Role of Protein Dynamics in Modulating the Thermodynamic Linkages in Allostery

蛋白质动力学在调节变构热力学联系中的作用

• 批准号: 7367841
• 负责人: JAMES C LEE
• 金额: $ 28.59万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7367841
• 关键词: Address; Affect; Algorithms; Allosteric Regulation; Amino Acid Sequence; Be++ element; Beryllium; Binding; Binding Sites; Biological; Complement Factor B; Coupled; Cyclic AMP; Cyclic AMP Receptor Protein; Cyclic GMP; Cyclic Nucleotides; DNA Binding Domain; Data; Elements; Entropy; Family; Helix (Snails); Induced Mutation; Knowledge; Laboratory Research; Lead; Ligand Binding; Ligands; Link; Literature; Maps; Mass Spectrum Analysis; Measurement; Modeling; Motion; Mutagenesis; Mutation; Pattern; Peptide Sequence Determination; Play; Progress Reports; Property; Protein Dynamics; Proteins; Range; Research; Roentgen Rays; Role; Site; Solvents; Specificity; Structure; Sucrose; System; Testing; Thermodynamics; Time; Urea; base; ear helix; mutant; physical property; polypeptide; protein structure; prototype; success; transcription factor

cAMP receptor protein (CRP) is the prototype of a super-family of transcription factors. One key feature of CRP's biological activity that has been the focus of this laboratory's research is the fundamental rules that govern the allosteric activation by cAMP of its sequence-specific DMA binding. Recently we established, for the first time in any allosteric system, that the energetics of allosteric parameters are linear functions of protein dynamics. Using protein dynamics as the focus, there is a convergence among the results derived from crystallographic, computational and functional energetic data. This preliminary success enables us to focus our research effort on protein dynamics as the fundamental physical property to establish quantitative linkages between CRP structure and mechanism(s) of allostery. For coming years we will address these issues: 1. Is protein dynamics one of the fundamental properties of protein that modulate allostery? Two strategies will be employed to test the validity of this relationship: a) loops which are solvent accessible are targeted as structural elements to alter protein dynamics and allostery; b) Osmolytes, which affect dynamic motions of proteins, will be employed as solvent additives to perturb protein dynamics and allostery. 2. Is there a network of structural elements that is particularly affected by allosteric regulation? The structural elements the perturbation of which can lead to changes in protein dynamics and allostery will be identified by: a) changes in the thermal B-factors in X-ray data due to mutation; b) H/D exchange measurements coupled with mass spectrometry; and c) computation evidence for structural connectivity. 3. Can one use the knowledge gained in Aims 1 and 2 to assist in defining the structural elements that exert long range effects in defining specificity of the activating effectors? CRP is activated most efficiently by cAMP, although it can bind other cNMP without being activated. Literature results imply that polypeptide outside of the cAMP binding site, specifically the DNA binding domain, dictates the ability of CRP to respond to the bound ligand. Based on the preliminary computation results which show connectivity between the cAMP binding site and the DNA binding domain, mutants will be used to test the connectivity pattern. X-ray structural data of the cAMP binding domain will be used to test if the presence of the DNA binding domain alters the structural connectivity pattern in the cAMP binding domain.

CAMP受体蛋白（CRP）是转录因子超家族的原型。一个关键特征的一个 CRP一直是该实验室研究重点的生物学活动是基本规则 控制cAMP的变构激活其序列特异性DMA结合。最近我们建立了 任何变构系统中的第一次，变构参数的能量是线性函数 蛋白质动力学。将蛋白质动力学作为焦点，在得出的结果之间存在收敛性 来自晶体学，计算和功能能量数据。这种初步的成功使我们能够 将我们的研究工作集中在蛋白质动力学上，作为建立定量的基本物理特性 CRP结构与变构机理之间的联系。未来几年，我们将解决这些问题 问题：1。蛋白质动力学是调节蛋白质的基本特性之一吗？二 将采用策略来测试这种关系的有效性：a）可访问溶剂的循环是 作为改变蛋白质动力学和变构的结构元素； b）渗透压，会影响动态 蛋白质的运动将被用作溶剂添加剂来扰动蛋白质动力学和变构。 2。是否有一个特别受变构调节影响的结构元素网络？这 结构元素的扰动可能导致蛋白质动力学和变构的变化 识别为：a）由于突变引起的X射线数据中热B因子的变化； b）H/D交换 测量与质谱法结合； c）结构连通性的计算证据。 3。可以使用目标1和2中获得的知识来帮助定义结构元素 那在定义激活效应子的特异性方面发挥了远距离影响？ CRP最激活 通过cAMP有效，尽管它可以结合其他CNMP而不会被激活。文献结果暗示 多肽在营地结合位点外，特别是DNA结合结构域，决定了 CRP应对绑定的配体。基于显示连接性的初步计算结果 在营地结合位点和DNA结合域之间，将使用突变体来测试连通性 图案。 CAMP结合结构域的X射线结构数据将用于测试DNA的存在 结合域改变了cAMP结合域中的结构连接模式。