The effect of carbon sources and lactate dehydrogenase deletion on 1,2-propanediol production in Escherichia coli
The effect of carbon sources and lactate dehydrogenase deletion on 1,2-propanediol production in Escherichia coli
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DOI：
10.1007/s10295-002-0006-0
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发表时间：
2003-01-01
影响因子：
3.4
通讯作者：
Bennett, GN
中科院分区：
文献类型：
--
作者：
Berríos-Rivera, SJ;San, KY;Bennett, GN
在之前的研究中，我们发现辅因子操作可以潜在地用作代谢工程的工具。在这项研究中，类似葡萄糖的糖被用作底物，这些糖可以用于糖酵解和丙酮酸的生产，但具有不同的氧化态。这提供了一种简单的方法来测试操纵NADH/NAD+比率或NADH的可用性对大肠杆菌在厌氧条件下的代谢模式和1,2-丙二醇（1,2- pd）的生产的影响，后者的合成需要NADH。通过过表达乙酰丁酸梭菌的甲基乙二醛合成酶和大肠杆菌的甘油脱氢酶，实现了1,2- pd的生产。此外，我们还研究了通过使用ldh（-）菌株（不具有乳酸脱氢酶活性）消除NADH竞争途径对1,2- pd生产的影响。碳源的氧化状态显著影响代谢物的产量，如乙醇、醋酸盐和乳酸盐。然而，添加更多的还原碳源并没有提高1,2- pd的产率。以葡萄糖为碳源的1,2- pd的生产通过加入ldh突变得到了改善。这些实验结果表明，我们目前的1,2- pd生产系统不受NADH的限制，而是受甲基乙二醛形成后的途径的限制。
In previous studies, we showed that cofactor manipulations can potentially be used as a tool in metabolic engineering. In this study, sugars similar to glucose, that can feed into glycolysis and pyruvate production, but with different oxidation states, were used as substrates. This provided a simple way of testing the effect of manipulating the NADH/NAD+ ratio or the availability of NADH on the metabolic patterns of Escherichia coli under anaerobic conditions and on the production of 1,2-propanediol (1,2-PD), which requires NADH for its synthesis. Production of 1,2-PD was achieved by overexpressing the two enzymes methylglyoxal synthase from Clostridium acetobutylicum and glycerol dehydrogenase from E. coli. In addition, the effect of eliminating a pathway competing for NADH by using a ldh(-) strain (without lactate dehydrogenase activity) on the production of 1,2-PD was investigated. The oxidation state of the carbon source significantly affected the yield of metabolites, such as ethanol, acetate and lactate. However, feeding a more reduced carbon source did not increase the yield of 1,2-PD. The production of 1,2-PD with glucose as the carbon source was improved by the incorporation of a ldh- mutation. The results of these experiments indicate that our current 1,2-PD production system is not limited by NADH, but rather by the pathways following the formation of methylglyoxal.