Mechanistic and spectroscopic investigation of sulfur-oxidizing non-heme iron enzymes

硫氧化非血红素铁酶的机理和光谱研究

• 批准号: 1213655
• 负责人: Brad Pierce
• 金额: $ 30万
• 依托单位: University of Texas at Arlington
• 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: 2012-08-15 至 2016-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1213655&HistoricalAwards=false
• 关键词: Mechanistic; spectroscopic; investigation; sulfur; oxidizing

In this award from the Chemistry of Life Processes Program in the Division of Chemistry, Dr. Brad Pierce, from the University of Texas at Arlington, will study the mechanism and regulation of mammalian thiol dioxygenase (TDO) enzymes. Characterization of transient non-heme iron [Fe-O] intermediates has historically attracted considerable interest within the area of bioinorganic chemistry. This intense focus of research efforts can largely be explained by the vast number of functionally (and structurally) diverse non-heme iron enzymes and the incredible versatility exhibited in chemical oxidations they initiate. In contrast to the canonical 2-His-1-carboxylate facial triad motif exhibited by nearly all members of this enzyme class, the active site of mammalian TDO enzymes utilize a neutral, all-His (3-His) facial triad motif. Furthermore, TDO enzymes also have an unusual covalently cross-linked cysteine-tyrosine pair in close proximity to the non-heme iron active site. One objective for this proposal is to identify and spectroscopically characterize transient intermediates produced during TDO-catalyzed thiol-oxidation. An additional area of focus is the role of a rare post-transcriptional cysteine-tyrosine covalent modification within the active site of TDO enzymes. Specifically, steady-state O2-coupling efficiency of TDO enzymes will be determined for comparison to C93 and Y157 mutants. These results will be correlated to specific molecular interactions within the active site as observed by spectroscopy to evaluate the role of outer-sphere interactions on native TDO catalysis. In contrast to oxygenase/oxidase enzymes, synthetic non-heme iron model complexes are unable to gate O2-reactivity through substrate-binding. As a consequence, model systems are poor catalysts for oxidative transformations. Currently, this poorly understood aspect of non-heme iron catalysis represents one of the most significant hurdles in the development of industrially useful O2-dependent iron catalysts. Therefore, an additional area of investigation will focus on the mechanism by which TDO enzymes modulate O2-reactivity through protein-substrate interactions. Dr. Pierce's research is potentially technology-enabling in that results obtained could be applied to the design of biologically inspired catalysts for oxidative transformations. The award will contribute significantly to the University of Texas at Arlington's strategic plan of becoming a national research university and a source of well-prepared STEM students. Furthermore, given the high population of first-generation students at the University, a significant impact can be made toward increasing diversity within STEM fields by sponsoring undergraduate and graduate research opportunities. Mentoring future scientists is a significant priority, and this award will provide continued research opportunities and support for both local area high school students and undergraduates at the University. The interdisciplinary nature of the proposed research will provide a wide breadth of research training for students at every level (high school, undergraduate, and graduate).

在化学系生命过程化学项目的奖项中，来自阿灵顿德克萨斯大学的Brad Pierce博士将研究哺乳动物硫醇双加氧酶（TDO）酶的机制和调控。瞬态非血红素铁[Fe-O]中间体的表征历来在生物无机化学领域引起了相当大的兴趣。大量功能（和结构）多样化的非血红素铁酶以及它们引发的化学氧化所表现出的令人难以置信的多功能性，在很大程度上可以解释这种研究努力的高度集中。与该酶类几乎所有成员都表现出的典型的2- his -1-羧酸面部三基序相反，哺乳动物TDO酶的活性位点利用中性的全his （3-His）面部三基序。此外，TDO酶在非血红素铁活性位点附近也有一个不寻常的共价交联半胱氨酸-酪氨酸对。本提案的一个目标是鉴定和光谱表征在tdo催化的硫醇氧化过程中产生的瞬态中间体。另一个重点领域是TDO酶活性位点内罕见的转录后半胱氨酸-酪氨酸共价修饰的作用。具体来说，将确定TDO酶的稳态o2偶联效率，并与C93和Y157突变体进行比较。这些结果将与光谱观察到的活性位点内的特定分子相互作用相关联，以评估外球相互作用对天然TDO催化的作用。与加氧酶/氧化酶相比，合成的非血红素铁模型复合物不能通过底物结合来控制o2的反应性。因此，模型系统是氧化转化的不良催化剂。目前，对非血红素铁催化的这一知之甚少的方面代表了工业上有用的依赖于o2的铁催化剂发展的最大障碍之一。因此，一个额外的研究领域将集中在TDO酶通过蛋白质-底物相互作用调节o2反应性的机制上。皮尔斯博士的研究是潜在的技术支持，其结果可以应用于氧化转化的生物启发催化剂的设计。该奖项将为德克萨斯大学阿灵顿分校成为一所全国性研究型大学的战略计划做出重大贡献，并成为准备充分的STEM学生的来源。此外，鉴于该大学的第一代学生人数众多，通过赞助本科生和研究生的研究机会，可以对增加STEM领域的多样性产生重大影响。指导未来的科学家是一个重要的优先事项，该奖项将为当地高中生和大学本科生提供持续的研究机会和支持。拟议研究的跨学科性质将为各个层次（高中、本科和研究生）的学生提供广泛的研究训练。