Unsaturated Phosphatidylcholine Acyl Chain Structure Affects the Size of Ordered Nanodomains (Lipid Rafts) Formed by Sphingomyelin and Cholesterol

Abstract

How lipid composition affects the size of ordered domains (lipid rafts) in ternary lipid mixtures with cholesterol is unclear. We used FRET, DPH anisotropy, and quenching by the nitroxide-bearing molecule tempo to study raft size and raft thermal stability. Because FRET pairs with a relatively long distance range were used, FRET was not be able to detect very small nanodomains. In contrast, such domains could be detected by nitroxide quenching, which is of very short range, and anisotropy, which measures order of the lipids in immediate contact with the fluorescent probe. Ordered domain formation in lipid vesicles containing 1:1:1 sphingomyelin/DOPC/cholesterol or sphingomyelin/POPC/cholesterol were compared. These mixtures form co-existing ordered and disordered domains at lower temperatures and homogeneous disordered fluid domains at high temperature. The melting temperature, above which ordered domains disappear, was similar for these mixtures as measured by anisotropy and tempo quenching, but much higher for sphingomyelin/DOPC/cholesterol than for sphingomyelin/POPC/cholesterol when measured by FRET. This was true for more than one donor/acceptor FRET pair. We conclude that these mixtures form ordered domains with a similar stability. However, while domains large enough to detect by FRET form in sphingomyelin/DOPC/cholesterol under a wide variety of conditions, sphingomyelin/POPC/cholesterol has a tendency to form very small nanodomains. Because POPC is an abundant lipid in mammalian cells, this may be one reason that cellular ordered domains/rafts are very small.