Interactions of the D- and L-Forms of Winter Flounder Antifreeze Peptide with the {201} Planes of Ice

Madura, Jeffry D.; Wierzbicki, Andrzej; Harrington, John P.; Maughon, Richard H.; Raymond, James A.; Sikes, C. Steven

doi:10.1021/ja00080a066

Cited by 64 publications

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“…It is reasonable to assume that the Thr adopt a similar rotameric state once they are bound to ice, since a random distribution of all possible Thr conformations would not produce the desired lattice matching distance of 16.7 Å. Regardless of the rotameric state of Thr, most computational studies place these residues above the water molecules that form the ridges of the {202 h1} surface (the {202 h1} surface has a regularly spaced ridge and valley topology) (8,10,12,35). This seems reasonable since short Thr side chain is probably not able to interact directly with those water molecules that lie in the valleys.…”

Section: Discussionmentioning

confidence: 99%

“…Although all of these models share the Thr hydrogen-bonding match, there is no consensus about the role of Asx. In two reports these residues were not featured in the ice-binding model (8,9); in others they were hydrogen bonded to different ranks of oxygen atoms in the lattice (10, 11) or they occupied a cage position within the lattice without hydrogen bonding (12).Throughout all of these deliberations there has been concern that the number and strength of the available hydrogen bonds (with or without a contribution from the Asx residues) may be inadequate to explain tight binding of the antifreeze to ice. At least two imaginative ideas have been put forward to reinforce the hydrogen-bonding hypothesis by suggesting ways in which more hydrogen bonds can be brought to bear on the binding interaction.…”

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A Diminished Role for Hydrogen Bonds in Antifreeze Protein Binding to Ice

et al. 1997

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The most abundant isoform (HPLC-6) of type I antifreeze protein (AFP 1 ) in winter flounder is a 37-amino-acid-long, alanine-rich, R-helical peptide, containing four Thr spaced 11 amino acids apart. It is generally assumed that HPLC-6 binds ice through a hydrogen-bonding match between the Thr and neighboring Asx residues to oxygens atoms on the {202 h1} plane of the ice lattice. The result is a lowering of the nonequilibrium freezing point below the melting point (thermal hysteresis). HPLC-6, and two variants in which the central two Thr were replaced with either Ser or Val, were synthesized. The Ser variant was virtually inactive, while only a minor loss of activity was observed in the Val variant. CD, ultracentrifugation, and NMR studies indicated no significant structural changes or aggregation of the variants compared to HPLC-6. These results call into question the role of hydrogen bonds and suggest a much more significant role for entropic effects and van der Waals interactions in binding AFP to ice.Type I AFP 1 is the smallest and arguably the simplest of the four macromolecular antifreeze types characterized to date (1). It is in effect a single, long R-helix and therefore lacks tertiary structure (2). The most abundant isoform of this AFP (HPLC-6) from winter flounder (Pleuronectes americanus) is 37 amino acids long, contains three complete 11-amino-acid repeats of Thr-X 2 -Asx-X 7 , where X is generally alanine, and ends with the start of a fourth repeat. This helical periodicity that places the Thr and Asx residues on the same face of the helix, suggested a mechanism for adsorption of the AFP to ice in which these regularly spaced hydrophilic groups would hydrogen bond to oxygen atoms in the ice lattice (3). Adsorption leads to inhibition of ice crystal growth (4) because ice is forced to grow with a surface curvature between the bound AFP, which in turn results in a lowering of the nonequilibrium freezing point below the melting point (5, 6). The difference in these two temperatures is termed thermal hysteresis and is used as a measure of antifreeze activity.At very low concentrations, AFP bind to ice but do not stop its growth. Under these conditions bound AFP is frozen into the ice rather than excluded by the advancing ice front. The protein binding planes in these crystals have been made visable by sublimation (ice etching) and determined to be the {202 h1} pyramidal plane of hexagonal ice (I h ) for type I AFP (5). Moreover, because this antifreeze is a nonglobular, extended molecule it was possible to establish a direction 〈011 h2〉 of binding on the plane. An elegant proof of this resulted from the synthesis of an all D-type I AFP, which was shown by the ice etching method to bind to the same plane but in the mirror image direction (7). This information was used to suggest a hydrogen-bonding match between the i, i + 11 threonines spaced 16.5 Å apart along the helix and accessible ice lattice oxygens spaced 16.7 Å apart along the 〈011 h2〉 direction of the {202 h1} binding plane.On the ba...

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Section: Discussionmentioning

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A Diminished Role for Hydrogen Bonds in Antifreeze Protein Binding to Ice

et al. 1997

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“…Such a convenient coordination arrangement is not available for planar amide groups (Asn, Gln); consequently, modeling studies (Wen & Laursen, 1992b;Madura et al, 1994) indicate other preferred positions for these side chains. These differences in H bonding possibilities between the hydroxyl groups and amide groups might help explain the large reductions in activity shown by mutants N14S, T18N, and Q44T, where hydroxyl and amide groups have been exchanged.…”

Section: Comparison Of the Mutations Of Residues N14 Tis And Q44mentioning

confidence: 99%

Structure‐function relationship in the globular type III antifreeze protein: Identification of a cluster of surface residues required for binding to ice

et al. 1994

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Antifreeze proteins (AFPs) depress the freezing point of aqueous solutions by binding to and inhibiting the growth of ice. Whereas the ice-binding surface of some fish AFPs is suggested by their linear, repetitive, hydrogen bonding motifs, the 66-amino-acid-long Type 111 AFP has a compact, globular fold without any obvious periodicity. In the structure, 9 P-strands are paired to form 2 triple-stranded antiparallel sheets and 1 double-stranded antiparallel sheet, with the 2 triple sheets arranged as an orthogonal &sandwich (Sonnichsen FD, Sykes BD, Chao H, Davies PL, 1993, Science 2591154-1157). Based on its structure and an alignment of Type 111 AFP isoform sequences, a cluster of conserved, polar, surface-accessible amino acids (N14, T18, 444, and N46) was noted on and around the triple-stranded sheet near the C-terminus. At 3 of these sites, mutations that switched amide and hydroxyl groups caused a large decrease in antifreeze activity, but amide to carboxylic acid changes produced AFPs that were fully active at pH 3 and pH 6. This is consistent with the observation that Type 111 AFP is optimally active from pH 2 to pH 11. At a concentration of 1 mg/mL, Q44T, N14S, and T18N had 50'70, 25%, and 10% of the activity of wild-type antifreeze, respectively. The effects of the mutations were cumulative, such that the double mutant N14S/Q44T had 10% of the wild-type activity and the triple mutant N14S/T18N/Q44T had no activity. All mutants with reduced activity were shown to be correctly folded by NMR spectroscopy. Moreover, a complete characterization of the triple mutant by 2-dimensional NMR spectroscopy indicated that the individual and combined mutations did not significantly alter the structure of these proteins. These results suggest that the C-terminal @-sheet of Type 111 AFP is primarily responsible for antifreeze activity, and they identify N14, T18, and 444 as key residues for the AFP-ice interaction.Keywords: NMR; site-directed mutagenesis; thermal hysteresis Some cold water marine fishes produce proteins or glycoproteins that lower the freezing point of their blood without significantly increasing its osmolarity (DeVries, 1983;Davies & Hew, 1990). These antifreeze proteins or glycoproteins bind to and halt the growth of seed ice crystals that form in solution at or below Reprint requests to: Peter L. Davies, Department of Biochemistry, Queen's University, Kingston, Ontario K7L 3N6, Canada.Abbreviations: AFGP, antifreeze glycoprotein; AFP, antifreeze protein; FPLC, fast protein liquid chromatography; H bond, hydrogen bond; NOESY, nuclear Overhauser effect spectroscopy; QAE, quaternary aminoethyVAFP isoform that binds QAE-Sephadex; TOCSY, total correlation spectroscopy; lD, one-dimensional; 2D, two-dimensional; 3D, three-dimensional.

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“…Thus far, several research groups have used computer simulation for investigating the stable binding conformations of AFP at an ice crystal plane. [30][31][32][33][34][35][36][37] Recently, we also used MD simulation to analyze the molecular-scale growth kinetics at an ice-water interface to which AFP was bound. 38,39 These computer simulation studies have contributed to the understanding of the molecular-scale mechanism of ice growth inhibition by AFPs.…”

Section: Introductionmentioning

confidence: 99%

Antifreeze proteins: computer simulation studies on the mechanism of ice growth inhibition

Nada

Furukawa

2012

Polym J

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Antifreeze proteins (AFPs), which are present in the bodily fluids of organisms inhabiting cold environments, function as inhibitors of ice growth by binding to certain planes of ice crystals. However, the exact mechanism of ice growth inhibition is still poorly understood as it is exceedingly difficult to experimentally analyze the molecular-scale growth kinetics of ice crystals at the planes to which the proteins bind. This review paper focuses on computer simulation studies on the mechanism of ice growth inhibition by AFPs. Recent molecular dynamics simulations of a growing ice-water interface to which protein is bound have indicated that, when the protein is stably bound to the interface, the growth rate of ice surrounding the protein decreases drastically, owing to depression of the ice melting point through the Gibbs-Thomson effect. The observed decrease in growth rate is expected to correspond to ice growth inhibition in real-world systems. In addition to presenting published simulation studies on the mechanism of ice growth inhibition by AFPs, this review also outlines the direction of future simulation studies in this field.

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Interactions of the D- and L-Forms of Winter Flounder Antifreeze Peptide with the {201} Planes of Ice

Cited by 64 publications

References 0 publications

A Diminished Role for Hydrogen Bonds in Antifreeze Protein Binding to Ice

A Diminished Role for Hydrogen Bonds in Antifreeze Protein Binding to Ice

Structure‐function relationship in the globular type III antifreeze protein: Identification of a cluster of surface residues required for binding to ice

Antifreeze proteins: computer simulation studies on the mechanism of ice growth inhibition

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