SINGLE CRYSTAL XAS STUDIES OF NITROGENASE PROTEINS

固氮酶蛋白的单晶 XAS 研究

• 批准号: 8169950
• 负责人: BRITT HEDMAN
• 金额: $ 0.34万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8169950
• 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; 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利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 固氮酶系统催化生物固氮的基本过程。它由发生底物还原的MoFe蛋白和作为MoFe蛋白的特定电子供体的Fe蛋白组成。FeMoco是MoFe蛋白中的一个[MoFe7S6X]簇，被认为是氮催化的位置。结晶学最近发现了位于FeMoco核心的中心低Z原子X(O.Einsle等人)。科学297,1696(2002))。FeMoco中间隙原子的存在提出了关于这个簇的结构、生物合成和反应活性的新问题，我们希望用单晶XAS来解决这些问题。极化单晶XAS的优点是提供关于蛋白质中与已知蛋白质三级结构相关的金属位置的局部结构和电子细节，并且进一步地，可以用于通过与光束极化矢量的晶体对准来选择性地增强特定的分子载体。我们的初步结果表明，应用这一技术来询问FeMoco的中心原子是可行的。本研究的目的是进一步发展和利用单晶XAS来研究固氮酶体系中的重要问题，包括：(1)FeMoco中间隙原子的检测和定量；(2)通过对催化不活跃的MoFe蛋白的研究，在没有N_2催化的情况下测定FeMoco的结构，特别是关于间隙原子的存在；(3)全铁蛋白质中Fe-Fe距离与全局蛋白质结构的关联，以了解这个不同的簇如何在催化中发挥作用；(4)通过对辐射还原、底物孵育的MoFe蛋白的研究，阐明底物和抑制剂与FeMoco的相互作用机理。