Electric field-driven transformations of a supported model biological membrane--an electrochemical and neutron reflectivity study

Abstract

A mixed bilayer of cholesterol and dimyristoylphosphatidylcholine has been formed on a gold-coated block of quartz by fusion of small unilamellar vesicles. The formation of this bilayer lipid membrane on a conductive surface allowed us to study the influence of the support's surface charge on the structure and hydration of the bilayer lipid membrane. We have employed electrochemical measurements and the specular reflection of neutrons to measure the thickness and water content in the bilayer lipid membrane as a function of the charge on the support's surface. When the surface charge density is close to zero, the lipid vesicles fuse directly on the surface to form a bilayer with a small number of defects and hence small water content. When the support's surface is negatively charged the film swells and incorporates water. When the charge density is more negative than -8 micro C cm(-2), the bilayer starts to detach from the metal surface. However, it remains in a close proximity to the metal electrode, being suspended on a thin cushion of the electrolyte. The field-driven transformations of the bilayer lead to significant changes in the film thicknesses. At charge densities more negative than -20 micro C cm(-2), the bilayer is approximately 37 A thick and this number is comparable to the thickness determined for hydrated multilayers of dimyristoylphosphatidylcholine from x-ray diffraction experiments. The thickness of the bilayer decreases at smaller charge densities to become equal to approximately 26 A at zero charge. This result indicates that the tilt of the acyl chains with respect to the bilayer normal changes from approximately 35 degrees to 59 degrees by moving from high negative charges (and potentials) to zero charge on the metal.

FIGURE 1

Schematic of the electrochemical cell configuration used in the neutron reflectivity experiments.

FIGURE 2

X-ray reflectivity as a function of momentum transfer vector for the Au/Cr-covered quartz block. Curve corresponds to the measurement made before electrochemistry. The curve for the postelectrochemical measurement is essentially identical and is omitted for clarity. The dotted line represents the results of the modeling. The inset shows the electron density profile for the model that best fits the data.

FIGURE 3

Differential capacitance curves obtained for a Au(111) single crystal using an AC perturbation of 5 mV, 25 Hz for 50 mM NaF only (———), and vesicles of a 7:3 mixture of DMPC and cholesterol in 50 mM NaF (- - - -).

FIGURE 4

The surface charge density on the gold electrode surface plotted versus the electrode potential for ▪, the background (50 mM NaF supporting electrolyte), and •, the mixed 7:3 DMPC/cholesterol bilayer spread from the vesicle solution. (Inset) The surface pressure versus electrode potential plot as calculated from the charge density data.

FIGURE 5

Experimentally determined reflectivity curves (points with associated error bars) for E = 50 mV in 50 mM NaF in D₂O. (Curve 1) The film-free electrode surface. (Curve 2) The electrode covered by a bilayer of the 7:3 mixture of h-DMPC and cholesterol. (Inset) Plot of RQ⁴ vs. Q_z calculated from the data presented in the main section of this figure. Solid line shows the calculated reflectivity curves from the best-fit model whose parameters are given in Tables 2 and 3.

FIGURE 6

(A–C) RQ⁴ vs. Q_z plots for a bilayer of the 7:3 mixture of h-DMPC and cholesterol in 50 mM NaF in D₂O; (A) at E = −375 mV; (B) at E = −500 mV; and (C) at E = −600 mV. Points with associated error bars show the experimental data. Solid lines show the reflectivity calculated from the parameters obtained from the fitting procedure. (D) The SLD profiles for the interface for E = 50 mV (▪); E = −375 mV (•); E = −500 mV (▴); and E = −600 mV (▾). The best-fit model parameters corresponding to the SLD profiles are listed in Tables 2 and 3.

FIGURE 7

(A–C) RQ⁴ vs. Q_z plots for a bilayer of the 7:3 mixture of h-DMPC and cholesterol in 50 mM NaF in D₂O, (A) at E = −700 mV; (B) at E = −800 mV; and (C) at E = −950 mV. Points with associated error bars show the experimental data. Solid lines show the reflectivity calculated from the parameters obtained from the fitting procedure. (D) The SLD profiles for the interface for E = −700 mV (▪); E = −800 mV (•); and E = −950 mV (▴). The best-fit model parameters corresponding to the SLD profiles are listed in Tables 2 and 3.

FIGURE 8

(A) RQ⁴ vs. Q_z plot for a bilayer of the 7:3 mixture of d-DMPC and cholesterol in 50 mM NaF in H₂O at E = −800 mV. Points with associated error bars show the experimental data. Solid line shows the reflectivity calculated from the SLD profile presented in B. The best-fit model parameters corresponding to the SLD profile are listed in Tables 2 and 3.

FIGURE 9

Pictorial description of the changes in the structure of the mixed DMPC-cholesterol bilayer deposited at the electrode surface as a function of the applied potential. We emphasize that this figure is not intended as a model of the biomimetic film. Note that the bilayer is in equilibrium with vesicles in the solution.

FIGURE 10

Comparison of the change in hydrocarbon tilt angle as a function of the electrode potential for a mixed DMPC/cholesterol (7:3) film spread on a gold electrode surface from a vesicle solution. The tilt angles were measured in the present work using neutron reflectivity (▴), and independently evaluated (X. Bin, I. Zawisza, S. Horswell, and J. Lipkowski, unpublished material) from IR results (▪).

FIGURE 11

Schematic representation of packing of the DMPC molecules, (A), at E < −600 mV, where the bilayer is suspended on a cushion of solvent, and (B), at E > −500 mV, where the bilayer is directly adsorbed at the metal surface (adopted from Hauser et al., 1981).