Abstract
We have characterized two BHK cell lines (BHK-21B and BHK-21A) with different glycosylation properties. N-glycans synthesized by BHK-21B cells contain the typical N-acetyllactosamine motif (Gal(βl-4)GlcNAc-R) whereas BHK-21 A cells bear to a high amount nonsialylated terminal GalNAc(βl-4)GlcNAc-R moieties [1]. Due to the incapability of the endogenous oc2, 3-sialyltransferase to transfer NeuAc to the GalNAc(βl-4) GlcNAc-R structure recombinant glycoproteins produced by BHK-21 A cells are under-sialylated.
Therefore we have transfected BHK-21 A cells harbouring a plasmid encoding human EPO with the human Golgi enzyme CMP-NeuAc:Gal(βl-4)GlcNAc-R a2,6-sialyltransferase (ST6N). Detailed structural analysis of Oligosaccharides from the affinity purified recombinant EPO (HPAEC-PAD-mapping and MALDI/TOF-MS) revealed a significant increased NeuAc content when compared to the parent BHK-21 A cells without ST6N activity. Methylation analysis corroborated these results. The newly introduced α2,6-sialyltransferase recognizes the terminal GlcNAc-R motif as a substrate. The cell line obtained thus exhibits a ‘human kidney-type’ glycosylation characteristic [2].
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Preview
Unable to display preview. Download preview PDF.
Similar content being viewed by others
References
Grabenhorst, E., Hoffmann, A., Nimtz, M., Zettlmeissl, G., Conradt, H.S. (1995) Eur. J. Biochem. 232, 718–725.
Bergwerff, A.A., Van Oostrum, J., Kamerling, J.P., Vliegenthart, J.F.G. (1995) Eur. J. Biochem. 228, 113–126.
Nimtz, M., Martin, W., Wray, V., Klöppel, K.-D., Augustin, J., Conradt, H.S. (1993) Eur. J. Biochem. 213: 39–56.
Takeuchi, M., Takasaki, S., Miyazaki, H., Kato, T., Hoshi, S., Kochibe, N., Kobata, A. (1988) J. Biol. Chem. 263: 3657–3663.
Goldwasser, E., Kung, K.H., Eliasson, J. (1974) J. Biol. Chem. 249, 4202–4206.
Fukuda, M.N., Sasaki, H., Lopez, L., Fukuda, M. (1989) Blood 73, 84–89.
Spivak, J.L., Hogans, B.B. (1989) Blood 73, 90–99.
Fibi, M.R., Hermentin, P., Pauly, J.U., Lauffer, L., Zettlmeissl, G. (1995) Blood 85(5), 1229–1236.
Vara, J. A., Pórtela, A., Ortin, J., Jiménez, A. (1986) Nucleic Acids Res. 14, 4617–4624.
Nemansky, M., Van den Eijnden, D.H. (1992) Biochem. J. 287, 311–318.
Weisshaar, G., Hiyama, J., Renwick, A.G.C., Nimtz, M. (1991) Eur. J. Biochem 95, 257–268.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1997 Springer Science+Business Media Dordrecht
About this chapter
Cite this chapter
Schlenke, P., Grabenhorst, E., Wagner, R., Nimtz, M., Conradt, H.S. (1997). Expression of human α2, 6-Sialyltransferase in BHK-21A cells increases the sialylation of coexpressed human erythropoietin: NeuAc-transfer onto GalNAc(βl-4)GlcNAc-R motives. In: Carrondo, M.J.T., Griffiths, B., Moreira, J.L.P. (eds) Animal Cell Technology. Springer, Dordrecht. https://doi.org/10.1007/978-94-011-5404-8_76
Download citation
DOI: https://doi.org/10.1007/978-94-011-5404-8_76
Publisher Name: Springer, Dordrecht
Print ISBN: 978-94-010-6273-2
Online ISBN: 978-94-011-5404-8
eBook Packages: Springer Book Archive