Dynamics of protein hydration water

We present the frequency- and temperature-dependent dielectric properties of lysozyme solutions in a broad concentration regime, measured at subzero temperatures, and compare the results with measurements above the freezing point of water and on hydrated lysozyme powder. Our experiments allow examining the dynamics of unfreezable hydration water in a broad temperature range. The obtained results prove the bimodality of the hydration shell dynamics. In addition, we find indications of a fragile-to-strong transition of hydration water.

Figure 1

Schematic plot of the dielectric loss of protein solutions above (a) and below (b) the freezing point of water (the labeling of the different processes follows the biophysical nomenclature). The circles show the loss after subtraction of the dc conductivity and EP contributions (grey regions). Frame (a) comprises the major processes hitherto found in protein solutions [15]: The commonly observed $\beta$ and $\gamma$ relaxations are due to the reorientational motions of the dipolar protein and water molecules, respectively. The $\delta$ relaxations are ascribed to hydration water. Additional contributions may arise from conformational sampling (sub-$\beta$). (b) Frozen lysozyme solutions show three intrinsic relaxations: The “ice” process results from proton hopping in the ice matrix. In the frequency range of the $\delta$ dispersion, two relaxations are depicted (${\delta }_1$, ${\delta }_2$).

Figure 2

Dielectric spectra of frozen protein solutions. (a) Dielectric constant of a 10 mmol/l lysozyme solution measured at different temperatures below 273 K. The solid lines are fits using the sum of five Cole-Cole functions. (b),(c) Zoomed view of ${\varepsilon }^{\prime }\left(\nu \right)$ (a) and ${\varepsilon }^{″}\left(\nu \right)$ of differently concentrated protein solutions ($c=10$, 20, and 100 mmol/l) measured at 250 K. Lines are fits using five (solid lines) or four (dashed lines) Cole-Cole functions in total. Pluses in (c) represent the dielectric loss of a hydrated lysozyme powder ($h=30$ wt%).

Figure 3

Relaxation times of the ${\delta }_1$ and ${\delta }_2$ relaxations of differently concentrated protein solutions ($c=3–100$ mmol/l) below the freezing point obtained in the present work. For comparison, literature data on ${\tau }_{{\delta }_1}$ [13, 14, 55] and ${\tau }_{{\delta }_2}$ [12, 13] at $T>273$ K are included (Refs. [13, 14, 55]: lysozyme solutions; Ref. [12]: ribonuclease A). The full circles and triangles are the relaxation times of the two intrinsic relaxations found in a hydrated lysozyme powder [15] ($h=30$ wt%). The asterisks and crosses are the relaxation times of water (own data) and supercooled water [56], respectively. The lines are master curves using the VFT formula.

Figure 4

Dielectric loss of a 10 mmol/l lysozyme solution measured at different temperatures below 273 K. Solid lines are fits using the sum of five Cole-Cole functions.

Figure 5

(a) Frequency dependence of the dielectric loss of the 5 mmol/l lysozyme solution for temperatures below the freezing point. (b) First derivative $\partial {log}_{10}\left({\varepsilon }^{″}\right)/\partial {log}_{10}\left(\nu \right)$ of the data shown in (a). To avoid excessive scattering, before calculating the derivatives, the data in (a) were smoothed by Savitzky-Golay filtering using 5th-order polynomials.