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Tunable switch mediated shikimate biosynthesis in an engineered non-auxotrophic Escherichia coli

Shikimate is a key intermediate in the synthesis of neuraminidase inhibitors. Compared with traditional methods, microbial production of shikimate has the advantages of environmental friendliness, low cost, feed stock renewability, and product selectivity and diversity. Despite these advantages, shi...

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Autores principales: Gu, Pengfei, Su, Tianyuan, Wang, Qian, Liang, Quanfeng, Qi, Qingsheng
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4942831/
https://www.ncbi.nlm.nih.gov/pubmed/27406890
http://dx.doi.org/10.1038/srep29745
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author Gu, Pengfei
Su, Tianyuan
Wang, Qian
Liang, Quanfeng
Qi, Qingsheng
author_facet Gu, Pengfei
Su, Tianyuan
Wang, Qian
Liang, Quanfeng
Qi, Qingsheng
author_sort Gu, Pengfei
collection PubMed
description Shikimate is a key intermediate in the synthesis of neuraminidase inhibitors. Compared with traditional methods, microbial production of shikimate has the advantages of environmental friendliness, low cost, feed stock renewability, and product selectivity and diversity. Despite these advantages, shikimate kinase I and II respectively encoded by aroK and aroL are inactivated in most shikimate microbial producers, thus requiring the addition of aromatic compounds during the fermentation process. To overcome this problem, we constructed a non-auxotrophic, shikimate-synthesising strain of Escherichia coli. By inactivation of repressor proteins, blocking of competitive pathways and overexpression of key enzymes, we increased the shikimate production of wild-type E. coli BW25113 to 1.73 g/L. We then designed a tunable switch that can conditionally decrease gene expression and substituted it for the original aroK promoters. Expression of aroK in the resulting P-9 strain was maintained at a high level during the growth phase and then reduced at a suitable time by addition of an optimal concentration of inducer. In 5-L fed-batch fermentation, strain P-9 produced 13.15 g/L shikimate without the addition of any aromatic compounds. The tunable switch developed in this study is an efficient tool for regulating indispensable genes involved in critical metabolic pathways.
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spelling pubmed-49428312016-07-20 Tunable switch mediated shikimate biosynthesis in an engineered non-auxotrophic Escherichia coli Gu, Pengfei Su, Tianyuan Wang, Qian Liang, Quanfeng Qi, Qingsheng Sci Rep Article Shikimate is a key intermediate in the synthesis of neuraminidase inhibitors. Compared with traditional methods, microbial production of shikimate has the advantages of environmental friendliness, low cost, feed stock renewability, and product selectivity and diversity. Despite these advantages, shikimate kinase I and II respectively encoded by aroK and aroL are inactivated in most shikimate microbial producers, thus requiring the addition of aromatic compounds during the fermentation process. To overcome this problem, we constructed a non-auxotrophic, shikimate-synthesising strain of Escherichia coli. By inactivation of repressor proteins, blocking of competitive pathways and overexpression of key enzymes, we increased the shikimate production of wild-type E. coli BW25113 to 1.73 g/L. We then designed a tunable switch that can conditionally decrease gene expression and substituted it for the original aroK promoters. Expression of aroK in the resulting P-9 strain was maintained at a high level during the growth phase and then reduced at a suitable time by addition of an optimal concentration of inducer. In 5-L fed-batch fermentation, strain P-9 produced 13.15 g/L shikimate without the addition of any aromatic compounds. The tunable switch developed in this study is an efficient tool for regulating indispensable genes involved in critical metabolic pathways. Nature Publishing Group 2016-07-13 /pmc/articles/PMC4942831/ /pubmed/27406890 http://dx.doi.org/10.1038/srep29745 Text en Copyright © 2016, Macmillan Publishers Limited http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
spellingShingle Article
Gu, Pengfei
Su, Tianyuan
Wang, Qian
Liang, Quanfeng
Qi, Qingsheng
Tunable switch mediated shikimate biosynthesis in an engineered non-auxotrophic Escherichia coli
title Tunable switch mediated shikimate biosynthesis in an engineered non-auxotrophic Escherichia coli
title_full Tunable switch mediated shikimate biosynthesis in an engineered non-auxotrophic Escherichia coli
title_fullStr Tunable switch mediated shikimate biosynthesis in an engineered non-auxotrophic Escherichia coli
title_full_unstemmed Tunable switch mediated shikimate biosynthesis in an engineered non-auxotrophic Escherichia coli
title_short Tunable switch mediated shikimate biosynthesis in an engineered non-auxotrophic Escherichia coli
title_sort tunable switch mediated shikimate biosynthesis in an engineered non-auxotrophic escherichia coli
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4942831/
https://www.ncbi.nlm.nih.gov/pubmed/27406890
http://dx.doi.org/10.1038/srep29745
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