Ice recrystallization is strongly inhibited when antifreeze proteins bind to multiple ice planes

Abstract

Ice recrystallization is a phenomenon observed as the increase in ice crystal size within an already frozen material. Antifreeze proteins (AFPs), a class of proteins capable of arresting ice crystal growth, are known to inhibit this phenomenon even at sub milli-molar concentrations. A tremendous range in the possible applications of AFPs is hence expected in both medical and industrial fields, while a key determinant of the ice recrystallization inhibition (IRI) is hardly understood. Here, IRI efficiency and ice plane affinity were examined for the wild-type AFPI–III, a defective AFPIII isoform, and a fungal AFP isoform. To simplify the IRI analysis using the formal representation of Ostwald-ripening (r3 = r03 + kt), we monitored specific ice grains exhibiting only uniform growth, for which maximum Feret diameter was measured. The cube of an ice grain’s radius (r3) increased proportionately with time (t), and its slope gave the recrystallization rate (k). There was a significant difference in the IRI efficiency between the samples, and the fungal AFP possessing the activity with the smallest amount (0.27 μM) exhibited an affinity to multiple ice planes. These results suggest that the IRI efficiency is maximized when AFPs bind to a whole set of ice planes.

Ice Crystal Growth in Sucrose Solutions Containing Kappa‐ and Iota‐Carrageenans

“…For instance, in the literature, the TH activity of AFGP purchased from the same company, Nichirei Co. Ltd., is described to have been measured using a different technique, i.e., the capillary technique and in 0.1 M ammonium bicarbonate (pH 7.9). The results are lower than the results obtained in this study [20].…”

“…Therefore, preliminary experiments have been carried out to reveal whether TH activities of AFP I and AFGP can be detected with this system. AFP I is a low-molecular-weight protein (< 30 kDa) obtained from the barfin plaice (bp), also called the native bpAFP [19] with a molecular weight of 3.5 kDa [20]. AFGP was provided by Nichirei Co. Ltd. as a purified sample of AFGP from Eleginus gracilis [21] with an assumed molecular weight of 15 kDa [20].…”

“…AFP I is a low-molecular-weight protein (< 30 kDa) obtained from the barfin plaice (bp), also called the native bpAFP [19] with a molecular weight of 3.5 kDa [20]. AFGP was provided by Nichirei Co. Ltd. as a purified sample of AFGP from Eleginus gracilis [21] with an assumed molecular weight of 15 kDa [20]. The TH activity of AFP I was stated in 0.1 M ammonium bicarbonate (pH 7.9) at a concentration of 10 mg mL -1 .…”

Mimicking the Ice Recrystallization Activity of Biological Antifreezes. When is a New Polymer “Active”?

“…This small set alone shows the importance of screening with a negative control, such as PEG, as essentially any material can claim to have IRI activity but the magnitude of this is the crucial parameter. A detailed discussion of AFGP/AFP's high activity is beyond the scope of this feature article, but in both cases it appears to be due to specific recognition of the primary prism face of ice, which is the fastest growing face under standard conditions . There is still uncertainty as to how exactly AFGPs bind ice though, either through their glycans, or the peptide backbone .…”

Fish-Derived Antifreeze Proteins and Antifreeze Glycoprotein Exhibit a Different Ice-Binding Property with Increasing Concentration

“…For the FIPA experiment, several single ice crystals with a cylindrical shape (Ø = 2-3 cm) were initially prepared within the plastic pipes (l = 4 cm) according to a previously described procedure [27][28][29]. After determination of their c-axis on a polarizer, each cylindrical ice was half-cut and mounted on a hollow copper tube (Ø = 15 mm), in which −0.8 • C coolant was circulated by a refrigerant pump (Hitachi AMS-007; Hitachi, Tokyo, Japan).…”