High-resolution iron X-ray absorption spectroscopic and computational studies of non-heme diiron peroxo intermediates
High-resolution iron X-ray absorption spectroscopic and computational studies of non-heme diiron peroxo intermediates
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DOI：
10.1016/j.jinorgbio.2019.110877
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发表时间：
2020-02-01
影响因子：
3.9
通讯作者：
DeBeer, Serena
中科院分区：
文献类型：
--
作者：
Cutsail, George E., III;Blaesi, Elizabeth J.;DeBeer, Serena
铁蛋白样羧酸桥联非血红素二铁酶可激活 O-2，以应对自然界中的各种困难反应。这些反应通常是从强 C-H 键中夺取氢开始的。这些酶在其不同的辅助因子上激活 O-2，形成典型的二铁过氧中间体，具有一系列可能的配位模式。在此，我们探索了高能分辨率荧光检测 X 射线吸收光谱 (HERFD XAS) 的能力，以深入了解非血红素二铁蛋白中过氧水平中间体的性质。研究了来自对氨基苯甲酸 N-加氧酶 (AurF)、醛脱甲酰加氧酶 (ADO) 和大肠杆菌 (Ec beta) 的 Ia 类核糖核苷酸还原酶 β 亚基的冷冻猝灭 (FQ) 过氧中间体。所有三种中间体均建议采用不同的过氧结合模式，并且各自表现出不同的 Fe K α HERFD XAS 前边缘特征和强度。由于这些 FQ 捕获的样品由多个物种组成，因此还需要根据穆斯堡尔光谱确定的物种形成对 HERFD XAS 光谱进行解卷积，从而从稀释的蛋白质样品中产生“纯”二铁过氧 HERFD XAS 光谱。最后，通过一系列假设模型复合物上的 XAS 光谱的时间相关密度泛函理论 (TDDFT) 计算，评估给定过氧配位模式对 HERFD XAS 前边缘能量和强度的影响，这些模型复合物跨越了所有可能的过氧配位模式到二铁核。讨论了 HERFD XAS 在酶中间体未来研究中的实用性。
Ferritin-like carboxylate-bridged non-heme diiron enzymes activate O-2 for a variety of difficult reactions throughout nature. These reactions often begin by abstraction of hydrogen from strong C-H bonds. The enzymes activate O-2 at their diferrous cofactors to form canonical diferric peroxo intermediates, with a range of possible coordination modes. Herein, we explore the ability of high-energy resolution fluorescence detected X-ray absorption spectroscopy (HERFD XAS) to provide insight into the nature of peroxo level intermediates in non-heme diiron proteins. Freeze quenched (FQ) peroxo intermediates from p-aminobenzoate N-oxygenase (AurF), aldehyde-deformylating oxygenase (ADO), and the beta subunit of class Ia ribonucleotide reductase from Escherichia coli (Ec beta) are investigated. All three intermediates are proposed to adopt different peroxo binding modes, and each exhibit different Fe K alpha HERFD XAS pre-edge features and intensities. As these FQ-trapped samples consist of multiple species, deconvolution of HERFD XAS spectra based on speciation, as determined by Mossbauer spectroscopy, is also necessitated - yielding 'pure' diferric peroxo HERFD XAS spectra from dilute protein samples. Finally, the impact of a given peroxo coordination mode on the HERFD XAS pre-edge energy and intensity is evaluated through time-dependent density functional theory (TDDFT) calculations of the XAS spectra on a series of hypothetical model complexes, which span a full range of possible peroxo coordination modes to a diferric core. The utility of HERFD XAS for future studies of enzymatic intermediates is discussed.