Nickel and cobalt affect galactosylation of recombinant IgG expressed in CHO cells
- 286 Downloads
Glycosylation is an important product quality attribute of antibody biopharmaceuticals. It involves enzymatic addition of oligosaccharides on proteins by sequential action of glycosyltransferases and glycosidases in the endoplasmic reticulum and golgi. Some of these enzymes like galactosyltransferase and N-acetylglucosaminyltransferase-I require trace metal cofactors. Variations in trace metal availability during production can thus affect glycosylation of recombinant glycoproteins such as monoclonal antibodies. Variability in trace metal concentrations can be introduced at multiple stages during production such as due to impurities in raw materials for culture medium and leachables from bioreactors. Knowledge of the effect of various trace metals on glycosylation can help in root-cause analysis of unintended variability in glycosylation. In this study, we investigated the effect of nickel and cobalt on glycosylation of recombinant IgG expressed in Chinese hamster ovary cells. Nickel concentrations below 500 µM did not affect glycosylation, but above 500 µM it significantly decreases galactosylation of IgG. Cobalt at 50 µM concentration causes slight increase in G1F glycans (mono galactosylated) as previously reported. However, higher concentrations result in a small increase in G0F (non galactosylated) glycans. This effect of nickel and cobalt on galactosylation of recombinant IgG can be reversed by supplementation of uridine and galactose which are precursors to UDP-Galactose, a substrate for the enzymatic galactosylation reaction.
KeywordsGlycosylation Trace metals Process variability Nickel Cobalt Galactosylation
MG acknowledges funding from the Department of Biotechnology, Government of India. The authors are thankful to the MALDI-MS facility at CSIR-NCL and to Dr. Gadre for help with HPLC-based galactosyltransferase assay.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
- Campbell C, Stanley P (1984) A dominant mutation to ricin resistance in Chinese hamster ovary cells induces UDP-GlcNAc: glycopeptide beta-4-N-acetylglucosaminyltransferase III activity. J Biol Chem 259:13370–13378Google Scholar
- Clincke M-F, Guedon E, Yen FT, Ogier V, Goergen J-L (2011) Effect of iron sources on the glycosylation macroheterogeneity of human recombinant IFN-γ produced by CHO cells during batch processes. In: BMC proceedings. BioMed Central, London, p P114Google Scholar
- Goh JB, Ng SK (2017) Impact of host cell line choice on glycan profile. Crit Rev Biotechnol 38:1–17Google Scholar
- Hossler P, Racicot C, McDermott S (2014) Targeted shifting of protein glycosylation profiles in mammalian cell culture through media supplementation of cobalt. J Glycobiol 3:108Google Scholar
- Kuhn NJ, Ward S, Leong WS (1991) Submicromolar manganese dependence of Golgi vesicular galactosyltransferase (lactose synthetase). FEBS J 195:243–250Google Scholar
- Powell JT, Brew K (1976) Metal ion activation of galactosyltransferase. J Biol Chem 251:3645–3652Google Scholar
- Salnikow K, Su W, Blagosklonny MV, Costa M (2000) Carcinogenic metals induce hypoxia-inducible factor-stimulated transcription by reactive oxygen species-independent mechanism. Cancer Res 60:3375–3378Google Scholar
- Wentz AE, Hemmavanh D, Matuck JG (2015) Use of metal ions for modulation of protein glycosylation profiles of recombinant proteins. US Patent 9,598,667, B2Google Scholar