Purification of antifreeze proteins by adsorption to ice

Kuiper, Michael J.; Lankin, Christopher; Gauthier, Sherry Y.; Walker, Virginia K.; Davies, Peter L.

doi:10.1016/s0006-291x(02)02900-5

Cited by 108 publications

(109 citation statements)

References 9 publications

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“…Midge AFP was purified from the soluble fraction of adult midge homogenates by sequential ice affinity purification (20). Antifreeze activity (TH) and ice crystal morphology were monitored using a nanoliter osmometer (21).…”

Section: Methodsmentioning

confidence: 99%

Flies expand the repertoire of protein structures that bind ice

Basu

Graham

Campbell

et al. 2015

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

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An antifreeze protein (AFP) with no known homologs has been identified in Lake Ontario midges (Chironomidae). The midge AFP is expressed as a family of isoforms at low levels in adults, which emerge from fresh water in spring before the threat of freezing temperatures has passed. The 9.1-kDa major isoform derived from a preproprotein precursor is glycosylated and has a 10-residue tandem repeating sequence xxCxGxYCxG, with regularly spaced cysteines, glycines, and tyrosines comprising one-half its 79 residues. Modeling and molecular dynamics predict a tightly wound left-handed solenoid fold in which the cysteines form a disulfide core to brace each of the eight 10-residue coils. The solenoid is reinforced by intrachain hydrogen bonds, side-chain salt bridges, and a row of seven stacked tyrosines on the hydrophobic side that forms the putative ice-binding site. A disulfide core is also a feature of the similar-sized beetle AFP that is a β-helix with seven 12-residue coils and a comparable circular dichroism spectrum. The midge and beetle AFPs are not homologous and their ice-binding sites are radically different, with the latter comprising two parallel arrays of outward-pointing threonines. However, their structural similarities is an amazing example of convergent evolution in different orders of insects to cope with change to a colder climate and provide confirmation about the physical features needed for a protein to bind ice.

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Section: Methodsmentioning

confidence: 99%

Flies expand the repertoire of protein structures that bind ice

Basu

Graham

Campbell

et al. 2015

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

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“…Thus, successive rounds of freezing permit the purification of THFs to homogeneity (21). The extractions and washes were aliquotted into 50-ml centrifuge tubes and placed in an alcohol bath set to Ϫ3.6°C.…”

Section: Methodsmentioning

confidence: 99%

“…THFs present in each extract (R1, soluble fraction; R2, first membraneassociated fraction; R3, second membrane-associated fraction) were separated from other solutes by ice affinity (21). The isolated THFs in the R1, R2, and R3 fractions weighed Ϸ125 g, Ϸ100 g, and Ϸ60 g, respectively, and exhibited Ϸ3.7 Ϯ 0.3°C of TH at 5 mg/ml, a value comparable to that of the most active insect antifreeze proteins (12).…”

Section: Thf Isolated From U Ceramboides Is Highly Active and Contaimentioning

confidence: 99%

A nonprotein thermal hysteresis-producing xylomannan antifreeze in the freeze-tolerant Alaskan beetle Upis ceramboides

Walters¹,

Serianni

Sformo

et al. 2009

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

117

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Thermal hysteresis (TH), a difference between the melting and freezing points of a solution that is indicative of the presence of large-molecular-mass antifreezes (e.g., antifreeze proteins), has been described in animals, plants, bacteria, and fungi. Although all previously described TH-producing biomolecules are proteins, most thermal hysteresis factors (THFs) have not yet been structurally characterized, and none have been characterized from a freeze-tolerant animal. We isolated a highly active THF from the freeze-tolerant beetle, Upis ceramboides, by means of ice affinity. Amino acid chromatographic analysis, polyacrylamide gel electrophoresis, UV-Vis spectrophotometry, and NMR spectroscopy indicated that the THF contained little or no protein, yet it produced 3.7 ؎ 0.3°C of TH at 5 mg/ml, comparable to that of the most active insect antifreeze proteins. Compositional and structural analyses indicated that this antifreeze contains a ␤-mannopyranosyl-(134) ␤-xylopyranose backbone and a fatty acid component, although the lipid may not be covalently linked to the saccharide. Consistent with the proposed structure, treatment with endo-␤-(134)xylanase ablated TH activity. This xylomannan is the first TH-producing antifreeze isolated from a freeze-tolerant animal and the first in a new class of highly active THFs that contain little or no protein.antifreeze protein ͉ insect cold tolerance ͉ glycolipid ͉ membrane associated antifreeze

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“…Contre toute attente, le degré de HT était supérieur à 2 o C [7], soit plus de deux fois l'activité maximale de la PAG de « type I ». De plus, les cristaux de glace présents dans le plasma étaient d'une forme fuselée complexe alors que les cristaux très particuliers dus à la présence de PAG de «type I » ont une forme de bipyramide hexagonale (Figure 1 [8].…”

Section: Nouvelleunclassified