A sucrose-binding protein and ß-conglycinins regulate soybean seed protein content and control multiple seed traits.

Expanded agriculture production is required to support the world's population but can impose substantial environmental and climate change costs, particularly with intensifying animal production and protein demand. Shifting from an animal- to a plant-based protein diet has numerous health benefits. Soybean (Glycine max (L.) Merr.) is a major source of protein for human food and animal feed; improved soybean protein content and amino acid composition could provide high-quality soymeal for animal feed, healthier human foods, and a reduced carbon footprint. Nonetheless, during the soybean genome evolution, a balance was established between the amount of seed protein, oil, and carbohydrate content, burdening the development of soybean cultivars with high proteins. We isolated two high-seed protein (HP) soybean mutants, HP1 and HP2, with improved seed amino acid composition and stachyose content, pointing to their involvement in controlling seed rebalancing phenomenon. HP1 encodes ß-conglycinin (GmCG-1) and HP2 encodes Sucrose Binding Protein (GmSBP-1), which are both highly expressed in soybean seeds. Mutations in GmSBP-1, GmCG-1, and the paralog GmCG-2 resulted in increased protein levels, confirming their role as general regulators of seed protein content, amino acid seed composition, and seed vigor. Biodiversity analysis of GmCG and GmSBP across 108 soybean accessions revealed haplotypes correlated with protein and seed carbohydrate content. Furthermore, our data revealed an unprecedented role of GmCG and GmSBP proteins in improving seed vigor, crude protein, and amino acid digestibility. Since GmSBP and GmCG are present in most seed plants analyzed, these genes could be targeted to improve multiple seed traits.

Commensal Bacillus siamensis LF4 ameliorates ß-conglycinin induced inflammation in intestinal epithelial cells of Lateolabrax maculatus.

ß-conglycinin and glycinin, two major heat-stable anti-nutritional factors in soybean meal (SM), have been suggested as the key inducers of intestinal inflammation in aquatic animals. In the present study, a spotted seabass intestinal epithelial cells (IECs) were used to compare the inflammation-inducing effects of ß-conglycinin and glycinin. The results showed that IECs co-cultured with 1.0 mg/mL ß-conglycinin for 12 h or 1.5 mg/mL glycinin for 24 h significantly decreased the cell viability (P < 0.05), and overstimulated inflammation and apoptosis response by significantly down-regulating anti-inflammatory genes (IL-2, IL-4, IL-10 and TGF-ß1) expressions and significantly up-regulated pro-inflammatory genes (IL-1ß, IL-8 and TNF-α) and apoptosis genes (caspase 3, caspase 8 and caspase 9) expressions (P < 0.05). Subsequently, a ß-conglycinin based inflammation IECs model was established and used for demonstrating whether commensal probiotic B. siamensis LF4 can ameliorate the adverse effects of ß-conglycinin. The results showed ß-conglycinin-induced cell viability damage was completely repaired by treated with 109 cells/mL heat-killed B. siamensis LF4 for ≥12 h. At the same time, IECs co-cultured with 109 cells/mL heat-killed B. siamensis LF4 for 24 h significantly ameliorated ß-conglycinin-induced inflammation and apoptosis by up-regulating anti-inflammatory genes (IL-2, IL-4, IL-10 and TGF-ß1) expressions and down-regulated pro-inflammatory genes (IL-1ß, IL-8 and TNF-α) and apoptosis genes (caspase 3, caspase 8 and caspase 9) expressions (P < 0.05). In summary, both ß-conglycinin and glycinin can lead to inflammation and apoptosis in spotted seabass IECs, and ß-conglycinin is more effective; commensal B. siamensis LF4 can efficiently ameliorate ß-conglycinin induced inflammation and apoptosis in IECs.

Probiotic components of Bacillus siamensis LF4 mitigated ß-conglycinin caused cell injury via modulating TLR2/MAPKs/NF-κB signaling in Lateolabrax maculatus.

ß-conglycinin is a recognized factor in leading to intestinal inflammation and limiting application of soybean meal in aquaculture. Our previous study reported that heat-killed B. siamensis LF4 could effectively mitigate inflammatory response and apoptosis caused by ß-conglycinin in spotted seabass (Lateolabrax maculatus) enterocytes, but the mechanisms involved are not fully understood. In the present study, therefore, whole cell wall (CW), peptidoglycan (PG) and lipoteichoic acid (LTA) and cell-free supernatant (CFS) have been collected from B. siamensis LF4 and their mitigative function on ß-conglycinin-induced adverse impacts and mechanisms underlying were evaluated. The results showed that ß-conglycinin-induced cell injury, characterized with significantly decreased cell viability and increased activities of lactate dehydrogenase, glutamic oxaloacetic transaminase, glutamic propylic transaminase (P < 0.05), were reversed by subsequent heat-killed B. siamensis LF4 and its CW, LTA, PG and CFS treatment. Enterocytes co-cultured with heat-killed B. siamensis LF4 and its CW, LTA, PG and CFS (especially PG) significantly increased expressions of anti-inflammatory genes (IL-2, IL-4, IL-10 and TGF-ß1), tight junction proteins (ZO-1, occludin and claudin-b) and antimicrobial peptides (ß-defensin, hepcidin-1, NK-lysin and piscidin-5), and decreased expressions of pro-inflammatory genes (IL-1ß, IL-8 and TNF-α) and apoptosis-related genes (caspase 3, caspase 8 and caspase 9) (P < 0.05), indicating their excellent mitigation effects on ß-conglycinin-induced cell damages. In addition, heat-killed B. siamensis LF4 and its CW, LTA, PG and CFS significantly increased TLR2 mRNA level (especially in PG treatment), and decreased MAPKs (JNK, ERK, p38 and AP-1) and NF-κB related genes expressions. In conclusion, heat-killed B. siamensis LF4 and its CW, LTA, PG and CFS could modulating TLR2/MAPKs/NF-κB signaling and alleviating ß-conglycinin-induced enterocytes injury in spotted seabass (L. maculatus), and PG presented the best potential.

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

PURPOSE: Soybean glycinin (11S) and ß-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. METHODS: 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 1 week. 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. RESULTS: 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-1ß, and IL-18 were observed in the duodenum, jejunum, and ileum. CONCLUSION: 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.

Peptides Derived from Soybean ß-Conglycinin Induce the Migration of Human Peripheral Polymorphonuclear Leukocytes.

Food-derived peptides have various biological activities. When food proteins are ingested orally, they are digested into peptides by endogenous digestive enzymes and absorbed by the immune cell-rich intestinal tract. However, little is known about the effects of food-derived peptides on the motility of human immune cells. In this study, we aimed to understand the effects of peptides derived from a soybean protein ß-conglycinin on the motility of human peripheral polymorphonuclear leukocytes. We illustrated that MITL and MITLAIPVNKPGR, produced by digestion using in-vivo enzymes (trypsin and pancreatic elastase) of ß-conglycinin, induces the migration of dibutyryl cAMP (Bt2 cAMP)-differentiated human promyelocytic leukemia 60 (HL-60) cells and human polymorphonuclear leukocytes in a dose- and time-dependent manner. This migration was more pronounced in Bt2 cAMP-differentiated HL-60 cells; mRNA expression of formyl peptide receptor (FPR) 1 increased significantly than in all-trans-retinoic acid (ATRA)-differentiated HL-60 cells. This migration was inhibited by tert-butoxycarbonyl (Boc)-MLP, an inhibitor of FPR, and by pretreatment with pertussis toxin (PTX). However, the effect was weak when treated with WRW4, a selective inhibitor of the FPR2. We then demonstrated that MITLAIPVNKPGR induced intracellular calcium responses in human polymorphonuclear leukocytes and Bt2 cAMP-HL60 cells. Furthermore, pre-treatment by fMLP desensitized the calcium response of MITLAIPVNKPGR in these cells. From the above, MITLAIPVNKPGR and MITL derived from soybean ß-conglycinin induced polymorphonuclear leukocyte migration via the FPR1-dependent mechanism. We found chemotactic peptides to human polymorphonuclear leukocytes, which are the endogenous enzyme digests of soybean protein.

Soybean Antigen Protein-Induced Intestinal Barrier Damage by Trigging Endoplasmic Reticulum Stress and Disordering Gut Microbiota in Weaned Piglets.

Endoplasmic reticulum (ER) stress is a crucial factor in the pathogenesis of intestinal diseases. Soybean antigenic proteins (ß-conglycinin and soy glycinin) induce hypersensitivity reactions and intestinal barrier damage. However, whether this damage is associated with ER stress, autophagy, and the gut microbiome is largely unclear. Therefore, in this study, we aimed to investigate the effect of dietary supplementation with soy glycinin (11S glycinin) and ß-conglycinin (7S glycinin) on intestinal ER stress, autophagy, and flora in weaned piglets. Thirty healthy 21-day-old weaned "Duroc × Long White × Yorkshire" piglets were randomly divided into three groups and fed a basic, 7S-supplemented, or 11S-supplemented diet for one week. The results indicated that 7S/11S glycinin disrupted growth performance, damaged intestinal barrier integrity, and impaired goblet cell function in piglets (p < 0.05). Moreover, 7S/11S glycinin induced ER stress and blocked autophagic flux in the jejunum (p < 0.05) and increased the relative abundance of pathogenic flora (p < 0.01) and decreased that of beneficial flora (p < 0.05). In conclusion, 7S/11S glycinin induces intestinal ER stress, autophagic flux blockage, microbiota imbalance, and intestinal barrier damage in piglets.

Controlling the assembly of soy ß-conglycinin to fabricate heat-stable particles for high protein liquid systems.

BACKGROUND: Recently, there is a growing interest in developing protein-fortified liquid systems, which are formulated to provide special nutrient combinations to those with special dietary needs. The fabrication of heat-stable protein for protein-fortified liquid systems relies heavily on precise control of the edible protein-building process. RESULTS: Results suggested that heat-stable 7S protein particles (7SPPs) could be obtained by preheating at 100 °C for an extended time, whereas 7S proteins with better gelling properties were discovered after preheating at lower temperatures. According to the findings of the protein conformational and morphological characterization, the 7SPPs showed rather stable tertiary and secondary structures as well as size distributions, which might be responsible for their heat stability. Additionally, during the reheating test, suspensions of 7SPPs showed no signs of gelation and had a low viscosity even though the protein content was as high as 120 mg mL-1 . However, 7S proteins with improved gelling properties were found to show rising aggregate size, higher susceptibility and larger conformational structure changing rates upon reheating treatment. CONCLUSION: Soy ß-conglycinin (7S) proteins with tunable heat stability were successfully prepared by preheating 10 mg mL-1 protein dispersions at various temperatures (80-120 °C) and durations (15-120 min). These findings provide fundamental insights for developing 7S-based protein-fortified systems. © 2023 Society of Chemical Industry.

Confocal Fluorescence Microscopy Investigation for the Existence of Subdomains within Protein Storage Vacuoles in Soybean Cotyledons.

In legumes, the seed storage proteins accumulate within specialized organelles called protein storage vacuoles (PSVs). In several plant species, PSVs are differentiated into subdomains that accumulate different kinds of proteins. Even though the existence of subdomains is common in cereals and legumes, it has not been reported in soybean PSVs. The two most abundant seed proteins of soybean, 7S and 11S globulins, have different temporal accumulation patterns and exhibit considerable solubility differences that could result in differential accretion of these proteins within the PSVs. Here, we employed confocal fluorescent microscopy to examine the presence or absence of subdomains within the soybean PSVs. Eosin-stained sections of FAA-fixed paraffin embedded soybean seeds, when viewed by confocal fluorescence microscopy, revealed the presence of intricate subdomains within the PSVs. However, fluorescence immunolabeling studies demonstrated that the 7S and 11S globulins were evenly distributed within the PSVs and failed to corroborate the existence of subdomains within the PSVs. Similarly, confocal scanning microscopy examination of free-hand, vibratome and cryostat sections also failed to demonstrate the existence of subdomains within PSVs. The subdomains, which were prominently seen in PSVs of FAA-fixed soybean seeds, were not observed when the seeds were fixed either in glutaraldehyde/paraformaldehyde or glutaraldehyde. Our studies demonstrate that the apparent subdomains observed in FAA-fixed seeds may be a fixation artifact.

High hydrostatic pressure treatment reduces the potential antigenicity of ß-conglycinin by changing the protein structure during in vitro digestion.

BACKGROUND: High hydrostatic pressure (HHP) treatment has been used to alleviate the allergenicity of soybeans, but there are little data about the potential antigenicity of ß-conglycinin after HHP treatment. RESULTS: We examined the effects of HHP treatment on the antigenicity and structure of ß-conglycinin. When the pressure was 300 and 400 MPa, HHP treatment reduced the immunoglobulin (Ig)G binding capacity of ß-conglycinin, while its IgE binding capacity did not change significantly. After in vitro digestion, both the IgE and IgG binding of ß-conglycinin was obviously inhibited after HHP treatment at 400 MPa and 60 °C, although its binding capacity with linear epitope antibodies increased. Moreover, HHP treatment changed the secondary structure of ß-conglycinin, the content of α-helix and random coils increased, while the ß-sheet and ß-turn decreased. After HHP treatment, the conformational structure was unfolded so that a large number of hydrophobic regions were exposed. CONCLUSION: HHP treatment alleviated the potential antigenicity of ß-conglycinin by modifying its structure, which facilitated in vitro digestion and destroyed epitopes. This research provides a new insight into the mechanism of HHP treatment that affects the sensitization of soy protein allergens. © 2022 Society of Chemical Industry.

The role of the extension region on the structural and physicochemical characteristics of the α-subunit of ß-conglycinin: implications of pH value and ionic strength.

BACKGROUND: To clarify the role of the extension region on the structure-functional relationship of the α-subunit of ß-conglycinin, α-subunit and its segment of the core region (αc-subunit) were expressed via an Escherichia coli system. Their physicochemical properties were compared under acid, neutral or alkaline conditions (pH 4.0, 7.0, and 8.0) and high or low ionic strength (µ = 0.05 and 0.5), respectively. RESULTS: The results showed that the extension region contributed to increasing thermal stability, especially at low ionic strength under acidic and neutral conditions. The extension region stabilized the α-subunit with high solubility, low turbidity, and small particle size under neutral and alkaline conditions, whereas these impacts were suppressed at a high ionic strength and acidic conditions. Surface hydrophobicity of the α-subunit decreased under acidic and alkaline conditions without being interfered with by ionic strength. CONCLUSION: It can be concluded that the extension region played different roles under different pH and ionic strength conditions. These factors should be specified carefully and speculated individually to explore the more detailed and profound nature of ß-conglycinin at the submolecular level. The results could benefit a better understanding of the relationship between domain structure and functions of soybean protein. © 2022 Society of Chemical Industry.

Antigenic activity and epitope analysis of ß-conglycinin hydrolyzed by pepsin.

BACKGROUND: Soybean is among the 'big eight' allergenic foods, and ß-conglycinin, the main antigenic protein of soybean, has high levels of antigenic activity. Why the antigenic activity of soybean ß-conglycinin is not eliminated by enzymatic hydrolysis is not clear. In this study, changes in the molecular composition and antigenicity of ß-conglycinin hydrolyzed by pepsin were analyzed and it was determined whether complete sequential epitopes exist in the resulting hydrolysates. The nature and antigenic activity of protein subunits obtained after ß-conglycinin hydrolysis were also assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and competitive enzyme-linked immunosorbent assay, respectively. RESULTS: The residual antigenic activity of ß-conglycinin was 52%, α'- and α-subunits completely disappeared, the 49 kDa fraction partially disappeared, and peptides measuring 27 and 23 kDa were newly formed after 60 min of enzymatic hydrolysis. Prolonged enzymatic hydrolysis did not result in remarkable changes in these peptides; thus, the peptides show some resistance to enzymatic hydrolysis. The amino acid sequences of the peptide chains were analyzed by matrix-assisted laser desorption / ionization-time of flight mass spectrometry and aligned with the related sequences in the corresponding protein and antigen databases. Ten complete sequential epitopes were identified in the residual 49 kDa fraction, of these epitopes, two were from α-subunits and eight were from ß-subunits. CONCLUSION: The presence of complete sequential epitopes in hydrolysates obtained from the enzymatic hydrolysis of soybean is an important reason for the incomplete disappearance of the antigenic activity of ß-conglycinin. © 2020 Society of Chemical Industry.

[Effect of Cooking and Processing on Quantitation of Soybean Proteins].

We examined the effects of cooking and processing on the quantitation of soy protein in various soy-based foods. For the phosphate-buffered saline (PBS) extraction, the total protein content was measured by bicinchoninic acid assay, and the buffer extraction containing sodium dodecyl sulfate (SDS) and 2-mercaptoethanol (ME) was measured by the 2-D Quant Kit, and SDS-polyacrylamide gel electrophoresis analysis (SDS-PAGE) of each extraction was performed. Furthermore, measurements were performed by various ELISAs. During the tofu cooking process, the protein concentrations of soaked soybeans and Seigo (soybean homogenized with water) fluctuated- the change in protein solubility due to the amount of water during sample homogenization was considered to be a factor. It was thought that the decrease in protein concentration due to heating of Seigo during soymilk production was considered to indicate thermal denaturation of the protein, and that SDS and ME extraction in tofu may affect the measurement system. In cooking excluding roasted beans, proteins with a mass of 50 kDa or above and around 20 kDa were denatured, and in twice-fried tofu, proteins around 40 kDa were denatured, but the protein concentration excluding boiled soybeans did not decreased. Furthermore, the protein concentration from roasted beans, yuba, roasted okara and fried tofu increased with the cooking time, suggesting that the denaturation temperature of the protein shifted to a high temperature as the water content decreased. Both of the two types of ELISAs that comply with the official labeling system of foods containing allergenic substances were useful for detecting soybean protein by detecting proteins and peptides in processed soybean products, fermented foods excluding natto, and health foods.

The dynamic process of dietary soybean ß-conglycinin in digestion, absorption, and metabolism among different intestinal segments in grass carp (Ctenopharyngodon idella).

The present study aimed to investigate the dynamic process of soybean ß-conglycinin in digestion, absorption, and metabolism in the intestine of grass carp (Ctenopharyngodon idella). Fish fed with 80 g ß-conglycinin/kg diet for 7 weeks, the intestinal digestive enzyme was extracted to hydrolyze ß-conglycinin in vitro, the free amino acid and its metabolism product contents in intestinal segments were analyzed. The present study first found that ß-conglycinin cannot be thoroughly digested by fish intestine digestive enzyme and produces new products (about 60- and 55-kDa polypeptides). The indigestible ß-conglycinin further caused the free amino acid imbalance, especially caused free essential amino acid deficiency in the proximal intestine but excess in the distal intestine. Moreover, these results might be partly associated with the effect of ß-conglycinin in amino acid transporters and tight junction-regulated paracellular pathway. Finally, dietary ß-conglycinin increased the content of amino acid catabolism by-product ammonia while decreased the amino acid anabolism product carnosine content in the proximal intestine and distal intestine. Thus, the current study first and systemically explored the dynamic process of ß-conglycinin in digestion, absorption, and metabolism, which further supported our previous study that dietary ß-conglycinin suppressed fish growth and caused intestine injure.

Effects of glycinin and ß-conglycinin on growth performance and intestinal health in juvenile Chinese mitten crabs (Eriocheir sinensis).

This study investigates the effects of two soybean antigens (glycinin and ß-conglycinin) as an antinutritional substance in the diet on the growth, digestive ability, intestinal health and microbiota of juvenile Chinese mitten crabs (Eriocheir sinensis). The isonitrogenous and isolipidic diets contained two soybean antigens at two levels each (70 and 140 g/kg ß-conglycinin, 80 and 160 g/kg glycinin) and a control diet without ß-conglycinin or glycinin supplementation, and were used respectively to feed juvenile E. sinensis for seven weeks. Dietary inclusion of either glycinin or ß-conglycinin significantly reduced crab survival and weight gain. The crabs fed diets containing soybean antigens had higher malondialdehyde concentrations and lower catalase activities in the intestine than those in the control. The activities of trypsin and amylase in the intestine were suppressed by dietary ß-conglycinin and glycinin. Dietary glycinin or ß-conglycinin impaired the immunity and morphological structure of intestine, especially the peritrophic membrane. The mRNA expression of constitutive and inducible immune responsive genes (lipopolysaccharide-induced TNF-α factor and interleukin-2 enhancer-binding factor 2) increased while the mRNA expression of the main genes related to the structural integrity peritrophic membrane (peritrophin-like gene and peritrophic 2) significantly decreased in the groups with soybean antigen addition. Soybean antigen could also change the intestinal microbial community. The abundance of pathogenic bacteria (Ochrobactrum, Burkholderia and Pseudomonas) increased significantly in both soybean antigen groups. Although pathogenic bacteria Vibrio were up-regulated in the glycinin group, the abundance of Dysgonomonas that degraded lignocellulose and ameliorated the gut environment decreased in the glycinin group. This study indicates that existence of soybean antigens (glycinin or ß-conglycinin) could induce gut inflammation, reshape the community of gut microbiota, and cause digestive dysfunction, ultimately leading to impaired growth in crabs.

Soya protein ß-conglycinin ameliorates fatty liver and obesity in diet-induced obese mice through the down-regulation of PPARγ.

Diets high in fat can result in obesity and non-alcoholic fatty liver disease (NAFLD). The improvement of obesity and NAFLD is an important issue. ß-Conglycinin, one of the soya proteins, is known to prevent hyperlipidaemia, obesity and NAFLD. Therefore, we aimed to investigate the effects of ß-conglycinin on the improvement of obesity and NAFLD in high-fat (HF) diet-induced obese (DIO) mice and clarify the mechanism underlying these effects in liver and white adipose tissue (WAT). DIO male ddY mice were divided into six groups: HF, medium-fat (MF) and low-fat (LF) groups fed casein, and HF, MF and LF groups in all of which the casein was replaced by ß-conglycinin. A period of 5 weeks later, the ß-conglycinin-supplemented group resulted in lower body weight, relative weight of subcutaneous WAT, and hepatic TAG content (P=0·001). Furthermore, ß-conglycinin suppressed the hepatic expression of Pparγ2 in the HF dietary group, sterol regulatory element-binding protein-1c and the target genes. The expressions of inflammation-related genes were significantly low in the epididymal and subcutaneous WAT from the mice fed ß-conglycinin compared with those fed casein in the HF dietary group. Moreover, the expressions of Pparγ1 and Pparγ2 mRNA were suppressed in subcutaneous WAT in the HF dietary group but not in epididymal WAT. The concentrations of insulin and leptin were low in the serum of the mice fed ß-conglycinin. In conclusion, ß-conglycinin effectively improved obesity and NAFLD in DIO mice, and it appears to be a promising dietary protein for the amelioration of NAFLD and obesity.

The α' subunit of ß-conglycinin and various glycinin subunits of soy are not required to modulate hepatic lipid metabolism in rats.

PURPOSE: This study examined the effect of soy proteins with depletion of different subunits of the two major storage proteins, ß-conglycinin and glycinin, on hepatic lipids and proteins involved in lipid metabolism in rats, since the bioactive component of soy responsible for lipid-lowering is unclear. METHODS: Weanling Sprague Dawley rats were fed diets containing either 20% casein protein in the absence (casein) or presence (casein + ISF) of isoflavones or 20% alcohol-washed soy protein isolate (SPI) or 20% soy protein concentrates derived from a conventional (Haro) or 2 soybean lines lacking the α' subunit of ß-conglycinin and the A1-3 (1TF) or A1-5 (1a) subunits of glycinin. After 8 weeks, the rats were necropsied and liver proteins and lipids were extracted and analysed. RESULTS: The results showed that soy protein diets reduced lipid droplet accumulation and content in the liver compared to casein diets. The soy protein diets also decreased the level of hepatic mature SREBP-1 and FAS in males, with significant decreases in diets 1TF and 1a compared to the casein diets. The effect of the soy protein diets on female hepatic mature SREBP-1, FAS, and HMGCR was confounded since casein + ISF decreased these levels compared to casein alone perhaps muting the decrease by soy protein. A reduction in both phosphorylated and total STAT3 in female livers by ISF may account for the gender difference in mechanism in the regulation and protein expression of the lipid modulators. CONCLUSIONS: Overall, soy protein deficient in the α' subunit of ß-conglycinin and A1-5 subunits of glycinin maintain similar hypolipidemic function compared to the conventional soy protein. The exact bioactive component(s) warrant identification.

Effective Food Ingredients for Fatty Liver: Soy Protein ß-Conglycinin and Fish Oil.

Obesity is prevalent in modern society because of a lifestyle consisting of high dietary fat and sucrose consumption combined with little exercise. Among the consequences of obesity are the emerging epidemics of hepatic steatosis and nonalcoholic fatty liver disease (NAFLD). Sterol regulatory element-binding protein-1c (SREBP-1c) is a transcription factor that stimulates gene expression related to de novo lipogenesis in the liver. In response to a high-fat diet, the expression of peroxisome proliferator-activated receptor (PPAR) γ2, another nuclear receptor, is increased, which leads to the development of NAFLD. ß-Conglycinin, a soy protein, prevents NAFLD induced by diets high in sucrose/fructose or fat by decreasing the expression and function of these nuclear receptors. ß-Conglycinin also improves NAFLD via the same mechanism as for prevention. Fish oil contains n-3 polyunsaturated fatty acids such as eicosapentaenoic acid and docosahexaenoic acid. Fish oil is more effective at preventing NAFLD induced by sucrose/fructose because SREBP-1c activity is inhibited. However, the effect of fish oil on NAFLD induced by fat is controversial because fish oil further increases PPARγ2 expression, depending upon the experimental conditions. Alcohol intake also causes an alcoholic fatty liver, which is induced by increased SREBP-1c and PPARγ2 expression and decreased PPARα expression. ß-Conglycinin and fish oil are effective at preventing alcoholic fatty liver because ß-conglycinin decreases the function of SREBP-1c and PPARγ2, and fish oil decreases the function of SREBP-1c and increases that of PPARα.

Quantification of Gly m 5.0101 in Soybean and Soy Products by Liquid Chromatography-Tandem Mass Spectrometry.

Gly m 5.0101, the alpha subunit of ß-conglycinin, is one of the major allergens found in soybeans that has been identified as causing an allergic reaction. Here, we developed a quantification method of Gly m 5.0101 with multiple reaction monitoring using the synthetic peptide 194NPFLFGSNR202 as the external standard. Firstly, the ground soybean was defatted and extracted with a protein extraction buffer. Then the crude extract was on-filter digested by trypsin and analyzed by liquid chromatography-tandem mass spectrometry. The selected peptide exhibited a detection limit of 0.48 ng/mL and a linear relationship in a concentration range from 1.6 to 500 ng/mL (r² > 0.99). The developed method was successfully applied to quantify the Gly m 5.0101 level in dozens of soybean varieties from different sources and soybean products derived from different processing techniques. The developed method could be used to further analyze ß-conglycinin in soybean seeds combined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis.

High hydrostatic pressure (HHP) effects on antigenicity and structural properties of soybean ß-conglycinin.

In this study, the effect of high hydrostatic pressure (HHP) on antigenicity, free sulfhydryl group (SH) content, hydrophobicity (Ho), fluorescence intensity and circular dichroism data of soybean ß-conglycinin was studied. The antigenicity of soybean ß-conglycinin was decreased significantly at pressures 200-400 MPa. The antigenicity inhibition rate of ß-conglycinin declined from 92.72 to 55.15%, after being treated at 400 MPa for 15 min. Results indicated that free sulphydryl (SH) groups and surface Ho of ß-conglycinin were significantly increased at pressures 200-400 MPa and 5-15 min, whereas these properties decreased at the treatments above 400 MPa and 15 min. The maximum fluorescence intensity was noticed at 400 MPa and 15 min. The circular dichroism data analysis revealed that the amount of ß-turns and unordered structure significantly increased, while the content of α-helix1 and ß-strand1 noticeably decreased. These results provide evidence that HHP-induced the structural modification of ß-conglycinin and could alter the antigenicity of ß-conglycinin.

Dietary ß-conglycinin prevents acute ethanol-induced fatty liver in mice.

Alcoholic fatty liver is the earliest stage of alcohol-induced liver disease leading to liver cirrhosis. ß-Conglycinin, one of the soy proteins, is known to prevent non-alcoholic fatty liver, hyperlipidemia and obesity. Therefore, we examined whether ß-conglycinin feeding has an effect on the prevention of acute ethanol-induced fatty liver in mice. Male C57BL/6J mice were fed with 20 energy% ß-conglycinin or casein for 4 weeks prior to ethanol administration and were then given ethanol or glucose, as a control, by gavage. Ethanol significantly increased liver triglyceride (TG) in mice fed casein due to the activation of peroxisome proliferator-activated receptor (PPAR) γ2, a nuclear transcription factor known for regulating lipid metabolism and de novo lipogenesis. The liver TG of ethanol-administered ß-conglycinin-fed mice was significantly lower than that in those fed casein, although ethanol increased the amount of liver TG in mice fed ß-conglycinin. The increased levels of PPARγ2 protein and its target gene CD36 in response to an ethanol were not observed in mice fed ß-conglycinin. Moreover, ß-conglycinin decreased the basal expression of de novo lipogenesis-related genes such as stearoyl-CoA desaturase-1, and therefore, the expressions of these genes were lower in the ethanol-administered ß-conglycinin-fed mice than in the casein-fed mice. In conclusion, ß-conglycinin supplementation appears to prevent the development of fatty liver in mice caused by ethanol consumption via the suppression of alcohol-induced activation of PPARγ2 and the downregulation of the basal expression of de novo lipogenesis.