Biochemistry and molecular physiology of the protein ABA3

本课题旨在研究拟南芥调控蛋白ABA3的详细功能。在我们之前的工作中，我们已经证明ABA3催化了一种新型的翻译后蛋白修饰：硫酸化激活。ABA3是一种钼辅酶硫酶，可激活两种含钼酶醛氧化酶（AO）和黄嘌呤脱氢酶（XDH）。AO催化脱落酸（ABA）生物合成的最后一步，AO和XDH都产生ROS。在ABA3蛋白的催化下，通过改变无活性XDH和AO分子与活性XDH分子的比值，ABA和ROS的浓度可以迅速增加。本项目的目的有两个：第一部分我们想要详细描述ABA3催化的调节性硫转移的反应机理。这种分子内和分子间硫转移的反应步骤序列是新颖的，没有先例。因此，它将为其他研究小组在哺乳动物、低等真核生物和细菌中破译这种翻译后激活反应提供一个原型，这些反应对它一无所知。在第二部分中，我们打算阐明ABA3的生理作用，同时我们小组已经制定了一些工具和检测方法来研究低丰度蛋白ABA3的表达和活性。我们想要展示ABA3的哪些功能与AO和XDH的激活有关，哪些功能与AO/XDH的激活无关。在这种情况下，我们将分析ABA3的选择性剪接和核定位的重要性，我们已经有了第一个实验适应症。

It is the aim of this project to study the detailed function of the Arabidopsis regulatory protein ABA3. In our previous work, we have shown that ABA3 catalyzes a novel type of posttranslational protein modification: activation by sulfuration. ABA3 is a molybdenum cofactorsulfurase that activates the two Mo-containing enzymes aldehyde oxidase (AO) and xanthine dehydrogenase (XDH). AO catalyzes that last step of abscisic acid (ABA) biosynthesis, and both AO and XDH produce ROS. The concentration of ABA and ROS can be rapidly increased by changing the ratio between inactive and active XDH and AO molecules, as catalyzed by the ABA3 protein. - The aim of this project is twofold: In the first part we want to delineate the detailed reaction mechanism of regulatory sulfur transfer catalyzed by ABA3. This sequence of reaction steps of intra- and intermolecular sulfur-transfer is novel and has no precedent. Thus it will constitute the prototype for other groups to decipher this kind of post-translational activation reactions in mammals, lower eukaryotes and bacteria where nothing is known about it. In the second part, we intend to clarify the physiological role of ABA3 as meanwhile a number of tools and assays has been worked out in our group for studying expression and activity of the low-abundant protein ABA3. We want to show which functions of ABA3 are related to the activation of AO and XDH and which functions are not connected to the activation of AO/XDH. In this context, we will analyze the importance of alternative splicing and of nuclear localization of ABA3 for which we have first experimental indications.