碱性氨基酸对β-乳球蛋白结构和致敏性的影响

Effect of basic amino acids on structure and allergenicity ofβ-lactoglobulin

碱性氨基酸是一种自身带正电荷的小分子,可与蛋白质上的负电残基发生静电吸引,也可与蛋白侧链上某些基团构成空间位阻,进而导致蛋白质结构的改变。β-乳球蛋白(β-lactoglobulin,β-Lg)可引发过敏反应,通过不同碱性氨基酸对β-Lg进行处理,研究其结构和致敏性的变化规律。以L-精氨酸(L-Arg)、L-赖氨酸(L-Lys)和L-组氨酸(L-His)单独或混合处理的β-Lg为研究对象,通过电泳、圆二色谱、光谱等技术分析其结构的变化;采用酶联免疫吸附法、细胞实验等方法评价其抗氧化活性、免疫球蛋白E(immune globulin E,IgE)/免疫球蛋白G(immune globulin G,IgG)结合能力和KU812细胞释放组胺和白介素-6的能力。结果表明,L-Arg、L-Lys和L-His处理β-Lg后,改变了β-Lg的二级结构、紫外吸收强度、内源荧光强度和表面疏水性,同时增加其ABTS阳离子自由基清除能力、降低IgE/IgG结合能力以及KU812细胞中组胺和白细胞介素-6的分泌能力。因此,L-Arg、L-Lys和L-His破坏β-Lg的构象过敏表位,降低其致敏性,顺序依次为L-His、L-Lys和L-Arg。研究结果为低致敏性牛乳制品的研发提供重要的理论指导。

Basic amino acids are small molecules with positive charges,which can generate electrostatic attraction with negatively charged residues on proteins and steric hindering with certain groups on protein side chains,thus leading to changes in protein structure.β-lactoglobulin(β-Lg)causes allergic reactions.The changes in the structure and allergenicity ofβ-Lg were studied by treatingβ-Lg with different basic amino acids.Takingβ-Lg treated with L-Arg,L-Lys,and L-His alone or in combination as research objects,the changes in the structure of the treatedβ-Lg were analyzed by electrophoresis,circular dichroism,and spectroscopy.The antioxidant activity,IgE/IgG binding capacity,and the secretory ability of histamine and interleukin-6 in KU812 cells were evaluated by enzyme-linked immunosorbent assay and cell assay.Results showed that afterβ-Lg was treated with L-Arg,L-Lys,and L-His,its secondary structure,UV absorption intensity,fluorescence intensity,and surface hydrophobicity were changed,the ABTS radical scavenging ability increased,and the IgE/IgG binding ability,the secretory ability of histamine and interleukin-6 in KU812 cells decreased.Thus,L-Arg,L-Lys,and L-His reduce the allergenicity ofβ-Lg by disrupting the conformational epitopes,in order of L-His,L-Lys,and L-Arg.The results provide important theoretical guidance for the development of hypoallergenic milk products.