Synthetic Models and Spectroscopy of Nonheme Diiron Enzymes

非血红素二铁酶的合成模型和光谱学

• 批准号: 7811796
• 负责人: LAWRENCE QUE
• 金额: $ 16.21万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7811796
• 关键词: Achievement; Active Sites; Alkanes; Anti-HIV Therapy; Bacteria; Biological; Biomimetics; Cell Proliferation; Chemicals; Chlamydia trachomatis; Complex; DNA biosynthesis; Development; Dioxygen; Electrochemistry; Electronics; Elongation Factor; Enzymes; Escherichia coli; Fatty Acid Desaturases; Ferritin; Funding; Goals; Histidine; Human; Immune response; Iron; Lead; Ligands; Mammalian Cell; Mammals; Mass Spectrum Analysis; Metabolic; Metalloproteins; Metals; Methane hydroxylase; Methodology; Mixed Function Oxygenases; Modeling; Nature; Oxidants; Oxidation-Reduction; Oxygen; Parasites; Peroxides; Pharmaceutical Preparations; Property; Proteins; Research; Ribonucleotide Reductase; Ribonucleotides; Sampling; Site; Spectrum Analysis; Stimulus; Structural Models; Structure; Techniques; Therapeutic Studies; Work; X-Ray Crystallography; adduct; analog; base; carboxylate; complex IV; deoxyhypusine monooxygenase; desaturase; human disease; inositol oxygenase; insight; interest; microorganism; novel; oxidation; programs; public health relevance; toluene 2-xylene monooxygenase; tumor

DESCRIPTION (provided by applicant): The overall goal of this proposal is to understand how dioxygen is activated by biological diiron centers in metabolically critical transformations. Nonheme diiron enzymes perform a variety of essential functions involving dioxygen, including DNA biosynthesis (ribonucleotide reductase), iron storage (ferritin), and oxidations of organic substrates (methane monooxygenase, fatty acid desaturases, alkane and arene hydroxylases, myo-inositol oxygenase, deoxyhypusine hydroxylase). In general, dioxygen activation is proposed to entail a common mechanism involving diiron(III)-peroxo intermediates and high-valent iron-oxo species derived therefrom. The project goals will be accomplished using a combination of biomimetic and spectroscopic approaches. Building on past accomplishments in modeling structural and spectroscopic properties of such sites, it is proposed to synthesize precursor complexes of tripodal ligands, to react them with O2 or peroxides, and to characterize the metastable intermediates derived therefrom. Of great interest are intermediates such as O2 adducts of diiron(II) complexes (either iron(II)iron(III)-superoxo or diiron(III)-peroxo species), and species with Fe(III)Fe(IV) and Fe(IV)Fe(IV) oxidation states. These complexes will be characterized by X-ray crystallography whenever possible and by a variety of techniques such as NMR, EPR, UV-vis-NIR, Raman, M"ssbauer, electrospray mass spectrometry, electrochemistry, and EXAFS. Parallel to these efforts, our spectroscopic expertise will be applied to elucidating the diiron site structures of methane monooxygenase intermediates and human deoxyhypusine hydroxylase. In this competitive revision, a new specific aim for GM-38767 is added because of the recent discovery that ribonucleotide reductase (RNR) of the parasite Chlamydia trachomatis uses as the oxidant needed to initiate ribonucleotide reduction a Fe(III)-O-Mn(IV) center, rather than the diiron(III)/tyrosyl radical combination characterized in E. coli and mammalian RNRs. It is suggested that this metal substitution allows this parasitic bacterium to circumvent the sensitivity of diiron RNR to NO produced in the typical immune response of mammalian cells. For the new specific aim, it is proposed to use methodologies developed in our ongoing work on synthetic diiron intermediates to obtain and characterize corresponding FeMn analogs, namely Fe(III)Mn(III)-peroxo, Fe(III)-O-Mn(IV), and Fe(IV)-O-Mn(IV) species. The properties of these novel FeMn complexes will be compared with those of their diiron counterparts to assess the chemical basis for Nature's choice of metal centers in this crucial enzyme. PUBLIC HEALTH RELEVANCE: Nonheme diiron enzymes perform a variety of metabolically critical functions that require dioxygen activation. Understanding how these enzymes work can lead to the development of new drug strategies for treating some human diseases. For example, ribonucleotide reductase is a key enzyme that controls DNA biosynthesis, while deoxyhypusine hydroxylase is required for the formation of mature eukaryotic elongation factor 5a that is essential for cell proliferation; thus both enzymes may serve as targets for anti-tumor or anti-HIV therapy.

描述(由申请人提供)：本提案的总体目标是了解在代谢关键的转化过程中，生物双铁中心是如何激活氧气的。非血红素二铁酶具有多种涉及氧气的基本功能，包括DNA生物合成(核糖核苷酸还原酶)、铁储存(铁蛋白)和有机底物的氧化(甲烷单加氧酶、脂肪酸去饱和酶、烷烃和芳烃羟基酶、肌醇加氧酶、脱氧亚硫氨酸羟基酶)。一般来说，氧的活化被认为需要一个共同的机制，涉及二铁(III)-过氧基中间体及其衍生的高价铁氧物种。该项目的目标将使用仿生学和光谱相结合的方法来实现。在过去对这些中心的结构和光谱性质进行模拟的基础上，建议合成三脚架配体的前体络合物，将它们与O2或过氧物反应，并表征由此衍生的亚稳中间体。感兴趣的是中间体，如二铁(II)络合物的O2加合物(铁(II)铁(III)-超氧或二铁(III)-过氧基物种)，以及具有Fe(III)Fe(IV)和Fe(IV)Fe(IV)氧化态的物种。这些络合物将尽可能通过X射线结晶学和各种技术进行表征，如核磁共振、EPR、UV-VIS-NIR、拉曼光谱、穆斯堡尔谱、电喷雾质谱学、电化学和EXAFS。在这些工作的同时，我们的光谱专业知识将被用于阐明甲烷单加氧酶中间体和人脱氧次氨酸羟基酶的双铁中心结构。在这次竞争性修订中，增加了GM-38767的一个新的特异性目标，因为最近发现沙眼衣原体的核糖核苷酸还原酶(RNR)作为氧化剂，启动Fe(III)-O-Mn(IV)中心的核苷酸还原，而不是大肠杆菌和哺乳动物RNRs所特有的二铁(III)/酪氨酸自由基结合。这表明，这种金属取代使这种寄生细菌能够绕过哺乳动物细胞典型免疫反应中产生的二铁RNR对NO的敏感性。对于新的具体目标，建议使用我们正在进行的合成二铁中间体工作中开发的方法来获得和表征相应的FeMn类似物，即Fe(III)Mn(III)-Peroxo，Fe(III)-O-Mn(IV)和Fe(IV)-O-Mn(IV)物种。这些新型的FeMn络合物的性质将与它们的双铁配合物进行比较，以评估大自然在这种关键酶中选择金属中心的化学基础。 公共卫生相关性：非血红素二铁酶执行各种代谢关键功能，需要氧气激活。了解这些酶是如何工作的，可以导致开发治疗某些人类疾病的新药策略。例如，核糖核苷酸还原酶是控制DNA生物合成的关键酶，而脱氧亚硫氨酸羟化酶是形成成熟的真核细胞延伸因子5a所必需的，这是细胞增殖所必需的；因此，这两种酶都可能成为抗肿瘤或抗HIV治疗的靶点。