Substrate binding specificity of enzymatic components of the N-end rule pathway

蛋白质而不是基因是细胞内多种功能的直接执行者。因此，作为所有活细胞的主要成分之一的蛋白质的分析是生物学、生物技术和医学中最重要的任务之一。蛋白质发挥核心作用，例如作为支架或储存组分（细胞骨架、能量储存），作为生物化学催化剂以大力促进生物化学反应，以及作为许多其他蛋白质及其修饰以及信号分子的“生物合成工厂”。保持细胞内所有蛋白质的实体（蛋白质组）处于生理功能平衡至关重要。这意味着1）必须调节蛋白质浓度和丰度，以及2）必要时它们的降解。在我们的实验室里，我们专注于一种非常特殊的降解途径，实际上是一种处理蛋白质的方法。蛋白质识别和降解的整个过程包括细胞生物学和生物化学途径以及各种额外的酶活性蛋白质，并消耗大部分可用的细胞内能量。例如，所谓的N末端规则特异性地识别需要降解的蛋白质，因为它们的作用时间已经过去或者它们变成细胞毒性的。我们专注于这个信号和降解途径，这是泛素蛋白酶体系统的一部分，因此蛋白质的质量控制。对于我们的工作，我们使用目前最好理解的模式植物，小十字花科拟南芥。目前，植物中参与N-末端规则途径蛋白质降解的酶促组分知之甚少，被降解的蛋白质靶点（底物）也很少。目前的研究出版物暗示，这些调节蛋白质稳定性的过程在发育和细胞生物学中起着核心作用。这些过程包括植物的内源分子通讯和环境分子通讯。例如，植物感染病原体，在洪水的情况下，在细胞分裂，种子萌发和储存蛋白质和脂质的分解，即与基础和应用植物科学高度相关的领域，也与植物生物技术和农业高度相关。这里提出的项目涉及分析所谓的E3泛素蛋白连接酶，蛋白水解1，它识别底物并将其降解。我们的研究旨在表征N-末端规则途径的酶的分子识别机制，并首次对植物中这种蛋白质修饰途径的酶组分进行功能分析。

Proteins rather than genes are the direct executers of manifold functions within a cell. The analysis of proteins as one of the main constituents of all living cells is therefore one of the most important tasks in biology, biotechnology and medicine. Proteins play central roles e.g. as scaffold or storage components (cytoskeleton, energy storage), as biochemical catalysts to energetically facilitate biochemical reactions and as "biosynthetic factories" for numerous additional proteins and their modifications as well as signaling molecules. It is crucial to keep the entity of all proteins within a cell (the proteome) in a physiologically functional balance. This implies that 1) the protein concentration and abundance must be regulated and 2) their degradation if necessary. In our lab, we focus on a very specific degradation pathway virtually a way to dispose of proteins. The overall process of protein recognition and degradation comprises cell biological and biochemical pathways and diverse additional enzymatically active proteins and consume a lion's share of available intracellular energy. For example, the so-called N-end rule recognizes specifically proteins that need to be degraded because their time-of-action is past or they turned to be cytotoxic. We focus on this signaling and degradation pathway which is part of the Ubiquitin proteasome system and thus of the protein quality control. For our work, we use the currently best-understood model plant, the small crucifer Arabidopsis thaliana. Currently, only very few in known about enzymatic components involved in protein degradation via the N-end rule pathway in plants and only a small number of protein targets to be degraded, the so-called substrates. Current research publications give hints that these processes regulating protein stability play central roles in developmental and cellular biology. These processes comprise both the endogenous as well as environmental molecular communication of the plant. Examples are amongst plant infection by pathogens, during flooding situations, in cell division, seed germination and breakdown of storage proteins and lipids, i.e. altogether areas with high relevance for basic and applied plant science but also for plant-based biotechnology and agriculture. The here proposed projects deal with the analysis of a so-called E3 Ubiquitin protein ligase, PROTEOLYSIS 1, which recognizes substrates and routing them for degradation. Our studies aim at characterizing the molecular recognition mechanism by enzymes of the N-end rule pathway and by this deliver for the first time a functional analysis of enzymatic components of this protein modification pathway in plants.