Parallel Temperature Replica-Exchange Molecular Dynamics\nSimulations Capture the Observed Impact of Stapling on Coiled-Coil\nConformational Stability

Macrocyclization or stapling is an\nimportant strategy for increasing\nthe conformational stability and target-binding affinity of peptides\nand proteins, especially in therapeutic contexts. Atomistic simulations\nof such stapled peptides and proteins could help rationalize existing\nexperimental data and provide predictive tools for the design of new\nstapled peptides and proteins. Standard approaches exist for incorporating\nnonstandard amino acids and functional groups into the force fields\nrequired for MD simulations and have been used in the context of stapling\nfor more than a decade. However, enthusiasm for their use has been\nlimited by their time-intensive nature and concerns about whether\nthe resulting simulations would be physically realistic. Here, we\nreport the development of force field parameters for two unnatural\ntriazole staples, which we have incorporated into implicit-solvent\nparallel temperature replica-exchange molecular dynamics simulations\nof several stapled coiled-coil variants and their nonstapled counterparts.\nWe used these simulations to calculate melting temperatures (<i>T</i><sub>m</sub>) of each variant along with the impact of\nstapling on the conformational stability of each variant relative\nto its nonstapled counterpart (ΔΔ<i>G</i>).\nTrends among these simulated <i>T</i><sub>m</sub> and ΔΔ<i>G</i> values closely match those observed in previous experiments,\nsuggesting that the parameters we developed for these staples are\nsufficiently realistic to be useful in predicting the impact of stapling\non the protein/peptide conformational stability in other contexts.