Genetically Incorporating Two Distinct Post-translational Modifications into One Protein Simultaneously,ACS Synthetic Biology

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Genetically Incorporating Two Distinct Post-translational Modifications into One Protein Simultaneously
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2018-01-17 00:00:00 , DOI: 10.1021/acssynbio.7b00408
Sumana Venkat ₁ , Jourdan Sturges ₁ , Alleigh Stahman ₁ , Caroline Gregory ₁ , Qinglei Gan ₁ , Chenguang Fan ₁

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Post-translational modifications (PTMs) play important roles in regulating a variety of biological processes. To facilitate PTM studies, the genetic code expansion strategy has been utilized to cotranslationally incorporate individual PTMs such as acetylation and phosphorylation into proteins at specific sites. However, recent studies have demonstrated that PTMs actually work together to regulate protein functions and structures. Thus, simultaneous incorporation of multiple distinct PTMs into one protein is highly desirable. In this study, we utilized the genetic incorporation systems of phosphoserine and acetyllysine to install both phosphorylation and acetylation into target proteins simultaneously in Escherichia coli. And we used this system to study the effect of coexisting acetylation and phosphorylation on malate dehydrogenase, demonstrating a practical application of this system in biochemical studies. Furthermore, we tested the mutual orthogonality of three widely used genetic incorporation systems, indicating the possibility of incorporating three distinct PTMs into one protein simultaneously.

中文翻译：

将两个不同的翻译后修饰同时遗传整合到一个蛋白质中

翻译后修饰（PTM）在调节各种生物学过程中起着重要作用。为了促进PTM研究，已使用遗传密码扩展策略将特定的PTM（例如乙酰化和磷酸化）共翻译纳入特定部位的蛋白质中。但是，最近的研究表明，PTM实际上可以协同调节蛋白质的功能和结构。因此，非常希望将多个不同的PTM同时掺入一种蛋白质中。在这项研究中，我们利用磷酸丝氨酸和乙酰赖氨酸的遗传掺入系统将磷酸化和乙酰化同时安装到大肠杆菌中的目标蛋白中。并且我们使用该系统研究了乙酰化和磷酸化共存对苹果酸脱氢酶的影响，证明了该系统在生化研究中的实际应用。此外，我们测试了三个广泛使用的基因整合系统的相互正交性，表明了将三个不同的PTM同时整合到一个蛋白质中的可能性。

更新日期：2018-01-17

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