Biophysical investigation of interactions between SARS‐CoV‐2 spike protein and neuropilin‐1

Abstract Recent studies have suggested that neuropilin‐1 (NRP1) may serve as a potential receptor in severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) infection. However, the biophysical characteristics of interactions between NRP1 and SARS‐CoV‐2 remain unclear. In this study, we examined the interactions between NRP1 and various SARS‐CoV‐2 spike (S) fragments, including the receptor‐binding domain (RBD) and the S protein trimer in a soluble form or expressed on pseudovirions, using atomic force microscopy and structural modeling. Our measurements shows that NRP1 interacts with the RBD and trimer at a higher binding frequency (BF) compared to ACE2. This NRP1‐RBD interaction has also been predicted and simulated via AlphaFold2 and molecular dynamics simulations, and the results indicate that their binding patterns are very similar to RBD‐ACE2 interactions. Additionally, under similar loading rates, the most probable unbinding forces between NRP1 and S trimer (both soluble form and on pseudovirions) are larger than the forces between NRP1 and RBD and between trimer and ACE2. Further analysis indicates that NRP1 has a stronger binding affinity to the SARS‐CoV‐2 S trimer with a dissociation rate of 0.87 s −1 , four times lower than the dissociation rate of 3.65 s −1 between NRP1 and RBD. Moreover, additional experiments show that RBD‐neutralizing antibodies can significantly reduce the BF for both ACE2 and NRP1. Together, the study suggests that NRP1 can be an alternative receptor for SARS‐CoV‐2 attachment to human cells, and the neutralizing antibodies targeting SARS‐CoV‐2 RBD can reduce the binding between SARS‐CoV‐2 and NRP1.