TIME-RESOLVED MAGNETIC CIRCULAR DICHROISM

该项目将应用快速时间分辨磁圆二色性（TRMCD）和磁旋光色散（TRMORD）光谱技术来研究血红素蛋白的功能和折叠。 采用的新颖光学方法使用准零椭圆偏振和偏振测量来研究含有磁光活性发色团（例如血红素和芳香氨基酸）的生物分子中的快速动力学过程（纳秒到秒时间尺度）。 功能性配体重新结合研究的总体计划是研究配体光解后的蛋白质松弛，目的是了解蛋白质结构如何调节血红素辅基的反应性并引发血红素蛋白在氧化代谢中发挥的各种功能。 拟议的氧转运蛋白血红蛋白动力学中间体的 TRMCD 研究的目的是更好地了解该协作系统中变构 R 产生 T 转变的动力学。 将检查近紫外色氨酸带和基于血红素的 Soret 和可见带的 TRMCD 光谱，以了解纳秒和微秒时间尺度上的 MCD 瞬态，这是 R 态羧基加合物光解后珠蛋白三级和四级结构变化的特征。同样，TRMCD/MORD 对肌红蛋白中配体光解作用的研究将致力于解决有关这种储氧蛋白功能背后的血红素袋动力学的突出问题。分子氧最终在细胞中通过细胞色素 c 氧化酶中铜-血红素铁位点催化的氧化还原反应被消耗。 这里提出的双金属位点配体动力学的 TRMCD 研究最终解决了生物物理学中的一个主要难题：细胞色素氧化酶如何将这种氧化还原化学中释放的能量与逆梯度泵浦质子耦合起来？ 最后，在提出的另一个主要研究方向中，TRMCD/MORD 技术将用于监测血红素蛋白折叠反应中的超快（亚毫秒）事件。 特别是，这项工作将寻找偏向扩散动力学类型的直接光谱动力学证据，该类型被认为可以表征新能源景观观点中蛋白质链折叠的最早事件。

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.