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KMID : 0380020170320030193
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Korean Journal of Biotechnology and Bioengineering
2017 Volume.32 No. 3 p.193 ~ p.198
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Engineering Human-like Sialylation in CHO Cells Producing hCTLA4-Ig by Overexpressing α2,6-Sialyltransferase
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Lim Jin-Hyuk
Cha Hyun-Myoung
Park Hea-Jin
Kim Ha-Hyung
Kim Dong-Il
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Abstract
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Sialylation is important in producing therapeutic proteins such as antibody, cytokine and fusion protein. Thus, enhancement of sialylation is usually performed in CHO cell cultures. α2,6-Sialyltransferase (ST), which plays a key role in the attachment of α2,6-sialic acid, is present in human cells but not in Chinese hamster ovary (CHO) cells. Overexpression of α2,6-ST can be used for enhancing the degree of sialylation and achieving human-like glycosylation. In this study, we constructed CHO cells producing human cytotoxic T-lymphocyte antigen4-immunoglobulin (hCTLA4-Ig) as well as α2,6-ST. Transfected CHO cells were selected using G418 and stable cell line was established. Profiles of viable cell density and hCTLA4-Ig titer in an overexpressed cell line were similar to those of a wild-type cell line. It was confirmed that the total amount of sialic acid was increased and α2,6-sialic acid was attached to the terminal residues of N-glycan of hCTLA4-Ig by ESI-LC-MS. Compared to 100% of α2,3-sialic acid in wild type cells, 70.9% of total sialylated N-glycans were composed of α2,6-sialic acid in transfected cells. In conclusion, overexpression of α2,6-ST in CHO cells led to the increase of both the amount of total sialylated N-glycan and the content of α2,6-sialic acid, which is more resemble to human-like structure of glycosylation.
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KEYWORD
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α2,6-Sialyltransferase, Chinese hamster ovary cells, hCTLA4-Ig, Sialylation
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