Enhanced succinic acid production under acidic conditions by introduction of glutamate decarboxylase system in E-coli AFP111

Wu Mingke; Li Xiaozhan; Guo Shunfeng; Lemma Wubliker Dessie; Zhang Wenming; Ma Jiangfeng; Jia Honghua; Wu Hao; Jiang Min; Ouyang Pingkai

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Enhanced succinic acid production under acidic conditions by introduction of glutamate decarboxylase system in E-coli AFP111

机译：通过在大肠杆菌AFP111中引入谷氨酸脱羧酶系统提高酸性条件下的琥珀酸生产

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Biological synthesis of succinic acid at low pH values was favored since it not only decreased investment cost but also simplified downstream purification process. In this study, the feasibility of using glutamate decarboxylase system to improve succinic acid production of Escherichia coli AFP111, a succinate-producing candidate with mutations in pfl, ldhA, and ptsG, under acidic conditions was investigated. By overexpressing gadBC operon in AFP111, a recombinant named as BA201 (AFP111/pMD19T-gadBC) was constructed. Fermentation at pH 5.6 showed that 30 g L-1 glucose was consumed and 26.58 g L-1 succinic acid was produced by BA201, which was 1.22- and 1.32-fold higher than that by the control BA200 (AFP111/pMD19T) containing the empty vector. Analysis of intracellular enzymes activities and ATP concentrations revealed that the activities of key enzymes involved in glucose uptake and products synthesis and intracellular ATP levels were all increased after overexpression of gadBC which were benefit for cell metabolism under weak acidic conditions. To further improve the succinic acid titer by recombinant BA201 at pH 5.6, the extracellular glutamate concentration was optimized and the final succinic acid titer increased 20.4% to 32.01 g L-1. Besides, the fermentation time was prolonged by repetitive fermentation and additional 15.78 g L-1 succinic acid was produced by recovering cells into fresh medium. The results here demonstrated a potential strategy of overexpressing gadBC for increased succinic acid production of E. coli AFP111 under weak acidic conditions.

机译：低pH值的琥珀酸的生物合成是有利的，因为它不仅降低了投资成本，而且简化了下游纯化过程。在这项研究中，研究了在酸性条件下使用谷氨酸脱羧酶系统改善琥珀酸生产大肠杆菌AFP111的可行性，大肠杆菌AFP111是产生pfl，ldhA和ptsG突变的琥珀酸生产候选物。通过在AFP111中过量表达gadBC操纵子，构建了名为BA201的重组体（AFP111 / pMD19T-gadBC）。在pH 5.6的发酵条件下，BA201消耗了30 g L-1葡萄糖，产生了26.58 g L-1琥珀酸，比空白对照BA200（AFP111 / pMD19T）高出1.22倍和1.32倍。向量。细胞内酶活性和ATP浓度的分析表明，过量表达gadBC后，参与葡萄糖吸收和产物合成的关键酶的活性以及细胞内ATP水平均增加，这有利于弱酸性条件下的细胞代谢。为了通过pH 5.6的重组BA201进一步提高琥珀酸效价，优化了细胞外谷氨酸的浓度，最终琥珀酸效价提高了20.4％，达到32.01 g L-1。此外，通过重复发酵延长了发酵时间，并且通过将细胞回收到新鲜培养基中而另外产生了15.78g L-1琥珀酸。此处的结果证明了在弱酸性条件下过表达gadBC的潜在策略，以提高大肠杆菌AFP111的琥珀酸产量。

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来源
《Bioprocess and Biosystems Engineering》 |2017年第4期|549-557|共9页
作者
Wu Mingke; Li Xiaozhan; Guo Shunfeng; Lemma Wubliker Dessie; Zhang Wenming; Ma Jiangfeng; Jia Honghua; Wu Hao; Jiang Min; Ouyang Pingkai;
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作者单位

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

Nanjing Tech Univ, Coll Biotechnol & Pharmaceut Engn, State Key Lab Mat Oriented Chem Engn, 30 Puzhu South Rd, Nanjing 211816, Jiangsu, Peoples R China;

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关键词
Low pH fermentation; E. coli AFP111; Glutamate decarboxylase system; Succinic acid;

机译：低pH发酵;E.大肠杆菌AFP111;谷氨酸脱羧酶系统;琥珀酸;

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1. Coproduction of succinic acid and cadaverine using lysine as a neutralizer and CO2 donor with L-lysine decarboxylase overexpressed Escherichia coli AFP111 [J] . Wang Jing, Mao Jingwen, Tian Weilong, Green chemistry . 2018,第12期

机译：用赖氨酸作为中和剂和CO2供体与L-赖氨酸脱羧酶过表达大肠杆菌AFP111的琥珀酸和野兔用赖氨酸和CO2供体
2. Evaluating the Contribution of Acid Resistance Systems and Probing the Different Roles of the Glutamate Decarboxylases of Listeria monocytogenes Under Acidic Conditions [J] . Journal of international management . 2020,第2期

机译：在酸性条件下评估酸性抗性系统的贡献并探测谷氨酸单核细胞增生的谷氨酸脱羧酶的不同作用
3. Succinic acid monomethyl ester protects rat pancreatic islet secretory potential against interleukin‐1β (IL‐1β) without affecting glutamate decarboxylase expression or nitric oxide production [J] . Eizirik Deacute, cio L., Malaisse Willy J., FEBS Letters . 1994,第3期

机译：琥珀酸单甲酯可保护大鼠胰岛分泌潜力，抵抗白介素-1β（IL-1β），而不会影响谷氨酸脱羧酶的表达或一氧化氮的产生
4. Properties of Glutamate Decarboxylase of Aspergillus oryzae and Its Application for Biotransformation of Glutamate into gamma-Aminobutyric Acid [C] . Lu Weiguo, Sun Liwei, Zhang Xiuqing, 2010 4th International Conference on Bioinformatics and Biomedical Engineering . 2010

机译：米曲霉谷氨酸脱羧酶的性质及其在谷氨酸向γ-氨基丁酸生物转化中的应用
5. Part one. Synthesis of phosphorus-containing amino acid analogues derived from trans-4-hydroxyproline and the study of their bioactivity in the glutamate excitatory neurotransmitter system. Part two. Synthesis, structural modifications and in vitro pharmacology of quinoline-2,4-dicarboxylic acids as proposed inhibitors of the glutamate vesicular transporter. [D] . Carrigan, Christina Nicole. 2000

机译：第一部分。反式-4-羟基脯氨酸衍生的含磷氨基酸类似物的合成及其在谷氨酸兴奋性神经递质系统中的生物活性的研究。第二部分。喹啉-2,4-二羧酸作为谷氨酸囊泡转运蛋白的抑制剂的合成，结构修饰和体外药理作用。
6. Disruption of pknG enhances production of gamma-aminobutyric acid by Corynebacterium glutamicum expressing glutamate decarboxylase [O] . Naoko Okai, Chihiro Takahashi, Kazuki Hatada, 2014

机译：pknG的破坏增强了表达谷氨酸脱羧酶的谷氨酸棒杆菌对γ-氨基丁酸的生产
7. Succinic acid monomethyl ester protects rat pancreatic islet secretory potential against interleukin-1 beta (IL-1 beta) without affecting glutamate decarboxylase expression or nitric oxide production. [O] . Eizirik, Decio L., Welsh, Nils, Niemann, A, 1994

机译：琥珀酸单甲酯可保护大鼠胰岛的分泌能力，使其免受白介素1β（IL-1 beta）的影响，而不会影响谷氨酸脱羧酶的表达或一氧化氮的产生。

Enhanced succinic acid production under acidic conditions by introduction of glutamate decarboxylase system in E-coli AFP111

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