title: Effect of different cell culture conditions on the polypeptide integrity and N-glycosylation of a recombinant model glycoprotein.
abstract: The effect of different short-term controlled cell culture conditions on the product quality of a genetically engineered human interleukin-2 N-glycosylation variant protein expressed from a baby hamster kidney cell line (BHK-21) has been investigated. A perfused 2-L stirred tank reactor was used. Products purified from the culture supernatant of cells grown under experimentally initiated nutrient limitations (glucose, amino acids, pO(2)) were characterized by their HPLC-elution profile, SDS-PAGE and western blotting, amino acid sequencing as well as for their N-linked carbohydrates, using "HPAEC-PAD fingerprinting" and methylation analysis. The glycoprotein products secreted from cells under the different culture conditions (kept for 24 h, after an adaption time period of 48 h) showed an almost identical oligosaccharide pattern. By contrast, short-term changes of the culture condition led to considerable differences in the ratio of glycosylated to unglycosylated protein forms. Significant amounts of NH(2)-terminally truncated polypeptide forms were observed. They lacked proponderantly the first two amino acids; however, under certain culture conditions forms lacking up to eight NH(2)-terminal amino acids were detected. (
authors.affiliation: Abteilung Zellkulturtechnik, Gesellschaft für Biotechnologische Forschung mbH, Mascheroder Weg 1, D-38124 Braunschweig, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Gawlitzek
authors.forename: H S
authors.initials: HS
authors.lastname: Conradt
authors.forename: R
authors.initials: R
authors.lastname: Wagner