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利用改良的四酶合成生物体系从乙醇生产 2,3-丁二醇的效率。

Efficient 2,3-Butanediol Production from Ethanol by a Modified Four-Enzyme Synthetic Biosystem.

机构信息

College of Chemical Engineering, Huaqiao University, Xiamen 361021, China.

Institute of Edible Fungi, Fujian Academy of Agricultural Sciences, Fuzhou 350012, China.

出版信息

Molecules. 2024 Aug 20;29(16):3934. doi: 10.3390/molecules29163934.

Abstract

2,3-butanediol (2,3-BD) is a versatile bio-based platform chemical. An artificial four-enzyme synthetic biosystem composed of ethanol dehydrogenase, NADH oxidase, formolase and 2,3-butanediol dehydrogenase was designed for upgrading ethanol to 2,3-BD in our previous study. However, a key challenge in developing in vitro enzymatic systems for 2,3-BD synthesis is the relatively sluggish catalytic efficiency of formolase, which catalyzes the rate-limiting step in such systems. Herein, this study reports how engineering the tunnel and substrate binding pocket of FLS improved its catalytic performance. A series of single-point and combinatorial variants were successfully obtained which displayed both higher catalytic efficiency and better substrate tolerance than wild-type FLS. Subsequently, a cell-free biosystem based on the FLS:I28V/L482E enzyme was implemented for upgrading ethanol to 2,3-BD. Ultimately, this system achieved efficient production of 2,3-BD from ethanol by the fed-batch method, reaching a concentration of 1.39 M (124.83 g/L) of the product and providing both excellent productivity and yield values of 5.94 g/L/h and 92.7%, respectively. Taken together, this modified enzymatic catalysis system provides a highly promising alternative approach for sustainable and cost-competitive production of 2,3-BD.

摘要

2,3-丁二醇(2,3-BD)是一种用途广泛的生物基平台化学品。在我们之前的研究中,设计了一种由乙醇脱氢酶、NADH 氧化酶、甲醛酶和 2,3-丁二醇脱氢酶组成的人工四酶合成生物系统,用于将乙醇升级为 2,3-BD。然而,开发用于 2,3-BD 合成的体外酶系统的一个关键挑战是甲醛酶的催化效率相对较慢,甲醛酶催化该系统中的限速步骤。在此,本研究报告了如何通过工程改造 FLS 的隧道和底物结合口袋来提高其催化性能。成功获得了一系列单点和组合变体,它们显示出比野生型 FLS 更高的催化效率和更好的底物耐受性。随后,基于 FLS:I28V/L482E 酶的无细胞生物系统被用于将乙醇升级为 2,3-BD。最终,该系统通过分批补料法实现了从乙醇到 2,3-BD 的高效生产,达到了 1.39 M(124.83 g/L)的产物浓度,提供了 5.94 g/L/h 和 92.7%的出色生产力和产率值。总之,这种经过修饰的酶催化系统为可持续和具有成本竞争力的 2,3-BD 生产提供了一种极具前景的替代方法。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b349/11357561/00a5eb76378e/molecules-29-03934-g001.jpg

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