Monounsaturated PE Does Not Phase-Separate from the Lipid Raft Molecules Sphingomyelin and Cholesterol: Role for Polyunsaturation?

We investigated interactions of the lipid raft molecules sphingomyelin (SM) and cholesterol (CHOL) in monolayers and bilayers composed of 1-palmitoyl-2-oleoyl-sn-glycerophosphatidylethanolamine (POPE) or 1-palmitoyl-2-docosahexaenoyl-sn-glycerophosphatidylethanolamine (PDPE) at 35 °C. Techniques employed were pressure-area (π-A) isotherms generated from Langmuir-Blodgett films, solidstate ²H and ³¹P NMR spectroscopies, and differential scanning calorimetry (DSC). Condensation calculated from Ï€-A isotherms and reduction in the enthalpy of the gel-liquid-crystalline transition in DSC scans showed CHOL has a strong affinity for POPE, comparable to that observed between SM-CHOL. Order parameters derived from ²H NMR spectra of the perdeuterated sn-1 chain of POPE-d31 increased by >50% upon addition of equimolar CHOL to POPE-d₃₁/SM (1:1 mol) bilayers. Close proximity of CHOL to POPE even in the presence of SM is indicated. Chemical shift anisotropy (Δσ_csa) measured from ¹H-decoupled ³¹P NMR spectra also implied intimate lipid mixing in POPE/SM/CHOL (1:1:1 mol). In contrast, π-A isotherms and corroborating DSC studies of PDPE/SM (1:1 mol) indicate phase separation between SM and PDPE, which was maintained in the presence of CHOL. The cholesterol-associated increase in order of the perdeuterated sn-1 chain of PDPE determined by ²H NMR was 2-fold less for PDPE-d₃₁/SM/CHOL (1:1:1 mol) than POPE-d₃₁/SM/CHOL (1:1:1 mol). Our findings support the notion that acyl chain dependent lateral phase separation occurs in the presence of a docosahexaenoic acid (DHA)-containing phospholipid (PDPE), but not an oleic acid-containing phospholipid (POPE). We propose that monounsaturated lipids do not promote formation of stable lipid rafts and that polyunsaturation may be important for raft stability.