Abstract
Cryo-transmission electron microscopy has been used to investigate the phase behavior and aggregate structure in dilute aqueous mixtures of dioleoylphosphatidylethanolamine (DOPE) and poly(ethylene glycol)-phospholipids (PEG-lipids). It is shown that PEG-lipids (micelle-forming lipids) induce a lamellar phase in mixtures with DOPE (inverted hexagonal forming lipid). The amount of PEG-lipid that is needed to induce a pure dispersed lamellar phase, at physiological conditions, depends on the size of the PEG headgroup. In the transition region between the inverted hexagonal phase and the lamellar phase, particles with dense inner textures are formed. It is proposed that these aggregates constitute dispersed cubic phase particles. Above bilayer saturating concentration of PEG-lipid, small disks and spherical micelles are formed. The stability of DOPE/PEG-lipid liposomes, prepared at high pH, against a rapid drop of the pH was also investigated. It is shown that the density of PEG-lipid in the membrane, sufficient to prevent liposome aggregation and subsequent phase transition, depends on the size of the PEG headgroup. Below a certain density of PEG-lipid, aggregation and phase transition occurs, but the processes involved proceed relatively slow, over the time scale of weeks. This allows detailed studies of the aggregate structure during membrane fusion.
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Selected References
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- Allen T. M., Hong K., Papahadjopoulos D. Membrane contact, fusion, and hexagonal (HII) transitions in phosphatidylethanolamine liposomes. Biochemistry. 1990 Mar 27;29(12):2976–2985. doi: 10.1021/bi00464a013. [DOI] [PubMed] [Google Scholar]
- BLIGH E. G., DYER W. J. A rapid method of total lipid extraction and purification. Can J Biochem Physiol. 1959 Aug;37(8):911–917. doi: 10.1139/o59-099. [DOI] [PubMed] [Google Scholar]
- Basáez G., Ruiz-Argüello M. B., Alonso A., Goñi F. M., Karlsson G., Edwards K. Morphological changes induced by phospholipase C and by sphingomyelinase on large unilamellar vesicles: a cryo-transmission electron microscopy study of liposome fusion. Biophys J. 1997 Jun;72(6):2630–2637. doi: 10.1016/S0006-3495(97)78906-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Belsito S., Bartucci R., Montesano G., Marsh D., Sportelli L. Molecular and mesoscopic properties of hydrophilic polymer-grafted phospholipids mixed with phosphatidylcholine in aqueous dispersion: interaction of dipalmitoyl N-poly(ethylene glycol)phosphatidylethanolamine with dipalmitoylphosphatidylcholine studied by spectrophotometry and spin-label electron spin resonance. Biophys J. 2000 Mar;78(3):1420–1430. doi: 10.1016/S0006-3495(00)76695-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chernomordik L. Non-bilayer lipids and biological fusion intermediates. Chem Phys Lipids. 1996 Jul 15;81(2):203–213. doi: 10.1016/0009-3084(96)02583-2. [DOI] [PubMed] [Google Scholar]
- Cullis P. R., de Kruijff B. The polymorphic phase behaviour of phosphatidylethanolamines of natural and synthetic origin. A 31P NMR study. Biochim Biophys Acta. 1978 Oct 19;513(1):31–42. doi: 10.1016/0005-2736(78)90109-8. [DOI] [PubMed] [Google Scholar]
- Edwards K., Johnsson M., Karlsson G., Silvander M. Effect of polyethyleneglycol-phospholipids on aggregate structure in preparations of small unilamellar liposomes. Biophys J. 1997 Jul;73(1):258–266. doi: 10.1016/S0006-3495(97)78066-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ellens H., Bentz J., Szoka F. C. Destabilization of phosphatidylethanolamine liposomes at the hexagonal phase transition temperature. Biochemistry. 1986 Jan 28;25(2):285–294. doi: 10.1021/bi00350a001. [DOI] [PubMed] [Google Scholar]
- Ellens H., Siegel D. P., Alford D., Yeagle P. L., Boni L., Lis L. J., Quinn P. J., Bentz J. Membrane fusion and inverted phases. Biochemistry. 1989 May 2;28(9):3692–3703. doi: 10.1021/bi00435a011. [DOI] [PubMed] [Google Scholar]
- Gerasimov OV, Boomer JA, Qualls MM, Thompson DH. Cytosolic drug delivery using pH- and light-sensitive liposomes. Adv Drug Deliv Rev. 1999 Aug 20;38(3):317–338. doi: 10.1016/s0169-409x(99)00035-6. [DOI] [PubMed] [Google Scholar]
- Gustafsson J, Nylander T, Almgren M, Ljusberg-Wahren H. Phase Behavior and Aggregate Structure in Aqueous Mixtures of Sodium Cholate and Glycerol Monooleate. J Colloid Interface Sci. 1999 Mar 15;211(2):326–335. doi: 10.1006/jcis.1998.5996. [DOI] [PubMed] [Google Scholar]
- Helfrich W. Elastic properties of lipid bilayers: theory and possible experiments. Z Naturforsch C. 1973 Nov-Dec;28(11):693–703. doi: 10.1515/znc-1973-11-1209. [DOI] [PubMed] [Google Scholar]
- Holland J. W., Cullis P. R., Madden T. D. Poly(ethylene glycol)-lipid conjugates promote bilayer formation in mixtures of non-bilayer-forming lipids. Biochemistry. 1996 Feb 27;35(8):2610–2617. doi: 10.1021/bi951999j. [DOI] [PubMed] [Google Scholar]
- Holland J. W., Hui C., Cullis P. R., Madden T. D. Poly(ethylene glycol)--lipid conjugates regulate the calcium-induced fusion of liposomes composed of phosphatidylethanolamine and phosphatidylserine. Biochemistry. 1996 Feb 27;35(8):2618–2624. doi: 10.1021/bi952000v. [DOI] [PubMed] [Google Scholar]
- Kenworthy A. K., Simon S. A., McIntosh T. J. Structure and phase behavior of lipid suspensions containing phospholipids with covalently attached poly(ethylene glycol). Biophys J. 1995 May;68(5):1903–1920. doi: 10.1016/S0006-3495(95)80368-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kirpotin D., Hong K., Mullah N., Papahadjopoulos D., Zalipsky S. Liposomes with detachable polymer coating: destabilization and fusion of dioleoylphosphatidylethanolamine vesicles triggered by cleavage of surface-grafted poly(ethylene glycol). FEBS Lett. 1996 Jun 17;388(2-3):115–118. doi: 10.1016/0014-5793(96)00521-2. [DOI] [PubMed] [Google Scholar]
- Siegel D. P., Banschbach J. L. Lamellar/inverted cubic (L alpha/QII) phase transition in N-methylated dioleoylphosphatidylethanolamine. Biochemistry. 1990 Jun 26;29(25):5975–5981. doi: 10.1021/bi00477a014. [DOI] [PubMed] [Google Scholar]
- Siegel D. P., Epand R. M. The mechanism of lamellar-to-inverted hexagonal phase transitions in phosphatidylethanolamine: implications for membrane fusion mechanisms. Biophys J. 1997 Dec;73(6):3089–3111. doi: 10.1016/S0006-3495(97)78336-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Siegel D. P., Green W. J., Talmon Y. The mechanism of lamellar-to-inverted hexagonal phase transitions: a study using temperature-jump cryo-electron microscopy. Biophys J. 1994 Feb;66(2 Pt 1):402–414. doi: 10.1016/s0006-3495(94)80790-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Siegel D. P. The modified stalk mechanism of lamellar/inverted phase transitions and its implications for membrane fusion. Biophys J. 1999 Jan;76(1 Pt 1):291–313. doi: 10.1016/S0006-3495(99)77197-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Slepushkin V. A., Simões S., Dazin P., Newman M. S., Guo L. S., Pedroso de Lima M. C., Düzgüneş N. Sterically stabilized pH-sensitive liposomes. Intracellular delivery of aqueous contents and prolonged circulation in vivo. J Biol Chem. 1997 Jan 24;272(4):2382–2388. doi: 10.1074/jbc.272.4.2382. [DOI] [PubMed] [Google Scholar]
- Szleifer I., Gerasimov O. V., Thompson D. H. Spontaneous liposome formation induced by grafted poly(ethylene oxide) layers: theoretical prediction and experimental verification. Proc Natl Acad Sci U S A. 1998 Feb 3;95(3):1032–1037. doi: 10.1073/pnas.95.3.1032. [DOI] [PMC free article] [PubMed] [Google Scholar]