Antifreeze glycoproteins: relationship between molecular weight, thermal hysteresis and the inhibition of leakage from liposomes during thermotropic phase transition.

Antifreeze glycoproteins (AFGP) were isolated and purified from the blood plasma of rock cod (Gadus ogac), using DEAE-Bio-gel ion exchange chromatography, followed by high performance liquid chromatography (HPLC). The purified proteins were analyzed using polyacrylamide gel electrophoresis (PAGE), and electrospray mass spectrometry. The results indicated that rock cod synthesize seven size classes of glycoproteins, ranging from 2.6 to 24 kDa, with each size class containing multiple isoforms. Antifreeze activity, as determined by thermal hysteresis, indicated that the AFGP could be separated into two groups, with the larger size classes (molecular mass>13 kDa) having approximately 3-4 times the activity of the smaller, proline containing, size classes (molecular mass<10 kDa). All of the AFGP size classes prevented leakage from dielaidoylphosphatidylcholine (DEPC) liposomes as they were cooled through their phase transition temperature, with the larger size classes being approximately 4 times as effective as the smaller ones. It is hypothesized that AFGP prevent liposomes from leaking as they pass through the phase transition temperature by binding to the phospholipid membrane.