Project R-10251

Development of microdevices for screening membrane protein realtime structure in physiological conditions and investigation of multidrug resistance transporter LmrP (Research)

Membrane proteins reside in the plasma membrane, the thin fatty layer that wraps around cells. Their 3D molecular structure provides clues to their function and is a promising handle for compound screening. Yet, membrane protein structure is understudied because mimicking the complex asymmetry over the membrane in a controlled in vitro context is next to impossible. In my project I will develop novel microdevices and light microscopy assays that allow quantifying membrane protein structure in a physiologically relevant context. Practically, I will use the bacterial protein LmrP as a model system. LmrP and other members of its superfamily actively transport substrates over the membrane and play important roles in metabolite transport and different diseases in humans and in drug resistance development in bacteria. I will develop a fluorescence assay based on single-molecule Förster resonance energy transfer, a method that can measure the 1-10 nm distance between two fluorescent probes with sub-nanometer resolution, allowing fast and sensitive imaging of protein structure in real time. In parallel, I will develop different novel microdevices with fluorescent LmrP embedded in miniaturized plasma membrane arrays over which an electrochemical potential can be applied. By smFRET imaging of the microdevices, I will provide novel insights in the workings of LmrP and generate a solid methodological basis for superior screening assays targeting membrane proteins in general.

01 November 2019 - 31 October 2021