The antifreeze potential of the spruce budworm thermal hysteresis protein

Abstract

Antifreeze proteins (AFP) inhibit ice growth by surface adsorption that results in a depression of the freezing point below the melting point. The maximum level of this thermal hysteresis shown by the four structurally unrelated fish AFP is approximately 1.5 degrees C. In contrast, hemolymph and crude extracts from insects can have 5 degrees to 10 degrees C of thermal hysteresis. Based on the isolation, cloning, and expression of a thermal hysteresis protein (THP) from spruce budworm (Choristoneura fumiferana), the vastly greater activity is attributable to a 9 kDa protein. This novel, threonine- and cysteine-rich THP has striking effects on ice crystal morphology, both before and during freezing. It is also 10 to 30 times more active than any known fish AFP, offering the prospect of superior antifreeze properties in cryoprotective applications.

Interactions of β-Helical Antifreeze Protein Mutants with Ice

“…1 The hyperactive AFPs have TH activity 2 orders of magnitudes higher, reaching similar TH values at tens of micromolar concentrations and up to 6°C at millimolar concentrations. [2][3][4] AFPs are markedly different in sequence and structure, even within each TH activity group. For example, type I AFP is a repetitive, small, alanine-rich, amphipathic R-helix containing four regularly spaced threonine residues; the lectin-like type II AFP and the type III AFPs are nonrepetitive globular proteins; the antifreeze glycoproteins are composed of repeats of AlaAla-Thr, with disaccharide moieties decorating the Thr side chains.…”

“…5 All these proteins belong to the moderately active AFPs and have similar specific and maximal TH activities. 1 Only a few hyperactive antifreeze proteins are known, including two nonhomologous -helical proteins from insects, 2,4,6 an elongated, Ala-rich, R-helical fish AFP, 3,7 a Gly-rich AFP from snow flea, 8 and a calcium-dependent bacterial AFP predicted to have a third class of -helical structure. 9 The commonly held theory for how AFPs depress freezing is the adsorption-inhibition mechanism.…”

Mimicking biopolymers on a molecular scale: nano(bio)technology based on engineered proteins

“…Additional AFP isoforms from insects with interesting properties have been discovered (Duman 2001). Insect AFPs display a higher thermal hysteresis compared with fish AFPs and can be up to 20 times more effective, as shown for the spruce budworm Choristoneura fumiferana (Tyshenko et al 1997). Structural analysis of this protein suggested an unusual structure of a left-handed b-helix with 15 amino acid residues per turn (Li et al 2005), while that from the beetle Tenebrio molitor displays also a regular b-helix with 12 amino acid loops and tandem 12-residue repeats (Liou et al 2000).…”

Animal ice-binding (antifreeze) proteins and glycolipids: an overview with emphasis on physiological function

“…Another insect AFP type is represented by larvae of the spruce budworm Choristoneura fumiferana (Tyshenko et al, 1997;Gauthier et al, 1998) and related Choristoneura species (Tyshenko et al, 2005), as well as the somewhat different inchworm Campanea perlata AFP (Lin et al, 2011). These insects have evolved AFPs with four disulfide bridges where 15-mer repeats produce a left-handed β-helix with T-X-T regions forming the ice-binding site on one side of the protein.…”