Multifaceted folding–unfolding landscape of the TrpZip2 β-hairpin and the role of external sub-piconewton mechanical tensions

Abstract

Proteins can experience uneven tensions of the order of tens of piconewtons when exposed to different solvent environment due to the thermal motion of the solvent. It is also true that biomolecules, especially proteins, are subjected to a variety of mechanical tensions generated by several factors, including mechanically assisted translocation and pressure gradients within living systems. Here, we use metadynamics simulations to revisit the folding–unfolding of the TrpZip2 β-hairpin and redefine it from the perspective of an external force of a sub-piconewton magnitude acting on the ends of the hairpin. The chosen forces, while preserving the morphology of the β-hairpin chain when it is pulled, are capable of influencing the conformational behavior of the chain during folding and unfolding. Our investigations confirm that the TrpZip2 β-hairpin exhibits a zipper (zip-out) mechanism for folding–unfolding in both mechanically unbiased and biased (with a 30 pN end force) situations. However, it is important to note that they present marked differences in their folding and unfolding paths, with the mechanically biased system capable of becoming trapped in various intermediate states. Both unbiased and biased scenarios of the hairpin indicate that the hairpin turn is highly stable during the folding–unfolding event and initiates folding. More importantly we confirm that the existing heterogeneity in the TrpZip2 β-hairpin folding–unfolding is a consequence of the wide range of conformations observed, owing to the different trapped intermediates caused by the uneven forces it may experience in solution.