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Engineering cushioned bilayers for enhanced biophysical characterisation of membrane proteins
Secondary Supervisor(s): Dr Naomi Pollock, Dr. Manish Singh Kushwah
University of Registration: Aston University
BBSRC Research Themes:
Project Outline
Integral membrane proteins (IMPs) control vital cellular processes and are the target of most modern therapeutics. Despite this, characterisation of these proteins and development of therapeutics for their disease states remains technically challenging. Traditionally IMPs were extracted from bilayers using detergents which disrupted their native state and function. The importance of lipid bilayers to maintain membrane proteins in their functional states and complexes is an emerging area of research which will address the current limitations in the field. This project will meet that challenge by developing polymer-cushioned supported bilayers with tunable heights, creating versatile model membranes for studying IMPs in environments that closely mimic the cell.
The project will combine polymer chemistry, membrane biochemistry, and advanced biophysical techniques to engineer bilayers supported by substrate bound polymer cushions of varying thickness. Lipid mobility in these bilayers will be characterised using fluorescence recovery after photobleaching (FRAP) allowing lipid composition and polymer properties to be probed in detail. Protein insertion into and diffusion within the bilayers will be measured using dynamic mass photometry (DMP). This cutting-edge label-free method simultaneously measures protein mass and diffusion at the single-molecule level. In this way we will develop bilayers tuned to the requirements of model proteins including aquaporins, GPCRs, and ATP-binding cassette (ABC) transporters. The proteins will be reconstituted into bilayers from native nanodiscs, enabling direct analysis of insertion, oligomerization, and lateral diffusion.
By linking polymer design to bilayer properties and protein behaviour, this project will provide fundamental insights into the co-dependence of membranes and IMPs. This will establish new opportunities for drug and antibody screening against this critical but underexplored class of therapeutic targets.