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

Abstract

The concentration of a protein is highly related to its biochemical properties, and is a key determinant for its biotechnological applications. Antifreeze proteins (AFPs) and antifreeze glycoproteins (AFGPs) are structurally diverse macromolecules that are capable of binding to embryonic ice crystals below 0 °C, making them useful as protectants of ice-block formation. In this study, we examined the maximal solubility of native AFP I–III and AFGP with distilled water, and evaluated concentration dependence of their ice-binding property. Approximately 400 mg/mL (AFP I), 200 mg/mL (AFP II), 100 mg/mL (AFP III), and >1800 mg/mL (AFGP) of the maximal solubility were estimated, and among them AFGP’s solubility is much higher compared with that of ordinary proteins, such as serum albumin (~500 mg/mL). The samples also exhibited unexpectedly high thermal hysteresis values (2–3 °C) at 50–200 mg/mL. Furthermore, the analysis of fluorescence-based ice plane affinity showed that AFP II binds to multiple ice planes in a concentration-dependent manner, for which an oligomerization mechanism was hypothesized. The difference of concentration dependence between AFPs and AFGPs may provide a new clue to help us understand the ice-binding function of these proteins.

Characterization of microbial antifreeze protein with intermediate activity suggests that a bound-water network is essential for hyperactivity

“…Similar observations have been reported for hyperactive AFPs, including Tis AFP8 22 , Col AFP 20 Mp AFP_RIV 31 , sbwAFP ( C. fumiferana ) 37 , and Tm AFP ( Tenebrio molitor ) 37 . Some moderately active AFPs, when assayed at a relatively high concentration (0.1 mg/ml), also exhibit whole-hemisphere binding, e.g., BpAFP 14 and AFPII 38 ( Brachyopsis rostratus ), with a concentration-dependent multiple-plane binding. In the current study, FIPA analysis of Tis AFP7 was performed using a low protein concentration (0.007 mg/ml); the protein’s multiple-plane binding ability was higher than those of the typical moderately active AFPs.…”

Transcriptome analyses and weighted gene coexpression network analysis reveal key pathways and genes involved in the rapid cold resistance of the Chinese white wax scale insect

“…AFPs can prevent the growth of ice crystals and can decrease the freezing point of the solution. Because of their THA, AFPs are the most important factor in the antifreeze process of cells and organisms (Inagawa et al, 2020; Tsuda et al, 2020; Ye et al, 2020). We identified one AFP with a THA of 0.97°C in a previous study (Yu et al, 2016).…”

An Ice-Binding Protein from an Antarctic Ascomycete Is Fine-Tuned to Bind to Specific Water Molecules Located in the Ice Prism Planes

“…This observation was interpreted as indicating that this IBP binds only to the pyramidal plane at 0.01 mg/mL, whereas it covers the whole ice crystal, including the basal plane, when its concentration is 10 times higher [40]. A similar change was observed for type II IBP from longsnout poacher (LpIBP) [41]. We performed a concentration dependence experiment for AnpIBP between 0.01 and 0.15 mg/mL.…”

“…It has been shown that the FIPA pattern is sometimes changed with increases in the concentration of IBP [40,41]. For example, the pattern was observed in a limited area for a 0.01-mg/mL solution of fish type I IBP from barfin plaice, and it was overcast by the entire illumination when the protein concentration was increased to 0.1 mg/mL.…”