A detailed study on the structure, dynamics, and thermodynamic behavior of phosphatidylcholine/cholesterol (PC/CHOL) mixtures was undertaken using differential scanning calorimetry (DSC) and solid-state nuclear magnetic resonance (NMR) spectroscopy. DSC thermograms of mixtures of cholesterol (CHOL) with 1,2-dipalmitoyl-sn-phosphatidylcholine (DPPC), 1,2-distearoyl-sn-phosphatidylcholine (DSPC), and 1,2-diarachidoyl-sn-phosphatidylcholine (DAPC) showed a broadening of the first-order gel-->liquid crystalline transition and a decrease in the transition enthalpy, indicating a gradual loss of cooperativity for high CHOL concentrations. DPPC and DSPC were labeled with 13C at the carbonyl group of the sn-2 chain and 2H was introduced into the middle of the sn-2 chain at the 6- and 12-position for DPPC and DSPC, respectively. The 13C and 2H NMR spectra of each labeled lipid were studied as a function of temperature and CHOL concentration. The residual quadrupole splitting in the 2H NMR spectra, delta nu Q perpendicular, was analyzed as a function of temperature and composition. For CHOL concentrations less than 30 mol %, a precipitous change in delta nu Q perpendicular occurs near the chain melting temperature of the phospholipid. Further increases in CHOL concentration broaden the transition and shift the midpoint to higher temperature, indicating the presence of a new phase at higher CHOL contents. Moreover, at a given temperature, delta nu Q perpendicular increases with increasing cholesterol content, which indicates a more ordered structure. The 13C NMR spectra in the gel state consisted of a superposition of two components which can be attributed to both gel-like and fluid phospholipid domains in the bilayer. This two-component spectrum can be simulated quantitatively with a two-parameter chemical exchange model, which permits the fraction of each form and the exchange rate to be determined as a function of temperature and composition. At high CHOL contents the line width of the fluid component broadens, suggesting an increase in the exchange rate between the domains. These results were interpreted in terms of a temperature composition diagram with one region L beta', two regions LGI and LGII, and one liquid crystalline region L alpha, with LG denoting "liquid-gel" type phases. Liquid-gel phases correspond to phases with increased order in the hydrocarbon chains (in comparison to that of the pure PC bilayer in the L alpha phase) combined with fast limit axial diffusion that averages the 13C NMR spectrum to a "fluidlike" line.(ABSTRACT TRUNCATED AT 400 WORDS)
Solid-state nuclear magnetic resonance (NMR) spectroscopy and X-ray powder diffraction were used to investigate the mechanism of trehalose (TRE) stabilization of lipid bilayers. Calorimetric investigation of dry TRE-stabilized bilayers reveals a first-order phase transition (L kappa----L lambda) at temperatures similar to the L beta'----(P beta')----L alpha transition of hydrated lipid bilayers. X-ray diffraction studies show that dry mixtures of TRE and 1,2-dipalmitoyl-sn-phosphatidylcholine (DPPC) have a lamellar structure with excess crystalline TRE being present. The L kappa phase shows typical gel-phase X-ray diffraction patterns. In contrast, the L lambda-phase diffraction patterns indicate disordered hydrocarbon chains. 2H NMR of specifically 2H chain-labeled DPPC confirmed that the acyl chains are disordered in the L lambda phase over their entire lengths. 2H spectra of the choline headgroup show hindered molecular motions as compared to dry DPPC alone, and 13C spectra of the sn-2-carbonyl show rigid lattice powder patterns indicating very little motion at the headgroup and interfacial regions. Thus, the sugar interacts extensively with the hydrophilic regions of the lipid, from the choline and the phosphate moieties in the headgroup to the glycerol and carbonyls in the interfacial region. We postulate that the sugar and the lipid form an extensive hydrogen-bonded network with the sugar acting as a spacer to expand the distance between lipids in the bilayer. The fluidity of the hydrophobic region in the L lambda phase together with the bilayer stabilization at the headgroup contributes to membrane viability in anhydrobiotic organisms.
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