15N-NMR STUDIES OF RHODOSPIRILLUM RUBRUM CYTOCHROME C2

红螺菌细胞色素 C2 的 15N-NMR 研究

• 批准号: 3284763
• 负责人: GARY M SMITH
• 金额: $ 4.51万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-12-01 至 1987-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3284763
• 关键词: Rhodospirillum; acidity /alkalinity; cytochrome c; hemoprotein structure; lysine; microorganism culture; nuclear magnetic resonance spectroscopy; oxidation reduction reaction; temperature

The structure and function of heme proteins has been an area of intense investigation for decades because of their involvement in the most fundamental biological processes such as oxygen transport, oxidative phosphorylation and photosynthesis. A large number of cytochromes, one-electron carrying heme proteins, are known that are involved in electron transport chains of all that the most anaerobic of organisms. Among these proteins, the cytochromes C constitute a group having very similar structure, with a few exceptions, but very different properties. It is a long range goal of this laboratory to determine how the structure of different cytochromes allow them to function optimally in their respective systems. The specificity involves both the matching of redox potentials to those of the other electron carriers in the system and the specificity of the binding interactions between them. Another interest of this laboratory is to develop 15N-NMR as a tool for the study of small proteins. Cytochromes C are appropriately sized proteins in which nitrogen atoms play unusually important roles in the structure and surface charge distribution of the protein. 15N can be incorporated biosynthetically into the cytochrome c2 of Rhodospirillum rubrum by supplying the organism with 15NH4 ion. The uniformly enriched cytochrome will be purified by existing procedures and its N-NMR spectrum examined as a function of pH, temperature and oxidation state. Resonance will be assigned as specifically as possible using known shifts of amino acid groups and double resonance techniques. Changes in chemical shifts of 15N resonances with pH will be correlated with ionizations known to occur in the molecule that are detected by changes in redox potential and visible absorption spectrum to determine which groups are capable of altering these properties. The time scale of the motions of resolved 15N resonances can be estimated from the value of the 1H-15N nuclear Overhouser effect. This information will be used to determine the rigidity of the different structural elements of the molecule. The local motion of the surface lysine residues might be used in future experiments to map out the site of interaction between the cytochrome and its redox partners.

血红素蛋白的结构和功能一直是研究的热点 调查了几十年，因为他们参与了最 基本的生物过程，如氧的运输，氧化 磷酸化和光合作用。 大量的细胞色素， 一种携带一个电子的血红素蛋白，已知参与 电子传递链的所有最厌氧的有机体。 在这些蛋白质中，细胞色素C构成一组具有非常 类似的结构，除了少数例外，但性质非常不同。 这个实验室的长期目标是确定 不同的细胞色素使它们能够在各自的细胞中发挥最佳功能。 各自的系统。 特异性包括氧化还原的匹配 系统中其他电子载流子的电势， 它们之间相互作用的特异性。 该实验室的另一个兴趣是开发15 N-NMR作为分析的工具。 研究小分子蛋白质。 细胞色素C是细胞中适当大小的蛋白质， 其中氮原子在结构中起着非常重要的作用， 蛋白质的表面电荷分布。 15 N可以合并 生物合成为红杜鹃的细胞色素c2， 为生物体提供15 NH 4离子。 均匀富集的细胞色素 将通过现有程序纯化，并检查其N-NMR光谱， pH值、温度和氧化状态的函数。 共振将是 使用已知的氨基酸位移尽可能具体地指定 组和双共振技术。 15 N化学位移的变化 与pH值的共振将与已知发生在 通过氧化还原电位的变化检测到的分子， 吸收光谱，以确定哪些基团能够改变这些 特性. 分辨的15 N共振的运动的时间尺度可以 根据1H-15 N核Overhouser效应的值估计。 这 信息将被用来确定不同的刚性 分子的结构元素。 表面的局部运动 赖氨酸残基可能会在未来的实验中使用，以绘制出的网站 细胞色素和它的氧化还原伙伴之间的相互作用。