Multiscale elucidation of ergothioneine-soy protein interactions: From molecular docking and binding thermodynamics to conformational rearrangement and functional augmentation

This study systematically investigated the interaction mechanism between ergothioneine (EGT) and soy protein isolate (SPI) and its effects on protein structure and functionality by integrating multi-spectroscopic techniques with computational simulation. The results demonstrated that EGT quenches the intrinsic fluorescence of SPI through a static quenching mechanism, with a binding constant (K A ) of 2.242 × 10 5  L/mol and approximately one binding site per molecule at 298 K. Thermodynamic parameters (ΔH = −35.01 kJ/mol, ΔS = −41.48 J/(mol·K)) indicated that the spontaneous interaction was primarily driven by hydrogen bonding and van der Waals forces. Molecular docking confirmed these findings, revealing that EGT binds specifically within the hydrophobic grooves of SPI subunits (7S and 11S) via hydrogen bonds and hydrophobic interactions. The binding induced significant conformational changes in SPI, evidenced by redshifted synchronous fluorescence peaks (Tyr: 282 → 284 nm; Trp: 289 → 296 nm) and alterations in FTIR amide bands, which collectively enhanced molecular flexibility and surface hydrophilicity. These structural modifications directly improved the functional properties of SPI, including significant reductions in particle size and turbidity, along with marked enhancements in solubility, emulsifying activity index (EAI), and emulsion stability index (ESI). Furthermore, the antioxidant capacity of the complexes-assessed by DPPH and ABTS radical scavenging assays and reducing power analysis-exhibited a concentration-dependent increase. This work provides a comprehensive elucidation of the EGT-SPI interaction mechanism and offers a solid theoretical foundation for developing SPI-based delivery systems to enhance the stability and functionality of bioactive compounds in functional foods.