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].
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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
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DOI: https://doi.org/10.1007/978-94-011-5404-8_76
Publisher Name: Springer, Dordrecht
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