Institute of Biophysics, Academy of Sciences of the Czech Republic, Brno, Czech Republic.
J Phys Chem B. 2013 May 9;117(18):5540-55. doi: 10.1021/jp401482m. Epub 2013 Apr 24.
The L1 stalk is a prominent mobile element of the large ribosomal subunit. We explore the structure and dynamics of its non-canonical rRNA elements, which include two kink-turns, an internal loop, and a tetraloop. We use bioinformatics to identify the L1 stalk RNA conservation patterns and carry out over 11.5 μs of MD simulations for a set of systems ranging from isolated RNA building blocks up to complexes of L1 stalk rRNA with the L1 protein and tRNA fragment. We show that the L1 stalk tetraloop has an unusual GNNA or UNNG conservation pattern deviating from major GNRA and YNMG RNA tetraloop families. We suggest that this deviation is related to a highly conserved tertiary contact within the L1 stalk. The available X-ray structures contain only UCCG tetraloops which in addition differ in orientation (anti vs syn) of the guanine. Our analysis suggests that the anti orientation might be a mis-refinement, although even the anti interaction would be compatible with the sequence pattern and observed tertiary interaction. Alternatively, the anti conformation may be a real substate whose population could be pH-dependent, since the guanine syn orientation requires protonation of cytosine in the tertiary contact. In absence of structural data, we use molecular modeling to explore the GCCA tetraloop that is dominant in bacteria and suggest that the GCCA tetraloop is structurally similar to the YNMG tetraloop. Kink-turn Kt-77 is unusual due to its 11-nucleotide bulge. The simulations indicate that the long bulge is a stalk-specific eight-nucleotide insertion into consensual kink-turn only subtly modifying its structural dynamics. We discuss a possible evolutionary role of helix H78 and a mechanism of L1 stalk interaction with tRNA. We also assess the simulation methodology. The simulations provide a good description of the studied systems with the latest bsc0χOL3 force field showing improved performance. Still, even bsc0χOL3 is unable to fully stabilize an essential sugar-edge H-bond between the bulge and non-canonical stem of the kink-turn. Inclusion of Mg(2+) ions may deteriorate the simulations. On the other hand, monovalent ions can in simulations readily occupy experimental Mg(2+) binding sites.
L1 茎是大亚基核糖体的一个突出的移动元件。我们探索其非规范 rRNA 元件的结构和动力学，其中包括两个扭结-转角、一个内部环和一个四联体环。我们使用生物信息学来识别 L1 茎 RNA 的保守模式，并对一组系统进行了超过 11.5 μs 的 MD 模拟，这些系统范围从孤立的 RNA 构建块到 L1 茎 rRNA 与 L1 蛋白和 tRNA 片段的复合物。我们表明，L1 茎四联体环具有不寻常的 GNNA 或 UNNG 保守模式，偏离了主要的 GNRA 和 YNMG RNA 四联体家族。我们认为这种偏离与 L1 茎内的一个高度保守的三级接触有关。现有的 X 射线结构仅包含 UCCG 四联体环，此外，它们在鸟嘌呤的取向（反式与顺式）上也有所不同。我们的分析表明，反式取向可能是一种错误的细化，尽管即使是反式相互作用也与序列模式和观察到的三级相互作用兼容。或者，反式构象可能是一种真实的亚稳态，其种群可能依赖于 pH，因为在三级接触中，鸟嘌呤顺式取向需要胞嘧啶的质子化。在没有结构数据的情况下，我们使用分子建模来探索在细菌中占主导地位的 GCCA 四联体环，并表明 GCCA 四联体环在结构上与 YNMG 四联体环相似。扭结-转角 Kt-77 因其 11 个核苷酸的凸起而不同寻常。模拟表明，长凸起是茎特异性的 8 个核苷酸插入到共识扭结中，仅略微改变其结构动力学。我们讨论了螺旋 H78 的可能进化作用和 L1 茎与 tRNA 相互作用的机制。我们还评估了模拟方法。模拟提供了对研究系统的良好描述，最新的 bsc0χOL3 力场显示出改进的性能。尽管如此，即使是 bsc0χOL3 也不能完全稳定扭结-转角的凸起和非规范茎之间的必需糖边缘氢键。加入镁离子可能会使模拟恶化。另一方面，单价离子在模拟中可以很容易地占据实验性镁离子结合位点。
