McGarry Kevin G, Walker Sarah E, Wang Huanyu, Fredrick Kurt
Department of Microbiology, The Ohio State University, Columbus, Ohio 43210, USA.
Mol Cell. 2005 Nov 23;20(4):613-22. doi: 10.1016/j.molcel.2005.10.007.
Retention of the reading frame in ribosomal complexes after single-round translocation depends on the acylation state of the tRNA. When tRNA lacking a peptidyl group is translocated to the P site, the mRNA slips to allow re-pairing of the tRNA with a nearby out-of-frame codon. Here, we show that this ribosomal activity results from movement of tRNA into the P/E hybrid state. Slippage of mRNA is suppressed by 3' truncation of the translocated tRNA, increased MgCl2 concentration, and mutation C2394A of the 50S E site, and each of these conditions inhibits P/E-state formation. Mutation G2252U of the 50S P site stimulates mRNA slippage, suggesting that decreased affinity of tRNA for the P/P state also destabilizes mRNA in the complex. The effects of G2252U are suppressed by C2394A, further implicating the P/E state in mRNA destabilization. This work uncovers a functional attribute of the P/E state crucial for understanding translation.
单轮转位后核糖体复合物中阅读框的保留取决于tRNA的酰化状态。当缺乏肽基的tRNA转位到P位点时,mRNA会滑动,以使tRNA与附近的框外密码子重新配对。在此,我们表明这种核糖体活性是由tRNA移动到P/E杂合状态引起的。转位tRNA的3'端截短、MgCl2浓度增加以及50S E位点的C2394A突变会抑制mRNA的滑动,并且这些条件中的每一个都抑制P/E状态的形成。50S P位点的G2252U突变会刺激mRNA滑动,这表明tRNA对P/P状态的亲和力降低也会使复合物中的mRNA不稳定。C2394A抑制了G2252U的作用,进一步表明P/E状态与mRNA不稳定有关。这项工作揭示了P/E状态对于理解翻译至关重要的功能属性。