Probing the Effect of the Ribosome on the Protein Folding Pathway using Single-Molecule Chemo-Mechanical Folding

Abstract

In vivo, proteins function in a complex environment where they are subject to many stresses that can modulate the protein's energy landscape. Using a combination of force and chemical denaturant (chemo-mechanical unfolding), we have demonstrated that a simple, two-state folding protein can fold through multiple parallel pathways and that seemingly small changes in force, denaturant concentration and protein sequence can strongly modulate the flux between the pathways. This result suggests that in vivo, the heterogeneous crowded cellular environment could strongly influence the mechanisms of protein folding and unfolding. Another perturbant that could affect the folding pathway in vivo is the ribosome since many proteins fold while they are being translated. To explore how the ribosome affects the folding mechanism, we use chemo-mechanical unfolding to characterize folding pathways both on and off the ribosome.