Airas R K
Department of Biochemistry, University of Turku, Finland.
Eur J Biochem. 1996 Aug 15;240(1):223-31. doi: 10.1111/j.1432-1033.1996.0223h.x.
The magnesium dependences of the ATP/PPi exchange and tRNA aminoacylation of reactions were measured for six aminoacyl-tRNA synthetases (isoleucyl-, tyrosyl- and arginyl-tRNA synthetases from class I, and histidyl-, lysyl- and phenylalanyl-tRNA synthetases from class II). The measured values were subjected to best-fit analyses using sum square error calculations between the data and the calculated curves in order to find the mode of participation of the Mg2+ and to optimize the sets of the kinetic constants. The following four dependences were observed: the class II synthetases require three Mg2+ for the activation reaction (including the one in MgATP), but the class I synthetases require only one Mg2+ (in MgATP); in class II synthetases both MgPPi and Mg2PPi participate in the pyrophosphorolysis of the aminoacyl adenylate. Arginyl-tRNA synthetase from class I also shows a better fit if also Mg2PPi reacts, but in the isoleucyl- and tyrosyl-tRNA synthetases only MgPPi but not Mg2PPi is used in the pyrophosphorolysis. Different synthetases have different requirements for the tRNA-bound Mg2+ and spermidine, independent of the enzyme class. 1-4 Mg2+ or spermidines are required in the best fit models. At the end of the reaction in all the synthetases analysed the dissociation of Mg2+ from the product aminoacyl-tRNA essentially enhances the subsequent dissociation of the aminoacyl-tRNA from the enzyme. The binding of ATP to the E. aminoacyl-tRNA complex also speeds up the dissociation of the aminoacyl-tRNA from most of these enzymes.
针对六种氨酰 - tRNA合成酶（I类的异亮氨酰 - 、酪氨酰 - 和精氨酰 - tRNA合成酶，以及II类的组氨酰 - 、赖氨酰 - 和苯丙氨酰 - tRNA合成酶），测定了ATP/PPi交换反应和tRNA氨酰化反应的镁依赖性。使用数据与计算曲线之间的均方误差计算对测量值进行最佳拟合分析，以确定Mg2+的参与模式并优化动力学常数集。观察到以下四种依赖性：II类合成酶在激活反应中需要三个Mg2+（包括MgATP中的一个），但I类合成酶仅需要一个Mg2+（在MgATP中）；在II类合成酶中，MgPPi和Mg2PPi都参与氨酰腺苷酸的焦磷酸解。如果Mg2PPi也发生反应，I类的精氨酰 - tRNA合成酶也显示出更好的拟合，但在异亮氨酰 - 和酪氨酰 - tRNA合成酶的焦磷酸解中仅使用MgPPi而不使用Mg2PPi。不同的合成酶对tRNA结合的Mg2+和亚精胺有不同的要求，与酶的类别无关。在最佳拟合模型中需要1 - 4个Mg2+或亚精胺。在所有分析的合成酶反应结束时，Mg2+从产物氨酰 - tRNA上的解离基本上增强了随后氨酰 - tRNA从酶上的解离。ATP与E.氨酰 - tRNA复合物的结合也加速了氨酰 - tRNA从这些酶中的大多数上的解离。
