Probing the Dimerization of Transmembrane Helices in Eukaryotic and Lipid Bilayer Membranes

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Lateral interactions between hydrophobic transmembrane (TM) helices in cellular membranes underlie the folding of multi-span membrane proteins and signal transduction mediated by receptor tyrosine kinases (RTKs). Quantitative measurements of dimerization energetics in membranes, required for deciphering these processes, are challenging to perform due to many limitations of current experimental methodologies. The PI will develop novel versatile methodologies for studies of TM helix dimerization in biological membranes. The PI will carry out measurements in eukaryotic membranes and introduce a FRET-Western assay that probes both the stability and the structure of membrane proteins in the context of the native cellular membrane. The PI will also develop surface-supported lipid bilayers that mimic the plasma membrane by integrating asymmetry in lipid composition and uni-directional orientation of the proteins. This work will reveal the role of the complex plasma membrane environment in membrane protein interactions. The FRET-Western will allow the identification of mutations in TM domains that alter TM dimer stability or TM dimer structure, thus providing new insights into the structural determinants of protein dimerization in membranes. The new methods will be a significant improvement over the methodology that is currently available to study association between membrane proteins. Thus, the project will provide the broad scientific community with better tools to study membrane proteins folding, structure, and function. Ultimately, improvement in methods will lead to discoveries that elucidate the role of membrane protein interactions in cell life and human disease. Furthermore, this project will support the funding of graduate students and enhance the educational experience of undergraduate students from diverse backgrounds. This project is jointly supported by Molecular Biophysics in the Division of Molecular and Cellular Biosciences in the Directorate for Biological Sciences and the Experimental Physical Chemistry Program in the Division of Chemistry in the Mathematical and Physical Sciences Directorate.