Topological frustration leading to backtracking in a coupled folding-binding process

Intrinsically disordered proteins (IDPs) are abundant in all species. Their discovery challenges the traditional "sequence-structure-function" paradigm of protein science because IDPs play important roles in various biological processes without preformed folded structures. Bioinformatic analysis reveals that the intrinsically conformational disorder of IDPs as well as their conformational transition upon binding to their targets is encoded by their amino acid sequences. The rRNase domain of colicin E3 and the immunity protein Im3 are a pair of proteins involved in bacterial survival. While the N-terminal segment and the central segment of E3 make comparable intermolecular contacts with Im3 in the bound state, binding of E3 with Im3 is dominantly triggered by the central segment of E3. In this work, to further investigate the binding mechanism of disordered E3 with Im3, we performed systematic free energy and transition path analysis through coarse-grained molecular dynamics simulations. We observed backtracking of the N-terminal segment of E3 in the binding process, whose occurrence depends on salt concentration. Conformational analysis revealed that initial binding of the N-terminal segment of E3 to Im3 usually leads to misorientation of a central hairpin of E3 on Im3, which generates topological frustration and results in backtracking of the N-terminal segment. Our results not only provide deeper mechanistic insights into the coupled folding-binding process of the E3/Im3 complex, but also suggest that topological frustration could be present in the coupled folding-binding process of IDPs and play an important role in regulating the binding transition pathways.