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工程大肠杆菌全细胞生物催化剂从乳酸高效生产乙酰丁酮。

Efficient production of acetoin from lactate by engineered Escherichia coli whole-cell biocatalyst.

机构信息

Department of Biochemical Engineering, School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.

Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (MOE), School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.

出版信息

Appl Microbiol Biotechnol. 2023 Jun;107(12):3911-3924. doi: 10.1007/s00253-023-12560-x. Epub 2023 May 13.

Abstract

Acetoin, an important and high-value added bio-based platform chemical, has been widely applied in fields of foods, cosmetics, chemical synthesis, and agriculture. Lactate is a significant intermediate short-chain carboxylate in the anaerobic breakdown of carbohydrates that comprise ~ 18% and ~ 70% in municipal wastewaters and some food processing wastewaters, respectively. In this work, a series of engineered Escherichia coli strains were constructed for efficient production of acetoin from cheaper and abundant lactate through heterogenous co-expression of fusion protein (α-acetolactate synthetase and α-acetolactate decarboxylase), lactate dehydrogenase and NADH oxidase, and blocking acetate synthesis pathways. After optimization of whole-cell bioconversion conditions, the engineered strain BL-11 produced 251.97 mM (22.20 g/L) acetoin with a yield of 0.434 mol/mol in shake flasks. Moreover, a titer of 648.97mM (57.18 g/L) acetoin was obtained in 30 h with a yield of 0.484 mol/mol lactic acid in a 1-L bioreactor. To the best of our knowledge, this is the first report on the production of acetoin from renewable lactate through whole-cell bioconversion with both high titer and yield, which demonstrates the economy and efficiency of acetoin production from lactate. Key Points • The lactate dehydrogenases from different organisms were expressed, purified, and assayed. • It is the first time that acetoin was produced from lactate by whole-cell biocatalysis. • The highest titer of 57.18 g/L acetoin was obtained with high theoretical yield in a 1-L bioreactor.

摘要

乙酰丙酮是一种重要的高附加值生物基平台化学品,广泛应用于食品、化妆品、化学合成和农业等领域。乳酸是碳水化合物厌氧分解的重要中间短链羧酸,分别占城市废水中的18%和一些食品加工废水中的70%。在这项工作中,构建了一系列工程大肠杆菌菌株,通过异源共表达融合蛋白(α-乙酰乳酸合酶和α-乙酰乳酸脱羧酶)、乳酸脱氢酶和 NADH 氧化酶,并阻断乙酸合成途径,从更便宜、更丰富的乳酸高效生产乙酰丙酮。在优化全细胞生物转化条件后,工程菌株 BL-11 在摇瓶中产生 251.97mM(22.20g/L)乙酰丙酮,产率为 0.434mol/mol。此外,在 1L 生物反应器中,以乳酸计,乙酰丙酮的浓度达到 648.97mM(57.18g/L),产率为 0.484mol/mol,反应时间为 30h。据我们所知,这是首次通过全细胞生物转化从可再生乳酸生产高浓度和高得率乙酰丙酮的报道,证明了从乳酸生产乙酰丙酮的经济性和高效性。

关键点

  • 不同来源的乳酸脱氢酶被表达、纯化并进行了酶活测定。

  • 这是首次通过全细胞生物催化从乳酸生产乙酰丙酮。

  • 在 1L 生物反应器中获得了 57.18g/L 的最高乙酰丙酮浓度和高理论产率。

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