Sum Lai Lozada , Jose Alberto Gómez , Katherine Menéndez , Tania Gómez , Daidee Montes de Oca , Jose L. Durán , Olga Lidia Fernández , Yoel Perera , Gabriela Rivas , Tammy Boggiano-Ayo , Nuris Ledon , Tania Carmenate
{"title":"通过铜催化空气氧化进行氧化重折叠可持续提高新型白细胞介素-2静音素的均一性和活性。","authors":"Sum Lai Lozada , Jose Alberto Gómez , Katherine Menéndez , Tania Gómez , Daidee Montes de Oca , Jose L. Durán , Olga Lidia Fernández , Yoel Perera , Gabriela Rivas , Tammy Boggiano-Ayo , Nuris Ledon , Tania Carmenate","doi":"10.1016/j.jbiotec.2024.07.013","DOIUrl":null,"url":null,"abstract":"<div><p>Interleukin-2 (IL-2) has been used in cancer treatment for over 30 years. However, due to its high toxicity, new mutant variants have been developed. These variants retain some of the biological properties of the original molecule but offer other therapeutic advantages. At the Center of Molecular Immunology, the IL-2 no-alpha mutein, an IL-2 agonist with lower toxicity than wtIL-2, has been designed, produced, and is currently being evaluated in a Phase I/II clinical trial. The mutein is produced in <em>E. coli</em> as an insoluble material that must be refolded <em>in vitro</em> to yield a fully active protein. Controlled oxidation steps are essential in the purification process of recombinant proteins produced in <em>E. coli</em> to ensure the proper formation of the disulfide bonds in the molecules. In this case, the new purification process includes a copper-catalyzed air oxidation step to induce disulfide bond establishment. The optimal conditions of pH, copper, protein and detergent concentration for this step were determined through screening. The produced protein demonstrated a conserved 3D structure, higher purity, and greater biological activity than the obtained by established process without the oxidation step. Four batches were produced and evaluated, demonstrating the consistency of the new process.</p></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"393 ","pages":"Pages 81-90"},"PeriodicalIF":4.1000,"publicationDate":"2024-07-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Oxidative refolding by Copper-catalyzed air oxidation consistently increases the homogeneity and activity of a Novel Interleukin-2 mutein\",\"authors\":\"Sum Lai Lozada , Jose Alberto Gómez , Katherine Menéndez , Tania Gómez , Daidee Montes de Oca , Jose L. Durán , Olga Lidia Fernández , Yoel Perera , Gabriela Rivas , Tammy Boggiano-Ayo , Nuris Ledon , Tania Carmenate\",\"doi\":\"10.1016/j.jbiotec.2024.07.013\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>Interleukin-2 (IL-2) has been used in cancer treatment for over 30 years. However, due to its high toxicity, new mutant variants have been developed. These variants retain some of the biological properties of the original molecule but offer other therapeutic advantages. At the Center of Molecular Immunology, the IL-2 no-alpha mutein, an IL-2 agonist with lower toxicity than wtIL-2, has been designed, produced, and is currently being evaluated in a Phase I/II clinical trial. The mutein is produced in <em>E. coli</em> as an insoluble material that must be refolded <em>in vitro</em> to yield a fully active protein. Controlled oxidation steps are essential in the purification process of recombinant proteins produced in <em>E. coli</em> to ensure the proper formation of the disulfide bonds in the molecules. In this case, the new purification process includes a copper-catalyzed air oxidation step to induce disulfide bond establishment. The optimal conditions of pH, copper, protein and detergent concentration for this step were determined through screening. The produced protein demonstrated a conserved 3D structure, higher purity, and greater biological activity than the obtained by established process without the oxidation step. Four batches were produced and evaluated, demonstrating the consistency of the new process.</p></div>\",\"PeriodicalId\":15153,\"journal\":{\"name\":\"Journal of biotechnology\",\"volume\":\"393 \",\"pages\":\"Pages 81-90\"},\"PeriodicalIF\":4.1000,\"publicationDate\":\"2024-07-18\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of biotechnology\",\"FirstCategoryId\":\"5\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0168165624002025\",\"RegionNum\":2,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"BIOTECHNOLOGY & APPLIED MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of biotechnology","FirstCategoryId":"5","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0168165624002025","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
Oxidative refolding by Copper-catalyzed air oxidation consistently increases the homogeneity and activity of a Novel Interleukin-2 mutein
Interleukin-2 (IL-2) has been used in cancer treatment for over 30 years. However, due to its high toxicity, new mutant variants have been developed. These variants retain some of the biological properties of the original molecule but offer other therapeutic advantages. At the Center of Molecular Immunology, the IL-2 no-alpha mutein, an IL-2 agonist with lower toxicity than wtIL-2, has been designed, produced, and is currently being evaluated in a Phase I/II clinical trial. The mutein is produced in E. coli as an insoluble material that must be refolded in vitro to yield a fully active protein. Controlled oxidation steps are essential in the purification process of recombinant proteins produced in E. coli to ensure the proper formation of the disulfide bonds in the molecules. In this case, the new purification process includes a copper-catalyzed air oxidation step to induce disulfide bond establishment. The optimal conditions of pH, copper, protein and detergent concentration for this step were determined through screening. The produced protein demonstrated a conserved 3D structure, higher purity, and greater biological activity than the obtained by established process without the oxidation step. Four batches were produced and evaluated, demonstrating the consistency of the new process.
期刊介绍:
The Journal of Biotechnology has an open access mirror journal, the Journal of Biotechnology: X, sharing the same aims and scope, editorial team, submission system and rigorous peer review.
The Journal provides a medium for the rapid publication of both full-length articles and short communications on novel and innovative aspects of biotechnology. The Journal will accept papers ranging from genetic or molecular biological positions to those covering biochemical, chemical or bioprocess engineering aspects as well as computer application of new software concepts, provided that in each case the material is directly relevant to biotechnological systems. Papers presenting information of a multidisciplinary nature that would not be suitable for publication in a journal devoted to a single discipline, are particularly welcome.