Integral membrane proteins are important components of many biological processes and comprise ~30 % of all cellular proteins. Surprisingly little is known about how these proteins fold and assemble to their final structure. This information could potentially be obtained through biophysical approaches (structural biology, enzyme kinetics, fluorescence methods etc.), but this has proven challenging for membrane proteins. Suitable in vitro systems are difficult to establish because simple artificial solvents cannot replicate the complex heterogenous lipid environment that membrane proteins normally experience.    

Bacteriorhodopsin
My recent work has focused on the light-activated proton pump bacteriorhodpsin (structure shown at top right). bR can be reversibly unfolded under equilibrium conditions by using mixed micelles composed of stabilizing and denaturing detergents. Through a combined kinetic and thermodynamic analysis, we showed that the free energy of unfolding was linear with denaturant concentration, and that folding could be considered as a simple two-state process. This allowed us to apply the analytical toolkit developed for soluble protein folding studies to a membrane protein for the first time. This revealed that the folding transition state is close to the unfolded state and that the spontaneous unfolding of bR in the absence of denaturant is very rare. A paper on this is available here. Further work (with Paula Booth) will continue to investigate the folding of bR. 

Previously, I studied the reconsitutiton of the multidrug transporter EmrE. The relatively simple sequence and structure of EmrE - each monomer comprises 110 amino acids arranged in four transmembrane helices (shown on the right) - make it a promising candidate for understanding the basic apparatus of multidrug transport. We demonstrated that EmrE could be reconstituted into artificial lipid vesicles. Reconstitution was efficient and the protein was able to bind substrate and transport it across the lipid bilayer against a proton electrochemical gradient. Altering the lipid composition of the vesicles influenced the behaviour of EmrE in a predictable manner; when the lipid lateral pressure was increased by introducing non-lamellar lipids, substrate transport was dramatically impaired. The details can be found here. Work with EmrE is ongoing within the Booth group.