We have previously demonstrated that at temperatures where cell growth occurs (Tg) membrane lipids form a unique unilamellar structure whose properties depend only on the lipid composition. This structure was shown to form in both prokaryotic and eukaryotic cells whose Tg's span virtually the entire range of temperatures for life on Earth. We have also shown that when the lipid composition becomes inappropriate for a specific Tg as in Alzheimer's disease, and metachromatic leukodystrophy, this unique unilamellar lipid structure degenerates to another lipid state that does not support cell growth. Although some progress has been made in previous years to characterize the physical properties of the membrane lipids at Tg, we have focussed this year's research to provide details of the physical nature of this unique membrane lipid state. Since diffusion is a sensitive monitor of physical states, we have measured lateral diffusion of a fluorescent lipid analog, NBD-PE, in lipid bilayers prepared from E. coli grown at three different temperatures, 25, 29, and 32 deg C. In general, the diffusion coefficient D of the probe increases with temperature. However, there was a sharp jump in D over a narrow temperature interval (about 1 deg) at the corresponding growth temperatures of the cellular source of the lipids. Thus, D measured in extracts of cells grown at 25+/-0.5 deg C peaked at 24.6 deg, D measured in extracts of cells grown at 29+/-0.5 deg C peaked at 29.2 deg C, and D measured in extracts of cells grown at 32+/-0.5 deg C peaked at 32.5 deg C. When temperatures are further increased D falls sharply to lower values. This temperature dependent diffusion behavior is similar to observations others have made for fluid systems exhibiting critical phenomena. The presence of a critical state in lipid membranes indicates that lipids will cluster to form domains of varying size, and will undergo relatively large fluctuations of molecular area in the plane of the membrane at Tg. The significance of a critical state lies in the uniqueness of the interactions in this state that are a superposition of the classical short-range, molecule-molecule interactions and the long range interactions that are characteristic of the critical state. We have observed that relatively small changes in the membrane composition of specific lipids profoundly influences the stability of the critical unilamellar state at Tg. Maintaining the membrane lipids in a critical state may be important for the activity of membrane enzymes and for the control of vesicle fusion. An analytical function relating lipid composition and heats of mixing has been derived that will be the basis for a systematic description of the role of specific lipids on destabilizing the critical bilayer structure in membranes.
