Determination of the protonation state of the Asp dyad: conventional molecular dynamics versus thermodynamic integration.

The protonation state of the Asp dyad is important as it can reveal enzymatic mechanisms, and the information this provides can be used in the development of drugs for proteins such as memapsin 2 (BACE-1), HIV-1 protease, and rennin. Conventional molecular dynamics (MD) simulations have been successfully used to determine the preferred protonation state of the Asp dyad. In the present work, we demonstrate that the results obtained from conventional MD simulations can be greatly influenced by the particular force field applied or the values used for control parameters. In principle, free-energy changes between possible protonation states can be used to determine the protonation state. We show that protonation state prediction by the thermodynamic integration (TI) method is insensitive to force field version or to the cutoff for calculating nonbonded interactions (a control parameter). In the present study, the protonation state of the Asp dyad predicted by TI calculations was the same regardless of the force field and cutoff value applied. Contrary to the intuition that conventional MD is more efficient, our results clearly show that the TI method is actually more efficient and more reliable for determining the protonation state of the Asp dyad.