Structure,dynamics and thermodynamic stability of high-pressure ices and clathrate hydrates

We present the recent development of simulation studies on structure and dynamics of high-pressure ices and filled ices. After surveying the representative structures of ices, focus is placed on some properties of one of the ice polymorphs, plastic ice, which has been theoretically predicted but not yet been found experimentally. Its intervention between ice VII and liquid water enables to account for large discrepancies among various experimental melting curves of ice VII. We also examine the dynamical properties of the filled ice in which hydrogen molecules are contained in void space of the low-pressure cubic ice. In contrast to the plastic ice, which has a bearing on ice VII, it exhibits a gradual change to each rotator phase of guest hydrogen and/or host water molecules.     

Liquid-like water confined in stacks of biological membranes at 200 k and its relation to protein dynamics

Confined water is of considerable current interest owing to its biophysical importance and relevance to cryopreservation. It can be studied in its amorphous or supercooled state in the "no-man's land", i.e., in the temperature range between 150 and 235 K, in which bulk water is always crystalline. Amorphous deuterium oxide (D(2)O) was obtained in the intermembrane spaces of a stack of purple membranes from Halobacterium salinarum by flash cooling to 77 K. Neutron diffraction showed that upon heating to 200 K the intermembrane water space decreased sharply with an associated strengthening of ice diffraction, indicating that water beyond the first membrane hydration layer flowed out of the intermembrane space to form crystalline ice. It was concluded that the confined water undergoes a glass transition at or below 200 K to adopt an ultraviscous liquid state from which it crystallizes to form ice as soon as it finds itself in an unconfined, bulk-water environment. Our results provide model-free evidence for translational diffusion of confined water in the no-man's land. Potential effects of the confined-water glass transition on nanosecond membrane dynamics were investigated by incoherent elastic neutron scattering experiments. These revealed no differences between flash-cooled and slow-cooled samples (in the latter, the intermembrane space at temperatures <250 K is occupied only by the first membrane hydration layers), with dynamical transitions at 150 and 260 K, but not at 200 K, suggesting that nanosecond membrane dynamics are not sensitive to the state of the water beyond the first hydration shell at cryotemperatures.     

A Detailed Study of the Amino Acids Produced from the Vacuum UV Irradiation of Interstellar Ice Analogs

We present a detailed analysis of the variety, quantity and distribution of the amino acids detected in organic residues after acid hydrolysis. Such organic residues are produced in the laboratory after the vacuum ultraviolet (VUV) irradiation of several astrophysically relevant ice mixtures containing H(2)O, CO, CO(2), CH(3)OH, CH(4) and NH(3) at low temperature (10-80 K), and subsequent warm-up to room temperature. We explore five experimental parameters: the irradiation time, the temperature, the ice mixture composition, the photon dose per molecule and the substrate for the ice deposition. The amino acids were detected and identified by ex-situ liquid chromatography analysis of the organic residues formed after warming the photolysed ices up to room temperature. This study shows that in all experiments amino acids are formed. Their total quantities and distribution depend slightly on the experimental parameters explored in the present work, the important requirement to form such molecules being that the starting ice mixtures must contain the four elements C, H, O and N. We also discuss the effects of the chemical treatment needed to detect and identify the amino acids in the organic residues. Finally, these results are compared with meteoritic amino acid data from the carbonaceous chondrite Murchison, and the formation processes of such compounds under astrophysical conditions are discussed.     

Atomic force microscopy of hydrated phosphatidylethanolamine bilayers.

We present images of the polar or headgroup regions of bilayers of dimyristoyl-phosphatidylethanolamine (DMPE), deposited by Langmuir-Blodgett deposition onto mica substrates at high surface pressures and imaged under water at room temperature with the optical lever atomic force microscope. The lattice structure of DMPE is visualized with sufficient resolution that the location of individual headgroups can be determined. The forces are sufficiently small that the same area can be repeatedly imaged with a minimum of damage. The DMPE molecules in the bilayer appear to have relatively good long-range orientational order, but rather short-range and poor positional order. These results are in good agreement with x-ray measurements of unsupported lipid monolayers on the water surface, and with electron diffraction of adsorbed monolayers.     

Freezing avoidance by supercooling in Olea europaea cultivars: the role of apoplastic water,solute content and cell wall rigidity

Plants can avoid freezing damage by preventing extracellular ice formation below the equilibrium freezing temperature (supercooling). We used Olea europaea cultivars to assess which traits contribute to avoid ice nucleation at sub‐zero temperatures. Seasonal leaf water relations, non‐structural carbohydrates, nitrogen and tissue damage and ice nucleation temperatures in different plant parts were determined in five cultivars growing in the Patagonian cold desert. Ice seeding in roots occurred at higher temperatures than in stems and leaves. Leaves of cold acclimated cultivars supercooled down to ?13 °C, substantially lower than the minimum air temperatures observed in the study site. During winter, leaf ice nucleation and leaf freezing damage (LT₅₀) occurred at similar temperatures, typical of plant tissues that supercool. Higher leaf density and cell wall rigidity were observed during winter, consistent with a substantial acclimation to sub‐zero temperatures. Larger supercooling capacity and lower LT₅₀ were observed in cold‐acclimated cultivars with higher osmotically active solute content, higher tissue elastic adjustments and lower apoplastic water. Irreversible leaf damage was only observed in laboratory experiments at very low temperatures, but not in the field. A comparative analysis of closely related plants avoids phylogenetic independence bias in a comparative study of adaptations to survive low temperatures.     

Calorimetric study of crystal growth of ice in hydrated methemoglobin and of redistribution of the water clusters formed on melting the ice.

Calorimetric studies of the melting patterns of ice in hydrated methemoglobin powders containing between 0.43 and 0.58 (g water)/(g protein), and of their dependence on annealing at subzero temperatures and on isothermal treatment at ambient temperature are reported. Cooling rates were varied between approximately 1500 and 5 K min-1 and heating rate was 30 K min-1. Recrystallization of ice during annealing is observed at T > 228 K. The melting patterns of annealed samples are characteristically different from those of unannealed samples by the shifting of the melting temperature of the recrystallized ice fraction to higher temperatures toward the value of "bulk" ice. The "large" ice crystals formed during recrystallization melt on heating into "large" clusters of water whose redistribution and apparent equilibration is followed as a function of time and/or temperature by comparison with melting endotherms. We have also studied the effect of cooling rate on the melting pattern of ice with a methemoglobin sample containing 0.50 (g water)/(g protein), and we surmise that for this hydration cooling at rates of > or = approximately 150 K min-1 preserves on the whole the distribution of water molecules present at ambient temperature.     

The influence of hydroxyethyl starch on ice formation in aqueous solutions

Differential scanning calorimetry, and, in some supplementary experiments, X-ray diffractometry and cryomicroscopy, were applied to study the influence of concentration (< 70 wt%) and cooling/warming rates (< 320 K/min) on ice formation in aqueous solutions of HES. The calorimetric measurements of the quantity of crystallizing water indicated that a mass fraction ? = 0.522 (i.e., grams water per gram HES) remained unfrozen. These results are in good agreement with our earlier extrapolations from ternary phase diagram data and tend to support the proposed cryoprotective mechanism. The value of ? determined during warming was essentially independent of composition up to the corresponding saturation concentration. It was observed that solutions containing 60 wt% HES or more remained wholly amorphous during cooling even at rates as low as 2.5 K/min (down to 120 K). Such glassy solutions are subject to devitrification at temperatures T_d which depend on the warming rate. The concentrations close to 55 wt% HES mark a transitional range exhibiting two crystallization peaks, probably due to different mechanisms of nucleation, the portion of ice formed during cooling being related to the imposed cooling rate. All samples showed a recrystallization transition at 257.5 K which was also observed cryomicroscopically. Glass transitions, however, could not be detected by the methods applied in this study. The X-ray diffraction patterns contained the structure of only one solid phase, namely hexagonal ice. A comparison of various modifications of HES, PEG, and PVP involving bound water and melting temperature did not reveal marked differences. Minimum initial HES concentrations preventing lethal salt enrichment were computed for both binary and ternary mass fractions of NaCl as biologically relevant parameters, yielding 24.1 and 10.8 wt% HES, respectively.     

Structural Transformations of Ice at High Pressures Via Molecular Dynamics Simulations

Abstract

A simple classical model is used for the study of the structural transformations of ice under high pressures, such as ice VIII to VII and X, via classical molecular dynamics (MD) simulation. In the present MD simulation, pair potentials of a simple form between pair of atoms and a thee-body potential representing the H-O-H angle dependence, originally developed by Kawamura et al., were used. Starting with a stable ice VIII at low pressure and low temperature, we have carried out two different MD runs, one with increasing pressure keeping the temperature constant (simulation I) and the other with increasing temperature under constant pressure (simulation II). From these MD simulations we have obtained the structural transformations from ice VIII to VII for both simulations; the former was finally transformed into ice X for the simulation I. The present results are compatible with recent experiments on high pressure ices.     

Role of hydration water in protein unfolding

In this paper, following our work on the two-state outer neighbor mixed bonding model of water, it is proposed that polar groups promote the formation of the low density ice Ih-type bonding in their neighborhood, whereas nonpolar groups tend to promote the higher density ice II-type structure. In a protein, because of the large numbers of exposed polar and nonpolar groups, large changes in the neighboring water structure can occur. These changes, of course, depend on whether the protein is in its native or its unfolded state and will be shown here to have a direct impact on the thermodynamics of protein unfolding at both high and low temperatures. For example, it is known that the polar hydration entropies become rapidly more negative with increasing temperature. This very unusual behavior can be directly related to the promotion in the outer bulk liquid of the more stable Ih-type bonding at the expense of II-type bonding by polar groups of the protein. In contrast, nonpolar groups have an opposite effect on the thermodynamics. It is the delicate balance created by these outer hydration contributions, mixed with ordinary thermodynamic contributions from the inner hydration shell and those from hydrogen-bond and van der Waals forces within the protein molecule itself that is responsible for both heat and cold denaturation of proteins.     

Ice segregation in the crown of winter cereals: Evidence for extraorgan and extratissue freezing

Meaningful improvements in winter cereal cold hardiness requires a complete model of freezing behaviour in the critical crown organ. Magnetic resonance microimaging diffusion‐weighted experiments provided evidence that cold acclimation decreased water content and mobility in the vascular transition zone (VTZ) and the intermediate zone in rye (Secale cereale L. Hazlet) compared with wheat (Triticum aestivum L. Norstar). Differential thermal analysis, ice nucleation, and localization studies identified three distinct exothermic events. A high‐temperature exotherm (?3°C to ?5°C) corresponded with ice formation and high ice‐nucleating activity in the leaf sheath encapsulating the crown. A midtemperature exotherm (?6°C and ?8°C) corresponded with cavity ice formation in the VTZ but an absence of ice in the shoot apical meristem (SAM). A low‐temperature exotherm corresponded with SAM injury and the killing temperature in wheat (?21°C) and rye (?27°C). The SAM had lower ice‐nucleating activity and freezing survival compared with the VTZ when frozen in vitro. The intermediate zone was hypothesized to act as a barrier to ice growth into the SAM. Higher cold hardiness of rye compared with wheat was associated with higher VTZ and intermediate zone desiccation resulting in the formation of ice barriers surrounding the SAM.     

Conformational transition in DNA on a cold surface

The contour length of DNA fragments, deposited and imaged on mica under buffer, was measured as a function of deposition temperature. Extended DNA molecules (on Ni- and silane-treated surfaces) contract rapidly with falling temperature, approaching the contour length of A-DNA at 2°C. The contraction is not unique to a specific sequence and does not occur in solution at 2°C or on a surface at 25°C, indicating that it arises from a combination of low temperature and surface contact. It is probably a consequence of reduced water activity at a cold surface.     

Temperature dependence of the generalized frequency distribution of water molecules: comparison of experiments and molecular dynamics simulations

The results of inelastic neutron scattering experiments on water in the temperature interval 300–623 K along the coexistence curve are compared with data obtained from molecular dynamics simulations. In general, a good agreement between experiments and calculations is observed and it serves as a satisfactory test of the potential models employed. The temperature dependence of the generalized frequency distribution of water molecules obtained by both experiment and computer simulation demonstrates the accordance with the temperature evolution of the water structure, extracted from neutron and X-ray diffraction measurements.     

Molecular dynamics simulations of a cyclic-DP-240 amylose fragment in a periodic cell: Glass transition temperature and water diffusion

Molecular dynamics simulations using AMB06C, an in-house carbohydrate force field, (NPT ensembles, 1 atm) were carried out on a periodic cell that contained a cyclic 240 glucose residue amylose fragment (c-DP-240) and TIP3P water molecules. Molecular conformation and movement of the amylose fragment and water molecules at different temperatures were examined. The periodic cell volume, density, and potential energy were determined at temperatures above and below the glass transition temperature (T_g) in 25 K increments_. The amorphous cell is constructed through successive dynamic equilibration steps at temperatures above the assumed T_g value and the temperature successively lowered until several temperature points were obtained below T_g. Molecular dynamics simulations were continued for at least 500 ps or until the volume drift stopped and remained constant for several hundred picoseconds. The T_g values were found by noting the discontinuity in slope of the volume (V), potential energy (PE), or density (ρ) versus 1/T. The changes in flexibility and motion of the amylose chain as well as differences in self diffusion coefficients of water molecules are described. The final average T_g value found (316 K) is in agreement with experimental values, i.e. 320 K.     

Temperature dependence of protein-hydration hydrodynamics by molecular dynamics simulations

The dynamics of water molecules near the protein surface are different from those of bulk water and influence the structure and dynamics of the protein itself. To elucidate the temperature dependence hydration dynamics of water molecules, we present results from the molecular dynamic simulation of the water molecules surrounding two proteins (Carboxypeptidase inhibitor and Ovomucoid) at seven different temperatures (T=273 to 303 K, in increments of 5 K). Translational diffusion coefficients of the surface water and bulk water molecules were estimated from 2 ns molecular dynamics simulation trajectories. Temperature dependence of the estimated bulk water diffusion closely reflects the experimental values, while hydration water diffusion is retarded significantly due to the protein. Protein surface induced scaling of translational dynamics of the hydration waters is uniform over the temperature range studied, suggesting the importance protein-water interactions.     

Thermal properties of water in myoglobin crystals and solutions at subzero temperatures.

The water of hydration in myoglobin crystals and solutions was studied at subzero temperatures by calorimetry and infrared spectroscopy (ir). For comparison we also investigated glycine, DL-alanine and DL-valine solutions. The hydration water remains amorphous at low temperatures. We find a broad glass transition between 180 and 270 K depending on the degree of hydration. The ice component shows a noncolligative melting point depression that is attributed to a finite conformational flexibility. The ir spectrum and the specific heat of water in myoglobin crystals was determined for the first time between 180 and 290 K. The glass transition in crystals is qualitatively similar to what is found in amorphous samples at the same water content. These data are compared with M?ssbauer experiments and dielectric relaxation of water in myoglobin crystals. The similar temperature dependencies suggest a cross correlation between structural fluctuations and the thermal motion of crystal water. A hydrogen bond network model is proposed to explain these features. The essential ingredients are cooperativity and a distribution of hydrogen-bonded clusters.     

Self-diffusion of nonfreezing water in porous carbohydrate polymer systems studied with nuclear magnetic resonance

Water is an integral part of the structure in biological porous materials such as wood and starch. A problem often encountered in the preparation of samples for, e.g., electron microscopy is that removal of water leads to a decreasing distance between supermolecular structural elements and a distortion of the structure. It is, therefore, of interest to find methods to investigate these materials in the native water-swollen state. We present a method to study water-swollen biological porous structures using NMR to determine the amount and self-diffusion of water within the porous objects. The contribution of bulk water to the NMR signal is eliminated by performing experiments below the bulk freezing temperature. Further decrease of the temperature leads to a gradual freezing of water within the porous objects. The contribution of the freezing water fraction to the migration of water through the porous network is, thus, estimated. The results are rationalized in terms of the ultrastructure of the samples studied, namely, wood pulp fibers and potato starch granules.     

Solution and surface effects on plasma fibronectin structure

As assessed by electron microscopy, the reported shape of the plasma fibronectin molecule ranges from that of a compact particle to an elongated, rod-like structure. In this study, we evaluated the effects of solution and surface conditions on fibronectin shape. Freeze-dried, unstained human plasma fibronectin molecules deposited at pH 7.0-7.4 onto carbon films and examined by scanning transmission electron microscopy appeared relatively compact and pleiomorphic, with approximate average dimensions of 24 nm X 16 nm. Negatively stained molecules also had a similar shape but revealed greater detail in that we observed irregular, yarn-like structures. Glutaraldehyde-induced intramolecular cross-linking did not alter the appearance of plasma fibronectin. Molecules deposited at pH 2.8, pH 9.3, or after succinylation were less compact than those deposited at neutral pH. In contrast, fibronectin molecules sprayed onto mica surfaces at pH 7, rotary shadowed, and examined by transmission electron microscopy were elongated and nodular with a contour length of 120-130 nm. Sedimentation velocity experiments and electron microscopic observations indicate that fibronectin unfolds when it is succinylated, when the ionic strength is raised at pH 7, or when the pH is adjusted to 9.3 or 2.8. Greater unfolding is observed at pH 2.8 at low ionic strength (less than 0.01) compared with material at that pH in 0.15 M NaCl solution. We conclude that (a) the shape assumed by the fibronectin molecule can be strongly affected by solution conditions and by deposition onto certain surfaces; and that (b) the images of fibronectin seen by scanning transmission electron microscopy at neutral pH on carbon film are representative of molecules in physiologic solution.     

Understanding the mechanism of ice binding by type III antifreeze proteins

Type III antifreeze proteins (AFPs) are present in the body fluids of some polar fishes where they inhibit ice growth at subzero temperatures. Previous studies of the structure of type III AFP by NMR and X-ray identified a remarkably flat surface on the protein containing amino acids that were demonstrated to be important for interaction with ice by mutational studies. It was proposed that this protein surface binds onto the (1 0 [\bar 1] 0) plane of ice with the key amino acids interacting directly with the water molecules in the ice crystal. Here, we show that the mechanism of type III AFP interaction with ice crystals is more complex than that proposed previously. We report a high-resolution X-ray structure of type III AFP refined at 1.15 A resolution with individual anisotropic temperature factors. We report the results of ice-etching experiments that show a broad surface coverage, suggesting that type III AFP binds to a set of planes that are parallel with or inclined at a small angle to the crystallographic c-axis of the ice crystal. Our modelling studies, performed with the refined structure, confirm that type III AFP can make energetically favourable interactions with several ice surfaces.     

Change of supercooling capability in solutions containing different kinds of ice nucleators by flavonol glycosides from deep supercooling xylem parenchyma cells in trees

Deep supercooling xylem parenchyma cells (XPCs) in Katsura tree contain flavonol glycosides with high supercooling-facilitating capability in solutions containing the ice nucleation bacterium (INB) Erwinia ananas, which is thought to have an important role in deep supercooling of XPCs. The present study, in order to further clarify the roles of these flavonol glycosides in deep supercooling of XPCs, the effects of these supercooling-facilitating (anti-ice nucleating) flavonol glycosides, kaempferol 3-O-β-d-glucopyranoside (K3Glc), kaempferol 7-O-β-d-glucopyranoside (K7Glc) and quercetin 3-O-β-d-glucopyranoside (Q3Glc), in buffered Milli-Q water (BMQW) containing different kinds of ice nucleators, including INB Xanthomonas campestris, silver iodide and phloroglucinol, were examined by a droplet freezing assay. The results showed that all of the flavonol glycosides promoted supercooling in all solutions containing different kinds of ice nucleators, although the magnitudes of supercooling capability of each flavonol glycoside changed in solutions containing different kinds of ice nucleators. On the other hand, these flavonol glycosides exhibited complicated nucleating reactions in BMQW, which did not contain identified ice nucleators but contained only unidentified airborne impurities. Q3Glc exhibited both supercooling-facilitating and ice nucleating capabilities depending on the concentrations in such water. Both K3Glc and K7Glc exhibited only ice nucleation capability in such water. It was also shown by an emulsion freezing assay in BMQW that K3Glc and Q3Glc had no effect on homogeneous ice nucleation temperature, whereas K7Glc increased ice nucleation temperature. The results indicated that each flavonol glycoside affected ice nucleation by very complicated and varied reactions. More studies are necessary to determine the exact roles of these flavonol glycosides in deep supercooling of XPCs in which unidentified heterogeneous ice nucleators may exist.     

Physical properties of oil-particle aggregate (OPA)-containing sediments

ABSTRACT

Sediments composed of oil-particle aggregates (OPAs) have unique physical characteristics. These in situ deposited sediments develop at locations where a continual or nearly continual discharge of non-aqueous phase liquids (NAPLs) have occurred, or are occurring through time. The NAPL discharged into the surface water body interacts with suspended particles in the water column. The particles adhere to the suspended NAPL, which generally is in the form of a bead, and produce a discrete aggregate. As the aggregate grows in response to additional particle adherence, the density of the unit increases and deposition occurs. The resulting sediment consists of a collection of discrete OPAs that form a network with small pores, where oil is tightly bound and/or contained. Porosity, water content, and dry bulk density measurements indicate the sediment formed by OPA deposition is physically unique. Although the sediment consists of a very open pore structure, the pore openings are relatively small, typically being less than 5 microns in diameter. These small pores inhibit fluid movement. Results of physical property testing suggest the OPA structure is retained upon deposition. Although the sediment contains NAPL, this original OPA structure inhibits the oil beads from coalescing, which would enable NAPL flow.