Influence of tea-derived polyphenols architecture on the structural, functional and allergenicity properties of β-lactoglobulin: A comparison of non-covalent and covalent interactions.

The relative effects of non-covalent complexes and covalent conjugates of β-lactoglobulin (β-LG) with three distinct polyphenolic structures isolated from teas (TDPs; EC, EGC, EGCG) on the properties of β-LG were studied comprehensively. Formation of β-LG-TDPs complexes/conjugates was determined through analysis of polyphenol content, SDS-PAGE, and thermodynamics. The molecular docking and dynamics studies revealed that the non-covalent complexation of TDPs involved hydrogen bonds and van der Waals interaction with binding of all TDPs inside the hydrophobic calyx structure of β-LG. The multi-spectroscopy analysis indicated that both types of interactions resulted in unfolding of β-LG, leading to decrease in β-sheets and increase in β-turns/random coils. Most importantly, covalent conjugation resulted in greater restructuring than complexation, particularly for covalent β-LG-EGCG. The observed changes in protein structure have resulted in improved functionality, including enhanced emulsifying and antioxidant properties and lower allergenicity of β-LG. Structure-dependent differences were observed, with covalently bound compounds displaying better functionality than those associated through non-covalent interactions. In particular, a significant improvement in the functionality of the covalent β-LG-EGCG conjugate has been observed. Compared to native β-LG, there was a 1.65-fold increase in emulsifying properties (ESI), a 3.25-fold increase in emulsifying activity index (EAI), a 9.25-fold increase in DPPH free radical scavenging ability, and a 6.62-fold increase in ABTS radical cation decolorization. Additionally, it had the least binding of IgG (74.10 ± 0.91%) and IgE (60.68 ± 0.69%). The results prove that the ability of TDPs to act as effective regulators depends on their chemical structures, especially the presence of phenolic hydroxyl groups and galloyl units. The research indicates that using covalent modification is the best way to improve protein performance and reduce allergenicity.