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Overexpression and characterization of two types of nitrile hydratases from Rhodococcus rhodochrous J1
Nitrile hydratase (NHase) from Rhodococcus rhodochrous J1 is widely used for industrial production of acrylamide and nicotinamide. However, the two types of NHases (L-NHase and H-NHase) from R. rhodochrous J1 were only slightly expressed in E. coli by routine methods, which limits the comprehensive...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Public Library of Science
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5482477/ https://www.ncbi.nlm.nih.gov/pubmed/28644864 http://dx.doi.org/10.1371/journal.pone.0179833 |
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author | Lan, Yao Zhang, Xiaohuan Liu, Zhongmei Zhou, Li Shen, Ruihua Zhong, Xianping Cui, Wenjing Zhou, Zhemin |
author_facet | Lan, Yao Zhang, Xiaohuan Liu, Zhongmei Zhou, Li Shen, Ruihua Zhong, Xianping Cui, Wenjing Zhou, Zhemin |
author_sort | Lan, Yao |
collection | PubMed |
description | Nitrile hydratase (NHase) from Rhodococcus rhodochrous J1 is widely used for industrial production of acrylamide and nicotinamide. However, the two types of NHases (L-NHase and H-NHase) from R. rhodochrous J1 were only slightly expressed in E. coli by routine methods, which limits the comprehensive and systematic characterization of the enzyme properties. We successfully expressed the two types of recombinant NHases in E. coli by codon-optimization, engineering of Ribosome Binding Site (RBS) and spacer sequences. The specific activity of the purified L-NHase and H-NHase were 400 U/mg and 234 U/mg, respectively. The molecular mass of L-NHase and H-NHase was identified to be 94 kDa and 504 kDa, respectively, indicating that the quaternary structure of the two types of NHases was the same as those in R. rhodochrous J1. H-NHase exhibited higher substrate and product tolerance than L-NHase. Moreover, higher activity and shorter culture time were achieved in recombinant E. coli, and the whole cell catalyst of recombinant E. coli harboring H-NHase has equivalent efficiency in tolerance to the high-concentration product relative to that in R. rhodochrous J1. These results indicate that biotransformation of nitrile by R. rhodochrous J1 represents a potential alternative to NHase-producing E. coli. |
format | Online Article Text |
id | pubmed-5482477 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-54824772017-07-06 Overexpression and characterization of two types of nitrile hydratases from Rhodococcus rhodochrous J1 Lan, Yao Zhang, Xiaohuan Liu, Zhongmei Zhou, Li Shen, Ruihua Zhong, Xianping Cui, Wenjing Zhou, Zhemin PLoS One Research Article Nitrile hydratase (NHase) from Rhodococcus rhodochrous J1 is widely used for industrial production of acrylamide and nicotinamide. However, the two types of NHases (L-NHase and H-NHase) from R. rhodochrous J1 were only slightly expressed in E. coli by routine methods, which limits the comprehensive and systematic characterization of the enzyme properties. We successfully expressed the two types of recombinant NHases in E. coli by codon-optimization, engineering of Ribosome Binding Site (RBS) and spacer sequences. The specific activity of the purified L-NHase and H-NHase were 400 U/mg and 234 U/mg, respectively. The molecular mass of L-NHase and H-NHase was identified to be 94 kDa and 504 kDa, respectively, indicating that the quaternary structure of the two types of NHases was the same as those in R. rhodochrous J1. H-NHase exhibited higher substrate and product tolerance than L-NHase. Moreover, higher activity and shorter culture time were achieved in recombinant E. coli, and the whole cell catalyst of recombinant E. coli harboring H-NHase has equivalent efficiency in tolerance to the high-concentration product relative to that in R. rhodochrous J1. These results indicate that biotransformation of nitrile by R. rhodochrous J1 represents a potential alternative to NHase-producing E. coli. Public Library of Science 2017-06-23 /pmc/articles/PMC5482477/ /pubmed/28644864 http://dx.doi.org/10.1371/journal.pone.0179833 Text en © 2017 Lan et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Lan, Yao Zhang, Xiaohuan Liu, Zhongmei Zhou, Li Shen, Ruihua Zhong, Xianping Cui, Wenjing Zhou, Zhemin Overexpression and characterization of two types of nitrile hydratases from Rhodococcus rhodochrous J1 |
title | Overexpression and characterization of two types of nitrile hydratases from Rhodococcus rhodochrous J1 |
title_full | Overexpression and characterization of two types of nitrile hydratases from Rhodococcus rhodochrous J1 |
title_fullStr | Overexpression and characterization of two types of nitrile hydratases from Rhodococcus rhodochrous J1 |
title_full_unstemmed | Overexpression and characterization of two types of nitrile hydratases from Rhodococcus rhodochrous J1 |
title_short | Overexpression and characterization of two types of nitrile hydratases from Rhodococcus rhodochrous J1 |
title_sort | overexpression and characterization of two types of nitrile hydratases from rhodococcus rhodochrous j1 |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5482477/ https://www.ncbi.nlm.nih.gov/pubmed/28644864 http://dx.doi.org/10.1371/journal.pone.0179833 |
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