Artificial Lipid Membranes: Past, Present, and Future

Abstract

The multifaceted role of biological membranes prompted early the development of artificial lipid-based models with a primary view of reconstituting the natural functions in vitro so as to study and exploit chemoreception for sensor engineering. Over the years, a fair amount of knowledge on the artificial lipid membranes, as both, suspended or supported lipid films and liposomes, has been disseminated and has helped to diversify and expand initial scopes. Artificial lipid membranes can be constructed by several methods, stabilized by various means, functionalized in a variety of ways, experimented upon intensively, and broadly utilized in sensor development, drug testing, drug discovery or as molecular tools and research probes for elucidating the mechanics and the mechanisms of biological membranes. This paper reviews the state-of-the-art, discusses the diversity of applications, and presents future perspectives. The newly-introduced field of artificial cells further broadens the applicability of artificial membranes in studying the evolution of life.

Keywords: Langmuir-Blodgett films; artificial cells; artificial lipid membranes; biosensors; drug delivery; ion channel monitoring; liposomes; nanoelectrodes; self-assembly; tethered membranes.

Figure 1

The movement of lipids within a bilayer takes many forms, energy-driven or spontaneous. The rates for rotational and lateral movement depend on the biophysics of the membrane and the lipids. Lipid translocation from one leaflet to the other can be protein-mediated and energy independent (e.g., via scramblases) or energy-dependent (e.g., via translocases).

Figure 2

Schematic of conventional approaches for preparing freely-suspended bilayer lipid membranes: (1) Painting technique: A droplet of lipid solution is painted into a small aperture in a hydrocarbon or Teflon partition; the thinning of the lipids into a bilayer occurs spontaneously (black lipid film). (2) Folding or monolayer opposition technique: (a) A hydrophobic septum, punched to produce an aperture of a few millimeters, separates two electrolyte compartments. (b) The electrolyte is removed from both compartments; lipids are added and accumulate at the surface of the aqueous phase. When the electrolyte level is raised in one compartment, one lipid monolayer is forced to develop around the aperture. (c) Replenishing the other compartment with electrolyte, attaches the second monolayer (like zipping); during zipping the solvent entrapped into the hydrophobic area is squeezed by the hydrostatic pressure applied on both sides of the bilayer towards the rim of the aperture (Plateau-Gibbs border). (3) Finally, an equilibrium is reached between membrane thinning (due to hydrostatic pressure) and unzipping (due to the solvent that tries to relocate at the middle of the bilayer). As evident, the slightest vibration disturbs this equilibrium and the bilayer collapses.

Figure 3

Some basic biochemical systems that can be reconstituted within an artificial lipid bilayer and further engineered into a diagnostic system (biosensor). The transduction of the biochemical information into a detector signal can be achieved with electrochemical, optical, piezoelectric or magnetic sensors. (a) bioaffinity interfaces can be constructed using enzymes or receptors, adsorbed on the membrane surface; the whole system can be optimized electrochemically (using redox amplifiers) or optically (using fluorescent tags). (b) The monitoring of immunochemical reactions follows similar methodology and further allows for the development of more advanced and rapid signal propagation systems (e.g., using enzymes, tagging or radiochemistry). (c) Many channel-forming proteins have been reconstituted within a bilayer; small molecules can flow through the channel, but in most cases some selectivity rules apply that make possible the development of a detection system (e.g., gramicidin channels transport potassium ions faster than sodium while valinomycin channels transport only potassium ions). (d) Single- or double-stranded DNA can adsorb on the membrane surface to investigate the effects of various adducts or to detect mutagens.

Figure 4

Schematic of two simple approaches in supporting the bilayer: self-assembly (a) and tethering (b). (a) Self-assembly: (1) A metal wire (freshly cut) is dipped into lipid solution; when withdrawn, a small drop of lipid solution is attached around the tip. (2) The electrode is immersed in electrolyte solution. (3) On dipping into the aqueous phase, lipids gather spontaneously at the tip of the electrode (pushing along electrolyte molecules) to form a monolayer that drives other lipid molecules to cover it at the top; the assembly is finally thinned to a bilayer. (b) Tethering: Thiolipids (1) or hydrogels (2) can be used as the anchoring layer. Alternatively, proteins can be used either as a lattice (3) or as a layer (4).

Figure 5

Droplet Interface Bilayers (DIBs) can be formed by two techniques, lipid-out and lipid-in. (a) Lipid-out technique: Ag/AgCl electrodes coated with agarose are loaded with aqueous droplets and dipped in oil-lipid solution (system setting phase). A 30-min stabilization phase is necessary for the formation of monolayers at the oil-water interface around the droplets. When the monolayers collide, they form a bilayer at the contact point (bilayer lipid formation phase). (b) Lipid-in technique: Similarly, the electrodes are loaded with aqueous droplets that contain vesicles and dipped in oil solution (system setting phase). A 5-min stabilization phase is necessary for the vesicles to fuse with the oil-water interface and form the monolayers, which subsequently brought into contact to form the bilayers.

Figure 6

Simplified schematic of liposome-mediated V. harveyi bioluminescence induction. The increase in the pH value at the suspension medium triggers the production of sugars inside the vesicles, which cross the membrane and diffuse into the suspension medium. There, they are readily detected by V. harveyi; the detection provokes the Vibrio cellular mechanism for the production of bioluminescent proteins.