Methods to directly inhibit gene expression using small molecules hold promise for the development of new therapeutics targeting proteins that have evaded previous attempts at drug discovery. Among these, small molecules including the drug-like compound PF- 06446846 (PF846) selectively inhibit the synthesis of specific proteins, by stalling translation elongation. These molecules also inhibit translation termination by an unknown mechanism. Using cryo-electron microscopy (cryo-EM) and biochemical approaches, we show that PF846 arrests translation at the stop codon by slowing hydrolysis of the protein nascent chain (NC) from peptidyl-site (P-site) tRNA by eukaryotic release factor 1 (eRF1). After NC hydrolysis from the P-site tRNA, PF846 traps the NC in the ribosome exit tunnel in a compact α-helical conformation that induces 28S rRNA nucleotide rearrangements propagating back to the ribosome peptidyl transferase center (PTC). Mutational analyses and human cell-based experiments elucidate the pivotal amino acids of the NC required for PF846-dependent termination arrest, all of which face the PF846 side of the ribosome exit tunnel. The structural and functional data support a model in which PF846 inhibits translation termination by inducing allosteric conformational rearrangements in the NC and PTC that suppress peptidyl-tRNA hydrolysis promoted by eRF1, and trap the NC in the ribosome exit tunnel. This unprecedented mechanism of action reveals new principles of translation termination and lays the foundation for new therapeutic strategies.

中文翻译：

---

通过捕获核糖体上终止的蛋白质新生链的类药物化合物对人类翻译的选择性抑制

使用小分子直接抑制基因表达的方法有望开发出针对靶向蛋白质的新疗法，而这种疗法已避免了以往的药物发现尝试。其中，包括药物样化合物PF-06446846（PF846）在内的小分子通过阻止翻译伸长来选择性抑制特定蛋白质的合成。这些分子还通过未知机制抑制翻译终止。使用低温电子显微镜（cryo-EM）和生化方法，我们显示PF846通过减慢真核生物释放因子1从肽基位点（P-位点）tRNA的蛋白质新生链（NC）的水解来阻止终止密码子的翻译。 （eRF1）。从P位点tRNA进行NC水解后，PF846以紧凑的α螺旋构型将NC捕获在核糖体出口通道中，该构型诱导了28S rRNA核苷酸重排，并传播回核糖体肽基转移酶中心（PTC）。突变分析和基于人体细胞的实验阐明了PF846依赖性终止终止所需的NC关键氨基酸，所有这些氨基酸都面对核糖体出口通道的PF846一侧。结构和功能数据支持一个模型，其中PF846通过在NC和PTC中诱导变构构象重排来抑制翻译终止，从而抑制eRF1促进的肽基tRNA水解，并将NC捕获在核糖体出口通道中。这种前所未有的作用机制揭示了翻译终止的新原理，并为新的治疗策略奠定了基础。