OPTICAL STUDIES OF HEME PROTEIN DYNAMICS

建议研究的具体目标是确定主导 利用肌红蛋白和血红蛋白刺激蛋白质运动的机制 蛋白质作为模型系统。 配体解离后， 血红素蛋白的反应功能是进化到其脱氧三级 结构 这种运动涉及到数千人的相关位移 原子的自由度。 蛋白质系统是如何进化的， 传播的结构变化是核心的一般理解 功能相关的蛋白质运动和分子协同性。 的 原子位移产生的力来自于 配体解离产生的能量梯度。 的长度尺度 这些力量分布的区域，路径的多样性， 和运动的能量学是关键问题。 在拟议的研究中，CO的光解将被用作一种 光学触发器来启动结构变化。 的重点 研究是直接监测结构松弛与各种 蛋白质运动的光学探针。 CO被选为主要的 配体，因为它表现出最小的重组的时间 感兴趣的尺度使结构弛豫动力学复杂化。 到 解决诱导的作用力的长度尺度问题， 运动时，需要使用 对蛋白质的不同长度尺度敏感的探针。 的 全局运动（长尺度运动）将使用 皮秒/飞秒相位光栅光谱，而运动局部 在铁-一氧化碳结合点的力量的中心将集中在 近端组氨酸运动的时间分辨拉曼探针。这些 研究将确定集体原子位移的程度 在触发蛋白质反应的初始阶段。 一个新 基于飞秒光致伸缩散射和光克尔技术 效应检测将提供对低频的直接测量 集体模式耦合到结构弛豫坐标。 的 不同阶段的总能量或驱动力 运动之后将进行相位光栅光谱学修改， 选择性地研究热致密度变化。 这个热阶段 光栅法在时间上处于基本极限 用于生物能量学测定的分辨率。 其具有足够 时间分辨率和灵敏度，以区分集体模式 和构象亚态模型作为初始阶段的主导阶段 蛋白质结构变化 相位光栅的组合使用 光谱学、飞秒自由散射和能量学给出了 研究蛋白质力学的综合实验方法 议案 这些研究将扩展到微秒范围， 可以从初始的飞秒/皮秒 在较长的时间尺度上启动运动的动力学结构 相对于功能性的松弛。

The specific goal of the proposed research is to determine the dominant mechanism for stimulated protein motion using myoglobin and hemoglobin proteins as model systems. Following ligand dissociation, the ensuing response function of heme proteins is to evolve to its deoxy tertiary structure. This motion involves the correlated displacement of thousands of atomic degrees of freedom. Exactly how the protein system evolves and propagates the structural changes is central to a general understanding of functionally relevant protein motion and molecular cooperativity. The forces that develop for the atomic displacements arise from the potential energy gradients that develop with ligand dissociation. The length scale over which these forces are distributed, the multiplicity of pathways, and the energetics for the motion are the key issues. In the proposed studies, the photodissociation of CO will be used as an optical trigger to initiate the structural changes. The emphasis of the research is on directly monitoring the structural relaxation with various optical probes to the protein motion. CO was chosen as the primary ligand for these studies as it exhibits minimal recombination on the time scale of interest to complicate the structural relaxation dynamics. To address the issue of length scale for the acting forces on the induced motion, one needs to determine the structural relaxation dynamics using probes that are sensitive to different length scales of the protein. The global motion (long length scale motion) will be followed using picosecond/femtosecond phase grating spectroscopy, while motion local to the epicenter for the forces at the Fe-CO binding site will focus on time-resolved Raman probes of the proximal histidine motion. These studies will determine the degree of collective atomic displacements during the initial phase of the triggered protein response. A new technique based on femtosecond librational scattering and optical Kerr effect detection will provide a direct measurement of the low frequency collective modes coupled to the structural relaxation coordinate. The overall energetics or driving force for the different phases of the motion will be followed by phase grating spectroscopy modified to selectively study thermally induced density changes. This thermal phase grating method is at the fundamental limit with respect to time resolution for the determination of bioenergetics. It has sufficient time resolution and sensitivity to distinguish between collective mode and conformational substate models as the dominant phase for the initial protein structural changes. The combined use of phase grating spectroscopy, femtosecond librational scattering, and energetics give a comprehensive experimental approach for studying the mechanics of protein motion. These studies will be extended to the microsecond range so that a complete connection can be made from the initial femtosecond/picosecond dynamics that initiate the motion to the longer time scale structural relaxations relative to functionality.