smFRET study of rRNA dimerization at the peptidyl transfer center.
smFRET study of rRNA dimerization at the peptidyl transfer center.
复制标题
DOI：
10.1016/j.bpc.2021.106657
复制
发表时间：
2021-10
影响因子：
3.8
通讯作者：
Wang Y
中科院分区：
文献类型：
--
作者：
Xu D;Wang Y
核糖体是一种核酶。在180 nt的肽基转移中心（PTC）处，两个环（A-环和P-环）与tRNA结合，并将它们定位成紧密接近以进行有效的肽基连接。PTC中也存在2重旋转对称性，这表明现代核糖体的前体可能通过二聚化和基因融合而出现。然而，证明可能的二聚化的实验尚未发表。在我们的研究中，我们报告了两个RNA片段的单分子FRET研究，产生高FRET值。通过在3 '-末端标记5'-生物素化的rRNA分子，或标记三种不同类型的tRNA样寡核苷酸，我们观察到RNA支架可以组装并使几个短的tRNA受体结构域类似物，但不是全长tRNA，非常接近。Mg 2+和连续的3-way连接基序对这个过程是必不可少的，但不需要将氨基酸加载到tRNA类似物上。我们通过天然凝胶迁移实验观察到RNA二聚体。这些实验支持可能存在RNA二聚体或多聚体形式的原核糖体。
The ribosome is a ribozyme. At the peptidyl transfer center (PTC) of 180 nt, two loops (the A- and P- loops) bind to tRNAs and position them in close proximity for efficient peptidyl ligation. There is also a 2-fold rotational symmetry in the PTC, which suggests that the precursor of the modern ribosome possibly emerged through dimerization and gene fusion. However, experiments that demonstrate the possible dimerization have not yet been published. In our investigation, we reported single molecule FRET studies of two RNA fragments that generated high FRET values. By labeling the 5’-biotinylated rRNA molecules at the 3’- terminals, or labeling three different types of tRNA-like oligos, we observed that RNA scaffolds can assemble and bring several short tRNA-acceptor-domain analogs, but not full-length tRNAs, to close proximity. Mg2+ and continuous 3-way junction motifs are essential to this process, but amino acid charging to the tRNA analogs is not required. We observed RNA dimers via native gel-shifting experiments. These experiments support the possible existence of a proto-ribosome in the form of an RNA dimer or multimer.