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タイトル: | Domain swapping of complementarity-determining region in nanobodies produced by Pichia pastoris |
著者: | Miura, Natsuko Miyamoto, Kana Ohtani, Yuta Yaginuma, Kenshi Aburaya, Shunsuke Kitagawa, Yoshinori Aoki, Wataru https://orcid.org/0000-0002-3118-9390 (unconfirmed) Ueda, Mitsuyoshi |
著者名の別形: | 三浦, 夏子 宮本, 佳奈 大谷, 優太 柳沼, 謙志 油屋, 駿介 北川, 義康 青木, 航 植田, 充美 |
キーワード: | CDR swapping CD4 nanobody GFP nanobody Relative binding ability Pichia pastoris |
発行日: | 15-Jul-2019 |
出版者: | Springer Science and Business Media LLC |
誌名: | AMB Express |
巻: | 9 |
論文番号: | 107 |
抄録: | Easy preparation of chimeric nanobodies with various scaffolds is important for customizing abilities of nanobodies toward practical utilization. To accomplish high-throughput production of various nanobodies, utilization of microbes is an attractive option. In the present study, various chimeric nanobodies were prepared using the methylotrophic yeast Pichia pastoris. We designed chimeric nanobodies with complementarity-determining regions (CDRs) against green fluorescent protein (GFP) or cluster of differentiation 4 (CD4) based on the scaffold of GFP-nanobody. FLAG-tagged chimeric nanobodies were prepared by one-step cloning and produced using P. pastoris. Secreted chimeric nanobodies were purified from the culture media of P. pastoris transformants. Relative binding abilities of purified chimeric nanobodies to GFP and CD4 was tested using a BIACORE T-200. P. pastoris successfully produced a high yield of FLAG-tagged chimeric nanobodies. FLAG-tagged GFP- and CD4-nanobodies were shown to specifically bind to GFP and CD4, respectively. Chimeric nanobodies, in which the CDR2 or 3 of GFP-nanobody was replaced with CDRs of CD4-nanobody, acquired the ability to bind to CD4 without binding to GFP. These results demonstrate successful production of functional chimeric nanobodies using P. pastoris. These results also suggest that swapping of CDRs, especially CDRs 2 or 3, potentially enables a novel method of creating nanobodies. |
著作権等: | © The Author(s) 2019. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. |
URI: | http://hdl.handle.net/2433/250107 |
DOI(出版社版): | 10.1186/s13568-019-0833-2 |
PubMed ID: | 31309388 |
出現コレクション: | 学術雑誌掲載論文等 |
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