{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,18]],"date-time":"2026-08-18T04:34:23Z","timestamp":1787027663554,"version":"build-2736575974"},"reference-count":38,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2023,10,11]],"date-time":"2023-10-11T00:00:00Z","timestamp":1696982400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Research Foundation of Korea","award":["2021RIS-001"],"award-info":[{"award-number":["2021RIS-001"]}]},{"name":"National Research Foundation of Korea","award":["2017R1D1A1B03032185"],"award-info":[{"award-number":["2017R1D1A1B03032185"]}]},{"name":"National Research Foundation of Korea","award":["2020R1I1A3071938"],"award-info":[{"award-number":["2020R1I1A3071938"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biomolecules"],"abstract":"<jats:p>Although single-chain variable fragment (scFv) is recognized as a highly versatile scaffold of recombinant antibody fragment molecules, its overexpression in Escherichia coli often leads to the formation of inclusion bodies. To address this issue, we devised and tested four different constructs, named v21, v22, v23 and v24, for producing anti-human epidermal growth factor receptor 2 (HER2) scFv. Among them, the v24 construct obtained from N-terminal fusion of maltose-binding protein (MBP) and subsequent tobacco etch virus protease (TEV) was identified as the most efficient construct for the production of anti-HER2 scFv. Aided by an MBP tag, high-yield soluble expression was ensured and soluble scFv was liberated in cells via autonomous proteolytic cleavage by endogenously expressed TEV. The isolated scFv containing a C-terminal hexahistidine tag was purified through a one-step purification via nickel-affinity chromatography. The purified scFv exhibited a strong (nanomolar Kd) affinity to HER2 both in vitro and in cells. Structural and functional stabilities of the scFv during storage for more than one month were also assured. Given the great utility of anti-HER2 scFv as a basic platform for developing therapeutic and diagnostic agents for cancers, the v24 construct and methods presented in this study are expected to provide a better manufacturing system for producing anti-HER2 scFv with various industrial applications.<\/jats:p>","DOI":"10.3390\/biom13101508","type":"journal-article","created":{"date-parts":[[2023,10,12]],"date-time":"2023-10-12T03:07:47Z","timestamp":1697080067000},"page":"1508","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":15,"title":["Development of an Anti-HER2 Single-Chain Variable Antibody Fragment Construct for High-Yield Soluble Expression in Escherichia coli and One-Step Chromatographic Purification"],"prefix":"10.3390","volume":"13","author":[{"given":"Kyu Tae","family":"Byun","sequence":"first","affiliation":[{"name":"Department of Biotechnology, Research Institute (RIBHS), College of Biomedical and Health Science, Konkuk University, Chungju 27478, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Boram","family":"Kim","sequence":"additional","affiliation":[{"name":"Department of Biotechnology, Research Institute (RIBHS), College of Biomedical and Health Science, Konkuk University, Chungju 27478, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Junmin","family":"Cho","sequence":"additional","affiliation":[{"name":"Department of Biotechnology, Research Institute (RIBHS), College of Biomedical and Health Science, Konkuk University, Chungju 27478, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Inbeom","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Biotechnology, Research Institute (RIBHS), College of Biomedical and Health Science, Konkuk University, Chungju 27478, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Myung Gu","family":"Lee","sequence":"additional","affiliation":[{"name":"Konkukbio Inc., Konkuk University, Chungju 27478, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dongsun","family":"Park","sequence":"additional","affiliation":[{"name":"Department of Biology Education, Korea National University of Education, Cheongju 28173, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1441-9470","authenticated-orcid":false,"given":"Tae-Bong","family":"Kang","sequence":"additional","affiliation":[{"name":"Department of Biotechnology, Research Institute (RIBHS), College of Biomedical and Health Science, Konkuk University, Chungju 27478, Republic of Korea"},{"name":"BK21 Project Team, Department of Applied Life Science, Graduate School, Konkuk University, Chungju 27478, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hyung-Sik","family":"Won","sequence":"additional","affiliation":[{"name":"Department of Biotechnology, Research Institute (RIBHS), College of Biomedical and Health Science, Konkuk University, Chungju 27478, Republic of Korea"},{"name":"BK21 Project Team, Department of Applied Life Science, Graduate School, Konkuk University, Chungju 27478, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6045-6868","authenticated-orcid":false,"given":"Chan Gil","family":"Kim","sequence":"additional","affiliation":[{"name":"Department of Biotechnology, Research Institute (RIBHS), College of Biomedical and Health Science, Konkuk University, Chungju 27478, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,10,11]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"143","DOI":"10.1016\/bs.apcsb.2018.03.002","article-title":"Chimeric Small Antibody Fragments as Strategy to Deliver Therapeutic Payloads","volume":"112","author":"Aguiar","year":"2018","journal-title":"Adv. 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