Department of Chemistry, Faculty of Science, University of Zagreb, Zagreb 10000, Croatia.
Institute for Clinical Sciences, Faculty of Medicine, Imperial College London and MRC London Institute of Medical Sciences, London, SW7 2AZ, UK.
Nucleic Acids Res. 2022 Apr 22;50(7):4029-4041. doi: 10.1093/nar/gkac207.
Aminoacyl-tRNA synthetases (AARS) translate the genetic code by loading tRNAs with the cognate amino acids. The errors in amino acid recognition are cleared at the AARS editing domain through hydrolysis of misaminoacyl-tRNAs. This ensures faithful protein synthesis and cellular fitness. Using Escherichia coli isoleucyl-tRNA synthetase (IleRS) as a model enzyme, we demonstrated that the class I editing domain clears the non-cognate amino acids well-discriminated at the synthetic site with the same rates as the weakly-discriminated fidelity threats. This unveiled low selectivity suggests that evolutionary pressure to optimize the rates against the amino acids that jeopardize translational fidelity did not shape the editing site. Instead, we propose that editing was shaped to safeguard cognate aminoacyl-tRNAs against hydrolysis. Misediting is prevented by the residues that promote negative catalysis through destabilisation of the transition state comprising cognate amino acid. Such powerful design allows broad substrate acceptance of the editing domain along with its exquisite specificity in the cognate aminoacyl-tRNA rejection. Editing proceeds by direct substrate delivery to the editing domain (in cis pathway). However, we found that class I IleRS also releases misaminoacyl-tRNAIle and edits it in trans. This minor editing pathway was up to now recognized only for class II AARSs.
氨酰-tRNA 合成酶（AARS）通过将 tRNA 与对应的氨基酸结合来翻译遗传密码。在 AARS 编辑结构域中，通过水解错误氨酰-tRNA 来清除氨基酸识别错误。这确保了忠实的蛋白质合成和细胞适应性。我们使用大肠杆菌异亮氨酰-tRNA 合成酶（IleRS）作为模型酶，证明了 I 类编辑结构域能够以与弱分辨保真度威胁相同的速率清除在合成部位得到很好区分的非对应氨基酸。这种低选择性揭示了进化压力并没有塑造编辑部位，以优化对危及翻译保真度的氨基酸的反应速率。相反，我们提出编辑是为了保护对应的氨酰-tRNA 免受水解。错误编辑是通过促进负催化的残基来防止的，通过使包含对应的氨基酸的过渡态不稳定来实现。这种强大的设计允许编辑结构域广泛接受底物，同时对对应的氨酰-tRNA 具有极高的特异性。编辑通过直接将底物递送至编辑结构域（顺式途径）进行。然而，我们发现 I 类 IleRS 还会释放错误氨酰-tRNAIle 并以反式方式对其进行编辑。这种次要的编辑途径到目前为止仅被认为存在于 II 类 AARS 中。
