Molecular Mechanisms of Protein Arginylation

描述（由申请人提供）：信号通路中蛋白质的翻译后修饰是体内几乎所有生理过程中控制因子活化、失活和降解的最关键控制点之一。该提案的目标是定义蛋白质酰基化的分子机制和体内功能，这是一种非常不了解的翻译后修饰，影响估计25%的细胞蛋白质组。精氨酸化在细胞迁移和发育形态发生中起关键作用，并且与细胞代谢、生理学和人类疾病的重大事件有关。我们的初步数据有力地表明，乙酰基化通过快速， 它们的活性和蛋白质相互作用的可逆调节，并构成了一个真正必要的和动态的生物调节剂，但几乎没有什么是已知的底物位点特异性和这种反应的分子机制。该提案将通过一种综合方法解决乙酰基转移酶（ATE 1）的机制及其底物特异性的调节，该方法将为这一神秘的生物学过程提供第一个机制见解。 翻译后修饰我们提出了以下具体目标：（1）确定N-末端N-酰基化的位点特异性和性质;（2）测试完整蛋白质中内部蛋白位点N-酰基化的机制;（3）测试细胞组分对不同ATE 1亚型N-酰基化的速率和位点特异性的影响，并调节其体内活性。总之，拟议的研究将阐明一种新的翻译后修饰的分子机制，具有重要的生物学意义。这些研究将最终实现ATE 1活性和生物靶向的差异调节，这对于探索关键的磷酸化依赖性人类疾病（包括心脏病，癌症，神经退行性疾病和衰老）的治疗潜力至关重要。我们的研究结果将打开新的可能性，功能性的乙酰化分析和靶向操纵的关键蛋白质的乙酰化在正常生理和疾病的基本过程。

DESCRIPTION (provided by applicant): Posttranslational modification of proteins in signal pathways is one of the most pivotal control points in vivo for the activation, inactivation, and degradation of factors governing nearly all physiological processes. The goal of this proposal is to define the molecular mechanism and in vivo functions of protein arginylation, a very poorly understood post-translational modification that affects an estimated 25% of the cellular proteome. Arginylation plays a key role in cell migration and developmental morphogenesis, and is implicated in major events of cell metabolism, physiology, and human disease. Our preliminary data strongly suggest that arginylation regulates its protein targets through rapid and reversible modulation of their activity and protein interactions and constitutes a truly essential and dynamic biological regulator, however virtually nothing is known about the substrate site specificity and the molecular mechanisms of this reaction. This proposal will address the mechanisms of the arginyl transfer enzyme (ATE1) and the regulation of its substrate specificity through an integrated approach that will provide the first mechanistic insights into this enigmatic posttranslational modification. We propose the following specific aims: (1) To determine site specificity and properties of N- terminal arginylation; (2) To test the mechanisms of arginylation at internal protein sites in intact proteins; and (3) To test the effect of cellular components on he rate and site specificity of arginylation by different ATE1 isoforms and modulate its activity in vivo. Together, the proposed studies will elucidate the molecular mechanism of a novel posttranslational modification with major biological significance. These studies will ultimately enable differential modulation of ATE1 activity and biological targeting, essential for exploring is therapeutic potential in critical arginylation-dependent human conditions, including heart disease, cancer, neurodegeneration, and aging. The results of our studies will open new possibilities of functional arginylation analysis and targeted manipulation of arginylation of key proteins during essential processes in normal physiology and disease.