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Langmuir : the ACS Journal of Surfaces... Sep 2019Lateral segregation of lipids in model and biological membranes has been studied intensively in the last decades using a comprehensive set of experimental techniques....
Lateral segregation of lipids in model and biological membranes has been studied intensively in the last decades using a comprehensive set of experimental techniques. Most methods require a probe to report on the biophysical properties of a specific molecule in the lipid bilayer. Because such probes can adversely affect the results of the measurement and perturb the local membrane structure and dynamics, a detailed understanding of probe behavior and its influence on the properties of its direct environment is important. Membrane phase-selective and lipid-mimicking molecules represent common types of probes. Here, we have studied how the fluorescent probes -parinaric acid (tPA), diphenylhexatriene (DPH), and 1-oleoyl-2-propionyl[DPH]--glycero-3-phosphocholine (O-DPH-PC) affect the membrane properties of 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC) bilayers using H and P NMR spectroscopy in the solid state. In addition, using 2D H magic-angle spinning (MAS) nuclear Overhauser enhancement spectroscopy (NOESY) NMR, we have determined the distribution of the probe moieties in the POPC membrane parallel to the membrane normal. We found that the different probes exhibit distinct membrane localizations and distributions, . tPA is located parallel to the membrane normal while DPH predominantly exist in two orientations. Further, tPA was conjugated to sphingomyelin (tPA-SM) as a substitute for the acyl chain in the SM. H NOESY NMR was used to probe the interaction of the tPA-SM with cholesterol as dominant in liquid ordered membrane domains in comparison to POPC-cholesterol interaction in membranes composed of ternary lipid mixtures. We could show that tPA-SM exhibited a strong favorable and very temperature-dependent interaction with cholesterol in comparison to POPC. In conclusion, the NMR techniques can explain probe behavior but also be used to measure lipid-specific affinities between different lipid segments and individual molecules in complex bilayers, relevant to understanding nanodomain formation in biological membranes.
PubMed: 31424941
DOI: 10.1021/acs.langmuir.9b01202 -
Biophysical Journal Jul 2019Ceramide-1-phosphate is a minor sphingolipid with important functions in cell signaling. In this study, we examined the propensity of palmitoyl ceramide-1-phosphate...
Ceramide-1-phosphate is a minor sphingolipid with important functions in cell signaling. In this study, we examined the propensity of palmitoyl ceramide-1-phosphate (Cer-1P) to segregate laterally into ordered domains in different bilayer compositions at 23 and 37°C and compared this with segregation of palmitoyl ceramide (PCer) and palmitoyl sphingomyelin (PSM). The ordered-domain formation in the fluid phosphatidylcholine bilayers was determined using the emission lifetime changes of trans-parinaric acid and from differential scanning calorimetry thermograms. The lateral segregation of Cer-1P was examined when hydrated to bilayers in Tris buffer (50 mM Tris, 140 mM NaCl (pH 7.4)). At this pH, Cer-1P was negatively charged. The lateral segregation propensity of Cer-1P in 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) bilayers was intermediate between PCer and PSM. Based on differential scanning calorimetry analysis, we observed that the gel domains formed by Cer-1P in POPC bilayers (POPC:Cer-1P 70:30 by mol) were less stable (melting interval 16-37°C) than the corresponding POPC and PCer gel domains at equal composition (melting interval 20-55°C). The gel-phase melting enthalpy was also much lower in Cer-1P (1.5 kcal/mol) than in the PCer-containing POPC bilayers (9 kcal/mol). Cer-1P appeared to be at least partially miscible with PCer domains in POPC bilayers. Cer-1P domains were stabilized in the presence of PSM (POPC:PSM 85:15), similarly as seen with PCer-rich domains. In bilayers at 37°C, with an approximate outer-leaflet cell membrane composition (sphingomyelin and cholesterol enriched, aminophospholipid poor), Cer-1P segregation did not lead to the formation of ordered domains, at least when compared with PCer segregation. In bilayers with an approximate inner-leaflet composition (sphingomyelin poor, cholesterol and aminophospholipid enriched), Cer-1P also failed to form ordered domains. PCer segregated into ordered domains only after the PCer/cholesterol ratio exceeded an approximate equimolar ratio.
Topics: Ceramides; Cholesterol; Lipid Bilayers; Liposomes; Phosphatidylcholines; Thermodynamics
PubMed: 31133285
DOI: 10.1016/j.bpj.2019.05.015