In bacteria, trans-translation is the major quality control system for rescuing stalled ribosomes. It is mediated by tmRNA, a hybrid RNA with properties of both a tRNA and a mRNA, and the small protein SmpB. Because trans-translation is absent in eukaryotes but necessary for bacterial fitness or survival, it is a promising target for the development of novel antibiotics. To facilitate screening of chemical libraries, various reliable in vitro and in vivo systems have been created for assessing trans-translational activity. However, the aim of the current work was to permit the safe and easy in vitro evaluation of trans-translation from pathogenic bacteria, which are obviously the ones we should be targeting. Based on green fluorescent protein (GFP) reassembly during active trans-translation, we have created a cell-free assay adapted to the rapid evaluation of trans-translation in ESKAPE bacteria, with 24 different possible combinations. It can be used for easy high-throughput screening of chemical compounds as well as for exploring the mechanism of trans-translation in these pathogens.

中文翻译：

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ESKAPE 致病菌 tmRNA-SmpB 介导的转译的安全、简便的体外评价

在细菌中，反式翻译是挽救停滞的核糖体的主要质量控制系统。它由 tmRNA（一种兼具 tRNA 和 mRNA 特性的杂合 RNA）和小蛋白 SmpB 介导。因为在真核生物中不存在反式翻译，但对于细菌适应性或生存是必需的，因此它是开发新型抗生素的有希望的目标。为了促进化学文库的筛选，已经创建了各种可靠的体外和体内系统来评估反式翻译活性。然而，当前工作的目的是允许对反式进行安全和简单的体外评估- 病原菌的翻译，这显然是我们应该针对的。基于主动反式翻译过程中的绿色荧光蛋白 (GFP) 重组，我们创建了一种无细胞测定法，适用于 ESKAPE 细菌中反式翻译的快速评估，具有 24 种不同的可能组合。它可用于简单的化合物的高通量筛选以及探索这些病原体的反式翻译机制。