Presence of fucosylated triantennary, tetraantennary and pentaantennary glycans in transferrin synthesized by the human hepatocarcinoma cell line Hep G2

Abstract

Transferrin synthesized by a human hepatocellular carcinoma cell line Hep G2 (called Hep G2 transferrin) was purified from culture media by immunoaffinity chromatography on a rabbit anti-(human serotransferrin) IgG column. The eluted transferrin was then resolved into five peaks on a cation-exchange column using the fast protein liquid chromatography system. The major fraction, named Hep G2 transferrin fraction C, having a molecular mass of 82.5 kDa was found to be homogeneous in polyacrylamide gel electrophoresis and in concanavalin-A-affinity crossed immunoelectrophoresis. A comparative analysis of the molar carbohydrate composition of normal human serotransferrin and of Hep G2 transferrin fraction C shows an increase in the latter in the number of galactose and N-acetylglucosamine residues and in the presence of fucose, which is absent in normal transferrin. By a combination of methylation analysis and NMR spectroscopy, the primary structure of the oligosaccharide alditols released from Hep G2 transferrin fraction C by reductive alkaline cleavage has been established as triantennary, tetraantennary and pentaantennary N-acetyllactosaminic structures with fucose residues (.alpha.1-3)-linked to peripheral N-acetylglucosamine residues. These results indicate that the increase in the number of antennae in transferrin glycans synthesized by the hepatocarcinoma cell line is much more pronounced than in liver diseases such as alcoholic cirrhosis and that, in addition, the malignant transformation of human liver induces the presence of fucose.