Synthetic Models and Spectroscopy of Nonheme Diiron Enzymes

描述（由申请人提供）：项目摘要。这个提议的目的是了解生物双铁中心在代谢关键转化中如何激活双氧。非血红素二铁蛋白和酶执行涉及二氧的多种基本功能，包括二氧转运（血红素蛋白）、DMA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、烷烃和芳烃羟化酶、肌醇加氧酶、脱氧羟腐胺赖氨酸羟化酶）。一般而言，提出双氧活化需要涉及二铁（III）-过氧中间体和由其衍生的高价铁-氧代物质的共同机制。该项目的目标将使用仿生和光谱方法相结合来实现。建立在过去的成就，这些网站的结构和光谱特性建模，建议合成的三脚架配体的前体配合物，使它们与O2或过氧化物反应，并表征从中衍生的亚稳中间体。非常感兴趣的是中间体，例如二铁（II）络合物（铁（II）铁（III）-超氧或二铁（III）-过氧物质）的O2加合物，以及具有Fe（III-）Fe（-IV）和Fe（-IV）Fe（-IV）氧化态的物质。这些络合物将尽可能通过X射线晶体学和各种技术如NMR、EPR、UV-vis-NIR、拉曼、穆斯堡尔、电喷雾质谱、电化学和EXAFS表征。停流和传统的动力学方法将被用来表征其形成和分解的机制。这些瞬态复合物对一系列底物的氧化反应性将进行研究，并与酶活性位点进行比较。也要合成的是复合物，可以作为先例的新的氧活化机制，最近提出的肌醇加氧酶需要一个铁（II）铁（III）中心，结合O2和二铁（III）-superoxo物种，作为初始氧化剂。平行于这些努力，我们的光谱专业知识将被应用于阐明甲烷单加氧酶中间体和脱氧羟腐胺赖氨酸羟化酶的二铁网站结构。本案无关非血红素二铁酶执行各种代谢关键功能，需要双氧激活。了解这些酶如何工作可以导致治疗某些人类疾病的新药策略的开发。例如，肌醇加氧酶可能与糖尿病相关的许多并发症有关，而脱氧羟腐胺赖氨酸羟化酶是形成成熟的真核细胞延伸因子5a所必需的，该因子对细胞增殖是必需的，因此可以作为抗肿瘤或抗HIV治疗的靶标。

DESCRIPTION (provided by applicant): Project Summary. The goal of this proposal is to understand how dioxygen is activated by biological diiron centers in metabolically critical transformations. Nonheme diiron proteins and enzymes perform a variety of essential functions involving dioxygen, including dioxygen transport (hemerythrin), DMA 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(lll)-peroxo intermediates and high-valent iron-oxo species derived therefrom. The project goal 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(ll) complexes (either iron(ll)iron(lll)-superoxo or diiron(lll)-peroxo species), and species with Fe(lll-)Fe(-IV) and Fe(-IV)Fe(-lV) 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, Mossbauer, electrospray mass spectrometry, electrochemistry, and EXAFS. Both stopped-flow and conventional kinetic methods will be used to characterize their mechanisms of formation and decomposition. The oxidative reactivities of these transient complexes towards a range of substrates will be investigated and compared with those of enzyme active sites. Also to be synthesized are complexes that can serve as precedents for the novel oxygen activation mechanism recently proposed for myo-inositol oxygenase entailing an iron(ll)iron(lll) center that binds O2 and a diiron(lll)-superoxo species that acts as the initial oxidant. Parallel to these efforts, our spectroscopic expertise will be applied to elucidating the diiron site structures of methane monooxygenase intermediates and deoxyhypusine hydroxylase. 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, myo-inositol oxygenase may be connected to the many complications associated with diabetes mellitus, while deoxyhypusine hydroxylase is required for the formation of mature eukaryotic elongation factor 5a that is essential for cell proliferation and may thus serve as the target for anti-tumor or anti-HIV therapy.