Mechanistic Insights into the interaction between aldehyde aroma compounds and β-Casein through Multi-Spectroscopy and molecular dynamics

The interaction between proteins and aroma compounds significantly impacts cheese flavor retention during processing. However, it is still unknown how cheese proteins and the aldehyde aroma compounds (AACs) interact. This study aims to clarify the interaction mechanisms between the AACs (benzaldehyde, 2-methylpropanal, 2-methylbutanal and 3-methylbutanal) and β-casein (β-CN) using SPME-GC/MS, multi-spectroscopy techniques, and molecular dynamics simulations. The results reveal notable variations in the binding abilities of the four AACs and β-CN, with the strongest binding observed for 3-methylbutanal. Specifically, the binding affinity (Ka) values between β-casein and benzaldehyde, 2-methylpropanal, 2-methylbutanal, and 3-methylbutanal are 2.26 × 10³, 1.78 × 10³, 2.03 × 10³, and 2.52 × 10³ M^-1, respectively, indicating moderate binding affinity. Additionally, the quenching rate constants (Kq) for interactions with these compounds are 2.57 × 10¹¹, 2.92 × 10¹¹, 3.74 × 10¹¹, and 4.81 × 10¹¹ M^-1s^-1, significantly exceeding the collisional quenching limit, suggesting specific interactions. The interactions between the four AACs and β-CN occur through irreversible covalent bonding, primarily involving hydrogen bonds and hydrophobic interactions. The quenching mechanism of β-CN and the four AACs is static, which leads to changes in the secondary structure and microenvironment of β-CN. Molecular docking and dynamics simulations confirm that hydrogen bonds and hydrophobic interactions are the key driving forces for the binding of β-CN with the four AACs, and contribute to the stability of the composite system.