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• W2128761336 abstract "Asparagine linked (N-linked) glycosylation is an important modification of recombinant proteins, because the attached oligosaccharide chains can significantly alter protein properties. Potential glycosylation sites are not always occupied with oligosaccharide, and site occupancy can change with the culture environment. To investigate the relationship between metabolism and glycosylation site occupancy, we studied the glycosylation of recombinant human interferon-γ (IFN-γ) produced in continuous culture of Chinese hamster ovary cells. Intracellular nucleotide sugar levels and IFN-γ glycosylation were measured at different steady states which were characterized by central carbon metabolic fluxes estimated by material balances and extracellular metabolite rate measurements. Although site occupancy varied over a rather narrow range, we found that differences correlated with the intracellular pool of UDP-N-acetylglu- cosamine + UDP-N-acetylgalactosamine (UDP-GNAc). Measured nucleotide levels and estimates of central carbon metabolic fluxes point to UTP depletion as the cause of decreased UDP-GNAc during glucose limitation. Glucose limited cells preferentially utilized available carbon for energy production, causing reduced nucleotide biosynthesis. Lower nucleoside triphosphate pools in turn led to lower nucleotide sugar pools and reduced glycosylation site occupancy. Subsequent experiments in batch and fed-batch culture have confirmed that UDP-sugar concentrations are correlated with UTP levels in the absence of glutamine limitation. Glutamine limitation appears to influence glycosylation by reducing amino sugar formation and hence UDP-GNAc concentration. The influence of nucleotide sugars on site occupancy may only be important during periods of extreme starvation, since relatively large changes in nucleotide sugar pools led to only minor changes in glycosylation. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 62: 336–347, 1999." @default.
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• W2128761336 date "1999-02-05" @default.
• W2128761336 modified "2023-10-18" @default.
• W2128761336 title "Metabolic effects on recombinant interferon-? glycosylation in continuous culture of Chinese hamster ovary cells" @default.
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• W2128761336 doi "https://doi.org/10.1002/(sici)1097-0290(19990205)62:3<336::aid-bit10>3.0.co;2-n" @default.
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