Soybean Glycinin Research Articles

Soybean Glycinin’s Antibacterial Properties Provide a Feasible Natural Alternative for Improving the Overall Quality and Shelf-Life of Beef Steaks and Combating Foodborne Pathogens

Soybean Glycinin’s Antibacterial Properties Provide a Feasible Natural Alternative for Improving the Overall Quality and Shelf-Life of Beef Steaks and Combating Foodborne Pathogens

Enhanced foaming and surface properties of soybean glycinin via hydrothermal treatment at acidic conditions

Plant-based proteins have evolved into a sustainable and health-conscious dietary option that consumers prioritize. However, plant proteins, especially globulins, typically exhibit poor functional properties due to their limited molecular flexibility and rigid conformation. In this study, we devised an enzyme-free approach to enhance the functional properties of soybean glycinin (11S). Under high-temperature (121 °C) and acidic conditions (pH 3.0), we modified soybean glycinin via hydrothermal treatment, primarily targeting selective breakdown at the aspartyl site. The obtained glycinin hydrolysates (11SH) formed micellar nanoparticles in solution and exhibited excellent foaming capacity, achieving up to a 600% overrun, surpassing that of egg white and albumins (typically 400–500%). And we successfully prepared gel-like foams using glycinin hydrolysates, demonstrating high viscoelasticity and uniform microstructure. Furthermore, the surface properties of glycinin hydrolysates could be further enhanced with the addition of NaCl. This study proposed an effective hydrothermal treatment to enhance the foaming capacity of soybean glycinin, and the resulting gel-like foams with high viscoelasticity and rigidity hold promising commercial application prospects.

Study of the binding interaction between modified glycinin and d-galactose and functions of the resulting complexes via multi-spectroscopy and molecular dynamics simulation

This study evaluated the effects of d-galactose (DG) concentration on the functional characteristics, physicochemical properties, structure and non-covalent binding ability of pH shifting treated soybean glycinin (S–11S). Multispectral and microscopic imaging analysis showed that the pH shifting induced soybean glycinin self-assembly into more stable small particle size aggregates, and the functional properties of S–11S were significantly enhanced. The introduction of DG further enhances the functional properties of S–11S, and the solubility and emulsification activity of the composite system are inversely proportional to the concentration of DG. In order to clarify the reason for the change, multispectral analysis found that the tertiary structure of S–11S was de-folded and the disordered structure was increased, which induced S–11S and DG to form smaller and stable nanoparticles than S–11S through hydrophobic forces and hydrogen bonding, and the optimal binding ratio was 10.8:1. In addition, the simulation results of molecular dynamics and molecular docking are in agreement with the experimental results. Therefore, the addition of DG can stabilize the overly discrete S–11S conformation and improve its functional characteristics. These results provide a theoretical basis for the application of modified protein and monosaccharide composite products in food and materials.

MicroRNA-223 Inhibits Soybean Glycinin- and β-Conglycinin-Induced Apoptosis of IPEC-J2 Cells by Targeting NLRP-3 in the IEL/IPEC-J2 Co-culture System.

The apoptosis of intestinal porcine epithelial cells induced by soybean antigen protein allergy is one of the most important mechanisms responsible for enteritis. MicroRNAs (miRNAs) affect the cellular and physiological functions of all multicellular organisms. We hypothesize that microRNA-223 inhibits soybean glycinin- and β-conglycinin-induced apoptosis of intestinal porcine enterocytes (IPEC-J2) by targeting the NLR family pyrin domain containing 3 (NLRP-3). Using the intestinal interepithelial lymphocyte (IEL)/IPEC-J2 co-culture system as an in vitro model, we investigate the role of microRNA-223 in the regulation of soybean glycinin- and β-conglycinin-induced apoptosis. In co-cultured IEL/IPEC-J2 cells incubated with glycinin or β-conglycinin, microRNA-223 decreased NLRP-3, ASC, caspase-1, caspase-3, FAS, BCL-2, and APAF-1 expressions in IPEC-J2 cells; decreased cytokine and cyclooxygenase-2 levels; significantly increased cell activity; and inhibited apoptosis. These data supported a novel antiallergic mechanism to mitigate the sensitization of soybean antigenic protein, which involves the upregulation of microRNA-223-targeting NLRP-3.

Molecular interaction of soybean protein and piperine by computational docking analyses

This study investigated the interaction of soybean glycinin (11S) and β-conglycinin (7S) proteins with piperine (PIP) and compared the differences between them. Molecular docking results showed that PIP bound to 11S and 7S mainly through electrostatic, hydrogen bonding, and hydrophobic interactions to form complexes with desirable microstructures, leading to a decrease in the surface hydrophobicity of 7S/11S protein. In particular, the decrease for 11S protein was more pronounced than that for 7S protein, and the affinity of PIP for 11S protein was higher than that for 7S protein. Molecular dynamics (MD) results showed that the binding of PIP to soybean protein was spontaneous. The addition of PIP could cause different changes in the secondary structure of 7S/11S protein, mainly a decrease in β-sheet and an increase in unordered coil, leading to the unfolding of soybean protein. This study provides a theoretical reference and research basis for elucidating the interaction mechanism between PIP and soybean protein in food systems, which is conducive to the potential application of soybean protein-PIP binding system in the food field.

Soybean glycinin and β-conglycinin damage the intestinal barrier by triggering oxidative stress and inflammatory response in weaned piglets.

Soybean glycinin (11S) and β-conglycinin (7S) are major antigenic proteins in soybean and can induce a variety of allergic reactions in the young animals. This study aimed to investigate the effect of 7S and 11S allergens on the intestine of piglets. Thirty healthy 21-day-old weaned "Duroc × Long White × Yorkshire" piglets were randomly divided into three groups fed with the basic diet, the 7S supplemented basic diet, or the 11S supplemented basic diet for 1week. Allergy markers, intestinal permeability, oxidative stress, and inflammatory reactions were detected, and we observed different sections of intestinal tissue. The expressions of genes and proteins related to NOD-like receptor thermal protein domain associated protein 3 (NLRP-3) signaling pathway were detected by IHC, RT-qPCR, and WB. Severe diarrhea and decreased growth rate were observed in the 7S and 11S groups. Typical allergy markers include IgE production and significant elevations of histamine and 5-hydroxytryptamine (5-HT). More aggressive intestinal inflammation and barrier dysfunction were observed in the experimental weaned piglets. In addition, 7S and 11S supplementation increased the levels of 8-hydroxy-2 deoxyguanosine (8-OHdG) and nitrotyrosine, triggering oxidative stress. Furthermore, higher expression levels of NLRP-3 inflammasome ASC, caspase-1, IL-1β, and IL-18 were observed in the duodenum, jejunum, and ileum. We confirmed that 7S and 11S damaged the intestinal barrier of weaned piglets and may be associated with the onset of oxidative stress and inflammatory response. However, the molecular mechanism underlying these reactions deserves further study.

Molecular interactions between soybean glycinin (11S) and genistein using spectroscopic and in silico analyses

Interactions and binding mechanisms between soybean glycinin (11S) and genistein were investigated at near neutral pH (7.4) and ambient temperature using various spectroscopic techniques and molecular docking. Ultra-violet visible spectroscopy revealed non-covalent bonds were responsible for the stabilisation of the complex. Following complexation, minor reductions in the alpha helix and beta sheet content of the protein were confirmed by circular dichroism (CD) and infrared spectroscopy (FTIR). Fluorescence quenching and molecular docking predicted a 11S trimer and genistein complex with a 1:1 stoichiometric ratio, with the binding location predominantly on the interior portion of the protein, stabilised in majority by hydrogen bonds. Although binding was determined to be of moderate strength, secondary structure characterisation suggested that the structural components of the protein remained largely unchanged. This outcome indicates that the ligand can be accommodated in the interior of the 11S trimer, forming a stable complex within the present conditions of neutral pH and ambient temperature.

Amino acids induce high seed-specific expression driven by a soybean (Glycine max) glycinin seed storage protein promoter.

We characterize GFP expression driven by a soybean glycinin promoter in transgenic soybean. We demonstrate specific amino acid-mediated induction of this promoter in developing soybean seeds in vitro. In plants, gene expression is primarily regulated by promoter regions which are located upstream of gene coding sequences. Promoters allow transcription in certain tissues and respond to environmental stimuli as well as other inductive phenomena. In soybean, seed storage proteins (SSPs) accumulate during seed development and account for most of the monetary and nutritional value of this crop. To better study the regulatory functions of a SSP promoter, we developed a cotyledon culture system where media and media addenda were evaluated for their effects on cotyledon development and promoter activity. Stably transformed soybean events containing a glycinin SSP promoter regulating the green fluorescent protein (GFP) were generated. Promoter activity, as visualized by GFP expression, was only observed in developing in planta seeds and in vitro-cultured isolated embryos and cotyledons from developing seeds when specific media addenda were included. Asparagine, proline, and especially glutamine induced glycinin promoter activity in cultured cotyledons from developing seeds. Other amino acids did not induce the glycinin promoter. Here, we report, for the first time, induction of a reintroduced glycinin SSP promoter by specific amino acids in cotyledon tissues during seed development.

Nanoencapsulation of antioxidant peptides from Lupinus mutabilis in chitosan nanoparticles obtained by ionic gelling and spray freeze drying intended for colonic delivery

Oxidative stress has long been associated with the pathogenesis of inflammatory bowel disease (IBD), and it has been suggested that the combined administration of antioxidants and anti-inflammatory agents may be helpful for its treatment. Biopeptides from plant proteins, such as soybean glycinin tripeptide VPY, effectively inhibit pro-inflammatory mediators in intestinal epithelial and immune cells, which are both involved in the pathogenesis of IBD. This study aimed to produce antioxidant biopeptides from proteins of Lupinus mutabilis seeds and then encapsulate them in chitosan nanoparticles (NPs) for colonic delivery. An antioxidant peptide fraction of less than three kDa (UF3) was obtained and added at different concentrations (0.1–0.4 mg/ml) in chitosan solutions. The NPs were prepared by gelation with tripolyphosphate (CTPP-UF3) or spray freeze-drying of the chitosan solution (SFDC-UF3). Sizes of 332 ± 13 and 465 ± 58 nm and maximum encapsulation efficiencies of 63.80 and 71.75%, respectively, were obtained. UF3 maintained its antioxidant capacity (>80%) and showed different release profiles in 1X PBS buffer at pH 7.4, depending on the encapsulation method. FT-IR showed hydrogen bonding and electrostatic interactions between the peptide and chitosan. Both nanosystems maintained cell viability greater than 70% in colonic cell lines HT-29. These results show that both methods are appropriate for the nanoencapsulation of UF3 and can be used to design nanoparticles for colonic delivery.

Quantum Dot Nanobead-Based Fluorescence-Linked Immunosorbent Assay for Detection of Glycinin in Soybeans and Soy Products.

Soybean glycinin, as a major soybean allergen, is difficult to accurately quantify due to its large molecular weight and complex structure. CdSe/ZnS quantum dot nanobead (QB) is a core/shell fluorescent nanomaterial with strong fluorescent signals and high sensitivity at 630 nm. An immunosorbent assay based on CdSe/ZnS quantum dot nanobeads (QBs-FLISA) was developed for the glycinin quantification in soybean and soybean products. Here, the purified glycinin was coated on the microporous plate to serve as the coating antigen, and CdSe/ZnS nanobead conjugated with anti-glycinin polyclonal antibodies was used as fluorescent detection probe. The target glycinin in the sample and the coated antigen on the plate competitively adsorbed the antibody labeled the CdSe/ZnS QBs probes. The limits of detection and quantitation for glycinin were 0.035 and 0.078 μg mL−1, respectively. The recoveries of the spiked samples ranged from 89.8% to 105.6%, with relative standard deviation less than 8.6%. However, compared with ELISA, the sensitivities of QBs-FLISA for the detection of glycinin were increased by 7 times, and the detection time was shortened by two-thirds. This QBs-FLISA method has been effectively applied to the detection of soybean seeds with different varieties and soy products with different processing techniques, which will provide a rapid screening method for soybean and soybean products with low allergens.

Effects of hydrodynamic cavitation at different pH values on the physicochemical properties and aggregation behavior of soybean glycinin

Soybean protein, especially glycinin, easily forms aggregates, limiting its application in some scenarios. In this research, the effects of different hydrodynamic cavitation (HC; 550 W for 0, 10 and 30 min) treatments on soybean glycinin aggregation at different pH values (2, 4, 6, 8 and 10) were investigated. At the five pH values, HC decreased the emulsifying activity index with increasing HC treatment time; however, the effects on the particle size, surface hydrophobicity (H0), solubility and emulsifying stability index (ESI) varied. Notably, HC had more significant effect on the physicochemical structure and functional properties at pH 6 than the other four pH because of different conformational structure, enhancing glycinin's H0 (from 850.95 ± 59.03 to 988.43 ± 64.68), solubility (from 21.85 ± 0.23 to 63.62 ± 1.18 g/100 g), and ESI (from 58.39 ± 7.46 to 127.74 ± 3.43 min). Moreover, HC influenced glycinin aggregation, as demonstrated by the intrinsic fluorescence spectra and circular dichroism results. In conclusion, HC changed the aggregation behavior and improved the functional properties of glycinin, indicating that HC is a promising technique to improve functional properties and provides a reference for the application of soybean protein in the food industry.

Gel-Forming of Self-Assembling Peptides Functionalized with Food Bioactive Motifs Modulate DPP-IV and ACE Inhibitory Activity in Human Intestinal Caco-2 Cells.

Food bioactive peptides are increasingly used for formulating food products, nutraceuticals, and functional food, since they are generally considered safe for human consumption and metabolic syndrome prevention. They are also becoming popular as sustainable sources of novel functional biomaterials such as hydrogels, edible nanonutraceuticals, delivery systems, and packing materials. However, such food peptides are mostly unstable, and degrade during food processing, or in a gastrointestinal environment, thus resulting in low bioavailability precluding their practical applications. Here, we decided to functionalize the well-known and characterized self-assembling peptide RADA16 with two synthetic analogues of food bioactive peptides deriving from the hydrolysis of soybean glycinin and lupin β-conglutin (namely IAVPTGVA and LTFPGSAED) for control of and improvement in their gel-forming nanostructures, biomechanics, and biological features. Extensive characterization was performed via Circular Dichroism (CD) spectroscopy, Fourier Transform Infrared spectroscopy (FT-IR), Thioflavin T (ThT) binding assay, rheological measurements, and Atomic Force Microscopy (AFM) analysis. Lastly, since self-assembling peptides (SAPs) can be co-assembled with diluent SAPs (without a bioactive epitope) as an approach to control the density of biological signals and therefore attain enhanced bioactivity, we investigated the effect of the co-assembly of RADA16 and functionalized food bioactive SAPs (dubbed cAP-Soy1 and cAP-Lup1) for the growth of Caco-2 human intestinal cells and contextually we characterized their biological activities as DPP-IV and ACE inhibitors, in order to demonstrate their potential use for the prevention of metabolic syndrome.

The effects of thermal treatments on the antigenicity and structural properties of soybean glycinin.

The effects of thermal treatments on the antigenicity and structural properties of soybean glycinin were investigated. An indirect enzyme-linked immunosorbent assay (iELISA) result indicated that the antigenicity of the soybean glycinin decreased to the lowest level when the heat treatment time was 50min and the temperature was set at 110°C. The reducing sodium dodecyl sulfate-Gel electrophoresis results showed that the heat treatment had promoted the formation of small molecular weight protein subunits and concentrated protein. The free sulfhydryl (-SH) group of glycinin had increased significantly (p<.05) under the conditions of temperature between 90 and 120°C and 40 to 50min heat treatments. The maximum fluorescence wavelength of the intrinsic fluorescence also changed significantly when compared with the control. The circular dichroism showed that the number of disordered structures had also increased significantly. These results provided evidence that the heat-induced structural modifications of glycinin will alter the antigenicity of soybean glycinin. PRACTICAL APPLICATIONS: The relationship between antigenicity and structural properties of heat-treated soybean globulin was studied by different protein structural research methods. The results showed that the antigenicity of the protein was minimized at 110°C 50min, which could be used in food safety production. This study chose to analyze the changes in the antigenicity and structural characteristics of soybean glycinin, both prior to and after heat treatments, for the purpose of promoting the development of low allergenic food products.

Enzymatic Preparation of Bioactive Peptides Exhibiting ACE Inhibitory Activity from Soybean and Velvet Bean: A Systematic Review.

The Angiotensin-I-converting enzyme (ACE) is a peptidase with a significant role in the regulation of blood pressure. Within this work, a systematic review on the enzymatic preparation of Angiotensin-I-Converting Enzyme inhibitory (ACEi) peptides is presented. The systematic review is conducted by following PRISMA guidelines. Soybeans and velvet beans are known to have high protein contents that make them suitable as sources of parent proteins for the production of ACEi peptides. Endopeptidase is commonly used in the preparation of soybean-based ACEi peptides, whereas for velvet bean, a combination of both endo- and exopeptidase is frequently used. Soybean glycinin is the preferred substrate for the preparation of ACEi peptides. It contains proline as one of its major amino acids, which exhibits a potent significance in inhibiting ACE. The best enzymatic treatments for producing ACEi peptides from soybean are as follows: proteolytic activity by Protease P (Amano-P from Aspergillus sp.), a temperature of 37 °C, a reaction time of 18 h, pH 8.2, and an E/S ratio of 2%. On the other hand, the best enzymatic conditions for producing peptide hydrolysates with high ACEi activity are through sequential hydrolytic activity by the combination of pepsin-pancreatic, an E/S ratio for each enzyme is 10%, the temperature and reaction time for each proteolysis are 37 °C and 0.74 h, respectively, pH for pepsin is 2.0, whereas for pancreatin it is 7.0. As an underutilized pulse, the studies on the enzymatic hydrolysis of velvet bean proteins in producing ACEi peptides are limited. Conclusively, the activity of soybean-based ACEi peptides is found to depend on their molecular sizes, the amino acid residues, and positions. Hydrophobic amino acids with nonpolar side chains, positively charged, branched, and cyclic or aromatic residues are generally preferred for ACEi peptides.

Oxidation induced by dielectric barrier discharge (DBD) plasma treatment reduces IgG/IgE binding capacity and improves the functionality of glycinin

The effect of dielectric barrier discharge (DBD) plasma treatment times from 2 to 5 min at 40 kV on IgG/IgE binding capacity and functionality of soybean glycinin was examined. A substantial reduction in the binding capacity (91.64% for IgG and 81.49% for IgE) was obtained after 5 min of plasma treatment, as determined by western-blot and ELISA analyses. Further studies demonstrated that the elimination of antigenicity and allergenicity of glycinin was directly related to plasma-induced structural changes on two aspects. A conformational alteration caused by oxidation of peptide bond amino groups, accompanied with an oxidation of Trp, Tyr, and Phe amino acid residues, which was confirmed by surface hydrophobicity, multi-spectroscopic analysis, and amino acid analysis. The cleavage of polypeptide chains inevitably partially diminished the linear epitopes, resulting in a primary decline in IgG/IgE binding capacity. Additionally, an increase in the solubility from 10.78 ± 0.35 to 65.96 ± 1.86% and significant increase in the emulsifying ability from 21.08 ± 2.64 to 160.29 ± 4.12 m2/g were observed after treatment of the plasma for 2 min. The present results confirm the potential use of DBD for the production of hypoallergenic soy protein-based products and improving their technical functions such as solubility and emulsifying ability.

The effect of trehalose on the thermodynamic stability and emulsification of soybean 11S globulin in the molten globule state

The molten globule (MG) state of soybean glycinin (11S) is partially folded at pH 1.0, so various polyhydroxyl osmotic agents are often used to maintain the structural stability of the MG state under acidic conditions. In this study trehalose was used as a penetrant, and intrinsic fluorescence spectroscopy and circular dichroism were performed to investigate the effects of a gradient concentration of trehalose on the structure of soybean 11S. The MG soybean 11S maintained a more α-helix structure (94.7%) and a larger hydrophobic area when treated with a high concentration of trehalose (1.3 mol/L) than when untreated, and retained a typical compact MG structure despite the lack of tertiary structural elements. A linear extrapolation model showed that the Gibbs free energy of the high-concentration trehalose-induced compact MG state of soybean 11S was reduced by 3 times, compared with the untreated globulin. This improvement of thermodynamic stability indicated that trehalose stabilized this structure by forming a stable trehalose–soybean 11S MG state system. With the addition of gradient trehalose, the surface tension of the solution was significantly reduced and the viscosity of the solution was increased. The emulsifying properties were optimized when the trehalose concentration was 0.5 mol/L, the emulsifying activity (EAI) and emulsifying stability (ESI) were 76.67 m2/g and 76.67%. The improvement of emulsifying property of 11S MG-trehalose system was further proved by observing the microstructure of emulsion by Cryo-SEM. This study advances the understanding of the 11S MG-trehalose system and provides a theoretical basis to produce proteins with excellent emulsifying properties.

Linkage and association study discovered loci and candidate genes for glycinin and β-conglycinin in soybean (Glycine max L. Merr.).

Linkage mapping and GWAS identified 67 QTLs related to soybean glycinin, β-conglycinin and relevant traits. Polymorphisms of the candidate gene Gy1 promoter were associated with the glycinin content in soybean. The major components of storage proteins in soybean seeds are glycinin and β-conglycinin, which play important roles in determining protein nutrition and soy food processing properties. Increasing the protein content while improving the ratio of glycinin to β-conglycinin is substantially important for soybean protein improvement. To investigate the genetic mechanism of storage protein subunits, 184 recombinant inbred lines (RILs) derived from a cross of Kefeng No. 1 and Nannong 1138-2 and 211 diverse soybean cultivars were used to detect loci related to glycinin (11S), β-conglycinin (7S), the sum of glycinin and β-conglycinin (SGC), and the ratio of glycinin to β-conglycinin (RGC). Sixty-seven QTLs and 11 hot genomic regions were identified as affecting the four traits. One genetic region (q10-1) on chromosome 10 was associated with multiple traits by both linkage and association analysis. Eight genes in 11 hot genomic regions might be related to soybean protein subunit. The candidate gene analysis showed that polymorphisms in Gy1 promoters were significantly correlated with the 11S content. The QTLs and candidate genes identified in the present study allow for further understanding the genetic basis of 11S and 7S regulation and provide useful information for marker-assisted selection (MAS) in soybean quality improvement.

Soybean glycinin disrupted intestinal structural integrity related to aggravation of apoptosis and downregulated transcription of tight junction proteins in the intestine of juvenile grass carp (Ctenopharyngodon idella)

Glycinin is a major anti-nutritional factor of soybean. Glycinin inhibits growth and impairs intestinal health in grass carp and Jian carp, but the underlying mechanisms are largely unknown. This study used grass carp to examine whether the negative influences of dietary glycinin are related to intestinal apoptosis signalling and tight junctions (TJs). The results showed that 8% dietary glycinin caused higher diamine oxidase activities and D-lactate content in the serum than did the control, indicating impairment of intestinal structural integrity. For the apoptosis indices, in the midgut and hindgut, glycinin aggravated the DNA fragmentation phenomenon, promoted caspase-3, −8, and − 9 activities, and increased cytochrome c protein abundance in the cytoplasm (apart from mitochondria) compared to those the control. In the midgut, glycinin increased the mRNA levels of pro-apoptotic molecules, including caspase-3, tumour necrosis factor-α (TNF-α), B-cell lymphoma 2 (Bcl-2)-associated X (Bax), and apoptotic protease activating factor 1 (Apaf-1), and decreased the mRNA levels of the anti-apoptotic molecule Bcl-2 compared to those in the control. In the hindgut, glycinin increased caspase-3, −8, and − 9, TNF-α, Fas-ligand (FasL), Bax, and Apaf-1 mRNA levels and decreased Bcl-2, myeloid cell leukaemia 1 (Mcl-1), and inhibitors of apoptosis proteins (IAP) mRNA levels compared to those in the control. For TJs, compared to that in the control, glycinin reduced the mRNA abundance of barrier-forming TJs—including claudin-c in the foregut; ZO-1, ZO-2b, occludin, and claudin-7a in the midgut; and ZO-1, ZO-2b, occludin, claudin-3c, claudin-7a, claudin-7b, and claudin-11 in the hindgut. In contrast, glycinin increased the mRNA abundance of pore-forming TJs, including claudin-15a and claudin-15b, in the midgut and hindgut. As an important nuclear factor of TJ transcription, hepatic nuclear factor 4 alpha (HNF-4α) protein abundance was reduced by glycinin in the midgut and hindgut. Glutamine alleviated apoptosis in the midgut. Glycinin aggravated apoptosis associated with both extrinsic and intrinsic signalling and disrupted TJ related to HNF-4α signalling in the midgut and hindgut, ultimately disrupting the intestinal structural integrity of grass carp.

Soybean glycinin impaired immune function and caused inflammation associated with PKC-ζ/NF-κb and mTORC1 signaling in the intestine of juvenile grass carp (Ctenopharyngodon idella)

Glycinin is a major protein and antinutritional factor of soybean. However, how dietary glycinin affect intestinal immune function of fish were largely unknown. In this study, we used juvenile grass carp as a model to investigate the impacts of glycinin on intestinal immune function of fish and involved mechanisms. We set three treatments including control, glycinin and glycinin + glutamine in this trial. For immune components, results revealed that compared with control group, glycinin group had lower acid phosphatase activities in the foregut, midgut and hindgut, lower C3 and C4 content, and lower mRNA abundances of IgM, IgZ, hepcidin, LEAP-2A, LEAP-2B and β-defensin-1 in the midgut and hindgut rather than foregut of grass carp. For pro-inflammatory cytokines and relevant signaling, glycinin elevated mRNA abundances of IL-1β, IL-8, IL-12p35, IL-12p40 and IL-17D in the midgut and IL-1β, IFN-γ2, IL-6, IL-8, IL-12p35, IL-12p40 and IL-17D in the hindgut, and increased protein abundances of PKC-ζ and nuclear NF-κB p65 in the midgut and hindgut in comparison to control. For anti-inflammatory cytokines and relevant signaling, glycinin reduced mRNA abundances of TGF-β1, TGF-β2, IL-4/13B (rather than IL-4/13A), IL-10 and IL-11 in the midgut and hindgut, and reduced p-mTOR (Ser 2448), p-S6K1 (Thr 389) and p-4EBP1 (Thr 37/46) protein abundances in the midgut and hindgut rather than foregut. Co-administration of glutamine with glycinin could partially enhance intestinal function and reduce intestinal inflammation compared with glycinin treatment. Concluded, glycinin decreased intestinal immune components and caused intestinal inflammation associated with PKC-ζ/NF-κB and mTORC1 signaling.

Molecular interaction of soybean glycinin and β-conglycinin with (−)-epigallocatechin gallate induced by pH changes

The effect of pH on interactions of soybean glycinin (11S) and β-conglycinin (7S) with (−)-epigallocatechin- 3-gallate (EGCG) were determined by turbidity. Results confirmed that the protein structure and interactions of 11S and 7S with EGCG depended largely on the pH. The binding affinity between 11S, 7S proteins and EGCG was strongest at pH 7.0 among the three pH levels and 7S showed a greater affinity to EGCG compared to 11S. The interaction mechanisms and structures of 11S-EGCG and 7S-EGCG complexes were further determined by multiple spectroscopic technique and molecular docking. Results showed that EGCG induced changes in the microenvironment of tryptophan, tyrosine, and C–H bending vibration at different pH levels. Binding to EGCG altered the secondary structures of the 11S and 7S proteins, inducing an increase in β-sheet at the cost of α-helix at pH7.0 and α-helix at pH5.0, respectively. The 11S and 7S presented more compact microstructure with the addition of EGCG. Electrostatic interaction, hydrogen bonding and hydrophobic interaction were all involved in the formation of complexes, where the hydrogen bond was the dominant one. These results might be helpful for the development of 11S-EGCG and 7S-EGCG as a bioactive food additive in the food industry.