Abstract
Broadly effective and safe anti-coronavirus agent is existentially needed. Major protease (3CL pro ) is a highly conserved enzyme of betacoronaviruses. The enzyme plays pivotal role in the virus replication cycle. Thus, it is a good target of a broadly effective anti- Betacoronavirus agent. In this study, human single-chain antibodies (HuscFvs) of the SARS-CoV-2 3CL pro were generated using phage display technology. The 3CL pro -bound phages were used to infect Escherichia coli host for the production the 3CL pro -bound HuscFvs. Computerized simulation was used to guide the selection of the phage infected- E. coli clones that produced HuscFvs with the 3CL pro inhibitory potential. HuscFvs of three phage infected- E. coli clones were predicted to form contact interface with residues for 3CL pro catalytic activity, substrate binding, and homodimerization. These HuscFvs were linked to a cell-penetrating peptide to make them cell-penetrable, i.e., became superantibodies. The superantibodies blocked the 3CL pro activity in vitro, were not toxic to human cells, traversed across membrane of 3CL pro -expressing cells to co-localize with the intracellular 3CL pro and most of all, they inhibited replication of authentic SARS-CoV-2 Wuhan wild type and α, β, δ, and Omicron variants that were tested. The superantibodies should be investigated further towards clinical application as a safe and broadly effective anti- Betacoronavirus agent.
Keywords: SARS-CoV-2; cell-penetrating antibody; human single-chain antibody variable fragments (HuscFvs); major protease (3CLpro); superantibody.
【저자키워드】 SARS-CoV-2, cell-penetrating antibody, human single-chain antibody variable fragments (HuscFvs), major protease (3CLpro), superantibody., 【초록키워드】 variant, peptide, 3CL pro, 3CLpro, protease, in vitro, omicron, Omicron variants, Betacoronavirus, Replication, Wuhan, membrane, virus replication, wild type, binding, betacoronaviruses, Contact, Escherichia coli, Safe, Major, E. coli, enzyme, residue, human cells, single-chain antibody, residues, clone, phage, catalytic activity, inhibitory, infect, Host, effective, Cell, produced, tested, predicted, blocked, was used, conserved, investigated, inhibited, were used, the SARS-CoV-2, 【제목키워드】 Human,