Unfolding, hydration contribution

The partial molar heat capacity can be considered to be composed of intrinsic and hydration contributions. The intrinsic component contains contributions from covalent and non-covalent interactions. It has been shown that about 85% of the total heat capacity of the native state of a protein in solution is due to the covalent structure [72]. Changes in the heat capacity upon unfolding are therefore primarily interpreted as due to changes in the hydration. A physical picture of energy fluctuations means changing the conformation between ordered and less ordered structures. This can be achieved by hindered internal rotations, low frequency... 

We can reasonably assume two major contributions to the difference in specific volume between the unfolded and folded states of a protein. The first contribution is that arising from the decrease in solvent-excluded volume when the tightly, but of course not perfectly, packed protein folded structure is disrupted. Water molecules enter this volume, thereby decreasing the overall volume of the protein solvent system. The magnitude of this contribution is a specific property of the protein, both in its folded and unfolded state. The second contribution arises from the change in the volume of the water molecules that hydrate the newly exposed protein surface area, relative to their volume in the bulk. Much of our present understanding of the contribution of differential hydration volume has come from recent studies of model compounds and proteins based on PPC. This technique, developed by Brandts and coworkers [17] and recently reviewed by us [16,18], is based on the measurement of the heat released or absorbed upon small (e.g., 0.5 MPa) pressure... 

It can be seen in Fig. 9.8 that the transition shifts, as expected, to higher temperature as a function of increasing osmolyte concentration. The AV decreases in absolute value from —19 to —5 ml mol-1. This is in part due to the increase in the transition temperature, and because of a positive Aa (see Fig. 9.5) the volume between the unfolded and folded state decreases in absolute value. There may be a contribution of the effect of the osmolyte to the structure of the unfolded state as well. The value of a at low temperature increases with increasing osmolyte as a result of the preferential hydration effect. At high temperature, the differences in the expansivity of the bulk water... 

Recent progress in X-ray diffraction of protein crystals in the diamond anvil cell will also make it possible to obtain quantitative information on the cavities [42, 43]. Optical spectroscopy [44] and neutron scattering [45] should also be valuable tools to probe the role of cavities. High-pressure molecular dynamics simulations should also allow estimating the contributions of the hydration and the cavities. High-pressure simulations on the small protein, bovine pancreatic trypsin inhibitor, indicate an increased insertion of water into the protein interior before unfolding starts to occur [46,47]. 

An adhesive-cohesive model for protein compressibility has been proposed by Dadarlat and Post [57]. This model assumes that the compressibility is a competition between adhesive protein-water interactions and cohesive protein-protein interactions. Computer simulations suggest that the intrinsic compressibility largely accounts for the experimental compressibilities indicating that the contribution of hydration water is small. The model also accounts for the correlation between the compressibility of the native state and the change in heat capacity upon unfolding for nine single chain proteins. 

Well different is, instead, the situation observed when the exploration was extended well inside the protein irreversible denaturation region. Two Lorentzians, appear just after the crossing of the border of the ID —> D phases, that is, where both the external protein hydration water and the internal one are detectable. When proteins unfold in an open polymeric structure, the internal water (also considering the effective high T) can easily break the HBs that link it to the protein residuals and can diffuse and interact with the external one. This reason explains the presence of two proton water NMR signals inside the phase D. One contribution for continuity is related with the protein hydration water whereas the second component with the internal water one. Both the components will survive in the measured spectra upto the end of the cooling phase. After the denaturation these two water forms are present in the system and can interact with each other or with the open biopolymer, in a complete different physical scenario if compared with the folded protein native state. 

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