NMR STUDIES OF RESPIRATORY PROTEINS AND MODEL SYSTEMS

描述：此应用程序的广泛、长期目标是 利用核磁共振波谱研究呼吸系统的结构 蛋白质；特别是O2和CO如何与肌红蛋白和血红蛋白结合 以及模型系统，例如“尖桩栅栏”和“带帽的”或 受保护的金属卟啉。这项工作与健康相关的是 充分了解蛋白质是如何控制O2和CO结合的 对于理解氧气是如何在血液中运输和 肌肉，氧气亲和力在各种呼吸系统疾病中是如何改变的， 以及从长远来看，血液替代品可能会被设计出来。这个 具体目标是首先，构建一个600 MHz的多探头宽带 核磁共振光谱仪，它将允许对 至少两个不同的样品，同时在同一磁铁中。 这将增加样品吞吐量，降低总体有效成本 昂贵的核磁共振资源。第二，将在#年研究CO键合 血红素模型体系，使用13C和17O化学位移和17O四重 耦合信息和密度泛函理论(DFT)。这里的目标是 是将观察到的光谱参数与结构(配体)联系起来 倾斜、弯曲、静电场、血红素褶皱/穹顶/鞍形，近端 侧面互动)。电子关联和交换的影响 将使用DFT形式主义来处理。第三个目标是澄清 血红素蛋白中FeCo/远端/近端相互作用的性质 他们自己。将特别强调获得第一名 ~(13)C、~(17)O屏蔽场的原理、定量分析 肌红蛋白A0、A1和A3底物中的梯度张量 血红蛋白。第四个目标是研究氧气是如何与铁结合的。 模型血红素系统，使用为CO.最终目标是 用~(17)O核磁共振和核磁共振仪研究血红素蛋白本身的铁-氧相互作用 DFT。与CO的早期报告一样，已报道的FeOO几何构型变化很大， 从~115度到159度。然而，很可能有很大一部分 角度的范围实际上更窄，因为能量 与这些扭曲相关的是非常大的。固态~(170)O核磁共振 将能够对配体几何构型的假设进行测试 接近130度。氧气的稳定程度将是 研究(例如，与远端组氨酸的氢键)，再次使用密度泛函理论 方法：研究方法。其目标是详细了解CO和 O2与血红素蛋白中的Fe结合，以澄清FeCo和FeO2的主题 几何结构，并提供电场/氢键相互作用的探针。 了解配体如何与金属蛋白结合；例如，O2如何结合 并由血红蛋白稳定，具有相当长期的健康- 相关含义，因为如果重要的相互作用是未知的， 那么，改进的治疗方法将更难设计。

DESCRIPTION: The broad, long term objective of this application is to use NMR spectroscopy to investigate the structures of respiratory proteins; in particular, how O2 and CO bind to myoglobin and hemoglobin as well as model systems, such as "picket fence" and "capped" or protected metalloporphyrins. The health relatedness of this work is that a full understanding of how O2 vs. CO binding is controlled by proteins is important for understanding how O2 is transported in blood and muscle, how O2 affinities are changed in various respiratory diseases, as well as how in the long term, blood substitutes may be designed. The specific aims are first, to construct a multiple probe 600 MHz widebore NMR spectrometer, which will permit routine observations to be made on at least two different samples, simultaneously, in the same magnet. This will increase sample throughput, reducing the overall effective cost of expensive NMR resources. Second, CO bonding will be investigated in heme model systems, using 13C and 17O chemical shift and 17O quadruple coupling information and density functional theory (DFT). The goal here is to relate observed spectroscopic parameters to structure (ligand tilt, bend, electrostatic fields, heme ruffling/doming/saddling, proximal side interactions). The effects of electron correlation and exchange will be handled using the DFT formalism. The third aim is to clarify the nature of FeCO/distal/proximal interactions in heme proteins themselves. Particular emphasis will be placed on obtaining first principles, quantitative analyses of 13C, 17O shielding and field gradient tensors in A0, A1, and A3 substrates of myoglobins and hemoglobins. The fourth goal is to investigate how O2 binds to Fe in model heme systems, using methods developed for CO. The final goal is to study Fe-O2 interaction in heme proteins themselves, using 17O NMR and DFT. As with early reports for CO, reported FeOO geometries vary widely, from ~115 degrees to 159 degrees. However, it is likely that a much narrower range of angles is actually present, since the energies associated with these distortions are very large. Solid-state 17O NMR will enable a test of the hypothesis that the ligand geometries are much closer to 130 degrees. The extent of O2 stabilization will be investigated (e.g. H-bonding to the distal histidine), again using DFT methods. The goal is to obtain a detailed understanding of how CO and O2 bind to Fe in heme proteins, to clarify the topic of FeCO and FeO2 geometries, and to provide probes of E-field/H-bonding interactions. Understanding how ligands bind to metalloproteins; e.g., how O2 binds to and is stabilized by hemoglobin, has considerable long-term health- related implications, since if the important interactions are not known, then improved therapies will be more difficult to engineer.