Molecular-scale structural and functional characterization of sparsely tethered bilayer lipid membranes

Abstract

Surface-tethered biomimetic bilayer membranes (tethered bilayer lipid membranes (tBLMs)) were formed on gold surfaces from phospholipids and a synthetic 1-thiahexa(ethylene oxide) lipid, WC14. They were characterized using electrochemical impedance spectroscopy, neutron reflection (NR), and Fourier-transform infrared reflection-absorption spectroscopy (FT-IRRAS) to obtain functional and structural information. The authors found that electrically insulating membranes (conductance and capacitance as low as 1 microS cm(-2) and 0.6 microF cm(-2), respectively) with high surface coverage (>95% completion of the outer leaflet) can be formed from a range of lipids in a simple two-step process that consists of the formation of a self-assembled monolayer (SAM) and bilayer completion by "rapid solvent exchange." NR provided a molecularly resolved characterization of the interface architecture and, in particular, the constitution of the space between the tBLM and the solid support. In tBLMs based on SAMs of pure WC14, the hexa(ethylene oxide) tether region had low hydration even though FT-IRRAS showed that this region is structurally disordered. However, on mixed SAMs made from the coadsorption of WC14 with a short-chain "backfiller," beta-mercaptoethanol, the submembrane spaces between the tBLM and the substrates contained up to 60% exchangeable solvent by volume, as judged from NR and contrast variation of the solvent. Complete and stable "sparsely tethered" BLMs (stBLMs) can be readily prepared from SAMs chemisorbed from solutions with low WC14 proportions. Phospholipids with unsaturated or saturated, straight or branched chains all formed qualitatively similar stBLMs.

Coarse-grained modeling of interactions of lipid bilayers with supports

“…To address this problem several methods have been developed to increase the spacing between the bilayer and substrate. 15,18,[28][29][30] Nonetheless, measurements of pure and mixed lipid membranes performed directly on supports remain a valuable source of information on the nature of lipid-lipid and lipid-solvent interactions.…”

“…7,8 Even interactions with whole cells have been studied. 9,10 Characterization of supported lipid bilayers has been accomplished with numerous methods including atomic force microscopy, 11 x-ray and neutron scattering and reflectometry, [12][13][14][15] nuclear magnetic resonance, 16 quartz crystal microbalance, 17 surface plasmon resonance, [18][19][20][21] ellipsometry, 6 electrical impedance spectroscopy, 15 and many versions of fluorescence microscopy such as fluorescence resonance energy transfer, 22 fluorescence recovery after photobleaching, 23 fluorescence interference contrast, 24,25 fluorescence correlation spectroscopy, 26 and total internal reflection fluorescence. 27 Phospholipid bilayers adsorb to hydrophilic surfaces to create a supported bilayer leaving a layer of water up to 5 Å thick-in addition to the hydration shell of the head groups-between the substrate and the head groups of the lipids in the proximal ͑closest to the solid͒ leaflet.…”

Myristoylation Restricts Orientation of the GRASP Domain on Membranes and Promotes Membrane Tethering

“…Formation of stBLMs-stBLMs were formed on solid substrates as described previously (19,20). The substrates, No.…”

“…Freshly gold-coated substrates were immersed in an ethanolic solution of 30% HC18 and 70% ␤-mercaptoethanol, at a total concentration of 0.2 mmol/liter, to form a mixed self-assembly monolayer. Rapid solvent exchange (19,22) or vesicle fusion was used to complete the formation of stBLMs.…”

New sources and instrumentation for neutrons in biology

“…If the SAM consists of pure WC14, the hexa(ethylene oxide) tether region was shown by NR to have low levels of hydration. However, if a mixed SAM is formed by coadsorption of WC14 with a short-chain "backfiller", β-mercaptoethanol, the open spaces between the tBLM and the substrates were shown to contain up to 60% exchangeable solvent by volume [76]. This made possible the functional reconstitution of the Staphylococcus aureus α-hemolysin channel protein in these model membranes [95].…”