SYNTHESIS AND ANALYSIS OF FLUORESCENT ANTIFREEZE GLYCOPROTEIN MIMIC PEPTIDES FOR DETERMINATION OF THE ICE BINDING MECHANISM

Abstract

Properties of antifreeze glycoproteins (AFGPs) are well known; however, it is unclear which molecular components of the proteins are directly responsible for binding to the ice surface. Previous molecular dynamics studies have implicated hydrophobic methyl groups in ice binding, while microscopy experiments have implicated polar hydroxyl groups from the galactosamine units. This thesis describes the synthesis of non-glycosylated AFGP mimic sequences which have been labeled with a water-sensitive naphthalimide fluorophore at several locations. In principle, the emission intensity of these systems should change if ice binding removes the fluorophore from contact with liquid water. Circular dichroism analysis suggests that the presence of the fluorescent label at the N-terminal amino acid induces polyproline II structure to the peptide. C-terminal labeling does not induce the same structuring. Results from fluorescence spectroscopy are consistent with the formation of microscopic ice crystals at or below 0[degrees]C, which serve to draw the peptides out of the sample beam and thereby decrease the fluorescence intensity. To our knowledge, this is the first example of bulk solution fluorescence being used to demonstrate ice binding through hydrophobic groups.