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通过T7RNA聚合酶引导基编辑在Corynebacterium glutamicum中的蛋白质的持续进化

Qing Wang^1,2, Jiajia You^1,2, Yichen Li^1,2

¹The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.

ACS synthetic biology

|December 19, 2024

概括

一个新的突变发生系统,CgMutaT7,可以在Corynebacterium glutamicum中实现高效的,有针对性的基因进化. 这个系统显著增加了突变频率,用于指导蛋白质进化和功能改进.

科学领域:

分子生物学分子生物学
生物技术是生物技术.
合成生物学合成生物学

背景情况:

定向进化对于蛋白质工程至关重要.
有针对性的突变发生系统对于有效的蛋白质修饰至关重要.
开发强大的系统,以持续进化的细菌,如Corynebacterium glutamicum是必要的.

研究的目的:

开发一种高效和有针对性的突变发生系统,用于Corynebacterium glutamicum的持续基因进化.
为了优化系统的高突变频率和低非目标效应.
为了证明该系统在改善蛋白质功能的实用性.

主要方法:

细胞氨酸脱氨酶和 uracil-DNA 糖酶抑制剂对 T7 RNA 聚合酶的序列融合,以创建 CgMutaT7 系统.
优化了CgMutaT7系统 (结果是CgMutaT7^4).
高通量测序用于分析突变概况和频率.

主要成果:

优化的CgMutaT7^4系统实现了1.12 × 10^4倍的目标基因突变频率的增加.
观察到低的非目标突变率.
在1.8 kb的DNA区域中证实了高效和均的C → T转换.
已经成功地证明了西洛斯异构酶的持续演变,以提高西洛斯的利用率.

结论:

在Corynebacterium glutamicum中,CgMutaT7系统提供了一种有效的工具,用于针对性的,持续的突变发生.
该系统显示了蛋白质功能和表达元件进化的重大潜力.
这项技术可以加速改进酶和生物元件的开发.

关键词:

在Corynebacterium glutamicum的研究中,我们发现在T7RNA聚合酶中,持续的进化,不断的演变.赛蒂丁脱胺酶的作用克西洛酶异构酶是什么?

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