Proton Translocation and Metal Chelation in Ferrochelatase

铁螯合酶中的质子易位和金属螯合

• 批准号: 0843532
• 负责人: William Lanzilotta
• 金额: $ 55.74万
• 依托单位: University of Georgia Research Foundation Inc
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0843532&HistoricalAwards=false
• 关键词: Proton; Translocation; Metal; Chelation; Ferrochelatase

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).Intellectual MeritHeme is an essential cofactor for a wide variety of biologically important proteins including globins, cytochromes, transcription factors, peroxidases, catalases, and others. With few exceptions, organisms that possess heme-containing proteins possess the biosynthetic machinery to synthesize their own heme and do not acquire heme from their diet. Inability to synthesize heme is lethal for most organisms. The importance of the metabolic synthesis of heme is underscored by recent evidence revealing that heme is also a regulator of a number of other metabolic processes. In this exciting new role, these "heme sensors" must detect and respond to heme but not bind it permanently. The terminal step in the seven step biosynthetic pathway, the insertion of ferrous iron into protoporphyrin to make protoheme (heme), is catalyzed by the enzyme ferrochelatase. In animals this membrane-associated enzyme is located on the matrix side of the inner mitochondrial membrane with its active site facing into the membrane. This is in contrast to the bacterial ferrochelatases that are quite soluble. At the heart of this investigation is the development of biophysical tools and techniques to directly monitor proton translocation as well as metal binding and chelation in porphyrin macrocycles. The implications of such technology and experimental findings have much broader implications because proton binding/transfer is a critical element of the vast majority of biochemical reactions and many essential biochemical processes. In addition, it is now widely recognized that metals play a critical role in approximately 30 percent of all known enzymatic reactions. Taking advantage of the catalytic diversity found in nature will be critical to sustaining the quality of life humanity currently enjoys in the absence of abundant fossil fuels.Due to the broad importance of the chelation reaction under investigation here, the experiments outlined in this project will significantly advance our understanding of how metals are inserted into porphyrin cofactors in general. In addition, a unique aspect of this project is the application of organic synthesis and neutron diffraction to test specific hypotheses regarding the atomic details of porphyrin deprotonation, metal dehydration, and metal insertion. Given the established and emerging roles for porphyrin cofactors in numerous enzymes, metabolic pathways, as well as clinical applications, this work will have far reaching implications. In addition, the application of both neutron diffraction methods and organic synthesis to probe enzyme mechanism makes this project truly multidisciplinary and places this work at the cutting edge of scientific discovery that will lead to and support new technology development.Broader ImpactIn addition to the broader technological impacts, this research project offers an exceptional educational opportunity in the context of a multi-disciplinary approach to advancing discovery and understanding of structure-function relationships in enzyme function. Funding will support training of two PhD candidates and several undergraduate students in biophysical chemistry, structural biology, biochemistry, molecular biology and organic chemistry. In addition, this project will further advance the attraction of underrepresented minority students to the physical sciences through an affiliation with the Peach State Louis Stokes Alliance for Minority Participation (LSAMP) as well as the Center for Undergraduate Research Opportunities (CURO) programs. Both of these programs have proven track records in helping these students find success in the professional fields of their choice.

该奖项由2009年《美国复苏和再投资法案》(Public Law 111-5)资助。智力优秀血红素是多种生物重要蛋白质的基本辅因子，包括珠蛋白、细胞色素、转录因子、过氧化物酶、过氧化氢酶等。几乎没有例外的是，拥有含血红素蛋白质的生物体拥有合成自身血红素的生物合成机制，而不是从饮食中获得血红素。不能合成血红素对大多数生物体来说是致命的。最近的证据显示，血红素也是许多其他代谢过程的调节剂，这突显了血红素代谢合成的重要性。在这一令人兴奋的新角色中，这些“血红素传感器”必须检测并对血红素做出反应，但不能永久地束缚它。在七步生物合成途径中的最后一步，即将亚铁插入原卟啉以生成原血红素(血红素)，是由铁络合酶催化的。在动物体内，这种膜相关酶位于线粒体内膜的基质侧，其活性部位面向膜。这与细菌铁络合酶形成鲜明对比，细菌铁络合酶是非常可溶的。这项研究的核心是生物物理工具和技术的发展，以直接监测质子转移以及卟啉大环中的金属结合和螯合。这种技术和实验结果的影响具有更广泛的影响，因为质子结合/转移是绝大多数生化反应和许多基本生化过程的关键要素。此外，现在人们普遍认为，在所有已知的酶反应中，金属在大约30%的反应中起着关键作用。利用自然界中发现的催化多样性将是维持人类目前在缺乏丰富化石燃料的情况下享受的生活质量的关键。由于这里研究的螯合反应具有广泛的重要性，本项目中概述的实验将极大地促进我们对金属是如何插入到卟啉辅因子中的理解。此外，该项目的一个独特方面是应用有机合成和中子衍射来测试关于卟啉去质子化、金属脱水和金属插入的原子细节的具体假设。鉴于卟啉辅因子在许多酶、代谢途径以及临床应用中已经确立和正在出现的作用，这项工作将具有深远的影响。此外，应用中子衍射法和有机合成来探测酶的机制，使这一项目真正成为多学科的项目，并将这项工作置于科学发现的前沿，这将导致并支持新技术的发展。广泛的影响除了更广泛的技术影响外，这项研究项目在推进酶功能中结构-功能关系的发现和理解的多学科方法的背景下，提供了一个特殊的教育机会。资金将用于支持两名博士生和几名本科生在生物物理化学、结构生物学、生物化学、分子生物学和有机化学方面的培训。此外，该项目将通过与桃州路易斯·斯托克斯少数民族参与联盟(LSAMP)以及本科生研究机会中心(CURO)计划的联系，进一步提高未被充分代表的少数族裔学生对物理科学的吸引力。这两个项目在帮助这些学生在他们选择的专业领域取得成功方面都有经过证明的记录。