Fine-Tuning of the Fatty Acid Pathway by Synthetic Antisense RNA for Enhanced (2S)-Naringenin Production from L-Tyrosine in Escherichia coli
Fine-Tuning of the Fatty Acid Pathway by Synthetic Antisense RNA for Enhanced (2S)-Naringenin Production from L-Tyrosine in Escherichia coli
复制标题
通过合成反义 RNA 微调脂肪酸途径以增强大肠杆菌中 L-酪氨酸的 (2S)-柚皮素生产
DOI：
10.1128/aem.02411-14
复制
发表时间：
2014-12-01
影响因子：
4.4
通讯作者：
Chen, Jian
中科院分区：
文献类型：
--
作者：
Wu, Junjun;Yu, Oliver;Chen, Jian
丙二醇辅酶A （Malonyl - coa）是合成多酮类、黄酮类等天然产物的重要前体。这种辅助因子的大部分通常被消耗用于产生脂肪酸和磷脂，只留下少量的细胞丙二酰辅酶a用于产生目标化合物。调节丙二酰辅酶a进入异源通路会产生显著的表型效应，如生长迟缓甚至细胞死亡。在这项研究中，提出了利用反义RNA （asRNA）微调大肠杆菌的脂肪酸途径，以平衡靶产物合成和细胞健康对丙二酰辅酶a的需求。为了建立一个高效的asRNA系统，我们探索了asRNA序列与功能之间的关系。结果表明，通过将asRNA定向到靶基因5 ' -UTR（非翻译区）的不同位置，可以调节asRNA的基因沉默效应。基于这一原理，我们定量地调整了asRNA的活性，以平衡细胞生长对丙二酰辅酶a的需求和主要类黄酮前体(2S)-柚皮素的产生。适当的抗fabb /fabF asRNA抑制效率可使生产滴度提高431% （391 mg/l）。因此，本研究提出的策略为细菌内源基因表达的微调提供了一个有用的工具。
ABSTRACT Malonyl coenzyme A (malonyl-CoA) is an important precursor for the synthesis of natural products, such as polyketides and flavonoids. The majority of this cofactor often is consumed for producing fatty acids and phospholipids, leaving only a small amount of cellular malonyl-CoA available for producing the target compound. The tuning of malonyl-CoA into heterologous pathways yields significant phenotypic effects, such as growth retardation and even cell death. In this study, fine-tuning of the fatty acid pathway in Escherichia coli with antisense RNA (asRNA) to balance the demands on malonyl-CoA for target-product synthesis and cell health was proposed. To establish an efficient asRNA system, the relationship between sequence and function for asRNA was explored. It was demonstrated that the gene-silencing effect of asRNA could be tuned by directing asRNA to different positions in the 5′-UTR (untranslated region) of the target gene. Based on this principle, the activity of asRNA was quantitatively tailored to balance the need for malonyl-CoA in cell growth and the production of the main flavonoid precursor, (2S)-naringenin. Appropriate inhibitory efficiency of the anti-fabB/fabF asRNA improved the production titer by 431% (391 mg/liter). Therefore, the strategy presented in this study provided a useful tool for the fine-tuning of endogenous gene expression in bacteria.