A functional relationship between helix 1 and the 900 tetraloop of 16S ribosomal RNA within the bacterial ribosome
A functional relationship between helix 1 and the 900 tetraloop of 16S ribosomal RNA within the bacterial ribosome
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DOI：
10.1261/rna.2160405
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发表时间：
2005-06-01
期刊：
影响因子：
4.5
通讯作者：
Brakier-Gingras, L
中科院分区：
文献类型：
--
作者：
Bélanger, F;Théberge-Julien, G;Brakier-Gingras, L
覆盖 16S 核糖体 RNA (rRNA) 螺旋 27 的保守 900 四环与 16S rRNA 螺旋 24 以及 23S rRNA 螺旋 67 相互作用，形成靠近解码中心的亚基间桥 B2c。在之前的研究中，我们研究了 900 四环和螺旋 24 之间的相互作用如何参与亚基关联和翻译保真度。在本研究中，我们研究了 900 四环是否参与了与大肠杆菌 16S rRNA 不同区域的其他未检测到的相互作用。使用遗传互补方法，我们选择了 16S rRNA 中的突变来补偿 900 四环突变 A900G，该突变严重损害亚基关联和翻译保真度。使用诱变 XL1-Red 大肠杆菌菌株或易错 PCR 策略，在 16S rRNA 中随机引入突变。使用专门的核糖体系统在体内选择功能获得突变。因此，在 16S rRNA 的螺旋 1 中独立选择了两个突变，即 U12 缺失和 U12C 取代。该螺旋位于27号螺旋附近，但不直接接触核糖体晶体结构中的900四环。这两种突变都纠正了 A900G 突变引起的亚基关联和翻译保真度缺陷，揭示了 16S rRNA 这两个区域之间意想不到的功能相互作用。
The conserved 900 tetraloop that caps helix 27 of 16S ribosomal RNA (rRNA) interacts with helix 24 of 16S rRNA and also with helix 67 of 23S rRNA, forming the intersubunit bridge B2c, proximal to the decoding center. in previous studies, we investigated how the interaction between the 900 tetraloop and helix 24 participates in subunit association and translational fidelity. In the present study, we investigated whether the 900 tetraloop is involved in other undetected interactions with different regions of the Escherichia coli 16S rRNA. Using a genetic complementation approach, we selected mutations in 16S rRNA that compensate for a 900 tetraloop mutation, A900G, which severely impairs subunit association and translational fidelity. Mutations were randomly introduced in 16S rRNA, using either a mutagenic XL1-Red E. coli strain or an error-prone PCR strategy. Gain-of-function mutations were selected in vivo with a specialized ribosome system. Two mutations, the deletion of U12 and the U12C substitution, were thus independently selected in helix 1 of 16S rRNA. This helix is located in the vicinity of helix 27, but does not directly contact the 900 tetraloop in the crystal structures of the ribosome. Both mutations correct the subunit association and translational fidelity defects caused by the A900G mutation, revealing an unanticipated functional interaction between these two regions of 16S rRNA.