{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T04:43:39Z","timestamp":1760244219127,"version":"build-2065373602"},"reference-count":39,"publisher":"MDPI AG","issue":"11","license":[{"start":{"date-parts":[[2022,11,7]],"date-time":"2022-11-07T00:00:00Z","timestamp":1667779200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Life"],"abstract":"<jats:p>The production of recombinant proteins in Escherichia coli cells is often hampered by aggregation of newly synthesized proteins and formation of inclusion bodies. Here we propose the use of transverse urea gradient electrophoresis (TUGE) in testing the capability of folding of a recombinant protein from inclusion bodies dissolved in urea. A plasmid encoding the amino acid sequence 55\u2013224 of TcpA pilin (C-terminal globular domain: TcpA-C) from Vibrio cholerae El Tor enlarged by a His-tag on its N-terminus was expressed in E. coli cells. The major fraction (about 90%) of the target polypeptide was detected in cell debris. The polypeptide was isolated from the soluble fraction and recovered from inclusion bodies after their urea treatment. Some structural properties of the polypeptide from each sample proved identical. The refolding protocol was developed on the basis of TUGE data and successfully used for the protein large-scale recovery from inclusion bodies. Spectral, hydrodynamic, and thermodynamic characteristics of the recombinant TcpA recovered from inclusion bodies indicate the presence of a globular conformation with a pronounced secondary structure and a rigid tertiary structure, which is promising for the design of immunodiagnostics preparations aimed to assess the pilin level in different strains of V. cholerae and to develop cholera vaccines.<\/jats:p>","DOI":"10.3390\/life12111802","type":"journal-article","created":{"date-parts":[[2022,11,8]],"date-time":"2022-11-08T12:11:00Z","timestamp":1667909460000},"page":"1802","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Recombinant Globular Domain of TcpA Pilin from Vibrio cholerae El Tor: Recovery from Inclusion Bodies and Structural Characterization"],"prefix":"10.3390","volume":"12","author":[{"given":"Victor","family":"Marchenkov","sequence":"first","affiliation":[{"name":"Institute of Protein Research, Russian Academy of Sciences, Institutskaya Street 4, 142290 Pushchino, Russia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Elena","family":"Dubovitskya","sequence":"additional","affiliation":[{"name":"Russian Antiplague Scientific Research Institute \u201cMicrobe\u201d of Rospotrebnadzor, Universitetskaya Street 46, 410005 Saratov, Russia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nina","family":"Kotova","sequence":"additional","affiliation":[{"name":"Russian Antiplague Scientific Research Institute \u201cMicrobe\u201d of Rospotrebnadzor, Universitetskaya Street 46, 410005 Saratov, Russia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Igor","family":"Tuchkov","sequence":"additional","affiliation":[{"name":"Russian Antiplague Scientific Research Institute \u201cMicrobe\u201d of Rospotrebnadzor, Universitetskaya Street 46, 410005 Saratov, Russia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nina","family":"Smirnova","sequence":"additional","affiliation":[{"name":"Russian Antiplague Scientific Research Institute \u201cMicrobe\u201d of Rospotrebnadzor, Universitetskaya Street 46, 410005 Saratov, Russia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2967-4677","authenticated-orcid":false,"given":"Natalia","family":"Marchenko","sequence":"additional","affiliation":[{"name":"Institute of Protein Research, Russian Academy of Sciences, Institutskaya Street 4, 142290 Pushchino, Russia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alexey","family":"Surin","sequence":"additional","affiliation":[{"name":"Institute of Protein Research, Russian Academy of Sciences, Institutskaya Street 4, 142290 Pushchino, Russia"},{"name":"Branch of the Shemyakin\u2013Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Prospekt Nauki 6, 142290 Pushchino, Russia"},{"name":"State Research Centre for Applied Microbiology and Biotechnology, 142279 Obolensk, Russia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Vladimir","family":"Filimonov","sequence":"additional","affiliation":[{"name":"Institute of Protein Research, Russian Academy of Sciences, Institutskaya Street 4, 142290 Pushchino, Russia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4311-3884","authenticated-orcid":false,"given":"Gennady","family":"Semisotnov","sequence":"additional","affiliation":[{"name":"Institute of Protein Research, Russian Academy of Sciences, Institutskaya Street 4, 142290 Pushchino, Russia"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2022,11,7]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"48","DOI":"10.1128\/CMR.8.1.48","article-title":"Cholera","volume":"8","author":"Kaper","year":"1995","journal-title":"Clin. 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