In-plane homogeneity and lipid dynamics in tethered bilayer lipid membranes (tBLMs).

Siddharth Shenoy,David J Vanderah,James Fitzpatrick,Markus Deserno,Radu Moldovan,Mathias Lösche

doi:10.1039/b919988h

Abstract

Tethered bilayer lipid membranes (tBLMs) were prepared by the self-assembly of thiolated lipidic anchor molecules on gold, followed by phospholipid precipitation via rapid solvent exchange. They were characterized by their in-plane structure, dynamics and dielectric properties. We find that the in-plane homogeneity and resistivity of the tBLMs depend critically on a well-controlled sample environment during the rapid solvent-exchange procedure. The in-plane dynamics of the systems, assessed by fluorescence correlation spectroscopy (FCS) as the diffusivity of free, labeled phospholipid dissolved in the membrane, depend on the density of the lipidic anchors in the bilayer leaflet proximal to the substrate as well as on details of the molecular structure of the anchor lipid. In DOPC tBLMs in which tethers are laterally dilute (sparsely tethered bilayer lipid membranes, stBLMs), measured diffusivities, D ≈ 4 μm(2) s(-1), are only slightly greater than those reported in physisorbed bilayers (M. Przybylo, J. Sykora, J. Humpolíckova, A. Benda, A. Zan and M. Hof, Langmuir, 2006, 22, 9096-9099). However, when we distinguish label diffusion in the proximal and in the distal bilayer leaflets, we observe distinct diffusivities, D ≈ 2 μm(2) s(-1) and 7 μm(2) s(-1), respectively. The value observed in the distal leaflet is identical to that in free membranes. stBLMs completed with phytanoyl lipids (DPhyPC) show consistently lower label diffusivity than those completed with unsaturated chains (DOPC). As the length of the tether chain increases, a reduction in the apparent diffusivity is observed, which we interpret as an increased propensity of the proximal bilayer leaflet to host free lipid. We also investigated preparation conditions that control whether the tBLMs are laterally homogeneous, as assessed by optical microscopy. In laterally heterogeneous bilayers, the label diffusivity varies only by a factor of ~2 to 4, indicating that the regions in the bilayers with different label solubilities do not correspond to distinct phases, such as a fluid phase coexisting with a gel phase.

Highlights

Lipid membranes separate the interior space of cells from the outside world and compartmentalize the cytoplasm into organelles
Because DPhyPC and diffusivity than those completed with unsaturated chains (DOPC) Tethered bilayer lipid membranes (tBLMs) are frequently used in studies of protein–membrane interactions, we explored in this work conditions that lead to homogeneous bilayers in tBLMs composed of these lipids, and have characterized their in-plane dynamics
Supported bilayers prepared on various substrates by vesicle fusion can be in-plane homogeneous,[32] which is useful for patterning or for studies of phase separation induced by chemical heterogeneities. dtBLMS and sparsely tethered bilayer lipid membrane (stBLM) prepared from DOPC or DPhyPC solutions by rapid solvent exchange compare well with these alternate preparations

Summary

Introduction

Lipid membranes separate the interior space of cells from the outside world and compartmentalize the cytoplasm into organelles. Among the different model membranes that have been developed, solid supported membranes[3] are robust and are amenable to many physical characterization techniques due to their This journal is a The Royal Society of Chemistry 2010 planar geometry. They are prepared by vesicle fusion[4] and conveniently studied with tools such as fluorescence microscopy,[5] X-ray or neutron reflection[6] for structural characterization while techniques such as surface-plasmon resonance (SPR)[7] are frequently used to quantify molecular interactions that occur at such membranes. It is straightforward to quantify the dielectric constant, 3, of the hydrophobic interior of the bilayers by a combination of EIS and neutron reflectometry.[9]

Results

Discussion

Conclusion