Please use this identifier to cite or link to this item: https://doi.org/10.1002/bit.20815
Title: Enhancing recombinant glycoprotein sialylation through CMP-sialic acid transporter over expression in Chinese hamster ovary cells
Authors: Wong, N.S.C.
Yap, M.G.S. 
Wang, D.I.C.
Keywords: CHO cells
CMP-sialic acid transporter
Glycosylation
Over expression
Protein quality
Sialylation
Issue Date: 5-Apr-2006
Source: Wong, N.S.C., Yap, M.G.S., Wang, D.I.C. (2006-04-05). Enhancing recombinant glycoprotein sialylation through CMP-sialic acid transporter over expression in Chinese hamster ovary cells. Biotechnology and Bioengineering 93 (5) : 1005-1016. ScholarBank@NUS Repository. https://doi.org/10.1002/bit.20815
Abstract: Glycosylation engineering strategies that are currently used to improve quality of recombinant glycoproteins involve the manipulation of glycosyltransferase and/or glycosidase expression. We explored the possibility that over expressing nucleotide sugar transporters, particularly the CMP-sialic acid transporter (CMP-SAT) would improve the sialylation process in Chinese hamster ovary cells (CHO). Our hypothesis was that increasing CMP-SAT in the cells through recombinant means would increase the transport of CMP-sialic acid into the Golgi, resulting in an increased CMP-sialic acid intra-lumenal pool and increased sialylation of the proteins produced. We report the construction of the CMP-SAT expression vector (pcDNA-SAT) using hamster CMP-SAT (GenBank accession number Y12074) and demonstrated its functionality using Lec2 CHO mutant cells. Transfection of pcDNA-SAT into CHO IFN-γ, a CHO cell line producing recombinant human interferon-gamma (IFN-γ) resulted in single clones that had 2-20 fold increase in total CMP-SAT expression atthetranscript level and 1.8-2.8 fold increase in CMP-SAT at the protein level when compared to untransfected parent CHO IFN-γ. This resulted in 4%-16% increase in site sialylation of IFN-γ. There was also a higher proportion of the more sialylated IFN-γ glycans produced by the clones. We have thus established a novel strategy for sialylation improvement in recombinant protein production that can be considered singly or along with existing glycosylation improvement strategies, including glycosyltransferase over expression and nucleotide sugar feeding. These multiprong approaches can possibly bring us closer toward the goal of maximum and consistent sialylation in glycoprotein production using mammalian cells. © 2006 Wiley Periodicals, Inc.
Source Title: Biotechnology and Bioengineering
URI: http://scholarbank.nus.edu.sg/handle/10635/63850
ISSN: 00063592
DOI: 10.1002/bit.20815
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