Proteins spread monolayers

It has been proposed that native protdn molecules are made up of parallel sheets or layers of peptide chains (72, 73, 74, 75). It is reasonable to believe that these layers have the same hydrophilic-hydrophobic characteristics as do spread films. Thus in a native molecule containing two such layers of peptide chains, the outer faces of the molecule would be predominantly hydrophilic with a hydrophoWc sandwich in between. Dervichian, for example, has drawn an anal< between a soap micelle and a protein molecule as far as the arrangement of the hydrophilic and hydrophobic groups are concerned. Certainly, there is much evidence to substantiate such a picture. While most of this evidence is of an indirect nature, it is no less compelling. Without the layer structure of the native molecule as the guiding idea, the formation of protein spread monolayers is difficult to understand. 

Fig. 3. A spread monolayer of protein decreases fe for the absorption of CO (from the gas phase at 1 atm. partial pressure) into water (28). The surface is cleaned thoroughly before the experiment, and the contra-rotating stirrers in the liquid are running at 437 r.p.m. The surface is nonrotating in aU the experiments described here, and when fct is reduced to 1.1 X lO"" cm. sec.", all random surface movements are also eliminated.

Fig. 9. Reduction in kt by spread monolayer of protein, due to prevention of transfer of momentum from ethylacetate, and to reduction in turbulence in the aqueous phase near the interface. Experimental data used here are taken from reference (60).

If a small amount of protein solution is suitably spread at the surface of an aqueous substrate, most of the protein will be surface-denatured, giving an insoluble monomolecular film before it has a chance to dissolve. The techniques already described for studying spread monolayers of insoluble material can, therefore, be used in... 

Tnterfacial phenomena play a fundamental role in biological systems. It A is important to know if surface energy and anisotropy affect the conformation of biological macromolecules. Well defined physicochemical models might simplify this problem (1- 8) spread monolayers at the air-water interface exemplify this kind of model. For polypeptides which are introduced as simple models of proteins, no surface denatura-tion of the spread macromolecules occurred (9, 10, 11). Protein structures are too complex to yield direct information about eventual changes of conformation, but one can detect the presence or the disappearance of biological activity—e.g., enzymic activity. The enzyme would be denatured if the conformation were modified by the anisotropy of the interface. 

A monolayer of protein spreads at the air-water interface. The amount of protein spread is equivalent to 0.80 mg m" and gives a surface tension lowering of 0.035 mN mWhat is the molecular weight of the protein ... 

Thirty milligrams of a protein spread at an air/water interface forms an insoluble monolayer. The interfacial pressure n is measured at different areas A available for the protein. The measurements are performed at 25°C. 

The above experiments on monolayers illustrate the strong dependence of desorption rates on n. In real systems stabilised by proteins, n for the film on average does not exceed a particular maximum value at which the rate of adsorption from solution is balanced by the rate of desorption. On perturbation from the equilibrium state of the film, such as a transient (local) expansion or compression a knowledge of both rates is important. Unfortunately, measurements of adsorption rates are not so straightforward since the surface concentration of protein, r, must be monitored with time and is not predetermined as in the spread monolayers. There is often disagreement between adsorption kinetic results obtained via different techniques - see below, for example. Relatively few measurements have been made of the adsorption kinetics of S-lactoglobulin at the A-W interface and for all proteins, because of experimental difficulties, there seem to be almost no direct measurements of r t) at O-W interfaces. 

Spread monolayers of proteins have been previously reviewed by Gorter (2) and by Neurath and Bull (3) and by Bateman (4). 

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. 

Proteins, like other macromolecules, can be made into monolayers at the air-water interface either by spreading, adsorption, or specific binding. Proteins, while complex polymers, are interesting because of their inherent surface activity and amphiphilicity. There is an increasing body of literature on proteins at liquid interfaces, and here we only briefly discuss a few highlights. 

FIG. 16 Fomation of a Langmuir lipid monolayer at the air/subphase interface and the subsequent crystallization of S-layer protein, (a) Amphiphilic lipid molecules are placed on the air/subphase interface between two barriers. Upon compression between the barriers, increase in surface pressure can be determined by a Wilhelmy plate system, (b) Depending on the final area, a liquid-expanded or liquid-condensed lipid monolayer is formed, (c) S-layer subunits injected in the subphase crystallized into a coherent S-layer lattice beneath the spread lipid monolayer and the adjacent air/subphase interface. 

Protein A is a cell-wall protein of Staphylococcus aureus with a molecular weight of 42,000. Since protein A binds specifically to the Fc part of IgG from various animals, it has been widely used in immunoassay and affinity chromatography. We found that protein A could be spread over the water surface to form a monolayer membrane by the LB method [21]. On the basis of this finding, an antibody array on the solid surface can be obtained by the following two steps. The first step is fabrication of an ordered protein A array on the solid surface by the LB method. The second step is self assembly of antibody molecules on the protein A array by biospecific affinity between protein A and the Fc of IgG as shown in Fig.34. 

As described here, the monolayer of a lipid can be formed by different spreading methods. The thermodynamics of the Ilcol analysis is given in the literature (Birdi, 1989). The monolayer collapse has been shown to provide much information also in the case of protein monolayers. 

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