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.

项目摘要 铁酶在生物学中的O2激活中起主要作用。这些根据 它们反映其O2激活模式的主动位点结构：非cactortor依赖性 单核非血红素铁（MNHFE）酶，辅因子（α-酮戊二酸（α-kg）和翼蛋白） 依赖的MNHFE酶，苯甲酸NHFE酶和O2/H2O2激活血红素 酶。所有晶体结构和氧反应中间体都存在于所有的金属酶 四个班级。多年来，我们开发了新的光谱方法，使 NHFEII活性位点以及其O2的几何和电子结构的详细研究 中间体，现在已经开发了一种定量研究铁中心的方法 高度共价和发色的血红素环境。在过去的5个成就中 年是：1）确定FEIIII-O2物种是所有子类中的反应性中间体 依赖性MNHFE酶的; 2）定义O2反应坐标以生成 Feiv = Oα-KG和翼蛋白依赖性酶中的中间体； 3）对于α-kg 依赖子类，定义了其Feiv = O酶的几何和电子结构 中间以及这些直接卤素如何超过热力学优先 卤代酶中的羟基化； 4）表明，与MNHFE酶形成对比， 氢过氧化物中间体在苯甲酸NHFE酶中活跃，以直接反应与 底物； 5）对于甲烷单加掺杂酶，其中过氧 - 果实中间迅速 转换为高价值2Feiv-oxo中间体Q，我们确定了Q的结构（a 当前辩论的主题），并洞悉了其对甲烷的高反应性； 6）使用了 光谱法我们现在已经在血红素环境中为铁开发了 在实验上，氧气血红蛋白的计算有争议的电子结构； 7）和 扩展了这种方法，以分析具有较强的Fe-oxo键的活性位点。我们的学习现在是 致力于完成四个类别及其子类的反应坐标， 了解在超氧，过氧和Feiv-oxo水平上的O2激活，确定 第二个铁中均质NHFE酶的反应性增强和理解 高价铁氧中间体的激活和选择性的差异 单核NH，Binclear NH和血红素活性位点。