Studies on the relationship between metabolism and control of N-linked Glycosylation in CHO cells

Abstract

The process of post-translational attachment of oligosaccharide structures to asparagine residues in nascent polypeptides is termed N-linked glycosylation. These glycan structures can influence the folding, stability, solubility, antigenicity, activity and rate of clearance of a glycoprotein from the bloodstream. Cellular metabolism for the generation of energy and biosynthetic intermediates is intrinsically linked to the process of N-linked glycosylation. Consequently, the mechanisms by which altered metabolism of Chinese hamster ovary cells (CHO) affected the N-linked glycosylation of recombinantly expressed human chorionic gonadotrophin (HCG) was explored in a batch and continuous mode of cultivation. The structures of the N-linked glycans attached to HCG were assessed with the use of IEF and HPLC, and, cellular metabolism was assessed by examining rates of consumption and production of metabolites and using metabolic flux analysis. The mechanism of any changes in glycosylation was determined by assaying the intracellular content of nueleotides and sugar nucleotides.