Spectroscopic Characterization of Oxygen Intermediates in Non-heme and Heme Iron Enzymes

非血红素和血红素铁酶中氧中间体的光谱表征

• 批准号: 10396809
• 负责人: EDWARD I SOLOMON
• 金额: $ 45.17万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10396809
• 关键词: Active Sites; Anabolism; Antibiotics; Biology; Bioremediations; Biotechnology; Chemistry; Coenzymes; Collagen; Crystallization; Disease; Engineering; Environment; Enzymes; Health; Heme; Heme Iron; Human; Hydrogen Peroxide; Hydroxylation; Hypoxia; Iron; Methane; Methane hydroxylase; Methods; Molecular; Mononuclear; Natural Products; Nature; Oxygen; Oxyhemoglobin; Play; Pterins; Reaction; Regulation; Research; Role; Structure; Thermodynamics; alpha ketoglutarate; base; catalyst; chromophore; drug metabolism; electronic structure; geometric structure; halogenation; insight; metalloenzyme; novel therapeutics; rational design

Project summary Iron enzymes play major roles in O2 activation in biology. These divide into four classes based on their active site structures that reflect their mode of O2 activation: the non-cofactor dependent mononuclear non-heme iron (MNHFe) enzymes, the cofactor (α-ketoglutarate (α-KG) and pterin) dependent MNHFe enzymes, the binuclear NHFe enzymes and the O2/H2O2 activating heme enzymes. Crystal structures and oxygen reaction intermediates exist for metalloenzymes in all four classes. Over the years, we have developed new spectroscopic methods enabling the detailed study of the NHFeII active sites, and the geometric and electronic structures of their O2 intermediates and have now developed a method to quantitatively study the iron center in the highly covalent and chromophoric heme environment. Among our accomplishments in the past 5 years are: 1) determined that FeIII-O2- species are the reactive intermediates in all the subclasses of non-cofactor dependent MNHFe enzymes; 2) defined the O2 reaction coordinates to generate the FeIV=O intermediates in both the α-KG and pterin dependent enzymes; 3) for the α-KG dependent subclass, defined the geometric and electronic structures of their FeIV=O enzyme intermediates and how these direct halogenation over the thermodynamically favored hydroxylation in the halogenases; 4) showed that in contrast to the MNHFe enzymes, hydroperoxide intermediates are active in the binuclear NHFe enzymes for direct reaction with substrates; 5) for methane monooxygenase, where the peroxo-biferric intermediate rapidly converts to a high-valent 2FeIV-oxo intermediate Q, we have determined the structure of Q (a topic of current debate) and provided insight into its high reactivity with methane; 6) used the spectroscopic method we have now developed for iron in heme environments to determine experimentally the computationally controversial electronic structure of oxyhemoglobin; 7) and extended this method to analyze active sites with strong Fe-oxo bonds. Our studies are now directed toward completing the reaction coordinates of the four classes and their subclasses, understanding O2 activation at the superoxo, peroxo and FeIV-oxo levels, determining the role of the second iron in the enhanced reactivity of the binuclear NHFe enzymes, and understanding the differences in the activation and selectivity of high-valent iron-oxo intermediates in mononuclear NH, binuclear NH and heme enzyme active sites.

项目总结 在生物体内，铁酶在氧活化中起着重要的作用。它们根据以下情况分为四类 反映其氧活化模式的活性部位结构：非辅因子依赖 单核非血红素铁(MNHFe)酶、辅因子(α-酮戊二酸(α-KG)和蝶呤) 依赖的MNHFe酶、双核NHFe酶和O2/H_2O_2激活的血红素 酵素。所有金属酶的晶体结构和氧反应中间体都存在 四节课。多年来，我们开发了新的光谱方法，使 NHFeII活性中心及其O2的几何和电子结构的详细研究 现在已经开发出一种方法来定量研究铁离子的中心。 高度共价和发色的血红素环境。在过去5年中，我们取得了以下成就 年：1)确定FeIII-O2-物种是所有亚类中的活性中间体 非辅因子依赖的MNHFe酶；2)定义O2反应生成的坐标 α-KG和蝶呤依赖酶中的FeIV=O中间体；3)α-KG 依赖亚类，定义了它们的FeIV=O酶的几何结构和电子结构 中间体以及这些直接卤化反应如何在热力学上受到青睐 4)结果表明，与MNHFe酶不同， 过氧化氢中间体在双核NHFe酶中具有活性，可直接与 底物；5)甲烷单加氧酶，过氧二铁中间体迅速 转化为高价的2FeIV-oxo中间体Q，我们确定了Q(A)的结构 当前辩论的主题)，并对其与甲烷的高反应性提供了洞察；6)使用了 我们现在开发的用于测定血红素环境中的铁的光谱方法 实验上计算上有争议的氧合血红蛋白电子结构；7)和 将该方法扩展到分析具有强Fe-OO键的活性中心。我们的研究现在是 针对完成四个类及其子类的反应坐标， 了解O2在超氧、过氧和FeIV-氧水平上的激活，确定氧的作用 第二铁在增强双核NHFe酶的反应性方面的认识 高价铁氧中间体的活性和选择性的差异 单核NH、双核NH和血红素酶活性部位。