神戸大学附属図書館デジタルアーカイブ
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https://hdl.handle.net/20.500.14094/90008321
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2024-04-26
05:09 集計
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90008321 (fulltext)
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メタデータID
90008321
アクセス権
open access
出版タイプ
Accepted Manuscript
タイトル
Production of an antibody Fab fragment using 2A peptide in insect cells
著者
Mizote, Yu ; Masumi-Koizumi, Kyoko ; Katsuda, Tomohisa ; Yamaji, Hideki
著者名
Mizote, Yu
著者名
Masumi-Koizumi, Kyoko
著者ID
A1208
研究者ID
1000050335460
KUID
https://kuid-rm-web.ofc.kobe-u.ac.jp/search/detail?systemId=0746680e0db53e41520e17560c007669
著者名
Katsuda, Tomohisa
勝田, 知尚
カツダ, トモヒサ
所属機関名
工学研究科
著者ID
A0921
研究者ID
1000040283874
KUID
https://kuid-rm-web.ofc.kobe-u.ac.jp/search/detail?systemId=87d20630482bf7df520e17560c007669
著者名
Yamaji, Hideki
山地, 秀樹
ヤマジ, ヒデキ
所属機関名
工学研究科
収録物名
Journal of Bioscience and Bioengineering
巻(号)
130(2)
ページ
205-211
出版者
Elsevier
刊行日
2020-08
公開日
2021-09-01
抄録
Antibody Fab fragments consist of heavy chain (Hc) and light chain (Lc) polypeptides assembled with a disulphide bond. The production of a recombinant Fab fragment requires the simultaneous expression of two genes encoding both an Hc and an Lc in the same host cell. In the present study, we investigated the production of Fab fragments in lepidopteran insect cells using a bicistronic plasmid vector carrying the Hc and Lc genes linked with a 2A self-cleaving peptide sequence from the porcine teschovirus-1. We also examined the arrangement of a GSG spacer sequence and a furin cleavage site sequence with the 2A sequence. Western blot analysis and enzyme-linked immunosorbent assay (ELISA) of culture supernatants showed that Trichoplusia ni BTI-TN-5B1-4 (High Five) cells transfected with a plasmid in which the Hc and Lc genes were joined by the 2A sequence successfully secreted Fab fragments with antigen-binding activity after self-cleavage of the 2A peptide. The GSG linker enhanced 2A cleavage efficiency, and the furin recognition site was useful for removal of 2A residues from the Hc. Transfection with a single plasmid that contained sequences for GSG, the furin cleavage site, GSG, and the 2A peptide between the Hc and Lc genes exhibited a higher productivity than co-transfection with a set of plasmids separately carrying the Hc or Lc gene. These results demonstrate that bicistronic expression with the appropriate combination of a furin recognition site, GSG linkers, and a 2A peptide may be an effective way to efficiently produce recombinant antibody molecules in insect cells.
キーワード
Insect cell
2A peptide
GSG linker
Furin cleavage site
Fab fragment
High Five cell
Recombinant protein production
カテゴリ
工学研究科
学術雑誌論文
権利
© 2020 The Society for Biotechnology, Japan.
This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/
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資源タイプ
journal article
言語
English (英語)
ISSN
1389-1723
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NCID
AA11307678
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関連情報
DOI
https://doi.org/10.1016/j.jbiosc.2020.03.009
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