Molecular Dynamics Insight Into Theanine-Induced Conformational Stabilization and Gelation of Walnut Protein Isolate

Walnut isolate protein (WPI) possesses high nutritional value and functional properties. However, its limited gelation ability due to weak intermolecular interactions constrains its application in the food industry. In the present study, theanine-mediated structural modulation strategy was employed to elucidate its multiscale mechanism for enhancing the gelation behavior of walnut isolate protein. Results indicated that the addition of theanine improved gel performance markedly, increasing water holding capacity from 56.7% to 82.2% and enhancing gel hardness by approximately 165%, indicating the formation of a denser and more stable network structure. Structural analyses revealed that theanine induced a conformational transition of WPI from α-helical to β-sheet structures and exposed more hydrophobic residues, thereby facilitating stronger intermolecular associations within the gel matrix. Molecular docking and molecular dynamics simulations further demonstrated that theanine formed stable protein-ligand complexes mainly through hydrogen bonding and hydrophobic interactions and WPI residues. Among the major components, theanine showed the strongest binding energy (-62.48 kcal/mol) with 11S globulin, thereby promoting the ordered aggregation and network formation of the protein. In addition, theanine significantly suppressed the intrinsic conformational fluctuations of 2S albumin, thereby enhancing the structural stability of the complex. This study revealed the molecular basis of theanine to enhance the gelation of the key components of WPI and provided a theoretical basis and technical approach for the development of high-value utilization of highly stable WPI gel foods.