Assessment of the Topology and Oligomerisation States of Coiled Coils Using Metadynamics with Conformational Restraints

Coiled-coil proteins provide an excellent scaffold for multi-state de novo protein design due to their established sequence-to-structure relationships and ability to switch conformations in response to external stimuli, such as changes in pH or temperature. However, the computational design of multi-state coiled-coil protein assemblies is challenging, as it requires accurate estimates of the free energy differences between multiple alternative coiled-coil conformations. Here, we demonstrate how this challenge can be tackled using metadynamics simulations with orientational, positional and conformational restraints. We show that, even for subtle sequence variations, our protocol can predict the preferred topology of coiled-coil dimers and trimers, the preferred oligomerisation states of coiled-coil dimers, trimers, and tetramers, as well as the switching behaviour of a pH-dependent multi-state system. Our approach provides a method for predicting the stability of coiled-coil designs and offers a new framework for computing binding free energies in protein-protein and multi-protein complexes.