Enhanced sialylation of recombinant erythropoietin in chinese hamster ovary cells = CHO 세포에서 재조합 Erythropoietin의 시알산 증진

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Sialic acid, the terminal sugar in N-linked complex glycans, is usually found in glycoproteins. Sialic acid residues play a major role in determining the circulatory lifespan of glycoproteins. Therefore, it is often desirable to maximize the sialic acid content of glycoproteins used as therapeutic agents to ensure their quality and consistency. In this study, we tried to enhance the sialylation of recombinant erythropoietin (EPO) in Chinese hamster ovary (CHO) cells. N-Acetylcysteine (NAC) is one of mild thiol reducing agent. We have previously shown that the anti-apoptotic reagent, NAC not only blocks apoptosis but also increases the production of recombinant erythropoietin. We also found that the productive effect of NAC obtained without the apoptotic condition. However, NAC resulted in decreasing the glycosylation performance, sialylation. Human α2,3-sialyltransferase (α2,3-ST) was introduced into CHO cells, which produce recombinant human EPO, in order to compensate for the reduced sialylation under supplementation of NAC. Sialyltransferase is responsible for terminal sialylation. When NAC was treated in culture media, EPO sialylation was reduced. However, when α2,3-ST was expressed under the supplementation of NAC, reduced sialylation was restored and even more sialylated glycans were produced. Thus, our study is significant in that it offers the possibility of preventing decreased sialylation of EPO while still allowing increased EPO production. Sialylation requires the metabolic generation of the nucleotide sugar, CMP-sialic acid, followed by the transfer of the sialic acid to an acceptor oligosaccharide in the Golgi apparatus by sialyltransferases. In the case of mammalian cells, CMP-sialic acid is generated from sialic acid by CMP-sialic acid synthase (CMP-SAS). To enhance EPO sialylation, we introduced human α2,3-ST and CMP-SAS into recombinant human EPO-producing CHO cells. The sialylation of EPO was increased by the expression of \alpha2...
Advisors
Kim, Jung-Hoeresearcher김정회researcher
Description
한국과학기술원 : 생명과학과,
Publisher
한국과학기술원
Issue Date
2008
Identifier
295331/325007  / 020015871
Language
eng
Description

학위논문(박사) - 한국과학기술원 : 생명과학과, 2008.2, [ ix, 97 p. ]

Keywords

Sialic Acid; Erythropoietin; Chinese Hamster Ovary Cells; Glycosylation; 시알산; 에리스로포이에틴; 중국햄스터난소세포; 당쇄합성; Sialic Acid; Erythropoietin; Chinese Hamster Ovary Cells; Glycosylation; 시알산; 에리스로포이에틴; 중국햄스터난소세포; 당쇄합성

URI
http://hdl.handle.net/10203/27637
Link
http://library.kaist.ac.kr/search/detail/view.do?bibCtrlNo=295331&flag=dissertation
Appears in Collection
BS-Theses_Ph.D.(박사논문)
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