Abstract: A series of mutants of the insect antifreeze protein(CfAFP) were designed, their best ice-binding modes were generated by using molecular dynamic simulations, and the interaction energy with ice was computed by semi-empirical molecular orbital methods AM1 and PM3. The results show that the contribution to the CfAFP-ice interaction of the threonine residues depends on the position and number in CfAFP, and Thr residues in TXT-motif play key roles on the recognition and binding of CfAFP and ice. Replacement of thr by other kinds of hydrophilic or hydrophobic amino residues reduced the interaction with ice, resulting in a moderate loss of antifreeze activity. Additionally, based on the structure complement of CfAFP and ice, increasing the size of hydrophobic residues could gain more stable interaction with ice and enhance the antifreeze activity.

Key words: Insect antifreeze protein, Mutagenesis, Hydrophilicity, Hydrophobicity, Molecular dynamics