SPECTROSCOPY OF BIOLOGICAL SATURATED PORPHYRINS

这个项目涉及详细的假肢的调查 二氢卟啉和饱和卟啉的基团结构 蛋白质和酶。 在过去的一段时间里，我们 确定的特征和确定的红外（IR）， 共振拉曼（RR）光谱性质的金属绿， 一直都是叶绿素大环的诊断指标， 生物和模型系统。 在目前的提案中，我们建议 将这些研究扩展到以下系统：1）绿色 血红素d末端氧化酶。大肠杆菌被认为是一种 二醇二氢卟酚或内酯二氢卟酚。 我们将检查RR光谱 纯化的E.大肠杆菌血红素d复合物在其各种自旋和 与二醇和内酯取代的氧化态相比 模型复合物，以确定体内结构的血红素d 大循环 2)肌红蛋白（Mb）的绿色形式由 暴露于硫化合物，sulfMb，具有硫改性的铁 二氢卟酚辅基。 哺乳动物的酶 乳过氧化物酶（LPO）和甲状腺过氧化物酶（TPO）被抑制 被一种叫做甲状腺肿素的硫类药物， 这导致sulfMb的形成。 我们建议确定 无论sulfLPO的辅基是否确实是硫二氢卟酚， 并阐明了所观察到的紫外-可见光谱的结构起源 sulfLPO和sulfMb的区别。 我们亦会延长 关于二氯亚砜系统的研究，包括稳定的、可提取的 来自sulfMb的硫代血红素，如果可能的话，还有sulfLPO，以及模型 磺二氢卟酚络合物。 3)我们将纯化细胞色素cd1 细菌亚硝酸盐还原酶铜绿假单胞菌和分离 绿色血红素d1辅基检测光谱 dioxoisobacteriochlorins（卟啉二酮）的特性。 我们 将比较纯化的CD 1酶及其 分离的d1辅基与各种位置的辅基 氧代二氢卟酚和二氧代iBC模型的取代异构体 配合物 4)我们计划将这些研究扩展到分析 西罗血红素和更高饱和度的卟啉，例如， 四氢卟啉和六氢卟啉，建立 光谱特性，这是具体的每一个水平的饱和 卟啉。5)肌红蛋白和其他血红素蛋白质 用铁二氢卟酚复合物重组。 我们会研究 这些二氢卟酚-蛋白质在所有可访问的自旋光谱特性， 和氧化态，以及配体络合物，相比之下， 与无蛋白质模型复合物。

This project concerns the investigation of the detailed prosthetic group structure of chlorin and saturated porphyrin-containing proteins and enzymes. In the previous grant period, we established characteristic and definitive infrared (IR) and resonance Raman (RR) spectral properties of metallochlorins that are consistently diagnostic for the chlorin macrocycle in biological and model systems. In the current proposal we propose to extend those studies to the following systems: 1) The green heme d terminal oxidase from E. coli is proposed to be either a diol chlorin or a lactone chlorin. We will examine the RR spectra of the purified E. coli heme d complex in its various spin- and oxidation-states in comparison with diol and lactone substituted model complexes to determine the in vivo structure of the heme d macrocycle. 2) The green form of myoglobin (Mb) induced by exposure to sulfur compounds, sulfMb, has a sulfur-modified iron chlorin prosthetic group. The mammalian enzymes lactoperoxidase (LPO) and thyroid peroxidase (TPO) are inhibited by sulfur drugs known as goitrogens, in a reaction similar to that which leads to the formation of sulfMb. We propose to determine whether the prosthetic group of sulfLPO is indeed a sulf-chlorin, and elucidate the structural origin of the observed UV-vis spectral differences between sulfLPO and sulfMb. We will also extend the studies on sulfchlorin systems to include the stable, extractable sulfhemes from sulfMb, and if possible sulfLPO, as well as model sulfchlorin complexes. 3) We will purify the cytochrome cd1 bacterial nitrite reductase from Ps. aeruginosa and isolate the green heme d1 prosthetic group to examine spectral characteristics of dioxoisobacteriochlorins (porphyrindione). We will compare the properties of the purified cd1 enzyme and its isolated d1 prosthetic group with those of a variety of positional and substitutional isomers of oxo-chlorin and dioxo-iBC model complexes. 4) We plan to extend these studies to the analysis of sirohemes and more highly saturated porphyrins, e.g., tetrahydroporphyrins and hexahydroporphyrins, to establish spectral properties which are specific for each level of saturated porphyrin. 5) Myoglobin and other heme proteins will be reconstituted with iron chlorin complexes. We will examine the spectral properties of these chlorin-proteins in all accessible spin- and oxidation-states, as well as ligand complexes, in comparison with the protein-free model complexes.