Functional Dynamics and Energy Coupling Mechanisms of Mitochondrial Membrane Proteins

线粒体膜蛋白的功能动力学和能量耦合机制

• 批准号: 1330695
• 负责人: Nathan Alder
• 金额: $ 42.14万
• 依托单位: University of Connecticut
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1330695&HistoricalAwards=false
• 关键词: Functional; Dynamics; Energy; Coupling; Mechanisms

INTELLECTUAL MERITMitochondria not only produce the vast majority of energy within eukaryotic cells, but also regulate cellular processes such as calcium homeostasis, lipid synthesis, and metabolite transport. The vital functions carried out by this organelle depend directly on the potential energy that is stored across its inner membrane in the form of a proton electrochemical gradient. Characteristic of all energy-conserving membranes, this potential is parsed as an electric field and a difference in proton activity across the lipid bilayer. This project will address two fundamental features of the mitochondrial inner membrane electrochemical potential. First, the changes in membrane protein structure that are driven by alterations in the energized state of the inner membrane will be investigated. Novel, high resolution fluorescence-based approaches will be employed to elucidate the manner in which alterations in the proton gradient and transmembrane electric field differentially change the conformation of a model mitochondria-resident protein. Elucidating the basis of such electromechanical coupling is vital to understanding how proteins in energy-conserving membranes harness the electrochemical potential to perform cellular work. Second, the energetic landscape of the topologically complex inner membrane will be investigated. Using precisely targeted pH-sensing and electrochromic probes to measure localized ion gradients and electric fields, the temporal and spatial heterogeneity of the inner membrane energetic profile will be measured with unprecedented resolution. By challenging the current dogma of electrochemical potential equilibrium across the membrane regions, this work is poised to create a new paradigm for understanding the energetic regulation of processes such as ATP production and local changes in the bilayer structure. These advances in the current understanding of membrane bioenergetics will be made possible by the technical innovations used in this research.BROADER IMPACTSThis NSF-sponsored work will involve two teams of researchers in a model designed to promote multidisciplinary interactions and development of leadership in the sciences, emphasizing the involvement of students traditionally underrepresented in the STEM fields. The work will also serve to further develop and expand novel research tools in the construction of model membrane systems and in the process of site-specific polypeptide labeling, all of which will be made available to the scientific community. The education outreach component of this project is based on the Biology Summer Institute series in cooperation with the University of Connecticut Early College Experience Program. This series of modular classroom and laboratory courses offers professional development to teachers from regional high schools, exposing them to advances in research and current scientific topics, which can be integrated into their own curricula during the subsequent academic year. As this program develops, the Institute will be expanded to include summer courses for high school students as well. Taken together, this education outreach will be highly integrated with the research activities of the project. Moreover, the outcomes of this pedagogical outreach model will be disseminated in educational publications and conference workshops as a means of effectively helping to raise scientific literacy of the public.

线粒体不仅在真核细胞中产生绝大多数的能量，而且还调节细胞过程，如钙稳态、脂质合成和代谢物运输。这种细胞器执行的重要功能直接依赖于以质子电化学梯度的形式储存在其内膜上的势能。作为所有节能膜的特征，这种电势被解析为电场和跨脂双层的质子活性的差异。这个项目将解决线粒体内膜电化学势的两个基本特征。首先，我们将研究由内膜通电状态变化所驱动的膜蛋白结构的变化。新的，高分辨率的基于荧光的方法将被用来阐明质子梯度和跨膜电场的变化如何不同地改变模型线粒体驻留蛋白的构象。阐明这种机电耦合的基础对于理解节能膜中的蛋白质如何利用电化学势来执行细胞工作至关重要。其次，将研究拓扑复杂的内膜的能量景观。使用精确定向的pH传感和电致变色探针来测量局域离子梯度和电场，将以前所未有的分辨率测量内膜能量分布的时间和空间异质性。通过挑战目前跨膜区域的电化学势平衡教条，这项工作有望为理解ATP产生和双层结构中的局部变化等过程的能量调节创造一个新的范式。这项研究中使用的技术创新将使目前对膜生物能量学的理解取得这些进展。BROADER IMPACTS由NSF赞助的工作将涉及两个研究团队，其模式旨在促进多学科互动和科学领导力的发展，强调传统上在STEM领域代表性较低的学生的参与。这项工作还将有助于在构建模型膜系统和特定部位多肽标记过程中进一步开发和扩大新的研究工具，所有这些都将向科学界提供。该项目的教育推广部分以生物学暑期学院系列为基础，与康涅狄格大学早期学院体验方案合作。这一系列的模块化课堂和实验室课程为地区高中的教师提供专业发展，使他们接触到研究进展和当前的科学主题，这些主题可以在下一学年纳入他们自己的课程。随着这一计划的发展，该学院将扩大到包括高中生的暑期课程。总而言之，这一教育推广活动将与该项目的研究活动高度结合。此外，这一教学推广模式的成果将在教育出版物和会议讲习班上传播，作为有效帮助提高公众科学素养的一种手段。