Key residues of the receptor binding motif in the spike protein of SARS-CoV-2 that interact with ACE2 and neutralizing antibodies

Chunyan Yi,Xiaoyu Sun,Jing Ye,Meiqin Liu,Aidong Qu,Liyang Ma,Zhuo Yang,Xiao Lu,Longfei Ding,Zhiyang Ling,Yaguang Zhang,Zhengli Shi,Bing Sun,Wangpeng Gu,Jianqing Xu

doi:10.1038/s41423-020-0458-z

Abstract

Coronavirus disease 2019 (COVID-19), caused by the novel human coronavirus SARS-CoV-2, is currently a major threat to public health worldwide. The viral spike protein binds the host receptor angiotensin-converting enzyme 2 (ACE2) via the receptor-binding domain (RBD), and thus is believed to be a major target to block viral entry. Both SARS-CoV-2 and SARS-CoV share this mechanism. Here we functionally analyzed the key amino acid residues located within receptor binding motif of RBD that may interact with human ACE2 and available neutralizing antibodies. The in vivo experiments showed that immunization with either the SARS-CoV RBD or SARS-CoV-2 RBD was able to induce strong clade-specific neutralizing antibodies in mice; however, the cross-neutralizing activity was much weaker, indicating that there are distinct antigenic features in the RBDs of the two viruses. This finding was confirmed with the available neutralizing monoclonal antibodies against SARS-CoV or SARS-CoV-2. It is worth noting that a newly developed SARS-CoV-2 human antibody, HA001, was able to neutralize SARS-CoV-2, but failed to recognize SARS-CoV. Moreover, the potential epitope residues of HA001 were identified as A475 and F486 in the SARS-CoV-2 RBD, representing new binding sites for neutralizing antibodies. Overall, our study has revealed the presence of different key epitopes between SARS-CoV and SARS-CoV-2, which indicates the necessity to develop new prophylactic vaccine and antibody drugs for specific control of the COVID-19 pandemic although the available agents obtained from the SARS-CoV study are unneglectable.

Highlights

Coronavirus disease 2019 (COVID-19) is a respiratory tract infection caused by a newly emergent coronavirus, SARS-CoV-2, which was first recognized in December 2019
The P values shown in the figures and figure legends were determined using unpaired two-tailed Student’s t-tests (*P < 0.05, **P < 0.01, and ***P < 0.001; not significant (NS)). Both the SARS-CoV and SARS-CoV-2 receptor-binding domain (RBD) bind to human ACE2 (hACE2) for virus entry To confirm that the infectivity of SARS-CoV and SARS-CoV-2 is dependent on hACE2, we constructed pseudo-typed SARS-CoV and SARS-CoV-2 by the co-transfection of a plasmid encoding Envdefective luciferase-expressing HIV-1 and a plasmid expressing the full-length S protein of SARS-CoV or SARS-CoV2 into HEK293T cells
Because syncytial formation has been observed in cultured Vero E6 cells infected with SARS-CoV,[16] we sought to determine whether HEK293T cells expressing the SARS-CoV-2 S protein could fuse with HEK293T cells expressing hACE2

Summary

Introduction

Coronavirus disease 2019 (COVID-19) is a respiratory tract infection caused by a newly emergent coronavirus, SARS-CoV-2, which was first recognized in December 2019. Many groups have shown that SARS-CoV-2 utilizes the homotrimeric spike (S) glycoprotein to bind to the functional receptor human ACE2 (hACE2); this mechanism for viral entry is used by SARS-CoV.[3,4] The RBD in the S protein mediates the binding of the virus to host cells, which is a critical step for the virus to enter target cells. According to the high-resolution crystal structure information acquired far,[5,6,7] the receptor-binding motif (RBM) is the main functional motif in RBD and is composed of two regions (region 1 and region 2) that form the interface between the S protein and hACE2.8 The region outside the RBM plays an important role in maintaining the structural stability of the RBD.[9]

Methods

Results

Conclusion