OPTICAL STUDIES OF HEME PROTEIN DYNAMICS

血红素蛋白动力学的光学研究

• 批准号: 2181119
• 负责人: R DWAYNE MILLER
• 金额: $ 11.53万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-12-01 至 1996-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2181119
• 关键词: Raman spectrometry; bioenergetics; biomechanics; carbon monoxide; chemical association; chemical models; conformation; heme; hemoglobin; ligands; molecular dynamics; myoglobin; optical polarization; protein structure; spectrometry; thermodynamics; time resolved data

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.

拟议研究的具体目标是确定主导 使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，随后发生 血红素蛋白的反应功能是进化到其脱氧三级 结构。 该运动涉及数千个相关位移 原子自由度。 蛋白质系统到底是如何进化的 传播结构变化是一般理解的核心 功能相关的蛋白质运动和分子协同性。 这 原子位移产生的力来自于势能 随着配体解离而形成的能量梯度。 长度尺度 这些力量分布在其上，路径的多样性， 运动的能量是关键问题。 在拟议的研究中，CO 的光解将被用作 光触发引发结构变化。 的重点是 研究是用各种方法直接监测结构松弛 蛋白质运动的光学探针。 选择 CO 作为主要 这些研究的配体，因为它在时间上表现出最小的重组 使结构松弛动力学复杂化的兴趣尺度。 到 解决感应力作用力的长度尺度问题 运动，需要使用确定结构松弛动力学 对蛋白质的不同长度尺度敏感的探针。 这 全局运动（长尺度运动）将使用 皮秒/飞秒相位光栅光谱，而局部运动 Fe-CO 结合位点的力中心将集中在 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 在触发的蛋白质反应的初始阶段。 一个新的 基于飞秒光学散射和光学克尔的技术 效果检测将提供低频的直接测量 与结构松弛坐标耦合的集体模式。 这 不同阶段的整体能量学或驱动力 运动将跟随相位光栅光谱修改为 有选择地研究热引起的密度变化。 这个热阶段 光栅方法在时间方面处于基本极限 用于测定生物能量学的分辨率。 它有足够的 时间分辨率和灵敏度区分集体模式 和构象亚状态模型作为初始的主导相 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 运动。 这些研究将扩展到微秒范围，以便 从最初的飞秒/皮秒就可以建立完整的连接 引发较长时间尺度结构运动的动力学 相对于功能的放松。