Abstract：
Isoleucyl-tRNA synthetase (IleRS) links isoleucine to cognate tRNA via the Ile-AMP intermediate. Non-cognate valine is often mistakenly recognized as the IleRS substrate; therefore, to maintain the accuracy of translation, IleRS hydrolyzes Val-AMP within the synthetic site (pre-transfer editing). As this activity is not efficient enough, Val-tRNAIle is formed and hydrolyzed in the distant post-transfer editing site. A strictly conserved synthetic site residue Gly56 was previously shown to safeguard Ile-to-Val discrimination during aminoacyl (aa)-AMP formation. Here, we show that the Gly56Ala variant lost its specificity in pre-transfer editing, confirming that this residue ensures the selectivity of all synthetic site reactions. Moreover, we found that the Gly56Ala mutation affects IleRS interaction with aa-tRNA likely by disturbing tRNA-dependent communication between the two active sites.
摘要：
异亮氨酸-tRNA合成酶(IleRS)通过Ile-AMP中间体将异亮氨酸与同源tRNA连接。非同源缬氨酸经常被错误地识别为IleRS底物；因此，为了保持翻译的准确性，IleRS在合成位点内水解Val-AMP(转移前编辑)。由于这项活动不够有效，Val-tRNAIle在远处的转移后编辑位点中形成并水解。先前已证明严格保守的合成位点残基Gly56可在氨酰基（aa）-AMP形成过程中保护Ile到Val的区别。这里,我们表明Gly56Ala变体在转移前编辑中失去了特异性，确认该残基确保所有合成位点反应的选择性。此外，我们发现Gly56Ala突变影响IleRS与aa-tRNA的相互作用,可能是通过干扰两个活性位点之间依赖tRNA的通讯.
