Recombinant Protein Expression and Purification of N, S1, and RBD of SARS-CoV-2 from Mammalian Cells and Their Potential Applications

SARS-CoV-2 N、S1 和 RBD 的哺乳动物细胞重组蛋白表达、纯化及其潜在应用

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作者:Julio García-Cordero, Juvenal Mendoza-Ramírez, David Fernández-Benavides, Daniela Roa-Velazquez, Jessica Filisola-Villaseñor, Sandra Paola Martínez-Frías, Erik Saul Sanchez-Salguero, Carlos E Miguel-Rodríguez, Jose L Maravillas Montero, Jose J Torres-Ruiz, Diana Gómez-Martín, Leopoldo Santos Argumed

Abstract

The coronavirus disease 2019 (COVID-19) pandemic has reached an unprecedented level. There is a strong demand for diagnostic and serological supplies worldwide, making it necessary for countries to establish their own technologies to produce high-quality biomolecules. The two main viral antigens used for the diagnostics for severe acute respiratory syndrome coronavirus (SARS-CoV-2) are the structural proteins spike (S) protein and nucleocapsid (N) protein. The spike protein of SARS-CoV-2 is cleaved into S1 and S2, in which the S1 subunit has the receptor-binding domain (RBD), which induces the production of neutralizing antibodies, whereas nucleocapsid is an ideal target for viral antigen-based detection. In this study, we designed plasmids, pcDNA3.1/S1 and pcDNA3.1/N, and optimized their expression of the recombinant S1 and N proteins from SARS-CoV-2 in a mammalian system. The RBD was used as a control. The antigens were successfully purified from Expi293 cells, with high yields of the S1, N, and RBD proteins. The immunogenic abilities of these proteins were demonstrated in a mouse model. Further, enzyme-linked immunosorbent assays with human serum samples showed that the SARS-CoV-2 antigens are a suitable alternative for serological assays to identify patients infected with COVID-19.

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