Antioxidant signaling by protein AMPylation

项目摘要 我们发现，预测的失活假激酶硒蛋白O（SelO）采用非典型蛋白质 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 称为AMPylation。我们的研究结果说明了蛋白激酶超家族的催化多功能性，并建议 AMPylation可能是比以前认识到的更广泛的翻译后修饰。Selo 定位于线粒体，使参与细胞代谢和氧化还原生物学的蛋白质腺苷酸化， 来调节一个古老而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，硒代半胱氨酸，我们建议作为氧化还原传感器的功能 调节SelO活性以应对氧化应激。 虽然活性氧是人体生物学的一个必要组成部分，但活性氧水平升高是 许多疾病状态。例如，活性氧物质的升高会导致DNA损伤， 引发致癌转化导致癌症。此外，氧化还原稳态的改变是 与诸如中风、心脏病发作和外周血管疾病等病症的病理学有关， 这是美国发病率和死亡率的主要原因。因此，机械的理解 保护细胞免受氧化应激的途径可能对人类健康和疾病产生重大影响。 该建议的主要目标是确定SelO依赖性腺苷酸化的分子机制， 线粒体蛋白质保护细胞免受氧化应激，并调节氧化还原稳态。作为其中的一部分 工作，我们将确定SelO催化的底物子集的AMP化的功能后果， 以及SelO活性的氧化还原依赖性调节的结构基础。我们预计， 将有可能定义细胞调节和氧化还原信号传导的新范例， 可能导致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.