Cargando…

Impairment of NADH dehydrogenase and regulation of anaerobic metabolism by the small RNA RyhB and NadE for improved biohydrogen production in Enterobacter aerogenes

BACKGROUND: Enterobacter aerogenes is a facultative anaerobe and is one of the most widely studied bacterial strains because of its ability to use a variety of substrates, to produce hydrogen at a high rate, and its high growth rate during dark fermentation. However, the rate of hydrogen production...

Descripción completa

Detalles Bibliográficos
Autores principales: Wu, Yan, Hao, Yaqiao, Wei, Xuan, Shen, Qi, Ding, Xuanwei, Wang, Liyan, Zhao, Hongxin, Lu, Yuan
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5663082/
https://www.ncbi.nlm.nih.gov/pubmed/29093752
http://dx.doi.org/10.1186/s13068-017-0938-2
_version_ 1783274760986165248
author Wu, Yan
Hao, Yaqiao
Wei, Xuan
Shen, Qi
Ding, Xuanwei
Wang, Liyan
Zhao, Hongxin
Lu, Yuan
author_facet Wu, Yan
Hao, Yaqiao
Wei, Xuan
Shen, Qi
Ding, Xuanwei
Wang, Liyan
Zhao, Hongxin
Lu, Yuan
author_sort Wu, Yan
collection PubMed
description BACKGROUND: Enterobacter aerogenes is a facultative anaerobe and is one of the most widely studied bacterial strains because of its ability to use a variety of substrates, to produce hydrogen at a high rate, and its high growth rate during dark fermentation. However, the rate of hydrogen production has not been optimized. In this present study, three strategies to improve hydrogen production in E. aerogenes, namely the disruption of nuoCDE, overexpression of the small RNA RyhB and of NadE to regulate global anaerobic metabolism, and the redistribution of metabolic flux. The goal of this study was to clarify the effect of nuoCDE, RyhB, and NadE on hydrogen production and how the perturbation of NADH influences the yield of hydrogen gas from E. aerogenes. RESULTS: NADH dehydrogenase activity was impaired by knocking out nuoCD or nuoCDE in E. aerogenes IAM1183 using the CRISPR-Cas9 system to explore the consequent effect on hydrogen production. The hydrogen yields from IAM1183-CD(∆nuoC/∆nuoD) and IAM1183-CDE (∆nuoC/∆nuoD/∆nuoE) increased, respectively, by 24.5 and 45.6% in batch culture (100 mL serum bottles). The hydrogen produced via the NADH pathway increased significantly in IAM1183-CDE, suggesting that nuoE plays an important role in regulating NADH concentration in E. aerogenes. Batch-cultivating experiments showed that by the overexpression of NadE (N), the hydrogen yields of IAM1183/N, IAM1183-CD/N, and IAM1183-CDE/N increased 1.06-, 1.35-, and 1.55-folds, respectively, compared with IAM1183. Particularly worth mentioning is that the strain IAM118-CDE/N reached 2.28 mol in H(2) yield, per mole of glucose consumed. IAN1183/R, IAM1183-CD/R, and IAM1183-CDE/R showed increasing H(2) yields in batch culture. Metabolic flux analysis indicated that increased expression of RyhB led to a significant shift in metabolic patterns. We further investigated IAM1183-CDE/N, which had the best hydrogen-producing traits, as a potential candidate for industry applications using a 5-L fermenter; hydrogen production reached up to 1.95 times greater than that measured for IAM1183. CONCLUSIONS: Knockout of nuoCD or nuoCDE and the overexpression of nadE in E. aerogenes resulted in a redistribution of metabolic flux and improved the hydrogen yield. Overexpression of RyhB had an significant change on the hydrogen production via NADH pathway. A combination of strategies would be a novel approach for developing a more economic and efficient bioprocess for hydrogen production in E. aerogenes. Finally, the latest CRISPR-Cas9 technology was successful for editing genes in E. aerogenes to develop our engineered strain for hydrogen production. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13068-017-0938-2) contains supplementary material, which is available to authorized users.
format Online
Article
Text
id pubmed-5663082
institution National Center for Biotechnology Information
language English
publishDate 2017
publisher BioMed Central
record_format MEDLINE/PubMed
spelling pubmed-56630822017-11-01 Impairment of NADH dehydrogenase and regulation of anaerobic metabolism by the small RNA RyhB and NadE for improved biohydrogen production in Enterobacter aerogenes Wu, Yan Hao, Yaqiao Wei, Xuan Shen, Qi Ding, Xuanwei Wang, Liyan Zhao, Hongxin Lu, Yuan Biotechnol Biofuels Research BACKGROUND: Enterobacter aerogenes is a facultative anaerobe and is one of the most widely studied bacterial strains because of its ability to use a variety of substrates, to produce hydrogen at a high rate, and its high growth rate during dark fermentation. However, the rate of hydrogen production has not been optimized. In this present study, three strategies to improve hydrogen production in E. aerogenes, namely the disruption of nuoCDE, overexpression of the small RNA RyhB and of NadE to regulate global anaerobic metabolism, and the redistribution of metabolic flux. The goal of this study was to clarify the effect of nuoCDE, RyhB, and NadE on hydrogen production and how the perturbation of NADH influences the yield of hydrogen gas from E. aerogenes. RESULTS: NADH dehydrogenase activity was impaired by knocking out nuoCD or nuoCDE in E. aerogenes IAM1183 using the CRISPR-Cas9 system to explore the consequent effect on hydrogen production. The hydrogen yields from IAM1183-CD(∆nuoC/∆nuoD) and IAM1183-CDE (∆nuoC/∆nuoD/∆nuoE) increased, respectively, by 24.5 and 45.6% in batch culture (100 mL serum bottles). The hydrogen produced via the NADH pathway increased significantly in IAM1183-CDE, suggesting that nuoE plays an important role in regulating NADH concentration in E. aerogenes. Batch-cultivating experiments showed that by the overexpression of NadE (N), the hydrogen yields of IAM1183/N, IAM1183-CD/N, and IAM1183-CDE/N increased 1.06-, 1.35-, and 1.55-folds, respectively, compared with IAM1183. Particularly worth mentioning is that the strain IAM118-CDE/N reached 2.28 mol in H(2) yield, per mole of glucose consumed. IAN1183/R, IAM1183-CD/R, and IAM1183-CDE/R showed increasing H(2) yields in batch culture. Metabolic flux analysis indicated that increased expression of RyhB led to a significant shift in metabolic patterns. We further investigated IAM1183-CDE/N, which had the best hydrogen-producing traits, as a potential candidate for industry applications using a 5-L fermenter; hydrogen production reached up to 1.95 times greater than that measured for IAM1183. CONCLUSIONS: Knockout of nuoCD or nuoCDE and the overexpression of nadE in E. aerogenes resulted in a redistribution of metabolic flux and improved the hydrogen yield. Overexpression of RyhB had an significant change on the hydrogen production via NADH pathway. A combination of strategies would be a novel approach for developing a more economic and efficient bioprocess for hydrogen production in E. aerogenes. Finally, the latest CRISPR-Cas9 technology was successful for editing genes in E. aerogenes to develop our engineered strain for hydrogen production. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13068-017-0938-2) contains supplementary material, which is available to authorized users. BioMed Central 2017-10-30 /pmc/articles/PMC5663082/ /pubmed/29093752 http://dx.doi.org/10.1186/s13068-017-0938-2 Text en © The Author(s) 2017 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
Wu, Yan
Hao, Yaqiao
Wei, Xuan
Shen, Qi
Ding, Xuanwei
Wang, Liyan
Zhao, Hongxin
Lu, Yuan
Impairment of NADH dehydrogenase and regulation of anaerobic metabolism by the small RNA RyhB and NadE for improved biohydrogen production in Enterobacter aerogenes
title Impairment of NADH dehydrogenase and regulation of anaerobic metabolism by the small RNA RyhB and NadE for improved biohydrogen production in Enterobacter aerogenes
title_full Impairment of NADH dehydrogenase and regulation of anaerobic metabolism by the small RNA RyhB and NadE for improved biohydrogen production in Enterobacter aerogenes
title_fullStr Impairment of NADH dehydrogenase and regulation of anaerobic metabolism by the small RNA RyhB and NadE for improved biohydrogen production in Enterobacter aerogenes
title_full_unstemmed Impairment of NADH dehydrogenase and regulation of anaerobic metabolism by the small RNA RyhB and NadE for improved biohydrogen production in Enterobacter aerogenes
title_short Impairment of NADH dehydrogenase and regulation of anaerobic metabolism by the small RNA RyhB and NadE for improved biohydrogen production in Enterobacter aerogenes
title_sort impairment of nadh dehydrogenase and regulation of anaerobic metabolism by the small rna ryhb and nade for improved biohydrogen production in enterobacter aerogenes
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5663082/
https://www.ncbi.nlm.nih.gov/pubmed/29093752
http://dx.doi.org/10.1186/s13068-017-0938-2
work_keys_str_mv AT wuyan impairmentofnadhdehydrogenaseandregulationofanaerobicmetabolismbythesmallrnaryhbandnadeforimprovedbiohydrogenproductioninenterobacteraerogenes
AT haoyaqiao impairmentofnadhdehydrogenaseandregulationofanaerobicmetabolismbythesmallrnaryhbandnadeforimprovedbiohydrogenproductioninenterobacteraerogenes
AT weixuan impairmentofnadhdehydrogenaseandregulationofanaerobicmetabolismbythesmallrnaryhbandnadeforimprovedbiohydrogenproductioninenterobacteraerogenes
AT shenqi impairmentofnadhdehydrogenaseandregulationofanaerobicmetabolismbythesmallrnaryhbandnadeforimprovedbiohydrogenproductioninenterobacteraerogenes
AT dingxuanwei impairmentofnadhdehydrogenaseandregulationofanaerobicmetabolismbythesmallrnaryhbandnadeforimprovedbiohydrogenproductioninenterobacteraerogenes
AT wangliyan impairmentofnadhdehydrogenaseandregulationofanaerobicmetabolismbythesmallrnaryhbandnadeforimprovedbiohydrogenproductioninenterobacteraerogenes
AT zhaohongxin impairmentofnadhdehydrogenaseandregulationofanaerobicmetabolismbythesmallrnaryhbandnadeforimprovedbiohydrogenproductioninenterobacteraerogenes
AT luyuan impairmentofnadhdehydrogenaseandregulationofanaerobicmetabolismbythesmallrnaryhbandnadeforimprovedbiohydrogenproductioninenterobacteraerogenes