Lupin is a leguminous plant whose edible seeds display amazing properties which is worth to prope... more Lupin is a leguminous plant whose edible seeds display amazing properties which is worth to properly exploit in view of the great demand of plant-derived food proteins. To discuss this matter and get the most updated advances in a disciplinary integrated way, the experts in the field met at the XIV edition of the International Lupin Conference last June in Milan, Italy. This is the report of the scientific outcomes of the conference and the satellite events, including two dedicated workshops and a unique Lupin banquet based on lupin foods. The results of a hedonic test performed on the lupin foods provided during the banquet are reported as well
Caco-2 cell monolayers to study the transport of lupin conglutin gamma across human intestinal ep... more Caco-2 cell monolayers to study the transport of lupin conglutin gamma across human intestinal epithelium / A. SCARAFONI, J. CAPRARO, C. MAGNI, A. CLEMENTE, M. DURANTI. - In: Proceedings of the 53rd National Meeting of the Italian Society of Biochemistry and Molecular Biology ( ...
gamma-Conglutin, a lupin seed glycoprotein, is capable of positively influencing glucose uptake b... more gamma-Conglutin, a lupin seed glycoprotein, is capable of positively influencing glucose uptake by various model cells and reducing glycaemia in animal models and human subjects. In particular, gamma-conglutin can activate in differentiating myocyte different signaling pathways that closely resembled those of insulin. A recent study aimed at identifying the metabolic fate of the protein, showed that it was transcytosed through a CaCo2 cell monolayer and crossed an ex-vivo intestinal barrier in an intact form. Beside, the protein displays strong resistance to endogenous and exogenous proteolytic enzymes. These findings encouraged us to further study the molecular mechanisms of action of this bioactive protein, which are still not completely elucidated. Aim of the present work was to study how gamma-conglutin interact with the target cells and show if the protein undergoes any covalent change during the process. 2D-electrophoresis of the gamma-conglutin treated cell lysates and antibody revelation of the blotted maps showed the presence of the intact protein inside the cells. Moreover, spots with an unexpectedly low pI, not present in the natural protein, were detected. These spots were submitted to MS/MS spectrometric analysis. Phosphorylated amino acids were evidenced in some gamma-conglutin peptides. To unveil gamma-conglutin cellular fate, HepG2 cells treated with the lupin protein were submitted to confocal microscopy and transmission electron microscopy (TEM). gamma-Conglutin was found accumulated inside the cells and spread as protein aggregates into the cytoplasm. Microvilli were involved in trapping the protein at the beginning of the process. A prerequisite for any biological activity is the interaction with or entry into the target cells. These findings, by showing that gamma-conglutin is uptaked by HepG2 cells in an intact form and is modified by multiple phosphorylation, open the way to new studies for the understanding of the described gamma-conglutin insulin-mimetic activity
Two-dimensional (2-D) IEF/SDS-PAGE is a powerful tool to get molecular “pictures” of food proteom... more Two-dimensional (2-D) IEF/SDS-PAGE is a powerful tool to get molecular “pictures” of food proteomes and monitor the processing effect(s) of a given food item on its protein profile. Still the use of 2-D approaches to this aim is rather uncommon.In this work, 2-D electrophoresis has been used to monitor the main steps of lupin-based gluten-free pasta production. Three different production
Citation: Improved lupin proteins isolation and molecular characterisation in view of their explo... more Citation: Improved lupin proteins isolation and molecular characterisation in view of their exploitation as food ingredients/M. Duranti, C. Magni, E. Sironi, F. Sessa, A. Scarafoni-In: Proceedings of the Final Conference of the European Project: Healthy ProFood/[a cura di] ...
2D electrophoretic mapping as a tool for the analysis of seed proteins in legume-based raw materi... more 2D electrophoretic mapping as a tool for the analysis of seed proteins in legume-based raw materials and novel foods / E. Sironi, A. Scarafoni, F. Sessa, C. Magni, V. Dani, M. Duranti - In: 5th European Conference on Grain Legumes and the 2nd International Conference on Legume ...
Adzuki seed β-vignin, a vicilin-like globulin, has proven to exert various health-promoting biolo... more Adzuki seed β-vignin, a vicilin-like globulin, has proven to exert various health-promoting biological activities, notably in cardiovascular health. A simple scalable enrichment procedure of this protein for further nutritional and functional studies is crucial. In this study, a simplified chromatography-independent protein fractionation procedure has been optimized and described. The electrophoretic analysis showed a high degree of homogeneity of β-vignin isolate. Furthermore, the molecular features of the purified protein were investigated. The adzuki bean β-vignin was found to have a native size of 146 kDa, and the molecular weight determined was consistent with a trimeric structure. These were identified in two main polypeptide chains (masses of 56–54 kDa) that are glycosylated polypeptides with metal binding capacity, and one minor polypeptide chain with a mass 37 kDa, wherein these features are absent. The in vitro analysis showed a high degree of digestibility of the protein ...
Biological activity, traceability and potential allergenicity of selected lupin proteins / M. Dur... more Biological activity, traceability and potential allergenicity of selected lupin proteins / M. Duranti, E. Sironi, C. Magni, F. Sessa, A. Scarafoni - In: Proceedings of the Final Conference of the European Project : Healthy ProFood / [a cura di] A. Arnoldi. - Roma : Aracne, 2005 ...
Biological activity, traceability and potential allergenicity of selected lupin proteins / M. Dur... more Biological activity, traceability and potential allergenicity of selected lupin proteins / M. Duranti, E. Sironi, C. Magni, F. Sessa, A. Scarafoni - In: Proceedings of the Final Conference of the European Project : Healthy ProFood / [a cura di] A. Arnoldi. - Roma : Aracne, 2005 ...
Food proteins and peptides are able to exert a variety of well-known bioactivities, some of which... more Food proteins and peptides are able to exert a variety of well-known bioactivities, some of which are related to well-being and disease prevention in humans and animals. Currently, an active trend in research focuses on chronic inflammation and oxidative stress, delineating their major pathogenetic role in age-related diseases and in some forms of cancer. The present study aims to investigate the potential effects of pseudocereal proteins and their derived peptides on chronic inflammation and oxidative stress. After purification and attribution to protein classes according to classic Osborne’s classification, the immune-modulating, antioxidant, and trypsin inhibitor activities of proteins from quinoa (Chenopodium quinoa Willd.), amaranth (Amaranthus retroflexus L.), and buckwheat (Fagopyrum esculentum Moench) seeds have been assessed in vitro. The peptides generated by simulated gastro-intestinal digestion of each fraction have been also investigated for the selected bioactivities. ...
γ-conglutin (γC) is a major protein of Lupinus albus seeds, but its function is still unknown. It... more γ-conglutin (γC) is a major protein of Lupinus albus seeds, but its function is still unknown. It shares high structural similarity with xyloglucan-specific endo-glucanase inhibitor proteins (XEGIPs) and, to a lesser extent, with Triticum aestivum endoxylanase inhibitors (TAXI-I), active against fungal glycoside hydrolases GH12 and GH11, respectively. However, γC lacks both these inhibitory activities. Since β-galactomannans are major components of the cell walls of endosperm in several legume plants, we tested the inhibitory activity of γC against a GH2 β-mannosidase (EC 3.2.1.25). γC was actually able to inhibit the enzyme, and this effect was enhanced by the presence of zinc ions. The stoichiometry of the γC/enzyme interaction was 1:1, and the calculated Ki was 1.55 μM. To obtain further insights into the interaction between γC and β-mannosidase, an in silico structural bioinformatic approach was followed, including some docking analyses. By and large, this work describes experim...
International journal of food sciences and nutrition, Jan 18, 2017
Fermentation represents a valuable and cost-effective approach for food stabilisation and nutriti... more Fermentation represents a valuable and cost-effective approach for food stabilisation and nutritional improvement. Tempeh is an example of soybean solid-state fermentation. In this work, we investigated the possibility of producing a tempeh analogue containing high amounts of vitamin B12 using seeds of three different species of the legume lupin, namely Lupinus albus, L. angustifolius and L. mutabilis, with Rhizopus oligosporus and Propionibacterium freudenreichii cofermentation. Synergic effects of Rhizopus and Propionibacterium in increasing vitamin B12 up to 1230 ng/g dw was observed. These findings indicate that this cofermentation can improve lupin nutritional quality and safety to provide a tempeh analogue with added value for vegan and vegetarian communities and low-income populations. The level of potentially toxic lupin alkaloids was also monitored during the tempeh preparation.
Biochemical and Biophysical Research Communications, 2013
Lupin seed γ-Conglutin is a protein capable of reducing glycaemia in mammalians and increasing gl... more Lupin seed γ-Conglutin is a protein capable of reducing glycaemia in mammalians and increasing glucose uptake by model cells. This work investigated whether γ-Conglutin is internalised into the target cells and undergoes any covalent change during the process, as a first step to understanding its mechanism of action. To this purpose, γ-Conglutin-treated and untreated HepG2 cells were submitted to confocal and transmission electron microscopy. Immune-revelation of γ-Conglutin at various intervals revealed its accumulation inside the cytosol. In parallel, 2D-electrophoresis of the cell lysates and antibody reaction of the blotted maps showed the presence of the protein intact subunits inside the treated cells, whilest no trace of the protein was found in the control cells. However, γ-Conglutin-related spots with an unexpectedly low pI were also observed in the maps. These spots were excised, trypsin-treated and submitted to MS/MS spectrometric analysis. The presence of phosphorylated amino acids was detected. These findings, by showing that γ-Conglutin is internalised by HepG2 cells in an intact form and is modified by multiple phosphorylation, open the way to the understanding of the lupin γ-Conglutin insulin-mimetic activity.
Lupin is a leguminous plant whose edible seeds display amazing properties which is worth to prope... more Lupin is a leguminous plant whose edible seeds display amazing properties which is worth to properly exploit in view of the great demand of plant-derived food proteins. To discuss this matter and get the most updated advances in a disciplinary integrated way, the experts in the field met at the XIV edition of the International Lupin Conference last June in Milan, Italy. This is the report of the scientific outcomes of the conference and the satellite events, including two dedicated workshops and a unique Lupin banquet based on lupin foods. The results of a hedonic test performed on the lupin foods provided during the banquet are reported as well
Caco-2 cell monolayers to study the transport of lupin conglutin gamma across human intestinal ep... more Caco-2 cell monolayers to study the transport of lupin conglutin gamma across human intestinal epithelium / A. SCARAFONI, J. CAPRARO, C. MAGNI, A. CLEMENTE, M. DURANTI. - In: Proceedings of the 53rd National Meeting of the Italian Society of Biochemistry and Molecular Biology ( ...
gamma-Conglutin, a lupin seed glycoprotein, is capable of positively influencing glucose uptake b... more gamma-Conglutin, a lupin seed glycoprotein, is capable of positively influencing glucose uptake by various model cells and reducing glycaemia in animal models and human subjects. In particular, gamma-conglutin can activate in differentiating myocyte different signaling pathways that closely resembled those of insulin. A recent study aimed at identifying the metabolic fate of the protein, showed that it was transcytosed through a CaCo2 cell monolayer and crossed an ex-vivo intestinal barrier in an intact form. Beside, the protein displays strong resistance to endogenous and exogenous proteolytic enzymes. These findings encouraged us to further study the molecular mechanisms of action of this bioactive protein, which are still not completely elucidated. Aim of the present work was to study how gamma-conglutin interact with the target cells and show if the protein undergoes any covalent change during the process. 2D-electrophoresis of the gamma-conglutin treated cell lysates and antibody revelation of the blotted maps showed the presence of the intact protein inside the cells. Moreover, spots with an unexpectedly low pI, not present in the natural protein, were detected. These spots were submitted to MS/MS spectrometric analysis. Phosphorylated amino acids were evidenced in some gamma-conglutin peptides. To unveil gamma-conglutin cellular fate, HepG2 cells treated with the lupin protein were submitted to confocal microscopy and transmission electron microscopy (TEM). gamma-Conglutin was found accumulated inside the cells and spread as protein aggregates into the cytoplasm. Microvilli were involved in trapping the protein at the beginning of the process. A prerequisite for any biological activity is the interaction with or entry into the target cells. These findings, by showing that gamma-conglutin is uptaked by HepG2 cells in an intact form and is modified by multiple phosphorylation, open the way to new studies for the understanding of the described gamma-conglutin insulin-mimetic activity
Two-dimensional (2-D) IEF/SDS-PAGE is a powerful tool to get molecular “pictures” of food proteom... more Two-dimensional (2-D) IEF/SDS-PAGE is a powerful tool to get molecular “pictures” of food proteomes and monitor the processing effect(s) of a given food item on its protein profile. Still the use of 2-D approaches to this aim is rather uncommon.In this work, 2-D electrophoresis has been used to monitor the main steps of lupin-based gluten-free pasta production. Three different production
Citation: Improved lupin proteins isolation and molecular characterisation in view of their explo... more Citation: Improved lupin proteins isolation and molecular characterisation in view of their exploitation as food ingredients/M. Duranti, C. Magni, E. Sironi, F. Sessa, A. Scarafoni-In: Proceedings of the Final Conference of the European Project: Healthy ProFood/[a cura di] ...
2D electrophoretic mapping as a tool for the analysis of seed proteins in legume-based raw materi... more 2D electrophoretic mapping as a tool for the analysis of seed proteins in legume-based raw materials and novel foods / E. Sironi, A. Scarafoni, F. Sessa, C. Magni, V. Dani, M. Duranti - In: 5th European Conference on Grain Legumes and the 2nd International Conference on Legume ...
Adzuki seed β-vignin, a vicilin-like globulin, has proven to exert various health-promoting biolo... more Adzuki seed β-vignin, a vicilin-like globulin, has proven to exert various health-promoting biological activities, notably in cardiovascular health. A simple scalable enrichment procedure of this protein for further nutritional and functional studies is crucial. In this study, a simplified chromatography-independent protein fractionation procedure has been optimized and described. The electrophoretic analysis showed a high degree of homogeneity of β-vignin isolate. Furthermore, the molecular features of the purified protein were investigated. The adzuki bean β-vignin was found to have a native size of 146 kDa, and the molecular weight determined was consistent with a trimeric structure. These were identified in two main polypeptide chains (masses of 56–54 kDa) that are glycosylated polypeptides with metal binding capacity, and one minor polypeptide chain with a mass 37 kDa, wherein these features are absent. The in vitro analysis showed a high degree of digestibility of the protein ...
Biological activity, traceability and potential allergenicity of selected lupin proteins / M. Dur... more Biological activity, traceability and potential allergenicity of selected lupin proteins / M. Duranti, E. Sironi, C. Magni, F. Sessa, A. Scarafoni - In: Proceedings of the Final Conference of the European Project : Healthy ProFood / [a cura di] A. Arnoldi. - Roma : Aracne, 2005 ...
Biological activity, traceability and potential allergenicity of selected lupin proteins / M. Dur... more Biological activity, traceability and potential allergenicity of selected lupin proteins / M. Duranti, E. Sironi, C. Magni, F. Sessa, A. Scarafoni - In: Proceedings of the Final Conference of the European Project : Healthy ProFood / [a cura di] A. Arnoldi. - Roma : Aracne, 2005 ...
Food proteins and peptides are able to exert a variety of well-known bioactivities, some of which... more Food proteins and peptides are able to exert a variety of well-known bioactivities, some of which are related to well-being and disease prevention in humans and animals. Currently, an active trend in research focuses on chronic inflammation and oxidative stress, delineating their major pathogenetic role in age-related diseases and in some forms of cancer. The present study aims to investigate the potential effects of pseudocereal proteins and their derived peptides on chronic inflammation and oxidative stress. After purification and attribution to protein classes according to classic Osborne’s classification, the immune-modulating, antioxidant, and trypsin inhibitor activities of proteins from quinoa (Chenopodium quinoa Willd.), amaranth (Amaranthus retroflexus L.), and buckwheat (Fagopyrum esculentum Moench) seeds have been assessed in vitro. The peptides generated by simulated gastro-intestinal digestion of each fraction have been also investigated for the selected bioactivities. ...
γ-conglutin (γC) is a major protein of Lupinus albus seeds, but its function is still unknown. It... more γ-conglutin (γC) is a major protein of Lupinus albus seeds, but its function is still unknown. It shares high structural similarity with xyloglucan-specific endo-glucanase inhibitor proteins (XEGIPs) and, to a lesser extent, with Triticum aestivum endoxylanase inhibitors (TAXI-I), active against fungal glycoside hydrolases GH12 and GH11, respectively. However, γC lacks both these inhibitory activities. Since β-galactomannans are major components of the cell walls of endosperm in several legume plants, we tested the inhibitory activity of γC against a GH2 β-mannosidase (EC 3.2.1.25). γC was actually able to inhibit the enzyme, and this effect was enhanced by the presence of zinc ions. The stoichiometry of the γC/enzyme interaction was 1:1, and the calculated Ki was 1.55 μM. To obtain further insights into the interaction between γC and β-mannosidase, an in silico structural bioinformatic approach was followed, including some docking analyses. By and large, this work describes experim...
International journal of food sciences and nutrition, Jan 18, 2017
Fermentation represents a valuable and cost-effective approach for food stabilisation and nutriti... more Fermentation represents a valuable and cost-effective approach for food stabilisation and nutritional improvement. Tempeh is an example of soybean solid-state fermentation. In this work, we investigated the possibility of producing a tempeh analogue containing high amounts of vitamin B12 using seeds of three different species of the legume lupin, namely Lupinus albus, L. angustifolius and L. mutabilis, with Rhizopus oligosporus and Propionibacterium freudenreichii cofermentation. Synergic effects of Rhizopus and Propionibacterium in increasing vitamin B12 up to 1230 ng/g dw was observed. These findings indicate that this cofermentation can improve lupin nutritional quality and safety to provide a tempeh analogue with added value for vegan and vegetarian communities and low-income populations. The level of potentially toxic lupin alkaloids was also monitored during the tempeh preparation.
Biochemical and Biophysical Research Communications, 2013
Lupin seed γ-Conglutin is a protein capable of reducing glycaemia in mammalians and increasing gl... more Lupin seed γ-Conglutin is a protein capable of reducing glycaemia in mammalians and increasing glucose uptake by model cells. This work investigated whether γ-Conglutin is internalised into the target cells and undergoes any covalent change during the process, as a first step to understanding its mechanism of action. To this purpose, γ-Conglutin-treated and untreated HepG2 cells were submitted to confocal and transmission electron microscopy. Immune-revelation of γ-Conglutin at various intervals revealed its accumulation inside the cytosol. In parallel, 2D-electrophoresis of the cell lysates and antibody reaction of the blotted maps showed the presence of the protein intact subunits inside the treated cells, whilest no trace of the protein was found in the control cells. However, γ-Conglutin-related spots with an unexpectedly low pI were also observed in the maps. These spots were excised, trypsin-treated and submitted to MS/MS spectrometric analysis. The presence of phosphorylated amino acids was detected. These findings, by showing that γ-Conglutin is internalised by HepG2 cells in an intact form and is modified by multiple phosphorylation, open the way to the understanding of the lupin γ-Conglutin insulin-mimetic activity.
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