SINGLE CRYSTAL XAS STUDIES OF NITROGENASE PROTEINS

固氮酶蛋白的单晶 XAS 研究

• 批准号: 7954220
• 负责人: BRITT HEDMAN
• 金额: $ 0.21万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-03-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7954220
• 关键词: Address; Anabolism; Biological Process; Catalysis; Computer Retrieval of Information on Scientific Projects Database; Crystallography; Detection; Electronics; Enzymes; Funding; Grant; Incubated; Institution; Metals; Molybdoferredoxin; Nitrogen; Nitrogen Fixation; Nitrogenase; Proteins; Research; Research Personnel; Resources; Science; Site; Source; Structure; System; Techniques; Tertiary Protein Structure; United States National Institutes of Health; electron donor; inhibitor/antagonist; interstitial; molecular vector; protein structure; structural biology; synchrotron radiation; vector

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The nitrogenase enzyme system catalyzes the essential process of biological nitrogen fixation. It is composed of the MoFe protein, where substrate reduction occurs, and the Fe protein, which is the specific electron donor to the MoFe protein. FeMoco, a [MoFe7S6X] cluster in the MoFe protein, is believed to be the site of nitrogen catalysis. The central low-Z atom, X, at the core of FeMoco was recently discovered by crystallography (O. Einsle, et al. Science 297, 1696 (2002)). The presence of an interstitial atom in FeMoco raises new questions about the structure, biosynthesis, and reactivity of this cluster that we hope to address using single crystal XAS. Polarized single crystal XAS has the advantage of providing both local structural and electronic detail about a metal site in a protein that is related to a known protein tertiary structure and, further, can be used to selectively enhance specific molecular vectors through crystallographic alignment with the beam polarization vector. We have preliminary results that indicate the feasibility of applying this technique to interrogate the central atom of FeMoco. The aim of this study is to further develop and utilize single crystal XAS for the study of important questions in the nitrogenase system including (1) detection and quantification of an interstitial atom in FeMoco; (2) determination of FeMoco structure, especially with regard to the presence of an interstitial atom, in the absence of N2 catalysis through the study of a catalytically inactive MoFe protein; (3) correlation of the Fe-Fe distances in the all-ferrous Fe protein with the global protein structure to gain an understanding of how this distinct cluster may function in catalysis; and (4) elucidation of the mechanism of substrate and inhibitor interaction with FeMoco through the study of radiolytically reduced, substrate-incubated MoFe proteins.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 固氮酶系统催化生物固氮的基本过程。它由MoFe蛋白和Fe蛋白组成，其中MoFe蛋白发生底物还原，Fe蛋白是MoFe蛋白的特异性电子供体。FeMoco是MoFe蛋白中的[MoFe 7S 6X]簇，被认为是氮催化的位点。最近通过晶体学发现了FeMoco核心的中心低Z原子X（O。Einsle等，Science 297，1696（2002））。FeMoco中间隙原子的存在提出了关于该簇的结构，生物合成和反应性的新问题，我们希望使用单晶XAS来解决这些问题。偏振单晶XAS具有提供关于蛋白质中的金属位点的局部结构和电子细节的优点，所述金属位点与已知蛋白质三级结构相关，并且还可以用于通过与光束偏振矢量的晶体学对准来选择性地增强特定分子矢量。我们有初步的结果表明，应用这种技术来询问FeMoco的中心原子的可行性。本研究的目的是进一步发展和利用单晶XAS来研究固氮酶系统中的重要问题，包括（1）FeMoco中间隙原子的检测和定量，（2）通过对无催化活性的MoFe蛋白的研究来确定FeMoco结构，特别是关于间隙原子的存在，在无N2催化的情况下;（3）全铁Fe蛋白中的Fe-Fe距离与整体蛋白质结构的相关性，以了解这种独特的簇如何在催化中起作用;以及（4）通过研究放射性还原的底物孵育的MoFe蛋白，阐明底物和抑制剂与FeMoco相互作用的机制。