Abstract: Proteins operate within an aqueous environment that influences their folding, stability, and activity. Membrane proteins have the added complication of being embedded in lipid bilayers that play an equally critical, yet significantly less understood, role. Much like solvent conditions modulate enzyme kinetics and protein interactions in solution, the lipid composition of the membrane performs regulatory functions for membrane proteins, affecting their organization, conformational dynamics, and catalytic output. To investigate these membrane-dependent phenomena, we employ time-resolved fluorescence spectroscopy techniques like pulsed interleaved excitation fluorescence cross-correlation spectroscopy (PIE-FCCS) in model membranes and live cells. We couple these spectroscopic probes with liposome-based activity assays in vitro. These complementary approaches allow us to quantitatively assess how specific lipid species influence the assembly and function of receptor tyrosine kinases (RTKs).
In this talk, I will highlight two key areas where lipid–protein interactions govern RTK behavior. First, I will describe how the anionic lipid phosphatidylinositol 4,5-bisphosphate (PIP₂) promotes multimeric assembly of EGFR and EphA2 in the plasma membrane, driving the formation of functionally distinct signaling complexes, as revealed by PIE-FCCS in live cells. Second, I will present our recent in vitro findings showing that both PIP₂ and phosphatidylserine (PS) act as reversible, noncompetitive inhibitors of EGFR catalytic activity in a reconstituted liposome system, demonstrating direct biochemical regulation of kinase function by anionic lipids. Together, these results reveal that the lipid bilayer is not merely a scaffold but an active participant in membrane protein regulation. By drawing parallels to the well-established influence of solvent on soluble proteins, we hope to center the membrane environment as a dynamic and tunable component of cell signaling.