Lipid Chain Unsaturation Research Articles

Effects of lipid chain unsaturation and headgroup type on molecular interactions between paclitaxel and phospholipid within model biomembrane

Molecular interactions between paclitaxel, an anticancer drug, and phospholipids of various chain unsaturations and headgroup types were investigated in the present study by Langmuir film balance and differential scanning calorimetry. Both the lipid monolayer at the air–water interface and the lipid bilayer vesicles (liposomes) were employed as model cell membranes. It was found that, regardless of the difference in molecular structure of the lipid chains and headgroup, the drug can form nonideal, miscible systems with the lipids at the air–water interface over a wide range of paclitaxel mole fractions. The interaction between paclitaxel and phospholipid within the monolayer was dependent on the molecular area of the lipids at the interface and can be explained by intermolecular forces or geometric accommodation. Paclitaxel is more likely to form thermodynamically stable systems with 1,2-dipalmitoyl- sn-glycerol-3-phosphocholine (DPPC) and 1,2-dielaidoyl- sn-glycero-3-phosphocholine (DEPC) than with 1,2-dipalmitoyl- sn-glycero-3-phosphoethanolamine (DPPE) and 1,2-distearoyl- sn-glycero-3-phosphocholine (DSPC). Investigation of the drug penetration into the lipid monolayer showed that DPPC and DEPC have higher incorporation abilities for the drug than DPPE and DSPC. A similar trend was also evidenced by DSC investigation with liposomes. While little change of DSC profiles was observed for the DPPE/paclitaxel and DSPC/paclitaxel liposomes, paclitaxel caused noticeable changes in the thermographs of DPPC and DEPC liposomes. Paclitaxel was found to cause broadening of the main phase transition without significant change in the peak melting temperature of the DPPC bilayers, which demonstrates that paclitaxel was localized in the outer hydrophobic cooperative zone of the bilayer, i.e., in the region of the C1–C8 carbon atoms of the acyl chain or binding at the polar headgroup site of the lipids. However, it may penetrate into the deeper hydrophobic zone of the DEPC bilayers. These findings provide useful information for liposomal formulation of anticancer drugs as well as for understanding drug–cell membrane interactions.

Influence of lipid chain unsaturation on membrane-bound melittin: a fluorescence approach

Melittin, a cationic hemolytic peptide, is intrinsically fluorescent due to the presence of a single functionally important tryptophan residue. The organization of membrane-bound melittin is dependent on the physical state and composition of membranes. In particular, polyunsaturated lipids have been shown to modulate the membrane-disruptive action of melittin. Phospholipids with polyunsaturated acyl chains are known to modulate a number of physical properties of membranes and play an important role in regulating structure and function of membrane proteins. In this study, we have used melittin to address the influence of unsaturated lipids in modulating lipid–protein interactions. Our results show that fluorescence parameters such as intensity, emission maximum, polarization, lifetime and acrylamide quenching of melittin incorporated in membranes are dependent on the degree of unsaturation of lipids in membranes. Importantly, melittin in membranes composed of various unsaturated lipids shows red edge excitation shift (REES) implying that melittin is localized in a motionally restricted region in membranes. The extent of REES was found to increase drastically in membranes with increasing unsaturation, especially when the lipids contained more than two double bonds. In addition, increasing unsaturation in membranes causes a considerable change in the secondary structure of membrane-bound melittin. Taken together, our results assume significance in the overall context of the role of unsaturated lipids in membranes in the organization and function of membrane proteins and membrane-active peptides.

Effect of Unsaturated Lipid Chains on Dimensions, Molecular Order and Hydration of Membranes

We conducted infrared linear dichroism and X-ray measurements to characterize membranes of sn-1 chain perdeuterated, polyunsaturated 1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine (18:0−22:6ω3 PC, SDPC-d35) and of monounsaturated 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (16:0−18:1 PC, POPC-d31) as a function of hydration and temperature. Our novel approach is to use IR order parameters of C−H and C−D vibrations of both sn-1 and sn-2 chains for a separate estimation of the mean length and effective cross sectional area of the saturated and unsaturated chains in the bilayers. In the gel phase, the thickness of the hydrophobic core of SDPC bilayers exceeded the thickness of POPC bilayers, because the polyunsaturated chains adopted an extended conformation. However, in the fluid phase, the hydrophobic thickness of SDPC membranes, with the longer 18:0 and 22:6 acyl chains, was almost equal to the thickness of the 16:0−18:1 chains in POPC membranes. The isothermal compressibility modulus of the docosahexaenoyl chains was smaller than the modulus of the stearoyl chains of SDPC, whereas the palmitoyl and oleoyl chains of POPC are similarly compressible. Analysis in terms of a simple polymer brush model showed that unsaturation increases chain flexibility and chain mean area in lipid membranes. Temperature dependent measurements indicated that polyunsaturation decreased the sensitivity of the bilayer to temperature changes. The study of lipid hydration-sensitive spectral parameters suggested that this tendency may be at least partially explained by a weakening of water binding to polar moieties of polyunsaturated lipids with increasing temperature. Although the influence of hydrocarbon chain polyunsaturation on hydration of phosphate groups and the entire lipid was minor, we observed a modification of lipid carbonyl group hydration. Hydration properties and phase behavior of SDPC suggest an influence from polyunsaturation on the lateral pressure profile across the bilayer.

Influence of lipid chain unsaturation on melittin-induced micellization

It is well known that melittin, an amphipathic helical peptide, causes the micellization of phosphatidylcholine vesicles. In the present work, we conclude that the extent of micellization is dependent on the level of unsaturation of the lipid acyl chains. We report the results obtained on two systems: dipalmitoylphosphatidylcholine (DPPC), containing 10(mol)% saturated or unsaturated fatty acid (palmitic, oleic, or linoleic), and DPPC, containing 10(mol)% positively charged diacyloxy-3-(trimethylammonio)propane bearing palmitic or oleic acyl chains. For both systems, the presence of unsaturation in the lipid acyl chains inhibits melittin-induced micellization. Conversely, the addition of saturated palmitic acid to the DPPC matrix enhances the micellization. This modulation is proposed to be associated with the cohesion of the hydrophobic core. When the lipid chain packing of the gel-phase bilayer is already perturbed by the presence of unsaturation, it seems easier for the membrane to accommodate melittin at the interface, and the distribution of the peptide in the bilayer could be the origin of the inhibition of the micellization. The cohesion of the apolar core is shown to play an unquestionable role in melittin-induced micellization; however, this contribution does not appear to be as important as the electrostatic interactions between melittin and positively or negatively charged lipids.

A small protein in model membranes: a time-resolved fluorescence and ESR study on the interaction of M13 coat protein with lipid bilayers.

Model membranes with unsaturated lipid chains containing various amounts of M13 coat protein in the alpha-helical form were studied using time-resolved fluorescence and ESR spectroscopy. The lipid-to-protein (L/P) ratios used were > 12 to avoid protein-protein contacts and irreversible aggregation leading to beta-polymeric coat protein. In the ESR spectra of the 12-SASL probe in dioleoyl phosphatidylcholine (DOPC) bilayers no second protein induced component is observed upon incorporation of M13 coat protein. However, strong effects are detected on the ESR lineshapes upon changing the protein concentration. The ESR lineshapes are simulated by assuming a fixed ratio between the parallel (D parallel) and perpendicular (D perpendicular) diffusion coefficients of 4, and an order parameter equal to zero. It is found that increasing the protein concentration from L/P infinity to L/P 15 results in a decrease of the rotational diffusion coefficient D perpendicular from 3.4 x 10(7) to 1.9 x 10(7) s-1. In the time-resolved fluorescence experiments with DPH-propionic acid as a probe, it is observed that increasing the M13 coat protein concentration causes an increase of the two fluorescent lifetimes, indicating an increase in bilayer order. Analysis of the time-resolved fluorescence anisotropy decay allows one to quantitatively determine the order parameters <P2> and <P4>, and the rotational diffusion coefficient D perpendicular of the fluorescent probe. The order parameters <P2> and <P4> increase from 0.34 to 0.55 and from 0.59 to 0.77, respectively, upon adding M13 coat protein to DOPC bilayers with an L/P ratio of 35.(ABSTRACT TRUNCATED AT 250 WORDS)

Dynamic structure of biological and model membranes: analysis by optical anisotropy decay measurement.

Rotational Brownian motion of molecules in membranes can be "visualized" by time-resolved measurement of the decay of anisotropy of various flash-induced optical signals, such as fluorescence, phosphorescence, delayed fluorescence, transient absorption, or fluorescence depletion. The basic principles of the various forms of anisotropy measurement are illustrated in a unified manner. In organized structures such as membranes, rotational motion is restricted in angular range. Methods of analysis of observed optical anisotropy decays for the case of restricted rotation are described; the emphasis is laid on the separate estimation of the two important parameters, the range and rate, that characterize the restricted rotation. Practical aspects of the analytical procedures are also discussed. As an example of application, recent work from the authors' laboratory is reviewed: dynamic structures of lipid hydrocarbon chain region of membranes have been revealed by time-resolved fluorescence depolarization studies. A lipophilic fluorescent probe 1,6-diphenyl-1,3,5-hexatriene was incorporated in model and biological membranes of known compositions. The decay of fluorescence anisotropy indicated that the rod-shaped probe molecules wobbled in the membranes with a wobbling diffusion constant around 0.1 rad2/nsec, presumably reflecting the dynamics of surrounding lipid chains. The effects of temperature, ions, lipid chain unsaturation, cholesterol, and protein on the range and rate of wobbling were examined with model membranes. The dynamic structure of biological membranes was found to be basically similar to that of the bilayer of unsaturated phospholipid; proteins and cholesterol act mainly as barriers that reduce the angular range of wobbling motion.