PDF(Pubmed)
Abstract：
The Ser/Leu-swapped genetic code can act as a genetic firewall, mitigating biohazard risks arising from horizontal gene transfer in genetically modified organisms. Our prior work demonstrated the orthogonality of this swapped code to the standard genetic code using a cell-free translation system comprised of 21 in vitro transcribed tRNAs. In this study, to advance this system for protein engineering, we introduce a natural/in vitro transcribed-hybrid tRNA set. This set combines natural tRNAs from Escherichia coli (excluding Ser, Leu, and Tyr) and in vitro transcribed tRNAs, encompassing anticodon-swapped tRNASerGAG and tRNALeuGGA. This approach reduces the number of in vitro transcribed tRNAs required from 21 to only 4. In this optimized system, the production of a model protein, superfolder green fluorescent protein, increases to 3.5-fold. With this hybrid tRNA set, the Ser/Leu-swapped cell-free translation system will stand as a potent tool for protein production with reduced biohazard concerns in future biological endeavors.
摘要：
Ser/Leu交换的遗传密码可以充当遗传防火墙，减轻转基因生物中水平基因转移引起的生物危害风险。我们先前的工作使用由21个体外转录的tRNA组成的无细胞翻译系统证明了这种交换的代码与标准遗传代码的正交性。在这项研究中,为了推进这个蛋白质工程系统，我们介绍了一个天然/体外转录-杂交的tRNA组。这一组结合了来自大肠杆菌的天然tRNA（不包括Ser，Leu,和Tyr）和体外转录的tRNA，包括反密码子交换的tRNASerGAG和tRNALeuGGA。这种方法将所需的体外转录tRNA的数量从21个减少到仅4个。在这个优化的系统中，模型蛋白质的生产，superfolder绿色荧光蛋白，增加到3.5倍。有了这个杂种tRNA集，Ser/Leu交换的无细胞翻译系统将成为蛋白质生产的有效工具，并减少未来生物学工作中的生物危害问题。
