Membrane bilayers, applications

There has been a surge of research activity in the physical chemistry of membranes, bilayers, and vesicles. In addition to the fundamental interest in cell membranes and phospholipid bilayers, there is tremendous motivation for the design of supported membrane biosensors for medical and pharmaceutical applications (see the recent review by Sackmann [64]). This subject, in particular its biochemical aspects, is too vast for full development here we will only briefly discuss some of the more physical aspects of these systems. The reader is referred to the general references and some additional reviews [65-69]. 

The first approach has successfully been applied to the study of amorphous as well as to macroscopically ordered solids. Examples of applications include the determination of backbone geometries in fibrous proteins [4] or the determination of protein-backbone, side-chain, and bound-ligand orientation with respect to the membrane normal in membrane-bound proteins [5-8]. Membranes, bilayers, bicelles, or liposomes are neither solid nor liquid systems but have aspects of both and are sometimes liquid crystalline. In most of these systems, time-independent anisotropic interactions play an important role,... 

Usually amphiphilic in nature, the latter form the transmembrane tunnel by recognition and organization within the membrane bilayer. Both systems require the application of principles evolved in studies of supramolecular chemistry. 

In conclusion, supported bilayers have evolved into a reliable model membrane system since their first inception almost a quarter century ago. Numerous basic research questions regarding the structure and function of biological membranes and applications that range from biosensing to proteomic analyses of membrane components have been addressed with this system. We anticipate more growth and an even more prominent role of this tool in basic and applied membrane research in the decades to come. 

Abstract EPR spectroscopy of site-directed spin labeled membrane proteins is at present a common and valuable biophysical tool to study structural details and conformational transitions under conditions relevant to function. EPR is considered a complementary approach to X-ray crystallography and NMR because it provides detailed information on (1) side chain dynamics with an exquisite sensitivity for flexible regions, (2) polarity and water accessibility profiles across the membrane bilayer, and (3) distances between two spin labeled side chains during protein functioning. Despite the drawback of requiring site-directed mutagenesis for each new piece of information to be collected, EPR can be applied to any complex membrane protein system, independently of its size. This chapter describes the state of the art in the application of site-directed spin labeling (SDSL) EPR to membrane proteins, with specific focus on the different types of information which can be obtained with continuous wave and pulsed techniques. 

Water soluble proteins can be triggered by a variety of factors to insert spontaneously in the membrane bilayer (e.g., toxins, apoptotic proteins, etc.). With SDSL EPR one can observe in real time the membrane insertion via mobility and accessibility changes. Moreover, interspin distances can be measured to model the conformation of the active membrane-bound state of the protein. The case presented here is the application of SDSL EPR to the proapoptotic Bcl-2 protein Bax, triggered to insert into the membrane bilayer by the proapoptotic protein Bid... 

Many complex systems have been spread on liquid interfaces for a variety of reasons. We begin this chapter with a discussion of the behavior of synthetic polymers at the liquid-air interface. Most of these systems are linear macromolecules however, rigid-rod polymers and more complex structures are of interest for potential optoelectronic applications. Biological macromolecules are spread at the liquid-vapor interface to fabricate sensors and other biomedical devices. In addition, the study of proteins at the air-water interface yields important information on enzymatic recognition, and membrane protein behavior. We touch on other biological systems, namely, phospholipids and cholesterol monolayers. These systems are so widely and routinely studied these days that they were also mentioned in some detail in Chapter IV. The closely related matter of bilayers and vesicles is also briefly addressed. 

The other class of phenomenological approaches subsumes the random surface theories (Sec. B). These reduce the system to a set of internal surfaces, supposedly filled with amphiphiles, which can be described by an effective interface Hamiltonian. The internal surfaces represent either bilayers or monolayers—bilayers in binary amphiphile—water mixtures, and monolayers in ternary mixtures, where the monolayers are assumed to separate oil domains from water domains. Random surface theories have been formulated on lattices and in the continuum. In the latter case, they are an interesting application of the membrane theories which are studied in many areas of physics, from general statistical field theory to elementary particle physics [26]. Random surface theories for amphiphilic systems have been used to calculate shapes and distributions of vesicles, and phase transitions [27-31]. 

The use of Upid bilayers as a relevant model of biological membranes has provided important information on the structure and function of cell membranes. To utilize the function of cell membrane components for practical applications, a stabilization of Upid bilayers is imperative, because free-standing bilayer lipid membranes (BLMs) typically survive for minutes to hours and are very sensitive to vibration and mechanical shocks [156,157]. The following concept introduces S-layer proteins as supporting structures for BLMs (Fig. 15c) with largely retained physical features (e.g., thickness of the bilayer, fluidity). Electrophysical and spectroscopical studies have been performed to assess the appUcation potential of S-layer-supported lipid membranes. The S-layer protein used in aU studies on planar BLMs was isolated fromB. coagulans E38/vl. 

The popular applications of the adsorption potential measurements are those dealing with the surface potential changes at the water/air and water/hydrocarbon interface when a monolayer film is formed by an adsorbed substance. " " " Phospholipid monolayers, for instance, formed at such interfaces have been extensively used to study the surface properties of the monolayers. These are expected to represent, to some extent, the surface properties of bilayers and biological as well as various artificial membranes. An interest in a number of applications of ordered thin organic films (e.g., Langmuir and Blodgett layers) dominated research on the insoluble monolayer during the past decade. 

The lipid molecule is the main constituent of biological cell membranes. In aqueous solutions amphiphilic lipid molecules form self-assembled structures such as bilayer vesicles, inverse hexagonal and multi-lamellar patterns, and so on. Among these lipid assemblies, construction of the lipid bilayer on a solid substrate has long attracted much attention due to the many possibilities it presents for scientific and practical applications [4]. Use of an artificial lipid bilayer often gives insight into important aspects ofbiological cell membranes [5-7]. The wealth of functionality of this artificial structure is the result of its own chemical and physical properties, for example, two-dimensional fluidity, bio-compatibility, elasticity, and rich chemical composition. 

Kunitake, T. (1992) Synthetic bilayer-membranes - molecular design, self-organization, and application. Angew. Chem. Int. Ed., 31, 709-726. 

The most successful continuum description of membrane elasticity, dynamics, and thermodynamics is based on the smectic bilayer model (for examples of different versions and applications of this approach see Ref. 76-82 and references therein). We introduce this model in conjunction with the question of membrane undulations. 

Other possible direct probes are optical experiments similar to studies [113] of vesicles but expanded towards shorter A (20-30 A). Alternatively neutron spin-echo studies of stacked bilayer arrays, which can probe the 10-30 A range [114], might possibly be applicable here. Finally, the x-ray grazing-incidence technique has been shown to be a powerful tool for studying short wavelength fluctuations at fluid interfaces [100]. The application of this technique to the investigation of membrane surface fluctuations can reasonably be expected in the near future [115,116]. 

The voltammetric method and concept are expected to be applicable also to the analysis of the oscillation at a very thin membrane such as a bilayer lipid membrane, since even the ion transfer through a bilayer lipid membrane can be observed as a vol-tammogram and interpreted by the way similar to that for a liquid membrane [20,21]. 

Here the basic concepts and the method for strictly determining DD sites in lipid bilayer membranes are viewed, together with some examples of their applications. 

The method utilizing ID NMR is simple and eonvenient. Henee the NMR method diseussed here ean be applied to the systematie investigation of the membrane irug inter-aetions, elosely related to the vital function in biomembranes. It is expected that the application can be extended to the lipid-peptide interaction and protein uptake. We are now applying the method to apolipoprotein binding with lipid bilayers and emulsions. Preferential protein binding sites in membranes can be specified by NMR on the molecular level. 

Amphipathic peptides contain amino acid sequences that allow them to adopt membrane active conformations [219]. Usually amphipathic peptides contain a sequence with both hydrophobic amino acids (e.g., isoleucine, valine) and hydrophilic amino acids (e.g., glutamic acid, aspartic acid). These sequences allow the peptide to interact with lipid bilayer. Depending on the peptide sequence these peptides may form a-helix or j6-sheet conformation [219]. They may also interact with different parts of the bilayer. Importantly, these interactions result in a leaky lipid bilayer and, therefore, these features are quite interesting for drug delivery application. Obviously, many of these peptides are toxic due to their strong membrane interactions. 

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