Electronic Structure of Heme Enzyme Intermediates from Resonant Inelastic X-ray Scattering and L-Edge X-ray Absorption Spectroscopy

共振非弹性 X 射线散射和 L 边 X 射线吸收光谱研究血红素酶中间体的电子结构

• 批准号: 9768514
• 负责人: LELAND BRUCE GEE
• 金额: $ 6.37万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-16 至 2021-09-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9768514
• 关键词: 3-Dimensional; Academia; Affect; Apoptosis; Biological; Biology; Cations; Characteristics; Chemistry; Complex; Coupled; Cysteine; Cytochrome P450; Data Analyses; Detection; Development; Dioxygen; Drug Metabolic Detoxication; Due Process; Electron Transport; Electrons; Electrostatics; Environment; Enzyme Reactivation; Enzymes; Exhibits; Fluorescence; Genes; Health; Heme; Heme Iron; Histidine; Hormones; Human; Human Genome; Hydrogen Bonding; Hydroxylation; Iron; Knowledge; Ligands; Ligation; Mechanics; Mentorship; Methionine; Methods; Modeling; Nature; Noise; Optics; Outcome; Oxygen; Photons; Play; Porphyrins; Property; Pterins; Reaction; Research; Resolution; Roentgen Rays; Role; Sampling; Site; Spectrum Analysis; Steroid biosynthesis; Structure; System; Techniques; Tyrosine; Vacuum; Work; X ray spectroscopy; absorption; alpha ketoglutarate; base; biological systems; career; cholesterol biosynthesis; circular magnetic dichroism; cofactor; computational chemistry; cytochrome c; detector; driving force; drug metabolism; electronic structure; frontier; heme a; innovation; insight; interest; molecular orbital; oxygen transport; porphyrin a; preference; quantum; sensor; simulation; steroid hormone; steroid metabolism; tool

Project Summary/Abstract Much work has been performed to study reactive electron orbitals in nonheme enzymes (NHE) and their activation of oxygen. Similar to NHEs, heme-containing enzymes play a diverse set of roles in the biosphere and factor heavily to human health: detoxification, oxygen transport, and hormone synthesis. It is important that heme enzymes be understood to a similar extent as NHEs, however the tools used to characterize NHE electron covalency of Fe, such as magnetic circular dichroism optical spectroscopy, are not as applicable to hemes due to the highly delocalized equatorial porphyrin ring that obscures the covalency of the Fe atom. The covalency of the low-lying valence orbitals of the heme site, actively tunes intermediates for their function in biology. To directly probe the Fe center in heme enzymes, this project will use the relatively new resonant inelastic X-ray scattering (RIXS) spectroscopy. 1s2p RIXS uses K-edge, hard X-ray, incident photons and detects the subsequent 2p to 1s hole-filling resulting in the same final state as L-edge X-ray absorption spectroscopy (XAS) – with different selection rules. RIXS yields information about the electronic structure of the frontier molecular orbitals (FMOs), specifically d orbital covalency, critical to studying oxygen activation by hemes, however without the weaknesses inherent in soft X-ray L-edge XAS such as high vacuum requirements, high sample concentrations, and fluorescence inhibition. However, a new ultra-low noise detector called the TES will finally allow Fe L-edge XAS spectra of dilute enzyme samples, and correlating 1s2p RIXS with L-edge XAS will afford the differential orbital covalency of the frontier molecular orbitals that are key to reactivity. Initially, the trainee will apply RIXS and L- edge XAS to nonheme and heme model complexes, particularly the well-understood nonheme models will allow development of the experimentally observed 4p orbital mixing into the simulation of RIXS spectra. The trainee will then explore the change of the Fe=O bond when going from a nonheme to a heme environment in an enzyme. With L-edge XAS and RIXS the reactive orbitals of cytochrome p450 compound II will reveal the driving forces for the “rebound mechanism” of hydroxylation. Next, this study will explore the change in the Fe=O bond upon conversion to the compound I radical cation intermediate and the implications for H-atom abstraction. Finally, the frontier molecular orbitals of compound I will be compared across the different trans axial ligations in heme enzymes. This interchange of the axial ligand (histidine, cysteine, and tyrosine) will quantitively identify the, currently, loosely defined “push” and “pull” effects on the heme electronic structure that allow heterolytic O2 cleavage. This project will further the fundamental knowledge of Fe oxygen chemistry in heme, and nonheme, enzymes. Providing new insights into the nature and reactivity of the Fe=O bond, the role that the porphyrin plays in hemes, and how that is adapted in NHEs where no such electron sink is available. This work will also develop the methods and modelling of 1s2p RIXS and TES detected L-edge XAS on bioinorganic systems.

项目概要/摘要 已经开展了大量工作来研究非血红素酶（NHE）中的反应电子轨道及其 与 NHE 类似，含血红素的酶在生物圈和生物圈中发挥着多种作用。 对人类健康至关重要的因素：解毒、氧气运输和激素合成 血红素非常重要。 酶的理解程度与 NHE 相似，但是用于表征 NHE 电子的工具 Fe 的共价性，例如磁圆二色性光谱，不适用于血红素，因为 高度离域的赤道卟啉环掩盖了铁原子的共价性。 血红素位点的低位价轨道，主动调节中间体的生物学功能。 探测血红素酶中的 Fe 中心，该项目将使用相对较新的共振非弹性 X 射线散射 (RIXS) 光谱学。 1s2p RIXS 使用 K 边缘、硬 X 射线、入射光子并检测后续的 2p 1s 空穴填充导致与 L 边 X 射线吸收光谱 (XAS) 相同的最终状态 – 不同 选择规则 RIXS 产生有关前沿分子轨道 (FMO) 电子结构的信息， 特别是 d 轨道共价性，对于研究血红素的氧活化至关重要，但没有缺点 软 X 射线 L 边 XAS 固有的特性，例如高真空要求、高样品浓度和 然而，一种称为 TES 的新型超低噪声检测器最终将允许 Fe L 边缘 XAS。 稀释酶样品的光谱，并将 1s2p RIXS 与 L 边缘 XAS 相关联将提供微分轨道 对反应性至关重要的前沿分子轨道的共价性最初，学员将应用 RIXS 和 L-。 边缘 XAS 到非血红素和血红素模型复合体，特别是易于理解的非血红素模型将允许 将实验观察到的 4p 轨道混合发展到 RIXS 光谱的模拟中 然后将探索酶中从非血红素环境转变为血红素环境时 Fe=O 键的变化。 利用 L-edge XAS 和 RIXS，细胞色素 p450 化合物 II 的反应轨道将揭示驱动力 接下来，本研究将探讨羟基化后Fe=O键的变化。 转化为化合物 I 自由基阳离子中间体以及对 H 原子提取的影响。 将比较血红素中不同跨轴连接的化合物 I 的前沿分子轨道 轴向配体（组氨酸、半胱氨酸和酪氨酸）的这种交换将定量识别， 目前，对血红素电子结构的松散定义的“推”和“拉”效应允许异解O2 该项目将进一步加深血红素和非血红素中铁氧化学的基础知识， 为 Fe=O 键的性质和反应性以及卟啉的作用提供了新的见解。 这项工作也将在血红素中发挥作用，以及如何在没有此类电子汇的 NHE 中进行调整。 开发 1s2p RIXS 和 TES 在生物无机系统上检测 L 边缘 XAS 的方法和建模。