Role of dynamics in type I antifreeze protein structure and function

This thesis investigates antifreeze proteins (AFP), which are found in various cold-surviving organisms and are capable of interacting with ice, inhibiting ice-crystal growth as well as bring about non-colligative freezing point depression, a property named thermal hysteresis (TH). The AFP-ice interaction is still poorly understood at the molecular level. In this project, the first successful bacterial expression and purification of a model protein for AFP family, HPLC6 isoform of type I AFP from the winter flounder was performed in minimal media. HPLC6 is a 3.2 kDa sized, alanine-rich, single alpha-helix. The cap structures formed by amidation of the last residue, Arg37, and the N-terminal aspartate stabilize the helix dipole of HPLC6, which is thought to enhance AFP activity. In this thesis, alterations in the C-terminal residue resulted in an increase in flexibility of the last few residues, making ice binding less favourable and, therefore, resulting in a loss of TH activity. Further studies investigated the alanine methyls in the AFP-ice interaction using deuterated rHPLC6 which showed a TH loss with rHPLC6 and deuterated rHPLC6 when the activity was measured in deuterated water.