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Thermodynamic Analysis of Protein Stability

蛋白质稳定性的热力学分析

基本信息

批准号：
09044222
负责人：
YUTANI Katsuhide
金额：
$ 2.18万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1999
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09044222/
关键词：
stability of proteins lysozyme DSC hydrogen bond hydrophobic interaction X-ray analysis prediction of tertiary structure stabilization factors of proteins

项目摘要

Understanding how tertiary structures of proteins are coded for in their amino acid sequences is one of the main targets in modern life science research. However, this goal has not yet been achieved. One of the reasons is that a protein structure is prescribed not only by a part of the amino acid sequence but also by an extensive interaction. In fact, the same kinds of substitutions give different changes in conformational stability. These results show that the effect of a single amino acid substitution on protein stability depends on the location of the mutation site and its environment in the protein structure. In order to estimate the contribution of each factor to protein stability, the data of structure and stability changes must be expanded by studying many mutant proteins with systematic and comprehensive substitutions. In this project, we tried to estimate some factors that contribute to conformational stability of a protein using data base of stability/structure, which are obtained from a series of mutant human lysozymes. Then, we constructed more than 100 mutant human lysozymes. Changes in stabilities and structures due to mutations were evaluated by DSC and X-ray analysis, respectively. Using database of structure and stability changes due to mutations, which are obtained in this project, the magnitude of hydrophobicity of carbon atom (C), and the hydrophobicity of nitrogen and neutral oxygen atoms (N/O) could be estimated to be 0.178 kJ/mol/ÅィイD12ィエD1, and -0.013 kJ/mol/ÅィイD12ィエD1, respectively. The net contribution of an intramolecular hydrogen bond, an intermolecular one between protein and ordered water molecules, and an intermolecular one between ordered water molecules was estimated to be 8.5, 5.2, and 5.0 kJ/mol, respectively, for a hydrogen bond with 3 Å length. The entropic cost due to the introduction of a water molecule could be also estimated to be about 8 kJ/mol.

了解蛋白质的三级结构是如何在其氨基酸序列中编码的是现代生命科学研究的主要目标之一。然而，这一目标尚未实现。其中一个原因是，蛋白质的结构不仅由部分氨基酸序列决定，而且由广泛的相互作用决定。事实上，同种取代对构象稳定性的影响是不同的。这些结果表明，单一氨基酸取代对蛋白质稳定性的影响取决于突变位置及其在蛋白质结构中的环境。为了估计每个因素对蛋白质稳定性的贡献，必须通过研究许多具有系统和全面替代的突变蛋白质来扩大结构和稳定性变化的数据。在这个项目中，我们试图利用从一系列突变的人溶菌酶中获得的稳定性/结构数据库来估计影响蛋白质构象稳定性的一些因素。然后，我们构建了100多个突变型人溶菌酶。分别用差示扫描量热法(DSC)和X射线分析(X-ray)评价突变后的稳定性变化和结构变化。利用本项目获得的突变引起的结构和稳定性变化的数据库，碳原子的疏水性大小(C)和氮原子和中性氧原子的疏水性(N/O)分别为0.178 kJ/ÅィイD12ィエd1和-0.013 kJ/mo1/ÅィイD12ィエd1。对于3Å长度的氢键，分子内氢键、蛋白质与有序水分子间的氢键和有序水分子之间的分子间氢键的净贡献率分别为8.5kJ/Å、5.2kJ/m ol和5.0kJ/m ol。由于水分子的引入，其熵成本也可以估计为约8kJ/mol。