Implementing and Assessing an Alchemical Method for Calculating Protein-Protein Binding Free Energy.

Protein-protein binding is fundamental to most biological processes. It is important to be able to use computation to accurately estimate the change in protein-protein binding free energy due to mutations in order to answer biological questions that would be experimentally challenging, laborious, or time-consuming. Although nonrigorous free-energy methods are faster, rigorous alchemical molecular dynamics-based methods are considerably more accurate and are becoming more feasible with the advancement of computer hardware and molecular simulation software. Even with sufficient computational resources, there are still major challenges to using alchemical free-energy methods for protein-protein complexes, such as generating hybrid structures and topologies, maintaining a neutral net charge of the system when there is a charge-changing mutation, and setting up the simulation. In the current study, we have used the package to generate hybrid structures and topologies, and a double-system/single-box approach to maintain the net charge of the system. To test the approach, we predicted relative binding affinities for two protein-protein complexes using a nonequilibrium alchemical method based on the Crooks fluctuation theorem and compared the results with experimental values. The method correctly identified stabilizing from destabilizing mutations for a small protein-protein complex, and a larger, more challenging antibody complex. Strong correlations were obtained between predicted and experimental relative binding affinities for both protein-protein systems.