Low force unfolding of a single-domain protein by parallel pathways

TL;DR

This study demonstrates that upward curvature in the logarithm of unfolding rates versus force indicates multiple unfolding pathways in single-domain proteins, supported by simulations showing pathway switches at different forces.

Contribution

The paper reveals that parallel unfolding pathways can be identified from force-dependent unfolding kinetics, highlighting structural changes in transition states with increasing force.

Findings

Upward curvature in $ extlog k_u(f)$ indicates multiple pathways.

Structural transition states change with force.

Parallel unfolding pathways are detectable from kinetic data.

Abstract

Deviations from linearity in the dependence of the logarithm of protein unfolding rates, $\log k_u(f)$, as a function of mechanical force, $f$, measurable in single molecule experiments, can arise for many reasons. In particular, upward curvature in $\log k_u(f)$ as a function of $f$ implies that the underlying energy landscape must be multidimensional with the possibility that unfolding ensues by parallel pathways. Here, simulations using the SOP-SC model of a wild type $\beta$-sandwich protein and several mutants, with immunoglobulin folds, show upward curvature in the unfolding kinetics. There are substantial changes in the structures of the transition state ensembles as force is increased, signaling a switch in the unfolding pathways. Our results, when combined with previous theoretical and experimental studies, show that parallel unfolding of structurally unrelated single domain proteins can be determined from the dependence of $\log k_u(f)$ as a function of force (or $\log k_u[C]$ where $[C]$ is the denaturant concentration).