Structural Basis for the Superior Activity of the Large Isoform of Snow Flea Antifreeze Protein

Mok, Yee-Foong; Lin, Feng-Hsu; Graham, Laurie A.; Celik, Yeliz; Braslavsky, Ido; Davies, Peter L.

doi:10.1021/bi901929n

Cited by 58 publications

(59 citation statements)

References 45 publications

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“…MpAFP_RIV tagged with GFP for fluorescence-based ice plane affinity (FIPA) analysis (27) turned single ice-crystal hemispheres uniformly fluorescent (Fig. 5 B and C), consistent with the protein's affinity toward the basal and primary prism planes of ice, as well as surfaces between these planes, a property also seen with the hyperactive AFP from snow fleas (28). For a comparison, FIPA analysis was also performed on type I AFP.…”

Section: Resultsmentioning

confidence: 67%

Anchored clathrate waters bind antifreeze proteins to ice

Garnham

Campbell

Davies

2011

Proc. Natl. Acad. Sci. U.S.A.

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344

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The mechanism by which antifreeze proteins (AFPs) irreversibly bind to ice has not yet been resolved. The ice-binding site of an AFP is relatively hydrophobic, but also contains many potential hydrogen bond donors/acceptors. The extent to which hydrogen bonding and the hydrophobic effect contribute to ice binding has been debated for over 30 years. Here we have elucidated the ice-binding mechanism through solving the first crystal structure of an Antarctic bacterial AFP. This 34-kDa domain, the largest AFP structure determined to date, folds as a Ca 2þ -bound parallel beta-helix with an extensive array of ice-like surface waters that are anchored via hydrogen bonds directly to the polypeptide backbone and adjacent side chains. These bound waters make an excellent three-dimensional match to both the primary prism and basal planes of ice and in effect provide an extensive X-ray crystallographic picture of the AFP∶ice interaction. This unobstructed view, free from crystal-packing artefacts, shows the contributions of both the hydrophobic effect and hydrogen bonding during AFP adsorption to ice. We term this mode of binding the "anchored clathrate" mechanism of AFP action.Ca2+ binding protein | repeats-in-toxin protein | thermal hysteresis | Antarctic bacterium | organized biohydration A ntifreeze proteins (AFPs) adsorb to the surface of ice crystals and prevent their growth (1). This adsorption lowers the freezing temperature of a solution below its melting point, enabling the survival of many organisms that inhabit ice-laden environments. Despite their common function, AFPs display remarkable diversity in their tertiary structures (2-7). This diversity results partly from their independent evolutionary origins (8, 9) and partly from the surface heterogeneity of their natural ligand, ice (10). Hexagonal ice presents many different planes (expressed as Miller indices) of water molecules to which an AFP can develop affinity. Although specificity toward different ice planes is a key determinant of antifreeze activity (11), the mechanism by which an AFP binds to ice remains undefined.Hydrogen bonds were originally proposed to be the main binding force between an AFP and ice (12). Yet this hypothesis was unable to explain how an AFP would preferentially bind ice when solvated by 55 M water. Subsequent studies proposed that the hydrophobic effect was the main ice-binding force, where constrained, clathrate-like water on the ice-binding site (IBS) is released into the solvent upon ice binding, resulting in a gain of entropy (13,14). However, several molecular dynamics (MD) simulations have indicated that the relatively hydrophobic IBS of an AFP is capable of ordering water molecules into an ice-like lattice (15-21) and, instead of shedding bound water molecules upon ice binding, the ordered waters might facilitate the AFP's interaction with ice by matching certain ice planes (15). Although intriguing, these simulations fall short of describing at the molecular level how an AFP might order water molecules into an ice-like ...

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Section: Resultsmentioning

confidence: 67%

Anchored clathrate waters bind antifreeze proteins to ice

Garnham

Campbell

Davies

2011

Proc. Natl. Acad. Sci. U.S.A.

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344

420

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“…Ice growth occurs from six prism surfaces and not out of the basal plane. However, when hyperactive AFPs are tested, most of them, including TmAFP, sfAFP (47), and MpAFP (7), bind to multiple planes as shown by adsorption to all surfaces of a single crystal hemisphere. GFP-tagged RiAFP also uniformly binds the entire surface of a single crystal hemisphere during fluorescence-based ice plane affinity analysis (Fig.…”

Section: Resultsmentioning

confidence: 99%

Crystal Structure of an Insect Antifreeze Protein and Its Implications for Ice Binding

Hakim

Nguyen

Basu

et al. 2013

Journal of Biological Chemistry

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103

121

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Background: Antifreeze proteins bind to ice crystals and inhibit their growth. Results: The crystal structure of a potent beetle antifreeze protein was determined by direct methods. Conclusion: Ordered crystallographic waters on the protein surface match several planes of hexagonal ice. Significance: The structure is the largest determined ab initio without heavy atoms, and its ordered waters suggest a molecular basis for ice binding.

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“…The primary sequence of the large isoform of sfAFP is different from the primary structure of the small sfAFP isoform. Mok et al (2010) reported that the large isoform consists of 203 residues with a high occurrence of glycine. As in the small isoform, a tripeptide repeat pattern is observed in which glycine occupies the first position and often also the second.…”

Section: Controlling the Freezing Process With Antifreeze Proteinsmentioning

confidence: 99%