex G-75 and DEAE-Sephadex. The enzyme exists in several glycosylated forms of 40-43 kDa, which can be converted to a single protein of 37 kDa by enzymatic deglycosylation. Activity of Lipase II is maximal at pH 7.0 and 40 degrees C. The enzyme is stable from pH 4.5 to 7.0. Activity is rapidly lost at temperatures above 50 degrees C. The enzyme specifically hydrolyzes monoAcylglycerols and diAcylglycerols, especially of medium chain fatty acids. The sequence of the 20 first amino acid residues is similar to the N-terminal region of P. camembertii Lipase and partially similar to Lipases from Humicola lanuginosa and Aspergillus oryzae, but is different from Penicillium cyclopium Lipase I. However, it can be observed that residues of valine and serine at positions 2 and 5 in Penicillium cyclopium Lipase II are conserved in Penicillium expansum Lipase, of which 16 out of the 20 first amino acid residues are similar to Penicillium cyclopium Lipase I.
G. Vanot - One of the best experts on this subject based on the ideXlab platform.
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Maximizing production of Penicillium cyclopium partial Acylglycerol Lipase
Applied Microbiology and Biotechnology, 2002Co-Authors: G. Vanot, D. Valérie, M.-c. Guilhem, R. Phan Tan Luu, L.-c. ComeauAbstract:Penicillium cyclopium partial Acylglycerol Lipase production was maximized in shaken batch culture. The effect of inoculum size and substrate concentration on the Lipase activity released in the culture medium was visualized using a surface response methodology based on a Doehlert experimental design. The main advantage of this approach is the low number of experiments required to construct a predictive model of the experimental domain. Substrate percentage (corn steep, w/v) ranged from 0.1% to 1.9% and inoculum from 100 spores/ml to 3,200 spores/ml. We determined that an optimal set of experimental conditions for high Lipase production was 1.0% substrate and 3,200 spores/ml, with initial pH 5.0, temperature 25 °C and shaking speed 120 rpm. Between the conditions giving the minimum and the maximum Lipase production, we observed a three-fold increase in both the predicted and the measured values.
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Production of extracellular Lipases by Penicillium cyclopium purification and characterization of a partial Acylglycerol Lipase.
Bioscience biotechnology and biochemistry, 2000Co-Authors: Henri Chahinian, G. Vanot, Aida Ibrik, Nathalie Rugani, Louis Sarda, L.-c. ComeauAbstract:Penicillium cyclopium, grown in stationary culture, produces a type I Lipase specific for triAcylglycerols while, in shaken culture, it produces a type II Lipase only active on partial Acylglycerols. Lipase II has been purified by ammonium sulfate precipitation and chromatographies on Sephadex G-75 and DEAE-Sephadex. The enzyme exists in several glycosylated forms of 40-43 kDa, which can be converted to a single protein of 37 kDa by enzymatic deglycosylation. Activity of Lipase II is maximal at pH 7.0 and 40°C. The enzyme is stable from pH 4.5 to 7.0. Activity is rapidly lost at temperatures above 50°C. The enzyme specifically hydrolyzes monoAcylglycerols and diAcylglycerols, especially of medium chain fatty acids. The sequence of the 20 first amino acid residues is similar to the N-terminal region of P. camembertii Lipase and partially similar to Lipases from Humicola lanuginosa and Aspergillus oryzae, but is different from Penicillium cyclopium Lipase I. However, it can be observed that residues of valine and serine at positions 2 and 5 in Penicillium cyclopium Lipase II are conserved in Penicillium expansum Lipase, of which 16 out of the 20 first amino acid residues are similar to Penicillium cyclopium Lipase I.
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production of extracellular Lipases by penicillium cyclopium purification and characterization of a partial Acylglycerol Lipase
Bioscience Biotechnology and Biochemistry, 2000Co-Authors: Henri Chahinian, G. Vanot, Aida Ibrik, Nathalie Rugani, Louis Sarda, L.-c. ComeauAbstract:Penicillium cyclopium, grown in stationary culture, produces a type I Lipase specific for triAcylglycerols while, in shaken culture, it produces a type II Lipase only active on partial Acylglycerols. Lipase II has been purified by ammonium sulfate precipitation and chromatographies on Sephadex G-75 and DEAE-Sephadex. The enzyme exists in several glycosylated forms of 40-43 kDa, which can be converted to a single protein of 37 kDa by enzymatic deglycosylation. Activity of Lipase II is maximal at pH 7.0 and 40 degrees C. The enzyme is stable from pH 4.5 to 7.0. Activity is rapidly lost at temperatures above 50 degrees C. The enzyme specifically hydrolyzes monoAcylglycerols and diAcylglycerols, especially of medium chain fatty acids. The sequence of the 20 first amino acid residues is similar to the N-terminal region of P. camembertii Lipase and partially similar to Lipases from Humicola lanuginosa and Aspergillus oryzae, but is different from Penicillium cyclopium Lipase I. However, it can be observed that residues of valine and serine at positions 2 and 5 in Penicillium cyclopium Lipase II are conserved in Penicillium expansum Lipase, of which 16 out of the 20 first amino acid residues are similar to Penicillium cyclopium Lipase I.
Bo Yang - One of the best experts on this subject based on the ideXlab platform.
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A novel strategy to improve the thermostability of Penicillium camembertii mono- and di-Acylglycerol Lipase.
Biochemical and biophysical research communications, 2018Co-Authors: Yanhua Liu, Bo Yang, Dongjuan Yuan, Zexin Zhao, Dongming Lan, Yonghua WangAbstract:Abstract Penicillium camembertii (PCL), a mono- and di-Acylglycerol Lipase (DGL), has the vital potential in the oil chemistry for food industry. However, known DGLs are mesophilic enzymes which restricts its application in the industry. To improve thermostability of PCL, we used amino acid substitution by comparison of amino acids compositions of PCL and protein sequences from typical thermophilic bacteria. Then, some conservative residues around active center were avoided to mutate according to homologous alignment analyses. Furthermore, the list was narrowed down to 28 candidate mutational sites of PCL by analyzing the hydrophobic interaction of amino acids in the structure. And among them only the mutant PCL-D25R had formed an additional salt bridge between R25-D32 and increased more hydrogen bonds interaction. Therefore, mutant PCL-D25R were constructed and expressed. Thermal inactivation assay showed that the half-life of mutant PCL-D25R at 45 °C increased 4-fold compared to that of PCL-WT. Melting temperature of mutant PCL-D25R increased to 49.5 °C from 46.5 °C by fluorescence-based thermal stability assay. This study provides a valuable strategy for engineering DGL thermostability.
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A mechanistic study into the epoxidation of carboxylic acid and alkene in a mono, di-Acylglycerol Lipase
Biochemical and biophysical research communications, 2015Co-Authors: Xuping Wang, Grzegorz M Popowicz, Bo Yang, Qingyun Tang, Yonghua WangAbstract:Abstract More and more industrial chemistry reactions rely on green technologies. Enzymes are finding increasing use in diverse chemical processes. Epoxidized vegetable oils have recently found applications as plasticizers and additives for PVC production. We report here an unusual activity of the Malassezia globosa Lipase (SMG1) that is able to catalyze epoxidation of alkenes. SMG1 catalyzes formation of peroxides from long chain carboxylic acids that subsequently react with double bonds of alkenes to produce epoxides. The SMG1 is selective towards carboxylic acids and active also as a mutant lacking hydrolase activity. Moreover we present previously unobserved mechanism of catalysis that does not rely on acyl–substrate complex nor tetrahedral intermediate. Since SMG1 Lipase is activated by allosteric change upon binding to the lipophilic–hydrophilic phase interface we reason that it can be used to drive the epoxidation in the lipophilic phase exclusively.
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biochemical properties of a new cold active mono and diAcylglycerol Lipase from marine member janibacter sp strain htcc2649
International Journal of Molecular Sciences, 2014Co-Authors: Dongjuan Yuan, Bo Yang, Yonghua WangAbstract:Mono- and di-Acylglycerol Lipase has been applied to industrial usage in oil modification for its special substrate selectivity. Until now, the reported mono- and di-Acylglycerol Lipases from microorganism are limited, and there is no report on the mono- and di-Acylglycerol Lipase from bacteria. A predicted Lipase (named MAJ1) from marine Janibacter sp. strain HTCC2649 was purified and biochemical characterized. MAJ1 was clustered in the family I.7 of esterase/Lipase. The optimum activity of the purified MAJ1 occurred at pH 7.0 and 30 °C. The enzyme retained 50% of the optimum activity at 5 °C, indicating that MAJ1 is a cold-active Lipase. The enzyme activity was stable in the presence of various metal ions, and inhibited in EDTA. MAJ1 was resistant to detergents. MAJ1 preferentially hydrolyzed mono- and di-Acylglycerols, but did not show activity to triAcylglycerols of camellia oil substrates. Further, MAJ1 is low homologous to that of the reported fungal diAcylglycerol Lipases, including Malassezia globosa Lipase 1 (SMG1), Penicillium camembertii Lipase U-150 (PCL), and Aspergillus oryzae Lipase (AOL). Thus, we identified a novel cold-active bacterial Lipase with a sn-1/3 preference towards mono- and di-acylglycerides for the first time. Moreover, it has the potential, in oil modification, for special substrate selectivity.
-
Enzymatic synthesis of extremely pure triAcylglycerols enriched in conjugated linoleic acids.
Molecules (Basel Switzerland), 2013Co-Authors: Yu Cao, Bo Yang, Weifei Wang, Yonghua WangAbstract:This work was objectively targeted to synthesize extremely pure triAcylglycerols (TAG) enriched in conjugated linoleic acids (CLAs) for medical and dietetic purposes. Extremely pure CLA-enriched TAG was successfully synthesized by using the multi-step process: TAG was primarily synthesized by Lipase-catalyzed esterification of CLA and glycerol and then the lower glycerides [monoAcylglycerol (MAG) and diAcylglycerol (DAG)] in the esterification mixtures was hydrolyzed to free fatty acids (FFAs) by a mono- and di-Acylglycerol Lipase (Lipase SMG1), finally, the FFAs were further separated from TAG by low temperature (150 °C) molecular distillation. The operation parameters for the Lipase SMG1-catalyzed hydrolysis were optimized using response surface methodology based on the central composite rotatable design (CCRD). The operation parameters included water content, pH and reaction temperature and all of these three parameters showed significant effects on the hydrolysis of lower glycerides. The optimal conditions were obtained with a water content of 66.4% (w/w, with respect to oil mass), pH at 5.7 and 1 h of reaction time at 19.6 °C. Under these conditions, the content of lower glycerides in the reaction mixture decreased from 45.2% to 0.3% and the purity of CLA-enriched TAG reached 99.7%. Further purification of TAG was accomplished by molecular distillation and the final CLA-enriched TAG product yielded 99.8% of TAG. These extremely pure CLA-enriched TAG would be used for in vivo studies in animals and humans in order to get specific information concerning CLA metabolism.