Spectroscopic Studies of Molybdoenzymes & Models

 描述（由申请人提供）：关于钼辅因子硫酸化酶C-末端（MOSC）结构域蛋白质结构和功能，以及复杂的吡喃蝶呤二硫纶配体在所有吡喃蝶呤钼酶的催化循环中的作用，知识库中存在根本性空白。我们的长期目标是开发一种分子电子结构，了解吡喃蝶呤钼酶的作用机制和功能，以便对人类健康质量产生积极的影响。我们的主要目标是确定1）电子结构对吡喃蝶呤二硫纶功能的贡献，2）几何和电子结构对MOSC结构域蛋白反应性的贡献，以及3）为什么二甲基亚砜还原酶家族酶需要两个吡喃蝶呤二硫纶才能发挥作用。我们将通过使用组合光谱方法（电子吸收、MCD、拉曼、XAS、EPR等）来实现这些目标。通过振动，光谱，键合和反应坐标计算来探测吡喃蝶呤和催化机制。对明智选择的小分子类似物的平行研究将补充这种方法。中心的假设是，钼网站的几何和电子结构，加上性质的吡喃蝶呤二硫纶，有助于这些酶的独特功能。这项研究的基本原理是，全面了解MOSC家族蛋白质以及Mo离子和吡喃蝶呤二硫烯之间的复杂相互作用将为疾病状态提供新的见解，并对人类健康产生积极影响。我们将测试我们的中心假设，以便通过成功追求三个特定目标来实现本提案的既定目标：1）确定吡喃蝶呤如何有助于黄嘌呤氧化酶和亚硫酸盐氧化酶家族酶中的电子转移和氧化还原过程，2）了解定义其功能的MOSC结构域蛋白之间的结构和机制关系，以及3）了解两种不同吡喃蝶呤之间的配位数和合作关系如何有助于二甲基亚砜还原酶家族酶的催化。我们的贡献，预计将提供一个详细的了解MOSC介导的催化在分子水平上，之间的联系的吡喃蝶呤二硫杂环戊烯氧化态和作用的吡喃蝶呤二硫杂环戊烯在催化，和协调环境之间的关系，需要两个吡喃蝶呤二硫杂环戊烯在二甲基亚砜还原酶家族的酶。拟议的研究是有意义的，因为它将推进我们的理解翻译后硫化过程和钼辅因子的贩运，前药活化和异生物质解毒的机制，以及在pyranopterin dithiolene的电子灵活性在pyranodoenzyme催化。

 DESCRIPTION (provided by applicant): There exists a fundamental gap in the knowledge base regarding molybdenum cofactor sulfurase C-terminal (MOSC) domain protein structure and function, as well as the role of the complex pyranopterin dithiolene ligand in the catalytic cycles of all pyranopterin molybdenum enzymes. Our long-term goal is to develop a molecular electronic structure understanding of pyranopterin molybdenum enzyme mechanism and function in order to have a positive impact on the quality of human health. Our primary objectives are to determine 1) electronic structure contributions to pyranopterin dithiolene function, 2) geometric and electronic structure contributions to the reactivity of MOSC domain proteins, and 3) why dimethyl sulfoxide reductase family enzymes require two pyranopterin dithiolenes in order to function. We will accomplish these objectives by using a combined spectroscopic approach (electronic absorption, MCD, Raman, XAS, EPR, etc.) augmented by vibrational, spectroscopic, bonding, and reaction coordinate computations to probe the pyranopterin and catalytic mechanisms. Parallel studies on judiciously chosen small molecule analogs will complement this approach. The central hypothesis is that Mo site geometric and electronic structure, coupled with the nature of the pyranopterin dithiolene, contributes to the unique function of these enzymes. The rationale for this research is that a comprehensive understanding of MOSC family proteins and the complex interplay between the Mo ion and the pyranopterin dithiolene will provide new insights into disease states and have a positive impact on human health. We will test our central hypothesis in order to accomplish the stated objective of this proposal through the successful pursuit of three Specific Aims 1) Determine how the pyranopterin contributes to electron transfer and redox processes in xanthine oxidase and sulfite oxidase family enzymes, 2) Understand structural and mechanistic relationships between MOSC domain proteins that define their function, and 3) Develop an understanding of how coordination number and the cooperative relationship between two different pyranopterins contribute to catalysis in dimethyl sulfoxide reductase family enzymes. Our contribution is expected to provide a detailed understanding of MOSC mediated catalysis at the molecular level, the link between pyranopterin dithiolene oxidation state and the role of the pyranopterin dithiolene in catalysis, and the relationship between coordination environment and the need for two pyranopterin dithiolenes in dimethyl sulfoxide reductase family enzymes. The proposed research is significant since it will advance our understanding of post-translational sulfuration processes and molybdenum cofactor trafficking, mechanisms of prodrug activation and xenobiotic detoxification, and the electronic flexibility of the pyranopterin dithiolene in molybdoenzyme catalysis.