Poly(ethylene glycol)−Lipid Conjugates Promote Bilayer Formation in Mixtures of Non-Bilayer-Forming Lipids

• Published in: Biochemistry (Easton) 1996-02, Vol.35 (8), p.2610-2617
• Main Authors: Holland, John W, Cullis, Pieter R, Madden, Thomas D
• Format: Article
• Language: English
• Subjects: Cholesterol - chemistry; Freeze Fracturing; In Vitro Techniques; Lipid Bilayers; Liposomes - chemistry; Magnetic Resonance Spectroscopy; Microscopy, Electron; Molecular Conformation; Phosphatidylethanolamines - chemistry; Phospholipids; Polyethylene Glycols - chemistry
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi951999j

The influence of poly(ethylene glycol)−lipid conjugates on phospholipid polymorphism has been examined using 31P-NMR and freeze−fracture electron microscopy. An equimolar mixture of dioleoylphosphatidylethanolamine (DOPE) and cholesterol adopts the hexagonal (HII) phase when hydrated under physiological conditions but can be stabilized in a bilayer conformation when a variety of PEG−lipid conjugates are included in the lipid mixture. These PEG conjugates produced an increase in the bilayer to hexagonal (HII) phase transition temperature and a broadening of the temperature range over which both phases coexisted. Further, the fraction of phospholipid adopting the bilayer phase increased with increasing mole fraction of PEG−lipid such that at 20 mole % DOPE−PEG2000 no HII phase phospholipid was observed up to at least 60 °C. Increasing the size of the PEG moiety from 2000 to 5000 Da (while maintaining the PEG−lipid molar ratio constant) increased the proportion of lipid in the bilayer phase. In contrast, varying the acyl chains of the PE anchor had no effect on polymorphic behavior. PEG−lipid conjugates in which ceramide provides the hydrophobic anchor also promoted bilayer formation in DOPE:cholesterol mixtures but at somewhat higher molar ratios compared to the corresponding PEG−PE species. The slightly greater effectiveness of the PE conjugates may result from the fact that these derivatives also possess a net negative charge. Phosphorus NMR spectroscopy indicated that a proportion of the phospholipid in DOPE:cholesterol:PEG−PE mixtures experienced isotropic motional averaging with this proportion being sensitive to both temperature and PEG molecular weight. Surprisingly, little if any isotropic signal was observed when PEG−ceramide was used in place of PEG−PE. Consistent with the 31P-NMR spectra, freeze−fracture electron microscopy showed the presence of small vesicles (diameter < 200 nm) and lipidic particles in DOPE:cholesterol mixtures containing PEG−PE. We conclude that the effects of PEG−lipid conjugates on DOPE:cholesterol mixtures are 2-fold. First, the complementary “inverted cone” shape of the conjugate helps to accommodate the “cone-shaped” lipids, DOPE and cholesterol, in the bilayer phase. Second, the steric hindrance caused by the PEG group inhibits close apposition of bilayers, which is a prerequisite for the bilayer to HII phase transition.