[关键词]
[摘要]
本文将乳糖通过糖基化反应引入到大豆分离蛋白(SPI)上制备大豆分离蛋白-乳糖复合物,采用间接竞争ELISA法测定不同温度、不同质量比、不同反应时间下大豆分离蛋白-乳糖复合物中β-伴大豆球蛋白的抗原性变化,并对糖基化产物进行了结构特性的研究。结果表明,糖基化能有效降低β-伴大豆球蛋白的抗原性,其抗原性从93.79%降到37.75%。糖基化改性后,大豆蛋白中游离氨基含量降低,在反应60 h时,游离氨基含量下降最大;傅里叶红外光谱结果表明,与原大豆分离蛋白相比,大豆分离蛋白-乳糖糖基化产物的α-螺旋、β-转角、无规卷曲的含量下降,而β-折叠的含量增加;SDS-PAGE电泳及PAS染色结果表明,随着糖基化反应的程度增加,SPI谱带逐渐的减弱,说明SPI与乳糖分子发生了共价连接。
[Key word]
[Abstract]
Lactose was introduced into soybean protein isolate (SPI) via glycosylation to produce an SPI-lactose conjugates, under different temperatures, protein and sugar mass ratios, as well as reaction durations. Subsequent changes in the antigenicity of β-conglycinin in the SPI-lactose conjugates was estimated by indirect-competition enzyme-linked immunosorbent assay (ic-ELISA). The structural properties of the SPI-lactose conjugates were also studied. The results indicated that glycosylation reduced the antigenicity of β-conglycinin from 93.79% (control) to 37.75% (conjugates). Compared to SPI, the content of free amino groups in the complex also decreased after glycosylation. The reduction was highest at 60 hours after the of the reaction. Fourier transformed infrared spectroscopy (FT-IR) showed that the quantity of α-helices, β-turns, and random coil structures were lower in the SPI-lactose glycosylation product than that in the SPI, while that of β-sheet increased. Results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and periodic acid-Schiff (PAS) staining showed that the SPI band gradually became weaker with the progress in glycosylation reaction, indicating that the glycosylation reaction occurred between SPI and lactose molecules.
[中图分类号]
[基金项目]
国家自然科学基金项目(31201293;21176058;31171790);国家863科技计划项目(2013AA102208-5);河南省教育厅科学技术研究重点项目(14B550013)