An understanding of the interaction between the antibody and its targeted antigen and knowing of the epitopes are critical for the development of monoclonal antibody drugs. Complement factor H (CFH) is implied to play a role in tumor growth and metastasis. An autoantibody to CHF is associated with anti-tumor cell activity. The interaction of a human monoclonal antibody Ab42 that was isolated from a cancer patient with CFH polypeptide (pCFH) antigen was analyzed by molecular docking, molecular dynamics (MD) simulation, free energy calculation, and computational alanine scanning (CAS). Experimental alanine scanning (EAS) was then carried out to verify the results of the theoretical calculation. Our results demonstrated that the Ab42 antibody interacts with pCFH by hydrogen bonds through the Tyr315, Ser100, Gly33, and Tyr53 residues on the complementarity-determining regions (CDRs), respectively, with the amino acid residues of Pro441, Ile442, Asp443, Asn444, Ile447, and Thr448 on the pCFH antigen. In conclusion, this study has explored the mechanism of interaction between Ab42 antibody and its targeted antigen by both theoretical and experimental analysis. Our results have important theoretical significance for the design and development of relevant antibody drugs.

SARS-CoV-2 is highly homologous to SARS-CoV. To date, the main protease (Mpro) of SARS-CoV-2 is regarded as an important drug target for the treatment of Coronavirus Disease 2019 (COVID-19). Some experiments confirmed that several HIV protease inhibitors present the inhibitory effects on the replication of SARS-CoV-2 by inhibiting Mpro. However, the mechanism of action has still not been studied very clearly. In this work, the interaction mechanism of four HIV protease inhibitors Darunavir (DRV), Lopinavir (LPV), Nelfinavir (NFV), and Ritonavire (RTV) targeting SARS-CoV-2 Mpro was explored by applying docking, molecular dynamics (MD) simulations, and MM–GBSA methods using the broad-spectrum antiviral drug Ribavirin (RBV) as the negative and nonspecific control. Our results revealed that LPV, RTV, and NFV have higher binding affinities with Mpro, and they all interact with catalytic residues His41 and the other two key amino acids Met49 and Met165. Pharmacophore model analysis further revealed that the aromatic ring, hydrogen bond donor, and hydrophobic group are the essential infrastructure of Mpro inhibitors. Overall, this study applied computational simulation methods to study the interaction mechanism of HIV-1 protease inhibitors with SARS-CoV-2 Mpro, and the findings provide useful insights for the development of novel anti-SARS-CoV-2 agents for the treatment of COVID-19.

1-17-2 is a rat anti-human DEC-205 monoclonal antibody that induces internalization and delivers antigen to dendritic cells (DCs). The potentially clinical application of this antibody is limited by its murine origin. Traditional humanization method such as complementarity determining regions (CDRs) graft often leads to a decreased or even lost affinity. Here we have developed a novel antibody humanization method based on computer modeling and bioinformatics analysis. First, we used homology modeling technology to build the precise model of Fab. A novel epitope scanning algorithm was designed to identify antigenic residues in the framework regions (FRs) that need to be mutated to human counterpart in the humanization process. Then virtual mutation and molecular dynamics (MD) simulation were used to assess the conformational impact imposed by all the mutations. By comparing the root-mean-square deviations (RMSDs) of CDRs, we found five key residues whose mutations would destroy the original conformation of CDRs. These residues need to be back-mutated to rescue the antibody binding affinity. Finally we constructed the antibodies in vitro and compared their binding affinity by flow cytometry and surface plasmon resonance (SPR) assay. The binding affinity of the refined humanized antibody was similar to that of the original rat antibody. Our results have established a novel method based on epitopes scanning and MD simulation for antibody humanization.

The COVID-19 epidemic is raging around the world. Neutralizing antibodies are powerful tools for the prevention and treatment of SARS-CoV-2 infection. Antibody CR3022, a SARS-CoV neutralizing antibody, was found to cross-react with SARS-CoV-2, but its affinity was lower than that of its binding with SARS-CoV, which greatly limited the further development of CR3022 against SARS-CoV-2. Therefore, it is necessary to improve its affinity to SARS-CoV-2 in vitro. In this study, the structure-based molecular simulations were utilized to virtually mutate the possible key residues in the complementarity-determining regions (CDRs) of the CR3022 antibody. According to the criteria of mutation energy, the mutation sites that have the potential to impact the antibody affinity were then selected. Then optimized CR3022 mutants with the enhanced affinity were further identified and verified by enzyme-linked immunosorbent assay (ELISA), surface plasma resonance (SPR) and autoimmune reactivity experiments. Finally, molecular dynamics (MD) simulation and binding free energy calculation (MM/PBSA) were performed on the wild-type CR3022 and its two double-site mutants to understand in more detail the contribution of these sites to the higher affinity. It was found that the binding affinity of the CR3022 antibody could be significantly enhanced more than ten times after the introduction of the S103F/Y mutation in HCDR–3 and the S33R mutation in LCDR–1. The additional hydrogen-bonding, hydrophobic interactions, as well as salt-bridges formed between the modified double-site mutated antibody and SARS-CoV-2 RBD were identified. The computational and experimental results clearly demonstrated that the affinity of the modified antibody has been greatly enhanced. This study indicates that CR3022 as a neutralizing antibody recognizing the conserved region of RBD against SARS-CoV with cross-reactivity with SARS-CoV-2, a different member in a large family of coronaviruses, could be improved by the computational and experimental approaches which provided insights for developing antibody drugs against SARS-CoV-2.

Aims & Objective: Coronavirus Disease 2019 (COVID-19) caused by the human coronavirus 2019 (HCoV-19, also known as SARS-CoV-2) infection is currently in a global outbreak. COVID-19 has posed a huge threat to public health and economic stability worldwide. CR3022, a human monoclonal neutralizing antibody isolated from a Severe Acute Respiratory Syndrome (SARS) recovery patient, was confirmed to be able to bind the S protein of HCoV-19 with a certain degree of neutralizing activity. Crystal structural information indicated that CR3022 could bind to the epitope on the receptor binding domain (RBD) of HCoV-19, whose epitope consists of 28 amino acids, and 24 of them are conserved in SARS-CoV of SARS. However, the crystal structure is only a static conformation at a certain moment in time, and it cannot provide dynamic details of the interaction between antigen and antibody. Methods: In this study, molecular dynamics (MD) simulation combined with MM/PBSA and CAS methods were performed to investigate the mechanism of binding of CR3022 against SARS-CoVRBD and HCoV-19-RBD in order to determine their holographic dynamic information. Results: It was found that the CR3022-SARS-CoV-RBD complex was more stable during 100ns MD run than that of the CR3022-HCoV-19-RBD system. There were common conservative amino acids on the β2 sheet of RBD, including Tyr369, Phe377, Lys378, Tyr380, Gly381, Lys386, Leu390 and others. These conservative amino acids play significant roles in the binding process of CR3022 antibody against SARS-CoV-RBD and HCoV-19-RBD. It was also found that the binding mode of CR3022 to its native target SARS-CoV-RBD is more comprehensive and uniform. Moreover, the β2 sheet residue Thr385 and non-β2 sheet residues Arg408 and Asp428 of the CR3022-SARS-CoV-RBD system were found to be crucial for their binding affinities, thus forming a special conformational epitope. However, these key amino acids are not present in the CR3022-HCoV-19-RBD system. The binding mode of CR3022 and HCoV-19-RBD is similar to that of SARS-CoV-RBD, but the deficiency of crucial hydrogen-bonds and salt-bridges. Therefore, the binding of CR3022 and HCoV-19-RBD only draws on the partial mode of the binding of CR3022 and SARS-CoV-RBD, so there is a loss of affinity. Conclusion: Thus, in order to better fight the epidemic of COVID-19 with the CR3022 antibody, this antibody needs to further improve the neutralization efficiency of HCoV-19 through mutation of it’s CDR region.

Protein folding is an important and yet challenging topic in current molecular biology. In this work, the folding dynamics and mechanisms of the Trp-cage mini-protein were studied with molecular dynamics simulations, in the absence and presence of water solvents. The important intermediates during the Trp-cage folding were determined by gradually decreasing the simulation temperature. The folding transition temperature was identified to be approximately 400 K, and the folding pathway was decomposed into six steps: U ↔ I ₁ ↔ I ₂ ↔ I ₃ ↔ I ₄ ↔ F ₁ ↔ F ₂, where U, I and F represent the unfolded, intermediate and folded states, respectively. The finding that the two helical subunits are successively formed is consistent with the experimental observations, and the Asp9/Arg16 salt bridge forms at the final stage and does not play a significant role during folding kinetics. The presence of water solvents induces hydrophobic collapse as the whole cage comparatively closes. Within aqueous solutions, the Trp-cage folding begins to contract into the meta-stable state, and by traversing the transition state it arrives at the native-like structure, which resembles the experimental structure closely.