Antioxidant signaling by protein AMPylation

项目摘要 我们发现，预测的失活的假激酶硒蛋白O(SELO)采用了一种非典型蛋白 激酶折叠，但在翻译后修饰中将AMP而不是磷酸转移到蛋白质底物 这就是所谓的AMP化。我们的结果说明了蛋白激酶超家族的催化多功能性，并提示 这种AMP化可能是一种比之前所认识到的更广泛的翻译后修饰。塞洛 定位于线粒体，AMPylates蛋白质参与细胞代谢和氧化还原生物学，并出现 调节一条古老且高度保守的细胞抗氧化信号通路。在高等真核生物中，Selo 含有第21个基因编码的氨基酸，硒半胱氨酸，我们认为它具有氧化还原感受器的功能。 调节SELO活性以应对氧化应激。 虽然活性氧物种是人类生物学的必备部分，但水平升高的特点是 许多疾病状态。例如，活性氧含量升高会导致DNA损伤，这可能 启动致癌转化，导致癌症。此外，氧化还原动态平衡的变化是 与中风、心脏病发作和外周血管疾病等疾病的病理有关，所有 它们是美国发病率和死亡率的主要贡献者。因此，机械地理解 保护细胞免受氧化应激影响的途径之一可能会对人类健康和疾病产生重大影响。 这一建议的主要目标是确定自我依赖的AMP化的分子机制 线粒体蛋白保护细胞免受氧化应激，并调节氧化还原稳态。作为这项工作的一部分 工作中，我们将确定自体催化的底物子集的AMP化的功能后果为 以及SELO活性依赖于氧化还原调节的结构基础。我们预计结果会是 在这里获得的将有可能定义新的细胞调节和氧化还原信号的范例和 可以带来创新的诊断工具或治疗人类疾病的新方法。

Project Summary We have discovered that the predicted inactive pseudokinase selenoprotein O (SelO) adopts an atypical protein kinase fold, yet transfers AMP instead of phosphate to protein substrates in a post translational modification known as AMPylation. Our results illustrate the catalytic versatility of the protein kinase superfamily and suggest that AMPylation may be a more widespread post translational modification than previously appreciated. SelO localizes to the mitochondria, AMPylates proteins involved in cellular metabolism and redox biology, and appears to regulate an ancient and highly conserved cellular antioxidant signaling pathway. In higher eukaryotes, SelO contains the 21st genetically encoded amino acid, selenocysteine, which we propose functions as a redox sensor to regulate SelO activity in response to oxidative stress. Although reactive oxygen species are an obligatory part of human biology, elevated levels are characteristic of many disease states. For example, elevated reactive oxygen species can lead to DNA damage, which can initiate oncogenic transformation leading to cancer. Furthermore, alterations in redox homeostasis are implicated in the pathology of conditions such as stroke, heart attack, and peripheral vascular disease, all of which are major contributors to morbidity and mortality in United States. Therefore, a mechanistic understanding of the pathways that protect cells from oxidative stress could have major impacts on human health and disease. The major goal of this proposal is to determine the molecular mechanisms by which SelO-dependent AMPylation of mitochondrial proteins protects cells from oxidative stress and regulates redox homeostasis. As part of this work, we will determine the functional consequences of SelO-catalyzed AMPylation of a subset of substrates as well as the structural basis for the redox-dependent regulation of SelO activity. We anticipate that the results obtained herein will have the potential to define new paradigms of cellular regulation and redox signaling and could lead to innovative diagnostic tools or novel approaches for the treatment of human diseases.