Pre-transfer editing by class II prolyl-tRNA synthetase - Role of aminoacylation active site in "selective release" of noncognate amino acids
Pre-transfer editing by class II prolyl-tRNA synthetase - Role of aminoacylation active site in "selective release" of noncognate amino acids
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DOI：
10.1074/jbc.m605856200
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发表时间：
2006-09-22
影响因子：
4.8
通讯作者：
Musier-Forsyth, Karin
中科院分区：
文献类型：
--
作者：
Hati, Sanchita;Ziervogel, Brigitte;Musier-Forsyth, Karin
氨酰-tRNA合成酶催化同源氨基酸与特定tRNA分子的连接。为了防止由非同源氨基酸的错误激活引起的蛋白质合成中的潜在错误，一些合成酶已经进化出编辑机制来水解错误激活的氨基酸（转移前编辑）或错误酰化的tRNA（转移后编辑）。在转移后编辑的情况下，合成酶采用与氨基酸活化位点不同的单独的编辑结构域，并且该机制被认为涉及tRNA的柔性CCA-3'末端从合成活性位点到水解位点的穿梭。转移前编辑的机制还不太清楚，在大多数情况下，转移前编辑的确切位点还没有最终确定。在这里，我们探测前转移编辑活动的II类脯氨酰-tRNA合成酶从五个物种代表所有三个王国的生活。为了定位转移前编辑的位点，通过缺失细菌脯氨酰-tRNA合成酶物种的插入结构域特征（其是转移后编辑的位点）或真核和古细菌酶的N-或C-末端延伸结构域来构建截短突变体。此外，还对大肠杆菌脯氨酰-tRNA合成酶的转移前编辑机制进行了详细的探讨。这些研究表明，脯氨酰-tRNA合成酶的转移前编辑不需要单独的编辑结构域。氨酰化活性位点通过充当选择性地将非同源腺苷酸释放到溶液中的过滤器，同时保护同源腺苷酸免于水解，在保持翻译的保真度方面起着重要作用。
Aminoacyl-tRNA synthetases catalyze the attachment of cognate amino acids to specific tRNA molecules. To prevent potential errors in protein synthesis caused by misactivation of noncognate amino acids, some synthetases have evolved editing mechanisms to hydrolyze misactivated amino acids (pre-transfer editing) or misacylated tRNAs (post-transfer editing). In the case of post-transfer editing, synthetases employ a separate editing domain that is distinct from the site of amino acid activation, and the mechanism is believed to involve shuttling of the flexible CCA-3' end of the tRNA from the synthetic active site to the site of hydrolysis. The mechanism of pre-transfer editing is less well understood, and in most cases, the exact site of pre-transfer editing has not been conclusively identified. Here, we probe the pre-transfer editing activity of class II prolyl-tRNA synthetases from five species representing all three kingdoms of life. To locate the site of pre-transfer editing, truncation mutants were constructed by deleting the insertion domain characteristic of bacterial prolyl-tRNA synthetase species, which is the site of post-transfer editing, or the N- or C-terminal extension domains of eukaryotic and archaeal enzymes. In addition, the pre-transfer editing mechanism of Escherichia coli prolyl-tRNA synthetase was probed in detail. These studies show that a separate editing domain is not required for pre-transfer editing by prolyl-tRNA synthetase. The aminoacylation active site plays a significant role in preserving the fidelity of translation by acting as a filter that selectively releases non-cognate adenylates into solution, while protecting the cognate adenylate from hydrolysis.