After successfully reporting the three-dimensional structure of full-length ACE2, the receptor of SARS-CoV-2, and full-length ACE2 in the complex with the receptor-binding domain (RBD) of the SARS-CoV-2 S protein, Qiang Zhou’s lab at Westlake University has again made a breakthrough in the research of the SARS-CoV-2 infection.

Qiang Zhou's team employed Cryo-EM to determine the structure of the S protein of SARS-CoV-2 and the neutralizing antibody 4A8 complex. 4A8 is one of the antibodies from the immune cells of recovered patients. These antibodies were screened and inspected by their collaborators, Chen Wei's group from the Beijing Institute of Biotechnology. 4A8 recognizes the N-terminal domain (NTD) of the S protein and shows high SARS-CoV-2 virus-neutralizing capabilities. This work has laid the foundation for drug designs and treatment strategies that target the NTD of the S protein.

The research was published on the preprint platform bioRxiv on May 9 (Beijing time).

In this project, the researchers at Chen Wei's group isolated selected antibodies from the memory B cells and plasma cells of recovered COVID-19 patients and obtained 399 monoclonal antibodies. After testing the affinities between antigens and antibodies in vitro and detecting the neutralization with the SARS-CoV-2 virus and the pseudovirus (viruses without the ability to replicate, but can infect cells), they discovered that 4A8 possess strong virus-neutralizing capabilities which means 4A8 could effectively suppress virus infection.

Subsequently, Qiang Zhou’s team employed Cryo-EM to determine the structure of the S protein of SARS-CoV-2 and the neutralizing antibodies 4A8 complex. The interaction interface has been clearly visible.

Illustration: (left) a Cryo-EM density map of the S protein of SARS-CoV-2 and the monoclonal antibodies 4A8 compounds, and a structural image)

The research has shown that 4A8 can bind to the NTD of the SARS-CoV-2 S protein through its heavy-chain. Researchers analyzed three flexible parts of the NTD of the complex (areas where the shape can easilychange). In previous studies of the S protein of SARS-CoV-2, these parts had been invisible because of the flexibility. Now, the structure of these parts revealed that the variable regions of the neutralizing antibody 4A8 interacted with two of the flexible parts, which can stabilize these parts.

The S protein exists in the form of homo-trimers, containing three protomers. The NTD of every S protein protomer has the ability to bind to 4A8, whereas there is no difference in each interaction interface. The binding site of the antibodies does not overlap with RBD and does not block the interaction between ACE2 and the S protein.

Illustration: Density map showing the interaction interface between the N-terminal domain (NTD) of the S protein of SARS-CoV-2 and the 4A8 heavy chain.

Previous researches focused on SARS-CoV-2, antibody designs and drug designs, and were mainly targeted at the RBD area of the SARS-CoV-2 S proteins There have been few treatment strategies aimed at other areas of the S protein.

Understanding the complex structure of 4A8 antibodies and the SARS-CoV-2 S-proteins has laid the foundation for drug designs and treatment strategies aimed at the NTD of SARS-CoV-2. Besides, 4A8 acts as a neutralizing antibody with a strong ability to suppress the virus infection and can be applied combined with antibodies to target other areas of the S protein of SARS-CoV-2, thus improving the chances of a successful treatment.

Co-first author of this study is postdoctoral researcher Renhong Yan at the Westlake University School of Life Sciences. Corresponding author is Qiang Zhou, distinguished researcher at the Westlake University School of Life Sciences.