Engineering glucose metabolism of Escherichia coli under nitrogen starvation.
Engineering glucose metabolism of Escherichia coli under nitrogen starvation.
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DOI：
10.1038/npjsba.2016.35
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发表时间：
2017
影响因子：
4
通讯作者：
Mukhopadhyay A
中科院分区：
文献类型：
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作者：
Chubukov V;Desmarais JJ;Wang G;Chan LJG;Baidoo EE;Petzold CJ;Keasling JD;Mukhopadhyay A
微生物代谢工程的一个主要方面是开发具有有利的整体代谢表型的底盘宿主，并且可以进一步工程化以产生多种化合物。在这项工作中，我们专注于模型细菌大肠杆菌中的解耦生长和生产的问题，特别是在氮限制的稳定期期间维持活性代谢。我们发现，通过过表达酶PtsI，葡萄糖摄取系统的一个组成部分，在氮限制期间被α-酮戊二酸抑制，我们能够实现代谢率增加四倍。还测试了替代系统：假设对α-酮戊二酸不敏感的嵌合PtsI蛋白在测试条件下没有改善代谢率，而基于半乳糖通透酶GalP的系统遭受能量应激和对表达水平的极端敏感性。PtsI的过表达很可能是代谢工程师研究中的一个有用的箭头，因为在稳定期生产过程中，工程化途径的生产率受到中心代谢率的限制。
A major aspect of microbial metabolic engineering is the development of chassis hosts that have favorable global metabolic phenotypes, and can be further engineered to produce a variety of compounds. In this work, we focus on the problem of decoupling growth and production in the model bacterium Escherichia coli, and in particular on the maintenance of active metabolism during nitrogen-limited stationary phase. We find that by overexpressing the enzyme PtsI, a component of the glucose uptake system that is inhibited by α-ketoglutarate during nitrogen limitation, we are able to achieve a fourfold increase in metabolic rates. Alternative systems were also tested: chimeric PtsI proteins hypothesized to be insensitive to α-ketoglutarate did not improve metabolic rates under the conditions tested, whereas systems based on the galactose permease GalP suffered from energy stress and extreme sensitivity to expression level. Overexpression of PtsI is likely to be a useful arrow in the metabolic engineer’s quiver as productivity of engineered pathways becomes limited by central metabolic rates during stationary phase production processes.
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