A unique insertion in the CP1 domain of Giardia lamblia leucyl-tRNA synthetase.
A unique insertion in the CP1 domain of Giardia lamblia leucyl-tRNA synthetase.
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DOI：
10.1021/bi801832j
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发表时间：
2009-02
期刊：
影响因子：
2.9
通讯作者：
Xiao-Long Zhou;P. Yao;Liang-Liang Ruan-Liang;B. Zhu;Jun Luo;L. Qu;E. Wang
中科院分区：
文献类型：
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作者：
Xiao-Long Zhou;P. Yao;Liang-Liang Ruan-Liang;B. Zhu;Jun Luo;L. Qu;E. Wang
亮氨酰-tRNA合成酶(Leucyl-tRNA synthetase，LEURS)催化tRNA(Leu)同工酶与亮氨酸的酯化反应。它含有一个大的插入结构域，连接肽1(CP1)，用于氨基酸编辑。在这里，我们克隆了蓝氏贾第鞭毛虫(GlLeuRS)的LEUR编码基因，这是最古老的真核生物之一。从一株大肠杆菌高产菌株中分离纯化了GlLeuRS，并对其性质进行了研究。GlLeuRS的CP1结构域(GlLeuRS-CP1)是一种编辑误充电蓝氏菌tRNA(Leu)(AAG)(GltRNA(Leu))的活性蛋白。在CP1结构域(所谓的49个氨基酸基序)中插入49个氨基酸残基对于GlLeuRS的最佳氨基酰化活性是重要的，并且对GlLeuRS-CP1的编辑能力至关重要。此外，该基序还可以赋予从大肠杆菌中分离的具有编辑缺陷的CP1结构域(EcLeuRS-CP1)的编辑活性。我们还发现GlLeuRS不能挽救酿酒酵母Leus零菌株，这表明这两种LeuRS对tRNA(Leu)的识别模式不同。
Leucyl-tRNA synthetase (LeuRS) catalyzes the esterification of the tRNA(Leu) isoacceptor with leucine. It contains a large insertion domain, connective peptide 1 (CP1), for amino acid editing. Here, we cloned the gene encoding LeuRS from Giardia lamblia (GlLeuRS), one of the most ancient eukaryotes. GlLeuRS was purified from an Escherichia coli overproduction strain, and its properties were investigated. The isolated CP1 domain of GlLeuRS (GlLeuRS-CP1) was an active protein for editing mischarged G. lamblia tRNA(Leu)(AAG) (GltRNA(Leu)). Insertion of 49 amino acid residues within the CP1 domain (the so-called 49-amino acid motif) was important for the optimal aminoacylation activity of GlLeuRS and was crucial for the editing capacity of GlLeuRS-CP1. Additionally, the motif can confer editing activity on the editing-defective isolated CP1 domain from E. coli LeuRS (EcLeuRS-CP1). We also found that GlLeuRS could not rescue a Saccharomyces cerevisiae leuS null strain, suggesting different recognition modes for these two LeuRSs with respect to tRNA(Leu).