Fast Kinetic Studies of Protein Folding and Function

蛋白质折叠和功能的快速动力学研究

• 批准号: 6874802
• 负责人: DAVID S. KLIGER
• 金额: $ 32.59万
• 依托单位: UNIVERSITY OF CALIFORNIA SANTA CRUZ
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6874802
• 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

DESCRIPTION (provided by applicant): This project applies techniques for fast time-resolved magnetic circular dichroism (TRMCD), natural circular dichroism (TRCD, and ordinary absorption spectroscopies to the study of function in heme proteins and folding in heme proteins and small peptides. The novel optical methods employed use near-null ellipsometry and polarimetry to study rapid kinetic processes (nanosecond to seconds) in biomolecules that contain magneto-optically active chromophores, such as heme and the aromatic amino acids, and naturally chiral chromophores, such as the amide groups of proteins and peptides. These techniques will be used to identify and study the earliest (submillisecond) events in the folding reactions of heme proteins such as cytochrome c. A major goal is the determination of a parameter that is fundamental to understanding the nature of protein folding: the speed with which the different unfolded conformations interconvert with one another. If this is slow compared to folding itself, then understanding protein folding will require more complicated theories (e.g., energy landscape) than the transition state theory used for typical chemical reactions. Such understanding may ultimately prove helpful in developing therapies for the many diseases associated with protein misfolding, such as cystic fibrosis, type 2 diabetes, and Alzheimer's, Parkinson's, and Creutzfeldt- Jakob disease. A major goal of the functional studies is to understand how the four subunits that make up the hemoglobin molecule cooperate with each other to transport oxygen more efficiently. A recent hypothesis about this cooperativity (Ackers symmetry rule), based originally on thermodynamic measurements, is tested by kinetic measurements in this project. A novel model for hemoglobin allostery, emerging from this linkage of thermodynamics and kinetics, holds promise for simplifying and systematizing our understanding of hemoglobin's dynamics and its control in the body by allosteric effectors such as organic phosphates. In addition, TRMCD studies of the aromatic amino acid residue tryptophan [337, positioned at a site critical for cooperativity, are intended to further clarify how hemoglobin's subunits work together as an efficient "molecular machine" for transporting oxygen from the lungs to the tissues.

描述（由申请人提供）：该项目应用快速时间分辨磁圆二色性（TRMCD）、天然圆二色性（TRCD）和普通吸收光谱技术来研究血红素蛋白的功能以及血红素蛋白和小肽的折叠。所采用的新颖光学方法使用近零椭圆偏振和偏振测量来研究快速动力学过程 （纳秒到秒）包含磁光活性发色团（例如血红素和芳香族氨基酸）以及天然手性发色团（例如蛋白质和肽的酰胺基团）的生物分子。这些技术将用于识别和研究血红素蛋白（如细胞色素 c）折叠反应中最早（亚毫秒）的事件。一个主要目标是 确定对于理解蛋白质折叠本质至关重要的参数：不同未折叠构象相互转化的速度。如果这比折叠本身慢，那么理解蛋白质折叠将需要比用于典型化学反应的过渡态理论更复杂的理论（例如能量景观）。这种理解最终可能有助于开发与蛋白质错误折叠相关的许多疾病的疗法，例如 如囊性纤维化、2 型糖尿病、阿尔茨海默病、帕金森病和克雅氏病。功能研究的一个主要目标是了解构成血红蛋白分子的四个亚基如何相互配合以更有效地运输氧气。关于这种协同性的最新假设（阿克斯对称规则）最初基于热力学 测量，在该项目中通过动力学测量进行测试。从热力学和动力学的这种联系中出现的一种新的血红蛋白变构模型有望简化和系统化我们对血红蛋白动力学及其通过有机磷酸盐等变构效应物在体内的控制的理解。此外，TRMCD 对芳香族氨基酸残基色氨酸的研究[337， 位于对协作性至关重要的位置，旨在进一步阐明血红蛋白的亚基如何作为有效的“分子机器”协同工作，将氧气从肺部输送到组织。