Influence of intracellular nucleotide and nucleotide sugar contents on recombinant interferon-γ glycosylation during batch and fed-batch cultures of CHO cells

✍ Scribed by N. Kochanowski; F. Blanchard; R. Cacan; F. Chirat; E. Guedon; A. Marc; J.-L. Goergen

• Publisher: John Wiley and Sons
• Year: 2008
• Tongue: English
• Weight: 314 KB
• Volume: 100
• Category: Article
• ISSN: 0006-3592
• DOI: 10.1002/bit.21816

✦ Synopsis

Abstract

Both the macroheterogeneity of recombinant human IFN‐γ produced by CHO cells and intracellular levels of nucleotides and sugar nucleotides, have been characterized during batch and fed‐batch cultures carried out in different media. Whereas PF‐BDM medium was capable to maintain a high percentage of the doubly‐ glycosylated glycoforms all over the process, mono‐glycosylated and non‐glycosylated forms increased during the batch culture using SF‐RPMI medium. Intracellular level of UTP was higher in PF‐BDM all over the batch culture compared to the SF‐RPMI process. UDP‐Gal accumulated only during the culture performed in PF‐BDM medium, probably as a consequence of the reduced UDP‐Glc synthesis flux in SF‐RPMI medium. When the recombinant CHO cells were cultivated in fed‐batch mode, the UTP level remained at a relatively high value in serum‐containing RPMI and its titer increased during the fed‐phase indicating an excess of biosynthesis. Besides, an accumulation of UDP‐Gal occurred as well. Those results all together indicate that UTP and UDP‐Glc syntheses in CHO cells cultivated in SF‐RPMI medium in batch process, could be limiting during the glycosylation processes of the recombinant IFN‐γ. At last, the determination of the energetic status of the cells over the three studied processes suggested that a relationship between the adenylate energy charge and the glycosylation macroheterogeneity of the recombinant IFN‐γ may exist. Biotechnol. Bioeng. 2008;100: 721–733. © 2008 Wiley Periodicals, Inc.