Effect of Glycosylation on Antifreeze Protein Activity

Abstract

Antifreeze proteins (AFPs) depress the freezing point of a solution which therefore produces a difference between the melting point and freezing point, so called thermal hysteresis (TH). AFPs have been known to be essential for organisms to survive subzero environments. So far, many AFPs have been characterized from fish, insects, plants, bacteria and microalgae. We isolated AFP from the culture supernatant of an Arctic yeast Leucosporidium sp. AY30 using ice affinity method. Native AY30 AFP was observed to be glycosylated by periodic acid Schiff staining. PNGase treatment indicated that AY30 AFP was N-glycosylated protein. We cloned and expressed recombinant AFP (rAY30 AFP) in BL21 (DE3). CD spectra data indicated that tertiary structures of native and rAFP were similar. The rAY30 AFP has shown to have lower TH activity than native AY30 AFP. The shape of ice crystal of rAFP was star shape meaning weak ice-binding activity. These results shows that glycosylation enhance the TH activity and ice-binding activity. Futher study, we plans to the X-ray crystallography for more accurate 3-D structure