Michael Nina, Huang Bridget Y, Ray Korak Kumar, Kinz-Thompson Colin D, Gonzalez Ruben L
Department of Chemistry, Columbia University, New York, NY 10027, USA.
Current address: MRC Laboratory of Medical Sciences, London, UK.
bioRxiv. 2025 Mar 13:2025.03.09.642146. doi: 10.1101/2025.03.09.642146.
Peptide release factor II (RF2) catalyzes the release of the nascent polypeptide from the bacterial ribosomal complex during translation termination and a subset of ribosome rescue pathways. Despite its critical role, the mechanisms that govern RF2 function and regulation remain elusive. Here, using single-molecule fluorescence energy transfer (smFRET), we characterize the conformational landscape that RF2 explores on the ribosomal complex and show that RF2 binding and dissociation from the ribosome follows a series of conformational rearrangements which depend on its ribosomal binding platform. We also show how further interactions with the ribosomal complex are necessary to properly position RF2 for polypeptide release. This work investigates not only the dynamics RF2 undergoes while in complex with the ribosome, but also identifies a potential mechanism by which the regulation of these dynamics may be disrupted, which may be exploited for future development of broad-spectrum antibiotics.
肽释放因子II（RF2）在翻译终止及核糖体拯救途径的一个子集中，催化新生多肽从细菌核糖体复合物上释放。尽管其作用关键，但调控RF2功能和调节的机制仍不清楚。在此，我们使用单分子荧光能量转移（smFRET）技术，描绘了RF2在核糖体复合物上探索的构象景观，并表明RF2与核糖体的结合和解离遵循一系列构象重排，这取决于其核糖体结合平台。我们还展示了与核糖体复合物的进一步相互作用如何对于将RF2正确定位以进行多肽释放是必要的。这项工作不仅研究了RF2与核糖体结合时所经历的动力学，还确定了一种可能扰乱这些动力学调节的潜在机制，这可能被用于未来广谱抗生素的开发。
