The focus in protein folding has been very much on the protein backbone and sidechains. However, hydration waters make comparable contributions to the structure and energy of proteins. The coupling between fast hydration dynamics and protein dynamics is considered to play an important role in protein folding. Fundamental questions of protein hydration include, how far out into the solvent does the influence of the biomolecule reach, how is the water affected, and how are the properties of the hydration water influenced by the separation between protein molecules in solution? We show here that Terahertz spectroscopy directly probes such solvation dynamics around proteins, and determines the width of the dynamical hydration layer. We also investigate the dependence of solvation dynamics on protein concentration. We observe an unexpected nonmonotonic trend in the measured terahertz absorbance of the five helix bundle protein * 6 -85 as a function of the protein: water molar ratio. The trend can be explained by overlapping solvation layers around the proteins. Molecular dynamics simulations indicate water dynamics in the solvation layer around one protein to be distinct from bulk water out to Ϸ10 Å. At higher protein concentrations such that solvation layers overlap, the calculated absorption spectrum varies nonmonotonically, qualitatively consistent with the experimental observations. The experimental data suggest an influence on the correlated water network motion beyond 20 Å, greater than the pure structural correlation length usually observed.solvation dynamics ͉ THz spectroscopy ͉ lambda repressor ͉ molecular modeling W ater molecules interact with proteins on many length and time scales. Although the dynamics of the hydration water occurs on the picosecond time scale, ''slaving'' to fast solvent modes profoundly affects the slower but larger-scale protein motions (1). In return the protein influences the structure and dynamics of surrounding water molecules (2). X-ray crystallography has revealed ordered water structure around polar and charged sidechains (3), as well as cooperative insertion of water into hydrophobic cavities (4). Dielectric spectroscopy extends the time scale from microseconds down to 0.1 ns (5). Experiments have been extended to the THz range in films and crystals, probing motions on the picosecond time scale (6, 7). Hydrated protein powders probed by inelastic neutron scattering (0.1-100 ps) or solid-state NMR (nanoseconds) reveal that slower protein time scales and faster solvent time scales indeed show correlated dynamics (8). On the fastest time scales, 2D infrared spectroscopy and fluorescence of surface residues provide local probes of the dynamics in the femtosecond to picosecond range (9, 10). Coupling of modeling with experiments has revealed complex solvation structure around small biomolecules (11, 12), bridging our microscopic structural and thermodynamic understanding of biosolvation.Terahertz absorption spectroscopy of biomolecules fully solvated in water yields direct informa...

Hydrogen bond rearrangement in water occurs on the picosecond time scale, so relevant experiments must access these times. Here, we show that terahertz spectroscopy can directly investigate hydration layers. By a precise measurement of absorption coefficients between 2.3 THz and 2.9 THz we could determine the size and the characteristics of the hydration shell. The hydration layer around a carbohydrate (lactose) is determined to extend to 5.13 ؎ 0.24 Å from the surface corresponding to Ϸ123 water molecules beyond the first solvation shell. Accompanying molecular modeling calculations support this result and provide a microscopic visualization. Terahertz spectroscopy is shown to probe the collective modes in the water network. The observed increase of the terahertz absorption of the water in the hydration layer is explained in terms of coherent oscillations of the hydration water and solute. Simulations also reveal a slowing down of the hydrogen bond rearrangement dynamics for water molecules near lactose, which occur on the picosecond time scale. The present study demonstrates that terahertz spectroscopy is a sensitive tool to detect solute-induced changes in the water network.hydration water dynamics ͉ molecular dynamics simulations of biomolecules ͉ solvated lactose T he many unusual properties of water combined with its importance as the solvent of life account for its continued study by numerous researchers. Indeed, the properties of water are essential in the behavior of all biological systems. For example, water plays a central role in the folding and function of proteins, and the function of carbohydrates. The dynamics of water surrounding a solute is of fundamental importance in a wide range of processes in solution. In particular, the properties of water molecules near the surface of a biomolecule have been the subject of numerous and sometimes controversial experimental and theoretical studies (1-5). The characteristics and role of this ''biological'' water, with properties that differ considerably from those of bulk water, are still not fully understood (6).The importance of the hydration layer or the biological water is obvious in specific protein processes, such as opening and closing of channels, the kinetics of which has been found to be coupled with the solvent fluctuations (1). Important questions that remain so far unanswered include the following: How does solvation water differ from bulk water, and how large is the solvation layer that can be attributed to solvent water? Simulations suggest the existence of rather rigid water structures around proteins (5) and carbohydrates (7), but experimental studies that characterize the hydration layer are limited. Hydrogen bond rearrangements in water occur on the picosecond time scale (8), so that a detailed understanding of the relevant processes at a molecular level requires experimental techniques that are able to probe the hydration layers on this time scale. However, experimental investigations of fast dynamics of hydration water still remain a ch...

Energy flows anisotropically through the residues and vibrational states of globular proteins. A variety of experimental and computational studies have identified energy transport channels traversing many residues, in some cases connecting functional regions, potentially important in allostery, and in other cases having no apparent function. This property and the diffusion of energy in proteins are mimicked by transport on a percolation cluster. I review work that addresses connections between globular proteins, percolation clusters, and the similarity of energy flow and thermal transport in these systems. I also review experimental and theoretical studies of the anisotropic flow of energy through the vibrational states of a protein, a property that also can be understood by comparison with simple model disordered systems.

We present new terahertz (THz) spectroscopic measurements of solvated sugars and compare the effect of two disaccharides (trehalose and lactose) and one monosaccharide (glucose) with respect to the solute-induced changes in the sub-picosecond network dynamics of the hydration water. We found that the solute affects the fast collective network motions of the solvent, even beyond the first solvation layer. For all three carbohydrates, we find an increase of 2-4% in the THz absorption coefficient of the hydration water in comparison to bulk water. Concentration-dependent changes in the THz absorption between 2.1 and 2.8 THz of the solute-water mixture were measured with a precision better than 1% and were used to deduce a dynamical hydration shell, which extends from the surface up to 5.7 +/- 0.4 and 6.5 +/- 0.9 A for the disaccharides lactose and trehalose, respectively, and 3.7 +/- 0.9 A for the glucose. This exceeds the values for the static hydration shell as determined, for example, by scattering, where the long-range structure was found to be not significantly affected by the solute beyond the first hydration shell. When comparing all three carbohydrates, we found that the solute-induced change in the THz absorption depends on the product of molar concentration of the solute and the number of hydrogen bonds between the carbohydrate and water molecules. We can conclude that the long-range influence on the sub-picosecond collective water network motions of the hydration water is directly correlated with the average number of hydrogen bonds between the molecule and adjacent water molecules for carbohydrates. This implies that monosaccharides have a smaller influence on the surrounding water molecules than disaccharides. This could explain the bioprotection mechanism of sugar-water mixtures, which has been found to be more effective for disaccharides than for monosaccharides.

Antifreeze proteins (AFPs) are specific proteins that are able to lower the freezing point of aqueous solutions relative to the melting point. Hyperactive AFPs, identified in insects, have an especially high ability to depress the freezing point by far exceeding the abilities of other AFPs. In previous studies, we postulated that the activity of AFPs can be attributed to two distinct molecular mechanisms: (i) short-range direct interaction of the protein surface with the growing ice face and (ii) long-range interaction by protein-induced water dynamics extending up to 20 Å from the protein surface. In the present paper, we combine terahertz spectroscopy and molecular simulations to prove that long-range protein-water interactions make essential contributions to the high antifreeze activity of insect AFPs from the beetle Dendroides canadensis. We also support our hypothesis by studying the effect of the addition of the osmolyte sodium citrate.hydration dynamics | THz spetroscopy A ntifreeze proteins (AFPs) and antifreeze glycoproteins (AFGPs) are classes of proteins that suppress ice growth in organisms and thereby enable their survival in subfreezing habitats (1). Despite their similar function, many distinct structures have been identified so far. AFPs have been identified in several organisms, including polar fish (2), insects (3), bacteria (4), and plants (5). Their common characteristic is the depression of the freezing temperatures of ice growth of a solution without depressing the melting point equilibrium of protein solutions. This nonequilibrium phenomenon leads to a difference between the freezing and melting temperature, which is referred to as thermal hysteresis (TH). TH is used as a characteristic measure for antifreeze activity of an AFP (6). AFGPs and AFPs, as extracted from the blood of polar fish, usually exhibit up to 2°of TH activity and are termed moderately active AFPs, whereas insect AFPs can exhibit over 5°of TH and therefore, are referred to as hyperactive AFPs. The work by Raymond and DeVries (7,8) proposed a mechanism in which freezing point depression is achieved by an adsorption-inhibition mechanism, in which the proteins recognize and bind "quasiirreversibly" to an ice surface, thereby preventing growth of ice crystals. The adsorption of the protein is thought to prevent macroscopic ice growth in the hysteresis gap, but microscopic growth occurs at the interface in the form of highly curved fronts between adsorbed antifreeze molecules. This effect will cause a decrease of the local freezing temperature because of the Kelvin effect, while leaving the melting temperature relatively unaffected (7). As recently pointed out in the work by Sharp (9), antifreeze activity involves one of the most difficult recognition problems in biology, the distinction between water as liquid and ice. The initially proposed mechanism builds on a local mechanism. In particular, threonine (Thr) residues were proposed to play a decisive role: their hydroxyl groups were thought to be responsible for the high af...

The normal modes of myoglobin, their lifetimes, the speed of sound, and mean free path are calculated to determine the coefficient of thermal conductivity and thermal diffusivity for the protein. A propensity is found for frequency differences of pairs of normal modes localized to nearby regions of the protein to be several hundred cm-1. As a result, the anharmonic decay rate of higher frequency, localized normal modes, calculated by perturbation theory, is typically nearly independent of temperature, consistent with results of pump−probe studies on myoglobin. The thermal diffusivity of myoglobin at 300 K is calculated to be 14 Å2 ps-1, the same as the value for water. The thermal conductivity at 300 K is found to be 2.0 mW cm-1 K-1, about one-third the value for water.

We present terahertz (THz) measurements of salt solutions that shed new light on the controversy over whether salts act as kosmotropes (structure makers) or chaotropes (structure breakers), which enhance or reduce the solvent order, respectively. We have carried out precise measurements of the concentration-dependent THz absorption coefficient of 15 solvated alkali halide salts around 85 cm(-1) (2.5 THz). In addition, we recorded overview spectra between 30 and 300 cm(-1) using a THz Fourier transform spectrometer for six alkali halides. For all solutions we found a linear increase of THz absorption compared to pure water (THz excess) with increasing solute concentration. These results suggest that the ions may be treated as simple defects in an H-bond network. They therefore cannot be characterized as either kosmotropes or chaotropes. Below 200 cm(-1), the observed THz excess of all salts can be described by a linear superposition of the water absorption and an additional absorption that is attributed to a rattling motion of the ions within the water network. By providing a comprehensive set of data for different salt solutions, we find that the solutions can all be very well described by a model that includes damped harmonic oscillations of the anions and cations within the water network. We find this model predicts the main features of THz spectra for a variety of salt solutions. The assumption of the existence of these ion rattling motions on sub-picosecond time scales is supported by THz Fourier transform spectroscopy of six alkali halides. Above 200 cm(-1) the excess is interpreted in terms of a change in the wing of the water network librational mode. Accompanying molecular dynamics simulations using the TIP3P water model support our conclusion and show that the fast sub-picosecond motions of the ions and their surroundings are almost decoupled. These findings provide a complete description of the solute-induced changes in the THz solvation dynamics for the investigated salts. Our results show that THz spectroscopy is a powerful experimental tool to establish a new view on the contributions of anions and cations to the structuring of water.

We apply multireference ab initio quantum chemistry and microcanonical transition state (RRKM) theory with quantum energy flow corrections from local random matrix theory (LRMT) to determine the kinetics of trans-stilbene photoisomerization. With a single ab initio potential energy surface and no adjustable parameters, simultaneous agreement with experiment of the microcanonical isomerization rates for the d 0 , d 2 , d 10 , and d 12 isotopomers is obtained. We are also able to reproduce the pressure dependence of the thermal rate. Laser cooling effects on the isomerization rate are calculated and found to be quite small. The S 1 /S 2 energy gap at the transition state is found to be quite large (0.86 eV), suggesting that nonadiabatic effects are negligible. Using the ab initio results in a simple RRKM theory without corrections for finite quantum energy flow does not lead to agreement with experiment. We conclude that non-RRKM effects are essential to understand photoisomerization of trans-stilbene and that these can be predicted using LRMT.