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Production of ORF8 protein from SARS-CoV-2 using an inducible virus-mediated expression system in suspension-cultured tobacco BY-2 cells

View ORCID ProfileTomohiro Imamura, Noriyoshi Isozumi, Yasuki Higashimura, Shinya Ohki, Masashi Mori
doi: https://doi.org/10.1101/2020.10.07.325910
Tomohiro Imamura
1Research Institute for Bioresources and Biotechnology, Ishikawa Prefectural University, 308-1 Suematsu, Nonoichi, Ishikawa 921-8836, Japan
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  • ORCID record for Tomohiro Imamura
Noriyoshi Isozumi
2Center for Nano Materials and Technology (CNMT), Japan Advanced Institute of Science and Technology (JAIST), 1-1 Asahidai, Nomi, Ishikawa 923-1292, Japan
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Yasuki Higashimura
3Department of Food Science, Ishikawa Prefectural University, 308-1 Suematsu, Nonoichi, Ishikawa 921-8836, Japan
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Shinya Ohki
2Center for Nano Materials and Technology (CNMT), Japan Advanced Institute of Science and Technology (JAIST), 1-1 Asahidai, Nomi, Ishikawa 923-1292, Japan
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Masashi Mori
1Research Institute for Bioresources and Biotechnology, Ishikawa Prefectural University, 308-1 Suematsu, Nonoichi, Ishikawa 921-8836, Japan
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  • For correspondence: mori@ishikawa-pu.ac.jp
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Abstract

COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which spread worldwide in 2020, is an urgent problem to be overcome. The ORF8 of SARS-CoV-2 has been suggested to be associated with the symptoms of COVID-19, according to reports of clinical studies. However, little is known about the function of ORF8. As one of the ways to advance the functional analysis of ORF8, mass production of ORF8 with the correct three-dimensional structure is necessary. In this study, we attempted to produce ORF8 protein by chemically inducible protein production system using tobacco BY-2 cells. An ORF8-producing line was generated by the Agrobacterium method. As a result, the production of ORF8 of 8.8 ± 1.4 mg/L of culture medium was confirmed. SDS-PAGE and nuclear magnetic resonance (NMR) analysis confirmed that the ORF8 produced by this system is a dimeric form with a single structure, unlike that produced in Escherichia coli. Furthermore, it was suggested that the ORF8 produced by this system was glycosylated. Through this study, we succeeded in producing ORF8 folded into a single structure in a chemically inducible protein production system using tobacco BY-2 cells. It is expected that the functional analysis of ORF8 will be advanced using the ORF8 produced by this system and that it will greatly contribute to the development of antibodies and therapeutic agents targeting ORF8.

Competing Interest Statement

The authors have declared no competing interest.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted December 07, 2020.
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Production of ORF8 protein from SARS-CoV-2 using an inducible virus-mediated expression system in suspension-cultured tobacco BY-2 cells
Tomohiro Imamura, Noriyoshi Isozumi, Yasuki Higashimura, Shinya Ohki, Masashi Mori
bioRxiv 2020.10.07.325910; doi: https://doi.org/10.1101/2020.10.07.325910
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Production of ORF8 protein from SARS-CoV-2 using an inducible virus-mediated expression system in suspension-cultured tobacco BY-2 cells
Tomohiro Imamura, Noriyoshi Isozumi, Yasuki Higashimura, Shinya Ohki, Masashi Mori
bioRxiv 2020.10.07.325910; doi: https://doi.org/10.1101/2020.10.07.325910

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