Electric field-induced reorganization of two-component supported bilayer membranes

Abstract

Application of electric fields tangent to the plane of a confined patch of f luid bilayer membrane can create lateral concentration gradients of the lipids. A thermodynamic model of this steady-state behavior is developed for binary systems and tested with experiments in supported lipid bilayers. The model uses Flory's approximation for the entropy of mixing and allows for effects arising when the components have different molecular areas. In the special case of equal area molecules the concentration gradient reduces to a Fermi-Dirac distribution. The theory is extended to include effects from charged molecules in the membrane. Calculations show that surface charge on the supporting substrate substantially screens electrostatic interactions within the membrane. It also is shown that concentration profiles can be affected by other intermolecular interactions such as clustering. Qualitative agreement with this prediction is provided by comparing phosphatidylserine-and cardiolipin-containing membranes.Electric fields can be used to reorganize molecules in fluid lipid bilayer membranes. In 1977 Poo and Robinson (1) observed that application of an electric field caused the membrane-bound protein, Con A, to congregate on one side of a living cell. The distribution of protein at steady-state results from a balance between field-induced drift and diffusion within the confined membrane area (2, 3). The shape of a field-induced concentration profile thus contains information about the forces affecting molecules in the membrane. However, the small size and complexity of native cell membranes hinders analysis of these profiles. Recently, electric fields have been used to reorganize and concentrate lipids and proteins in supported bilayers where quantitative analysis is greatly simplified (4, 5).Supported bilayers can be created by spontaneous fusion of unilamellar phospholipid vesicles with an appropriate substrate such as silica (6, 7). The resulting membrane is typically separated from the solid substrate by a thin (10 Å) layer of water (8-10) and retains many of the properties of free membranes, including lateral fluidity. The fluidity is macroscopically long-range with mobile components of both leaflets of the bilayer freely diffusing over the entire surface of the support. The diffusive mixing and flow of molecules within the membrane can be confined by imposing barriers on this lateral motion. Manually scratching the membrane-coated surface can effectively create such barriers. Alternatively, more precisely partitioned membranes have been formed by using pre-patterned substrates to impose structure onto the membrane (11). Application of electric fields tangent to the plane of supported membranes partitioned by either method causes molecules to reorganize within the confined membrane corrals. The size and geometry of these corrals can be tailored to allow precise analysis of the field-induced concentration profiles.In this work, a general description of the steady-state electric field-induced reorganization of ...

Polymer-Supported Lipid Bilayers on Benzophenone-Modified Substrates

“…Under normal conditions, the electric-field-induced molecular reorganization and the thermal energy of the dye molecule result in a smooth distribution of fluorescence intensity between the two barriers after reaching the steady state. 35 The fluorescence signal at the barrier closer to the cathode (negative end) should be negligible, if all lipids in the distal layer are mobile. However, it appears that there is a significant immobile fraction in the polymer-supported lipid layers.…”

Micropattern Formation in Supported Lipid Membranes

“…These results suggest a critical demixing coefficient for cardiolipin/egg-PC mixtures corresponding to a critical temperature for spontaneous lateral phase separation, T c , of 75 K. It is interesting to compare this result to similar experiments carried out on mixtures of phosphatidylserine (PS) and egg-PC where nearly ideal mixing (T c ) 0 K) was observed. 39 An advantage of the dilute probe method is that subtle changes in the concentration profiles of the major components can produce exaggerated differences in the probe concentration profile. This effect is particularly prominent in the case at hand.…”

“…32,39 The rearrangement of membrane components after deposition offers a number of capabilities including directed concentration of lipids or proteins 40 and generation of continuous concentration gradients.…”

Single-particle tracking of murine polyoma virus-like particles on live cells and artificial membranes

“…Artificial lipid bilayers were prepared by the vesicle drop method (23). Briefly, dioleoylphosphatidylcholine, f luorescein-dipalmitoylphosphatidylethanolamine (Fl-DPPE) (Avanti Polar Lipids) and GD1a (Hytest, Turku, Finland) were dissolved in chloroform:methanol (4:1), dried under nitrogen flow, and kept under vacuum for at least 2 h. The dried lipid mixture was resuspended in 50 mM Tris͞0.2 M NaCl͞2 mM CaCl 2 , pH 7.4.…”

“…The bilayers were generated on the surface of glass coverslips by deposition of small unilamellar liposomes (23). The liposomes were composed of dioleoylphosphatidylcholine, with 0.07 mole % of GD1a and 1 mole % of FL-DPPE.…”