In vitro methods for measuring lipid binding, transfer and competitive displacement reactions by peripheral membrane proteins.

Biological membranes represent versatile platforms that reduce the dimensionality of biochemical reactions involved in signal transduction. In so doing, membranes effectively concentrate proteins and other molecules to ensure that biochemical reactions can take place even in the face of relatively weak interactions between binding partners. Key factors in specifying organelle identities in the eukaryotic cell are the unique lipid compositions of the cytoplasmic leaflets of organelle membranes. It is these compositions, and their modifications, that determine the peripheral membrane protein cohort that will be recruited to each membrane system. In this chapter, we describe a set of quantitative approaches to characterize lipid binding, transfer and displacement reactions for soluble lipid transfer proteins (LTPs) or for soluble domains of otherwise insoluble (e.g., contact site) proteins. LTPs execute trafficking and sorting lipids between intracellular compartments and are essential for cells to achieve organelle identity. The presented approaches provide a template that can be more generally applied to experimental inquires of peripheral membrane proteins that are involved in lipid metabolism, trafficking and lipid-mediated signal transduction.