NMR SPECTROSCOPY OF IRON-SULFUR PROTEINS

铁硫蛋白的核磁共振波谱

• 批准号: 7007744
• 负责人: JOHN LUTE MARKLEY
• 金额: $ 24.74万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-02-01 至 2007-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7007744
• 关键词: NMR; SPECTROSCOPY; IRON; SULFUR; PROTEINS

DESCRIPTION (provided by applicant): Work carried out in previous project periods has demonstrated that the combination of NMR spectroscopy, X-ray crystallography, and quantum chemical calculations serves as a highly productive approach to understanding sequence-structure-function relationships in iron-sulfur proteins. NMR spectra contain exquisitely sensitive information about electron nuclear interactions. This information, which is unavailable from X-ray crystal structures, provides insights into the chemical properties of the iron centers, details of their geometry, strengths of hydrogen bonds, and patterns of electron delocalization. NMR thus serves as a window for viewing the properties of the cluster that control redox potentials and regulate pathways of electron transfer. We have shown that high-level quantum chemical approaches can successfully correlate experimental NMR observables with geometric structures provided by high-resolution X-ray crystallography, and thus provide detailed insight to the factors tuning the properties of the metal site. In turn, NMR data and quantum calculations can provide structural constraints in cases where X-ray crystallography has not been feasible at sufficient resolution. We propose to use this coordinated approach to investigate a series of 1Fe and [2Fe-2S] proteins that are model systems in the study of electron transfer, with the aim of answering a number of questions regarding their structure-function relationships. The proteins to be studied include: a series of clostridial rubredoxins (monomeric 1Fe proteins) that have altered redox potentials, desulforedoxin (dimeric 1Fe protein), and four prototypical [2Fe-2S] proteins that have distinct spectral and functional properties (Anabaena vegetative ferredoxin, human ferredoxin, a Rieske protein, and Aquifex aeolicus ferredoxin). We propose, not only to collect and analyze NMR data, but also to collaborate with others to obtain crystal structures and electron-nuclear double resonance (ENDOR) data. An X-ray structure for reduced rubredoxin recently became available, but currently no structures are available for any reduced [2Fe-2S] proteins. Effort will be expended toward improving the calculations for 1Fe proteins and extending the methodology to [2Fe-2S] proteins. Effort will also be expended toward developing new methodology for paramagnetic NMR, in particular, for systems that are not amenable to established approaches.

描述（由申请人提供）：在之前的项目期间开展的工作已经证明，核磁共振波谱，x射线晶体学和量子化学计算的结合可以作为一种高效的方法来理解铁硫蛋白的序列-结构-功能关系。核磁共振波谱包含有关电子核相互作用的非常敏感的信息。这些信息是无法从x射线晶体结构中获得的，它提供了对铁中心的化学性质、几何形状细节、氢键强度和电子离域模式的深入了解。因此，核磁共振可以作为一个窗口，用于观察控制氧化还原电位和调节电子转移途径的簇的性质。我们已经证明，高水平的量子化学方法可以成功地将实验核磁共振观测结果与高分辨率x射线晶体学提供的几何结构相关联，从而提供了对调整金属位点性质的因素的详细见解。反过来，核磁共振数据和量子计算可以在x射线晶体学在足够分辨率下不可行的情况下提供结构约束。我们建议使用这种协调的方法来研究一系列的1Fe和[2Fe-2S]蛋白质，它们是电子转移研究中的模型系统，目的是回答一些关于它们的结构-功能关系的问题。要研究的蛋白质包括：一系列具有改变氧化还原电位的梭菌氧化还原蛋白（单体铁蛋白），脱硫还蛋白（二聚体铁蛋白），以及四种具有不同光谱和功能特性的原型[2Fe-2S]蛋白（Anabaena vegetative铁氧还蛋白，人铁氧还蛋白，Rieske蛋白和Aquifex aeolicus铁氧还蛋白）。我们建议，不仅要收集和分析核磁共振数据，而且要与他人合作，以获得晶体结构和电子-核双共振（ENDOR）数据。最近有了还原红氧还蛋白的x射线结构，但目前没有任何还原[2Fe-2S]蛋白的x射线结构。将努力改进1Fe蛋白的计算，并将方法扩展到[2Fe-2S]蛋白。还将努力开发顺磁核磁共振的新方法，特别是那些不适合现有方法的系统。