Prediction of allosterically acting mutations using MD simulations and Rosetta

Abstract

Computationally modeling how mutations affect protein-protein binding not only helps uncover the biophysics of protein interfaces, but also enables the redesign and optimization of protein interactions. Traditional high-throughput methods for estimating binding free energy changes are currently limited to mutations directly at the interface due to difficulties in accurately modeling how long-distance mutations propagate their effects through the protein structure. However, the modeling and design of such mutations is of substantial interest as it allows for greater control and flexibility in protein design applications. We have developed a method that combines high-throughput Rosetta-based side-chain optimization with conformational sampling using classical molecular dynamics simulations, finding significant improvements in our ability to accurately predict long-distance mutational perturbations to protein binding. Our approach uses an analytical framework grounded in alchemical free energy calculations while enabling exploration of a vastly larger sequence space. When comparing to experimental data, we find that our method can predict internal long-distance mutational perturbations with a level of accuracy similar to that of traditional methods in predicting the effects of mutations at the protein-protein interface. This work represents a new and generalizable approach to optimize protein free energy landscapes for desired biological functions. Author SummaryProtein-protein interactions are vital to almost all biological processes, and therefore the ability to accurately and efficiently predict how mutations alter protein binding has far-reaching applications in protein analysis and design. Current approaches to predict such mutational free energy changes are limited to mutations directly at the interaction interface. Much research has underlined the prevalence of allosteric protein regulation in biological processes, indicating the importance of understanding and predicting the effects of protein perturbations which act over long distances. In this work we develop a novel method based on molecular dynamics simulations, the Rosetta macromolecular modeling suite, and an analytical framework from alchemical free energy calculations which can predict the effects of long-distance mutations with levels of accuracy rivaling state of the art interface-specific methods. We hope that our method will serve as a novel framework for high throughput mutational analysis and therefore benefit future protein design efforts.