CELLULAR & MOLECULAR BIOLOGY LETTERS Volume 8, (2003) pp 147 ��� 159 http://www.cmbl.org.pl Received 24 September 2002 Accepted 10 February 2003 Abbreviations used: EPR - electron paramagnetic resonance PC - phospatidylcholine DLPC - dilauroylphospatidylcholine DMPC - dimyristoylphospatidylcholine DPPC - dipalmitoylphospatidylcholine DSPC - distearoylphospatidylcholine DOPC - dioleoylphospatidylcholine DEPC - dielaidoylphospatidylcholine EYPC - egg-yolk phospatidylcholine GPI - glycosylphosphatidylinositol DRM - detergent-resistant membrane DOT - discrimaination by oxygen transport SLOT - slow oxygen transport BR - bacteriorodopsin IFV - influenza virus IS A FLUID-MOSAIC MODEL OF BIOLOGICAL MEMBRANES FULLY RELEVANT? STUDIES ON LIPID ORGANIZATION IN MODEL AND BIOLOGICAL MEMBRANES ANNA WI��NIEWSKA1, JOLANTA DRAUS1 and WITOLD K. SUBCZYNSKI2 1Biophysics Department, Institute of Molecular Biology and Biotechnology, Jagiellonian University, Krakow, Poland, 2Biophysics Research Institute, Medical College of Wisconsin, Milwaukee, Wisconsin, USA Abstract: The basic concept of the fluid-mosaic model of Singer and Nicolson, an essential point of which is that the membrane proteins are floating in a sea of excess lipid molecules organized in the lipid bilayer, may be misleading in understanding the movement of membrane components in biological membranes that show distinct domain structure. It seems that the lipid bilayer is an active factor in forming the membrane structure, and the lipid composition is responsible for the presence of domains in the membrane. The main role in the process of domain formation is played by cholesterol and sphingolipids. The results presented here show that in a binary mixture of cholesterol and unsaturated phospholipids, cholesterol is segregated out from the bulk unsaturated liquid-crystalline phase. This forms cholesterol-enriched domains or clustered cholesterol domains due to the lateral nonconformability between the rigid planar ring structure of cholesterol and the rigid bend of the unsaturated alkyl chain at double bond position. These cholesterol-enriched domains may be stabilized by the presence of saturated alkyl chains of sphingomyelin or glycosphingolipids, and also by specific proteins which selectively locate in these domains and stabilize them as a result of protein-protein interaction. Such lipid domains are called ���rafts��� and have been shown to be responsible both for signal transduction to and from the cell and for protein sorting. We also looked

CELL. MOL. BIOL. LETT. Vol. 8. No. 1. 2003 148 at whether polar carotenoids, compounds showing some similarities to cholesterol and affecting membrane properties in a similar way, would also promote domain formation and locate preferentially in one of the lipid phases. Our preliminary data show that in the presence of cholesterol, lutein (a polar carotenoid) may segregate out from saturated lipid regions (liquid-ordered phase) and accumulate in the regions rich in unsaturated phospholipids forming carotenoid-rich domains there. Conventional and pulse EPR (electron paramagnetic resonance) spin labeling techniques were employed to assess the molecular organization and dynamics of the raft-constituent molecules and of the raft itself in the membrane. Key Words: Lipid Unsaturation, Cholesterol, Lutein, Membrane Domains, Raft Domains, Lipid Bilayer INTRODUCTION The basic concept of the fluid-mosaic model of a biological membrane is that the membrane proteins are floating in a sea of excess lipid molecules organized in a bilayer [1]. According to this model, lipids are mainly responsible for isolating the cell interior from the outside world and providing the environment for membrane proteins that regulate the exchange of substances and communicate between the cell and its surroundings. However, the wide variety of membrane lipids (more than 2000 different lipids in membranes of mammalian cells, including sphingolipids and sterols [2]), suggests more than a simple barrier function for a lipid bilayer. This lipid heterogeneity must have some functional consequences. One of them is non-random mixing of lipid molecules in a bilayer, resulting in phase separation and formation of lipid domains. It seems, therefore, that the lipid bilayer is an active factor in the formation of the membrane structure and the lipid composition is responsible for the presence of domains in the membrane. Lipid bilayers can exist in two main phases, depending on temperature. At low temperatures (below the melting temperature (Tm) characteristic of each lipid bilayer, which depends on the alkyl chains length and degree of unsaturation), the bilayer is present in a gel phase. In the gel phase, lipid alkyl chains are ordered and their mobility is restricted. Above Tm, the bilayer exists in a phase, termed liquid-crystalline (lc) or liquid-disordered (ld), in which alkyl chains are fluid and disordered. When a bilayer is formed of two lipids with different Tm, a phase separation may occur, and the gel and liquid-crystalline phase domains coexist. The gel phase does not appear to exist in biological membranes, but phase separation between two fluid phases may occur when cholesterol is present in the membrane [3,4]. In this case, a liquid-ordered (lo) phase can separate from a liquid-disordered phase as the amount of cholesterol is increased. Lipid alkyl chains in the lo phase have intermediate properties between those of gel and ld phases: they are extended and ordered, as in the gel