The study of hydration, folding, and interaction of proteins by volumetric measurements has been promoted by recent advances in the development of highly sensitive instrumentations. However, the separation of the measured apparent volumes into contributions from the protein and the hydration water, V(app) = V(int) + ΔV, is still challenging, even with the detailed microscopic structural information from molecular simulations. By the examples of the amyloidogenic polypeptides hIAPP and Aβ42 in aqueous solution, we analyze molecular dynamics simulation runs for different temperatures, using the Voronoi-Delaunay tessellation method. This method allows a parameter free determination of the intrinsic volume V(int) of complex solute molecules without any additional assumptions. For comparison, we also use fused sphere calculations, which deliver van der Waals and solute accessible surface volumes as special cases. The apparent volume V(app) of the solute molecules is calculated by different approaches, using either a traditional distance based selection of hydration water or the construction of sequential Voronoi shells. We find an astonishing coincidence with the predictions of a simple empirical approach, which is based on experimentally determined amino acid side chain contributions (Biophys. Chem.1999, 82, 35). The intrinsic volumes of the polypeptides are larger than their apparent volumes and also increase with temperature. This is due to a negative contribution of the hydration water ΔV to the apparent volume. The absolute value of this contribution is less than 10% of the intrinsic volume for both molecules and decreases with temperature. Essential volumetric differences between hydration water and bulk water are observed in the nearest neighborhood of the solute only, practically in the first two Delaunay sublayers of the first Voronoi shell. This also helps to understand the pressure dependence of the partial molar volumes of proteins.

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The volumetric properties of human islet amyloid polypeptide (hIAPP) in water were studied in a wide temperature range by computer simulations. The intrinsic density rho(p) and the intrinsic thermal expansion coefficient alpha(p) of hIAPP were evaluated by taking into account the difference between the volumetric properties of hydration and bulk water. The density of hydration water rho(h) was found to decrease almost linearly with temperature upon heating and its thermal expansion coefficient was found to be notably higher than that of bulk water. The peptide surface exposed to water is more hydrophobic and its rho(h) is smaller in conformation with a larger number of intrapeptide hydrogen bonds. The two hIAPP peptides studied (with and without disulfide bridge) show negative alpha(p), which is close to zero at 250 K and decreases to approximately -1.5 x 10(-3) K(-1) upon heating to 450 K. The analysis of various structural properties of peptides shows a correlation between the intrinsic peptide volumes and the number of intrapeptide hydrogen bonds. The obtained negative values of alpha(p) can be attributed to the shrinkage of the inner voids of the peptides upon heating.

The effect of temperature on the connectivity of hydrogen bonds in the hydration shells of the islet amyloid polypeptides (IAPPs) is studied by means of computer simulations. The hydrogen-bonded network of hydration water homogeneously envelopes a peptide at low temperature and breaks into an ensemble of small clusters upon heating. This thermal break occurs via a percolation transition, which is not found to be sensitive to the chemical modifications of IAPP (IAPP with and without a disulfide bridge, human and rat IAPP). The radius of gyration of IAPP starts to increase when the hydration water network breaks upon heating. The fluctuations of the number of intra-peptide hydrogen bonds show negative correlation with the fraction of molecules in the largest cluster of hydration water. The thermal stability of the network of hydration water is enhanced upon increasing number of intra-peptide hydrogen bonds, which makes the peptide surface more hydrophobic. The thermal stabilities of the hydrogen-bonded water networks in the hydration shells of IAPPs and of several other biomolecules are found to be rather similar: the network breaks between 300 and 330 K, i.e., in the temperature interval where the biological activity of living organisms is maximal.

u sol describes the energy of H-bonds to solvent. These three model parameters are obtained by fitting to experimental C p curves. Best fits on 19 different lysozyme mutants under the same thermodynamic conditions (pH, ionic strength, etc.) reveals that u sol can be treated as constant. Moreover, u sol can be robustly parameterized with as few as five experimental mutants (the standard error of 100 random quintets is <12%). It was observed that a second degree of freedom could be removed due to a linear relationship (R¼0.86) between the remaining two parameters (d nat and v dha ) indicating that a global balance in enthalpy-entropy compensation must be maintained. Consequently, over a fairly wide range of d nat values {0.4, 1.6}, the correlation between the experimental and theoretical T m 's is nearly constant (ranging from 0.68 to 0.72). Using the best parameter set, T m can be predicted for new lysozyme mutants. Results on a validation set of an additional 81 lysozyme point mutations will be presented. This work is supported by NIH R01 GM073082.