Hyperactive antifreeze protein in flounder species

Abstract

Antifreeze proteins (AFPs) are functionally defined by their ability to bind to the surface of ice and inhibit its growth, which causes a lowering of the freezing temperature below the equilibrium freezing ⁄ melting point [1]. This thermal hysteresis (TH) effect of AFPs enables teleost fishes to live in ice-laden polar and subpolar oceans where temperatures can reach the freezing point of the seawater ()1.9°C), which is over 1 C°colder than the freezing temperature of their hypotonic body fluids ()0.7 to )0.9°C). Without the protection of AFPs, fish are effectively supercooled in these waters and will freeze on contact with ice [2,3]. Thus, the recent acquisition of AFPs during the late stages of the teleost radiation has enabled some species to survive in and ⁄ or expand into the relatively new niche created by sea-level glaciation 1-20 million years ago [4]. Type I AFPs are small, monomeric, alanine-rich single a-helices that have an 11-amino-acid periodicity. They are one of five distinct nonhomologous types of AFP found in fishes [5] and they are present in some righteye flounders, including the winter flounder (Pseudopleuronectes americanus), yellowtail flounder (Limanda ferruginea) and Alaskan plaice (Pleuronectes quadritaberulatus) [6][7][8]. The AFP isoforms in the winter flounder have been particularly well characterized. One of them, the 37-amino-acid HPLC-6, was the first AFP to have its structure solved [9,10] and to have the ice plane to which it binds defined by ice-etching [8]. The differences between isoforms mainly lie in their length (the number of 11-amino-acid repeats being either three or four) and in the amino acid replacements on the less well conserved hydrophilic side of

Isolation and characterization of type I antifreeze proteins from cunner,Tautogolabrus adspersus, order Perciformes

“…CD measurements indicate an α‐helix content of over 99% for cunner AFP based on its predicted size of 45 amino acids (Table 1), which is close to values obtained for other type I AFPs [14,19,26,50]. Similar to the type I AFP variant from winter flounder [26] cunner AFP exhibited a reversible thermal denaturation in the range 1–70 °C, with a loss of less than 10% of the original helicity when the AFP solution was cooled back to 1 °C.…”

“…The CD spectra of a cunner AFP sample at low temperature resembled that of other alanine‐rich α‐helical proteins [14,19,26,29,30] with well‐defined minima near 208 and 222 nm (Fig. 5A).…”

“…This periodicity is reflected in the sequences, which contain 11 amino acid repeats (ThrX 10 ) where X is frequently alanine [10]. Since their initial discovery these type I AFPs have been characterized and sequenced in members of two distinct teleost orders: Pleuronectiformes [family Pleuronectidae (seven species [11–15])] and Scorpaeniformes [family Cottidae (four species [10,16–18]) and family Cyclopteridae (two species [19])].…”

Design and Synthesis of Antifreeze Glycoproteins and Mimics

“…Fish, bacteria, insects and other organisms produce diverse classes of antifreeze proteins that appear to act through a common mechanism and prevent the damaging effects of ice crystal growth and allow survival in subzero temperatures. [1][2][3][4][5][6][7][8] These unique properties have attracted significant interest due to their potential applications in a wide variety of industries from medicine and veterinary science (cryopreservation), through agriculture (genetic engineering) to the frozen-food industry. [9][10] The identification of antifreeze proteins (AFPs) and antifreeze glycoproteins (AFGPs) in the blood serum of fish that survive in the sub-zero Arctic and Antarctic oceans was first investigated by Scholander [11][12][13] and De Vries.…”

Solution structure of hyperactive type I antifreeze protein

“…A coiledcoil structure was ruled out based on the paucity of hydrophobic amino acid residues (e.g., Leu, Ile, Val), thereby explaining the instability of the protein (T denat y9 uC). 8,10 Also, a coiled-coil structure would interfere with regularly positioned threonine residues appearing on a flat ice-binding site along the length of the protein. The complex was therefore modelled as an antiparallel dimer composed of two straight helices with relatively weak interactions between the alanine (Ala) side chains, i.e., as an Ala-zipper, resulting in a closer packing with L = 27.5 nm and d = 1.6 nm giving an axial ratio of L/d = 17.…”