Quantifying Protein and Lipid Accumulation at Sites of Membrane Curvature

Protein and lipid organization is tightly regulated on the cellular plasma membrane. This process is driven by intermolecular interactions and leads to nanoscale domain formation where specific biochemical reactions can occur. One feature of membranes that plays a less obvious role is the shape of the lipid membrane. This aspect is difficult to measure in living cells, making biochemical assays a useful tool for quantifying curvature dependent sorting. In our work, we use nanoscale patterning to create substrates that introduce curvature into supported lipid bilayers with the goal of separately assessing the role of shape and chemical composition on the organization of proteins and lipids. To create substrates, nanoparticles are deposited onto glass surfaces prior to lipid bilayer formation. Confocal and total internal reflection fluorescence microscopies, combined with colocalization, single molecule imaging and tracking methods, are used to assess protein and lipid accumulation at regions of curvature. With these methods, we demonstrate that a peripheral membrane binding protein, C-reactive protein, binds highly curved membranes and this interaction depends on the state of the protein. Other proteins, like Cholera Toxin B, avoid curvature. Our membrane curvature assay would be highly useful for assessing the interplay between curved membranes and lipid/protein accumulation.