Antifreeze proteins (AFPs) and antifreeze glycoproteins (AFGPs) enable the survival of organisms living in subfreezing habitats and serve as preservatives. Although their function is known, the underlying molecular mechanism was not understood. Mutagenesis experiments questioned the previous assumption of hydrogen bonding as the dominant mechanism. We use terahertz spectroscopy to show that antifreeze activity is directly correlated with long-range collective hydration dynamics. Our results provide evidence for a new model of how AFGPs prevent water from freezing. We suggest that antifreeze activity may be induced because the AFGP perturbs the aqueous solvent over long distances. Retarded water dynamics in the large hydration shell does not favor freezing. The complexation of the carbohydrate cis-hydroxyl groups by borate suppresses the long-range hydration shell detected by terahertz absorption. The hydration dynamics shift toward bulk water behavior strongly reduces the AFGP antifreeze activity, further supporting our model.
"This mechanism was proposed previously as underlying the interactions in carbohydrate or glycopeptide solutions, which displayed IRI without influencing the dynamics of ice shaping . Recent studies have showed that AFGPs affect water molecules in the far hydration shell and, therefore, support this possible mechanism . "
[Show abstract][Hide abstract] ABSTRACT: The control over ice crystal growth, melting, and shaping is important in a variety of fields, including cell and food preservation and ice templating for the production of composite materials. Control over ice growth remains a challenge in industry, and the demand for new cryoprotectants is high. Naturally occurring cryoprotectants, such as antifreeze proteins (AFPs), present one solution for modulating ice crystal growth; however, the production of AFPs is expensive and inefficient. These obstacles can be overcome by identifying synthetic substitutes with similar AFP properties. Zirconium acetate (ZRA) was recently found to induce the formation of hexagonal cavities in materials prepared by ice templating. Here, we continue this line of study and examine the effects of ZRA and a related compound, zirconium acetate hydroxide (ZRAH), on ice growth, shaping, and recrystallization. We found that the growth rate of ice crystals was significantly reduced in the presence of ZRA and ZRAH, and that solutions containing these compounds display a small degree of thermal hysteresis, depending on the solution pH. The compounds were found to inhibit recrystallization in a manner similar to that observed in the presence of AFPs. The favorable properties of ZRA and ZRAH suggest tremendous potential utility in industrial applications.
PLoS ONE 03/2013; 8(3):e59540. DOI:10.1371/journal.pone.0059540 · 3.23 Impact Factor
"The elongated TxTxTxT-motifs seen in these longhorn beetle AFPs is in accordance with this concept. Recently, Ebbinghaus et al. (2010, 2012) proposed that the AFPs might alternatively act over distance by perturbing the aqueous solvent. The Ramashandran plots generated from the molecular models (Fig. 5G and Supplemental material, S3) suggest that our simulated structures have some less common phi and psi combinations. "
[Show abstract][Hide abstract] ABSTRACT: This study reports on structural characteristics of hyperactive antifreeze proteins (AFPs) from two species of longhorn beetles. In Rhagium mordax, eight unique mRNAs coding for five different mature AFPs were identified from cold-hardy individuals. These AFPs are apparently homologues to a previously characterized AFP from the closely related species Rhagium inquisitor, and consist of six identifiable repeats of a putative ice binding motif TxTxTxT spaced irregularly apart by segments varying in length from 13 to 20 residues. Circular dichroism spectra show that the AFPs from both species have a high content of β-sheet and low levels of α-helix and random coil. Theoretical predictions of residue-specific secondary structure locate these β-sheets within the putative ice-binding motifs and the central parts of the segments separating them, consistent with an overall β-helical structure with the ice-binding motifs stacked in a β-sheet on one side of the coil. Molecular dynamics models based on these findings show that these AFPs would be energetically stable in a β-helical conformation.
"or whether the protein inhibits growth of ice locally at the protein-water interface or over a larger number of water layers near the protein. Recent THz studies  indicate that, at least for AFP in winter flounder, the effect appears to be delocalized. "
[Show abstract][Hide abstract] ABSTRACT: We examine dynamics of water molecules and hydrogen bonds at the water-protein interface of the wild-type antifreeze protein from spruce budworm
and a mutant that is not antifreeze active by all-atom molecular dynamics simulations. Water dynamics in the hydration layer around the protein is analyzed by calculation of velocity autocorrelation functions and their power spectra, and hydrogen bond time correlation functions are calculated for hydrogen bonds between water molecules and the protein. Both water and hydrogen bond dynamics from subpicosecond to hundred picosecond time scales are sensitive to location on the protein surface and appear correlated with protein function. In particular, hydrogen bond lifetimes are longest for water molecules hydrogen bonded to the ice-binding plane of the wild type, whereas hydrogen bond lifetimes between water and protein atoms on all three planes are similar for the mutant.
Journal of Atomic Molecular and Optical Physics 03/2012; 6(1687-9228). DOI:10.1155/2012/125071
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