NMR and DFT Investigation of Porphyrin Conformation in Cytochromes c

细胞色素 c 中卟啉构象的 NMR 和 DFT 研究

• 批准号: 7871457
• 负责人: Matthew D Liptak
• 金额: $ 4.76万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7871457
• 关键词: Biochemical Process; Biological; Biological Models; Biological Process; Biology; Catalysis; Cell Nucleus; Chemicals; Circular Dichroism; Computing Methodologies; Crystallization; DNA Sequence Rearrangement; Data; Data Reporting; Development; Disease; Electron Spin Resonance Spectroscopy; Electron Transport; Electronics; Family; Future; Gases; Goals; Heme; Hemeproteins; Human; Hydrogen Bonding; Investigation; Label; Link; Measures; Methods; Molecular Conformation; Mutation; NMR Spectroscopy; Nuclear Magnetic Resonance; Organism; Oxygen; Porphyrins; Positioning Attribute; Process; Propionates; Protein Family; Proteins; Pyrroles; Raman Spectrum Analysis; Research Proposals; Sampling; Shapes; Side; Solutions; Structure; System; Techniques; United States National Institutes of Health; Variant; X-Ray Crystallography; absorption; base; biological systems; cofactor; covalent bond; cytochrome c; density; electronic structure; oxygen transport; polypeptide; public health relevance; research study; small molecule; structural biology; theories; tool

DESCRIPTION (provided by applicant): The long-term objective of this project is to develop the first experimental tool capable of quantitatively measuring the conformation of heme in solution. This tool will be an important contribution to the structural biology initiative of the NIH roadmap, which could be used in future studies that determine the nuclear magnetic resonance (NMR) solution structure of heme proteins. Heme proteins have a diverse set of biological functions, including: electron transport, gas transport, small molecule sensing, and catalysis. Each heme protein optimizes the reactivity of the heme cofactor to achieve its specific function and heme conformation is arguably the most under-investigated strategy available. It is essential to understand this determinant of heme electronic structure and reactivity to fully comprehend the biochemical processes involving heme proteins, and diseases associated with their malfunction. The cytochrome c family of proteins will be used as a model system to develop this tool because this family utilizes a nonplanar heme cofactor and the protein matrix provides a strategy to alter the heme conformation in a systematic way that is unlikely to introduce large-scale protein conformational changes. Paramagnetic 1H and 13C NMR spectroscopy will be used as a spectroscopic probe because this technique can simultaneously examine the electronic structure of heme at several positions on the porphyrin macrocycle. Multi-dimensional NMR experiments and selective isotopic enrichment of the heme cofactor will be used to make heme paramagnetic 1H and 13C resonance assignments. Density functional theory (DFT) calculations, initially assisted by X-ray crystallography and resonance Raman spectroscopy, will be used to interpret the paramagnetic NMR data in terms of out-of-plane distortions of the heme cofactor. Ultimately, this strategy will produce a DFT-based correlation between paramagnetic NMR data and heme conformation. This new tool can be applied to a wide range of heme proteins that have important functions within biological systems. The DFT-based correlation will also identify the relationship between heme conformation, electronic structure, and reactivity. PUBLIC HEALTH RELEVANCE: Biological organisms, including humans, use heme to carry out a diverse set of fundamental processes. To maximize the efficiency of a specific process, biological systems manipulate heme and its surroundings. Arguably, the most under-investigated manipulation available to heme proteins is changing the shape, and this proposal will develop the first method to measure the shape of heme in solution.

描述（由申请人提供）：本项目的长期目标是开发第一个能够定量测量溶液中血红素构象的实验工具。该工具将对NIH路线图的结构生物学倡议做出重要贡献，该路线图可用于确定血红素蛋白质的核磁共振（NMR）溶液结构的未来研究。血红素蛋白具有多种生物学功能，包括：电子传输、气体传输、小分子传感和催化。每种血红素蛋白优化血红素辅因子的反应性以实现其特定功能，血红素构象可以说是研究最少的策略。理解血红素电子结构和反应性的决定因素对于充分理解涉及血红素蛋白的生化过程以及与其功能障碍相关的疾病至关重要。细胞色素c家族的蛋白质将被用作一个模型系统来开发这个工具，因为这个家庭利用一个非平面血红素辅因子和蛋白质基质提供了一个策略，以改变血红素构象的系统的方式，是不太可能引入大规模的蛋白质构象变化。顺磁1H和13 C NMR光谱将被用作光谱探针，因为这种技术可以同时检查在卟啉大环上的几个位置的血红素的电子结构。多维NMR实验和选择性同位素富集的血红素辅因子将被用来使血红素顺磁1H和13 C共振分配。密度泛函理论（DFT）计算，最初由X射线晶体学和共振拉曼光谱学的协助下，将被用来解释顺磁NMR数据的血红素辅因子的平面外扭曲。最终，这种策略将产生基于DFT的顺磁NMR数据和血红素构象之间的相关性。这种新工具可以应用于在生物系统中具有重要功能的各种血红素蛋白。基于DFT的相关性还将确定血红素构象、电子结构和反应性之间的关系。公共卫生相关性：生物有机体，包括人类，使用血红素进行一系列不同的基本过程。为了使特定过程的效率最大化，生物系统操纵血红素及其周围环境。可以说，血红素蛋白质最缺乏研究的操纵是改变形状，这项提议将开发第一种测量溶液中血红素形状的方法。