Antifreeze proteins (AFPs) and antifreeze glycoproteins (AFGPs) enable the survival of various organisms in freezing or subfreezing habitats. Even very low concentrations of these proteins are sufficient to lower the freezing temperature by several degrees. To reach a similar freezing point depression using sodium chlori common car antifreeze (polyethylene glycol) would require much higher concentrations. AF(G)Ps already find applications in the food industry, for instance to prevent recrystallization in ice cream and have great potential in medical applications (cell & organ storage, operations at low temperature). The lowering of the freezing point of water by adding solutes is a quite general phenomenon. For practically all solutes (salts, sugars) this effect results from the entropy of mixing and does not depend on the nature of the solute (colligative effect). However, for AFPs and AFGPs the effect on the freezing point is much larger (up to 500 times) than the colligative effect and shows a strong hysteresis, meaning that only the freezing point and not the melting point is being depressed. Therefore the depression of the freezing point due to AF(G)Ps is believed to result from highly specific interactions be-tween these proteins and nucleating ice crystals. As such, the freezing point depression by AF(G)Ps constitutes a highly non-colligative effect. Protein-water interactions are of general interest owing to the importance of protein hydration for protein function; AFPs and AFGPs form an extraordinary example of this coupling that is sufficiently strong and specific that the protein controls macroscopic thermodynamic properties of water. Despite their importance in nature and their industrial relevance, the mechanisms by which AF(G)Ps depress the freezing point are still poorly understood. Although substantial information presently exists on the static protein structures and thermodynamic properties of these systems, molecular scale information on the dynamics of the conformations of the AF(G)Ps, their hydration shells and their binding to ice, is extremely scarce. Within the project AntWatFre (654936) the molecular mechanism by which AF(G)Ps lower the freezing temperature were studied with advanced (nonlinear) spectroscopic techniques like 2D polarization-resolved vibrational spectroscopy and surface sum-frequency generation
