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Regulation of ATP levels in Escherichia coli using CRISPR interference for enhanced pinocembrin production

BACKGROUND: Microbial biosynthesis of natural products holds promise for preclinical studies and treating diseases. For instance, pinocembrin is a natural flavonoid with important pharmacologic characteristics and is widely used in preclinical studies. However, high yield of natural products product...

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Autores principales: Tao, Sha, Qian, Ying, Wang, Xin, Cao, Weijia, Ma, Weichao, Chen, Kequan, Ouyang, Pingkai
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6142380/
https://www.ncbi.nlm.nih.gov/pubmed/30227873
http://dx.doi.org/10.1186/s12934-018-0995-7
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author Tao, Sha
Qian, Ying
Wang, Xin
Cao, Weijia
Ma, Weichao
Chen, Kequan
Ouyang, Pingkai
author_facet Tao, Sha
Qian, Ying
Wang, Xin
Cao, Weijia
Ma, Weichao
Chen, Kequan
Ouyang, Pingkai
author_sort Tao, Sha
collection PubMed
description BACKGROUND: Microbial biosynthesis of natural products holds promise for preclinical studies and treating diseases. For instance, pinocembrin is a natural flavonoid with important pharmacologic characteristics and is widely used in preclinical studies. However, high yield of natural products production is often limited by the intracellular cofactor level, including adenosine triphosphate (ATP). To address this challenge, tailored modification of ATP concentration in Escherichia coli was applied in efficient pinocembrin production. RESULTS: In the present study, a clustered regularly interspaced short palindromic repeats (CRISPR) interference system was performed for screening several ATP-related candidate genes, where metK and proB showed its potential to improve ATP level and increased pinocembrin production. Subsequently, the repression efficiency of metK and proB were optimized to achieve the appropriate levels of ATP and enhancing the pinocembrin production, which allowed the pinocembrin titer increased to 102.02 mg/L. Coupled with the malonyl-CoA engineering and optimization of culture and induction condition, a final pinocembrin titer of 165.31 mg/L was achieved, which is 10.2-fold higher than control strains. CONCLUSIONS: Our results introduce a strategy to approach the efficient biosynthesis of pinocembrin via ATP level strengthen using CRISPR interference. Furthermore coupled with the malonyl-CoA engineering and induction condition have been optimized for pinocembrin production. The results and engineering strategies demonstrated here would hold promise for the ATP level improvement of other flavonoids by CRISPRi system, thereby facilitating other flavonoids production. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12934-018-0995-7) contains supplementary material, which is available to authorized users.
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spelling pubmed-61423802018-09-20 Regulation of ATP levels in Escherichia coli using CRISPR interference for enhanced pinocembrin production Tao, Sha Qian, Ying Wang, Xin Cao, Weijia Ma, Weichao Chen, Kequan Ouyang, Pingkai Microb Cell Fact Research BACKGROUND: Microbial biosynthesis of natural products holds promise for preclinical studies and treating diseases. For instance, pinocembrin is a natural flavonoid with important pharmacologic characteristics and is widely used in preclinical studies. However, high yield of natural products production is often limited by the intracellular cofactor level, including adenosine triphosphate (ATP). To address this challenge, tailored modification of ATP concentration in Escherichia coli was applied in efficient pinocembrin production. RESULTS: In the present study, a clustered regularly interspaced short palindromic repeats (CRISPR) interference system was performed for screening several ATP-related candidate genes, where metK and proB showed its potential to improve ATP level and increased pinocembrin production. Subsequently, the repression efficiency of metK and proB were optimized to achieve the appropriate levels of ATP and enhancing the pinocembrin production, which allowed the pinocembrin titer increased to 102.02 mg/L. Coupled with the malonyl-CoA engineering and optimization of culture and induction condition, a final pinocembrin titer of 165.31 mg/L was achieved, which is 10.2-fold higher than control strains. CONCLUSIONS: Our results introduce a strategy to approach the efficient biosynthesis of pinocembrin via ATP level strengthen using CRISPR interference. Furthermore coupled with the malonyl-CoA engineering and induction condition have been optimized for pinocembrin production. The results and engineering strategies demonstrated here would hold promise for the ATP level improvement of other flavonoids by CRISPRi system, thereby facilitating other flavonoids production. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12934-018-0995-7) contains supplementary material, which is available to authorized users. BioMed Central 2018-09-18 /pmc/articles/PMC6142380/ /pubmed/30227873 http://dx.doi.org/10.1186/s12934-018-0995-7 Text en © The Author(s) 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research
Tao, Sha
Qian, Ying
Wang, Xin
Cao, Weijia
Ma, Weichao
Chen, Kequan
Ouyang, Pingkai
Regulation of ATP levels in Escherichia coli using CRISPR interference for enhanced pinocembrin production
title Regulation of ATP levels in Escherichia coli using CRISPR interference for enhanced pinocembrin production
title_full Regulation of ATP levels in Escherichia coli using CRISPR interference for enhanced pinocembrin production
title_fullStr Regulation of ATP levels in Escherichia coli using CRISPR interference for enhanced pinocembrin production
title_full_unstemmed Regulation of ATP levels in Escherichia coli using CRISPR interference for enhanced pinocembrin production
title_short Regulation of ATP levels in Escherichia coli using CRISPR interference for enhanced pinocembrin production
title_sort regulation of atp levels in escherichia coli using crispr interference for enhanced pinocembrin production
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6142380/
https://www.ncbi.nlm.nih.gov/pubmed/30227873
http://dx.doi.org/10.1186/s12934-018-0995-7
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