Functional analysis of the receptor binding domain of SARS coronavirus S1 region and its monoclonal antibody

Abstract

Severe acute respiratory syndrome (SARS) is caused by the SARS coronavirus (CoV). The spike protein of SARS-CoV consists of S1 and S2 domains, which are responsible for virus binding and fusion, respectively. The receptor-binding domain (RBD) positioned in S1 can specifically bind to angiotensin-converting enzyme 2 (ACE2) on target cells, and ACE2 regulates the balance between vasoconstrictors and vasodilators within the heart and kidneys. Here, a recombinant fusion protein containing 193-amino acid RBD (residues 318-510) and glutathione S-transferase were prepared for binding to target cells. Additionally, monoclonal RBD antibodies were prepared to confirm RBD binding to target cells through ACE2. We first confirmed that ACE2 was expressed in various mouse cells such as heart, lungs, spleen, liver, intestine, and kidneys using a commercial ACE2 polyclonal antibody. We also confirmed that the mouse fibroblast (NIH3T3) and human embryonic kidney cell lines (HEK293) expressed ACE2. We finally demonstrated that recombinant RBD bound to ACE2 on these cells using a cellular enzymelinked immunosorbent assay and immunoassay. These results can be applied for future research to treat ACE2related diseases and SARS.

COVID-19: The Immune Responses and Clinical Therapy Candidates

“…The S1 domain (amino acids 15–680 in SARS-CoV) is responsible for virus binding to its receptor on the target cell and comprises two sub-domains: the N-terminal and the C-terminal of RBD which consists of 193 amino acids (318–510 in SARS-CoV). The S2 domain of the spike protein (amino acids 681–1255 in SARS-CoV) participates in cell membrane fusion [ 17 , 21 ]. Cellular proteases including human airway trypsin-like protease (HAT), transmembrane protease serine 2 (TMPRSS2) and cathepsin trigger cleavage of the S protein trimer in SARS-CoV, while furin makes the MERS-CoV S protein competent for fusion.…”