Reversibility, of protein adsorption

Early experiments of MacRitchie Alexander (1963) showed that the adsorption behaviour of different proteins can be described by a simple reaction relation given by Eq. (4.80). Important investigation of the reversibility of protein adsorption... 

A number of important questions revolve around the central one of reversibility of protein adsorption. Are proteins, once adsorbed, able to desorb Why are proteins so difficult to remove from a surface What happens in the situation where there is competitive adsorption between different proteins or between proteins and other species Do the different points on an adsorption isotherm correspond to dynamic equilibria If a protein molecule can desorb, does it revert to its original solution configuration and recover its original biological activity Can the Gibbs Adsorption Equation be applied to protein adsorption Some of these questions may be effectively tackled by studies with the film balance. 

MacRitchie, F. 1998. Reversibility of protein adsorption. In Profezns at liquid interfaces, ed. D. Moebius and R. Miller, 149-177. Amsterdam Elsevier Science B.V. 

Norde, W. and C.A. Haynes. 1995. Reversibility and the mechanism of protein adsorption. In Proteins at Interfaces II Fundamentals and Applications. T.A. Hor-bett and J.L. Brash, editors. American Chemical Society, Washington, D.C., 26 40. 

Fig. 12. Hypothetical 3-D plot of protein adsorption isotherm (D plotted against [P]B) as a function of surface ligand concentration, [A]s. Note that the system is reversible only up to a critical [A]s and then behaves irreversibly for higher ligand surface concentrations. The right arrows (— ) denote adsorption the left-facing ones ( -), desorption...

Fig. 20. Schematic adsorption isotherms with a constant surface site concentration ([A]s in Fig. 12 is here constant), but with adsorption time as a variable. At very short times, adsorption is diffusion controlled. At short times, the protein has insufficient time to conformationally adjust to the interface, thus adsorption can be reversible and of the Langmuir type. At longer times, conformational adjustments begin leading to the commonly observed semi- orir-reversible behavior of protein adsorption. Other nomenclature same as Fig. 12...

Rejection of protein adsorption to the outermost grafted surface is attributed to a steric hinderance effect due to the tethered chains. A grafted surface in contact with an aqueous medium, a good solvent of the chains, has been identified to have a diffuse structure [57,151,152]. Reversible deformation of the tethered... 

Furthermore, the attachment of the hydrophobic tails will affect the surface activity more than by simply increasing hydrophobicity. As presented in Fig. 2, the attachment of hydrophobic tails will cause several domains of the protein to behave as a classical surfactant, and the overall effect will be the formation of a polymer-like surfactant. Such a polymer, which will be adsorbed at various hydrophobic interfaces, should have an increased surface activity due to the existence of many adsorption sites per molecule. This adsorption mechanism may affect the reversibility of the adsorption process and might even prevent surface denaturation. This latter effect is important in appli-... 

It is evident that elucidation of the interfacial behavior of proteins is not a simple matter and requires contributions from several disciplines. In recent years considerable progress has been made in applying spectroscopic techniques to proteins in the adsorbed state (e.g., 7,8,9). In such studies a (small) part of the molecule is analyzed in detail. In our laboratory we study protein adsorption from a more classical, colloid-chemical point of view. Arguments are derived from experimental data referring to whole protein molecules or to layers of them. Information is obtained from adsorption isotherms, proton titrations and both electrokinetic and thermochemical measurements. Recently, topical questions such as reversibility of the adsorption process and changes in the protein structure have been considered. This more holistic approach has produced some insights that could not easily be obtained otherwise. 

The in situ adsorption/desorption technique can be used to distinguish between irreversibly and reversibly adsorbed protein. The desorption/adsorption ratio depends on the nature of protein and surface. Further development of this technique would involve the design of a new type of measuring cell for adsorption/desorption measurements in flow conditions and extension of protein adsorption experiments to competitive protein adsorption measurement. Studies of albumin and fibrinogen competitive adsorption versus collagen 12) have recently been initiated. 

We begin with some comments on the various experimental methods used in our studies. Investigations of single proteins in buffer are then discussed, including kinetics and isotherms, reversibility, denaturation, and the structural status of adsorbed proteins. Results of competitive adsorption studies using mixtures of proteins in buffer are then described. We next discuss our more recent studies of protein adsorption from blood plasma using both radiolabeled proteins and elution techniques. Finally, data on the effect of red blood cells on protein adsorption are summarized. 

The electronically conducting polymers are particularly interesting because one can control the oxidation state of these polymers in a reversible and predictable manner by the application of a small potential or current. The implication is that one may be able to control the charge density on the surface of the polymer. The nature and density of surface charges play a very important role in defining the extent and nature of protein adsorption on a biomaterial surface, which in turn is key in determining cellular-biomaterial interactions. 

Abstract A dynamic model of protein adsorption on solid surfaces is presented. It is based on the assumption that a protein molecule may change conformation after adsorption. The computed adsorption isotherms are similar in form to experimentally observed isotherms. A model for the experimentally observed reversible potential changes at a metal electrode upon protein adsorption/de-sorption is also discussed. It is thus suggested that a second loosely bound protein layer may be present on several metal surfaces. Some notes are also made about the significance of the time scale and detailed method of performing an adsorption experiment. 

Brusatori and Van Tassel [20] presented a kinetic model of protein adsorption/surface-induced transition kinetics evaluated by the scale particle theory (SPT). Assuming that proteins (or, more generally, particles ) on the surface are at all times in an equilibrium distribution, they could express the probability functions that an incoming protein finds a space available for adsorption to the surface and an adsorbed protein has sufficient space to spread in terms of the reversible work required to create cavities in a binary system of reversibly and irreversibly adsorbed states. They foimd that the scale particle theory compared well with the computer simulation in the limit of a lower spreading rate (i.e., smaller surface-induced unfolding rate constant) and a relatively faster rate of surface filling. 

Figure 7.8. A representation of the collapse and charge reversal of proteins with change of pH and adsorption of surfactant of opposite charge.

Owing to the weak hydrophobicity of the PEO stationary phases and reversibility of the protein adsorption, some advantages of these columns could be expected for the isolation of labile and high-molecular weight biopolymers. Miller et al. [61] found that labile mitochondrial matrix enzymes — ornitine trans-carbomoylase and carbomoyl phosphate synthetase (M = 165 kDa) could be efficiently isolated by means of hydrophobic interaction chromatography from the crude extract. 

This precipitation process can be carried out rather cleverly on the surface of a reverse phase. If the protein solution is brought into contact with a reversed phase, and the protein has dispersive groups that allow dispersive interactions with the bonded phase, a layer of protein will be adsorbed onto the surface. This is similar to the adsorption of a long chain alcohol on the surface of a reverse phase according to the Langmuir Adsorption Isotherm which has been discussed in an earlier chapter. Now the surface will be covered by a relatively small amount of protein. If, however, the salt concentration is now increased, then the protein already on the surface acts as deposition or seeding sites for the rest of the protein. Removal of the reverse phase will separate the protein from the bulk matrix and the original protein can be recovered from the reverse phase by a separate procedure. 

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