| Peanut(Arachis hypogaea Linn)is a widely distributed and nutrient-rich plant resource.However,the peanut allergenic protein contained in it often causes food allergies.Peanut allergy is a typical Ig E-mediated allergic reaction and it can be life-threatening in severe cases.Symptoms often appears in skin,respiratory tract,digestive system,etc.At present,studies have shown that the combination of polyphenol and peanut protein can behave a reduced allergenicity.In order to further realize the mechanism of polyphenols alleviating the sensitization of peanut protein and understand the structure-activity relationship,we used multi-spectroscopy to study the spectral behavior changes of peanut protein after combining with polyphenols,and analyzed the changes in its structure and properties by measuring the particle size and potential,soluble protein content,surface hydrophobicity,scanning electron microscopy,and differential scanning calorimetry.In this study,we explored the interaction types,protein structure characteristics and desensitization effects after that peanut protein interacted with polyphenols of different structures under different p H conditions.This work will provide more theoretical basis for the in-depth development of hypoallergenic peanut products.The main research contents and results of this paper were as follows:(1)The interaction between peanut protein and six kinds of polyphenols was preliminarily explored by using ultraviolet spectrum,endogenous fluorescence spectrum and synchronous fluorescence spectrum.The type of interaction force between peanut protein and trans-cinnamic acid,trans-ferulic acid,naringenin,genistein,seperately,was hydrophobic interaction,and the type of interaction force between peanut protein and caffeic acid,quercetin,seperately,was hydrogen bond and van der Waals force.After peanut protein combined with trans-cinnamic acid and quercetin,separately,the surrounding environment of the chromogenic amino acid residues in peanut protein both had increased hydrophobicity and reduced polarity,and the protein structure might be more compact.In the complexes forming by peanut protein with trans-ferulic acid,caffeic acid,naringenin and genistein,separately,the surrounding environment of chromogenic amino acid residues in peanut protein all showed increased hydrophilicity and polarity,and the protein structure might become looser.(2)The peanut sensitizing protein detection kit was used to determine the Ig E binding inhibition ability of peanut protein combined with different polyphenols.Results showed that peanut protein combined with trans-cinnamon acid,trans-ferulic acid,caffeic acid,naringenin,genistein,and quercetin,respectively,all exhibited fantastic Ig E binding inhibition effects in vitro,and that of the phenolic acid treatment groups was stronger.With the declined soluble protein content,the results of SDS-PAGE also showed that the combination of peanut protein with six kinds of polyphenols caused some protein bands to disappear or decrease,which changed more in phenolic acid treatment groups.(3)Multi-spectroscopy,soluble protein content,particle size and potential,surface hydrophobicity,differential scanning calorimetry,scanning electron microscopy,etc.were used to characterize the interaction between polyphenols of different structures and peanut protein and the changes in protein structure.The results showed that all six polyphenols interacted with peanut protein,and the structure of peanut protein changed.Hydrogen bonding,electrostatic interaction and hydrophobic force were involved in the interaction between polyphenols and peanut protein,and the site was closer to tryptophan residues.In general,the addition of phenolic acid had a greater impact on peanut protein than the addition of flavonoids.The phenolic acid treatment group showed a significantly reduced soluble protein content,a smaller average particle size,a looser structure,and gully appeared on the surface of the complexes.On the whole,the interaction between polyphenols and peanut protein caused the secondary structure to shift from α-helix and β-turn to β-sheet.(4)Molecular docking was used to simulate the interaction between peanut allergenic protein Ara h 1 and polyphenols.The results showed that 6 kinds of polyphenols bind at different sites in the structure of the protein.Among them,tyrosine was involved in the interactions between Ara h 1 and trans-ferulic acid,genistein,and quercetin,separately.Moreover,Ala-502 and Arg-503,which are two key amino acids related to desensitization,were involved in the interaction between trans-ferulic acid and Ara h 1.(5)Multi-spectroscopy,soluble protein content,particle size and potential,surface hydrophobicity,differential scanning calorimetry,scanning electron microscopy,Ig E binding inhibition rate,etc.were used to characterize the interaction of quercetin/trans-cinnamic acid and peanut protein,the changes in protein structure and allergenicity at different p H.The results were as follows: the type of interaction between quercetin/trans-cinnamic acid and peanut protein at different p H differed from each other,which had different effects on the structure and allergenicity of peanut protein.The interaction of polyphenols and peanut protein led to the decrease in protein sensitization,which might be related to the decrease of protein content,the reduction of hydrophobicity of some amino acid residues,the unfolding of structure,the decrease ofα-helix and/or β-turn content,and the looser protein structure.Changes in allergenicity might be the result of a combination of many factors.(6)Molecular simulation results showed that high p H value was not conducive to the stability of Ara h 1,and it might also be detrimental to the binding of polyphenols to Ara h 1.This might be caused by the local structure of the protein becoming more compact.The results of molecular docking showed that when trans-cinnamic acid at p H7 and quercetin/trans-cinnamic acid at p H 9 interacted with Ara h 1,key amino acids related to desensitization were involved. |
PDF Full Text Request