Engineering dynamic pathway regulation using stress-response promoters
Engineering dynamic pathway regulation using stress-response promoters
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DOI：
10.1038/nbt.2689
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发表时间：
2013-11-01
影响因子：
46.9
通讯作者：
Keasling, Jay D.
中科院分区：
文献类型：
--
作者：
Dahl, Robert H.;Zhang, Fuzhong;Keasling, Jay D.
代谢工程中使用的异源途径可能会产生对细胞有毒的中间体。对途径酶的动态控制可以防止这些代谢物的积累，但这种策略需要传感器，这些传感器在很大程度上是未知的，可以检测并对代谢物做出反应。在这里，我们应用全基因组转录阵列来识别对有毒中间体的积累做出反应的启动子，然后使用这些启动子来控制中间体的积累并提高所需产品的最终效价。我们应用这种方法来调控大肠杆菌中类异戊二烯生物合成途径中焦磷酸法尼基(FPP)的产生。这一策略使最终产品氨甲二烯的产量提高了一倍，不再需要昂贵的诱导剂，减少了乙酸酯的积累，并促进了生长。我们将这种方法扩展到另一种有毒中间体，以展示识别用于动态调节的新型传感器-调节系统的广泛用途。
Heterologous pathways used in metabolic engineering may produce intermediates toxic to the cell. Dynamic control of pathway enzymes could prevent the accumulation of these metabolites, but such a strategy requires sensors, which are largely unknown, that can detect and respond to the metabolite. Here we applied whole-genome transcript arrays to identify promoters that respond to the accumulation of toxic intermediates, and then used these promoters to control accumulation of the intermediate and improve the final titers of a desired product. We apply this approach to regulate farnesyl pyrophosphate (FPP) production in the isoprenoid biosynthetic pathway in Escherichia coli. This strategy improved production of amorphadiene, the final product, by twofold over that from inducible or constitutive promoters, eliminated the need for expensive inducers, reduced acetate accumulation and improved growth. We extended this approach to another toxic intermediate to demonstrate the broad utility of identifying novel sensor-regulator systems for dynamic regulation.