Liposome Co-sedimentation and Co-flotation Assays to Study Lipid–Protein Interactions

A large proportion of proteins are expected to interact with cellular membranes to carry out their physiological functions in processes such as membrane transport, morphogenesis, cytoskeletal organization, and signal transduction. The recruitment of proteins at the membrane–cytoplasm interface and their activities are precisely regulated by phosphoinositides, which are negatively charged phospholipids found on the cytoplasmic leaflet of cellular membranes and play critical roles in membrane homeostasis and cellular signaling. Thus, it is important to reveal which proteins interact with phosphoinositides and to elucidate the underlying mechanisms. Here, we present two standard in vitro methods, liposome co-sedimentation and co-flotation assays, to study lipid–protein interactions. Liposomes can mimic various biological membranes in these assays because their lipid compositions and concentrations can be varied. Thus, in addition to mechanisms of lipid–protein interactions, these methods provide information on the possible specificities of proteins toward certain lipids such as specific phosphoinositide species and can hence shed light on the roles of membrane interactions on the functions of membrane-associated proteins.