UCLA

Membrane proteins are a neglected, but important class of proteins throughout the biological world. They carry out critical roles in the cell due to their unique location, such as transport across a membrane, transduction of exterior signals, and interaction between discrete aqueous regions. Despite the importance of these proteins, understanding of how they fold has lagged far behind that of soluble proteins. One of the primary challenges to studying membrane protein folding is developing methods that interrogate folding in the native environment of the lipid bilayer. Our lab has developed a method for measuring membrane protein stability under native conditions using a secondary protein that preferentially binds the unfolded state, obviating the need for harsh denaturants. Employing this method with a multimeric polytopic membrane protein, we measured an extremely slow unfolding rate, demonstrating that α-helical membrane proteins can have high kinetic stability under non-denaturing conditions. Efforts were made to expand the steric trap method for single-molecule fluorescence measurements in lipid vesicles, but were ultimately stymied by the inability to preserve the trapped complexes for measurement. Our lab has also applied single-molecule techniques to membrane protein folding. We were able to map the energy landscape of a membrane protein in a lipid bilayer using forced unfolding driven by magnetic tweezers. Further advancements to this technique simplified the attachment chemistry to ready the protein for tweezing. These techniques can be applied to a wide array of membrane proteins in a broad spectrum of membrane environments.