Didemnin B and ternatin-4 differentially inhibit conformational changes in eEF1A required for aminoacyl-tRNA accommodation into mammalian ribosomes.
Didemnin B and ternatin-4 differentially inhibit conformational changes in eEF1A required for aminoacyl-tRNA accommodation into mammalian ribosomes.
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DOI：
10.7554/elife.81608
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发表时间：
2022-10-20
期刊：
影响因子：
7.7
通讯作者：
Blanchard SC
中科院分区：
文献类型：
--
作者：
Juette MF;Carelli JD;Rundlet EJ;Brown A;Shao S;Ferguson A;Wasserman MR;Holm M;Taunton J;Blanchard SC
快速准确的mRNA翻译需要将正确的氨酰-tRNA（aa-tRNA）有效地密码子依赖性递送到核糖体A位点。在哺乳动物中，这一决定性反应由GTP延伸因子-1 α（eEF 1A）促进，其作为eEF 1A（GTP）-aa-tRNA三元复合物护送aa-tRNA进入核糖体。结构上不相关的环肽didemnin B和ternatin-4与eEF 1A（GTP）-aa-tRNA三元复合物结合并抑制翻译，但在体外和体内对蛋白质合成具有不同的影响。在这里，我们采用单分子荧光成像和低温电子显微镜来确定这些天然产物如何抑制哺乳动物核糖体的翻译延伸。通过与eEF 1A上的共同位点结合，didemnin B和ternatin-4将eEF 1A捕获在aa-tRNA选择的中间状态，阻止eEF 1A释放和核糖体上的aa-tRNA容纳。我们还表明，didemnin B和ternatin-4表现出不同的影响动态的aa-tRNA选择，通知观察到的差异，其抑制效果和生理影响。这些综合研究结果强调了动力学测量在评估小分子抑制机制方面的价值，并强调了单分子方法揭示不同天然产物如何差异化地影响人类翻译机制的潜力。
Rapid and accurate mRNA translation requires efficient codon-dependent delivery of the correct aminoacyl-tRNA (aa-tRNA) to the ribosomal A site. In mammals, this fidelity-determining reaction is facilitated by the GTPase elongation factor-1 alpha (eEF1A), which escorts aa-tRNA as an eEF1A(GTP)-aa-tRNA ternary complex into the ribosome. The structurally unrelated cyclic peptides didemnin B and ternatin-4 bind to the eEF1A(GTP)-aa-tRNA ternary complex and inhibit translation but have different effects on protein synthesis in vitro and in vivo. Here, we employ single-molecule fluorescence imaging and cryogenic electron microscopy to determine how these natural products inhibit translational elongation on mammalian ribosomes. By binding to a common site on eEF1A, didemnin B and ternatin-4 trap eEF1A in an intermediate state of aa-tRNA selection, preventing eEF1A release and aa-tRNA accommodation on the ribosome. We also show that didemnin B and ternatin-4 exhibit distinct effects on the dynamics of aa-tRNA selection that inform on observed disparities in their inhibition efficacies and physiological impacts. These integrated findings underscore the value of dynamics measurements in assessing the mechanism of small-molecule inhibition and highlight potential of single-molecule methods to reveal how distinct natural products differentially impact the human translation mechanism.