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Rapid, high-yield production of full-length SARS-CoV-2 spike ectodomain by transient gene expression in CHO cells.
- Source :
-
Journal of biotechnology [J Biotechnol] 2021 Jan 20; Vol. 326, pp. 21-27. Date of Electronic Publication: 2020 Dec 07. - Publication Year :
- 2021
-
Abstract
- Recombinant forms of the spike protein of SARS-CoV-2 and related viruses have proven difficult to produce with good yields in mammalian cells. Given the panoply of potential COVID-19 diagnostic tools and therapeutic candidates that require purified spike protein and its importance for ongoing SARS-CoV-2 research, we have explored new approaches for spike production and purification. Three transient gene expression methods based on PEI-mediated transfection of CHO or HEK293 cells in suspension culture in chemically-defined media were compared for rapid production of full-length SARS-CoV-2 spike ectodomain. A high-cell-density protocol using DXB11-derived CHO <superscript>BRI/55E1</superscript> cells gave substantially better yields than the other methods. Different forms of the spike ectodomain were expressed, including the wild-type SARS-CoV-2 sequence and a mutated form (to favor expression of the full-length spike ectodomain stabilized in pre-fusion conformation), with and without fusion to putative trimerization domains. An efficient two-step affinity purification method was also developed. Ultimately, we have been able to produce highly homogenous preparations of full-length spike, both monomeric and trimeric, with yields of 100-150 mg/L in the harvested medium. The speed and productivity of this method support further development of CHO-based approaches for recombinant spike protein manufacturing.<br /> (Crown Copyright © 2020. Published by Elsevier B.V. All rights reserved.)
Details
- Language :
- English
- ISSN :
- 1873-4863
- Volume :
- 326
- Database :
- MEDLINE
- Journal :
- Journal of biotechnology
- Publication Type :
- Academic Journal
- Accession number :
- 33301853
- Full Text :
- https://doi.org/10.1016/j.jbiotec.2020.12.005