Single-molecule force spectroscopy reveals a mechanically stable protein fold and the rational tuning of its mechanical stability.
It is recognized that shear topology of two directly connected force-bearing terminal beta-strands is a common feature among the vast majority of mechanically stable proteins known so far. However, these proteins belong to only two distinct protein folds, Ig-like beta sandwich fold and beta-grasp fold, significantly hindering delineating molecular determinants of mechanical stability and rational tuning of mechanical properties. Here we combine single-molecule atomic force microscopy and steered molecular dynamics simulation to reveal that the de novo designed Top7 fold [Kuhlman B, Dantas G, Ireton GC, Varani G, Stoddard BL, Baker D (2003) Science 302:1364-1368] represents a mechanically stable protein fold that is distinct from Ig-like beta sandwich and beta-grasp folds. Although the two force-bearing beta strands of Top7 are not directly connected, Top7 displays significant mechanical stability, demonstrating that the direct connectivity of force-bearing beta strands in shear topology is not mandatory for mechanical stability. This finding broadens our understanding of the design of mechanically stable proteins and expands the protein fold space where mechanically stable proteins can be screened. Moreover, our results revealed a substructure-sliding mechanism for the mechanical unfolding of Top7 and the existence of two possible unfolding pathways with different height of energy barrier. Such insights enabled us to rationally tune the mechanical stability of Top7 by redesigning its mechanical unfolding pathway. Our study demonstrates that computational biology methods (including de novo design) offer great potential for designing proteins of defined topology to achieve significant and tunable mechanical properties in a rational and systematic fashion.
Sharma
Deepak
D
Department of Chemistry, University of British Columbia, Vancouver, BC, Canada.
Perisic
Ognjen
O
Peng
Qing
Q
Cao
Yi
Y
Lam
Canaan
C
Lu
Hui
H
Li
Hongbin
H
D003198:Computer Simulation; D019563:Mechanics; D008958:Models, Molecular; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D013057:Spectrum Analysis
D011506:Proteins