{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,7,30]],"date-time":"2025-07-30T10:06:35Z","timestamp":1753869995667,"version":"3.41.2"},"reference-count":47,"publisher":"Wiley","issue":"1","license":[{"start":{"date-parts":[[2024,12,26]],"date-time":"2024-12-26T00:00:00Z","timestamp":1735171200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["JP21K05386","JP23H04559","JP24K08717","JP23KJ1820"],"award-info":[{"award-number":["JP21K05386","JP23H04559","JP24K08717","JP23KJ1820"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100007807","name":"Japan Science Society","doi-asserted-by":"publisher","award":["2022\u20104052"],"award-info":[{"award-number":["2022\u20104052"]}],"id":[{"id":"10.13039\/501100007807","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Protein Science"],"published-print":{"date-parts":[[2025,1]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Antibodies and antibody mimics are extensively used in the pharmaceutical industry, where stringent safety standards are required. Implementing heat sterilization during or after the manufacturing process could help prevent contamination by viruses and bacteria. However, conventional antibodies and antibody mimics are not suitable for heat sterilization because they irreversibly denature at high temperatures. In this study, we focused on the refolding property of the cold shock protein from the hyperthermophile <jats:italic>Thermotoga maritima<\/jats:italic> (TmCSP), which denatures at elevated temperatures but regains its native structure upon re\u2010cooling. We designed and constructed a mutant library of TmCSP in which amino acid residues in its three surface loops were diversified. From the library, mutant TmCSPs that bind to each of eight target proteins were selected by phage and yeast surface display methods. We confirmed that the secondary structure and binding affinity of all the selected mutants were restored after heat treatment followed by cooling. Additionally, freeze\u2010drying did not impair their binding affinity. The crystal structure of a mutant TmCSP in complex with its target, the esterase from <jats:italic>Alicyclobacillus acidocaldarius<\/jats:italic>, revealed specific interactions between them. These results clearly demonstrate the feasibility of creating heat\u2010sterilizable antibody mimics using TmCSP as a scaffold.<\/jats:p>","DOI":"10.1002\/pro.70018","type":"journal-article","created":{"date-parts":[[2024,12,26]],"date-time":"2024-12-26T14:11:02Z","timestamp":1735222262000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Heat\u2010sterilizable antibody mimics designed on the cold shock protein scaffold from hyperthermophile <i>Thermotoga maritima<\/i>"],"prefix":"10.1002","volume":"34","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1688-7796","authenticated-orcid":false,"given":"Hiroshi","family":"Amesaka","sequence":"first","affiliation":[{"name":"Graduate School of Life and Environmental Sciences Kyoto Prefectural University  Kyoto Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Marin","family":"Tachibana","sequence":"additional","affiliation":[{"name":"Graduate School of Life and Environmental Sciences Kyoto Prefectural University  Kyoto Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Mizuho","family":"Hara","sequence":"additional","affiliation":[{"name":"Graduate School of Life and Environmental Sciences Kyoto Prefectural University  Kyoto Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shuntaro","family":"Toya","sequence":"additional","affiliation":[{"name":"Graduate School of Life and Environmental Sciences Kyoto Prefectural University  Kyoto Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Haruki","family":"Nakagawa","sequence":"additional","affiliation":[{"name":"Graduate School of Life and Environmental Sciences Kyoto Prefectural University  Kyoto Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0361-3796","authenticated-orcid":false,"given":"Hiroyoshi","family":"Matsumura","sequence":"additional","affiliation":[{"name":"Department of Biotechnology College of Life Sciences, Ritsumeikan University  Kusatsu Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3928-5018","authenticated-orcid":false,"given":"Azumi","family":"Hirata","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Cell Biology, Faculty of Medicine Osaka Medical and Pharmaceutical University  Osaka Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6882-9697","authenticated-orcid":false,"given":"Masahiro","family":"Fujihashi","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Faculty of Medicine Osaka Medical and Pharmaceutical University  Osaka Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7548-7314","authenticated-orcid":false,"given":"Kazufumi","family":"Takano","sequence":"additional","affiliation":[{"name":"Graduate School of Life and Environmental Sciences Kyoto Prefectural University  Kyoto Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4270-0374","authenticated-orcid":false,"given":"Shun\u2010ichi","family":"Tanaka","sequence":"additional","affiliation":[{"name":"Graduate School of Life and Environmental Sciences Kyoto Prefectural University  Kyoto Japan"},{"name":"Department of Biotechnology College of Life Sciences, Ritsumeikan University  Kusatsu Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2024,12,26]]},"reference":[{"key":"e_1_2_8_2_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-024-07487-w"},{"key":"e_1_2_8_3_1","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444909052925"},{"key":"e_1_2_8_4_1","doi-asserted-by":"publisher","DOI":"10.1002\/pro.4961"},{"key":"e_1_2_8_5_1","doi-asserted-by":"publisher","DOI":"10.1021\/acschembio.4c00197"},{"key":"e_1_2_8_6_1","doi-asserted-by":"publisher","DOI":"10.1021\/acs.bioconjchem.3c00372"},{"key":"e_1_2_8_7_1","doi-asserted-by":"publisher","DOI":"10.1093\/protein\/15.12.1015"},{"key":"e_1_2_8_8_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2008.01.093"},{"key":"e_1_2_8_9_1","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444909042073"},{"key":"e_1_2_8_10_1","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444910007493"},{"key":"e_1_2_8_11_1","unstructured":"EvansR O'NeillM PritzelA AntropovaN SeniorA GreenT et al.Protein complex prediction with AlphaFold\u2010Multimer. Biorxiv.2022."},{"key":"e_1_2_8_12_1","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444913012274"},{"key":"e_1_2_8_13_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-023-39807-5"},{"key":"e_1_2_8_14_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.sbi.2007.08.012"},{"key":"e_1_2_8_15_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.0505379103"},{"key":"e_1_2_8_16_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-017-15711-z"},{"key":"e_1_2_8_17_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.sbi.2020.01.015"},{"key":"e_1_2_8_18_1","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/30.10.e43"},{"key":"e_1_2_8_19_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ymeth.2012.08.008"},{"key":"e_1_2_8_20_1","doi-asserted-by":"publisher","DOI":"10.1021\/acsmedchemlett.3c00342"},{"key":"e_1_2_8_21_1","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444909047337"},{"key":"e_1_2_8_22_1","doi-asserted-by":"publisher","DOI":"10.1002\/jps.20461"},{"key":"e_1_2_8_23_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2011.12.019"},{"key":"e_1_2_8_24_1","doi-asserted-by":"publisher","DOI":"10.1046\/j.1432-1327.2001.02127.x"},{"key":"e_1_2_8_25_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.82.2.488"},{"key":"e_1_2_8_26_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0969-2126(02)00913-9"},{"key":"e_1_2_8_27_1","first-page":"79","article-title":"FN3\u2010based monobodies selective for the receptor binding domain of the SARS\u2010CoV\u20102 spike protein","volume":"62","author":"Miller CJ","year":"2021","journal-title":"Nat Biotechnol"},{"key":"e_1_2_8_28_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbc.2023.105009"},{"key":"e_1_2_8_29_1","doi-asserted-by":"publisher","DOI":"10.1002\/pro.4813"},{"key":"e_1_2_8_30_1","doi-asserted-by":"publisher","DOI":"10.1021\/acscentsci.0c01708"},{"key":"e_1_2_8_31_1","doi-asserted-by":"publisher","DOI":"10.1093\/oxfordjournals.jbchem.a022684"},{"key":"e_1_2_8_32_1","doi-asserted-by":"publisher","DOI":"10.1110\/ps.051519805"},{"key":"e_1_2_8_33_1","doi-asserted-by":"publisher","DOI":"10.1002\/pro.3148"},{"key":"e_1_2_8_34_1","doi-asserted-by":"publisher","DOI":"10.1006\/jmbi.2000.4195"},{"key":"e_1_2_8_35_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.copbio.2007.04.010"},{"key":"e_1_2_8_36_1","doi-asserted-by":"publisher","DOI":"10.1021\/cr960065d"},{"key":"e_1_2_8_37_1","doi-asserted-by":"publisher","DOI":"10.1038\/nbt1218"},{"key":"e_1_2_8_38_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi400652c"},{"key":"e_1_2_8_39_1","doi-asserted-by":"publisher","DOI":"10.1021\/acschembio.5b01006"},{"key":"e_1_2_8_40_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.2204481119"},{"key":"e_1_2_8_41_1","doi-asserted-by":"publisher","DOI":"10.1006\/jmbi.1999.2772"},{"key":"e_1_2_8_42_1","doi-asserted-by":"publisher","DOI":"10.1110\/ps.8.2.394"},{"key":"e_1_2_8_43_1","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/btq066"},{"key":"e_1_2_8_44_1","doi-asserted-by":"publisher","DOI":"10.1107\/S2059798318004576"},{"key":"e_1_2_8_45_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2003.10.007"},{"key":"e_1_2_8_46_1","doi-asserted-by":"publisher","DOI":"10.1080\/19420862.2023.2213793"},{"key":"e_1_2_8_47_1","doi-asserted-by":"publisher","DOI":"10.1002\/prot.20264"},{"key":"e_1_2_8_48_1","doi-asserted-by":"publisher","DOI":"10.1111\/febs.13674"}],"container-title":["Protein Science"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/pro.70018","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2024,12,30]],"date-time":"2024-12-30T05:07:59Z","timestamp":1735535279000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1002\/pro.70018"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,12,26]]},"references-count":47,"journal-issue":{"issue":"1","published-print":{"date-parts":[[2025,1]]}},"alternative-id":["10.1002\/pro.70018"],"URL":"https:\/\/doi.org\/10.1002\/pro.70018","archive":["Portico"],"relation":{},"ISSN":["0961-8368","1469-896X"],"issn-type":[{"type":"print","value":"0961-8368"},{"type":"electronic","value":"1469-896X"}],"subject":[],"published":{"date-parts":[[2024,12,26]]},"assertion":[{"value":"2024-10-09","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2024-12-13","order":2,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2024-12-26","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e70018"}}