Enhancement of sialylation on humanized IgG-like bispecific antibody by overexpression of α2,6-sialyltransferase derived from Chinese hamster ovary cells.
Improvement of glycosylation is one of the most important topics in the industrial production of therapeutic antibodies. We have focused on terminal sialylation with alpha-2,6 linkage, which is crucial for anti-inflammatory activity. In the present study, we have successfully cloned cDNA of beta-galactosyl alpha-2,6 sialyltransferase (ST6Gal I) derived from Chinese hamster ovary (CHO) cells regardless of reports that stated this was not endogenously expressed in CHO cells. After expressing cloned ST6Gal I in Escherichia coli, the transferase activity was confirmed by HPLC and lectin binding assay. Then, we applied ST6Gal I to alpha-2,6 sialylation of the recombinant antibody; the ST6Gal I expression vector was transfected into the CHO cell line producing a bispecific antibody. The N-glycosylation pattern of the antibody was estimated by HPLC and sialidase digestion. About 70% of the total N-linked oligosaccharide was alpha-2,6 sialylated in the transfected cell line whereas no sialylation was observed in the non-transfected cell line. The improvement of sialylation would be of practical importance for the industrial production of therapeutic antibodies.
Institute of Technology and Science, The University of Tokushima, 2-1 Minamijosanjima-cho, Tokushima 770-8506, Japan.
Masayoshi
M
Onitsuka
Wook-Dong
WD
Kim
Hiroyuki
H
Ozaki
Akira
A
Kawaguchi
Kohsuke
K
Honda
Hiroyuki
H
Kajiura
Kazuhito
K
Fujiyama
Ryutaro
R
Asano
Izumi
I
Kumagai
Hisao
H
Ohtake
Takeshi
T
Omasa
D000818:Animals; D018033:Antibodies, Bispecific / Q000235:genetics / Q000378:metabolism*; D061067:Antibodies, Monoclonal, Humanized / Q000235:genetics / Q000378:metabolism*; D016466:CHO Cells; D006224:Cricetinae / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics / Q000378:metabolism*; D010053:Ovary / Q000201:enzymology*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012799:Sialyltransferases / Q000235:genetics* / Q000378:metabolism; D015854:Up-Regulation; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase
D018033:Antibodies, Bispecific; D061067:Antibodies, Monoclonal, Humanized; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D012799:Sialyltransferases; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase