在大肠杆菌中使用超级绿色光蛋白介导分泌系统进行非特异性过氧酶的工程Escherichia coli
Xingyu Yan1, Xiaodong Zhang1, Haoran Li1
1State Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, #368 Youyi Road, Wuhan 430062, P. R. China.
JACS Au
|April 26, 2024
概括
研究人员设计了一种新的真菌酶,非特异性过氧酶 (UPO),用于改进化学合成. 这种酶工程策略提高了氧功能化反应的选择性,为可持续的化学转化铺平了道路.
科学领域:
- 生物催化和酶工程 生物催化和酶工程
- 有机化学 有机化学
- 分子生物学分子生物学
背景情况:
- 非特异性过氧酶 (UPO) 是一种能够催化具有挑战性的选择性氧功能化的真菌酶.
- 由于UPO的有限可用性和高效的工程策略,阻碍了它们在化学合成中的更广泛应用.
- 开发强大的异质表达和工程平台对于扩大UPO实用性至关重要.
研究的目的:
- 为了识别和表征一种来自 *Coprinopsis marcescibilis* (*Cma*UPO) 的新型非特异性过氧酶.
- 在不对称的乙氧化中设计*Cma*UPO以增强或逆转酶选择性.
- 为UPO异质表达和工程建立一个高效的分泌系统.
主要方法:
- *Cma*UPO的识别和特征.
- 在大肠杆菌 (Escherichia coli) 中开发了一种超级绿色光蛋白 (GFP) 中介的分泌系统.
- 代和突变发生 (ISM) 针对*Cma*UPO的基质道.
主要成果:
- 两个工程 *Cma*UPO 变种 (T125A/A129G 和 T125A/A129V/A247H/T244A/F243G) 被生成.
- 这些变体显著提高了乙氧化的酶选择性,达到99%*ee* (R) 和84%*ee* (S).
- 通过GFP介导的分泌系统被证明适用于其他UPO (*Aae*UPO, *Cci*UPO, *Pab*UPO-I).
结论:
- 开发的GFP介导分泌系统为UPO异质表达和工程提供了一个强大的平台.
- 工程 *Cma*UPO 变种表现出显著增强的对不对称基化酶的酶选择性.
- 这种方法为UPO在化学转化中的可持续和高效应用提供了巨大的潜力.
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