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Florence Husson - One of the best experts on this subject based on the ideXlab platform.
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Predicted secondary structure of hydroperoxide lyase from Green Bell Pepper cloned in the yeast Yarrowia lipolytica
Journal of Molecular Catalysis B: Enzymatic, 2010Co-Authors: Mirna Santiago-gómez, Jean-marc Nicaud, Jean-marc Belin, Selim Kermasha, Florence HussonAbstract:Fatty acid hydroperoxide lyase (HPL) is a member of the cytochrome P450 family acting on fatty acid hydroperoxides in many organisms. The active Green Bell Pepper HPL, cloned and expressed in the yeast Yarrowia lipolytica, was purified by immobilized metal-ion affinity chromatography (IMAC) in the presence of 2% of Triton X-100R. The secondary structure prediction by bioinformatics servers of HPL was realized by ANTHEPROT software, using the GOR, DPM and Predator methods. The theoretical results which are average values obtained from three different calculation methods showed 33% α-helix, 18% β-sheet, 7% turn and 42% coil. On the other hand, the secondary structure approach of the purified active HPL (specific activity of 2.94 U/mg protein) was realized by differential scanning calorimetry (DSC) and circular dichroism (CD) spectroscopy, and showed 13% α-helix, 29% β-sheet, 5% turn and 53% random coil.
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modeling hexanal production in oxido reducing conditions by the yeast yarrowia lipolytica
Process Biochemistry, 2009Co-Authors: Mirna P Santiagogomez, Jean-marc Belin, Selim Kermasha, Hoa Tran Thanh, Joelle De Coninck, Remy Cachon, Patrick Gervais, Florence HussonAbstract:Hexanal produced by cells of a recombinant Yarrowia lipolytica yeast expressing the hydroperoxide lyase (HPL) from Green Bell Pepper fruit was studied under oxido-reducing conditions using the reducing dithiotreitol and oxidizing potassium ferricyanide compounds. The combined effect of pH, linoleic acid 13-hydroperoxides concentration, temperature and oxido-reducing molecules on the hexanal production was studied. Significant positive effects for the hexanal production were found using high concentrations of hydroperoxides (100 mM, 30 g/L). Adding reducing molecules enhanced significantly hexanal production while the oxidizing molecules had an inhibitory effect. Combined effects of 13-hydroperoxides and dithiotreitol were optimised by a central composite design and a model was proposed. Finally, 6 mM (600 mg/L) of hexanal was obtained when 119 mM of 13-hydroperoxides (37 g/L) and 50 mM of dithiotreitol were introduced directly in the biocatalytic medium of the yeast Y. lipolytica.
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Secondary structure conformation of hydroperoxide lyase from Green Bell Pepper, cloned in Yarrowia lipolytica, and its activity in selected media
Journal of Molecular Catalysis B: Enzymatic, 2008Co-Authors: Mirna Santiago-gómez, Jean-marc Nicaud, Jean-marc Belin, Selim Kermasha, Florence HussonAbstract:Circular dichroism (CD) spectroscopy of secondary structure conformation of the purified Green Bell Pepper hydroperoxide lyase (HPL), cloned in the yeast Yarrowia lipolytica, was investigated. The CD spectra of HPL in iso-octane, obtained at 60 °C, in the presence of the reducing agent dithiothreitol showed dramatic increase in α-helix content. The enzyme conformation remained unchanged over a range of pH values of 5.0–7.0. Using 13-hydroperoxide of linoleic acid (13-HPOD) as substrate, the biocatalysis of HPL in organic solvent media, including chloroform, dichloromethane, hexane, iso-octane, octane and toluene, was investigated. The results indicated an increase in HPL activity in the biphasic hexane medium.
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Toxicity of fatty acid hydroperoxides towards Yarrowia lipolytica: Implication of their membrane fluidizing action
Biochimica et Biophysica Acta:Biomembranes, 2007Co-Authors: Hoa Tran Thanh, Jean-marc Belin, Patrick Gervais, Laurent Beney, Hélène Simonin, Thi Xuan Sam Nguyen, Florence HussonAbstract:Linoleic acid hydroperoxide (HPOD), substrate of hydroperoxide lyase, an enzyme of the lipoxygenase pathway, can be transformed into many aromatic compounds, the so-called “Green notes”. The presence of linoleic acid hydroperoxide in the culture medium of Yarrowia lipolytica, the yeast expressing the cloned hydroperoxide lyase of Green Bell Pepper, undoubtedly exerted an inhibition on the growth and a toxic effect with 90% of yeast cells died after 120 min of exposition in 100 mM HPOD solution. The increase in cell membrane fluidity evaluated by measuring fluorescence generalized polarization with the increasing concentration of HPOD in the medium confirmed the fluidizing action of HPOD on yeast membrane. In addition, we determined by infrared spectroscopy measurement that this compound rapidly diffused into model phospholipids [1, 2-Dimyristoyl-D54-sn-Glycero-3-Phosphocholine (DMPC-D54)] bilayer, modifying their general physical state and their phase transition. In the presence of various concentrations of HPOD, the phase transition of DMPC-D54 occurred with an increase of both the corresponding wave number shift and the temperature range but the phase transition temperature was not modified. These results show that the toxic effects of HPOD on the yeast Yarrowia lipolytica may be initially linked to a strong interaction of this compound with the cell membrane phospholipids and components.
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Characterization of Purified Green Bell Pepper Hydroperoxide Lyase Expressed by Yarrowia Lipolytica: Radicals Detection during Catalysis
Enzyme and Microbial Technology, 2007Co-Authors: Mirna P. Santiago-gómez, Jean-marc Nicaud, Jean-marc Belin, Catherine Vergely, Clotilde Policar, Luc Rochette, Florence HussonAbstract:The optimization of the expression of recombinant 6-His-tagged HPO lyase in Yarrowia lipolytica is described: 1800U/L of culture was detected at 24h of culture on a medium containing olive oil as the sole carbon source. The enzyme was purified by IMAC and showed an optimal pH at 5.5, an optimal temperature at 20^\circC and a Km value of 9μM with 13-HPOD substrate. The participation of radicals during the catalysis of purified Bell Pepper fruit hydroperoxide lyase has been observed by electron paramagnetic resonance spectroscopy and the yet unidentified radical species might be an alkyl or alkoxyl radical linked to the enzyme.
Jean-marc Belin - One of the best experts on this subject based on the ideXlab platform.
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Predicted secondary structure of hydroperoxide lyase from Green Bell Pepper cloned in the yeast Yarrowia lipolytica
Journal of Molecular Catalysis B: Enzymatic, 2010Co-Authors: Mirna Santiago-gómez, Jean-marc Nicaud, Jean-marc Belin, Selim Kermasha, Florence HussonAbstract:Fatty acid hydroperoxide lyase (HPL) is a member of the cytochrome P450 family acting on fatty acid hydroperoxides in many organisms. The active Green Bell Pepper HPL, cloned and expressed in the yeast Yarrowia lipolytica, was purified by immobilized metal-ion affinity chromatography (IMAC) in the presence of 2% of Triton X-100R. The secondary structure prediction by bioinformatics servers of HPL was realized by ANTHEPROT software, using the GOR, DPM and Predator methods. The theoretical results which are average values obtained from three different calculation methods showed 33% α-helix, 18% β-sheet, 7% turn and 42% coil. On the other hand, the secondary structure approach of the purified active HPL (specific activity of 2.94 U/mg protein) was realized by differential scanning calorimetry (DSC) and circular dichroism (CD) spectroscopy, and showed 13% α-helix, 29% β-sheet, 5% turn and 53% random coil.
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modeling hexanal production in oxido reducing conditions by the yeast yarrowia lipolytica
Process Biochemistry, 2009Co-Authors: Mirna P Santiagogomez, Jean-marc Belin, Selim Kermasha, Hoa Tran Thanh, Joelle De Coninck, Remy Cachon, Patrick Gervais, Florence HussonAbstract:Hexanal produced by cells of a recombinant Yarrowia lipolytica yeast expressing the hydroperoxide lyase (HPL) from Green Bell Pepper fruit was studied under oxido-reducing conditions using the reducing dithiotreitol and oxidizing potassium ferricyanide compounds. The combined effect of pH, linoleic acid 13-hydroperoxides concentration, temperature and oxido-reducing molecules on the hexanal production was studied. Significant positive effects for the hexanal production were found using high concentrations of hydroperoxides (100 mM, 30 g/L). Adding reducing molecules enhanced significantly hexanal production while the oxidizing molecules had an inhibitory effect. Combined effects of 13-hydroperoxides and dithiotreitol were optimised by a central composite design and a model was proposed. Finally, 6 mM (600 mg/L) of hexanal was obtained when 119 mM of 13-hydroperoxides (37 g/L) and 50 mM of dithiotreitol were introduced directly in the biocatalytic medium of the yeast Y. lipolytica.
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Secondary structure conformation of hydroperoxide lyase from Green Bell Pepper, cloned in Yarrowia lipolytica, and its activity in selected media
Journal of Molecular Catalysis B: Enzymatic, 2008Co-Authors: Mirna Santiago-gómez, Jean-marc Nicaud, Jean-marc Belin, Selim Kermasha, Florence HussonAbstract:Circular dichroism (CD) spectroscopy of secondary structure conformation of the purified Green Bell Pepper hydroperoxide lyase (HPL), cloned in the yeast Yarrowia lipolytica, was investigated. The CD spectra of HPL in iso-octane, obtained at 60 °C, in the presence of the reducing agent dithiothreitol showed dramatic increase in α-helix content. The enzyme conformation remained unchanged over a range of pH values of 5.0–7.0. Using 13-hydroperoxide of linoleic acid (13-HPOD) as substrate, the biocatalysis of HPL in organic solvent media, including chloroform, dichloromethane, hexane, iso-octane, octane and toluene, was investigated. The results indicated an increase in HPL activity in the biphasic hexane medium.
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Toxicity of fatty acid hydroperoxides towards Yarrowia lipolytica: Implication of their membrane fluidizing action
Biochimica et Biophysica Acta:Biomembranes, 2007Co-Authors: Hoa Tran Thanh, Jean-marc Belin, Patrick Gervais, Laurent Beney, Hélène Simonin, Thi Xuan Sam Nguyen, Florence HussonAbstract:Linoleic acid hydroperoxide (HPOD), substrate of hydroperoxide lyase, an enzyme of the lipoxygenase pathway, can be transformed into many aromatic compounds, the so-called “Green notes”. The presence of linoleic acid hydroperoxide in the culture medium of Yarrowia lipolytica, the yeast expressing the cloned hydroperoxide lyase of Green Bell Pepper, undoubtedly exerted an inhibition on the growth and a toxic effect with 90% of yeast cells died after 120 min of exposition in 100 mM HPOD solution. The increase in cell membrane fluidity evaluated by measuring fluorescence generalized polarization with the increasing concentration of HPOD in the medium confirmed the fluidizing action of HPOD on yeast membrane. In addition, we determined by infrared spectroscopy measurement that this compound rapidly diffused into model phospholipids [1, 2-Dimyristoyl-D54-sn-Glycero-3-Phosphocholine (DMPC-D54)] bilayer, modifying their general physical state and their phase transition. In the presence of various concentrations of HPOD, the phase transition of DMPC-D54 occurred with an increase of both the corresponding wave number shift and the temperature range but the phase transition temperature was not modified. These results show that the toxic effects of HPOD on the yeast Yarrowia lipolytica may be initially linked to a strong interaction of this compound with the cell membrane phospholipids and components.
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Characterization of Purified Green Bell Pepper Hydroperoxide Lyase Expressed by Yarrowia Lipolytica: Radicals Detection during Catalysis
Enzyme and Microbial Technology, 2007Co-Authors: Mirna P. Santiago-gómez, Jean-marc Nicaud, Jean-marc Belin, Catherine Vergely, Clotilde Policar, Luc Rochette, Florence HussonAbstract:The optimization of the expression of recombinant 6-His-tagged HPO lyase in Yarrowia lipolytica is described: 1800U/L of culture was detected at 24h of culture on a medium containing olive oil as the sole carbon source. The enzyme was purified by IMAC and showed an optimal pH at 5.5, an optimal temperature at 20^\circC and a Km value of 9μM with 13-HPOD substrate. The participation of radicals during the catalysis of purified Bell Pepper fruit hydroperoxide lyase has been observed by electron paramagnetic resonance spectroscopy and the yet unidentified radical species might be an alkyl or alkoxyl radical linked to the enzyme.
Talwinder S. Kahlon - One of the best experts on this subject based on the ideXlab platform.
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In vitro Bile Acid Binding of Kale, Mustard Greens, Broccoli, Cabbage and Green Bell Pepper Improves with Microwave Cooking
Vegetos, 2012Co-Authors: Talwinder S. Kahlon, Rebecca R. Milczarek, Mei-chin M. ChillAbstract:Bile acid binding potential of foods and food fractions has been related to lowering the risk of heart disease and that of cancer. Sauteing or steam cooking has been observed to significantly improve bile acid binding of Green/leafy vegetables. It was hypothesized that microwave cooking could improve the bile acid binding of various vegetables. Microwave cooking with no added water resulted in bile acid binding measured on a dry matter basis relative to cholestyramine of 13% for kale and mustard Greens, 9% for broccoli and collard Greens, 4% for cabbage, and 2% for Green Bell Pepper. These results point to the significantly different (P ≤ 0.05) health promoting potential of kale = mustard Greens > broccoli = collard Greens > cabbage > Green Bell Pepper. Microwave cooking with no added water significantly improved in vitro bile acid binding of kale, mustard Greens, broccoli and cabbage compared with microwave cooking with added water or raw (uncooked). For Green Bell Pepper, binding values were similar when microwave cooked with or without added water and significantly higher than uncooked. In the case of collard Greens, bile acid binding values were similar for raw or microwave cooked with added water and were significantly lowered by microwave cooking with no added water. Microwave cooking with no added water should be used for kale, mustard Greens, broccoli and cabbage; for collard Greens and Green Bell Pepper microwave with added water as the cooking method. These Green/leafy vegetables, when consumed regularly after microwave cooking would promote a healthy lifestyle, keep dietary fat low, and have the potential to lower the risk of premature degenerative diseases.
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In vitro Bile Acid Binding of Mustard Greens, Kale, Broccoli, Cabbage and Green Bell Pepper Improves with Sautéing Compared with Raw or Other Methods of Preparation
Food and Nutrition Sciences, 2012Co-Authors: Talwinder S. Kahlon, Rebecca R. Milczarek, Mei-chin M. ChiuAbstract:Bile acid binding potential of foods and food fractions has been related to lowering the risk of heart disease and that of cancer. Steam cooking has been observed to significantly improve bile acid binding of Green/leafy vegetables. It was hypothesized that other cooking methods could further improve the bile acid binding of various vegetables. Sautee cooking resulted in in vitro bile acid binding measured on a dry matter basis relative to cholestyramine of 14% for mustard Greens and kale, 9% for broccoli, 8% for collard Greens, 6% for cabbage, and 5% for Green Bell Pepper. These results point to the significantly different (P ≤ 0.05) health promoting potential of mustard Greens = kale > broccoli > collard Greens > cabbage > Green Bell Pepper. Sauteing significantly improved in vitro bile acid binding of mustard Greens, kale, broccoli, cabbage and Green Bell Pepper compared with steaming, boiling or raw (uncooked). Collard Greens exhibited significantly higher bile acid binding by steaming compared with sauteing, boiling or raw. Data suggest that the cooking method with most heath promoting potential for mustard Greens, kale, broccoli, cabbage and Green Bell Pepper should be sauteing. Steaming should be used for collard Greens as the cooking method. These Green/leafy vegetables, when consumed regularly after sauteing, would promote a healthy lifestyle and have the potential to lower the risk of premature degenerative diseases.
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Steam cooking significantly improves in vitro bile acid binding of collard Greens, kale, mustard Greens, broccoli, Green Bell Pepper, and cabbage☆
Nutrition research (New York N.Y.), 2008Co-Authors: Talwinder S. Kahlon, Mei-chen M. Chiu, Mary H. ChapmanAbstract:Bile acid binding capacity has been related to the cholesterol-lowering potential of foods and food fractions. Lowered recirculation of bile acids results in utilization of cholesterol to synthesize bile acid and reduced fat absorption. Secondary bile acids have been associated with increased risk of cancer. Bile acid binding potential has been related to lowering the risk of heart disease and that of cancer. Previously, we have reported bile acid binding by several uncooked vegetables. However, most vegetables are consumed after cooking. How cooking would influence in vitro bile acid binding of various vegetables was investigated using a mixture of bile acids secreted in human bile under physiological conditions. Eight replicate incubations were conducted for each treatment simulating gastric and intestinal digestion, which included a substrate only, a bile acid mixture only, and 6 with substrate and bile acid mixture. Cholestyramine (a cholesterol-lowering, bile acid binding drug) was the positive control treatment and cellulose was the negative control. Relative to cholestyramine, in vitro bile acid binding on dry matter basis was for the collard Greens, kale, and mustard Greens, 13%; broccoli, 10%; Brussels sprouts and spinach, 8%; Green Bell Pepper, 7%; and cabbage, 5%. These results point to the significantly different (P broccoli > Brussels sprouts = spinach = Green Bell Pepper > cabbage as indicated by their bile acid binding on dry matter basis. Steam cooking significantly improved the in vitro bile acid binding of collard Greens, kale, mustard Greens, broccoli, Green Bell Pepper, and cabbage compared with previously observed bile acid binding values for these vegetables raw (uncooked). Inclusion of steam-cooked collard Greens, kale, mustard Greens, broccoli, Green Bell Pepper, and cabbage in our daily diet as health-promoting vegetables should be emphasized. These Green/leafy vegetables, when consumed regularly after steam cooking, would lower the risk of cardiovascular disease and cancer, advance human nutrition research, and improve public health.
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In vitro binding of bile acids by spinach, kale, brussels sprouts, broccoli, mustard Greens, Green Bell Pepper, cabbage and collards
Food Chemistry, 2007Co-Authors: Talwinder S. Kahlon, Mary H. Chapman, G.e. SmithAbstract:Abstract The in vitro binding of bile acids by spinach ( Spinacia oleracea ), kale ( Brassica oleracea acephala ), Brussels sprouts ( Brassica oleracea gemmifera ), broccoli ( Brassica oleracea italica ), mustard Greens ( Brassica juncea ), Green Bell Peppers ( Capsicum annuum ), cabbage ( Brassica oleracea capitala ) and collards ( Brassica oleracea acephala ) was determined using a mixture of bile acids secreted in human bile at a duodenal physiological pH of 6.3. Six treatments and two blank incubations were conducted testing various fresh raw Green vegetables on an equal dry matter basis. Considering cholestyramine (bile acid binding, cholesterol lowering drug) as 100% bound, the relative in vitro bile acid binding of various vegetables tested on equal dry matter and total dietary fibre basis was 2–9% and 6–32%, respectively. Bile acid binding for spinach, kale and brussels sprouts was significantly higher than for broccoli and mustard Greens. For broccoli and mustard Greens binding values were significantly higher those for cabbage, Bell Pepper and collards. These results point to the health promoting potential of spinach = kale = brussels sprouts > broccoli = mustard Greens > cabbage = Green Bell Peppers = collards, as indicated by their bile acid binding on dry matter basis.
Cornelis Van Leeuwen - One of the best experts on this subject based on the ideXlab platform.
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Impact of clonal variability in Vitis vinifera Cabernet franc on grape composition, wine quality, leafblade stilbene content, and downy mildew resistance
Journal of Agricultural and Food Chemistry, 2013Co-Authors: Cornelis Van Leeuwen, Jean Philippe Roby, Virginia Alonso-villaverde, Katia GindroAbstract:In this study, 10 clones of Vitis vinifera Cabernet franc (not yet commercial) have been phenotyped on precocity, grape composition, and assessment of wine quality made by microvinification in 2008–2010. Additionally, two original criteria have been considered: concentration of 3-isobutyl-2-methoxypyrazine (IBMP) in grapes and wines (the Green Bell Pepper flavor) and resistance of grapevines to downy mildew (Plasmopara viticola) by stilbene quantification upon infection. Precocity of veraison varied up to four days at veraison. Berry size and yield were highly variable among clones. However, these variables were not correlated. Tanins and anthocyanins varied among clones in grapes and wines. Variations in grape and wine IBMP were not significant. Some clones showed lower susceptibility for downy mildew on leaves. Lower susceptibility was linked to a higher production of stilbenic phytoalexins involved in downy mildew resistance mechanisms.
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Organoleptic Impact of 2-Methoxy-3-isobutylpyrazine on Red Bordeaux and Loire Wines. Effect of Environmental Conditions on Concentrations in Grapes during Ripening
Journal of agricultural and food chemistry, 2000Co-Authors: Dominique Roujou De Boubee, Cornelis Van Leeuwen, Denis DubourdieuAbstract:The 2-methoxy-3-isobutylpyrazine content in grapes and red wines was assayed by stable isotope dilution gas chromatography-mass spectrometry, following vapor extraction and purification on a cation resin microcolumn. The threshold beyond which the Green Bell Pepper character is marked in wines has been determined. From a comparison of the 2-methoxy-3-isobutylpyrazine concentrations of 50 red Bordeaux and Loire wines from different vintages and grape varieties (Cabernet Sauvignon, Cabernet franc, and Merlot) with the intensity of the Green Bell Pepper character as perceived on tasting, the threshold value was estimated to be 15 ng/L. Statistical analysis of the 2-methoxy-3-isobutylpyrazine concentrations of 89 red Bordeaux wines showed that Cabernet wines were more commonly affected by this vegetative character. Changes in the 2-methoxy-3-isobutylpyrazine concentration as the grapes ripen are affected by the environmental and cultural conditions (soil, climate, training system, etc.). A very good correlation was shown between the breakdown of malic acid and 2-methoxy-3-isobutylpyrazine as the grapes ripened, irrespective of grape variety, type of soil, or weather conditions.
Katia Gindro - One of the best experts on this subject based on the ideXlab platform.
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Impact of clonal variability in Vitis vinifera Cabernet franc on grape composition, wine quality, leafblade stilbene content, and downy mildew resistance
Journal of Agricultural and Food Chemistry, 2013Co-Authors: Cornelis Van Leeuwen, Jean Philippe Roby, Virginia Alonso-villaverde, Katia GindroAbstract:In this study, 10 clones of Vitis vinifera Cabernet franc (not yet commercial) have been phenotyped on precocity, grape composition, and assessment of wine quality made by microvinification in 2008–2010. Additionally, two original criteria have been considered: concentration of 3-isobutyl-2-methoxypyrazine (IBMP) in grapes and wines (the Green Bell Pepper flavor) and resistance of grapevines to downy mildew (Plasmopara viticola) by stilbene quantification upon infection. Precocity of veraison varied up to four days at veraison. Berry size and yield were highly variable among clones. However, these variables were not correlated. Tanins and anthocyanins varied among clones in grapes and wines. Variations in grape and wine IBMP were not significant. Some clones showed lower susceptibility for downy mildew on leaves. Lower susceptibility was linked to a higher production of stilbenic phytoalexins involved in downy mildew resistance mechanisms.
