Constructing a synthetic constitutive metabolic pathway in Escherichia coli for (R, R)-2,3-butanediol production

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• 作者：Ying-Jia Tong ; Xiao-Jun Ji ; Meng-Qiu Shen…
• 关键词：(R ; R) ; 2 ; 3 ; butanediol ; Promoter ; Escherichia coli ; Metabolic engineering
• 刊名：Applied Microbiology and Biotechnology
• 出版年：2016
• 出版时间：January 2016
• 年：2016
• 卷：100
• 期：2
• 页码：637-647
• 全文大小：907 KB
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• 作者单位：Ying-Jia Tong (1)
  Xiao-Jun Ji (1)
  Meng-Qiu Shen (1)
  Lu-Gang Liu (1)
  Zhi-Kui Nie (1)
  He Huang (1)
  
  1. State Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, No. 30 South Puzhu Road, Nanjing, 211816, People’s Republic of China
  
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Chemistry
  Biotechnology
  Microbiology
  Microbial Genetics and Genomics
  
• 出版者：Springer Berlin / Heidelberg
• ISSN：1432-0614

文摘

Many microorganisms could naturally produce (R, R)-2,3-butanediol ((R, R)-2,3-BD), which has unique applications due to its special chiral group and spatial configuration. But the low enantio-purity of the product hindered the development of large-scale production. In this work, a synthetic constitutive metabolic pathway for enantiomerically pure (R, R)-2,3-BD biosynthesis was constructed in Escherichia coli with vector pUC6S, which does not contain any lac sequences. The expression of this artificial constructed gene cluster was optimized by using two different strength of promoters (AlperPLTet01 (P₀₁) and AlperBB (P_BB)). The strength of P₀₁ is twice stronger than P_BB. The fermentation results suggested that the yield of (R, R)-2,3-BD was higher when using the stronger promoter. Compared with the wild type, the recombinant strain E. coli YJ2 produced a small amount of acetic acid and showed higher glucose consumption rate and higher cell density, which indicated a protection against acetic acid inhibition. In order to further increase the (R, R)-2,3-BD production by reducing the accumulation of its precursor acetoin, the synthetic operon was reconstructed by adding the strong promoter P₀₁ in front of the gene ydjL coding for the enzyme of (R, R)-2,3-BD dehydrogenase which catalyzes the conversion of acetoin to (R, R)-2,3-BD. The engineered strain E. coli YJ3 showed a 20 % decrease in acetoin production compared with that of E. coli YJ2. After optimization the fermentation conditions, 30.5 g/L of (R, R)-2,3-BD and 3.2 g/L of acetoin were produced from 80 g/L of glucose within 18 h, with an enantio-purity over 99 %. Keywords (R, R)-2,3-butanediol Promoter Escherichia coli Metabolic engineering