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Fast Kinetic Studies of Protein Folding and Function

蛋白质折叠和功能的快速动力学研究

基本信息

批准号：
7270641
负责人：
DAVID S. KLIGER
金额：
$ 31.42万
依托单位：
UNIVERSITY OF CALIFORNIA SANTA CRUZ
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-09-01 至 2009-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7270641
关键词：
Address Alzheimer&apos s Disease Amides Amino Acids Aromatic Amino Acids Biological Assay Cell Respiration Charge Circular Dichroism Cobalt Complement component C1s Condition Coupled Creutzfeldt-Jakob Syndrome Cyprinus carpio Cysteine Cystic Fibrosis Cytochromes b Diffusion Disease Electronics Equilibrium Equus caballus Event Free Energy Goals Heme Hemeproteins Hemoglobin Heterogeneity Histidine Hybrids Hydrogen Bonding Kinetics Lasers Ligand Binding Ligands Ligation Lung Measurement Measures Methionine Methods Modeling Molecular Conformation Molecular Machines Monitor Mutation Myoglobin NADH Nature Non-Insulin-Dependent Diabetes Mellitus Numbers Optical Methods Parkinson Disease Pathway interactions Peptides Phosphoric Acid Esters Plant Roots Positioning Attribute Process Proteins Range Rate Reaction Rehydrations Relaxation Reporting Site Spectrum Analysis Speed Structure Surface Techniques Temperature Testing Thermodynamics Time Tissues Tryptophan Tuna Variant Water Work absorption azobenzene base chemical property chemical reaction chromophore circular magnetic dichroism crosslink cytochrome c dimer fish Carp interfacial mutant nanosecond novel oxygen transport photolysis polarimetry polyol polypeptide protein folding protein function protein misfolding research study species difference theories

项目摘要

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.

项目描述（由申请人提供）：本项目应用快速时间分辨磁圆二色（TRMCD）、自然圆二色（TRCD）和普通吸收光谱技术研究血红素蛋白的功能和血红素蛋白和小肽的折叠。新的光学方法采用近零椭偏和偏振法来研究生物分子的快速动力学过程（纳秒到秒），这些生物分子含有磁光活性发色团，如血红素和芳香氨基酸，以及天然手性发色团，如蛋白质和肽的酰胺基团。这些技术将用于识别和研究血红素蛋白（如细胞色素c）折叠反应中的最早（亚毫秒）事件。一个主要目标是确定一个参数，这是理解蛋白质折叠本质的基础：不同的未展开构象相互转换的速度。如果这与折叠本身相比是缓慢的，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）。这样的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治疗方法，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组成血红蛋白分子的四个亚基如何相互合作以更有效地运输氧气。最近关于这种协同性的假设（Ackers对称规则），最初基于热力学测量，在本项目中通过动力学测量进行了测试。一个新的血红蛋白变构模型，从热力学和动力学的联系中出现，有望简化和系统化我们对血红蛋白动力学及其在体内由变构效应物（如有机磷酸盐）控制的理解。此外，TRMCD对位于协同作用关键位点的芳香氨基酸残基色氨酸[337]的研究，旨在进一步阐明血红蛋白亚基如何作为一个有效的“分子机器”共同工作，将氧气从肺部运输到组织。