Evidence for liquid water during the high-density to low-density amorphous ice transition

Abstract

Polymorphism of water has been extensively studied, but controversy still exists over the phase transition between high-density amorphous (HDA) and low-density amorphous (LDA) ice. We report the phase behavior of HDA ice inside high-pressure cryocooled protein crystals. Using X-ray diffraction, we demonstrate that the intermediate states in the temperature range from 80 to 170 K can be reconstructed as a linear combination of HDA and LDA ice, suggesting a first-order transition. We found evidence for a liquid state of water during the ice transition based on the protein crystallographic data. These observations open the possibility that the HDA ice induced by high-pressure cryocooling is a genuine glassy form of high-density liquid.liquid-liquid hypothesis ͉ supercooled water ͉ water phases ͉ high-density liquid S upercooled water shows anomalous thermodynamic behavior (1-3). Theories that account for these anomalous properties include the stability limit (4), the singularity-free (5, 6), and the liquid-liquid (LL) critical point (7) hypotheses. The latter 2 hypotheses propose the existence of 2 distinct forms of supercooled water: high-density liquid (HDL) and low-density liquid (LDL) water (8). In the singularity-free hypothesis, HDL transforms continuously to LDL. In the LL critical point theory, HDL undergoes a first-order phase transition to LDL (8). However, experimental study of the HDL-LDL phase transition is challenging as supercooled water spontaneously converts to crystalline forms below the homogeneous nucleation temperature (Ϸ235 K at 0.1 MPa). The transition between 2 glassy forms of water, high-density amorphous (HDA) and low-density amorphous (LDA) ice, has been extensively studied (9-18), as an analogue of the HDL-LDL transition. Controversy still exists as to whether the HDA-LDA ice transition is truly a first-order phase transition (10-13) or if it occurs because of a relaxation process of an unstable amorphous structure (15-18). More importantly, the connection between the HDA-LDA ice transition and the HDL-LDL phase transition, implied by thermodynamic and structural studies on water (19-22), remains challenging to prove experimentally (8).We used X-ray diffraction to study the transition of HDA to LDA ice in protein crystals. HDA ice was induced inside protein crystals by a high-pressure cryocooling method (23) originally developed for macromolecular crystallography (23-26). The Bragg diffraction from protein crystals mainly provides information on protein structure. The simultaneously recorded water diffuse diffraction (WDD) profile reports on the phase of the water in the protein crystal, which accounts for typically 40-60% of its volume. Fig. 1 shows diffraction images and the WDD profiles from a high-pressure cryocooled crystal of the globular protein thaumatin. As the crystal temperature is increased from 80 to 170 K, the primary WDD peak, corresponding to the mean distance between neighboring water molecule oxygen atoms, shifts to lower momentum transfer (Q) region [Q ϭ 4 sin( )/ , wh...

Two-Dimensional Infrared Spectroscopy of Isotope-Diluted Low Density Amorphous Ice

“…The structure of amorphous ice is a crucial aspect in a comprehensive understanding of the thermodynamic properties of water [7]; in particular in light of the liquid-to-liquid phase transition hypothesis put forward by Stanley and coworkers to account for the anomalous thermodynamic properties of water [8,9]. That is, two distinct liquid phases in the deeply supercooled regime are predicted (low-density liquid LDL and high-density liquid HDL), which are believed to be associated with the corresponding amorphous ice phases, LDA and HDA, respectively [10][11][12][13].…”

Protein dynamical transition at 110 K

“…The LL critical theory was originally proposed to explain the anomalous thermodynamic properties of the supercooled water (46)(47)(48). The theory predicts a first-order phase transition between HDA and LDA ice (35,46). The theory also requires the liquid state counterparts of HDA and LDA ice (46)(47)(48).…”

“…The theory also requires the liquid state counterparts of HDA and LDA ice (46)(47)(48). Based on the LL critical theory, the observed protein dynamical transition at cryogenic temperatures might be related to the two proposed liquid forms of water (highdensity liquid and low-density liquid) existing well below the homogeneous nucleation temperature of water (35,(46)(47)(48)(49). It remains to be seen how the cryogenic protein dynamical transition is related to the conventional protein dynamical transition at around 200 K, which is thought to be a precondition for a biologically active state of proteins.…”

“…It is important to note that the cryogenic protein dynamical transition in our study is observed along with the phase transition from HDA to LDA states of water. It has been proposed that the water phase transition from HDA to LDA may involve a high mobility, liquid form of water: The HDA ice first undergoes a glass-liquid transition, then continuously converts to LDA ice (35).…”

“…In this work, we studied a protein dynamical transition inside protein crystals using a high-pressure cryocooling method (34) and temperature-dependent X-ray diffraction (8,35,36). Protein crystals contain large amounts of water in solvent channels between the proteins in the crystal lattice; water fractions of greater than 50% are very common.…”