The systems for exploring the interaction between human angiotensin converting enzyme 2 (ACE2) and spike protein of SARS-CoV-2

Xin-Rui Yang (楊欣瑞) 1, Cheng-Han Lin (林承翰) 1, Ho-Ju Chang1, Jia-Rong Chang1, Yi-Ju Chen, Chih-Sheng Lin1

1 Department of Biological Science and Technology, National Yang Ming Chiao Tung University

In recent three years, SARS-CoV-2 has spread rapidly around the world, causing the collapse of medical systems around global world. The spike protein of SARS-CoV-2 and ACE2 on human cell membranes (hACE2) play a critical role in severe epidemics. The spike (S) protein binds to hACE2 to infected and damaged lung cells. Since the end of 2020, a series of mutations have made SARS-CoV-2 unpredictable. Numerous studies have shown that the neutralization of SARS-CoV-2 variants with serum by vaccine is gradually reduced. Therefore, this study aims to construct a system to explore the interaction between viral S protein and hACE2. In vitro study, HEK293T-hACE2 cell line was used. The packaging system of Lentiviral-S pseudovirus was established to express the SARS-CoV-2 S protein. The lentiviral transfer vector DNA with the reporter EGFP gene, together with packaging and envelope plasmid DNA were combined at a ratio of 10:9:1, respectively. In addition, in order to understand the effect of virus mutation on infection efficiency, we used various pseudoviral mutants for comparison, including Wuhan, Alpha, Beta, Delta, and Omicron BA.1, BA.2 and BA.4/5 strains. The experimental results show that the fluorescence intensity of BA.2 and BA.4/5 was significantly increased. It is indicated highly infective efficiency in the Omicron mutants. The drugs to block the interaction of S protein and hACE2 to reduce the efficiency of viral infection were performed. From the results, an effective in vitro system for the drugs screening, that are potential to reduce the SARS-CoV-2 infection, would be confirmed. In vivo study, the mouse with genetic modifications of hACE2(+)/mACE2(-) [hACE2 transgenic and mACE knock out] has been successfully bred. Our preliminary results show that the hACE2(+)/mACE2(-) mice could be effectively infected by SARS-CoV-2 pseudovirus and adaptably used to detect the drug efficacy of reducing lung infection by the pseudovirus.