, Volume 15, Issue 1–3, pp 209–215 | Cite as

Effect of lipid supplements on the production and glycosylation of recombinant interferon-γ expressed in CHO cells

  • Nigel Jenkins
  • Paula Castro
  • Sunitha Menon
  • Andrew Ison
  • Alan Bull


The effects of lipids on the glycosylation of recombinant human interferon-γ expressed in a Chinese Hamster Ovary cell line were investigated in batch culture. Lipids form an essential part of the N-glycosylation pathway, and have been shown to improve cell viability. In control (serum-free) medium the proportion of fully-glycosylated interferon-γ deteriorated reproducibly with time in batch culture, but the lipoprotein supplement ExCyte was shown to minimise this trend. Partially substituting the bovine serum albumin content of the medium with a fatty-acid free preparation also improved interferon-γ glycosylation, possibly indicating that oxidised lipids carried on Cohn fraction V albumin may damage the glycosylation process.

Key words

Glycosylation recombinant interferon CHO cells lipids 



bovine serum albumin


chinese hamster ovary


dihydrofolate reductase


foetal calf serum


human interferon-gamma


specific interferon production rate


specific growth rate








Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Bause E (1984) Model studies on N-glycosylation of proteins. Biochem. Soc. Trans. 12: 514–517.Google Scholar
  2. Bulleid NJ, Curling EM, Freedman RB and Jenkins N (1990) Source of heterogeneity in secreted interferon-gamma. A study on products of translation in vitro. Biochem. J, 268: 777–781.Google Scholar
  3. Bulleid NJ, Bassel-Duby RS, Freedman RB, Sambrook J and Gething MJ (1992) Cell-free synthesis of enzymically active tissue-type plasminogen activator. Protein folding determines the extent of N-linked gycosylation. Biochem. J. 286: 275–280.Google Scholar
  4. Cumming DA (1991) Glycosylation of recombinant protein therapeutics: control and functional implications. Glycobiology 1: 115–130.Google Scholar
  5. Curling EM, Hayter PM, Baines AJ, Bull AT, Gull K, Strange PG and Jenkins N (1990) Recombinant human interferon-gamma. Differences in glycosylation and proteolytic processing lead to heterogeneity in batch culture. Biochem J, 272: 333–337.Google Scholar
  6. Ealick SE, Cook WJ, Vijaykumar S, Carson M, Nagabhushan TL, Trotta PP and Bugg CE (1991) 3-dimensional structure of recombinant human interferon-gamma. Science 252: 698–702.Google Scholar
  7. Farrar MA and Schreiber RD (1993) The molecular cell biology of interferon-gamma and its receptor. Ann. Rev. Immunol. 11: 571–611.Google Scholar
  8. Finlayson JS (1980) Albumin products. Sem. Thromb. Hem. 6: 85–120.Google Scholar
  9. Gavel Y and von Heijne G (1990) Sequence differences between glycosylated and non-glycosylated Asn-X-Thr/Ser acceptor sites: implication for protein engineering. Protein Eng. 3: 433–442.Google Scholar
  10. Goochee CF, Gramer MJ, Andersen DC, Bahr JB and Rasmussen JR (1991) The oligosaccharides of glycoproteins-bioprocess factors affecting oligosaccharide structure and their effect on glycoprotein properties. Bio/technology 9: 1347–1355.Google Scholar
  11. Gramer MJ and Goochee CF (1993) Glycosidase activities in chinese hamster ovary cell lysate and cell culture supernatant. Biotechnol. Prog. 9: 366–373.Google Scholar
  12. Cizesiek S, Dobeli H, Gentz R, Garotta G, Labhardt AM and Bax A (1992)1H,13C, and15N NMR backbone assignments and secondary structure of human interferon-gamma. Biochemistry 31: 8180–8190.Google Scholar
  13. Hayer PM, Curling EM, Baines AJ, Jenkins N, Salmon I, Strange PG and Bull AT (1991) Chinese hamster ovary growth and interferon production kinetics in stirred batch culture. Appl. Microbiol. Biotechnol. 34: 559–564.Google Scholar
  14. Hayter PM, Curling EM, Baines AJ, Jenkins N, Salmon I, Strange PG, Tong JM and Bull AT (1992) Glucose-limited chemostat culture of chinese hamster ovary cells producing recombinant human interferon-gamma. Biotechnol. Bioeng. 39: 327–335.Google Scholar
  15. Hayter PM, Curling EM, Gould ML, Baines AJ, Jenkins N, Salmon I, Strange PG and Bull AT (1993) The effect of dilution rate on CHO cell physiology and recombinant interferon-gamma production in glucose-limited chemostat cultures. Biotechnol. Bioeng. 42: 1077–1085.Google Scholar
  16. Hewlett G, Duvinski MS and Montalto JG (1989) Pentex ExCyte growth enhancement media supplement as a lipoprotein additive for mammalian cell culture. Miles Science Journal 11: 9–14.Google Scholar
  17. James DC, Freedman RB, Hoare M and Jenkins N (1994) High resolution separation of recombinant human interferongamma by micellar electrokinetic capillary chromatography. Anal. Biochem. (in press).Google Scholar
  18. Jenkins N (1991) Growth Factors. In: Butler M. (ed.) Mammalian Cell Biotechnology: A Practical Approach. (pp. 39–55) Oxford University Press, Oxford, U.K.Google Scholar
  19. Jenkins N, Wingrove C, Strange PG, Baines AJ, Curling EM, Freedman RB and Pucci P (1993) Changes in the glycosylation pattern of interferon-gamma during batch culture. In: Kaminogawa S, Ametani A and Hachimura S (ed.) Animal Cell Technology: Basic and Applied Aspects Vol. 5 (pp. 231–235) Kluwer Academic Publishers, Dordecht, The Netherlands.Google Scholar
  20. Jenkins N and Curling EM (1994) Glycosylation of recombinant proteins: problems and prospects. Enzyme Microb. Technol. 16: 354–364.Google Scholar
  21. Kaiden A and Krag SS (1992) Dolichol metabolism in Chinese hamster ovary cells. Biochem. Cell. Biol. 70: 385–389.Google Scholar
  22. Kovar J and Franek F (1986) Serum-free medium for hybridoma and parental myeloma cell cultivation. Methods Enzymol. 121: 277–292.Google Scholar
  23. Lehrman MA (1991) Biosynthesis of N-acetylglucosamine-P-P-dolichol, the committed step of asparagine-linked oligosaccharide assembly. Glycobiology 1: 553–562.Google Scholar
  24. Lin AA, Kimura R and Miller WM (1993) Production of tPA in recombinant cho cells under oxygen-limited conditions. Biotechnol. Bioeng. 42: 339–350.Google Scholar
  25. Lund JT, Takahashi N, Hindley SA, Tyler R, Goodall M and Jefferis R (1993) Glycosylation of human IgG subclass and mouse IgG2b heavy chains secreted by mouse J558L transfectoma cell lines as chimeric antibodies. Human Antibodies and Hybridomas 4: 20–25.Google Scholar
  26. Maiorella BL, Winkelhake J, Young J, Moyer B, Bauer R, Hora M, Andya J, Thomson J, Patel T and Parekh RB (1993) Effect of culture conditions on IgM antibody structure, pharmacokinetics and activity. Bio/technology 11: 387–392.Google Scholar
  27. Minamoto Y, Ogawa K, Abe H, Iochi Y and Mitsugi K (1991) Development of a serum-free and heat sterilizable medium and continuous high density cell culture. Cytotechnology 5: 35–51.Google Scholar
  28. Patel TP, Parekh RB, Moellering BJ and Prior CP (1992) Different culture methods lead to diferences in glycosylation of a murine IgG monoclonal antibody. Biochem. J. 285: 839–845.Google Scholar
  29. Rosenwald AG, Stoll J and Krag SS (1990) Regulation of glycosylation. Three enzymes compete for a common pool of dolichyl phosphate in vivo. J. Biol. Chem. 265:14544–14553.Google Scholar

Copyright information

© Kluwer Academic Publishers 1994

Authors and Affiliations

  • Nigel Jenkins
    • 2
  • Paula Castro
    • 1
  • Sunitha Menon
    • 1
  • Andrew Ison
    • 2
  • Alan Bull
    • 1
  1. 1.Research School of BiosciencesUniversity of KentCanterburyUK
  2. 2.SERC Advanced Centre for Biochemical Engineering, University CollegeLondon UniversityLondonUK

Personalised recommendations