The role of side chain conformational flexibility in surface recognition by Tenebrio molitor antifreeze protein

Abstract

Two‐dimensional nuclear magnetic resonance spectroscopy was used to investigate the flexibility of the threonine side chains in the β‐helical Tenebrio molitor antifreeze protein (TmAFP) at low temperatures. From measurement of the 3 J αβ 1 H‐ 1 H scalar coupling constants, the χ 1 angles and preferred rotamer populations can be calculated. It was determined that the threonines on the ice‐binding face of the protein adopt a preferred rotameric conformation at near freezing temperatures, whereas the threonines not on the ice‐binding face sample many rotameric states. This suggests that TmAFP maintains a preformed ice‐binding conformation in solution, wherein the rigid array of threonines that form the AFP‐ice interface matches the ice crystal lattice. A key factor in binding to the ice surface and inhibition of ice crystal growth appears to be the close surface‐to‐surface complementarity between the AFP and crystalline ice, and the lack of an entropic penalty associated with freezing out motions in a flexible ligand.