Characterization of horizontal lipid bilayers as a model system to study lipid phase separation

Research output: Contribution to journalResearch articleContributedpeer-review

Contributors

  • Alf Honigmann - , University Osnabruck (Author)
  • Claudius Walter - , Ionovation GmbH (Author)
  • Frank Erdmann - , University Osnabruck, University of Münster (Author)
  • Christian Eggeling - , Max Planck Institute for Biophysical Chemistry (Karl Friedrich Bonhoeffer Institute) (Author)
  • Richard Wagner - , University Osnabruck (Author)

Abstract

Artificial lipid membranes are widely used as a model system to study single ion channel activity using electrophysiological techniques. In this study, we characterize the properties of the artificial bilayer system with respect to its dynamics of lipid phase separation using single-molecule fluorescence fluctuation and electrophysiological techniques. We determined the rotational motions of fluorescently labeled lipids on the nanosecond timescale using confocal time-resolved anisotropy to probe the microscopic viscosity of the membrane. Simultaneously, long-range mobility was investigated by the lateral diffusion of the lipids using fluorescence correlation spectroscopy. Depending on the solvent used for membrane preparation, lateral diffusion coefficients in the range Dlat = 10-25 μm2/s and rotational diffusion coefficients ranging from Drot = 2.8 - 1.4 × 107 s-1 were measured in pure liquid-disordered (Ld) membranes. In ternary mixtures containing saturated and unsaturated phospholipids and cholesterol, liquid-ordered (L0) domains segregated from the Ld phase at 23°0C. The lateral mobility of lipids in L0 domains was around eightfold lower compared to those in the Ld phase, whereas the rotational mobility decreased by a factor of 1.5. Burstintegrated steady-state anisotropy histograms, as well as anisotropy imaging, were used to visualize the rotational mobility of lipid probes in phase-separated bilayers. These experiments and fluorescence correlation spectroscopy measurements at different focal diameters indicated a heterogeneous microenvironment in the L0 phase. Finally, we demonstrate the potential of the optoelectro setup to study the influence of lipid domains on the electrophysiological properties of ion channels. We found that the electrophysiological activity of gramicidin A (gA), a well-characterized ion channel-forming peptide, was related to lipid-domain partitioning. During liquid-liquid phase separation, gA was largely excluded from L0 domains. Simultaneously, the number of electrically active gA dimers increased due to the increased surface density of gA in the L d phase.

Details

Original languageEnglish
Pages (from-to)2886-2894
Number of pages9
JournalBiophysical journal
Volume98
Issue number12
Publication statusPublished - 16 Jun 2010
Peer-reviewedYes
Externally publishedYes

External IDs

PubMed 20550901
ORCID /0000-0003-0475-3790/work/190134729

Keywords

ASJC Scopus subject areas