Institute for Organic Chemistry and Chemical Biology, Center for Biomolecular Magnetic Resonance (BMRZ), Johann Wolfgang Goethe-Universität, Frankfurt am Main 60438, Germany.
Institute of Chemistry and Department Life, Light & Matter, University of Rostock, Rostock 18059, Germany.
Nucleic Acids Res. 2022 Feb 28;50(4):2334-2349. doi: 10.1093/nar/gkac040.
A plethora of modified nucleotides extends the chemical and conformational space for natural occurring RNAs. tRNAs constitute the class of RNAs with the highest modification rate. The extensive modification modulates their overall stability, the fidelity and efficiency of translation. However, the impact of nucleotide modifications on the local structural dynamics is not well characterized. Here we show that the incorporation of the modified nucleotides in tRNAfMet from Escherichia coli leads to an increase in the local conformational dynamics, ultimately resulting in the stabilization of the overall tertiary structure. Through analysis of the local dynamics by NMR spectroscopic methods we find that, although the overall thermal stability of the tRNA is higher for the modified molecule, the conformational fluctuations on the local level are increased in comparison to an unmodified tRNA. In consequence, the melting of individual base pairs in the unmodified tRNA is determined by high entropic penalties compared to the modified. Further, we find that the modifications lead to a stabilization of long-range interactions harmonizing the stability of the tRNA's secondary and tertiary structure. Our results demonstrate that the increase in chemical space through introduction of modifications enables the population of otherwise inaccessible conformational substates.
大量修饰核苷酸扩展了天然 RNA 的化学和构象空间。tRNA 是修饰率最高的 RNA 类。广泛的修饰调节它们的整体稳定性、翻译的保真度和效率。然而，核苷酸修饰对局部结构动力学的影响还没有很好地描述。在这里，我们表明修饰核苷酸的掺入导致大肠杆菌中的 fMet-tRNA 局部构象动力学增加，最终导致整体三级结构稳定。通过 NMR 光谱方法分析局部动力学，我们发现，尽管修饰分子的 tRNA 整体热稳定性更高，但与未修饰 tRNA 相比，局部构象波动增加。因此，与修饰相比，未修饰 tRNA 中单个碱基对的熔解受到高熵罚的限制。此外，我们发现修饰导致长程相互作用稳定，协调 tRNA 的二级和三级结构的稳定性。我们的结果表明，通过引入修饰来增加化学空间可以使原本无法进入的构象亚稳态得以实现。
