The dynamics of the DNA hydration shell at gigahertz frequencies

We have used Brillouin scattering to measure the linewidths and frequencies of GHz acoustic phonons in Na- and Li-DNA films as a function of temperature between 300 and 140 K for samples that were dry, lightly, and heavily hydrated. The linewidths decrease with falling temperature and water contents, indicating that coupling to a water relaxation is the main source of phonon damping. The strength of the relaxation was determined using measurements of the phonon linewidth as a function of frequency, and confirmed by comparison of measured and calculated spectral profiles. The relaxation strength is anisotropic, being greater for phonons propagating perpendicular to the helix axis. The hydrated DNA exhibits both a rapid relaxation (≤ 10^?11 s per radian) giving rise to a classical f² damping, and a slower motion with a relaxation time that varies from ～ 4 × 10^?11 s per radian (primary hydration shell) to ～ 2 × 10^?12 s per radian (secondary hydration shell) at room temperature. In the frequency interval that bounds these relaxation times (～ 4 to 80 GHz) we expect degrees of freedom associated with the primary hydration shell to be important. The sample with primary hydration follows a simple Arrhenius behavior with ΔH ～ 5 kcal mole^?1. The effective activation energy for the sample with secondary hydration is somewhat higher (indicating a more cooperative water relaxation) and varies strongly with temperature. The elastic moduli change much more than can be accounted for by relaxation, indicating the importance of water motion in softening interatomic potentials. The extent of the softening caused by the “unfreezing” of water motion is similar to the degree of softening caused by hydrating the sample.     

Low-frequency Raman spectra of lysozyme crystals and oriented DNA films: dynamics of crystal water.

We observed low-frequency Raman spectra of tetragonal lysozyme crystals and DNA films, with varying water content of the samples. The spectra are fitted well by sums of relaxation modes and damped harmonic oscillators in the region from approximately 1 cm(-1) to 250 cm(-1). The relaxation modes are due to crystal water, and the distribution of relaxation times is determined. In wet samples, the relaxation time of a small part of the water molecules is a little longer than that of bulk water. The relaxation time of a considerable part of the crystal water, which belongs mainly to the secondary hydration shell, is an order of magnitude longer than that of bulk water. Furthermore, the relaxation time of some water molecules in the primary hydration shell of semidry samples is shorter than we expected. Thus we have shown that low-frequency Raman measurements combined with properly oriented samples can give specific information on the dynamics of hydration water in the ps range. On the other hand, we concluded, based on polarized Raman spectra of lysozyme crystals, that the damped oscillators correspond to essentially intramolecular vibrational modes.     

A Raman scattering study of the interactions of DNA with its water of hydration

Raman spectroscopy is used to probe the nature of the hydrogen bonds which hold the water of hydration to DNA. The ～ 3450?cm^?1 molecular O–H stretching mode shows that the first six water molecules per base pair of the primary hydration shell are very strongly bound to the DNA. The observed shift in the peak position of this mode permits a determination of the length of the hydrogen bonds for these water molecules. These hydrogen bonds appear to be about 0.3?Å shorter than the hydrogen bonds in bulk water. The linewidth of this mode shows no significant changes above water contents of about 15 water molecules per base pair. This technique of using a vibrational spectroscopy to obtain structural information about the hydration shells of DNA could be used to study the hydration shells of other biomolecules.     

Photo-induced electron transport and water state in Rhodospirillum rubrum chromatophores

It is shown that in bacterial chromatophores the pronounced changes in the free water content with a proton spin-spin relaxation time (T₂) of 10^?3—10^?2 s does not influence the efficiency of electron transfer from the photosynthetic reaction centre to the membrane pool of secondary acceptors. An abrupt inhibition of this process occurs only after the loss of the water with faster proton spin-spin relaxation time (T₂ of 10^?4 s). The process is reversible. The water fraction in question is obviously bound to the chromatophore proteins and forms the primary hydration layer.     

Rotational and translational water diffusion in the hemoglobin hydration shell: dielectric and proton nuclear relaxation measurements.

The dynamic properties of water in the hydration shell of hemoglobin have been studied by means of dielectric permittivity measurements and nuclear magnetic resonance spectroscopy. The temperature behavior of the complex permittivity of hemoglobin solutions has been measured at 3.02, 3.98, 8.59, and 10.80 GHz. At a temperature of 298 K the average rotational correlation time tau of water within a hydration shell of 0.5-nm thickness is determined from the activation parameters to be 68 +/- 10 ps, which is 8-fold the corresponding value of bulk water. Solvent proton magnetic relaxation induced by electron-nuclear dipole interaction between hemoglobin bound nitroxide spin labels and water protons is used to determine the translational diffusion coefficient D(T) of the hydration water. The temperature dependent relaxation behavior for Lamor frequencies between 3 and 90 MHz yields an average value D(298K) = (5 +/- 2) x 10(-10)m2 s-1, which is about one-fifth of the corresponding value of bulk water. The decrease of the water mobility in the hydration shell compared to the bulk is mainly due to an enhanced activation enthalpy.     

Conductometric evaluation of interactions of electrolytes with D-glucitol, D-glucose, glycerol, D-mannitol, and sucrose

By measurement of the conductance of various electrolytes in solutions of D-glucitol, D-glucose, glycerol, D-mannitol, and sucrose, the hydration of the polyhydroxy compounds and their association with electrolytes have been examined. The results have been analyzed in the light of conductance equations; it was found that D-glucose and sucrose associate with electrolytes, glycerol and D-mannitol are practically inert, and D-glucitol is only rarely bound. The association constants are of the order of 10^?1 to 10^?2. The results indicate that anions guide the process, with a free-energy change of 1.5–3.0 kcal.mole^-1, the order of the energy of hydrogen bonding. In the analysis of conductance data, neglect of association may introduce hydration numbers many times greater than are usual.     

The state of water in the outer barrier of the isolated frog skin

Summary The flux of water across the outer barrier of the frog skin is generally regarded as the rate-limiting step in the movement of water across the whole membrane. This paper presents some evidence that, at room temperature, the flux of water across the outer barrier occurs through water in a non-liquid state. The organization of water in a non-liquid state lowers the diffusion coefficient of water through water by several orders of magnitude. The study employs a method recently developed in this laboratory which permits measurement of unidirectional fluxes at the outermost part of an epithelial membrane mounted as a flat sheet. Only above 25°C is the activation energy for the flow of tritiated water (4.3 kcal mole⁻¹) similar to the one observed in free water (4.6 kcal mole⁻¹). At temperatures around 15°C, the energy of activation is 8.5 kcal mole⁻¹. At temperatures near 0°C, at which the frog lives only part of the year, the energy of activation is 16.7 kcal mole⁻¹.     

Theory of acoustic mode vibrations of DNA fibers

A previous model for acoustic mode vibrations of a DNA molecule in water is extended to the case of an array of many DNA molecules, as occurs in the fibers studied in most experimental work on DNA. The acoustic modes of this system are found to consist of coupled modes of water sound vibrations and DNA acoustic modes. This model is used to study the electrostatic coupling of acoustic vibrations to the relaxational modes of the orientational degrees of freedom of the water molecules. It is found that the long-range or macroscopic electric field generated by the acoustic mode vibrations of the water-DNA system gives too small a damping and frequency shift of the acoustic modes to account for the observations on DNA fibers. Therefore, the observed damping and frequency shifts are most likely due to either friction between the surrounding water and the vibrating DNA, or coupling to the water orientation degrees of freedom resulting from the short range (i.e., screened) Coulomb interaction. The latter explanation (which is most likely the correct one) implies that the relaxation time of the hydration shell water is longer than the observed relaxation time by a factor of the static dielectric constant of the hydration water.     

Microsolvation and hydration enthalpies of CaC2O4(H2O) n (n = 0-16) and C2O4 2-(H2O) n (n = 0-14): an ab initio study

We studied hydrated calcium oxalate and its ions at the restricted Hartree–Fock RHF/6-31G* level of theory. Performing a configurational search seems to improve the fit of the HF/6-31G* level to experimental data. The first solvation shell of calcium oxalate contains 13 water molecules, while the first solvation shell of oxalate ion is formed by 14 water molecules. The first solvation shell of Ca(II) is formed by six water molecules, while the second shell contains five. At 298.15 K, we estimate the asymptotic limits (infinite dilution) of the total standard enthalpies of hydration for Ca(II), oxalate ion and calcium oxalate as ?480.78, –302.78 and –312.73 kcal mol^?1, resp. The dissociation of hydrated calcium oxalate is an endothermic process with an asymptotic limit of +470.84 kcal mol^?1.

Hydration of Escherichia coli lipids. Deuterium T1 relaxation time studies of phosphatidylglycerol, phosphatidylethanolamine and phosphatidylcholine

The hydration properties of Escherichia coli lipids (phosphatidylglycerol, phosphatidylethanolamine) and synthetic 1,2-dioleoyl-sn-glycero-3-phosphocholine in H2O/2H2O mixtures (9:1, v/v) were investigated with 2H-NMR. Comparison of the 2H2O spin lattice relaxation time (T1) as a function of the water content revealed a remarkable quantitative similarity of all three lipid-H2O systems. Two distinct hydration regions could be discerned in the T1 relaxation time profile. (1) A minimum of 11-16 water molecules was needed to form a primary hydration shell, characterized by an average relaxation time of T1 approximately equal to 90 ms. (2) Additional water was found to be in exchange with the primary hydration shell. The exchange process could be described in terms of a two-site exchange model, assuming rapid exchange between bulk water with T1 = 500 ms and hydration water with T1 = 80-120 ms. Analysis of the linewidth and the residual quadrupole splitting (at low water content) confirmed the size of the primary hydration layer. However, each lipid-water system exhibited a somewhat different linewidth behavior, and a detailed molecular interpretation appeared to be preposterous.     

Structure and dynamics of primary hydration shell of phosphatidylcholine bilayers at subzero temperatures.

Deuterium NMR relaxation and intensity measurements of the 2H-labeled H2O/dimyristoyl phosphatidylcholine bilayer were performed to understand the molecular origin of the freezing event of phospholipid headgroup and the structure and dynamics of unfrozen water molecules in the interbilayer space at subzero temperatures. The results suggest that about one to two water molecules associated with the phosphate group freeze during the freezing event of phospholipid headgroups, whereas about five to six waters near the trimethylammonium group behave as a water cluster and remain unfrozen at temperatures as low as -70 degrees C. In addition, temperature-dependent T1 and T2 relaxation times suggest that dynamic coupling occurs not only between the phosphate group and its bound water, but also between the methyl group and the adjacent water molecules. Based on these observations, the primary hydration shell of phosphatidylcholine headgroup at subzero temperatures is suggested to consist of two distinct regions: a clathrate-like water cluster, most likely a water pentamer, near the hydrophobic methyl group, and hydration water molecules associated with the phosphate group.     

Hydrational and intrinsic compressibilities of globular proteins

Partial compressibilities of globular proteins in water are reviewed. Contribution of hydrational and of intrinsic compressibilities to experimental partial quantity have been evaluated from ultrasonic data using two independent methods: (a) additive calculation of the hydrational contributions of the surface atomic groups and (b) an analysis of correlation between partial compressibility and molecular surface area. The value (14 ± 3) × 10^?6 bar ^?1 for the isothermal compressibility coefficient of the protein interior at 25°C was obtained as an average value for variety of globular proteins. This value is similar to that of solid organic polymers. Possible relaxation contribution to partial compressibility is roughly estimated from comparison of thermodynamic with x-ray data on protein compressibility. The average compressibility of water in the hydration shell of proteins was found to be 35 × 10^?6 bar ^?1, which is 20% less than that of pure water. © 1993 John Wiley & Sons, Inc.     

A study of vibrational relaxation of B-state carbon monoxide in the heme pocket of photolyzed carboxymyoglobin.

The vibrational energy relaxation of dissociated carbon monoxide in the heme pocket of sperm whale myoglobin has been studied using equilibrium molecular dynamics simulation and normal mode analysis methods. Molecular dynamics trajectories of solvated myoglobin were run at 300 K for both the delta- and epsilon-tautomers of the distal histidine, His64. Vibrational population relaxation times were estimated using the Landau-Teller model. For carbon monoxide (CO) in the myoglobin epsilon-tautomer, for a frequency of omega0 = 2131 cm-1 corresponding to the B1 state, T1epsilon(B1) = 640 +/- 185 ps, and for a frequency of omega0 = 2119 cm-1 corresponding to the B2 state, T1epsilon(B2) = 590 +/- 175 ps. Although the CO relaxation rates in both the epsilon- and delta-tautomers are similar in magnitude, the simulations predict that the vibrational relaxation of the CO is faster in the delta-tautomer. For CO in the myoglobin delta-tautomer, it was found that the relaxation times were identical within error for the two CO substate frequencies, T1delta(B1) = 335 +/- 115 ps and T1delta(B2) = 330 +/- 145 ps. These simulation results are in reasonable agreement with experimental results of Anfinrud and coworkers (unpublished results). Normal mode calculations were used to identify the dominant coupling between the protein and CO molecules. The calculations suggest that the residues of the myoglobin pocket, acting as a first solvation shell to the CO molecule, contribute the primary "doorway" modes in the vibrational relaxation of the oscillator.     

Dynamics of uncrystallized water and protein in hydrated elastin studied by thermal and dielectric techniques

Dynamics of uncrystallized water and protein was studied in hydrated pellets of the fibrous protein elastin in a wide hydration range (0 to 23 wt.%), by differential scanning calorimetry (DSC), thermally stimulated depolarization current technique (TSDC) and dielectric relaxation spectroscopy (DRS). Additionally, water equilibrium sorption–desorption measurements (ESI) were performed at room temperature. The glass transition of the system was studied by DSC and its complex dependence on hydration water was verified. A critical water fraction of about 18 wt.% was found, associated with a reorganization of water in the material. Three dielectric relaxations, associated to dynamics related to distinct uncrystallized water populations, were recorded by TSDC and DRS. The low temperature secondary relaxation of hydrophilic polar groups on the protein surface triggered by hydration water for almost dry samples contains contributions from water molecules themselves at higher water fractions (ν relaxation). This particular relaxation is attributed to water molecules in the primary and secondary hydration shells of the protein fibers. At higher temperatures and for water fraction values equal to or higher than 10 wt.%, a local relaxation of water molecules condensed within small openings in the interior of the protein fibers was recorded. The evolution of this relaxation (w relaxation) with hydration level results in enhanced cooperativity at high water fraction values, implying the existence of “internal” water confined within the protein structure. At higher temperatures a relaxation associated with water dynamics within clusters between fibers (p relaxation) was also recorded, in the same hydration range.     

The enthalpy of oxidation of flavin mononucleotide

The oxidation enthalpy of reduced flavin mononucleotide at pH 7.0 in 0.2 m phosphate buffer has been studied by determining the heat associated with the reaction: FMNH₂ + 2 Fe(CN)^?3₆ ? FMN + 2 Fe(CN)^?4₆ + 2 H⁺. (a) (The quinone, semiquinone, and hydroquinone forms of FMN are represented as FMN, FMNH, and FMNH₂, respectively.) Calorimetric experiments were performed in a flow microcalorimeter which was modified to prevent sample contamination by oxygen. The enthalpy observed for reaction (a), after correction for dilution and buffer effects, was ?39.2 ± 0.4 kcal (mole FMNH₂)^?1 at 25 °C. The potential difference, ΔE′, developed by reaction (a) was determined potentiometrically and corresponded to a free energy change, ΔG′, of ?30.3 kcal (mole FMNH₂)^?1. The resulting entropy change, ΔS′, was thus calculated to be ?29.8 e.u. Reaction (a) was also studied at temperatures of 7 °C and 35.5 °C. ΔC_p′ for the reaction was calculated as ?155 ± 18 cal deg^?1 (mole FMNH₂)^?1 at 20 °C. ΔH′ for the reaction (b), FMNH₂ ? FMN + H₂, (b) was calculated as +14.2 ± 0.7 kcal mole^?1 at 25 °C, relative to the enthalpy of the hydrogen electrode being identically equal to zero at all values of pH and temperature. The free energy at pH 7.0 for reaction (b), calculated from the potential was found to be ?9.7 kcal mole^?1, which resulted in an entropy for reaction (b) of 80.2 e.u. A thermal titration of reaction (a) was used to calculate the thermodynamic parameters for the formation of semiquinone dimer according to the reaction FMNH₂ + FMN ? (·FMNH)₂. (c) The free energy, enthalpy, and entropy changes for reaction (c) were estimated to be ?6.1 kcal mole^?1, ?7 kcal mole^?1, and ?3 e.u., respectively.     

Hydration of Escherichia coli lipids: Deuterium T1 relaxation time studies of phosphatidylglycerol,phosphatidylethanolamine and phosphatidylcholine

The hydration properties of Escherichia coli lipids (phosphatidylglycerol, phosphatidylethanolamine) and synthetic $\text{1,2-}\text{dioleoyl}\text{-sn-}\text{glycero-3-phosphocholine}$ in H₂O/²H₂O mixtures (9:1, v/v) were investigated with ²H-NMR. Comparison of the ²H₂O spin lattice relaxation time ($\text{T}{}_1$) as a function of the water content revealed a remarkable quantitative similarity of all three lipid-H₂O systems. Two distinct hydration regions could be discerned in the $\text{T}{}_1$ relaxation time profile. (1) A minimum of 11–16 water molecules was needed to form a primary hydration shell, characterized by an average relaxation time of $\text{T}{}_1\text{≈ 90}\text{ms}$. (2) Additional water was found to be in exchange with the primary hydration shell. The exchange process could be described in terms of a two-site exchange model, assuming rapid exchange between bulk water with $\text{T}{}_1\text{= 500}\text{ms}$ and hydration water with $\text{T}{}_1\text{= 80–120}\text{ms}$. Analysis of the linewidth and the residual quadrupole splitting (at low water content) confirmed the size of the primary hydration layer. However, each lipid-water system exhibited a somewhat different linewidth behavior, and a detailed molecular interpretation appeared to be preposterous.     

The influence of a protein on water dynamics in its vicinity investigated by molecular dynamics simulation

A system containing the globular protein ubiquitin and 4,197 water molecules has been used for the analysis of the influence exerted by a protein on solvent dynamics in its vicinity. Using Voronoi polyhedra, the solvent has been divided into three subsets, i.e., the first and second hydration shell, and the remaining bulk, which is hardly affected by the protein. Translational motion in the first shell is retarded by a factor of 3 in comparison to bulk. Several molecules in the first shell do not reach the diffusive regime within 100 ps. Shell-averaged orientational autocorrelation functions, which are also subject to a retardation effect, cannot be modeled by a single exponential time law, but are instead well-described by a Kohlrausch-Williams-Watts (KWW) function. The underlying distribution of single-molecule rotational correlation times is both obtained directly from the simulation and derived theoretically. The temperature dependence of reorientation is characterized by a strongly varying correlation time, but a virtually temperature-independent KWW exponent. Thus, the coupling of water structure relaxation with the respective environment, which is characteristic of each solvation shell, is hardly affected by temperature. In other words, the functional form of the distributions of single-molecule rotational correlation times is not subject to a temperature effect. On average, a correlation between reorientation and lifetimes of neighborhood relations is observed. © 1996 Wiley-Liss, Inc.     

Divalent paramagnetic counterions site binding in polyelectrolyte solutions. Analysis of the frequency dependence of the water protons magnetic relaxation and the characteristic parameters of "site binding"

Chemical shift and relaxation time measurements on the water protons in polyelectrolyte solutions containing divalent paramagnetic counterions have shown the existence of three types of counterions: - site bound with loss of water molecules and partial or complete release of the electrostriction in the first hydration sphere, - atmospherically trapped with no change in hydration, - free. The overall stoichiometry of the two former is in agreement with Manning's fraction of condensed counterions. A complete analysis of the frequency dependent contribution of site bound counterions to the water protons relaxation times leads us to interesting conclusions on the modifications of the first hydration shell and on the life time of site binding.     

Effect of hydration upon the thermal stability of tropocollagen and its dependence on the presence of neutral salts

The endotherm enthalpy changes ΔH_D and temperatures T_D of thermal denaturation of tropocollagen fibers were measured by DSC calorimetry as functions of water content. The denaturation temperatures decrease with increasing water content. The enthalpy change values increase sharply in the range 0–28% of water content, where a maximum of 14.3 cal g^?1 is reached. The effect of water uptake on the enthalpy term is explained by water bridge formation within the collagen triple helix. Evidence is given for the existence of approximately three intercatenary water bridges per triplet at the enthalpy maximum, their H-bond energy amounting to approximately 4000 kcal/mol of protein. In the 30–60% range of water content, ΔH_D decreases by 2 cal^?1 probably due to interactions between secondary water structures and the stabilizing intrahelical water bonds. The influence of two neutral potassium salts, with a structure-stabilizing and a structure-breaking anion (F^? and I^?), on the hydration dependence of ΔH_D and T_D was also studied. It was shown that the primary hydration is not influenced by these ions, but that T_D and ΔH_D are altered in an ion specific way in the presence of interface and bulk water. Hydrophobic interactions do not explain the experimental results. A reaction mechanism of the effects of ions upon the structural stability of collagen is proposed and discussed in terms of interactions of the medium water molecules with the intrahelical water bonds, and in terms of proton-donor/proton-acceptor equilibria between peptide groups, hydrated ions, and intrahelical water molecules.     

Hydration dependent dynamics in RNA

The essential role played by local and collective motions in RNA function has led to a growing interest in the characterization of RNA dynamics. Recent investigations have revealed that even relatively simple RNAs experience complex motions over multiple time scales covering the entire ms–ps motional range. In this work, we use deuterium solid-state NMR to systematically investigate motions in HIV-1 TAR RNA as a function of hydration. We probe dynamics at three uridine residues in different structural environments ranging from helical to completely unrestrained. We observe distinct and substantial changes in ²H solid-state relaxation times and lineshapes at each site as hydration levels increase. By comparing solid-state and solution state ¹³C relaxation measurements, we establish that ns–μs motions that may be indicative of collective dynamics suddenly arise in the RNA as hydration reaches a critical point coincident with the onset of bulk hydration. Beyond that point, we observe smaller changes in relaxation rates and lineshapes in these highly hydrated solid samples, compared to the dramatic activation of motion occurring at moderate hydration.