Specific interactions of the L10(L12)4 ribosomal protein complex with mRNA, rRNA, and L11
Specific interactions of the L10(L12)4 ribosomal protein complex with mRNA, rRNA, and L11
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DOI：
10.1021/bi701838y
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发表时间：
2008-03-04
期刊：
影响因子：
2.9
通讯作者：
Draper, David E.
中科院分区：
文献类型：
--
作者：
Iben, James R.;Draper, David E.
大型核糖体亚基蛋白L10和L12形成五聚体蛋白复合物，L10(L12)(4)，密切参与核糖体延伸周期。它与rRNA或其他核糖体蛋白的接触仅被晶体学部分解决。在大肠杆菌中，L10和l12是由单个操纵子编码的，其中L10(l12)(4)是识别mRNA先导体中的二级结构的翻译抑制子。在本研究中，L10(L12)(4)从中等嗜热的脂嗜热芽孢杆菌中表达，以定量比较复合物与mRNA和核糖体靶点结合的策略。最小的mRNA识别结构广泛分布于细菌中，并有可能形成类似于rRNA中作为L10(L12)(4)结合位点一部分的扭结结构。两个序列之间相等位置的突变对L10(L12)(4)-RNA结合亲和力有相似的影响，并将kink-turn基序和loop AA序列确定为重要的识别元件。与较大的rRNA结构相反，mRNA明显定位扭结旋转基序和蛋白识别环，而没有依赖于Mg2+的三级结构的好处。mRNA和rRNA片段以相似的亲和力结合L10(L12)(4)（类似于10(8)M-1），但荧光结合研究表明，核糖体附近的蛋白质L11可将L10(L12)(4)的结合增强100倍。因此，mRNA和核糖体靶标使用相似的RNA特征，在不同的结构背景下，识别L10(L12)(4)，核糖体通过额外的蛋白质-蛋白质相互作用确保其L10(L12)(4)结合位点的饱和。
Large ribosomal subunit proteins L10 and L12 forma pentameric protein complex, L10(L12)(4), that is intimately involved in the ribosome elongation cycle. Its contacts with rRNA or other ribosomal proteins have been only partially resolved by crystallography. In Escherichia coli, L10 and L 12 are encoded from a single operon for which L10(L 12)(4) is a translational repressor that recognizes a secondary structure in the mRNA leader. In this study, L10(L12)(4) was expressed from the moderate thermophile Bacillus stearothermophilus to quantitatively compare strategies for binding of the complex to mRNA and ribosome targets. The minimal mRNA recognition structure is widely distributed among bacteria and has the potential to form a kink-turn structure similar to one identified in the rRNA as part of the L10(L12)(4) binding site. Mutations in equivalent positions between the two sequences have similar effects on L10(L12)(4)-RNA binding affinity and identify the kink-turn motif and a loop AA sequence as important recognition elements. In contrast to the larger rRNA structure, the mRNA apparently positions the kink-turn motif and loop for protein recognition without the benefit of Mg2+-dependent tertiary structure. The mRNA and rRNA fragments bind L10(L12)(4) with similar affinity (similar to 10(8) M-1), but fluorescence binding studies show that a nearby protein in the ribosome, L11, enhances L10(L12)(4) binding similar to 100-fold. Thus, mRNA and ribosome targets use similar RNA features, held in different structural contexts, to recognize L10(L12)(4), and the ribosome ensures the saturation of its L10(L12)(4) binding site by means of an additional protein-protein interaction.