The contributions of individual galactosyltransferases to protein specific N-glycan processing in Chinese Hamster Ovary cells

Journal of Biotechnology
Nina BydlinskiNicole Borth

Abstract

Galactosylation as part of N-glycan processing is conducted by a set of beta-1,4-galactosyltransferases (B4GALTs), with B4GALT1 as the dominant isoenzyme for this reaction. Nevertheless, the exact contributions of this key-player as well as of the other isoenzymes involved in N-glycosylation, B4GALT2, B4GALT3 and B4GALT4, have not been studied in-depth. To increase the understanding of the protein- and site-specific activities of individual galactosyltransferases in Chinese Hamster Ovary cells, a panel of triple deletion cell lines was generated that expressed only one isoform of B4GALT each. Two model proteins were selected for this study to cover a large spectrum of possible N-glycan structures: erythropoietin and deamine-oxidase. They were expressed as Fc-fusion constructs (EPO-Fc and Fc-DAO) and their N-glycan processing status was analyzed by site-specific mass spectrometry. The sole activity of B4GALT1 resulted in a decrease of 15-21 % of fully galactosylated structures for erythropoietin, emphasizing the involvement of other isoenzymes. Interestingly, the contributions of B4GALT2 and B4GALT3 differed for the two model proteins. Unexpectedly, removal of galactosyltransferases influenced the overall process of N-glycan mat...Continue Reading

Citations

Sep 3, 2019·Biotechnology Journal·Michael T CoatsNicole Borth
Feb 23, 2021·The Journal of Biological Chemistry·Yoshiki NarimatsuHenrik Clausen
Mar 12, 2021·Biochemical Society Transactions·Roberto DoniniCleo Kontoravdi
Oct 10, 2019·ACS Synthetic Biology·Christopher S StachMichael Smanski
Nov 11, 2020·Gerontology·Jennifer KremerHildegard Isolde Dietlinde Mack

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