phosphorus-radioisotopes and 1-palmitoyl-2-oleoylphosphatidylcholine

The synthetic estrogen diethylstilbestrol (DES) is used to treat metastatic carcinomas and prostate cancer. We studied its interaction with membranes and its localization to understand its mechanism of action and side-effects. We used differential scanning calorimetry (DSC) showing that DES fluidized the membrane and has poor solubility in DMPC (1,2-dimyristoyl-

Topics: Calorimetry, Differential Scanning; Carbon; Diethylstilbestrol; Fatty Acids; Lipid Bilayers; Magnetic Resonance Spectroscopy; Membranes, Artificial; Molecular Dynamics Simulation; Phase Transition; Phosphatidylcholines; Phospholipids; Phosphorus Radioisotopes; Protons; Solubility; Temperature; Thermodynamics; X-Ray Diffraction

Nonlamellar lipid membranes are frequently induced by proteins that fuse, bend, and cut membranes. Understanding the mechanism of action of these proteins requires the elucidation of the membrane morphologies that they induce. While hexagonal phases and lamellar phases are readily identified by their characteristic solid-state NMR line shapes, bicontinuous lipid cubic phases are more difficult to discern, since the static NMR spectra of cubic-phase lipids consist of an isotropic (31)P or (2)H peak, indistinguishable from the spectra of isotropic membrane morphologies such as micelles and small vesicles. To date, small-angle X-ray scattering is the only method to identify bicontinuous lipid cubic phases. To explore unique NMR signatures of lipid cubic phases, we first describe the orientation distribution of lipid molecules in cubic phases and simulate the static (31)P chemical shift line shapes of oriented cubic-phase membranes in the limit of slow lateral diffusion. We then show that (31)P T2 relaxation times differ significantly between isotropic micelles and cubic-phase membranes: the latter exhibit 2 orders of magnitude shorter T2 relaxation times. These differences are explained by the different time scales of lipid lateral diffusion on the cubic-phase surface versus the time scales of micelle tumbling. Using this relaxation NMR approach, we investigated a DOPE membrane containing the transmembrane domain (TMD) of a viral fusion protein. The static (31)P spectrum of DOPE shows an isotropic peak, whose T2 relaxation times correspond to that of a cubic phase. Thus, the viral fusion protein TMD induces negative Gaussian curvature, which is an intrinsic characteristic of cubic phases, to the DOPE membrane. This curvature induction has important implications to the mechanism of virus-cell fusion. This study establishes a simple NMR diagnostic probe of lipid cubic phases, which is expected to be useful for studying many protein-induced membrane remodeling phenomena in biology.

Topics: Deuterium; Lipids; Magnetic Resonance Spectroscopy; Membranes, Artificial; Micelles; Parainfluenza Virus 5; Phase Transition; Phosphatidylcholines; Phosphatidylethanolamines; Phosphatidylglycerols; Phosphorus Radioisotopes; Scattering, Small Angle; Temperature; Viral Fusion Proteins; X-Ray Diffraction

27Al and 31P nuclear magnetic resonance (NMR) spectroscopies were used to investigate aluminum interactions at pH 3.4 with model membranes composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC). A solution state 27Al NMR difference assay was developed to quantify aluminum binding to POPC multilamellar vesicles (MLVs). Corresponding one-dimensional (1D) fast magic angle spinning (MAS) 31P NMR spectra showed that aluminum induced the appearance of two new isotropic resonances for POPC shifted to -6.4 ppm and -9.6 ppm upfield relative to, and in slow exchange with, the control resonance at -0.6 ppm. Correlation of the (27)Al and (31)P NMR binding data revealed a 1:2 aluminum:phospholipid stoichiometry in the aluminum-bound complex at -9.6 ppm and a 1:1 aluminum:phospholipid stoichiometry in that at -6.4 ppm. Slow MAS 31P NMR spectra demonstrated shifts in the anisotropic chemical shift tensor components of the aluminum-bound POPC consistent with a close coordination of aluminum with phosphorus. A model of the aluminum-bis-phospholipid complex is proposed on the basis of these findings.

Topics: Aluminum; Binding Sites; Lipid Bilayers; Liposomes; Macromolecular Substances; Magnetic Resonance Spectroscopy; Membranes, Artificial; Models, Chemical; Models, Molecular; Phosphatidylcholines; Phosphorus Radioisotopes

The isothermal phase behavior of three gramicidin A'/phospholipid mixtures was investigated by an equilibrium Ca(2+)-binding technique. The phospholipid component was 1,2-dioleoyl-sn-glycero-3-phosphoserine (DOPS), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoserine (POPS), or POPS/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) at a constant mole ratio of 1/4. The bulk aqueous free Ca2+ concentration, [Ca2+]*f, in equilibrium with one or two gramicidin A'/phospholipid fluid phases and a small amount of the Ca (phosphatidylserine)2 gel phase, was measured as a function of composition at 20 degrees C by use of chromophoric high-affinity Ca2+ chelators. The coexistence of two gramicidin A'/phospholipid fluid phases was detected by an invariance in [Ca2+]*f over the range of compositions throughout which the two phases coexist. The compositions of the two coexisting phases are determined by the compositions at which the invariance in [Ca2+]*f begins and ends. With each of the gramicidin A'/phospholipid mixtures, we estimate that the composition of the gramicidin-poor phase is 0.03-0.04 mole fraction gramicidin A' and the composition of the gramicidin-rich phase is 0.13-0.14 mole fraction gramicidin A'. Characterization of these phases by low-angle X-ray diffraction revealed that, in each case, the gramicidin-poor phase is an L alpha phase and the gramicidin-rich phase is an HII phase. The isothermal phase behavior of gramicidin A'/POPC mixtures at approximately 23 degrees C, as determined by low-angle X-ray diffraction, was found to be similar to that of the other gramicidin A'/phospholipid mixtures.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Binding Sites; Calcium; Chelating Agents; Gramicidin; Kinetics; Magnetic Resonance Spectroscopy; Phosphatidylcholines; Phosphatidylserines; Phosphorus Radioisotopes; Reference Standards; X-Ray Diffraction