Tunable Gene Expression System Independent of Downstream Coding Sequence,ACS Synthetic Biology

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Tunable Gene Expression System Independent of Downstream Coding Sequence
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2020-10-14 , DOI: 10.1021/acssynbio.0c00029
Seung Min Yoo ₁ , Seung-Woon Jung ₁ , Jinho Yeom ₁ , Sang Yup Lee ₂ , Dokyun Na ₁

Affiliation

Fine control of the expression levels of proteins constitutes a major challenge in synthetic biology and metabolic engineering. However, the dependence of translation initiation on the downstream coding sequence (CDS) obscures accurate prediction of the protein expression levels from mRNA sequences. Here, we present a tunable gene-expression system comprising 24 expression cassettes that produce predefined relative expression levels of proteins ranging from 0.001 to 1 without being influenced by the downstream CDS. To validate the practical utility of the tunable expression system, it was applied to a synthetic circuit displaying three states of fluorescence depending on the difference in protein expression levels. To demonstrate the suitability of application to metabolic engineering, this system was used to diversify the levels of key metabolic enzymes. As a result, expression-optimized strains were capable of producing 2.25 g/L of cadaverine, 2.59 g/L of L-proline, and 95.7 mg/L of 1-propanol. Collectively, the tunable expression system could be utilized to optimize genetic circuits for desired operation and to optimize metabolic fluxes through biosynthetic pathways for enhancing production yields of bioproducts. This tunable system will be useful for studying basic and applied biological sciences in addition to applications in synthetic biology and metabolic engineering.

中文翻译：

独立于下游编码序列的可调基因表达系统

蛋白质表达水平的精细控制构成了合成生物学和代谢工程的主要挑战。然而，翻译起始对下游编码序列 (CDS) 的依赖性掩盖了从 mRNA 序列准确预测蛋白质表达水平。在这里，我们提出了一个包含 24 个表达盒的可调基因表达系统，这些表达盒产生范围从 0.001 到 1 的蛋白质的预定义相对表达水平，而不受下游 CDS 的影响。为了验证可调表达系统的实际效用，它被应用于一个合成电路，根据蛋白质表达水平的差异显示三种荧光状态。为了证明应用于代谢工程的适用性，该系统用于使关键代谢酶的水平多样化。结果，表达优化的菌株能够生产 2.25 g/L 尸胺、2.59 g/L L-脯氨酸和 95.7 mg/L 1-丙醇。总的来说，可调表达系统可用于优化所需操作的遗传回路，并通过生物合成途径优化代谢通量，以提高生物产品的产量。除了在合成生物学和代谢工程中的应用外，该可调系统还可用于研究基础和应用生物科学。可调表达系统可用于优化所需操作的遗传回路，并通过生物合成途径优化代谢通量，以提高生物产品的产量。除了在合成生物学和代谢工程中的应用外，该可调系统还可用于研究基础和应用生物科学。可调表达系统可用于优化所需操作的遗传回路，并通过生物合成途径优化代谢通量，以提高生物产品的产量。除了在合成生物学和代谢工程中的应用外，该可调系统还可用于研究基础和应用生物科学。

更新日期：2020-11-21

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