Designing Modular Cell-free Systems for Tunable Biotransformation of l-phenylalanine to Aromatic Compounds.
Designing Modular Cell-free Systems for Tunable Biotransformation of l-phenylalanine to Aromatic Compounds.
复制标题
设计模块化无细胞系统，将 L-苯丙氨酸生物转化为芳香族化合物
DOI：
10.3389/fbioe.2021.730663
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发表时间：
2021
影响因子：
5.7
通讯作者：
Li J
中科院分区：
文献类型：
--
作者：
Yang C;Liu Y;Liu WQ;Wu C;Li J
无细胞系统已被用于通过体外表达酶的重组来合成化学物质。然而，在一锅无细胞反应中，由于资源限制/竞争（例如，能量），多种酶的共表达来重建长酶途径经常是有问题的。为了解决这一限制，我们设计了一个模块化的无细胞平台，以(S)-1-苯基-1,2-乙二醇（(S)-PED）和2-苯乙醇（2-PE）为模型，构建可调合成增值芳香族化合物的长生物合成途径。最初，所有参与生物合成途径的酶都被大肠杆菌为基础的无细胞蛋白合成（CFPS）系统单独表达，并证实了它们的催化活性。然后，三组酶在三个无细胞模块中共表达，每个模块都具有完成部分途径的能力。最后，通过混合两个相关模块，重构完整的生物合成途径，分别合成(S)-PED和2-PE。优化后，以l-苯丙氨酸为起始底物，(S)-PED和2-PE的最终转化率分别达到100和82.5%。我们预计，模块化的无细胞方法将使高附加值化学品的高效高产生物合成成为可能。
Cell-free systems have been used to synthesize chemicals by reconstitution of in vitro expressed enzymes. However, coexpression of multiple enzymes to reconstitute long enzymatic pathways is often problematic due to resource limitation/competition (e.g., energy) in the one-pot cell-free reactions. To address this limitation, here we aim to design a modular, cell-free platform to construct long biosynthetic pathways for tunable synthesis of value-added aromatic compounds, using (S)-1-phenyl-1,2-ethanediol ((S)-PED) and 2-phenylethanol (2-PE) as models. Initially, all enzymes involved in the biosynthetic pathways were individually expressed by an E. coli-based cell-free protein synthesis (CFPS) system and their catalytic activities were confirmed. Then, three sets of enzymes were coexpressed in three cell-free modules and each with the ability to complete a partial pathway. Finally, the full biosynthetic pathways were reconstituted by mixing two related modules to synthesize (S)-PED and 2-PE, respectively. After optimization, the final conversion rates for (S)-PED and 2-PE reached 100 and 82.5%, respectively, based on the starting substrate of l-phenylalanine. We anticipate that the modular cell-free approach will make a possible efficient and high-yielding biosynthesis of value-added chemicals.
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影响因子：
8.4
作者：
Karim, Ashty S.;Jewett, Michael C.
通讯作者：
Jewett, Michael C.
影响因子：
4.7
作者：
Li, Jian;Lawton, Thomas J.;Jewett, Michael C.
通讯作者：
Jewett, Michael C.
无细胞的合成生物学，用于药物天然产物的体外生物合成。
影响因子：
4.8
作者：
Li J;Zhang L;Liu W
通讯作者：
Liu W
影响因子：
4.7
作者：
Moore SJ;Lai HE;Chee SM;Toh M;Coode S;Chengan K;Capel P;Corre C;de Los Santos EL;Freemont PS
通讯作者：
Freemont PS
DOI：
10.1002/cbic.201700653
发表时间：
2018-04-04
期刊：
Chembiochem : a European journal of chemical biology
影响因子：
--
作者：
Corrado ML;Knaus T;Mutti FG
通讯作者：
Mutti FG