摘要
大豆蛋白是一种优质蛋白资源,在保障我国膳食蛋白质供给方面发挥着重要作用。大豆蛋白及其降解产物具有调节胃肠道健康的功能。本研究室前期以碱性蛋白酶酶解大豆糖蛋白(8-伴大豆球蛋白),并从p-伴大豆球蛋白酶解物中分离大豆糖肽(SGP),且发现SGP具有抑制细菌粘附和调节肠粘膜屏障功能的活性,但是SGP具有抑制粘附作用的机制并不清楚,糖链在抑制粘附中发挥的作用也不明确。本课题在前期研究基础上,分析SGP与宿主细胞的相互作用,评价糖链在抑制粘附中发挥的作用,进一步探讨SGP抑制粘附的机制,为建立SGP的生理活性调控方法和应用技术创新提供理论依据；在明确理论问题的基础上,研究具有维持肠道健康作用的p-伴大豆球蛋白酶解物在模拟体外消化过程中以及在热处理条件下的稳定性,在技术层面上为β-伴大豆球蛋白酶解物作为食源肽类胃肠道调节剂或功能因子添加于各种类型的食品中提供基础。
本研究首先采用高碘酸盐氧化的方法制备了糖链破坏的大豆糖肽,并对破坏糖链的大豆糖肽性质进行了分析,结果表明高碘酸盐处理后的SGP中总糖相对含量由15.44%下降至10.93%,样品负电性增强,分子量相对较低的组分含量增加,疏水性相对较强的组分含量增加；高碘酸盐处理后,样品中芳香族氨基酸的暴露状态发生了变化,表明高碘酸盐氧化不仅可以部分破坏糖链,还会引起样品构象的变化。
以Caco-2细胞为模型,研究了SGP与Caco-2细胞的相互作用。结果表明,细胞表面Zeta电位值在添加SGP的30min内降低,后在24h内缓慢回复至加入肽前的水平；荧光结合实验结果显示,SGP能够结合于细胞膜表面,增加细胞膜的流动性。细胞膜上的脂筏区域被破坏后,SGP与细胞膜的结合作用明显减弱,表明SGP在细胞膜的脂筏区域与细胞膜发生结合作用。进一步比较SGP和periodate-treated SGP对致病菌粘附肠道细胞的抑制效果。结果表明,periodate-treatedSGP对实验所用Escherichia coli0114、Escherichia coli026、Salmonella enteritidis、S. typhimuriumLT2四株常见的肠道致病菌有一定的抑制粘附的活性。高碘酸盐氧化破坏糖链后对大肠杆菌粘附的抑制效果明显下降,对沙门氏菌粘附的抑制效果则影响不明显；且periodate-treated SGP对细胞膜流动性增加的程度小于完整的SGP。这些结果意味着糖链在大豆糖肽抑制大肠杆菌粘附细胞的作用中发挥了重要作用。
β-伴大豆球蛋白酶解物的稳定性是将其应用于食品生产的基础,本研究分析了其在热处理(85℃,30min)条件下的稳定性。结果表明加热会导致β-伴大豆球蛋白酶解物发生可溶性聚集,且聚集程度与溶液浓度有关；较低浓度时(小于200mg/mL),β-伴大豆球蛋白酶解物溶液有良好的耐热性；在较高浓度时,加热会导致β-伴大豆球蛋白解物在胃肠道消化酶的作用下的降解作用。结果表明,在模拟消化过程中,肽段发生了部分酶解物负电性增加,钙敏感性下降。进一步采用体外模拟消化的方式研究了β-伴大豆球蛋白酶降解,但是降解的程度较低,其中10-20kDa组分含量保持相对稳定。亲水性相对较强的组分含量略有减少,疏水性较强的组分略有增加,但整体表面疏水性指数没有显著变化(P<0.05)。说明β-伴大豆球蛋白酶解物中的10-20kDa有效功能组分对胃肠道消化酶的降解有较好的抵抗作用,在消化过程中能保持相对稳定。
Soybean protein is the quality resources of food protein, and plays an important role in protein supply in the daily diet for people in China. Soybean protein and its derivatives could regulate intestinal function. Soybean glycoprotein (β-conglycinin) was hydrolyzed using alcalase2.4L and soybean glycopeptides (SGP) was seperated from β-conglycinin hydrolysate in previous study. SGP could inhibit the adhesion of pathogens on human intestinal cells, but the mechanism of this effect and role of carbohydrate chain are still unclear. Based on these results, interactions between SGP and host cells and the role of carbohydrate chain in inhibiting adhesion were examined in this study. The mechanism of inhibiting adhesion was also analyzed in this study. The study provides theoretical foundation for regulation of SGP biological activity and application of the technical innovation. On the basis of theoretical research, the in vitro digestibility and thermal stability of β-conglycinin hydrolysates were also evaluated in this study, which could supply the technical foundation for the application of β-conglycinin hydrolysates as functional component for intestinal health in food industry.
The carbohydrate chain in SGP was destroyed partly by periodate treatment and properties of periodate-treated SGP were also examined in this study. Results showed that the carbohydrate content in SGP decreased from15.44%to10.93%after periodate treatment. The electronegativity, content of peptides with relatively low molecular weight and the content of component with relatively high hydrophobicity were increased in periodate-treated SGP. Moreover, the exposure of the aromatic amino acids in periodate-treated SGP changed. These results indicated that periodate treatment could destroy part of sugar chain and led to conformation changes in SGP.
The changes in Zeta potential and cell membrane fluidity of Caco-2cells with or without SGP and the binding interaction between SGP and cell membrane were examined using a Caco-2cell model. Results showed that the cell surface Zeta potential decreased after30min of incubation with SGP and then slowly restored to the level of Caco-2cells alone within24h. Fluorescent experimental results showed that SGP could bind to the cell membrane, and increase the membrane fluidity of Caco-2cells. The extent of binding decreased when the lipid rafts in the cell membrane were destroyed, indicating that SGP bound to the cell membrane on lipid rafts,
The inhibitory effects of SGP and periodate-treated the SGP on the pathogenic bacteria adhesion intestinal cells were compared. Results showed that SGP presented inhibition effect on the bacterial adhesion to the host cell. The inhibition effect of periodate-treated the SGP on Escherichia coli adhesion were significantly lower than SGP, but there was no significant differenece between the inhibition effects of SGP and periodate-treated the SGP on Salmonella adhesion. Moreover, the increasing extent of membrane fluidity of Caco-2cells affected by periodate-treated the SGP was lower than SGP. These results suggested that carbohydrate chain in SGP played an important role in inhibiting Escherichia coli adhesion to intestinal cells.
The stability of P-conglycinin hydrolysates was foundation of using it in food processing. The stability of soybean β-conglycinin hydrolysates in heat treatment (85℃,30min) was studied. Results indicated that heat treatment caused β-conglycinin hydrolysates soluble aggregation and the aggregation degree related to sample concentration. β-Conglycinin hydrolysates solution had good heat resistance at lower concentration (<200mg/mL), while heat treatment induced electronegativity increased and calcium sensitivity decreased at higher concentration. The digestibility in vitro of P-conglycinin hydrolysates was also studied. The results showed that β-conglycinin hydrolysates were degraded through in vitro digestion, but the degree of degradation was relatively low. The content of peptides with molecular weights from10kDa to20kDa remained stable during digestion in vitro. The proportion of highly hydrophobic peptides decreased through in vitro digestion, but no significant change in the surface hydrophobicity indices of digestion products was observed (P<0.05). These results indicate that peptides with molecular weights from10kDa to20kDa in the β-conglycinin hydrolysates resisted digestion by pepsin and trypsin and they remained stable during in vitro digestion.
引文
董群,郑丽伊.改良的苯酚-硫酸法测定多糖和寡糖含量的研究[J].中国药学杂志,1996,31(9)：550-553.
刘贺.大豆肽与钙结合机制及Caco-2细胞内肽促钙转运模式研究[博士学位论文].2013,北京：中国农业大学.
杨柏崇.大豆糖肽抑制致病菌粘附及调节肠粘膜屏障功能的研究[博士学位论文].2008,北京：中国农业大学.
袁德保.大豆蛋白热聚集行为及其机理研究[博士学位论文].2010,广东：华南理工大学.
张雪梅.大豆蛋白酶解产物中的糖肽及其对小鼠脾细胞增殖效果的研究[硕士学位论文].2003,北京：中国农业大学.
Abraham SN, Duncan MJ, Li G, et al. Bacterial penetration of the mucosal barrier by targeting lipid rafts [J]. Journal of Investigative Medicine,2005,53(6):318-321.
Abubakar A, Saito T, Kitazawa H, et al. Structural analysis of new antihypertensive peptides derived from cheese whey protein by proteinase K digestion [J]. Journal of Dairy Science,1998,81(12): 3131-3138.
Altankov G, Richau K, Groth T. The role of surface Zeta potential and substratum chemistry for regulation of dermal fibroblasts interaction [J]. Materialwissenschaft und Werkstofftechnik,2003, 34(12):1120-1128.
Amigo-Benavent M, Clemente A, Ferranti P et al. Digestibility and immunoreactivity of soybean β-conglycinin and its deglycosylated form [J]. Food Chemistry,2011,129(4):1598-1605.
Aoki S, Epand RM. Caveolin-1 hydrophobic segment peptides insertion into membrane mimetic systems:Role of Proline residue [J]. Biochimica et Biophysica Acta (BBA)-Biomembranes,2012, 1818(1):12-18.
Bayer AS, Prasad R, Chandra J, et al. In vitro resistance of Staphylococcus aureus to thrombin-induced platelet microbicidal protein is associated with alterations in cytoplasmic membrane fluidity [J]. Infection and Immunity,2000,68(6):3548-3553.
Beachey EH. Bacterial adherence:adhesin-receptor interactions mediating the attachment of bacteria to mucosal surfaces [J]. Journal of Infectious Diseases,1981,143(3):325-345.
Bennion BJ, Daggett V. The molecular basis for the chemical denaturation of proteins by urea [J]. Proceedings of the National Academy of Sciences,2003,100(9):5142-5147.
Beintema JJ. Do asparagine-linked carbohydrate chains in glycoproteins have a preference for β-bends? [J]. Bioscience reports,1986,6(8):709-714.
Berkes J, Viswanathan VK, Savkovic SD, et al. Intestinal epithelial responses to enteric pathogens: effects on the tight junction barrier, ion transport, and inflammation [J]. Gut,2003,52(3):439-451.
Bianchi MA, Del Rio D, Pellegrini N, et al. A fluorescence-based method for the detection of adhesive properties of lactic acid bacteria to Caco-2 cells [J]. Letters in Applied Microbiology,2004,39(3): 301-305.
Bishop JR, Gagneux P. Evolution of carbohydrate antigens—microbial forces shaping host glycomes? [J]. Glycobiology,2007,17(5):23R-34R.
Bosch Ojeda C, Sanchez Rojas F. Recent applications in derivative ultraviolet/visible absorption spectrophotometry:2009-2011:A review [J]. Microchemical Journal,2013,106:1-16.
Brashears MM, Reilly SS, Gilliland SE. Antagonistic action of cells of Lactobacillus lactis toward Escherichia coli 0157:H7 on refrigerated raw chicken meat [J]. Journal of Food Protection,1998, 61(2):166-170.
Breines DM, Burnham JC. Modulation of Escherichia coli type 1 fimbrial expression and adherence to uroepithelial cells following exposure of logarithmic phase cells to quinolones at subinhibitory concentrations [J]. Journal of Antimicrobial Chemotherapy,1994,34(2):205-221.
Briick WM, Kelleher SL, Gibson GR, et al. The effects of a-lactalbumin and glycomacropeptide on the association of Caco-2 cells by enteropathogenic Escherichia coli, Salmonella typhimurium and Shigella flexneri [J]. FEMS Microbiology Letters,2006,259(1):158-162.
Bosch Ojeda C, Sanchez Rojas F. Recent developments in derivative ultraviolet/visible absorption Spectrophotometry [J]. Analytica Chimica Acta,2004,518(1):1-24.
Burger O, Ofek I, Tabak M, et al. A high molecular mass constituent of cranberry juice inhibits Helicobacter pylori adhesion to human gastric mucus [J]. FEMS Immunology & Medical Microbiology,2000,29(4):295-301.
Burger O, Weiss E, Sharon N, et al. Inhibition of Helicobacter pylori adhesion to human gastric mucus by a high-molecular-weight constituent of cranberry juice [J]. Critical Reviews in Food Science and Nutrition,2002,42(S3):279-284.
Burstein EA, Emelyanenko VI. Log-normal description of fluorescence spectra of organic fluorophores [J]. Photochemistry and Photobiology,1996,64(2):316-320.
Celli J, Deng W, Finlay BB. Enteropathogenic Escherichia coli (EPEC) attachment to epithelial cells: exploiting the host cell cytoskeleton from the outside [J]. Cellular Microbiology,2000,2(1):1-9.
Chen HM, Muramoto K, Yamauchi F. Structural analysis of antioxidative peptides from soybean β-conglycinin [J]. Journal of Agricultural and Food Chemistry,1995,43:574-578.
Cherukuri A, Dykstra M, Pierce SK. Floating the raft hypothesis:lipid rafts play a role in immune cell activation [J]. Immunity,2001,14(6):657-660.
Cho JA, Chinnapen DJF, Aamar E, et al. Insights on the trafficking and retro-translocation of glycosphingolipid-binding bacterial toxins [J]. Frontiers in Cellular and Infection Microbiology, 2012,2(51):1-5.
Chrispeels MJ. Sorting of proteins in the secretory system [J]. Annual Review of Plant Biology,1991, 42(1):21-53.
Coppa GV, Zampini L, Galeazzi T, et al. Human milk oligosaccharides inhibit the adhesion to Caco-2 cells of diarrheal pathogens:Escherichia coli, Vibrio cholerae, and Salmonella fyris [J]. Pediatric Research,2006,59(3):377-382.
Cravioto A, Tello A, Villafan H, et al. Inhibition of localized adhesion of enteropathogenic Escherichia coli to HEp-2 cells by immunoglobulin and oligosaccharide fractions of human colostrum and breast milk[J]. Journal of Infectious Diseases,1991,163(6):1247-1255.
Daig R, Andus T, Aschenbrenner E, et al. Increased interleukin 8 expression in the colon mucosa of patients with inflammatory bowel disease [J]. Gut,1996,38(2):216-222.
Dal SM, Bovio C, Culici M, et al. The combination of the SH metabolite of erdosteine (a mucoactive drug) and ciprofloxacin increases the inhibition of bacterial adhesiveness achieved by ciprofloxacin alone [J]. Drugs under Experimental and Clinical Research,2002,28(2-3):75.
de Repentigny L, Aumont F, Bernard K, et al. Characterization of binding of Candida albicans to small intestinal mucin and its role in adherence to mucosal epithelial cells [J]. Infection and Immunity, 2000,68(6):3172-3179.
DeVinney R, Gauthier A, Abe A, et al. Enteropathogenic Escherichia coli:a pathogen that inserts its own receptor into host cells [J]. Cellular and Molecular Life Sciences CMLS,1999,55(6-7): 961-976.
Dickinson E, Stainsby G Colloids in food [M]. Applied Science Publishers,1982.
Donnenberg MS, Kaper JB, Finlay BB. Interactions between enteropathogenic Escherichia coli and host epithelial cells [J]. Trends in Microbiology,1997,5(3):109-114.
Donnenberg MS. Interactions between enteropathogenic Escherichia coli and epithelial cells [J]. Clinical Infectious Diseases,1999,28(3):451-455.
Dubois M, Gilles KA, Hamilton JK, et al. Colorimetric method for determination of sugars and related substances [J]. Analytical Chemistry,1956,28(3):350-356.
Eftink MR, Ghiron CA. Exposure of tryptophanyl residues in proteins. Quantitative determination by fluorescence quenching studies [J]. Biochemistry,1976,15(3):672-680.
Finlay BB. Interactions between Salmonella typhimurium, enteropathogenic Escherichia coli (EPEC), and host epithelial cells [J]. Advances in Dental Research,1995,9(1):31-36.
Finlay BB, Cossart P. Exploitation of mammalian host cell functions by bacterial pathogens [J]. Science, 1997,276(5313):718-725.
Finlay BB, Brumell JH. Salmonella interactions with host cells:in vitro to in vivo [J]. Philosophical Transactions of the Royal Society of London. Series B:Biological Sciences,2000,355(1397): 623-631.
Firon N, Ashkenazi S, Mirelman D, et al. Aromatic alpha-glycosides of mannose are powerful inhibitors of the adherence of type 1 fimbriated Escherichia coli to yeast and intestinal epithelial cells [J]. Infection and Immunity,1987,55(2):472-476.
Frankel G, Phillips AD, Rosenshine I, et al. Enteropathogenic and enterohaemorrhagic Escherichia coli: more subversive elements [J]. Molecular Microbiology,1998,30(5):911-921.
Gatellier P, Kondjoyan A, Portanguen S, et al. Determination of aromatic amino acid content in cooked meat by derivative spectrophotometry:Implications for nutritional quality of meat [J]. Food Chemistry,2009,114(3):1074-1078.
Goot FG, Harder T. Raft membrane domains:from a liquid-ordered membrane phase to a site of pathogen attack [J]. Seminars in Immunology,2001,13(2):89-97.
Haney EF, Lau F, Vogel HJ. Solution structures and model membrane interactions of lactoferrampin, an antimicrobial peptide derived from bovine lactoferrin [J]. Biochimica et Biophysica Acta (BBA)-Biomembranes,2007,1768(10):2355-2364.
Hartlova A, Cerveny L, Hubalek M, et al. Membrane rafts:a potential gateway for bacterial entry into host cells [J]. Microbiology and Immunology,2010,54(4):237-245.
Helenius A, Aebi M. Intracellular functions of N-linked glycans [J]. Science,2001,291(5512): 2364-2369.
Heron DS, Shinitzky M, Hershkowitz M, et al. Lipid fluidity markedly modulates the binding of serotonin to mouse brain membranes [J]. Proceedings of the National Academy of Sciences,1980, 77(12):7463-7467.
Holowka D, Baird B. Fc∈RI as a paradigm for a lipid raft-dependent receptor in hematopoietic cells [J]. Seminars in Immunology,2001,13(2):99-105.
Hosokawa T, Kusugami K, Ina K, et al. Interleukin-6 and soluble interleukin-6 receptor in the colonic mucosa of inflammatory bowel disease [J]. Journal of Gastroenterology and Hepatology,1999, 14(10):987-996.
Huang B, Zou GL, Yang TM. Studies on the interaction between adriamycin and bovine serum albumin [J].ACTA CHIMICA SINICA-CHINESE EDITION-,2002,60(10):1867-1871.
Hunter RJ. Foundations of colloidal science [M]. Oxford:Oxford University Press,1986.
Imberty A, Varrot A. Microbial recognition of human cell surface glycoconjugates [J]. Current Opinion in Structural Biology,2008,18(5):567-576.
Imperiali B, O'Connor SE. Effect of N-linked glycosylation on glycopeptide and glycoprotein structure [J]. Current Opinion in Chemical Biology,1999,3(6):643-649.
Ismaili A, Meddings JB, Ratnam S, et al. Modulation of host cell membrane fluidity:a novel mechanism for preventing bacterial adhesion [J]. American Journal of Physiology-Gastrointestinal and Liver Physiology,1999,277(1):G201-G208.
Janes PW, Ley SC, Magee AI, et al. The role of lipid rafts in T cell antigen receptor (TCR) signaling [J]. Seminars in Immunology,2000,12(1):23-34.
Jankowska A, Laubitz D, Antushevich H, et al. Competition of Lactobacillus paracasei with Salmonella enterica for adhesion to Caco-2 cells [J]. BioMed Research International,2008(2008):1-6.
Jourd'Heuil D, Vaananen P, Meddings J B. Lipid peroxidation of the brush-border membrane: membrane physical properties and glucose transport [J]. American Journal of Physiology-Gastrointestinal and Liver Physiology,1993,264(6):G1009-G1015.
Kariko K, Weissman D, Welsh FA. Inhibition of toll-like receptor and cytokine signaling-a unifying theme in ischemic tolerance [J]. Journal of Cerebral Blood Flow & Metabolism,2004,24(11): 1288-1304.
Katsube T, Kang IJ, Takenaka Y, et al. N-Glycosylation does not affect assembly and targeting of proglycinin in yeast [J]. Biochimica et Biophysica Acta (BBA)-General Subjects,1998,1379(1): 107-117.
Kim SY, Park PSW, Rhee KC. Functional properties of proteolytic enzyme modified soy protein isolate [J]. Journal of Agricultural and Food Chemistry,1990,38(3):651-656.
Kimura Y, Ohno A, Takagi S. Structural analysis of N-glycans of storage glycoproteins in soybean (Glycine max. L) seed [J]. Bioscience, Biotechnology, and Biochemistry,1997,61(11):1866-1871.
Kobayashi K, Hattori M, Hara-Kudo Y, et al. Glycopeptide derived from hen egg ovomucin has the ability to bind enterohemorrhagic Escherichia coli 0157:H7 [J]. Journal of Agricultural and Food Chemistry,2004,52(18):5740-5746.
Korhonen TK, Vaisanen-Rhen V, Rhen M, et al. Escherichia coli fimbriae recognizing sialyl galactosides [J]. Journal of Bacteriology,1984,159(2):762-766.
Koshiyama I. Carbohydrate component in 7S protein of soybean casein fraction [J]. Agricultural and Biological Chemistry,1966,30(7):646-650.
Krachler AM, Orth K. Targeting the bacteria-host interface [J]. Virulence,2013,4(4):284-294.
Kraft CA, Garrido JL, Leiva-Vega L, et al. Quantitative analysis of protein-lipid interactions using tryptophan fluorescence [J]. Science Signaling,2009,2(99):p14.
Kourie JI, Shorthouse AA. Properties of cytotoxic peptide-formed ion channels [J]. American Journal of Physiology-Cell Physiology,2000,278(6):C1063-C1087.
Labrecque J, Bodet C, Chandad F, et al. Effects of a high-molecular-weight cranberry fraction on growth, biofilm formation and adherence of Porphyromonas gingivalis [J]. Journal of Antimicrobial Chemotherapy,2006,58(2):439-443.
Ladokhin AS, Jayasinghe S, White SH. How to measure and analyze tryptophan fluorescence in membranes properly, and why bother? [J]. Analytical Biochemistry,2000,285(2):235-245.
Lafont F, Van Der Goot FG. Bacterial invasion via lipid rafts [J]. Cellular Microbiology,2005,7(5): 613-620.
Lowry OH, Rosebrough NJ, Farr A L, et al. Protein measurement with the Folin phenol reagent [J]. Journal of Biological Chemistry,1951,193(1):265-275.
Lee JH, Shim JS, Lee JS, et al. Inhibition of pathogenic bacterial adhesion by acidic polysaccharide from green tea (Camellia sinensis) [J]. Journal of Agricultural and Food Chemistry, 2006,54(23):8717-8723.
Lengsfeld C, Titgemeyer F, Faller G, et al. Glycosylated compounds from okra inhibit adhesion of Helicobacter pylori to human gastric mucosa [J]. Journal of Agricultural and Food Chemistry, 2004,52(6):1495-1503.
Li SC, Deber CM. A measure of helical propensity for amino acids in membrane environments [J]. Nature Structural & Molecular Biology,1994,1(6):368-373.
Liu SQ, Shibata A, Ueno S, et al. Investigation of interaction of Leu-enkephalin with lipid membranes [J]. Colloids and Surfaces B:Biointerfaces,2006,48(2):148-158.
Liu Y, Guo R. Aggregation properties of aqueous casein hydrolysate solutions at different pH [J]. International Dairy Journal,2008,18(10):1022-1027.
Mahmoud MI, Malone WT, Cordle CT. Enzymatic hydrolysis of casein:effect of degree of hydrolysis on antigenicity and physical properties [J]. Journal of food science,1992,57(5):1223-1229.
Malhotra A, Coupland JN. The effect of surfactants on the solubility, Zeta potential, and viscosity of soy protein isolates [J]. Food Hydrocolloids,2004,18(1):101-108.
Manjarrez-Hernandez HA, Aitken A, Baldwin TJ, et al. Intestinal epithelial cell protein phosphorylation in enteropathogenic Escherichia coli diarrhoea [J]. The Lancet,1992,339(8792):521-523.
Martinez-Villaluenga C, Dia VP, Berhow M, et al. Protein hydrolysates from β-conglycinin enriched soybean genotypes inhibit lipid accumulation and inflammation in vitro [J]. Molecular nutrition & food research,2009,53(8):1007-1018.
Maruyama N, Katsube T, Wada Y, et al. The roles of the N-linked glycans and extension regions of soybean beta-conglycinin in folding, assembly and structural features [J]. European Journal of Biochemistry.1998,258(2):854-862.
Maruyama N, Salleh MRM, Takahashi K, et al. The effect of the N-linked glycans on structural features and physicochemical functions of soybean β-conglycinin homotrimers [J]. Journal of the American Oil Chemists'Society,2002,79(2):139-144.
McCormick BA, Hofman PM, Kim J, et al. Surface attachment of Salmonella typhimurium to intestinal epithelia imprints the subepithelial matrix with gradients chemotactic for neutrophils [J]. The Journal of Cell Biology,1995,131(6):1599-1608.
McCormick BA, Parkos CA, Colgan SP, et al. Apical secretion of a pathogen-elicited epithelial chemoattractant activity in response to surface colonization of intestinal epithelia.by Salmonella typhimurium [J]. The Journal of Immunology,1998,160(1):455-466.
Miteva T, Palmer L, Kloppenburg L, et al. Interplay of thermochromicity and liquid crystalline behavior in poly (p-phenyleneethynylene) s:π-π Interactions or planarization of the conjugated backbone? [J]. Macromolecules,2000,33(3):652-654.
Morrissey PEW, Folan MA, Baird AW, et al. Effects of a partially digested whey protein concentrate on Salmonella enterica serotype Typhimurium adhesion to Caco-2 cells:Prevention of Salmonella adhesion to Caco-2 cells using whey [J]. Food Control,2010,21(8):1113-1120.
Nakajima K, Tamura N, Kobayashi-Hattori K, et al. Prevention of intestinal infection by glycomacropeptide [J]. Bioscience, Biotechnology, and Biochemistry,2005,69(12):2294-2301.
Nataro JP, Kaper JB. Diarrheagenic Escherichia coli [J]. Clinical Microbiology Reviews,1998,11(1): 142-201.
Nakamura S, Takasaki H, Kobayashi K, et al. Hyperglycosylation of hen egg white lysozyme in yeast [J]. Journal ofBiological Chemistry,1993,268(17):12706-12712.
O'Connor SE, Imperiali B. Modulation of protein structure and function by asparagine-linked glycosylation [J]. Chemistry & Biology,1996,3(10):803-812.
Oelschlaeger TA. Mechanisms of probiotic actions-a review [J]. International Journal of Medical Microbiology,2010,300(1):57-62.
Ofek I, Mirelman D, Sharon N. Adherence of Escherichia coli to human mucosal cells mediated by mannose receptors [J]. Nature,1977,265(5595):623-625.
Ofek I, Doyle RJ. Bacterial adhesion to cells and tissues [M]. New York:Chapman & Hall,1994.
Ofek I, Goldhar J, Keisari Y, et al. Nonopsonic phagocytosis of microorganisms [J]. Annual Reviews in Microbiology,1995,49(1):239-276.
Ofek I, Hasty DL, Sharon N. Anti-adhesion therapy of bacterial diseases:prospects and problems [J]. FEMS Immunology & Medical Microbiology,2003a,38(3):181-191.
Ofek I, Hasty DL, Doyle RJ, et al. Bacterial adhesion to animal cells and tissues [M]. Washington, DC: ASM press,2003b.
Ofek I, Sharon N, Abraham SN. Bacterial adhesion [M] The Prokaryotes. Springer New York,2006: 16-31.
Ogawa M, Shimizu K, Nomoto K, et al. Inhibition of in vitro growth of Shiga toxin-producing Escherichia coli 0157:H7 by probiotic Lactobacillus strains due to production of lactic acid [J]. International Journal of Food Microbiology,2001,68(1):135-140.
Paulson AT, Tung MA. Solubility, hydrophobicity and net charge of succinylated canola protein isolate [J]. Journal of Food Science,1987,52(6):1557-1561.
Radin NS. Preventing the binding of pathogens to the host by controlling sphingolipid metabolism [J]. Microbes and Infection,2006,8(3):938-945.
Rankin SE, Watts A, Pinheiro TJT. Electrostatic and hydrophobic contributions to the folding mechanism of apocytochrome c driven by the interaction with lipid [J]. Biochemistry,1998,37(36): 12588-12595.
Rhoades JR, Gibson GR, Formentin K, et al. Caseinoglycomacropeptide inhibits adhesion of pathogenic Escherichia coli strains to human cells in culture [J]. Journal of Dairy Science,2005,88(10): 3455-3459.
Rosenshine I, Donnenberg MS, Kaper JB, et al. Signal transduction between enteropathogenic Escherichia coli (EPEC) and epithelial cells:EPEC induces tyrosine phosphorylation of host cell proteins to initiate cytoskeletal rearrangement and bacterial uptake [J]. The EMBO Journal,1992, 11(10):3551.
Roufik S, Gauthier SF, Turgeon SL. In vitro digestibility of bioactive peptides derived from bovine β-lactoglobulin [J]. International Dairy Journal,2006,16(4):294-302.
Ryan M, McEvoy E, Duignan S, et al. Thermal stability of soy protein isolate and hydrolysate ingredients [J]. Food Chemistry,2008,108(2):503-510.
Sagle LB, Zhang Y, Litosh VA, et al. Investigating the hydrogen-bonding model of urea denaturation [J]. Journal of the American Chemical Society,2009,131(26):9304-9310.
Saito T, Nakamura T, Kitazawa H, et al. Isolation and structural analysis of antihypertensive peptides that exist naturally in Gouda cheese [J]. Journal of Dairy Science,2000,83(7):1434-1440.
Salyer JL, Bohnsack JF, Knape WA, et al. Mechanisms of tumor necrosis factor-alpha alteration of PMN adhesion and migration [J]. The American Journal of Pathology,1990,136(4):831-841.
Sato H, Feix JB. Peptide-membrane interactions and mechanisms of membrane destruction by amphipathic a-helical antimicrobial peptides [J]. Biochimica et Biophysica Acta (BBA)-Biomembranes,2006,1758(9):1245-1256.
Schagger H, Von Jagow G. Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa [J]. Analytical biochemistry,1987,166(2): 368-379.
Schagger H. Tricine-SDS-PAGE [J]. Nature protocols,2006,1(1):16-22.
Segura-Campos M, Chel-Guerrero L, Betancur-Ancona D, et al. Bioavailability of bioactive peptides [J]. Food Reviews International,2011,27(3):213-226.
Sharon N, Ofek I. Fighting infectious diseases with inhibitors of microbial adhesion to host tissues [J]. Critical Reviews in Food Science and Nutrition,2002,42(S3):267-272.
Sharon N. Carbohydrates as future anti-adhesion drugs for infectious diseases [J]. Biochimica et Biophysica Acta (BBA)-General Subjects,2006,1760(4):527-537.
Sheets ED, Holowka D, Baird B. Membrane organization in immunoglobulin E receptor signaling [J]. Current Opinion in Chemical Biology,1999,3(1):95-99.
Shmuely H, Ofek I, Weiss EI, et al. Cranberry components for the therapy of infectious disease [J]. Current Opinion in Biotechnology,2012,23(2):148-152.
Shinitzky, M. Membrane fluidity and cellular functions. In:Physiology of Membrane Fluidity, edited by M. Shinitzky. BocaRaton, FL:CRC,1984,1-51.
Simons K, Toomre D. Lipid rafts and signal transduction [J]. Nature Reviews Molecular Cell Biology, 2000,1(1):31-39.
Simons K, Ehehalt R. Cholesterol, lipid rafts, and disease [J]. Journal of Clinical Investigation,2002, 110(5):597-603.
Smith S, Elbein AD, Pan YT. Inhibition of bacterial binding by high-mannose oligosaccharides:U.S. Patent 5,939,279 [P].1999.
Spector AA, Yorek MA. Membrane lipid composition and cellular function [J]. Journal of Lipid Research,1985,26(9):1015-1035.
Steele-Mortimer O, Knodler LA, Finlay BB. Poisons, ruffles and rockets:bacterial pathogens and the host cell cytoskeleton [J]. Traffic,2000,1(2):107-118.
Stromquist M, Falk P, Bergstrom S, et al. Human milk κ-casein and inhibition of Helicobacter pylori adhesion to human gastric mucosa [J]. Journal of Pediatric Gastroenterology and Nutrition,1995, 21:288-296.
Sugita-Konishi Y, Sakanaka S, Sasaki K, et al. Inhibition of bacterial adhesion and Salmonella infection in BALB/c mice by sialyloligosaccharides and their derivatives from chicken egg yolk [J]. Journal of Agricultural and Food Chemistry,2002,50(12):3607-3613.
Sugita-Konishi Y, Kobayashi K, Sakanaka S, et al. Preventive Effect of Sialylglycopeptide Nondigestive Polysaccharide Conjugates on Salmonella Infection [J]. Journal of Agricultural and Food Chemistry,2004,52(17):5443-5448.
Suzuki T, Tanabe S, Hara H. Kaempferol enhances intestinal barrier function through the cytoskeletal association and expression of tight junction proteins in Caco-2 cells [J]. The Journal of Nutrition, 2011,141(1):87-94.
Svensson M, Frendeus B, Butters T, et al. Glycolipid depletion in antimicrobial therapy [J]. Molecular Microbiology,2003,47(2):453-461.
Thennarasu S, Tan A, Penumatchu R, et al. Antimicrobial and membrane disrupting activities of a peptide derived from the human cathelicidin antimicrobial peptide LL37 [J]. Biophysical Journal, 2010,98(2):248-257.
Tome'D, Ledoux N. Nutritional and physiological role of milk protein components [J]. International Dairy Federation,1998,336:11-16.
Tsuruki T, Kishi K, Takahashi M, et al. Soymetide, an immunostimulating peptide derived from soybean β-conglycinin, is an fMLP agonist [J].FEBS Letters,2003,540(1):206-210.
Tuomola EM, Ouwehand AC, Salminen SJ. The effect of probiotic bacteria on the adhesion of pathogens to human intestinal mucus [J]. FEMS Immunology & Medical Microbiology,1999,26(2): 137-142.
Uma L, Sharma Y, Balasubramanian D. A conformational study of corneal dermatan sulfate proteoglycan using fluorescence spectroscopy [J]. International Journal of Biological Macromolecules,1996,19(1):75-80.
Utsumi S, Matsumura Y, Mori T. Structure-function relationships of soy proteins [J]. Food Science and Technology-New York-Marcel Dekker-,1997:257-292.
Vallance B A, Finlay BB. Exploitation of host cells by enteropathogenic Escherichia coli [J]. Proceedings of the National Academy of Sciences,2000,97(16):8799-8806.
Vanapalli S, Coupland JN. Characterization of food colloids by phase analysis light scattering [J]. Food Hydrocolloids,2000,14(4):315-317.
Vandenbergh PA. Lactic acid bacteria, their metabolic products and interference with microbial growth [J]. FEMS Microbiology Reviews,1993,12(1):221-237.
Wallqvist A, Covell DG, Thirumalai D. Hydrophobic interactions in aqueous urea solutions with implications for the mechanism of protein denaturation [J]. Journal of the American Chemical Society,1998,120(2):427-428.
Wang C, Eufemi M, Turano C, et al. Influence of the carbohydrate moiety on the stability of glycoproteins [J]. Biochemistry,1996,35(23):7299-7307.
Wang T, Qin GX, Zhao Y, et al. Comparative study on the stability of soybean (Glycine max) β-conglycinin in vivo [J]. Food and Agricultural Immunology,2009,20(4):295-304.
Wojnicz D, Jankowski S. Effects of subinhibitory concentrations of amikacin and ciprofloxacin on the hydrophobicity and adherence to epithelial cells of uropathogenic Escherichia coli strains [J]. International Journal of Antimicrobial Agents,2007,29(6):700-704.
Wormald MR, Dwek RA. Glycoproteins:glycan presentation and protein-fold stability [J]. Structure, 1999,7(7):R155-R160.
Yamagishi T, Yamauchi F, Shibasaki K. State of aromatic amino acid residues in soybean 11S globulin heated in the presence of N-ethylmaleimide by derivative spectrophotometry [J]. Agricultural and Biological Chemistry,1981,45(2):459-467.
Yamagishi T, Ebina F, Yamauchi F. Analysis of the state of aromatic amino acid residues in heated soybean 7S globulin by absorption derivative spectrophotometry and spectrofluorimetry [J]. Agricultural and Biological Chemistry,1982,46(10):2441-2448.
Yamauchi F, Kawase M, Kanbe M, et al. Separation of the P-aspartamido-carbohydrate fractions from soybean 7s protein:protein-carbohydrate linkage [J]. Agricultural and Biological Chemistry,1975, 39(4):873-878.
Yamauchi F, Thanh VH, Kawase M, et al. Separation of the glycopeptides from soybean 7S protein: their amino acid sequences [J]. Agricultural and Biological Chemistry,1976,40(4):691-696.
Yamauchi F, Yamagishi T. Carbohydrate sequence of a soybean 7S protein [J]. Agricultural and Biological Chemistry,1979,43(3):505-510.
Yamul DK, Lupano CE. Whey protein concentrate gels with honey and wheat flour [J]. Food Research International,2005,38(5):511-522.
Yang BC, Zhang XM, Bao XL, et al. Glycopeptide derived from soybean P-conglycinin inhibits the adhesion of Escherichia coli arid Salmonella to human intestinal cells [J]. Food Research International,2008,41(6):594-599.
Yang BC, Lv Y, Chen Y, et al. Inhibitory Action of soybean β-conglycinin hydrolysates on Salmonella typhimurium translocation in Caco-2 epithelial cell monolayers [J]. Journal of Agricultural and Food Chemistry,2008,56(16):7522-7527.
Yang ST, Shin SY, Kim JI. Interaction mode of a symmetric Trp-rich undeca peptide PST11-RK with lipid bilayers [J]. FEBS Letters,2007,581(1):157-163.
Yuli I, Tomonaga A, Synderman R. Chemoattractant receptor functions in human polymorphonuclear leukocytes are divergently altered by membrane fluidizers [J]. Proceedings of the National Academy of Sciences,1982,79(19):5906-5910.
Zafriri D, Oron Y, Eisenstein BI, et al. Growth advantage and enhanced toxicity of Escherichia coli adherent to tissue culture cells due to restricted diffusion of products secreted by the cells [J]. Journal of Clinical Investigation,1987,79(4):1210.
Zhao Y, Qin GX, Sun ZW, et al. Stability and immunoreactivity of glycinin and β-conglycinin to hydrolysis in vitro [J]. Food and Agricultural Immunology,2010,21(3):253-263.
Zhang Y, Yang M, Portney NG, et al. Zeta potential:a surface electrical characteristic to probe the interaction of nanoparticles with normal and cancer human breast epithelial cells [J]. Biomedical Microdevices,2008,10(2):321-328.
Zhao HX, Lappalainen P. A simple guide to biochemical approaches for analyzing protein-lipid interactions [J]. Molecular Biology of the Cell,2012,23(15):2823-2830.