Pathway engineering of Escherichia coli for α-ketoglutaric acid production.,Biotechnology and Bioengineering

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Pathway engineering of Escherichia coli for α-ketoglutaric acid production.
Biotechnology and Bioengineering ( IF 3.5 ) Pub Date : 2020-06-12 , DOI: 10.1002/bit.27456
Xiulai Chen _{1,

2,

3} , Xiaoxiang Dong _{1,

2,

3} , Jia Liu _{1,

2,

3} , Qiuling Luo _{1,

4} , Liming Liu _{1,

2,

3}

Affiliation

α‐Ketoglutaric acid (α‐KG) is a multifunctional dicarboxylic acid in the tricarboxylic acid (TCA) cycle, but microbial engineering for α‐KG production is not economically efficient, due to the intrinsic inefficiency of its biosynthetic pathway. In this study, pathway engineering was used to improve pathway efficiency for α‐KG production in Escherichia coli. First, the TCA cycle was rewired for α‐KG production starting from pyruvate, and the engineered strain E. coli W3110Δ4‐PCAI produced 15.66 g/L α‐KG. Then, the rewired TCA cycle was optimized by designing various strengths of pyruvate carboxylase and isocitrate dehydrogenase expression cassettes, resulting in a large increase in α‐KG production (24.66 g/L). Furthermore, acetyl coenzyme A (acetyl‐CoA) availability was improved by overexpressing acetyl‐CoA synthetase, leading to α‐KG production up to 28.54 g/L. Finally, the engineered strain E. coli W3110Δ4‐P_(H)CAI_(H)A was able to produce 32.20 g/L α‐KG in a 5‐L fed‐batch bioreactor. This strategy described here paves the way to the development of an efficient pathway for microbial production of α‐KG.

中文翻译：

大肠杆菌生产α-酮戊二酸的途径工程。

α-酮戊二酸 (α-KG) 是三羧酸 (TCA) 循环中的多功能二羧酸，但由于其生物合成途径固有的低效率，用于 α-KG 生产的微生物工程并不经济有效。在本研究中，通路工程用于提高大肠杆菌中α-KG 生产的通路效率。首先，从丙酮酸和工程菌株大肠杆菌开始，重新连接 TCA 循环以生产 α-KGW3110Δ4-PCAI 产生 15.66 g/L α-KG。然后，通过设计不同强度的丙酮酸羧化酶和异柠檬酸脱氢酶表达盒来优化重新连接的 TCA 循环，从而导致 α-KG 产量大幅增加（24.66 g/L）。此外，通过过度表达乙酰辅酶 A 合成酶提高了乙酰辅酶 A（乙酰辅酶 A）的可用性，导致 α-KG 产量高达 28.54 g/L。最后，工程菌株大肠杆菌W3110Δ4-P _(H) CAI _(H) A 能够在 5-L 分批补料生物反应器中生产 32.20 g/L α-KG。这里描述的这种策略为开发微生物生产 α-KG 的有效途径铺平了道路。

更新日期：2020-08-14

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