Department of Chemical and Biomolecular Engineering, University of California, Los Angeles, CA 90095, USA.
Proc Natl Acad Sci U S A. 2012 Apr 17;109(16):6018-23. doi: 10.1073/pnas.1200074109. Epub 2012 Apr 2.
While conservation of ATP is often a desirable trait for microbial production of chemicals, we demonstrate that additional consumption of ATP may be beneficial to drive product formation in a nonnatural pathway. Although production of 1-butanol by the fermentative coenzyme A (CoA)-dependent pathway using the reversal of β-oxidation exists in nature and has been demonstrated in various organisms, the first step of the pathway, condensation of two molecules of acetyl-CoA to acetoacetyl-CoA, is thermodynamically unfavorable. Here, we show that artificially engineered ATP consumption through a pathway modification can drive this reaction forward and enables for the first time the direct photosynthetic production of 1-butanol from cyanobacteria Synechococcus elongatus PCC 7942. We further demonstrated that substitution of bifunctional aldehyde/alcohol dehydrogenase (AdhE2) with separate butyraldehyde dehydrogenase (Bldh) and NADPH-dependent alcohol dehydrogenase (YqhD) increased 1-butanol production by 4-fold. These results demonstrated the importance of ATP and cofactor driving forces as a design principle to alter metabolic flux.
虽然对于微生物生产化学品来说，ATP 的节约通常是一个理想的特性，但我们证明，额外消耗 ATP 可能有助于在非天然途径中驱动产物形成。虽然通过发酵辅酶 A (CoA) 依赖性途径利用β-氧化的逆转来生产 1-丁醇在自然界中存在，并且已经在各种生物体中得到证明，但该途径的第一步，即两个乙酰辅酶 A 分子的缩合形成乙酰乙酰辅酶 A，热力学上是不利的。在这里，我们通过途径修饰表明，人为设计的 ATP 消耗可以推动这一反应的进行，并首次使蓝藻 Synechococcus elongatus PCC 7942 能够直接从光合作用中生产 1-丁醇。我们进一步证明，用单独的丁醛脱氢酶 (Bldh) 和 NADPH 依赖性醇脱氢酶 (YqhD) 替代双功能醛/醇脱氢酶 (AdhE2)，可使 1-丁醇产量增加 4 倍。这些结果表明，ATP 和辅因子驱动力作为改变代谢通量的设计原则的重要性。
