Elucidating the interaction between pyrazine flavor compounds and coffee proteins: Insights from multiscale structural and molecular dynamics simulations

For roasted coffee, its distinctive aroma is imparted by pyrazine compounds. Understanding how they interact with coffee proteins (CP) is crucial for controlling coffee flavor retention. This study demonstrated that the type and number of alkyl groups determine the binding strength between pyrazines and CP. At 2.00 and 10.00 mg/L of the pyrazine compounds, the binding affinity of CP for the pyrazines followed the order: 2,3,5-trimethylpyrazine (TRP) > 2-ethyl-3,5-dimethylpyrazine (EDP) > 2-ethyl-3-methylpyrazine (EMP) > 2-ethylpyrazine (EP). Multi-spectroscopic analysis indicated that pyrazines quenched the fluorescence of CP through both dynamic and static quenching processes, primarily via binding near Tyr and Trp residues. Hydrophobic interactions primarily governed the binding of TRP and EDP to CP, whereas van der Waals forces and hydrogen bonding were dominant for EP and EMP. Pyrazine binding induced the aggregation of CP, with TRP exhibiting the most pronounced effect on secondary structure, including a decrease in α-helix content and an increase in β-sheet content indicative of partial protein unfolding. Molecular simulation confirmed that TRP exhibited the smallest highest occupied molecular orbital-lowest unoccupied molecular orbital (HOMO-LUMO) energy gap (5.338505 eV) with CP, indicating a greater propensity for interaction. The study revealed the relationship between pyrazine structure and its binding capacity to CP, providing a theoretical basis for flavor regulation during coffee processing and storage.