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Characterization of Single-Chain Fv Fragments of Neutralizing Antibodies to Rabies Virus Glycoprotein
| Title: | Characterization of Single-Chain Fv Fragments of Neutralizing Antibodies to Rabies Virus Glycoprotein |
| Authors: | Yumoto, Kohei Browse this author | | Arisaka, Tomoaki Browse this author | | Okada, Kazuma Browse this author | | Aoki, Kyosuke Browse this author | | Ose, Toyoyuki Browse this author →KAKEN DB | | Masatani, Tatsunori Browse this author →KAKEN DB | | Sugiyama, Makoto Browse this author →KAKEN DB | | Ito, Naoto Browse this author →KAKEN DB | | Fukuhara, Hideo Browse this author →KAKEN DB | | Maenaka, Katsumi Browse this author →KAKEN DB |
| Keywords: | rabies virus | | rabies virus glycoprotein | | single-chain Fv | | Fab | | neutralization antibody | | biolayer interferometry (BLI) | | differential scanning fluorometry (DSF) |
| Issue Date: | 19-Nov-2021 |
| Publisher: | MDPI |
| Journal Title: | Viruses-Basel |
| Volume: | 13 |
| Issue: | 11 |
| Start Page: | 2311 |
| Publisher DOI: | 10.3390/v13112311 |
| Abstract: | Rabies has almost a 100% case-fatality rate and kills more than 59,000 people annually around the world. There is no established treatment for rabies. The rabies virus (RABV) expresses only the glycoprotein (RABVG) at the viral surface, and it is the target for the neutralizing antibodies. We previously established mouse monoclonal antibodies, 15-13 and 12-22, which showed neutralizing activity against the RABV, targeting the sequential and conformational epitopes on the RABVG, respectively. However, the molecular basis for the neutralizing activity of these antibodies is not yet fully understood. In this study, we evaluated the binding characteristics of the Fab fragments of the 15-13 and 12-22 antibodies. The recombinant RABVG protein, in prefusion form for the binding analysis, was prepared by the silkworm-baculovirus expression system. Biolayer interferometry (BLI) analysis indicated that the 15-13 Fab interacts with the RABVG, with a K-D value at the nM level, and that the 12-22 Fab has a weaker binding affinity (K-D ～ μM) with the RABVG compared to the 15-13 Fab. Furthermore, we determined the amino acid sequences of both the antibodies and the designed single-chain Fv fragments (scFvs) of the 15-13 and 12-22 antibodies as another potential biopharmaceutical for targeting rabies. The 15-13 and 12-22 scFvs were successfully prepared by the refolding method and were shown to interact with the RABVG at the nM level and the μM level of the K-D, respectively. These binding characteristics were similar to that of each Fab. On the other hand, differential scanning fluorometry (DSF) revealed that the thermal stability of these scFvs decreases compared to their Fabs. While the improvement of the stability of scFvs will still be required, these results provide insights into the neutralizing activity and the potential therapeutic use of antibody fragments for RABV infection. |
| Type: | article |
| URI: | http://hdl.handle.net/2115/83836 |
| Appears in Collections: | 国際連携研究教育局 : GI-CoRE (Global Institution for Collaborative Research and Education : GI-CoRE) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)
薬学研究院 (Faculty of Pharmaceutical Sciences) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)
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