title: Giant unilamellar vesicles electroformed from native membranes and organic lipid mixtures under physiological conditions.
abstract: In recent years, giant unilamellar vesicles (GUVs) have become objects of intense scrutiny by chemists, biologists, and physicists who are interested in the many aspects of biological membranes. In particular, this "cell size" model system allows direct visualization of particular membrane-related phenomena at the level of single vesicles using fluorescence microscopy-related techniques. However, this model system lacks two relevant features with respect to biological membranes: 1), the conventional preparation of GUVs currently requires very low salt concentration, thus precluding experimentation under physiological conditions, and 2), the model system lacks membrane compositional asymmetry. Here we show for first time that GUVs can be prepared using a new protocol based on the electroformation method either from native membranes or organic lipid mixtures at physiological ionic strength. Additionally, for the GUVs composed of native membranes, we show that membrane proteins and glycosphingolipids preserve their natural orientation after electroformation. We anticipate our result to be important to revisit a vast variety of findings performed with GUVs under low- or no-salt conditions. These studies, which include results on artificial cell assembly, membrane mechanical properties, lipid domain formation, partition of membrane proteins into lipid domains, DNA-lipid interactions, and activity of interfacial enzymes, are likely to be affected by the amount of salt present in the solution.
authors: Unidad de Biofísica (Centro Mixto CSIC-UPV/EHU), Departamento de Bioquímica, Universidad del País Vasco, 48080 Bilbao, Spain.
authors: L-Ruth
authors: LR
authors: Montes
authors: Alicia
authors: A
authors: Alonso
authors: Felix M
authors: FM
authors: Goñi
authors: Luis A
authors: LA
authors: Bagatolli
mesh_terms: D001703:Biophysics / Q000379:methods*; D002118:Calcium / Q000378:metabolism; D004594:Electrophysiology / Q000379:methods; D004910:Erythrocyte Membrane / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry; D008055:Lipids / Q000737:chemistry*; D008081:Liposomes; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D010713:Phosphatidylcholines / Q000737:chemistry; D017434:Protein Structure, Tertiary; D012492:Salts / Q000494:pharmacology
chemical_list: D008051:Lipid Bilayers; D008055:Lipids; D008081:Liposomes; D010713:Phosphatidylcholines; D012492:Salts; D002118:Calcium
references: Chem Phys Lipids. 2000 Apr;105(2):135-47
references: 10823462
references: Biochim Biophys Acta. 1999 Dec 6;1473(1):247-66
references: 10580143
references: Chemphyschem. 2003 Jul 14;4(7):699-704
references: 12901301
references: Proc Natl Acad Sci U S A. 2003 Sep 16;100(19):10718-21
references: 12963816
references: Annu Rev Biophys Biomol Struct. 2003;32:257-83
references: 12543707
references: Annu Rev Biophys Biomol Struct. 2004;33:269-95
references: 15139814
references: J Biol Chem. 2004 Sep 24;279(39):40715-22
references: 15231828
references: Annu Rev Cell Dev Biol. 2004;20:839-66
references: 15473862
references: J Cell Physiol. 1969 Feb;73(1):49-60
references: 5765779
references: Science. 1972 Feb 18;175(4023):720-31
references: 4333397
references: Methods Enzymol. 1974;31:172-80
references: 4370662
references: Proc Natl Acad Sci U S A. 1987 Jul;84(13):4475-9
references: 3474616
references: J Biol Chem. 1994 Mar 4;269(9):6347-54
references: 8119984
references: Blood. 1995 Sep 1;86(5):1983-91
references: 7655025
references: Biophys J. 1996 Dec;71(6):3242-50
references: 8968594
references: Biophys J. 1997 Jun;72(6):2413-29
references: 9168019
references: Curr Opin Chem Biol. 1998 Dec;2(6):726-32
references: 9914194
references: Can J Biochem Physiol. 1959 Aug;37(8):911-7
references: 13671378
references: Biophys J. 2005 Sep;89(3):1874-81
references: 15994901
references: Biochim Biophys Acta. 2005 Dec 30;1746(3):172-85
references: 16043244
references: Mol Membr Biol. 2006 Jan-Feb;23(1):29-39
references: 16611578
references: Biophys J. 2006 Jun 15;90(12):4437-51
references: 16565051
references: Biochim Biophys Acta. 2006 Oct;1758(10):1541-56
references: 16854370
references: Proc Natl Acad Sci U S A. 2007 Feb 27;104(9):3165-70
references: 17360623
references: Biophys J. 2001 Mar;80(3):1417-28
references: 11222302