美国波士顿学院Abhishek Chatterjee课题组发现哺乳动物细胞中病毒辅助的强化抑制tRNA的定向演化。这一研究成果于2022年12月22日在线发表在国际学术期刊《自然—方法学》上。

Title: Virus-assisted directed evolution of enhanced suppressor tRNAs in mammalian cells

Author: Jewel, Delilah, Kelemen, Rachel E., Huang, Rachel L., Zhu, Zeyu, Sundaresh, Bharathi, Cao, Xiaofu, Malley, Kaitlin, Huang, Zeyi, Pasha, Muhammad, Anthony, Jon, van Opijnen, Tim, Chatterjee, Abhishek

Issue&Volume: 2022-12-22

Abstract: Site-specific incorporation of unnatural amino acids (Uaas) in living cells relies on engineered aminoacyl-transfer RNA synthetase–tRNA pairs borrowed from a distant domain of life. Such heterologous suppressor tRNAs often have poor intrinsic activity, presumably due to suboptimal interaction with a non-native translation system. This limitation can be addressed in Escherichia coli using directed evolution. However, no suitable selection system is currently available to do the same in mammalian cells. Here we report virus-assisted directed evolution of tRNAs (VADER) in mammalian cells, which uses a double-sieve selection scheme to facilitate single-step enrichment of active yet orthogonal tRNA mutants from naive libraries. Using VADER we developed improved mutants of Methanosarcina mazei pyrrolysyl-tRNA, as well as a bacterial tyrosyl-tRNA. We also show that the higher activity of the most efficient mutant pyrrolysyl-tRNA is specific for mammalian cells, alluding to an improved interaction with the unique mammalian translation apparatus.

DOI: 10.1038/s41592-022-01706-w

Source: https://www.nature.com/articles/s41592-022-01706-w