Interfacial thermodynamics and electrochemistry of protein partitioning in two-phase systems
The subject of this thesis is protein partition between an aqueous salt solution and a surface or an apolair liquid and the concomitant co-partition of small ions. The extent of co-partitioning determines the charge regulation in the protein partitioning process.Chapters 2 and 3 deal with phenomenological relations between the partition coefficient of the protein and the extent of the co- partition. The method of analysis is illustrated by some worked-out examples, using data taken either from literature or from chapter 5. The examples include proton titration curves, ion exchange chromatography, adsorption on colloidal particles and solubilization in reverse micelles. An important conclusion is that the partition process is subject to a rule, similar to the principle of Le Chatelier for chemical equilibria: if upon protein partitioning ions are expelled into the water phase, an increase of the ionic concentrations results in a decrease of the protein partition coefficient and conversely.A theory which allows for the prediction and molecular interpretation of the charge regulations is presented in chapter 4. The model describes the electrochemistry of a protein molecule through site binding of ions on a rigid surface. Although this is a considerably simplified picture of a real protein molecule, some aspects of the theory may be of general validity. One of them is the notion of the electrochemical adaptability of a charged colloidal particle, as measured by its intrinsic capacitance. In the case of a high intrinsic capacitance, a change in electrostatic interactions results in a large charge regulation whilst the surface potential remains almost constant. On the other hand, if the intrinsic capacitance is low, the particle resists externally imposed shifts in charge but does adapt its surface potential.Chapter 5 contains an experimental study towards understanding the mechanism of charge-regulation in protein adsorption. The system consists of crystals of the insoluble salt silver iodide as the adsorbent and the protein Bovine Serum Albumin as the adsorbate. By using a combined iodide and proton titration technique, the charges of the surface and the protein can be measured independently. We find that a negative surface induces a positive shift in the charge of the adsorbed protein. Opposed to intuitive expectation, the reverse is not always true: when the charge of the protein charge is maximallypositive, adsorption renders the silver iodide surface less negative.The anomalous charge regulation is explained in terms of the intrinsic capacitance of the adsorbed protein. The maximally positive protein cannot adapt its charge, and so the silver iodide surface is forced to adjust its charge completely to that of the protein. As the contact layer between adsorbed protein and the silver iodide crystal is electroneutral under almost all circumstances, the silver iodide surface must be as negative as the protein is positive. Hence, if the charge of the surface before adsorption is more negative than this value, adsorption of the protein is accompanied by a desorption of negative charge.The experimental results are well understood in view of the developed phenomenological theory and model analysis. Two thermodynamic relations are succesfully verified, indicating the internal consistency of the various experiments. Application of the model gives two independent estimates of the size of the adsorbed protein. It is concluded that the protein does not substantially modify its native structure upon adsorption.
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| Format: | Doctoral thesis biblioteca |
| Language: | English |
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Fraaije
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| Subjects: | boundary layer, capillaries, electrochemistry, fluid mechanics, proteins, surface phenomena, surface tension, thermal energy, thermodynamics, two-phase systems, capillairen, eiwitten, elektrochemie, grenslaag, oppervlaktespanning, oppervlakteverschijnselen, thermische energie, thermodynamica, tweefasesystemen, vloeistofmechanica, |
| Online Access: | https://research.wur.nl/en/publications/interfacial-thermodynamics-and-electrochemistry-of-protein-partit |
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| Summary: | The subject of this thesis is protein partition between an aqueous salt solution and a surface or an apolair liquid and the concomitant co-partition of small ions. The extent of co-partitioning determines the charge regulation in the protein partitioning process.Chapters 2 and 3 deal with phenomenological relations between the partition coefficient of the protein and the extent of the co- partition. The method of analysis is illustrated by some worked-out examples, using data taken either from literature or from chapter 5. The examples include proton titration curves, ion exchange chromatography, adsorption on colloidal particles and solubilization in reverse micelles. An important conclusion is that the partition process is subject to a rule, similar to the principle of Le Chatelier for chemical equilibria: if upon protein partitioning ions are expelled into the water phase, an increase of the ionic concentrations results in a decrease of the protein partition coefficient and conversely.A theory which allows for the prediction and molecular interpretation of the charge regulations is presented in chapter 4. The model describes the electrochemistry of a protein molecule through site binding of ions on a rigid surface. Although this is a considerably simplified picture of a real protein molecule, some aspects of the theory may be of general validity. One of them is the notion of the electrochemical adaptability of a charged colloidal particle, as measured by its intrinsic capacitance. In the case of a high intrinsic capacitance, a change in electrostatic interactions results in a large charge regulation whilst the surface potential remains almost constant. On the other hand, if the intrinsic capacitance is low, the particle resists externally imposed shifts in charge but does adapt its surface potential.Chapter 5 contains an experimental study towards understanding the mechanism of charge-regulation in protein adsorption. The system consists of crystals of the insoluble salt silver iodide as the adsorbent and the protein Bovine Serum Albumin as the adsorbate. By using a combined iodide and proton titration technique, the charges of the surface and the protein can be measured independently. We find that a negative surface induces a positive shift in the charge of the adsorbed protein. Opposed to intuitive expectation, the reverse is not always true: when the charge of the protein charge is maximallypositive, adsorption renders the silver iodide surface less negative.The anomalous charge regulation is explained in terms of the intrinsic capacitance of the adsorbed protein. The maximally positive protein cannot adapt its charge, and so the silver iodide surface is forced to adjust its charge completely to that of the protein. As the contact layer between adsorbed protein and the silver iodide crystal is electroneutral under almost all circumstances, the silver iodide surface must be as negative as the protein is positive. Hence, if the charge of the surface before adsorption is more negative than this value, adsorption of the protein is accompanied by a desorption of negative charge.The experimental results are well understood in view of the developed phenomenological theory and model analysis. Two thermodynamic relations are succesfully verified, indicating the internal consistency of the various experiments. Application of the model gives two independent estimates of the size of the adsorbed protein. It is concluded that the protein does not substantially modify its native structure upon adsorption. |
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