TIME-RESOLVED MAGNETIC CIRCULAR DICHROISM

时间分辨磁圆二色性

• 批准号: 6127433
• 负责人: DAVID S. KLIGER
• 金额: $ 27.97万
• 依托单位: UNIVERSITY OF CALIFORNIA SANTA CRUZ
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-07-01 至 2004-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6127433
• 关键词: allosteric site; chemical kinetics; circular magnetic dichroism; conformation; cytochrome c; cytochrome oxidase; electron transport; hemoglobin; hemoprotein structure; myoglobin; optical rotation; photolysis; protein folding; protein structure function; structural biology; time resolved data; ultraviolet radiation

This project will apply techniques for fast time-resolved magnetic circular dichroism (TRMCD) and magnetic optical rotatory dispersion (TRMORD) spectroscopies to the study of function and folding in heme proteins. The novel optical methods employed use quasi-null ellipsometry and polarimetry to study rapid kinetic processes (nanosecond to second time scales) in biomolecules that contain magneto-optically active chromophores such as heme and the aromatic amino acids. The overall program of the functional ligand-rebinding studies is to investigate protein relaxation after ligand photolysis with the goal of understanding how protein structure modulates the reactivity of the heme prosthetic group and elicits the variety of functions that heme proteins perform in oxidative metabolism. The proposed TRMCD studies of kinetic intermediates in the oxygen transport protein hemoglobin have the goal of better understanding the dynamics of the allosteric R yields T transition in this cooperative system. The TRMCD spectra of the near-UV tryptophan bands and the heme-based Soret and visible bands will be examined for MCD transients on the nanosecond and microsecond timescales that are characteristic of globin tertiary and quaternary structural changes after photolysis of the R-state carboxy adduct. Similarly, TRMCD/MORD studies of ligand photolysis in myoglobin will be directed toward resolving outstanding questions about the heme pocket dynamics underlying the function of this oxygen storage protein. Molecular oxygen is ultimately consumed in cells by redox reactions catalyzed at a copper-heme iron site in the enzyme cytochrome c oxidase. The TRMCD studies of ligand dynamics at the bimetallic site proposed here are ultimately addressed at a major puzzle in biophysics: How does cytochrome oxidase couple the energy released in this redox chemistry to the pumping of protons against a gradient? Finally, in the other major direction of investigation proposed, TRMCD/MORD techniques will be used to monitor ultrafast (submillisecond) events in the folding reactions of heme proteins. In particular, this work will look for direct spectrokinetic evidence for the type of biased diffusional dynamics thought to characterize the earliest events in the folding of protein chains in the new, energy landscape point of view.

该项目将对快速分辨的磁圆二色性（TRMCD）和磁光旋转分散（TRMORD）光谱应用技术应用技术来研究血红素蛋白的功能和折叠。 使用准无椭圆法和极化法研究的新型光学方法研究了含有磁性活性发色体（如Heme和芳香族氨基酸）的生物分子中的快速动力学过程（纳秒至第二次尺度）。 功能性配体重新结合研究的总体程序是研究配体光解时蛋白质松弛，目的是了解蛋白质结构如何调节血红素假体组的反应性，并引起血红素蛋白在氧化代谢中的功能。 拟议的TRMCD研究在氧转运蛋白血红蛋白中的动力学中间体的目的是更好地理解变构r的动力学在该合作系统中产生T转变。 将检查近紫外色氨酸带和血红素的SORET和可见条带的TRMCD光谱，以在纳秒和微秒的时间尺度上进行MCD瞬变，这些瞬态是环球蛋白第三纪念和Quaternary结构性变化的特征，并在光态型牛boxy添加后光解。同样，肌红蛋白中配体光解的TRMCD/MORD研究将针对解决该氧气储存蛋白功能的血红素袋动力学的出色问题。最终，通过在酶细胞色素C氧化酶的铜血红素位点催化的氧化还原反应最终在细胞中消耗分子氧。 最终在生物物理学的主要难题中解决了双金属部位的配体动力学的TRMCD研究：细胞色素氧化酶对氧化还原化学中释放的能量如何泵送质子泵送质子，以促进梯度？ 最后，在提出的另一个主要研究方向上，TRMCD/MORD技术将用于监测血红素蛋白折叠反应中的超快（亚颌骨）事件。 特别是，这项工作将寻找直接的光谱证据，以表明在新的能量景观观点中蛋白质链折叠中最早事件的偏置扩散动力学类型。