{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,24]],"date-time":"2026-03-24T20:08:55Z","timestamp":1774382935028,"version":"3.50.1"},"reference-count":29,"publisher":"MDPI AG","issue":"11","license":[{"start":{"date-parts":[[2021,11,19]],"date-time":"2021-11-19T00:00:00Z","timestamp":1637280000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100009619","name":"Japan Agency for Medical Research and Development","doi-asserted-by":"publisher","award":["JP20am0101093"],"award-info":[{"award-number":["JP20am0101093"]}],"id":[{"id":"10.13039\/100009619","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100009619","name":"Japan Agency for Medical Research and Development","doi-asserted-by":"publisher","award":["JP20ae0101047"],"award-info":[{"award-number":["JP20ae0101047"]}],"id":[{"id":"10.13039\/100009619","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001700","name":"Ministry of Education, Culture, Sports, Science and Technology","doi-asserted-by":"publisher","award":["JPMXS0420100119"],"award-info":[{"award-number":["JPMXS0420100119"]}],"id":[{"id":"10.13039\/501100001700","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001700","name":"Ministry of Education, Culture, Sports, Science and Technology","doi-asserted-by":"publisher","award":["15H02384"],"award-info":[{"award-number":["15H02384"]}],"id":[{"id":"10.13039\/501100001700","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100008373","name":"Takeda (Japan)","doi-asserted-by":"publisher","award":["research grant"],"award-info":[{"award-number":["research grant"]}],"id":[{"id":"10.13039\/100008373","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>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\u201313 and 12\u201322, 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\u201313 and 12\u201322 antibodies. The recombinant RABVG protein, in prefusion form for the binding analysis, was prepared by the silkworm\u2013baculovirus expression system. Biolayer interferometry (BLI) analysis indicated that the 15\u201313 Fab interacts with the RABVG, with a KD value at the nM level, and that the 12\u201322 Fab has a weaker binding affinity (KD ~ \u03bcM) with the RABVG compared to the 15\u201313 Fab. Furthermore, we determined the amino acid sequences of both the antibodies and the designed single-chain Fv fragments (scFvs) of the 15\u201313 and 12\u201322 antibodies as another potential biopharmaceutical for targeting rabies. The 15\u201313 and 12\u201322 scFvs were successfully prepared by the refolding method and were shown to interact with the RABVG at the nM level and the \u03bcM level of the KD, 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.<\/jats:p>","DOI":"10.3390\/v13112311","type":"journal-article","created":{"date-parts":[[2021,11,21]],"date-time":"2021-11-21T21:02:06Z","timestamp":1637528526000},"page":"2311","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["Characterization of Single-Chain Fv Fragments of Neutralizing Antibodies to Rabies Virus Glycoprotein"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-7282-0150","authenticated-orcid":false,"given":"Kohei","family":"Yumoto","sequence":"first","affiliation":[{"name":"Laboratory of Biomolecular Science, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tomoaki","family":"Arisaka","sequence":"additional","affiliation":[{"name":"Laboratory of Biomolecular Science, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7775-5984","authenticated-orcid":false,"given":"Kazuma","family":"Okada","sequence":"additional","affiliation":[{"name":"Faculty of Applied Biological Sciences, Gifu University, Gifu 501-1193, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kyosuke","family":"Aoki","sequence":"additional","affiliation":[{"name":"Laboratory of Biomolecular Science, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Toyoyuki","family":"Ose","sequence":"additional","affiliation":[{"name":"Laboratory of Biomolecular Science, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2347-4079","authenticated-orcid":false,"given":"Tatsunori","family":"Masatani","sequence":"additional","affiliation":[{"name":"Faculty of Applied Biological Sciences, Gifu University, Gifu 501-1193, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Makoto","family":"Sugiyama","sequence":"additional","affiliation":[{"name":"Faculty of Applied Biological Sciences, Gifu University, Gifu 501-1193, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0296-966X","authenticated-orcid":false,"given":"Naoto","family":"Ito","sequence":"additional","affiliation":[{"name":"Faculty of Applied Biological Sciences, Gifu University, Gifu 501-1193, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hideo","family":"Fukuhara","sequence":"additional","affiliation":[{"name":"Center for Research and Education on Drug Discovery, Hokkaido University, Sapporo 060-0812, Japan"},{"name":"Global Station for Biosurfaces and Drug Discovery, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5459-521X","authenticated-orcid":false,"given":"Katsumi","family":"Maenaka","sequence":"additional","affiliation":[{"name":"Laboratory of Biomolecular Science, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan"},{"name":"Center for Research and Education on Drug Discovery, Hokkaido University, Sapporo 060-0812, Japan"},{"name":"Global Station for Biosurfaces and Drug Discovery, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,11,19]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"754","DOI":"10.1128\/iai.16.3.754-759.1977","article-title":"Rabies virus glycoprotein. 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