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

¹ Institut de Biologie Structurale
² Inst. de Biologie Structurale CEA-CNRS

^* To whom correspondence should be addressed. E-mail: weik{at}ibs.fr.

Submitted on November 4, 2004
Revised on January 3, 2005
Accepted on 31 January 2005

	Abstract

Confined water is of considerable current interest owing to its biophysical importance and relevance to cryo-preservation. 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 (D2O) was obtained in the inter-membrane 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 inter-membrane water space decreased sharply with an associated strengthening of ice diffraction, indicating water beyond the first membrane hydration layer flowed out of the inter-membrane space to form crystalline ice. It was concluded that the confined water undergoes a glass transition at or below 200 K to adopt an ultra-viscous 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 inter-membrane space at temperatures below 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 nanosecond membrane dynamics are not sensitive to the state of the water beyond the first hydration shell at cryo-temperatures.

Key Words: Dynamical transition, Glass transition, Neutron scattering, Protein dynamics, Ultra-viscous water, cryo-biology

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