IRON-BOUND AZURIN AS MIMICS OF LOW COORDINATE IRON SITES OF NITROGENASES

作为固氮酶低配位铁位点模拟物的铁结合天青蛋白

• 批准号: 8362246
• 负责人: Robert Karoly Szilagyi
• 金额: $ 0.16万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-01 至 2012-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8362246
• 关键词: Azides; Azurin; Binding; Binding Sites; Bioinorganic Chemistry; Complex; Data; Electron Nuclear Double Resonance; Engineering; Environment; Funding; Grant; Iron; Ligands; Link; Metals; Modeling; Molybdoferredoxin; National Center for Research Resources; Nitrogenase; Principal Investigator; Proteins; Radiation; Research; Research Infrastructure; Resources; Site; Source; Spectrum Analysis; Structure; United States National Institutes of Health; absorption; cost; electronic structure; mutant; novel; oxidation; research study; small molecule; structural biology

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The mechanism of dinitrogen reduction at the iron-molybdenum cofactor (FeMoco) of nitrogenase is one of the great mysteries of bioinorganic chemistry, and recent evidence suggests that substrates bind at iron. However, nitrogenase has many iron atoms, making it difficult to analyze spectroscopically. There is an urgent need for one- and two-iron complexes with the same spin state and geometry as the iron atoms in nitrogenase, to use as spectroscopic models for interpretation of the data on nitrogenase. Our hypothesis is that reductive activation of the FeMoco generates transient unsaturated iron sites for substrate binding and reduction. These iron binding sites would have highspin iron in a ligand-poor environment, with a geometry quite different from that in most known iron complexes but similar to the FeMoco iron sites. Our preliminary evidence shows that the geometry of an iron complex has a dominant influence on its orbital energies, which in turn determine the strength of binding N2 and other small molecules. Therefore, the proposed research focuses on compounds in which the iron atom has similar geometry (trigonal pyramidal), electronic structure (high-spin with weak-field ligands) and spectroscopy as the FeMoco iron sites. The unusual coordination environment will be enforced by coordinating iron to mutants of the azurin apo-protein, which is known to constrain metals to a trigonal pyramidal geometry. The iron complexes will be evaluated by ENDOR, infrared, Raman, M¿ssbauer, and x-ray absorption. The spectroscopic results will provide a link between the structures of novel model compounds and the known data for nitrogenase, and the x-ray absorption experiments will show the oxidation level,geometry, and binding modes to the iron atom. Preliminary results from NMR indicate binding of azide to the iron atom in some engineered azurin-iron complexes, and we will use x-ray absorption to analyze this and other novel iron species.

该副本是利用资源的众多研究子项目之一 由NIH/NCRR资助的中心赠款提供。对该子弹的主要支持 而且，副投影的主要研究员可能是其他来源提供的 包括其他NIH来源。列出的总费用可能 代表subproject使用的中心基础架构的估计量， NCRR赠款不直接向子弹或副本人员提供的直接资金。 硝基化酶的铁溶剂辅因子（FEAMOCO）在硝基化合物（FEMOCO）上的二氮的机理是生物无机化学的最大奥秘之一，最近的证据表明底物在铁与铁结合。但是，氮酶具有许多铁原子，因此很难通过光谱镜进行分析。迫切需要具有与氮化合酶中的铁原子相同的单铁复合物，将其用作光谱模型，以解释硝化酶的数据。我们的假设是，减少了Femoco的激活会产生瞬态的不饱和铁位点，以实现底物结合和还原。这些铁结合位点将在配体贫乏的环境中具有高螺旋铁，其几何形状与大多数已知的铁配合物的几何形状完全不同，但类似于股骨铁位点。我们的初步证据表明，铁复合物的几何形状对其轨道能具有主要影响，这又决定了结合N2和其他小分子的强度。因此，提出的研究重点是铁原子具有相似的几何形状（三角形金字塔），电子结构（具有弱场配体的高旋转）和光谱与Femoco铁位点的化合物。不寻常的协调环境将通过协调的铁对硫蛋白apo蛋白的突变体强制执行，该突变体已知将金属限制为三角形金字塔几何形状。铁综合体将由Endor，Infrared，Raman，M ssbauer和X射线惊喜评估。光谱结果将提供新型模型化合物的结构与氮化合酶的已知数据之间的联系，X射线惊喜实验将显示与铁原子的氧化水平，几何形状和结合模式。 NMR的初步结果表明叠氮化物与某些工程化的Azurin-Iron配合物中的铁原子结合，我们将使用X射线惊喜来分析这种和其他新型的铁物种。