Metabolic engineering of Escherichia coli for the production of malic acid
Metabolic engineering of Escherichia coli for the production of malic acid
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DOI：
10.1016/j.bej.2008.01.001
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发表时间：
2008-06-01
影响因子：
3.9
通讯作者：
Lee, Sang Yup
中科院分区：
文献类型：
--
作者：
Moon, Soo Yun;Hong, Soon Ho;Lee, Sang Yup
苹果酸是一种C-4-二羧酸，是三羧酸（TCA）循环的中间体。它已广泛应用于聚合物、食品和制药工业。通过代谢流分析，寻找提高大肠杆菌苹果酸产量的策略。模拟结果表明，磷酸烯醇式丙酮酸（PEP）羧化通量的放大允许增加苹果酸的生产。由于E.大肠杆菌将PEP转化为草酰乙酸而不产生ATP，从而失去PEP的高能磷酸键，选择在此转化过程中产生ATP的PEP羧激酶。然而，E.大肠杆菌PEP羧激酶催化将草酰乙酸转化为PEP的反应，而不是期望的相反反应。因此，我们克隆了Mannheimia succiniciproducens的PEP羧激酶（由pckA基因编码），其将PEP转化为草酰乙酸作为有利的反应。pta突变体E.大肠杆菌WGS-10菌株，该菌株携带含有M.构建succiniciproducens pckA基因并在37 ℃下培养。好氧培养12 h后，苹果酸的最终浓度可达9.25 g/L。(C)2008 Elsevier B. V.保留所有权利。
Malic acid is a C-4-dicarboxylic acid and an intermediate of tricarboxylic acid (TCA) cycle. It has been widely used in the polymer, food and pharmaceutical industries. Metabolic flux analysis was performed to find a strategy for enhanced malic acid production in Escherichia coli. The simulation results suggested that the amplification of phosphoenolpyruvate (PEP) carboxylation flux allowed increased malic acid production. Since the PEP carboxylase of E. coli converts PEP to oxaloacetate without generating ATP, thus losing the high-energy phosphate bond of PEP, the PEP carboxykinase, which generates ATP during this conversion, was chosen. However, the E. coli PEP carboxykinase catalyzes the reaction that converts oxaloacetate to PEP rather than the desirable opposite reaction. Thus, we cloned the PEP carboxykinase (enconded by the pckA gene) of Mannheimia succiniciproducens, which converts PEP to oxaloacetate as a favorable reaction. The pta mutant E. coli strain WGS-10 harboring the plasmid p104ManPck containing the M. succiniciproducens pckA gene was constructed and cultured at 37 degrees C. The final malic acid concentration of 9.25 g/L could be obtained after 12 h of aerobic cultivation. (C) 2008 Elsevier B.V. All rights reserved.