A role for the 30S subunit E site in maintenance of the translational reading frame
A role for the 30S subunit E site in maintenance of the translational reading frame
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DOI：
10.1261/rna.1320109
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发表时间：
2009-02-01
期刊：
影响因子：
4.5
通讯作者：
Fredrick, Kurt
中科院分区：
文献类型：
--
作者：
Devaraj, Aishwarya;Shoji, Shinichiro;Fredrick, Kurt
出口 (E) 位点与多种核糖体活动有关，包括易位、解码和翻译阅读框的维护。在这里，我们通过在 rpsG 中引入缺失来靶向 30S 亚基 E 位点，从而截断核糖体蛋白 S7 的 β 发夹。这种突变 (S7 Delta R77-Y84) 会增加 -1 和 +1 移码，但不会增加错误编码，这提供了 30S E 位点在帧维护中发挥特定作用的证据。在许多合成环境中，突变 S7 Delta R77-Y84 还在 prfB'-lacZ 翻译过程中刺激 +1 程序性移码。然而，当移码位点的 E 密码子与自然界中发现的密码子相对应时，没有看到任何效果，这表明 E-tRNA 释放通常不会限制 prfB 移码速率。含有 S7 Delta R77-Y84 的核糖体表现出自发反向易位率升高和 E-tRNA 的 K-1/2 增加。这些影响的程度相似，表明两者都是 E-tRNA 不稳定的结果。最后，30S E 位点的突变不会抑制 EF-G 依赖性易位，这与 50S E 位点在该机制中的主要作用一致。
The exit (E) site has been implicated in several ribosomal activities, including translocation, decoding, and maintenance of the translational reading frame. Here, we target the 30S subunit E site by introducing a deletion in rpsG that truncates the beta-hairpin of ribosomal protein S7. This mutation (S7 Delta R77-Y84) increases both -1 and +1 frameshifting but does not increase miscoding, providing evidence that the 30S E site plays a specific role in frame maintenance. Mutation S7 Delta R77-Y84 also stimulates +1 programmed frameshifting during prfB'-lacZ translation in many synthetic contexts. However, no effect is seen when the E codon of the frameshift site corresponds to those found in nature, suggesting that E-tRNA release does not normally limit the rate of prfB frameshifting. Ribosomes containing S7 Delta R77-Y84 exhibit an elevated rate of spontaneous reverse translocation and an increased K-1/2 for E-tRNA. These effects are of similar magnitude, suggesting that both result from destabilization of E-tRNA. Finally, this mutation of the 30S E site does not inhibit EF-G-dependent translocation, consistent with a primary role for the 50S E site in the mechanism.