The Experts below are selected from a list of 234 Experts worldwide ranked by ideXlab platform
Patrick Fickers - One of the best experts on this subject based on the ideXlab platform.
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Waste soybean frying oil for the production, extraction, and characterization of cell-wall-associated lipases from Yarrowia Lipolytica
Bioprocess and Biosystems Engineering, 2021Co-Authors: Patrícia M. B. Nunes, Patrick Fickers, Jully L. Fraga, Rafael B. Ratier, Maria Helena M. Rocha-leão, Ana I. S. Brígida, Priscilla F. F. AmaralAbstract:The Lipolytic yeast Yarrowia Lipolytica produces cell-wall-associated lipases, namely Lip7p and Lip8p, that could have interesting properties as catalyst either in free (released lipase fraction—RLF) or cell-associated (cell-bound lipase fraction—CBLF) forms. Herein, a mixture of waste soybean frying oil, yeast extract and bactopeptone was found to favor the enzyme production. Best parameters for lipase activation and release from the cell wall by means of acoustic wave treatment were defined as: 26 W/cm^2 for 1 min for CBLF and 52 W/cm^2 for 2 min for RLF. Optimal pH and temperature values for lipase activity together with storage conditions were similar for both the free enzyme and cell-associated one: pH 7.0; T = 37 °C; and > 70% residual activity for 60 days at 4, − 4 °C and for 15 days at 30 °C.
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Methyl Oleate Modulates LIP2 Expression in the Lipolytic Yeast Yarrowia Lipolytica
Biotechnology Letters, 2005Co-Authors: Patrick Fickers, Jacqueline Destain, Philippe ThonartAbstract:Methyl oleate was used as a primary carbon source and as an alternative inducer for the production of an extracellular lipase, Lip2, in Y. Lipolytica strain LgX64.81 grown in a 20-l bioreactor. The lipase-encoding gene, LIP2 , was investigated during culture on methyl oleate using a p LIP2 – LacZ reporter fusion and we provide evidence for the involvement of methyl oleate in its regulation.
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Identification and characterisation of LIP7 and LIP8 genes encoding two extracellular triacylglycerol lipases in the yeast Yarrowia Lipolytica.
Fungal Genetics and Biology, 2005Co-Authors: Patrick Fickers, Philippe Thonart, Claude Gaillardin, Franck Fudalej, M-therese Le Dall, Serge Casaregola, Jean-marc NicaudAbstract:In the Lipolytic yeast Yarrowia Lipolytica, the LIP2 gene was previously reported to encode an extracellular lipase. The growth of a Deltalip2 strain on triglycerides as sole carbon source suggest an alternative pathway for triglycerides utilisation in this yeast. Here, we describe the isolation and the characterisation of the LIP7 and LIP8 genes which were found to encode a 366 and a 371-amino acid precursor protein, respectively. These proteins which belong to the triacylglycerol hydrolase family (EC 3.1.1.3) presented a high homology with the extracellular lipase CdLIP2 and CdLIP3 from Candida deformans. The physiological function of the lipase isoenzymes was investigated by creating single and multi-disrupted strains. Lip7p and Lip8p were found to correspond to active secreted lipases. The lack of lipase production in a Deltalip2 Deltalip7 Deltalip8 strain suggest that no additional extracellular lipase remains to be discovered in Y. Lipolytica. The substrate specificity towards synthetic ester molecules indicates that Lip7p presented a maximum activity centred on caproate (C6) while that of Lip8p is in caprate (C10).
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Carbon and nitrogen sources modulate lipase production in the yeast Yarrowia Lipolytica
Journal of Applied Microbiology, 2004Co-Authors: Patrick Fickers, Jacqueline Destain, Jean-marc Nicaud, Claude Gaillardin, Philippe ThonartAbstract:AIMS: To analyse the influence of nitrogen and carbon sources on extracellular lipase production by Yarrowia Lipolytica-overproducing mutant in order to optimize its production in large-scale bioreactors. METHODS AND RESULTS: The level of lipase production and LIP2 induction, measured using an LIP2-LacZ reporter gene, were compared for different carbon and nitrogen sources and for different concentrations. The localization of the enzyme during growth was also determined by Western blotting analysis using a six-histidine-tagged lipase. SIGNIFICANCE AND IMPACT OF THE STUDY: Tryptone N1 and oleic acid are the most suitable nitrogen and carbon sources for the production of the extracellular lipase by the Y. Lipolytica mutant. Higher levels of lipase production were obtained as the tryptone concentration increased in the culture medium. Such a positive correlation was not observed with oleic acid media where the highest Lipolytic productivities were obtained in the presence of low concentration. We also demonstrate that in the presence of oleic acid, lipase is cell-bound during the growth phase before being released in the media. CONCLUSIONS: This work provides a better understanding of the mechanism controlling LIP2 expression and, thus, extracellular lipase production in the yeast Y. Lipolytica.
Hugo S. Garcia - One of the best experts on this subject based on the ideXlab platform.
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extraction and fractionation of Lipolytic enzyme from viscera of monterey sardine sardinops sagax caerulea
International Journal of Food Science and Technology, 2009Co-Authors: J A Noriegarodriguez, O Anguloguerrero, Luis Angel Medinajuarez, Nohemi Gamezmeza, Argentina Alanisvilla, Armando Tejedamansir, Hugo S. GarciaAbstract:Summary In this study, Lipolytic activity of a semi-purified Lipolytic enzyme (SLE) from the viscera of sardine (Sardinops sagax caerulea) was screened on the lipolysis of olive, Menhaden and sardine oil. A Lipolytic enzyme was partially purified from the crude extract of sardine viscera by fractional precipitation followed by ultrafiltration (30 kDa). The main tissues found in sardine viscera were pyloric caeca (19.0% w/w), digestive tract (13.0% w/w), liver (4.8% w/w) and pancreas (1.5% w/w). Results show that pancreas had the highest Lipolytic activity. There were no significant differences in Lipolytic activity between pyloric caeca, intestine and liver (P < 0.05). Specific activity of the SLE increased 47.0-fold after extraction and fractionation, with a yield of 0.34% calculated for the whole viscera weight. Lipolytic activity of SLE from sardine viscera increased threefold when sardine oil was used as substrate. The results of this study confirm the potential importance of lipases from marine sources.
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Extraction and fractionation of Lipolytic enzyme from viscera of Monterey sardine (Sardinops sagax caerulea)
International Journal of Food Science & Technology, 2009Co-Authors: J. A. Noriega-rodriguez, Luis Ángel Medina-juárez, Nohemí Gámez-meza, Argentina Alanis-villa, Armando Tejeda-mansir, Ofelia Angulo-guerrero, Hugo S. GarciaAbstract:Summary In this study, Lipolytic activity of a semi-purified Lipolytic enzyme (SLE) from the viscera of sardine (Sardinops sagax caerulea) was screened on the lipolysis of olive, Menhaden and sardine oil. A Lipolytic enzyme was partially purified from the crude extract of sardine viscera by fractional precipitation followed by ultrafiltration (30 kDa). The main tissues found in sardine viscera were pyloric caeca (19.0% w/w), digestive tract (13.0% w/w), liver (4.8% w/w) and pancreas (1.5% w/w). Results show that pancreas had the highest Lipolytic activity. There were no significant differences in Lipolytic activity between pyloric caeca, intestine and liver (P
Philippe Thonart - One of the best experts on this subject based on the ideXlab platform.
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An enhanced process for the production of a highly purified extracellular lipase in the non-conventional yeast Yarrowia Lipolytica.
Applied Biochemistry and Biotechnology, 2010Co-Authors: Saoussen Turki, Philippe Thonart, Atef Ayed, Néjib Chalghoumi, Frederic Weekers, Héla KallelAbstract:Yarrowia Lipolytica LgX64.81 is a non-genetically modified mutant that was previously identified as a promising microorganism for extracellular lipase production. In this work, the development of a fed-batch process for the production of this enzyme in this strain was described. A Lipolytic activity of 2,145 U/mL was obtained after 32 h of batch culture in a defined medium supplemented with 10 g/L of tryptone, an enhancer of lipase expression. To maximize the volumetric productivity, two different fed-batch strategies had been investigated. In comparison to batch process, the intermittent fed-batch strategy had not improved the volumetric lipase productivity. In contrast, the stepwise feeding strategy combined with uncoupled cell growth and lipase production phases resulted in a 2-fold increase in the volumetric lipase productivity, namely, the lipase activity reached 10,000 U/mL after 80 h of culture. Furthermore, this lipase was purified to homogeneity by anion exchange chromatography on MonoQ resin followed by gel filtration on Sephacryl S-100. This process resulted in an overall yield of 72% and a 3.5-fold increase of the specific lipase activity. The developed process offers a great potential for an economic production of Lip2 at large scale in Y. Lipolytica LgX64.81.
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Methyl Oleate Modulates LIP2 Expression in the Lipolytic Yeast Yarrowia Lipolytica
Biotechnology Letters, 2005Co-Authors: Patrick Fickers, Jacqueline Destain, Philippe ThonartAbstract:Methyl oleate was used as a primary carbon source and as an alternative inducer for the production of an extracellular lipase, Lip2, in Y. Lipolytica strain LgX64.81 grown in a 20-l bioreactor. The lipase-encoding gene, LIP2 , was investigated during culture on methyl oleate using a p LIP2 – LacZ reporter fusion and we provide evidence for the involvement of methyl oleate in its regulation.
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Identification and characterisation of LIP7 and LIP8 genes encoding two extracellular triacylglycerol lipases in the yeast Yarrowia Lipolytica.
Fungal Genetics and Biology, 2005Co-Authors: Patrick Fickers, Philippe Thonart, Claude Gaillardin, Franck Fudalej, M-therese Le Dall, Serge Casaregola, Jean-marc NicaudAbstract:In the Lipolytic yeast Yarrowia Lipolytica, the LIP2 gene was previously reported to encode an extracellular lipase. The growth of a Deltalip2 strain on triglycerides as sole carbon source suggest an alternative pathway for triglycerides utilisation in this yeast. Here, we describe the isolation and the characterisation of the LIP7 and LIP8 genes which were found to encode a 366 and a 371-amino acid precursor protein, respectively. These proteins which belong to the triacylglycerol hydrolase family (EC 3.1.1.3) presented a high homology with the extracellular lipase CdLIP2 and CdLIP3 from Candida deformans. The physiological function of the lipase isoenzymes was investigated by creating single and multi-disrupted strains. Lip7p and Lip8p were found to correspond to active secreted lipases. The lack of lipase production in a Deltalip2 Deltalip7 Deltalip8 strain suggest that no additional extracellular lipase remains to be discovered in Y. Lipolytica. The substrate specificity towards synthetic ester molecules indicates that Lip7p presented a maximum activity centred on caproate (C6) while that of Lip8p is in caprate (C10).
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Carbon and nitrogen sources modulate lipase production in the yeast Yarrowia Lipolytica
Journal of Applied Microbiology, 2004Co-Authors: Patrick Fickers, Jacqueline Destain, Jean-marc Nicaud, Claude Gaillardin, Philippe ThonartAbstract:AIMS: To analyse the influence of nitrogen and carbon sources on extracellular lipase production by Yarrowia Lipolytica-overproducing mutant in order to optimize its production in large-scale bioreactors. METHODS AND RESULTS: The level of lipase production and LIP2 induction, measured using an LIP2-LacZ reporter gene, were compared for different carbon and nitrogen sources and for different concentrations. The localization of the enzyme during growth was also determined by Western blotting analysis using a six-histidine-tagged lipase. SIGNIFICANCE AND IMPACT OF THE STUDY: Tryptone N1 and oleic acid are the most suitable nitrogen and carbon sources for the production of the extracellular lipase by the Y. Lipolytica mutant. Higher levels of lipase production were obtained as the tryptone concentration increased in the culture medium. Such a positive correlation was not observed with oleic acid media where the highest Lipolytic productivities were obtained in the presence of low concentration. We also demonstrate that in the presence of oleic acid, lipase is cell-bound during the growth phase before being released in the media. CONCLUSIONS: This work provides a better understanding of the mechanism controlling LIP2 expression and, thus, extracellular lipase production in the yeast Y. Lipolytica.
Jean-marc Nicaud - One of the best experts on this subject based on the ideXlab platform.
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Identification and characterisation of LIP7 and LIP8 genes encoding two extracellular triacylglycerol lipases in the yeast Yarrowia Lipolytica.
Fungal Genetics and Biology, 2005Co-Authors: Patrick Fickers, Philippe Thonart, Claude Gaillardin, Franck Fudalej, M-therese Le Dall, Serge Casaregola, Jean-marc NicaudAbstract:In the Lipolytic yeast Yarrowia Lipolytica, the LIP2 gene was previously reported to encode an extracellular lipase. The growth of a Deltalip2 strain on triglycerides as sole carbon source suggest an alternative pathway for triglycerides utilisation in this yeast. Here, we describe the isolation and the characterisation of the LIP7 and LIP8 genes which were found to encode a 366 and a 371-amino acid precursor protein, respectively. These proteins which belong to the triacylglycerol hydrolase family (EC 3.1.1.3) presented a high homology with the extracellular lipase CdLIP2 and CdLIP3 from Candida deformans. The physiological function of the lipase isoenzymes was investigated by creating single and multi-disrupted strains. Lip7p and Lip8p were found to correspond to active secreted lipases. The lack of lipase production in a Deltalip2 Deltalip7 Deltalip8 strain suggest that no additional extracellular lipase remains to be discovered in Y. Lipolytica. The substrate specificity towards synthetic ester molecules indicates that Lip7p presented a maximum activity centred on caproate (C6) while that of Lip8p is in caprate (C10).
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Carbon and nitrogen sources modulate lipase production in the yeast Yarrowia Lipolytica
Journal of Applied Microbiology, 2004Co-Authors: Patrick Fickers, Jacqueline Destain, Jean-marc Nicaud, Claude Gaillardin, Philippe ThonartAbstract:AIMS: To analyse the influence of nitrogen and carbon sources on extracellular lipase production by Yarrowia Lipolytica-overproducing mutant in order to optimize its production in large-scale bioreactors. METHODS AND RESULTS: The level of lipase production and LIP2 induction, measured using an LIP2-LacZ reporter gene, were compared for different carbon and nitrogen sources and for different concentrations. The localization of the enzyme during growth was also determined by Western blotting analysis using a six-histidine-tagged lipase. SIGNIFICANCE AND IMPACT OF THE STUDY: Tryptone N1 and oleic acid are the most suitable nitrogen and carbon sources for the production of the extracellular lipase by the Y. Lipolytica mutant. Higher levels of lipase production were obtained as the tryptone concentration increased in the culture medium. Such a positive correlation was not observed with oleic acid media where the highest Lipolytic productivities were obtained in the presence of low concentration. We also demonstrate that in the presence of oleic acid, lipase is cell-bound during the growth phase before being released in the media. CONCLUSIONS: This work provides a better understanding of the mechanism controlling LIP2 expression and, thus, extracellular lipase production in the yeast Y. Lipolytica.
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identification of a triacylglycerol lipase gene family in candida deformans molecular cloning and functional expression
Yeast, 2003Co-Authors: Frederic Bigey, Jean-marc Nicaud, Karine Tuery, Daisy Bougard, Guy MoulinAbstract:The yeast Candida deformans CBS 2071 produces an extracellular lipase which was shown to catalyse the production of various esters by the esterification of free fatty acids, even in the presence of a large molar excess of water. To clone the gene encoding this extracellular lipase, Saccharomyces cerevisiae was transformed with C. deformans genomic libraries and screened for Lipolytic activity on a medium containing rapeseed oil emulsion and rhodamine B. Three members of a lipase gene family (CdLIP1, CdLIP2 and CdLIP3) were cloned and characterized. Each deduced lipase sequence has a Gly–His–Ser–Leu–Gly–(Gly/Ala)–Ala conserved motif, eight cysteine residues and encodes an N-terminal signal sequence. MALDI–TOF mass spectrometry analysis of a proteolytic digest of the lipase produced was used to obtain experimental evidence that the CdLIP1 gene encoded the extracellular lipase. Recombinant expression studies confirmed that the cloned genes encoded functional lipases. The three lipases are very similar to lipases from the related species Yarrowia Lipolytica. Significant homologies were also found with several yeast and fungal lipases. As C. deformans CBS 2071 was previously considered to be synonymous with Y. Lipolytica, the strains were compared for the extent of nucleotide divergence in the variable regions (D1/D2) at the 5′-end of the large-subunit (26S) ribosomal DNA (rDNA) gene. This rDNA region has diverged sufficiently to suggest that C. deformans is a separate species. The nucleotide sequences of the CdLIP1, CdLIP2 and CdLIP3 genes will appear in the EMBL nucleotide sequence database under Accession Nos AJ428393, AJ428394 and AJ428395, respectively. Copyright © 2003 John Wiley & Sons, Ltd.
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Identification of a triacylglycerol lipase gene family in Candida deformans: molecular cloning and functional expression
Yeast, 2003Co-Authors: Frederic Bigey, Jean-marc Nicaud, Karine Tuery, Daisy Bougard, Guy MoulinAbstract:The yeast Candida deformans CBS 2071 produces an extracellular lipase which was shown to catalyse the production of various esters by the esterification of free fatty acids, even in the presence of a large molar excess of water. To clone the gene encoding this extracellular lipase, Saccharomyces cerevisiae was transformed with C deformans genomic libraries and screened for Lipolytic activity on a medium containing rapeseed oil emulsion and rhodamine B. Three members of a lipase gene family (CdLIP1, CdLIP2 and CdLIP3) were cloned and characterized. Each deduced lipase sequence has a Gly-His-Ser-Leu-Gly-(Gly/Ala)-Ala conserved motif, eight cysteine residues and encodes an N-terminal signal sequence. MALDI-TOF mass spectrometry analysis of a proteolytic digest of the lipase produced was used to obtain experimental evidence that the CdLIP1 gene encoded the extracellular lipase. Recombinant expression studies confirmed that the cloned genes encoded functional lipases. The three lipases are very similar to lipases from the related species Yarrowia Lipolytica. Significant homologies were also found with several yeast and fungal lipases. As C. deformans CBS 2071 was previously considered to be synonymous with Y. Lipolytica, the strains were compared for the extent of nucleotide divergence in the variable regions (D1/D2) at the F-end of the large-subunit (26S) ribosomal DNA (rDNA) gene. This rDNA region has diverged sufficiently to suggest that C. deformans is a separate species. The nucleotide sequences of the CdLIP1, CdLIP2 and CdLIP3 genes will appear in the EMBL nucleotide sequence database under Accession Nos AJ428393, AJ428394 and AJ428395, respectively.
Juergen Wiegel - One of the best experts on this subject based on the ideXlab platform.
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thermosyntropha Lipolytica gen nov sp nov a Lipolytic anaerobic alkalitolerant thermophilic bacterium utilizing short and long chain fatty acids in syntrophic coculture with a methanogenic archaeum
International Journal of Systematic and Evolutionary Microbiology, 1996Co-Authors: Vitalii Svetlitshnyi, Fred A Rainey, Juergen WiegelAbstract:Three strains of an anaerobic thermophilic organoheterotrophic Lipolytic alkalitolerant bacterium, Thermosyntropha Lipolytica gen. nov., sp. nov. (type strain JW/VS-265T; DSM 11003), were isolated from alkaline hot springs of Lake Bogoria (Kenya). The cells were nonmotile, non-spore forming, straight or slightly curved rods. At 60°C the pH range for growth determined at 25°C [pH25°C] was 7.15 to 9.5, with an optimum between 8.1 and 8.9 (pH60°C of 7.6 and 8.1). At a pH25°C of 8.5 the temperature range for growth was from 52 to 70°C, with an optimum between 60 and 66°C. The shortest doubling time was around 1 h. In pure culture the bacterium grew in a mineral base medium supplemented with yeast extract, tryptone, Casamino Acids, betaine, and crotonate as carbon sources, producing acetate as a major product and constitutively a lipase. During growth in the presence of olive oil, free long-chain fatty acids were accumulated in the medium but the pure culture could not utilize olive oil, triacylglycerols, short- and long-chain fatty acids, and glycerol for growth. In syntrophic coculture (Methanobacterium strain JW/VS-M29) the Lipolytic bacteria grew on triacylglycerols and linear saturated and unsaturated fatty acids with 4 to 18 carbon atoms, but glycerol was not utilized. Fatty acids with even numbers of carbon atoms were degraded to acetate and methane, while from odd-numbered fatty acids 1 mol of propionate per mol of fatty acid was additionally formed. 16S rDNA sequence analysis identified Syntrophospora and Syntrophomonas spp. as closest phylogenetic neighbors.