Department of Chemical and Biomolecular Engineering, University of California, Los Angeles, Los Angeles, CA 90095, USA.
Appl Environ Microbiol. 2011 May;77(9):2905-15. doi: 10.1128/AEM.03034-10. Epub 2011 Mar 11.
1-Butanol, an important chemical feedstock and advanced biofuel, is produced by Clostridium species. Various efforts have been made to transfer the clostridial 1-butanol pathway into other microorganisms. However, in contrast to similar compounds, only limited titers of 1-butanol were attained. In this work, we constructed a modified clostridial 1-butanol pathway in Escherichia coli to provide an irreversible reaction catalyzed by trans-enoyl-coenzyme A (CoA) reductase (Ter) and created NADH and acetyl-CoA driving forces to direct the flux. We achieved high-titer (30 g/liter) and high-yield (70 to 88% of the theoretical) production of 1-butanol anaerobically, comparable to or exceeding the levels demonstrated by native producers. Without the NADH and acetyl-CoA driving forces, the Ter reaction alone only achieved about 1/10 the level of production. The engineered host platform also enables the selection of essential enzymes with better catalytic efficiency or expression by anaerobic growth rescue. These results demonstrate the importance of driving forces in the efficient production of nonnative products.
1-丁醇是一种重要的化学原料和先进的生物燃料，由梭菌属产生。人们已经做出了各种努力将梭菌的 1-丁醇途径转移到其他微生物中。然而，与类似的化合物相比，只能达到有限的 1-丁醇产量。在这项工作中，我们在大肠杆菌中构建了一个改良的梭菌 1-丁醇途径，提供了由反式烯酰辅酶 A（CoA）还原酶（Ter）催化的不可逆反应，并产生 NADH 和乙酰辅酶 A 驱动力来引导通量。我们实现了 1-丁醇的高产（30 克/升）和高收率（70%至 88%的理论值）的厌氧生产，与天然生产者的水平相当或超过。没有 NADH 和乙酰辅酶 A 驱动力，仅 Ter 反应的产量约为 1/10。工程化的宿主平台还可以通过厌氧生长拯救来选择具有更好催化效率或表达的必需酶。这些结果表明驱动力在非天然产物的高效生产中的重要性。
