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Quantifying the contribution of a new type of H-bond to protein stability

量化新型氢键对蛋白质稳定性的贡献

基本信息

批准号：
1716864
负责人：
Floyd Romesberg
金额：
$ 31.5万
依托单位：
The Scripps Research Institute
依托单位国家：
美国
项目类别：
Standard Grant
财政年份：
2017
资助国家：
美国
起止时间：
2017-08-01 至 2019-07-31
项目状态：
已结题

来源：
https://www.nsf.gov/awardsearch/showAward?AWD_ID=1716864&HistoricalAwards=false
关键词：
Quantifying contribution new type bond

项目摘要

Proteins are linear chains of amino acids that fold into complex three-dimensional structures by virtue of a variety of intramolecular interactions involving both their main chain (i.e. backbone) atoms, as well as the different, individual amino acid side chain atoms. One very well characterized interaction in proteins that provides structural stability is the hydrogen bond (H-bond), which is a type of electrostatic interaction formed between protein backbone atoms that have opposite polarity and positioned in a structurally accessible manner. This project investigates the role of an additional and previously underappreciated type of stabilizing interaction in proteins, H-bonds between backbone atoms of sequentially adjacent amino acids. This project will use an experimental approach that relies on the use of carbon-deuterium (C-D) bonds to visualize specific protein vibrations to investigate energetic contribution of this type of H-bond in protein stability. The interactions that define a protein's folding, structure, and stability are all important factors in activity and function, thus research to identify and quantify novel types of interactions have the potential to impact basic research into all aspects of protein science. This project will provide education and training for graduate, undergraduate and high school students by involving them in every aspect of the project. H-bonds formed between backbone N-H donor and CO acceptors have long been appreciated as dominant forces underlying protein structure and stability. However, many residues within proteins adopt conformations with phi and psi values of approximately ±90° and 0°, respectively, which are traditionally considered forbidden due to a steric clash between the amide nitrogen (Ni) and that of the following amino acid (Ni+1). This project will investigate the (Ni+1)-H---(Ni) interactions formed within these conformations which constitute a previously underappreciated type of H-bond. The objectives of this project is to focus specifically on determining the stabilization mediated by the (Ni+1)-H---(Ni) H-bonds in R67 DHFR. This project will compare the stability of the protein with its (Ni+)-H---(Ni) H-bonds intact with that of an isosteric CH2 analog [wherein one or more of th(Ni+1)-H---(Ni) H-bonds are disrupted]. In addition, model systems will be used to generate a calibration curve that will enable an interpretation of the shifts in IR spectra observed with the protein in terms of stabilization. Successful completion of the objectives will provide a convincing demonstration that (Ni+1-H)---(Ni) H-bonds are formed, and importantly, will provide a gauge of their contribution to protein stability.

蛋白质是一种线性氨基酸链，通过主链(即主链)原子和不同的氨基酸侧链原子之间的分子内相互作用，折叠成复杂的三维结构。蛋白质中一种非常典型的提供结构稳定性的相互作用是氢键(H键)，这是一种在蛋白质主干原子之间形成的静电相互作用，这些原子具有相反的极性，并以结构可及的方式定位。这个项目调查了蛋白质中一种额外的、以前被低估的稳定相互作用的作用，即顺序相邻氨基酸的主干原子之间的氢键。这个项目将使用一种实验方法，依赖于使用碳-氚(C-D)键来可视化特定的蛋白质振动，以研究这种类型的氢键对蛋白质稳定性的能量贡献。决定蛋白质折叠、结构和稳定性的相互作用都是活性和功能的重要因素，因此，识别和量化新型相互作用的研究有可能影响蛋白质科学各个方面的基础研究。该项目将为研究生、本科生和高中生提供教育和培训，让他们参与到项目的各个方面。长期以来，主链N-H供体和CO受体之间形成的氢键一直被认为是蛋白质结构和稳定性的主导力量。然而，蛋白质中的许多残基采用phi和psi分别约为±90°和0°的构象，这传统上被认为是禁止的，因为酰胺氮(Ni)和随后的氨基酸(Ni+1)之间存在空间碰撞。这个项目将研究在这些构象中形成的(Ni+1)-H-(Ni)相互作用，它构成了一种以前被低估的氢键类型。本项目的目标是明确R67 DHFR中(Ni+1)-H-(Ni)H-键所介导的稳定性。该项目将比较具有完整的(Ni+)-H-(Ni)H-键的蛋白质的稳定性与等量构型CH2类似物的稳定性[其中一个或多个(Ni+1)-H-(Ni)H-键被破坏]。此外，模型系统将被用来生成校准曲线，该曲线将能够从稳定性的角度解释观察到的蛋白质红外光谱的变化。这些目标的成功完成将提供令人信服的证据，证明(Ni+1-H)-(Ni)H-键已经形成，重要的是，将提供它们对蛋白质稳定性的贡献的衡量标准。