The Ins and Outs of Lipid Flip-Flop

Our current view of cellular membranes centers on the fluid mosaic model, which envisions the cellular membrane as a liquidlike bilayer of lipids, cholesterol, and proteins that freely diffuse in two dimensions. In stark contrast, the exchange of materials between the leaflets of a bilayer was presumed to be prohibited by the large enthalpic bather associated with translocating hydrophilic materials, such as a charged lipid headgroup, through the hydrophobic membrane core. This static picture with regard to lipid translocation (or flip-flop as it is affectionately known) has been a long-held belief in the study of membrane dynamics. The current accepted membrane model invokes specific protein flippase (inward moving), floppase (outward moving), and scramblase (bidirectional) enzymes that assist in the movement of lipids between the leaflets of cellular membranes. The low rate of protein-free lipid flip-flop has also been a cornerstone of our understanding of the bilateral organization of cellular membrane components, specifically the asymmetric distribution of lipid species found in the luminal and extracellular leaflets of the plasma membrane of eukaryotic cells. Much of the previous work contributing to our current understanding of lipid flip-flop has utilized fluorescent- or spin-labeled lipids. However, there is growing evidence that these lipid probes do not accurately convey the dynamics and thermodynamics of native (unlabeled) lipid motion. This Account summarizes our research efforts directed toward developing a deep physical and chemical understanding of protein-free lipid flip-flop in phospholipid membrane models using sum-frequency vibrational spectroscopy (SFVS). Our use of SFVS enables the direct measurement, of native lipid flip-flop in model membranes. In particular, we have explored the kinetic rates and activation thermodynamics of lipid translocation as a means of deciphering the underlying chemical and physical directors governing this process. By means of transition state theory, the contributions from enthalpy and entropy on the activation energy barrier to lipid flip-flop have been explored in detail for a variety of lipid species and membrane compositions. Specifically, the effect of lipid structure and packing and the inclusion of cholesterol and transmembrane peptides on the rates and thermodynamics of lipid translocation have been investigated in detail. It is our hope that these studies will provide a new perspective on lipid translocation in biological membranes and the role of lipid flip-flop in generating and maintaining cell membrane lipid asymmetry.