Biophys J, February 2001, p. 776-788, Vol. 80, No. 2

Fluid-Fluid Membrane Microheterogeneity: A Fluorescence Resonance Energy Transfer Study

Luís M. S. Loura,^* Aleksandre Fedorov,^* and Manuel Prieto^*

 ^*Centro de Química-Física Molecular, Instituto Superior Técnico, P-1049-001 Lisboa, Portugal,  Departamento de Química, Universidade de Évora, P-7000-671 Évora, Portugal, and  On leave from the Vavilov Optical Institute, St. Petersburg, Russia

Large unilamellar vesicles of dimyristoylphosphatidylcholine/cholesterol mixtures were studied using fluorescence techniques (steady-state fluorescence intensity and anisotropy, fluorescence lifetime, and fluorescence resonance energy transfer (FRET)). Three compositions (cholesterol mole fraction 0.15, 0.20, and 0.25) and two temperatures (30 and 40°C) inside the coexistence range of liquid-ordered (l_o) and liquid-disordered (l_d) phases were investigated. Two common membrane probes, N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-dimyristoylphosphatidylethanolamine (NBD-DMPE) and N-(lissamine^TM-rhodamine B)-dimyristoylphosphatidylethanolamine (Rh-DMPE), which form a FRET pair, were used. The l_o/l_d partition coefficients of the probes were determined by individual photophysical measurements and global analysis of time-resolved FRET decays. Although the acceptor, Rh-DMPE, prefers the l_d phase, the opposite is observed for the donor, NBD-DMPE. Accordingly, FRET efficiency decreases as a consequence of phase separation. Comparing the independent measurements of partition coefficient, it was possible to detect very small domains (<20 nm) of l_o in the cholesterol-poor end of the phase coexistence range. In contrast, domains of l_d in the cholesterol-rich end of the coexistence range have comparatively large size. These observations are probably related to different processes of phase separation, nucleation being preferred in formation of l_o phase from initially pure l_d, and domain growth being faster in formation of l_d phase from initially pure l_o.

Biophys J, February 2001, p. 776-788, Vol. 80, No. 2
© 2001 by the Biophysical Society   0006-3495/01/02/776/13  $2.00

This article has been cited by other articles:

				A. Hodzic, M. Rappolt, H. Amenitsch, P. Laggner, and G. Pabst Differential Modulation of Membrane Structure and Fluctuations by Plant Sterols and Cholesterol Biophys. J., May 15, 2008; 94(10): 3935 - 3944. [Abstract] [Full Text] [PDF]

				F. Fernandes, L. M. S. Loura, F. J. Chichon, J. L. Carrascosa, A. Fedorov, and M. Prieto Role of Helix 0 of the N-BAR Domain in Membrane Curvature Generation Biophys. J., April 15, 2008; 94(8): 3065 - 3073. [Abstract] [Full Text] [PDF]

				G. S. Longo, D. H. Thompson, and I. Szleifer Stability and Phase Separation in Mixed Monopolar Lipid/Bolalipid Layers Biophys. J., October 15, 2007; 93(8): 2609 - 2621. [Abstract] [Full Text] [PDF]

				R. F. M. de Almeida, J. Borst, A. Fedorov, M. Prieto, and A. J. W. G. Visser Complexity of Lipid Domains and Rafts in Giant Unilamellar Vesicles Revealed by Combining Imaging and Microscopic and Macroscopic Time-Resolved Fluorescence Biophys. J., July 15, 2007; 93(2): 539 - 553. [Abstract] [Full Text] [PDF]

				K. B. Towles, A. C. Brown, S. P. Wrenn, and N. Dan Effect of Membrane Microheterogeneity and Domain Size on Fluorescence Resonance Energy Transfer Biophys. J., July 15, 2007; 93(2): 655 - 667. [Abstract] [Full Text] [PDF]

				L. C. Silva, R. F. M. de Almeida, B. M. Castro, A. Fedorov, and M. Prieto Ceramide-Domain Formation and Collapse in Lipid Rafts: Membrane Reorganization by an Apoptotic Lipid Biophys. J., January 15, 2007; 92(2): 502 - 516. [Abstract] [Full Text] [PDF]

				E. Falck, J. T. Hautala, M. Karttunen, P. K. J. Kinnunen, M. Patra, H. Saaren-Seppala, I. Vattulainen, S. K. Wiedmer, and J. M. Holopainen Interaction of Fusidic Acid with Lipid Membranes: Implications to the Mechanism of Antibiotic Activity Biophys. J., September 1, 2006; 91(5): 1787 - 1799. [Abstract] [Full Text] [PDF]

				P. V. Nazarov, R. B. M. Koehorst, W. L. Vos, V. V. Apanasovich, and M. A. Hemminga FRET Study of Membrane Proteins: Simulation-Based Fitting for Analysis of Membrane Protein Embedment and Association Biophys. J., July 15, 2006; 91(2): 454 - 466. [Abstract] [Full Text] [PDF]

				F. Fernandes, L. M. S. Loura, A. Fedorov, and M. Prieto Absence of clustering of phosphatidylinositol-(4,5)-bisphosphate in fluid phosphatidylcholine J. Lipid Res., July 1, 2006; 47(7): 1521 - 1525. [Abstract] [Full Text] [PDF]

				S. L. Veatch, K. Gawrisch, and S. L. Keller Closed-Loop Miscibility Gap and Quantitative Tie-Lines in Ternary Membranes Containing Diphytanoyl PC Biophys. J., June 15, 2006; 90(12): 4428 - 4436. [Abstract] [Full Text] [PDF]

				V. Pata and N. Dan Effect of Membrane Characteristics on Phase Separation and Domain Formation in Cholesterol-Lipid Mixtures Biophys. J., February 1, 2005; 88(2): 916 - 924. [Abstract] [Full Text] [PDF]

				D. A. Redfern and A. Gericke Domain Formation in Phosphatidylinositol Monophosphate/Phosphatidylcholine Mixed Vesicles Biophys. J., May 1, 2004; 86(5): 2980 - 2992. [Abstract] [Full Text] [PDF]

				S. L. Veatch and S. L. Keller Separation of Liquid Phases in Giant Vesicles of Ternary Mixtures of Phospholipids and Cholesterol Biophys. J., November 1, 2003; 85(5): 3074 - 3083. [Abstract] [Full Text] [PDF]

				C. Madeira, L. M. S. Loura, M. R. Aires-Barros, A. Fedorov, and M. Prieto Characterization of DNA/Lipid Complexes by Fluorescence Resonance Energy Transfer Biophys. J., November 1, 2003; 85(5): 3106 - 3119. [Abstract] [Full Text] [PDF]

				R. F. M. de Almeida, A. Fedorov, and M. Prieto Sphingomyelin/Phosphatidylcholine/Cholesterol Phase Diagram: Boundaries and Composition of Lipid Rafts Biophys. J., October 1, 2003; 85(4): 2406 - 2416. [Abstract] [Full Text] [PDF]