Department of Biotechnology and Dr. B C Guha Centre for Genetic Engineering and Biotechnology, University of Calcutta, 35, Ballygunge Circular Road, Kolkata, 700 019, India.
Protein J. 2014 Feb;33(1):48-60. doi: 10.1007/s10930-013-9537-7.
Glutamyl-queuosine-tRNA(Asp) synthetase (Glu-Q-RS) is a paralog of glutamyl-tRNA synthetase (GluRS) and is found in more than forty species of proteobacteria, cyanobacteria, and actinobacteria. Glu-Q-RS shows striking structural similarity with N-terminal catalytic domain of GluRS (NGluRS) but it lacks the C-terminal anticodon binding domain (CGluRS). In spite of structural similarities, Glu-Q-RS and NGluRS differ in their functional properties. Glu-Q-RS glutamylates the Q34 nucleotide of the anticodon of tRNA(Asp) whereas NGluRS constitutes the catalytic domain of GluRS catalyzing the transfer of Glu on the acceptor end of tRNA(Glu). Since NGluRS is able to catalyze aminoacylation of only tRNA(Glu) the glutamylation capacity of tRNA(Asp) by Glu-Q-RS is surprising. To understand the substrate specificity of Glu-Q-RS we undertook a systemic approach by investigating the biophysical and biochemical properties of the NGluRS (1-301), CGluRS (314-471) and Glu-Q-RS-CGluRS, (1-298 of Glu-Q-RS fused to 314-471 from GluRS). Circular dichroism, fluorescence spectroscopy and differential scanning calorimetry analyses revealed absence of N-terminal domain (1-298 of Glu-Q-RS) and C-terminal domain (314-471 from GluRS) communication in chimera, in contrast to the native full length GluRS. The chimeric Glu-Q-RS is still able to aminoacylate tRNA(Asp) but has also the capacity to bind tRNA(Glu). However the chimeric protein is unable to aminoacylate tRNA(Glu) probably as a consequence of the lack of domain-domain communication.
谷氨酰-tRNA（Asp）合成酶（Glu-Q-RS）是谷氨酰-tRNA 合成酶（GluRS）的同源物，存在于四十多种变形菌、蓝藻和放线菌中。Glu-Q-RS 与 GluRS 的 N 端催化结构域（NGluRS）具有显著的结构相似性，但缺乏 C 端反密码子结合结构域（CGluRS）。尽管结构相似，但 Glu-Q-RS 和 NGluRS 在功能特性上有所不同。Glu-Q-RS 使 tRNA（Asp）的反密码子第 34 位核苷酸发生谷氨酰化，而 NGluRS 构成 GluRS 的催化结构域，催化 Glu 转移到 tRNA（Glu）的受体末端。由于 NGluRS 只能催化 tRNA（Glu）的氨酰化，因此 Glu-Q-RS 使 tRNA（Asp）发生谷氨酰化的能力令人惊讶。为了了解 Glu-Q-RS 的底物特异性，我们通过研究 NGluRS（1-301）、CGluRS（314-471）和 Glu-Q-RS-CGluRS（Glu-Q-RS 的 1-298 与 GluRS 的 314-471 融合）的生物物理和生化特性，采取了系统的方法。圆二色性、荧光光谱和差示扫描量热法分析表明，与天然全长 GluRS 相比，嵌合体中不存在 N 端结构域（Glu-Q-RS 的 1-298）和 C 端结构域（GluRS 的 314-471）的交流。该嵌合 Glu-Q-RS 仍能够氨酰化 tRNA（Asp），但也能够结合 tRNA（Glu）。然而，该嵌合蛋白不能氨酰化 tRNA（Glu），可能是由于缺乏结构域-结构域交流。
