Soybean beta-conglycinin peptone suppresses food intake and gastric emptying by increasing plasma cholecystokinin levels in rats.
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Cholecystokinin (CCK) is an important physiologic mediator that regulates satiety and gastric emptying. We demonstrated previously that soybean peptone acts directly on rat small intestinal mucosal cells to stimulate CCK release. In the present study, we examined the effects of beta-conglycinin, a major component of soy protein, and its peptone on food intake and gastric emptying after an intraduodenal infusion of beta-conglycinin peptone in relation to CCK release and interaction with the mucosal cell membrane. Intraduodenal infusion of beta-conglycinin peptone inhibited food intake in a dose-dependent manner, but that of whole soy peptone or camostat did not. The suppression of food intake by beta-conglycinin peptone was abolished by an intravenous injection of devazepide, a selective peripheral CCK receptor antagonist. The beta-conglycinin peptone infusion strongly suppressed gastric emptying with marked increases in portal CCK levels. We also observed that the beta-conglycinin peptone dose dependently and more potently stimulated CCK release from isolated dispersed mucosal cells of the rat jejunum than did beta-conglycinin itself. This stimulation corresponded to the binding activity of the peptide or protein to solubilized components of the rat jejunum membrane as evaluated by surface plasmon biosensor. These results indicate that beta-conglycinin peptone suppresses food intake, and this effect may be due to beta-conglycinin peptone in the lumen stimulating endogenous CCK release with direct acceptance to the intestinal cells. (+info)
Engineered recombinant peanut protein and heat-killed Listeria monocytogenes coadministration protects against peanut-induced anaphylaxis in a murine model.
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Peanut allergy (PNA) is the major cause of fatal and near-fatal anaphylactic reactions to foods. Traditional immunotherapy using peanut (PN) protein is not an option for PNA therapy because of the high incidence of adverse reactions. We investigated the effects of s.c. injections of engineered (modified) recombinant PN proteins and heat-killed Listeria monocytogenes (HKLM) as an adjuvant on anaphylactic reactions in a mouse model of PN allergy. PN-allergic C3H/HeJ mice were treated s.c. with a mixture of the three major PN allergens and HKLM (modified (m)Ara h 1-3 plus HKLM). The effects on anaphylactic reactions following PN challenge and the association with Ab levels and cytokine profiles were determined. Although all mice in the sham-treated groups exhibited anaphylactic symptoms with a median symptom score of 3, only 31% of mice in the mAra h 1-3 plus HKLM group developed mild anaphylaxis, with a low median symptom score of 0.5. Alterations in core body temperature, bronchial constriction, plasma histamine, and PN-specific IgE levels were all significantly reduced. This protective effect was markedly more potent than in the mAra h 1-3 protein alone-treated group. HKLM alone did not have any protective effect. Reduced IL-5 and IL-13, and increased IFN-gamma levels were observed only in splenocytes cultures from mAra h 1-3 plus HKLM-treated mice. These results show that immunotherapy with modified PN proteins and HKLM is effective for treating PN allergy in this model, and may be a potential approach for treating PNA. (+info)
Soymetide, an immunostimulating peptide derived from soybean beta-conglycinin, is an fMLP agonist.
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A tridecapeptide (MITLAIPVNKPGR) that stimulates phagocytosis of human neutrophils was isolated from a trypsin digest of soybean proteins. This peptide is derived from the soybean beta-conglycinin alpha' subunit and was named soymetide-13. The N-terminal methionine residue of soymetide-13 is essential for its activity, and removal of C-terminal residues revealed that soymetide-4 (MITL) is the minimal structure required for phagocytosis stimulation. Although they are not formylated at their N-termini, soymetides have a weak affinity for the N-formyl-methionyl-leucyl-phenylalanine (fMLP) receptor and their phagocytosis-stimulating activity is inhibited by the fMLP antagonist Boc-MLP. Interestingly, soymetide-4 promotes tumor necrosis factor alpha production at a higher level than soymetide-13 following oral administration in mice. (+info)
Storage and mobilization as antagonistic functional constraints on seed storage globulin evolution.
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When seeds germinate nearly all the proteins are degraded in senescing storage tissue cells. All these proteins act as amino acid reserves which are mobilized to nourish the seedling. Nevertheless, the major amount of the seeds' protein reserve consists of a few enzymatically inactive, abundant, genuine storage proteins. In their metabolism the conflicting processes of biosynthesis, protein turnover and breakdown, are temporally separated. No degradation of correctly formed storage proteins was observed at the time of synthesis and accumulation during seed maturation. Breakdown takes place after a (long) period of rest when seeds germinate and seedlings start growing. At that time genuine storage proteins are no longer synthesized. Genuine storage proteins have evolved structural features permitting controlled temporal patterns of protection and proteolysis. The acquisition of inserted sequence stretches as sites accessible to limited proteolysis played a key role in the evolution of this control system and happened in coevolution of genuine storage proteins with specific proteinases. This can be deduced from the results of current research on the mechanisms of limited and unlimited proteolysis of storage globulins and on storage globulin evolution. The evolved system of controlled structure-function interplay between storage globulins and proteinases is part of a syndrome that, in addition, comprises differential compartmentation and gene expression of storage proteins and proteinases for controlling the total spatial and temporal patterns of globulin storage and mobilization in maturing and germinating seeds. (+info)
Creation of soybean beta-conglycinin beta with strong phagocytosis-stimulating activity.
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beta-Conglycinin is composed of three kinds of subunit: alpha, alpha' and beta. A phagocytosis-stimulating peptide sequence (MITLAIPVNKPGR), soymetide, exists in the alpha' subunit of beta-conglycinin. Met at N terminus of the soymetide is essential for the activity. When Thr at the third residue from N terminus of the soymetide is replaced by Phe or Trp, the phagocytosis-stimulating activity greatly increases (ThrMet, Lys-->Thr, Phe, or Trp) into the beta subunit after confirmation of the effects of residue replacements by molecular modeling, suggesting that the introduced mutations might not prevent the correct folding. The studies of circular dichroism (CD), gel filtration and differential scanning calorimetry (DSC) of the mutants (I122M/K124T, I122M/K124F, I122M/K124W) expressed in E. coli demonstrated that they folded and self-assembled similarly to the wild type. This was confirmed by X-ray analysis of I122M/K124W crystal where the biggest residue tryptophane was introduced. The three mutants exhibited phagocytosis activities after digestion by trypsin, and the order was the wild type+info)
Subcellular localization of soybean 7S globulin in HepG2 cells and LDL receptor up-regulation by its alpha' constituent subunit.
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The aims of this work were to monitor the subcellular localization of soybean 7S globulin in HepG2 cells and determine its interaction with cell protein components, by using laser-induced fluorescence capillary electrophoresis (LIF-CE). Furthermore, we evaluated in the same cell line the involvement of the alpha' constituent subunit from 7S globulin in the modulation of LDL catabolism. The results indicated a main fluorescein isothiocyanate-tagged 7S globulin (FITC-7S) component in the cytosolic fraction, that was not present in the nuclear compartment. The electrophoretic mobility of this tagged component suggested either a dissociation of the 7S oligomer or its partial intracellular degradation. Interactions of soybean 7S globulin with FITC-thioredoxin 1 and FITC-cyclophilin B, HepG2 cell membrane proteins, were demonstrated in in vitro assays. In a separate experiment with HepG2 cells, the ability of the alpha' subunit purified from soybean 7S globulin to modulate the activity of the LDL receptors was evaluated by tracking the uptake and degradation of labeled LDL. The up-regulation of LDL receptors by the alpha' subunit, as further confirmed by a LDL receptor promoter assay, was significantly greater than that found in the control cells. In conclusion, this study, while confirming our previous indirect evidence of the key role of alpha' subunit on the cell cholesterol homeostasis, reveals a potentially interesting association of soybean 7S globulin with proteins, such as thioredoxin 1 and cyclophilin B, that are involved in cell protection against oxidative stress. (+info)
The soybean beta-conglycinin beta 51-63 fragment suppresses appetite by stimulating cholecystokinin release in rats.
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We previously demonstrated that soybean beta-conglycinin peptone suppresses food intake and gastric emptying by direct action on rat small intestinal mucosal cells to stimulate cholecystokinin (CCK) release. The aim of the present study was to define the active fragment in beta-conglycinin by using synthetic peptides chosen from the sequence of three beta-conglycinin subunits. We selected the fragments that had multiple nonadjacent arginine residues, and investigated their ability to bind to components of the rat intestinal brush border membrane as well as to stimulate CCK release and appetite suppression. The fragment from 51 to 63 of the beta subunit (beta 51-63) had the strongest binding activity. Intraduodenal infusion of beta 51-63 inhibited food intake and markedly increased portal CCK concentration. The threshold concentration of beta 51-63 to affect food intake was 3 micro mol/L. The CCK-A receptor antagonist abolished the beta 51-63-induced suppression of food intake. Three types of smaller fragments of beta 51-63 (beta 51-59, beta 53-63 and beta 53-59) and two types of fragments similar to beta 51-63 in the beta-conglycinin alpha and alpha' subunits (alpha 212-224 and alpha' 230-240) had less binding ability than did beta 51-63. Model peptides constructed with arginine (R) and glycine (G), such as GRGRGRG, had strong binding affinity, but peptides containing a single R or RR did not. These results indicate that the beta-conglycinin beta 51-63 fragment is the bioactive appetite suppressant in beta-conglycinin, and multiple arginine residues in the fragment may be involved in this effect. (+info)
Introduction of DPR, an enterostatin fragment peptide, into soybean beta-conglycinin alpha' subunit by site-directed mutagenesis.
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DPR, a fragment peptide of enterostatin (VPDPR) having hypocholesterolemic activity, was introduced into the three homologous sites, EPR, DYR, and DPI, in the soybean beta-conglycinin alpha' subunit by site-directed mutagenesis. The modified beta-conglycinin was expressed in Escherichia coli and recovered in the soluble fraction. After purification on ion-exchange HPLC, the modified beta-conglycinin was digested by trypsin to release integrated DPR. The yield of DPR from 1 mole of the modified beta-conglycinin was 1.2 mole. (+info)