Glycosylation by Chinese hamster ovary cells in dolichol phosphate-supplemented cultures.
N-linked glycosylation often imparts important properties to protein therapeutics. An essential step in this intracellular process is the transfer of oligosaccharide from dolichol monophosphate (Dol-P) to a potential glycosylation site. Variability in the success rate of this reaction affects the extent of protein glycosylation. The critical role of Dol-P suggests that its availability may influence the extent of glycosylation by limiting the pool of lipid-linked oligosaccharides (LLOs), the glycosyl donor. To test this hypothesis, the impact of Dol-P supplementation on protein glycosylation in Chinese hamster ovary (CHO) cells was investigated. Although exogenous Dol-P was incorporated by CHO cells and processed into LLOs in a dose-dependent manner, Dol-P supplementation had no marked effects on LLO or overall cellular glycosylation levels. While concentrations of exogenous Dol-P exceeding 100 microg/ml were detrimental to CHO cell viability, maximum non-toxic supplemental doses of Dol-P had no significant impact on the glycosylation of recombinant interferon- gamma produced by batch cultures of CHO cells. These results show that glycosylation in CHO cells cannot be readily enhanced by Dol-P feeding under normal culture conditions.
Yuk
Inn H Y
IH
Biotechnology Process Engineering Center and Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Wang
Daniel I C
DI
D000818:Animals; D016466:CHO Cells / Q000187:drug effects / Q000378:metabolism*; D018929:Cell Culture Techniques / Q000379:methods; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D003470:Culture Media; D004288:Dolichol Phosphates / Q000493:pharmacokinetics / Q000494:pharmacology* / Q000633:toxicity; D004305:Dose-Response Relationship, Drug; D005947:Glucose / Q000378:metabolism; D006031:Glycosylation / Q000187:drug effects; D007371:Interferon-gamma / Q000378:metabolism*; D007930:Leucine / Q000378:metabolism; D008070:Lipopolysaccharides / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism*
D003470:Culture Media; D004288:Dolichol Phosphates; D008070:Lipopolysaccharides; D011506:Proteins; D011994:Recombinant Proteins; C023023:lipid-linked oligosaccharides; D007371:Interferon-gamma; D007930:Leucine; D005947:Glucose