Single-Molecule Force Spectroscopy of Transmembrane beta-Barrel Proteins

Single-molecule force spectroscopy (SMFS) has been widely applied to study the mechanical unfolding and folding of transmembrane proteins. Here, we review the recent progress in characterizing bacterial and human transmembrane beta-barrel proteins by SMFS. First, we describe the mechanical unfolding of transmembrane beta-barrels, which follows a general mechanism dictated by the sequential unfolding and extraction of individual beta-strands and beta-hairpins from membranes. Upon force relaxation, the unfolded polypeptide can insert stepwise into the membrane as single beta-strands or beta-hairpins to fold as the native beta-barrel. The refolding can be followed at a high spatial and temporal resolution, showing that small beta-barrels are able to fold without assistance, whereas large and complex beta-barrels require chaperone cofactors. Applied in the dynamic mode, SMFS can quantify the kinetic and mechanical properties of single beta-hairpins and reveal complementary insight into the membrane protein structure and function relationship. We further outline the challenges that SMFS experiments must overcome for a comprehensive understanding of the folding and function of transmembrane beta-barrel proteins.