背景:酶交联是一种可用于使用多酚修饰印加花生白蛋白(IPA)的方法,并提供理想的功能;然而,多酚结构对共轭性质的影响尚不清楚。在这项研究中,我们选择了四种具有不同数量酚羟基的多酚[对羟基苯甲酸(HBA),原儿茶酸(PCA),没食子酸(GA),和表没食子儿茶素没食子酸酯(EGCG)]通过酶交联共价修饰IPA,并探讨了IPA-多酚共轭物的结构-功能变化。
结果:十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和基质辅助激光解吸/电离飞行时间质谱(MALDI-TOF-MS)分析表明,漆酶成功地促进了IPA与多酚的共价交联,共轭程度的顺序为EGCG>GA>PCA>HBA,IPA-EGCG缀合物显示出最高的多酚结合当量(98.35gkg-1蛋白),以及游离氨基含量的显著降低,巯基,和酪氨酸组。漆酶对多酚的氧化形成与IPA的反应性基团共价交联的醌或半醌自由基,导致IPA的二级和三级结构发生重大变化,球形结构转变为致密的层状结构。IPA-EGCG偶联物的2,2-二苯基-1-吡啶酰肼(DPPH)自由基清除能力和乳化稳定性提高了近6倍和2.7倍,分别,与未修饰的IPA相比。
结论:这些数据表明,多酚羟基的数量越高,IPA-多酚共轭程度越高;此外,酶交联可以显著提高IPA的功能性能。©2024化学工业学会。
BACKGROUND: Enzymatic crosslinking is a method that can be used to modify Inca peanut albumin (IPA) using polyphenols, and provides desirable functionalities; however, the effect of polyphenol structures on conjugate properties is unclear. In this study, we selected four polyphenols with different numbers of phenolic hydroxyl groups [para-hydroxybenzoic acid (HBA), protocatechuic acid (PCA), gallic acid (GA), and epigallocatechin gallate (EGCG)] for covalent modification of IPA by enzymatic crosslinking, and explored the structure-function changes of the IPA-polyphenol conjugates.
RESULTS: Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis showed that laccase successfully promoted covalent crosslinking of IPA with polyphenols, with the order of degree of conjugation as EGCG > GA > PCA > HBA, the IPA-EGCG conjugate showed the highest polyphenol binding equivalents (98.35 g kg-1 protein), and a significant reduction in the content of free amino, sulfhydryl, and tyrosine group. The oxidation of polyphenols by laccase forms quinone or semiquinone radicals that are covalently crosslinked to the reactive groups of IPA, leading to significant changes in the secondary and tertiary structures of IPA, with spherical structures transforming into dense lamellar structures. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging ability and emulsification stability of IPA-EGCG conjugates improved by almost 6-fold and 2.7-fold, respectively, compared with those of unmodified IPA.
CONCLUSIONS: These data suggest that the higher the number of polyphenol hydroxyl groups, the higher the degree of IPA-polyphenol conjugation; additionally, enzymatic crosslinking can significantly improve the functional properties of IPA. © 2024 Society of Chemical Industry.