Research & Teaching Faculty

Default Header Image

Single Molecule Force Spectroscopy Reveals Critical Roles of Hydrophobic Core Packing in Determining the Mechanical Stability of Protein GB1

TitleSingle Molecule Force Spectroscopy Reveals Critical Roles of Hydrophobic Core Packing in Determining the Mechanical Stability of Protein GB1
Publication TypeJournal Article
Year of Publication2012
AuthorsBu, T, Wang, H-CEileen, Li, H
JournalLANGMUIR
Volume28
Pagination12319-12325
Date PublishedAUG 21
ISSN0743-7463
Abstract

Understanding molecular determinants of protein mechanical stability is important not only for elucidating how elastomeric proteins are designed and functioning in biological systems but also for designing protein building blocks with defined nanomechanical properties for constructing novel biomaterials. GB1 is a small alpha/beta protein and exhibits significant mechanical stability. It is thought that the shear topology of GB1 plays an important role in determining its mechanical stability. Here, we combine single molecule atomic force microscopy and protein engineering techniques to investigate the effect of side chain reduction and hydrophobic core packing on the mechanical stability of GB1. We engineered seven point mutants and carried out mechanical phi-value analysis of the mechanical unfolding of GB1. We found that three mutations, which are across the surfaces of two subdomains that are to be sheared by the applied stretching force, in the hydrophobic core (F30L, Y45L, and F52L) result in significant decrease in mechanical unfolding force of GB1. The mechanical unfolding force of these mutants drop by 50-90 pN compared with wild-type GB1, which unfolds at around 180 pN at a pulling speed of 400 nm/s. These results indicate that hydrophobic core packing plays an important role in determining the mechanical stability of GB1 and suggest that optimizing hydrophobic interactions across the surfaces that are to be sheared will likely be an efficient method to enhance the mechanical stability of GB1 and GB1 homologues.

DOI10.1021/la301940g