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In vitro selection of RNAs aminoacylated by Escherichia coli leucyl-tRNA synthetase

机译：大肠杆菌亮氨酰tRNA合成酶氨基酰化的RNA的体外选择

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摘要

To investigate systematically the MA sequences necessary for aminoacylation by Escherichia coli leucyl-trRNA synthetase, RNAs with leucylation activity were isolated by in vitro selection from a library of tRNA(Leu) variants possessing randomized sequences in the D-loop, the variable arm, and the T-loop. After two rounds of selection, most of the selected variants showed the following features: (1) the tertiary interaction between nucleotides at positions 15 and 48 was A15-U48; (2) the continuous G18G19 sequence, which is invariant in canonical tRNAs, appeared at the fixed position in the D-loop; and (3) the nucleotide at position 20a in the D-loop was A. These selected nucleotides and their positions, concentrating on the hinge region of tRNA, were identical to those of native tRNA(Leu). In contrast, although the long variable arm is the most characteristic of the tRNA(Leu) structure, the primary and secondary structures were not correlated with the leucylation activity. These findings indicate that A15-U48, A20a, and G18G19 located at specific positions are involved in the tertiary folding of leucine-accepting tRNA molecules. With increases in the selection cycle, the D-loop sequence and the secondary structure of the variable arm became similar to those of tRNA(Leu), suggesting that tRNA(Leu) represents an optimized RNA sequence for leucylation. (C) 1998 Academic Press. [References: 29]
机译：为了系统地研究大肠杆菌亮氨酰-trRNA合成酶进行氨基酰化所必需的MA序列，通过体外选择在D环，可变臂和D环中具有随机序列的tRNA（Leu）变异体文库中分离具有亮氨酸化活性的RNA。 T环。经过两轮选择，大多数选择的变体表现出以下特征：（1）15和48位核苷酸之间的三级相互作用是A15-U48; （2）在标准tRNA中不变的连续G18G19序列出现在D环的固定位置; （3）D-环中20a位的核苷酸为A。这些选择的核苷酸及其位置集中在tRNA的铰链区，与天然tRNA（Leu）相同。相比之下，尽管长的可变臂是tRNA（Leu）结构的最大特征，但一级和二级结构与亮氨酰化活性无关。这些发现表明，位于特定位置的A15-U48，A20a和G18G19与接受亮氨酸的tRNA分子的三级折叠有关。随着选择周期的增加，D环序列和可变臂的二级结构变得与tRNA（Leu）相似，这表明tRNA（Leu）代表了一个优化的RNA序列，可用于白细胞化。 （C）1998年学术出版社。 [参考：29]

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