SUPPRESSION OF BETA-N-ACETYLGLUCOSAMINIDASE IN THE N-GLYCOSYLATION PATHWAY FOR COMPLEX GLYCOPROTEIN FORMATION IN DROSOPHILA S2 CELLS
SCIE
SCOPUS
- Title
- SUPPRESSION OF BETA-N-ACETYLGLUCOSAMINIDASE IN THE N-GLYCOSYLATION PATHWAY FOR COMPLEX GLYCOPROTEIN FORMATION IN DROSOPHILA S2 CELLS
- Authors
- Kim, YK; Kim, KR; Kang, DG; Jang, SY; Kim, YH; Cha, HJ
- Date Issued
- 2009-03
- Publisher
- OXFORD UNIV PRESS INC
- Abstract
- Most insect cells have a simple N-glycosylation process and consequently paucimannosidic or simple core glycans predominate. Previously, we have shown that paucimannosidic N-glycan structures are dominant in Drosophila S2 cells. It has been proposed that beta-N-acetylglucosaminidase (GlcNAcase), a hexosaminidase in the Golgi membrane which removes a terminal N-acetylglucosamine (GlcNAc), might contribute to simple N-glycosylation in several insects and insect-derived cells except S2 cells. In the present work, we investigated the substantial effects of GlcNAcase on N-glycan patterns in Drosophila S2 cells using two GlcNAcase suppression strategies: an mRNA-targeting approach using RNA interference (RNAi) and a protein-targeting approach using the specific chemical inhibitor 2-acetamido-1,2-dideoxynojirimycin (2-ADN). Using high-performance liquid chromatography (HPLC) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analyses, we found that the N-glycosylation patterns of human erythropoietin (hEPO) secreted by stably transfected S2 cells were more complex following GlcNAcase suppression, which generated N-glycan structures with a terminal GlcNAc and/or galactose. These data demonstrate that GlcNAcase may be an important factor in the formation of paucimannosidic core N-glycans in Drosophila S2 cells and suggest that it may be possible to express complex glycoproteins in engineered Drosophila S2 cells by suppressing GlcNAcase in the N-glycosylation pathway.
- Keywords
- SECRETED HUMAN ERYTHROPOIETIN; FUNCTIONAL-CHARACTERIZATION; STATISTICAL OPTIMIZATION; INSECT CELLS; PURIFICATION; GLYCANS; EXPRESSION; MEMBRANE; PROTEINS; ENCODES
- URI
- https://oasis.postech.ac.kr/handle/2014.oak/28830
- DOI
- 10.1093/glycob/cwn138
- ISSN
- 0959-6658
- Article Type
- Article
- Citation
- GLYCOBIOLOGY, vol. 19, no. 3, page. 301 - 308, 2009-03
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