Incorporation of alpha-hemolysin in different tethered bilayer lipid membrane architectures.

Tethered bilayer lipid membranes are stable solid supported model membrane systems. They can be used to investigate the incorporation and function of membrane proteins. In order to study ion translocation mediated via incorporated proteins, insulating membranes are necessary. The architecture of the membrane can have an important effect on both the electrical properties of the lipid bilayer as well as on the possibility to functionally host proteins. Alpha-hemolysin pores have been functionally incorporated into a tethered bilayer lipid membrane coupled to a gold electrode. The protein incorporation has been monitored optically and electrically and the influence of the molecular structure of the anchor lipids on the insertion properties has been investigated.

Anchor-lipid monolayers at the air-water interface; prearranging of model membrane systems.

Model membrane systems are gaining more and more interest both for basic studies of membrane-related processes as well as for biotechnological applications. Several different model systems have been reported among which the tethered bilayer lipid membranes (tBLMs) form a very attractive and powerful architecture. In all the proposed architectures, a control of the lateral organization of the structures at a molecular level is of great importance for an optimized preparation. For tBLMs, a homogeneous and not too dense monolayer is required to allow for the functional incorporation of complex membrane proteins. We present here an alternative approach to the commonly used self-assembly preparation. Lipids are spread on the air-water interface of a Langmuir film balance and form a monomolecular film. This allows for a better control of the lateral pressure and distribution for subsequent transfer to solid substrates. In this paper, we describe the properties of the surface monolayer, in terms of surface pressure, structure of the lipid molecule, content of lipid mixtures, temperature, and relaxations features. It is shown that a complete mixing of anchor-lipids and free lipids can be achieved. Furthermore, an increase of the spacer lengths and a decrease of the temperature lead to more compact films. This approach is a first step toward the fully controlled assembly of a model membrane system.

Functional incorporation of the pore forming segment of AChR M2 into tethered bilayer lipid membranes.

Tethered bilayer lipid membranes (tBLMs) are robust and flexible model platforms for the investigation of various membrane related processes. They are especially suited to study the incorporation and function of ion channel proteins, where a high background resistance of the membrane is essential. Synthetic M2 peptides, analogues of the transmembrane fragment of the acetylcholine receptor, could be incorporated into two different membrane architectures. The functional reconstitution and the formation of a conducting pore are shown by electrochemical impedance spectroscopy (EIS). The pore is selective for small monovalent cations, while bulky ions cannot pass. This is a significant step towards a novel biosensing approach. We envision a device, where a stable and insulating membrane would be attached to an electronic read-out unit and embedded proteins would serve as actual sensing units.

A molecular toolkit for highly insulating tethered bilayer lipid membranes on various substrates.

Tethered bilayer lipid membranes (tBLMs) are promising model architectures that mimic the structure and function of natural biomembranes. They provide a fluid, stable, and electrically sealing platform for the study of membrane related processes, specifically, the function of incorporated membrane proteins. This paper presents a generic approach toward the synthesis of functional tBLMs adapted for application to various surfaces. The central element of a tethered membrane consists of a lipid bilayer. Its proximal layer is covalently attached via a spacer unit to a solid support, either gold or silicon oxide. The membranes are characterized optically by using surface plasmon resonance spectroscopy (SPR) or ellipsometry and electrically by using electrochemical impedance spectroscopy (EIS). The bilayer membranes obtained show high electrical barrier properties and can be used to incorporate and study small membrane proteins in a functional form.