Production of soluble truncated spike protein of porcine epidemic diarrhea virus from inclusion bodies of Escherichia coli through refolding
The emergence of highly pathogenic variant porcine epidemic diarrhea virus (PEDV) strains, from 2013 to 2014, in North American and Asian countries have greatly threatened global swine industry. Therefore, development of effective vaccines against PEDV variant strains is urgently needed. Recently, i...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Elsevier Inc.
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7173294/ https://www.ncbi.nlm.nih.gov/pubmed/27260969 http://dx.doi.org/10.1016/j.pep.2016.05.018 |
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author | Piao, Da-Chuan Lee, Yoon-Seok Bok, Jin-Duck Cho, Chong-Su Hong, Zhong-Shan Kang, Sang-Kee Choi, Yun-Jaie |
author_facet | Piao, Da-Chuan Lee, Yoon-Seok Bok, Jin-Duck Cho, Chong-Su Hong, Zhong-Shan Kang, Sang-Kee Choi, Yun-Jaie |
author_sort | Piao, Da-Chuan |
collection | PubMed |
description | The emergence of highly pathogenic variant porcine epidemic diarrhea virus (PEDV) strains, from 2013 to 2014, in North American and Asian countries have greatly threatened global swine industry. Therefore, development of effective vaccines against PEDV variant strains is urgently needed. Recently, it has been reported that the N-terminal domain (NTD) of S1 domain of PEDV spike protein is responsible for binding to the 5-N-acetylneuraminic acid (Neu5Ac), a possible sugar co-receptor. Therefore, the NTD of S1 domain could be an attractive target for the development of subunit vaccines. In this study, the NTD spanning amino acid residues 25-229 (S25-229) of S1 domain of PEDV variant strain was expressed in Escherichia coli BL21 (DE3) in the form of inclusion bodies (IBs). S25-229 IBs were solubilized in 20 mM sodium acetate (pH 4.5) buffer containing 8 M urea and 1 mM dithiothreitol with 95% yield. Solubilized S25-229 IBs were refolded by 10-fold flash dilution and purified by one-step cation exchange chromatography with >95% purity and 20% yield. The CD spectrum of S25-229 showed the characteristic pattern of alpha helical structure. In an indirect ELISA, purified S25-229 showed strong reactivity with mouse anti-PEDV sera. In addition, immunization of mice with 20 μg of purified S25-229 elicited highly potent serum IgG titers. Finally, mouse antisera against S25-229 showed immune reactivity with native PEDV S protein in an immunofluorescence assay. These results suggest that purified S25-229 may have potential to be used as a subunit vaccine against PEDV variant strains. |
format | Online Article Text |
id | pubmed-7173294 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | Elsevier Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-71732942020-04-22 Production of soluble truncated spike protein of porcine epidemic diarrhea virus from inclusion bodies of Escherichia coli through refolding Piao, Da-Chuan Lee, Yoon-Seok Bok, Jin-Duck Cho, Chong-Su Hong, Zhong-Shan Kang, Sang-Kee Choi, Yun-Jaie Protein Expr Purif Article The emergence of highly pathogenic variant porcine epidemic diarrhea virus (PEDV) strains, from 2013 to 2014, in North American and Asian countries have greatly threatened global swine industry. Therefore, development of effective vaccines against PEDV variant strains is urgently needed. Recently, it has been reported that the N-terminal domain (NTD) of S1 domain of PEDV spike protein is responsible for binding to the 5-N-acetylneuraminic acid (Neu5Ac), a possible sugar co-receptor. Therefore, the NTD of S1 domain could be an attractive target for the development of subunit vaccines. In this study, the NTD spanning amino acid residues 25-229 (S25-229) of S1 domain of PEDV variant strain was expressed in Escherichia coli BL21 (DE3) in the form of inclusion bodies (IBs). S25-229 IBs were solubilized in 20 mM sodium acetate (pH 4.5) buffer containing 8 M urea and 1 mM dithiothreitol with 95% yield. Solubilized S25-229 IBs were refolded by 10-fold flash dilution and purified by one-step cation exchange chromatography with >95% purity and 20% yield. The CD spectrum of S25-229 showed the characteristic pattern of alpha helical structure. In an indirect ELISA, purified S25-229 showed strong reactivity with mouse anti-PEDV sera. In addition, immunization of mice with 20 μg of purified S25-229 elicited highly potent serum IgG titers. Finally, mouse antisera against S25-229 showed immune reactivity with native PEDV S protein in an immunofluorescence assay. These results suggest that purified S25-229 may have potential to be used as a subunit vaccine against PEDV variant strains. Elsevier Inc. 2016-10 2016-05-31 /pmc/articles/PMC7173294/ /pubmed/27260969 http://dx.doi.org/10.1016/j.pep.2016.05.018 Text en © 2016 Elsevier Inc. All rights reserved. Since January 2020 Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre - including this research content - immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active. |
spellingShingle | Article Piao, Da-Chuan Lee, Yoon-Seok Bok, Jin-Duck Cho, Chong-Su Hong, Zhong-Shan Kang, Sang-Kee Choi, Yun-Jaie Production of soluble truncated spike protein of porcine epidemic diarrhea virus from inclusion bodies of Escherichia coli through refolding |
title | Production of soluble truncated spike protein of porcine epidemic diarrhea virus from inclusion bodies of Escherichia coli through refolding |
title_full | Production of soluble truncated spike protein of porcine epidemic diarrhea virus from inclusion bodies of Escherichia coli through refolding |
title_fullStr | Production of soluble truncated spike protein of porcine epidemic diarrhea virus from inclusion bodies of Escherichia coli through refolding |
title_full_unstemmed | Production of soluble truncated spike protein of porcine epidemic diarrhea virus from inclusion bodies of Escherichia coli through refolding |
title_short | Production of soluble truncated spike protein of porcine epidemic diarrhea virus from inclusion bodies of Escherichia coli through refolding |
title_sort | production of soluble truncated spike protein of porcine epidemic diarrhea virus from inclusion bodies of escherichia coli through refolding |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7173294/ https://www.ncbi.nlm.nih.gov/pubmed/27260969 http://dx.doi.org/10.1016/j.pep.2016.05.018 |
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