Solute exclusion by polymer and protein-dominated water: correlation with results of nuclear magnetic resonance (NMR) and calorimetric studies and their significance for the understanding of the physical state of water in living cells.

According to the polarized multilayer (PM) theory of cell water proteins with their backbones fully extended and their NHCO groups directly exposed to bulk water, polarize water in multilayers. Experimental testing of the theory led to a new understanding of the uniqueness of gelatin, due to its permanently maintained fully extended conformation and its ability to polarize the bulk phase water in multilayers with reduced solubilities for solutes in a size dependent manner ("size rule"). Other models which behave like gelatin are urea-denatured proteins, synthetic polymers like polyethylene oxide (PEO), and polyvinylpyrrolidine (PVP), but not native proteins. NMR studies showed that the majority of water molecules dominated by these polymers does indeed suffer rotational (and translational) motional restriction as predicted by the PM theory. In conjunction with ultra-high frequency dielectric studies but particularly quasielastic neutron scattering of both model systems (e.g., PEO) and living cells (i.e., brine shrimp cysts and frog muscle), this finding offers confirmation of the PM theory of living cell water and model systems. Studies of the freezing point depression showed that the presence of as much as 50% of native proteins had no effect on the freezing point of water while inclusion of gelatin, PEO, etc., caused concentration-dependent lowering of the freezing temperature. These findings demonstrate the key role of polarized water in the phenomena of freezing point depression and the unusual ice forms seen in living cells.