FE-S ENSEMBLES RELATED TO HYDROGENASE ACTIVITY

与氢化酶活性相关的 FE-S 系综

• 批准号: 6089923
• 负责人: Thomas Rauchfuss
• 金额: $ 16.16万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-07-01 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6089923
• 关键词: chemical models; enzyme activity; enzyme mechanism; hydrogenase; iron sulfur protein; metalloenzyme; model design /development

The project goals are to develop molecular level understanding of the hydrogenase metalloenzymes, which effect the interconversion of H2 and H+. Now is the perfect time to attack this problem from the modeling perspective because the active sites have recently been elucidated by X-ray crystallography on four enzymes, two [Fe]- and two [NiFe]-hydrogenases. The structural similarities of these enzymes and their extraordinary features (with respect to other metal sites in biology) strongly suggests that mechanistic insights obtained for one metallo hydrogenase will be applicable to the others. Thus the study here is broadly applicable to the entire (metallo) hydrogenase problem. A specific impetus for this project is the pair of recent (1999, and late 1998) crystallographic results on the two Fe- hydrogenases. These reports describe a surprisingly simple core structure that is amenable to chemical synthesis from organometallic precursors, an area of specific expertise to the PI. Our plans for preparing active site models are supported by compelling preliminary evidence that the Fe2(SR)2(CO)2(CN)2 core of the [Fe]-hydrogenase can be efficiently synthesized. Crystallographic analysis of our model confirms that it displays many of the structural characteristics seen in the two most recent protein crystallography reports. Furthermore, the synthetic analogue reduces protons to dihydrogen. In other words, we begin the project with a model that has fidelity with nature in terms of both structure and function. It is therefore quite likely that we will be able to obtain considerable molecular detail about how nature processes hydrogen. The first part of the project involves the preparation of accurate structural models for the Fe-hydrogenase. This preparative effort is coupled to spectroscopic characterization as well as mechanistic studies on the hydrogenase activity of the models. The synthetic effort entails both stepwise methods and multicomponent assembly processes starting from the most primitive precursor reagents. These models are subjected to protonation studies to give H2. The second part of the project is broadly aimed at gaining an understanding of the electronic and structural factors that determine the function of nonheme Fe centers to bind and activate H2. These studies probe libraries of thiolato, amino, and cyano ligation to afford Fe centers capable of activating H2, with connections to the binding and activation of RCN, N2, and CO. Initial studies are already promising with respect to LFe(CN)3-, LFe(CN)2(CO), LFe(SR)(CO)2+, and LFe(SR)2(CO) in conjunction with diversity in L as well. These studies will define how nature uses Fe-S and Fe-Ni-S ensembles to activate weakly basic small molecule substrates.

该项目的目标是在分子水平上了解氢酶金属酶，它影响氢和H+的相互转化。现在是从建模的角度解决这个问题的完美时机，因为最近四种酶，两种[Fe]-氢酶和两种[NiFe]-氢酶的X-射线结晶学已经阐明了活性部位。这些酶的结构相似及其非凡的特征(相对于生物学中的其他金属位置)强烈表明，从一个金属氢酶获得的机械洞察力将适用于其他金属氢酶。因此，这里的研究广泛适用于整个(金属)氢酶问题。这个项目的一个特殊推动力是最近(1999年和1998年底)关于两个铁氢酶的结晶学结果。这些报告描述了一种令人惊讶的简单的核心结构，它可以从有机金属前体进行化学合成，这是PI的一个特殊专业领域。我们的制备活性中心模型的计划得到了令人信服的初步证据的支持，即可以有效地合成[Fe]-氢酶的Fe2(SR)2(CO)2(CN)2核心。对我们的模型进行的结晶学分析证实，它显示了最近两份蛋白质结晶学报告中看到的许多结构特征。此外，合成的类似物将质子还原为氢气。换句话说，我们从一个在结构和功能上都与自然保持逼真的模型开始这个项目。因此，我们很可能能够获得有关自然如何处理氢的相当大的分子细节。该项目的第一部分涉及为铁氢酶准备准确的结构模型。这一准备工作与光谱表征以及对模型氢酶活性的机理研究相结合。合成工作包括从最原始的前体试剂开始的分步方法和多组分组装过程。这些模型受到质子化研究的影响，以得到H2。该项目的第二部分主要是为了了解电子和结构因素，这些因素决定了非血红素铁中心结合和激活H2的功能。这些研究探索了硫代、氨基和氰基连接的文库，以提供能够激活H2的Fe中心，并与RCN、N2和CO的结合和激活有关。初步研究表明，LFe(CN)3-、LFe(CN)2(CO)、LFe(SR)(CO)2+和LFe(SR)2(CO)与L的多样性有关。这些研究将确定自然界如何使用Fe-S和Fe-Ni-S系综来激活弱碱性小分子底物。