Analytical Sciences

Targeted Proteo-Glycomics Analysis of Sialyl Lewis X Antigen Expressing Glycoproteins Secreted by Human Hepatoma Cell Line

Koji HIGAI,*  Kumiko SHIBUKAWA,* Satoshi MUTO,** and Kojiro MATSUMOTO*

*Department of Clinical Chemistry, School of Pharmaceutical Sciences, Toho University, Miyama 2-2-1, Funabashi, Chiba 274-8510, Japan
**Department of Disease Analysis, School of Pharmaceutical Sciences, Toho University, Miyama 2-2-1, Funabashi, Chiba 274-8510, Japan

Sialyl Lewis X (SLEX) antigen, Neu5Acalpha2-3Gal beta1-4 (Fucalpha1-3) GlcNAc-R, plays important roles in cell-to-cell interaction: for example, the E- and P-selectin-mediated influx of SLEX expressing leukocytes into inflamed areas. A human hepatocellular carcinoma cell line, HepG2 cells, was highly expressed SLEX on secreted glycoproteins and cell surface, in contrast with HuH-7 cells. We identified SLEX expressing glycoproteins in HepG2 cultured medium by two-dimensional polyacrylamide gel electrophoresis, followed by in gel digestion and peptide mass fingerprint using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOFMS), including transferrin, alpha₁-antitrypsin, alpha₂-HS glycoprotein and beta₂-glycoprotein. We analyzed N-glycans of these glycoproteins by MALDI-TOFMS in combination with exoglycosidase digestion; our results indicate increases in poly-fucosylated and high-branched N-glycans. High alpha1,3-fucosylation in glycoproteins would be caused by increased expression of alpha1,3-fucosyltransferase activities in HepG2 cells.

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