Critical Point Mutations in the RBD of SARS-COV-2 Involved in Binding to ACE2

Abstract

The spike protein of SARS-CoV-2 plays an essential role in viral pathogenesis. It binds to human cells' angiotensin-converting enzyme 2 (ACE2) receptor through the receptor-binding domain (RBD), mediating virus entry into the human host cell. The genomic changes of SARS-CoV-2 can affect its pathogenic potential, making the development of treatments and vaccines more challenging. The virus accumulates mutations to evade immune response while preserving or enhancing the binding feature to ACE2. In this study, we aimed to identify mutations in the RBD region of the spike gene from SARS-CoV-2 RNA samples taken from infected patients. We use two-step RT-PCR (cDNA synthesis followed by separate PCR amplification) reactions to amplify the RBD sequence. We aligned the sequencing data with the reference RBD sequence of the Wuhan-Hu-1 (wild-type virus) to identify the different mutations. In addition, further bioinformatic analyses were performed to evaluate the impact of the mutation(s) on the spike protein, including the prediction of protein structure and its binding affinity to ACE2. Our results show that the substitutions Y421I, Q493H, S494P, and F497S may decrease the stability of the spike protein due to the different physicochemical properties of the substituted amino acids, affecting their interactions with other RBD amino acids. Moreover, molecular docking results indicate that Q493H and S494P substitutions cause an increased binding affinity of spike protein to ACE2. At the same time, Y421I and F497S substitutions decrease the binding potential of these two proteins. Based on our findings, new mutations can cause the emergence of dangerous strains so continuous monitoring of the virus and ongoing research is crucial to prevent the spread of new, potentially dangerous strains.