ABSTRACT
The continuous mutation of SARS-CoV-2 has presented enormous challenges to global pandemic prevention and control. Recent studies have shown evidence that the genome sequence of SARS-CoV-2 nucleocapsid proteins is relatively conserved, and their biological functions are being confirmed. There is increasing evidence that the N protein will not only provide a specific diagnostic marker but also become an effective treatment target. In this study, 2G4, which specifically recognizes the N protein, was identified by screening a human phage display library. Based on the computer-guided homology modelling and molecular docking method used, the 3-D structures for the 2G4 scFv fragment (VH-linker-VL structure, with (G4S)3 as the linker peptide in the model), SARS-CoV-2 N protein and its complex were modelled and optimized with a suitable force field. The binding mode and key residues of the 2G4 and N protein interaction were predicted, and three mutant antibodies (named 2G4-M1, 2G4-M2 and 2G4-M3) with higher affinity were designed theoretically. Using directed point mutant technology, the three mutant antibodies were prepared, and their affinity was tested. Their affinity constants of approximately 0.19 nM (2G4-M1), 0.019 nM (2G4-M2) and 0.075 nM (2G4-M3) were at least one order of magnitude lower than that of the parent antibody (3 nM; 2G4, parent antibody), as determined using a biolayer interferometry (BLI) assay. It is expected that high-affinity candidates will be used for diagnosis and even as potential therapeutic drugs for the SARS-CoV-2 pandemic.
Subject(s)
COVID-19 , SARS-CoV-2 , Antibodies, Monoclonal , Antibody Affinity , Cell Surface Display Techniques , Humans , Molecular Docking Simulation , SARS-CoV-2/geneticsABSTRACT
The novel coronavirus (SARS-CoV-2) is highly infectious and can cause novel coronavirus pneumonia (COVID-19). Anti-SARSCoV-2 antibody has high safety, strong specificity, and its ability to neutralize SARS-CoV-2 is significant. It is considered to be an ideal drug for the treatment of COVID-19. In view of the core position of the SARS-CoV-2 protruding protein (S protein), the currently developed anti-SARS-CoV-2 antibodies basically target the S protein, and the recognition epitopes are mostly located in the receptor binding domain, and a few are located in the S2 subunit or S1/S2 proteolysis site. Using technologies such as single-cell sequencing, antibody libraries, and transgenic mice, polyclonal antibodies (peripheral blood sources), monoclonal antibodies, antibody fusion proteins, and single domain antibodies have all made significant progress. Multiple products...