Abstract SARS‐CoV‐2 is responsible for a disruptive worldwide viral pandemic, and renders a severe respiratory disease known as COVID‐19. Spike protein of SARS‐CoV‐2 mediates viral entry into host cells by binding ACE2 through the receptor‐binding domain (RBD). RBD is an important target for development of virus inhibitors, neutralizing antibodies, and vaccines. RBD expressed in mammalian cells suffers from low expression yield and high cost. E. coli is a popular host for protein expression, which has the advantage of easy scalability with low cost. However, RBD expressed by E. coli (RBD‐1) lacks the glycosylation, and its antigenic epitopes may not be sufficiently exposed. In the present study, RBD‐1 was expressed by E. coli and purified by a Ni Sepharose Fast Flow column. RBD‐1 was structurally characterized and compared with RBD expressed by the HEK293 cells (RBD‐2). The secondary structure and tertiary structure of RBD‐1 were largely maintained without glycosylation. In particular, the major β‐sheet content of RBD‐1 was almost unaltered. RBD‐1 could strongly bind ACE2 with a dissociation constant (K D ) of 2.98 × 10 –8 M. Thus, RBD‐1 was expected to apply in the vaccine development, screening drugs and virus test kit.
【저자키워드】 COVID‐19, SARS‐CoV‐2, Spike protein, RBD, purification, 【초록키워드】 ACE2, pandemic, Vaccines, Neutralizing antibodies, glycosylation, spike, drug, virus, inhibitors, viral entry, COVID‐19, SARS‐CoV‐2, Protein, binding, secondary structure, Flow, host cell, E. coli, severe respiratory disease, low expression, protein expression, receptor‐binding domain, mammalian cell, Host, responsible, lack, characterized, expressed, the vaccine, expected, purified, antigenic epitope, HEK293 cell, 【제목키워드】 SARS‐CoV‐2, Escherichia coli, receptor‐binding domain,