Analysis of Cell Surface N-glycosylation of the Human Embryonic Kidney 293T Cell Line

Abstract

Protein glycosylation is a prominent posttranslational modification and is involved in many biological functions. Human cell lines used for the expression of recombinant glycoproteins present variations in their cell surface N-glycosylation due to their cell type–specific origin. We therefore investigated the presence of specific glycosyltransferases by RT-PCR and the cell surface N-glycan structures of HEK293T cells by MALDI-TOF-MS and MALDI-TOF/TOF-MS analyses. Expression of N-acetylglucosaminyltransferase-III and fucosyltransferase-VIII were coincident with the presence of bisecting N-acetylglucosamine and high amounts of core-fucosylated N-glycans. Furthermore, a high overall amount of sialylated N-glycans and the expression of α2,3- and α2,8-specific, but not α2,6-specific, sialyltransferases were found.

Keywords:

• HEK293T
• N-glycosylation
• Glycosyltransferases
• RT-PCR
• MALDI- TOF-MS

ACKNOWLEDGEMENTS

This work was supported by the Bundesministerium für Bildung und Forschung by a grant to S.H. of the program “Arbeitsgruppenwettbewerb Glykobiotechnologie” and to M. Be. “InnoProfile 03IP511.” Further support by the “Europäischer Fonds für regionale Entwicklung” (S.H.) is acknowledged. We thank Elena Frisch and Detlef Grunow for their technical assistance in MALDI-TOF-MS experiments.