(442d) Dynamic Interactions between Intrinsically Disordered Proteins and Curved Membrane Surfaces

Lipid bilayers are the primary structural component of biological membranes and are inherently resistant to deformation, exhibiting elastic properties when they are bent. However, membrane curvature is ubiquitous in the cell as curved structures, such as endocytic and synaptic vesicles, are essential for proper cellular physiology. These structures are the result protein-lipid interactions that are facilitated by proteins that sense membrane curvature. The mechanisms of curvature sensing often rely on specific protein structural features. However, our research has recently revealed that Intrinsically Disordered Proteins (IDPs) are also able to sense membrane curvature. In the context of the structure-function paradigm, this finding is somewhat paradoxical as IDPs lack substantial secondary structure but are still able to sense membrane curvature. This curvature sensitivity arises from repulsive steric and electrostatic interactions with the membrane surface. These types of IDPs typically have an N-terminal region that has biochemical affinity for the membrane, while the C-terminus remains disordered and externally tethered. Therefore, highly curved surfaces minimize the free energy of the protein when it is in the membrane-bound state. Our previous work has demonstrated that endocytic IDPs are sensitive to membrane curvature. However, this previous work, as well as other existing work, examines curvature sensitivity at thermodynamic equilibrium, where the relative partitioning of proteins is monitored in regions of various curvature. While it can take minutes for proteins to reach thermodynamic equilibrium, the physiologically relevant timescale of cellular processes like endocytosis is on the order of milliseconds to seconds. This disparity in timescales makes it apparent that thermodynamic equilibrium measurements alone likely fail to capture dynamic curvature sensing events that may be essential to biological processes. Therefore, it is likely much more valuable to measure the impact of membrane curvature on kinetic variables such as rate constants, rather than measuring the impact of curvature on equilibrium partitioning. However, to our knowledge, no literature exists that examines the time-dependent evolution of equilibrium binding processes on curved membranes, likely owing to the difficulty associated with obtaining these measurements. Therefore, the goal of our work is to develop the experimental tools necessary for visualization and analysis of dynamic protein-lipid interactions.