Metabolic shifts do not influence the glycosylation patterns of a recombinant fusion protein expressed in BHK cells.

Autor: Cruz HJ; IBET/ITQB- Instituto de Biologia Experimental e Tecnológica / Instituto de Tecnologia Química e Biológica, Apartado 12, P-2781-901 Oeiras, Portugal., Peixoto CM, Nimtz M, Alves PM, Dias EM, Moreira JL, Carrondo MJ
Jazyk: angličtina
Zdroj: Biotechnology and bioengineering [Biotechnol Bioeng] 2000 Jul 20; Vol. 69 (2), pp. 129-39.
DOI: 10.1002/(sici)1097-0290(20000720)69:2<129::aid-bit2>3.0.co;2-5
Abstrakt: BHK-21 cells expressing a human IgG-IL2 fusion protein, with potential application in tumor-targeted therapy, were grown under different nutrient conditions in a continuous system for a time period of 80 days. At very low-glucose (< 0.5 mM) or glutamine (< 0. 2 mM) concentrations, a shift toward an energetically more efficient metabolism was observed. Cell-specific productivity was maintained under metabolically shifted growth conditions and at the same time an almost identical intracellular ATP content, obtained by in vivo (31)P NMR experiments, was observed. No significant differences in the oligosaccharide structures were detected from the IgG-IL2 fusion protein preparations obtained by growing cells under the different metabolic states. By using oligosaccharide mapping and MALDI/TOF-MS, only neutral diantennary oligosaccharides with or without core alpha1-6-linked fucose were detected that carried no, one or two beta1-4-linked galactose. Although the O-linked oligosaccharide structures that are present in the IL2 moiety of the protein were studied with less detail, the data obtained from the hydrazinolysis procedure point to the presence of the classical NeuAcalpha2-3Galbeta1-3GalNAc structure. Here, it is shown that under different defined cellular metabolic states, the quality of a recombinant product in terms of O- and N-linked oligosaccharides is stable, even after a prolonged cultivation period. Moreover, unaffected intracellular ATP levels under the different metabolic states were observed.
(Copyright 2000 John Wiley & Sons, Inc.)
Databáze: MEDLINE
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