Experimental models of biological membranes

In view of the complexity and diversity of the functions performed by the various proteins embedded in a biomembrane (the integral proteins), it has been found convenient to incorporate single integral proteins or smaller lipophihc biomolecules into experimental models of biological membranes, so as to isolate and investigate their functions. This serves to reduce complex membrane processes to well-defined interactions between selected proteins, lipids, and hgands. There is... 

The vast majority of biological membranes are in the liquid-crystalline phase. There are many experimental studies on model bilayer phase behavior [3]. Briefly, at low temperatures lipid bilayers form a gel phase, characterized by high order and rigidity and slow lateral diffusion. There is a main phase transition, as the temperature is increased, to the liquid-crystalline phase. The liquid-crystalline phase has more fluidity and fast lateral diffusion. 

It is possible that quite different molecular architectures may occur in membranes from different sources. Current research may result in a much more dramatic revision or complete rejection of the bilayer model for some membranes, especially in such systems as mitochondria (30) and chloroplasts (2). However, it is also possible that structural differences are only variations on the basic theme of the bilayer, from myelin at one extreme to mitochondria or chloroplasts on the other. One must not readily reject the fundamentals of the Danielli concept, especially in view of the present inadequate knowledge of the properties of phospholipids in water. Clearly the molecular architecture of membranes is speculative, but most aspects of the problem are amenable to direct experimental test by the new physical techniques. A consistent model for biological membranes will emerge quickly. 

The major discovery in the field of biologic membranes is undoubtedly the finding that the biomembrane is a liquid-crystal-line lipid bilayer with embedded proteins (1-3). This so-called fluid-mosaic model (1) has been the central paradigm in membrane biology for more than three decades and has been very successful in rationalizing a large body of experimental observations. The model includes two references to the lipid... 

Log Kow is the most widely used descriptor for baseline QSARs but has many disadvantages in the context of our model. First, because octanol does not perfectly mimic the physicochemical properties of biological membranes, there are two different QSAR equations for nonpolar and polar compounds. This complication can be overcome by using the liposome-water partition coefficient log Kupw as the descriptor instead [32]. We therefore recalculated the Kow-based QSARs from the EU Technical Guidance Documents [31] using relationships between log Kow and logKupw for nonpolar (Eq. 9) and polar compounds (Eq. 10), which had been experimentally determined by Vaes et al. [33,34]. 

Thus, a comparison between HPLC-derived lipophilicity indices and calculated log P values for a series of 8-substituted xanthines showed a clear influence of conformational effects. 8 In this case, Rekker s method was unable to take 3D effects into account, but the difference between experimental and predicted values was structure dependent rather than constant. Conformational analyses confirmed that a smaller than predicted lipophilicity was associated with folded conformers stabilized by hydrophobic and van der Waals forces and having part of their nonpolar surface masked from the aqueous phase. A 4D theoretical approach (log P calculations by MLP for conformers generated by high temperature molecular dynamics) suggests that these effects should be lower in an w-octanol/water system than in RP-HPLC. Indeed, the n-octanol/water system is not the most suitable model to study intramolecular interactions in nonpolar media because a surprisingly high proportion of water is dissolved in the w-octanol. Recall, however, that w-octanol, despite some limitations, was selected by many workers in the field as a model for biological membranes. 

Biological membranes provide the essential barrier between cells and the organelles of which cells are composed. Cellular membranes are complicated extensive biomolecular sheetlike structures, mostly fonned by lipid molecules held together by cooperative nonco-valent interactions. A membrane is not a static structure, but rather a complex dynamical two-dimensional liquid crystalline fluid mosaic of oriented proteins and lipids. A number of experimental approaches can be used to investigate and characterize biological membranes. However, the complexity of membranes is such that experimental data remain very difficult to interpret at the microscopic level. In recent years, computational studies of membranes based on detailed atomic models, as summarized in Chapter 21, have greatly increased the ability to interpret experimental data, yielding a much-improved picture of the structure and dynamics of lipid bilayers and the relationship of those properties to membrane function [21]. 

A final example of the simulation of a complex system is a series of MD simulations of bilayer membranes. Membranes are crucial constituents of living organisms they are the scene for many important biological processes. Experimental data are known for model systems for example for the system sodium decanoate, decanol and water that forms smectic liquid crystalline structures at room temperature, with the lipids organized in bilayers. 

Although for the moment this model is only partially supported by experimental data it offers the opportunity to design new experiments which will help to understand the mechanisms of pardaxin insertion and pore formation in lipid bilayers and biological membranes which at a molecular level are the events leading to shark repellency and toxicity of this marine toxin. 

Figure 9. A comparison of the order parameter profile as found by MC simulations [72] of model 9/10 cis unsaturated chains in a monolayer (the x-line is to guide the eye) with experimental data obtained from NMR experiments (o) on the same chains incorporated into a biological membrane. Redrawn from [72] by permission of the American Institute of Physics...

Meanwhile, computational methods have reached a level of sophistication that makes them an important complement to experimental work. These methods take into account the inhomogeneities of the bilayer, and present molecular details contrary to the continuum models like the classical solubility-diffusion model. The first solutes for which permeation through (polymeric) membranes was described using MD simulations were small molecules like methane and helium [128]. Soon after this, the passage of biologically more interesting molecules like water and protons [129,130] and sodium and chloride ions [131] over lipid membranes was considered. We will come back to this later in this section. 

As seen above (equation (5)), the basis of the simple bioaccumulation models is that the metal forms a complex with a carrier or channel protein at the surface of the biological membrane prior to internalisation. In the case of trace metals, it is extremely difficult to determine thermodynamic stability or kinetic rate constants for the adsorption, since for living cells it is nearly impossible to experimentally isolate adsorption to the membrane internalisation sites (equation (3)) from the other processes occurring simultaneously (e.g. mass transport complexation adsorption to other nonspecific sites, Seen, (equation (31)) internalisation). 

Until the 1950s, the rare periodic phenomena known in chemistry, such as the reaction of Bray [1], represented laboratory curiosities. Some oscillatory reactions were also known in electrochemistry. The link was made between the cardiac rhythm and electrical oscillators [2]. New examples of oscillatory chemical reactions were later discovered [3, 4]. From a theoretical point of view, the first kinetic model for oscillatory reactions was analyzed by Lotka [5], while similar equations were proposed soon after by Volterra [6] to account for oscillations in predator-prey systems in ecology. The next important advance on biological oscillations came from the experimental and theoretical studies of Hodgkin and Huxley [7], which clarified the physicochemical bases of the action potential in electrically excitable cells. The theory that they developed was later applied [8] to account for sustained oscillations of the membrane potential in these cells. Remarkably, the classic study by Hodgkin and Huxley appeared in the same year as Turing s pioneering analysis of spatial patterns in chemical systems [9]. 

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