Cargando…

Metabolic engineering of Bacillus subtilis for redistributing the carbon flux to 2,3-butanediol by manipulating NADH levels

BACKGROUND: Acetoin reductase (Acr) catalyzes the conversion of acetoin to 2,3-butanediol (2,3-BD) with concomitant oxidation of NADH to NAD(+). Therefore, intracellular 2,3-BD production is likely governed by the quantities of rate-limiting factor(s) Acr and/or NADH. Previously, we showed that a hi...

Descripción completa

Detalles Bibliográficos
Autores principales: Yang, Taowei, Rao, Zhiming, Hu, Guiyuan, Zhang, Xian, Liu, Mei, Dai, Yue, Xu, Meijuan, Xu, Zhenghong, Yang, Shang-Tian
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4549875/
https://www.ncbi.nlm.nih.gov/pubmed/26312069
http://dx.doi.org/10.1186/s13068-015-0320-1
_version_ 1782387369465348096
author Yang, Taowei
Rao, Zhiming
Hu, Guiyuan
Zhang, Xian
Liu, Mei
Dai, Yue
Xu, Meijuan
Xu, Zhenghong
Yang, Shang-Tian
author_facet Yang, Taowei
Rao, Zhiming
Hu, Guiyuan
Zhang, Xian
Liu, Mei
Dai, Yue
Xu, Meijuan
Xu, Zhenghong
Yang, Shang-Tian
author_sort Yang, Taowei
collection PubMed
description BACKGROUND: Acetoin reductase (Acr) catalyzes the conversion of acetoin to 2,3-butanediol (2,3-BD) with concomitant oxidation of NADH to NAD(+). Therefore, intracellular 2,3-BD production is likely governed by the quantities of rate-limiting factor(s) Acr and/or NADH. Previously, we showed that a high level of Acr was beneficial for 2,3-BD accumulation. RESULTS: Metabolic engineering strategies were proposed to redistribute carbon flux to 2,3-BD by manipulating NADH levels. The disruption of NADH oxidase (YodC, encoded by yodC) by insertion of a formate dehydrogenase gene in Bacillussubtilis was more efficient for enhancing 2,3-BD production and decreasing acetoin formation than the disruption of YodC by the insertion of a Cat expression cassette. This was because the former resulted in the recombinant strain AFY in which an extra NADH regeneration system was introduced and NADH oxidase was disrupted simultaneously. On fermentation by strain AFY, the highest 2,3-BD concentration increased by 19.9 % while the acetoin titer decreased by 71.9 %, relative to the parental strain. However, the concentration of lactate, the main byproduct, increased by 47.2 %. To further improve carbon flux and NADH to 2,3-BD, the pathway to lactate was blocked using the insertional mutation technique to disrupt the lactate dehydrogenase gene ldhA. The resultant engineered strain B. subtilis AFYL could efficiently convert glucose into 2,3-BD with little acetoin and lactate accumulation. CONCLUSIONS: Through increasing the availability of NADH and decreasing the concentration of unwanted byproducts, this work demonstrates an important strategy in the metabolic engineering of 2,3-BD production by integrative recombinant hosts.
format Online
Article
Text
id pubmed-4549875
institution National Center for Biotechnology Information
language English
publishDate 2015
publisher BioMed Central
record_format MEDLINE/PubMed
spelling pubmed-45498752015-08-27 Metabolic engineering of Bacillus subtilis for redistributing the carbon flux to 2,3-butanediol by manipulating NADH levels Yang, Taowei Rao, Zhiming Hu, Guiyuan Zhang, Xian Liu, Mei Dai, Yue Xu, Meijuan Xu, Zhenghong Yang, Shang-Tian Biotechnol Biofuels Research Article BACKGROUND: Acetoin reductase (Acr) catalyzes the conversion of acetoin to 2,3-butanediol (2,3-BD) with concomitant oxidation of NADH to NAD(+). Therefore, intracellular 2,3-BD production is likely governed by the quantities of rate-limiting factor(s) Acr and/or NADH. Previously, we showed that a high level of Acr was beneficial for 2,3-BD accumulation. RESULTS: Metabolic engineering strategies were proposed to redistribute carbon flux to 2,3-BD by manipulating NADH levels. The disruption of NADH oxidase (YodC, encoded by yodC) by insertion of a formate dehydrogenase gene in Bacillussubtilis was more efficient for enhancing 2,3-BD production and decreasing acetoin formation than the disruption of YodC by the insertion of a Cat expression cassette. This was because the former resulted in the recombinant strain AFY in which an extra NADH regeneration system was introduced and NADH oxidase was disrupted simultaneously. On fermentation by strain AFY, the highest 2,3-BD concentration increased by 19.9 % while the acetoin titer decreased by 71.9 %, relative to the parental strain. However, the concentration of lactate, the main byproduct, increased by 47.2 %. To further improve carbon flux and NADH to 2,3-BD, the pathway to lactate was blocked using the insertional mutation technique to disrupt the lactate dehydrogenase gene ldhA. The resultant engineered strain B. subtilis AFYL could efficiently convert glucose into 2,3-BD with little acetoin and lactate accumulation. CONCLUSIONS: Through increasing the availability of NADH and decreasing the concentration of unwanted byproducts, this work demonstrates an important strategy in the metabolic engineering of 2,3-BD production by integrative recombinant hosts. BioMed Central 2015-08-27 /pmc/articles/PMC4549875/ /pubmed/26312069 http://dx.doi.org/10.1186/s13068-015-0320-1 Text en © Yang et al. 2015 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 Article
Yang, Taowei
Rao, Zhiming
Hu, Guiyuan
Zhang, Xian
Liu, Mei
Dai, Yue
Xu, Meijuan
Xu, Zhenghong
Yang, Shang-Tian
Metabolic engineering of Bacillus subtilis for redistributing the carbon flux to 2,3-butanediol by manipulating NADH levels
title Metabolic engineering of Bacillus subtilis for redistributing the carbon flux to 2,3-butanediol by manipulating NADH levels
title_full Metabolic engineering of Bacillus subtilis for redistributing the carbon flux to 2,3-butanediol by manipulating NADH levels
title_fullStr Metabolic engineering of Bacillus subtilis for redistributing the carbon flux to 2,3-butanediol by manipulating NADH levels
title_full_unstemmed Metabolic engineering of Bacillus subtilis for redistributing the carbon flux to 2,3-butanediol by manipulating NADH levels
title_short Metabolic engineering of Bacillus subtilis for redistributing the carbon flux to 2,3-butanediol by manipulating NADH levels
title_sort metabolic engineering of bacillus subtilis for redistributing the carbon flux to 2,3-butanediol by manipulating nadh levels
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4549875/
https://www.ncbi.nlm.nih.gov/pubmed/26312069
http://dx.doi.org/10.1186/s13068-015-0320-1
work_keys_str_mv AT yangtaowei metabolicengineeringofbacillussubtilisforredistributingthecarbonfluxto23butanediolbymanipulatingnadhlevels
AT raozhiming metabolicengineeringofbacillussubtilisforredistributingthecarbonfluxto23butanediolbymanipulatingnadhlevels
AT huguiyuan metabolicengineeringofbacillussubtilisforredistributingthecarbonfluxto23butanediolbymanipulatingnadhlevels
AT zhangxian metabolicengineeringofbacillussubtilisforredistributingthecarbonfluxto23butanediolbymanipulatingnadhlevels
AT liumei metabolicengineeringofbacillussubtilisforredistributingthecarbonfluxto23butanediolbymanipulatingnadhlevels
AT daiyue metabolicengineeringofbacillussubtilisforredistributingthecarbonfluxto23butanediolbymanipulatingnadhlevels
AT xumeijuan metabolicengineeringofbacillussubtilisforredistributingthecarbonfluxto23butanediolbymanipulatingnadhlevels
AT xuzhenghong metabolicengineeringofbacillussubtilisforredistributingthecarbonfluxto23butanediolbymanipulatingnadhlevels
AT yangshangtian metabolicengineeringofbacillussubtilisforredistributingthecarbonfluxto23butanediolbymanipulatingnadhlevels