Identification and characterization of a tRNA decoding the rare AUA codon in Haloarcula marismortui
Identification and characterization of a tRNA decoding the rare AUA codon in Haloarcula marismortui
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DOI：
10.1261/rna.795508
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发表时间：
2008-01-01
期刊：
影响因子：
4.5
通讯作者：
Rajbhandary, Uttam L.
中科院分区：
文献类型：
--
作者：
Kohrer, Caroline;Srinivasan, Gayathri;Rajbhandary, Uttam L.
极端嗜盐古菌Haloarcula marismortui的完整基因组的注释不包括用于翻译AUA的tRNA，AUA是异亮氨酸的罕见密码子。这是迄今为止测序的大多数古细菌基因组的典型情况。基于计算分析，最近已经提出，单个含内含子的tRNA基因通过选择性剪接产生两种非常相似但功能不同的tRNA; UGG-解码tRNA（CCA）（Trp）和AUA-解码tRNA（UAU）（Ile）。通过对H. marismortui，我们已经证实了tRNA（CCA）（Trp）的存在，但没有发现tRNA（UAU）（Ile）存在的证据。相反，我们已经表明，一个tRNA，目前被注释为延长子甲硫氨酸tRNA，并含有CAU作为反密码子，在体内被异亮氨酸氨酰化，这种tRNA代表缺失的异亮氨酸tRNA。有趣的是，这种tRNA在反密码子中携带C34的碱基修饰，不同于在真细菌中发现的众所周知的赖氨酸，其将tRNA的氨基酸身份从甲硫氨酸转换为异亮氨酸，并且其解码特异性从AUG转换为AUA。本文所描述的方法用于鉴定H. marismortui提供了实验确认细胞中任何tRNA存在所必需的工具，从而测试tRNA基因的计算预测。
Annotation of the complete genome of the extreme halophilic archaeon Haloarcula marismortui does not include a tRNA for translation of AUA, the rare codon for isoleucine. This is a situation typical for most archaeal genomes sequenced to date. Based on computational analysis, it has been proposed recently that a single intron-containing tRNA gene produces two very similar but functionally different tRNAs by means of alternative splicing; a UGG-decoding tRNA(CCA)(Trp) and an AUA-decoding tRNA(UAU)(Ile). Through analysis of tRNAs from H. marismortui, we have confirmed the presence of tRNA(CCA)(Trp), but found no evidence for the presence of tRNA(UAU)(Ile). Instead, we have shown that a tRNA, currently annotated as elongator methionine tRNA and containing CAU as the anticodon, is aminoacylated with isoleucine in vivo and that this tRNA represents the missing isoleucine tRNA. Interestingly, this tRNA carries a base modification of C34 in the anticodon different from the well-known lysidine found in eubacteria, which switches the amino acid identity of the tRNA from methionine to isoleucine and its decoding specificity from AUG to AUA. The methods described in this work for the identification of individual tRNAs present in H. marismortui provide the tools necessary for experimentally confirming the presence of any tRNA in a cell and, thereby, to test computational predictions of tRNA genes.