title: Single-molecule chemo-mechanical unfolding reveals multiple transition state barriers in a small single-domain protein.
abstract: A fundamental question in protein folding is whether proteins fold through one or multiple trajectories. While most experiments indicate a single pathway, simulations suggest proteins can fold through many parallel pathways. Here, we use a combination of chemical denaturant, mechanical force and site-directed mutations to demonstrate the presence of multiple unfolding pathways in a simple, two-state folding protein. We show that these multiple pathways have structurally different transition states, and that seemingly small changes in protein sequence and environment can strongly modulate the flux between the pathways. These results suggest that in vivo, the crowded cellular environment could strongly influence the mechanisms of protein folding and unfolding. Our study resolves the apparent dichotomy between experimental and theoretical studies, and highlights the advantage of using a multipronged approach to reveal the complexities of a protein's free-energy landscape.
authors.affiliation: Institute for Quantitative Biosciences (QB3), University of California-Berkeley, Berkeley, California 94720-3220, USA.
authors.forename: Emily J
authors.initials: EJ
authors.lastname: Guinn
authors.affiliation: Institute for Quantitative Biosciences (QB3), University of California-Berkeley, Berkeley, California 94720-3220, USA.
authors.forename: Bharat
authors.initials: B
authors.lastname: Jagannathan
authors.affiliation: 1] Institute for Quantitative Biosciences (QB3), University of California-Berkeley, Berkeley, California 94720-3220, USA [2] Department of Molecular and Cell Biology, University of California-Berkeley, Berkeley, California 94720-3220, USA.
authors.forename: Susan
authors.initials: S
authors.lastname: Marqusee
mesh_terms: D000818:Animals; D002645:Chickens; D016297:Mutagenesis, Site-Directed; D052898:Optical Tweezers; D011489:Protein Denaturation; D058767:Protein Unfolding; D018909:src Homology Domains; D019061:src-Family Kinases / Q000737:chemistry*
chemical_list: D019061:src-Family Kinases