Chemical Biology of Energy-Dependent Proteolysis in Mitochondria

线粒体能量依赖性蛋白水解的化学生物学

• 批准号: 1213175
• 负责人: Irene Lee
• 金额: $ 27万
• 依托单位: Case Western Reserve University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-01 至 2015-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1213175&HistoricalAwards=false
• 关键词: Chemical; Biology; Energy; Dependent; Proteolysis

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.

来自凯斯西储大学的Irene Lee博士将开发一种化学生物学方法来评估ATP依赖性蛋白水解在维持线粒体稳态中的贡献翻译后水平。线粒体是一种重要的细胞器，为生命提供能量。其中一个关键的生化反应是氧化磷酸化，它经常产生活性氧化物质（ROS）作为副产物。如果允许其积累，这些ROS会通过破坏细胞器中脂质、核酸和蛋白质的化学完整性来威胁线粒体的完整性。在线粒体基质中，蛋白质质量控制大概由两种能量依赖性蛋白酶维持，其中一种称为ClpXP（酪蛋白分解蛋白酶复合物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中线粒体蛋白裂解物和完整线粒体中的蛋白酶功能。一旦被证明是有效的，在这项研究中提出的策略将发现广泛的应用在生成工具，以阐明所发挥的作用，非蛋白酶体ATP依赖性蛋白酶在维持细胞的稳态在未来。该项目采用多学科方法开发选择性活性探针和抑制剂，用于解决线粒体中蛋白酶机器的生理功能。学生将获得酶动力学，蛋白质化学，液相色谱，质谱，化学合成，固相肽合成，荧光光谱，蛋白质纯化和分子克隆的培训。这些研究活动为化学生物学的本科生和研究生提供了极好的培训机会。 李博士小组的每个研究生也将参与本科教育，因为每个学生将与一名本科研究人员一起培训和工作至少一个学期。这项奋进旨在为研究生提供教学和指导经验。 总的来说，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，在李氏集团工作的所有人员都是国家青年体育计划（NYSP）的志愿者，这是一个为大克利夫兰地区10至16岁的低收入家庭儿童提供的暑期计划。Lee小组通过家用化学品和干冰的实验来教授这些孩子日常生活中的化学。这项一年一度的外展活动已成为李氏集团的传统，本科生和研究生在这里学习更多地参与社区和弱势群体的教育。