Chemical Biology of Energy-Dependent Proteolysis in Mitochondria

在化学部生命过程化学项目的这一奖项中，来自凯斯西储大学的艾琳·李博士将开发一种化学生物学方法，在翻译后水平上评估依赖于ATP的蛋白分解在维持线粒体动态平衡方面的贡献。线粒体是维持生命所需能量的重要细胞器。关键的生化反应之一是氧化磷酸化，它经常产生活性氧化物种(ROS)作为副产物。如果任其积累，这些ROS会破坏细胞器中脂类、核酸和蛋白质的化学完整性，从而威胁线粒体的完整性。在线粒体的基质中，蛋白质的质量控制可能是由两种能量依赖的蛋白酶维持的，其中一种被称为ClpXP(酪蛋白溶解蛋白酶复合体XP)。Lee博士的工作将产生人类ClpXP的选择性底物和抑制剂，以评估模型细胞系HeLa中线粒体蛋白裂解产物和完整线粒体的蛋白酶功能。一旦被证明是有效的，这项研究中提出的策略将在生成工具方面获得广泛的应用，以阐明非蛋白酶体ATP依赖的蛋白水解酶在未来维持细胞内稳中所扮演的角色。该项目采用多学科方法来开发选择性活性探针和抑制剂，这些探针和抑制剂将用于解决线粒体中蛋白酶机的生理功能。学生将接受酶动力学、蛋白质化学、液相色谱、质谱学、化学合成、固相肽合成、荧光光谱学、蛋白质纯化和分子克隆方面的培训。这些研究活动为本科生和研究生提供了极好的化学生物学培训机会。李博士团队中的每一名研究生还将参与本科教育，因为每个学生都将与一名本科生研究员一起培训和工作至少一个学期。这一努力旨在为研究生提供教学和指导经验。综上所述，该项目将为学生提供进入研究或科学教育职业所需的技术和社交技能。此外，Lee小组的所有工作人员都是国家青少年体育计划(NYSP)的志愿者，该计划是为大克利夫兰地区10至16岁的低收入家庭的儿童提供的一个暑期计划。Lee小组通过实验家用化学品和干冰来教这些孩子日常生活的化学。这项一年一度的外展活动已经成为Lee小组的传统，本科生和研究生在那里学习更多地参与社区和弱势群体的教育。

In this award from the Chemistry of Life Processes Program in the Chemistry Division, Dr. Irene Lee, from Case Western Reserve University, will develop a chemical biology approach to evaluating the contribution of ATP-dependent proteolysis in the maintenance of mitochondria homeostasis at a post-translational level. The mitochondrion is an important organelle that provides energy to sustain life. One of the key biochemical reactions is oxidative phosphorylation, which frequently generates reactive oxidizing species (ROS) as by-products. If allowed to accumulate, these ROS threaten the integrity of mitochondria by damaging the chemical integrity of lipids, nucleic acids and proteins in the organelles. In the matrix of mitochondria, protein quality control is presumably maintained by two energy dependent proteases, one of which is known as ClpXP (caseinolytic protease complex XP). Dr. Lee's work will generate selective substrates and inhibitors of human ClpXP to allow for evaluation of protease functions in mitochondrial protein lysates and intact mitochondria in the model cell line HeLa. Once proven to be effective, the strategy proposed in this study will find broad applications in generating tools to elucidate the roles played by non-proteasomal ATP-dependent proteases in the maintenance of cellular homeostasis in the future. This project employs a multi-disciplinary approach to develop selective activity probes and inhibitors that will be used to address the physiological functions of a protease machine in mitochondria. Students will acquire training in enzyme kinetics, protein chemistry, liquid chromatography, mass spectrometry, chemical synthesis, solid phase peptide synthesis, fluorescence spectroscopy, protein purification and molecular cloning. These research activities provide excellent training opportunities to undergraduate and graduate students in chemical biology. Each graduate student in Dr. Lee's group will also be involved in undergraduate education, as each student will train and work with an undergraduate researcher for at least one semester. This endeavor is designed to provide graduate students teaching and mentoring experiences. Taken together, this project will provide students with the technical and social skills needed to enter a research or science education career. Additionally, all personnel working in the Lee group serve as volunteers to the National Youth Sports Program (NYSP), a summer program offered to children of low-income families in the Greater Cleveland area between the ages of 10 and 16. The Lee group teaches these children chemistry of daily life through experimentation with household chemicals and dry ice. This annual outreach activity has become a tradition in the Lee group, where undergraduate and graduate students learn to become more involved in the community and education for the underprivileged.