Peripheral insertion modulates the editing activity of the isolated CP1 domain of leucyl-tRNA synthetase
Peripheral insertion modulates the editing activity of the isolated CP1 domain of leucyl-tRNA synthetase
复制标题
外周插入调节亮氨酰-tRNA 合成酶分离 CP1 结构域的编辑活性
DOI：
10.1042/bj20111177
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发表时间：
2011-12-01
影响因子：
4.1
通讯作者：
Wang, En-Duo
中科院分区：
文献类型：
--
作者：
Liu, Ru-Juan;Tan, Min;Wang, En-Duo
在Ia类氨基酰基- trna合成酶中存在一个称为CP1（连接肽I）的大插入结构域，负责转移后编辑。风湿蜱和贾第鞭毛虫的LeuRS（亮氨酸- trna合成酶）在CP1中分别有20个和59个独特的氨基酸插入，这对编辑活性至关重要。通过解析AaLeuRS-CP1[2.4埃（1埃= 0.1 nm）]、G/LeuRS-CP1（2.6埃）和插入缺失突变体AaLeuRS-CP1 Delta 20（2.5埃）的晶体结构，了解这些插入在编辑中的作用。这两个插入都被折叠成位于CP1结构域活性位点入口蛋白对面的外周基序。对接模型和定点突变表明，插入物不与底物相互作用。分子动力学模拟结果表明，完整的CP1比缺乏插入基序的突变体更具活力。综上所述，这些数据表明，没有底物结合位点或活性位点主要结构作用的外周插入可能会调节蛋白质的催化功能，可能来自两个LeuRS CP1s的蛋白质动力学调节。脯氨酸和甘氨酸突变分析的进一步结果旨在降低或增加蛋白质的灵活性，与这一假设是一致的。
A large insertion domain called CP1 (connective peptide I) present in class Ia aminoacyl-tRNA synthetases is responsible for post-transfer editing. LeuRS (leucyl-tRNA synthetase) from Aquifex aeolicus and Giardia lamblia possess unique 20 and 59 amino acid insertions respectively within the CP1 that are crucial for editing activity. Crystal structures of AaLeuRS-CP1 [2.4 angstrom (1 angstrom = 0.1 nm)], G/LeuRS-CP1 (2.6 angstrom) and the insertion deletion mutant AaLeuRS-CP1 Delta 20 (2.5 angstrom) were solved to understand the role of these insertions in editing. Both insertions are folded as peripheral motifs located on the opposite side of the proteins from the active-site entrance in the CP1 domain. Docking modelling and site-directed mutagenesis showed that the insertions do not interact with the substrates. Results of molecular dynamics simulations show that the intact CP1 is more dynamic than its mutant devoid of the insertion motif. Taken together, the data show that a peripheral insertion without a substrate-binding site or major structural role in the active site may modulate catalytic function of a protein, probably from protein dynamics regulation in two respective LeuRS CP1s. Further results from proline and glycine mutational analyses intended to reduce or increase protein flexibility are consistent with this hypothesis.