sphingosine-phosphorylcholine and 1-2-oleoylphosphatidylcholine

We have demonstrated that dynamic phase separation is induced by coalescence of two self-spreading supported lipid bilayers (SLBs) with different components. Coalescence between a phosphocholine/sphingolipid SLB and a phosphocholine/cholesterol one forms raft-like liquid ordered (Lo) domains, which can be observed by fluorescence microscopy at the boundary of two phases. This phase separation process indicates that lipid molecules, such as sphingolipid and cholesterol, are intermixed. When saturated phospholipid is used instead of sphingolipid, small Lo domains are formed. Cholesterol is harder to incorporate with domains of saturated phospholipid than that of sphingolipid. This technique is very useful for observation of lipid-lipid interactions.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Cholesterol; Glycerylphosphorylcholine; Lipid Bilayers; Microscopy, Fluorescence; Models, Molecular; Phosphatidylcholines; Phosphorylcholine; Sphingosine

An interaction of glycyrrhizin (GC) with a lipid raft biomembrane model that consisted of N-palmitoyl-d-erythro-sphingosylphosphorylcholine (PSM), 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), and cholesterol (CHOL) was systematically studied using the Langmuir monolayer technique. To construct the lipid raft model, the surface pressure (π)-molecular area (A) and surface potential (ΔV)-A isotherms for three-component (PSM/DOPC/CHOL) systems on 0.02M Tris buffer with 0.13M NaCl (pH7.4) were primarily measured by changing their compositions. Thermodynamic and interaction parameters for binary PSM/DOPC and PSM/CHOL systems revealed that PSM interacts more strongly with CHOL than with DOPC. In addition, a morphological analysis performed with Brewster angle microscopy (BAM) and fluorescence microscopy (FM) revealed an optimal ratio of PSM/DOPC/CHOL (1/1/1, by mole) as a model of lipid rafts. Second, the interaction of GC with the ternary PSM/DOPC/CHOL monolayers was investigated on Tris buffer solutions containing different GC concentrations (1, 5, 10, 25, and 50μM). In BAM and FM images, microdomains were found to become smaller by increasing the GC concentration in the subphase, suggesting that GC regulates the size of raft domains, which provide dynamic scaffolding for numerous cellular processes. More interestingly, the distinctive GC striped regions were formed at the interface at 50μM, which shows that GC divides the ternary monolayer into pieces. This phenomenon was observed only in the presence of CHOL in the monolayer. These results suggest that CHOL plays an essential role in the interaction with GC, which results in one of the major activities associated with saponins' membrane disruption.

Topics: Cholesterol; Glycyrrhizic Acid; Membrane Microdomains; Microscopy, Fluorescence; Phosphatidylcholines; Phosphorylcholine; Sphingosine

We measured the nonradiative fluorescence resonance energy transfer between 7-nitro-2,1,3-benzoxadiazol-4-yl (NBD) labeled lipids (amine labeled phosphatidylethanolamine or acyl chain labeled phosphatidylcholine) and rhodamine labeled lipids in large unilamellar dioleoylphosphatidylcholine vesicles. Two new rhodamine labeled lipid analogues, one a derivative of monolauroylphosphatidylethanolamine and the other of sphingosylphosphorylcholine, were found to exchange through the aqueous phase between vesicle populations but not to be capable of rapid transbilayer movement between leaflets. Energy transfer from NBD to rhodamine was measured using liposomes with symmetric or asymmetric distributions of these new rhodamine labeled lipid analogues to determine the relative contributions of energy transfer between donor and acceptor fluorophores in the same (cis) and opposite (trans) leaflets. Since the characteristic R0 values for energy transfer ranged from 47 to 73 A in all cases, significant contributions from both cis and trans energy transfer were observed. Therefore, neither of these probes acts strictly as a half-bilayer quencher of NBD lipid fluorescence. The dependence of transfer efficiency on acceptor density was fitted to a theoretical treatment of energy transfer to determine the distances of closest approach for cis and trans transfer. These parameters set limits on the positions of the fluorescent groups relative to the bilayer center, 20-31 A for NBD and 31-55 A for rhodamine, and provide a basis for future use of these analogues in measurements of transbilayer distribution and transport.

Topics: 4-Chloro-7-nitrobenzofurazan; Energy Transfer; Fluorescent Dyes; Lipid Bilayers; Liposomes; Lysophospholipids; Models, Molecular; Molecular Structure; Phosphatidylcholines; Phosphatidylethanolamines; Phospholipids; Phosphorylcholine; Rhodamines; Spectrometry, Fluorescence; Sphingosine