Lipid domain nucleation, growth and phase behavior: Model systems for biological membranes

Over the past decade there has been an emergence of evidence indicating that the cell membrane may not exist as a homogeneous lipid matrix, but rather certain lipid constituents (glycosphingolipids, cholesterol, and sphingomyelin) phase separate into microdomains or rafts. Rafts are believed to serve several functions, including signaling, sorting and trafficking, and acting as attachment platforms for host pathogens and their toxins. A notable example of this later function is the sexual transmission of HIV, which occurs through rafts enriched in Galactosylceramide (GalCer). The existence of rafts is based on the notion that particular lipids can 'phase-separate' forming ordered domains within the less ordered fluid lipid matrix. Model membrane systems containing 'raft-like' lipids, fluid phase lipids and cholesterol (cholesterol is believed to play a role in raft formation) have been extensively employed to study this phenomenon. A specific model membrane system that has been used to mimic biological membranes is supported lipid bilayers (SLBs). The two-dimensional geometry of SLBs allows one to readily apply high-resolution surface probe microscopy techniques to characterize lipid domain formation and microstructure. GalCer was the primary 'raft-like' lipid studied in this dissertation due to its the biological relevance in relation to the sexual transmission of HIV. Using slow-cooled SLBs the effects of the fluid phase lipid component on lipid domain phase in ternary mixtures containing GalCer, cholesterol and one of the following fluid phase lipids 1,2-Dilauroyl-sn-Glycero-3-Phosphocholine (DLPC), 1-Palmitoyl-2-Oleoyl-sn-Glycero-3-Phosphocholine (POPC), 1,2-Dioleoyl-sn-Glycero-3-Phosphocholine (DOPC) was examined. From this work it was shown that the degree of fluid phase acyl chain unsaturation has dramatic effects on the partitioning of cholesterol between the 'raft phase' and fluid phase. In addition we have examined the process of lipid domain nucleation and growth as a function of temperature. Using this method in combination with classical nucleation and growth theories we calculated interfacial line tension between the 'raft-phase' and surrounding fluid phase as a function of domain symmetry and cholesterol for a wide range of lipid compositions. Theses measurements proved valuable in understanding the nano-scale nature of membrane rafts and providing insight into the energetic barriers cell membranes must overcome for lipid domain growth.