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Hydration Effects on the Structure and Thermodynamics of Proteins in Nonnative States

水合对非天然状态蛋白质结构和热力学的影响

基本信息

批准号：
06680646
负责人：
SODA Kunitsugu
金额：
$ 1.41万
依托单位：
Nagaoka University of Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1995
项目状态：
已结题

项目摘要

A quantitative measure of the structural stability of proteins is the difference in free energy DELTAG_d between their native (N) and denatured (D) states. Hence, to identify and estimate the factors contributing to DELTAG_d, it is indispensable to obtain structural information not only on the N state but also on the D atate. It has been established from studies using various structural probes that there exists some intermediate states such as the molten globule (MG) state between the N and the completely unfolded (U) states, and that the D-state structure is very diverse. Complementary to spectroscopic methods, the solution X-ray scattering (SXS) method is a powerful method for studies on the nonnative state which is essentially the multiconformational state, but it cannot determine protein structure by itself. In thisstudy, we developed methods to supplement the above weakness of SXS method, and analyzed the structures of proteins in the nonnative states :(1) Taking account of the hy … More dration of polar groups and the probability distribution of internal rotation angles of peptide chains, we devised a method for generating various conformations in computer.(2) Extending the above, we developed a method to generate conformations for the multipartite (MP) structure model of the nonnative state, where a polypeptide chain is composed of a segment having the structure identical to that in the N state and segments having totally unfolded structure.(3) We devised a new algorithm for the SXS simulation, where the X-ray scattering profile is evaluated by explicitly considering the contribution from solvent under the continuum approximation of solvent.(4) Applying the above method to several natural proteins and unfolded chains, we confirmed its performance and effectiveness, and showed the importance of accurately considering solvent influences.(5) Applying the MP model to three proteins in the MG state, we compared our prediction with reported data and have confirmed that their SXS profiles, especially that of cytochrome c, can be well explained by this model. However, slight difference has been found for alpha-lactalbumin and myoglobin, indicating necessity to refine the model for them. Less

蛋白质结构稳定性的定量测量是其天然(N)和变性(D)状态之间的自由能DELTAG_d的差异。因此，要识别和估计影响DELTAG_d的因素，不仅要获得N态的结构信息，还要获得D态的结构信息。各种结构探针的研究表明，在N态和完全展开的(U)态之间存在熔融球态（MG）等中间态，d态结构非常多样。溶液x射线散射（SXS）方法是光谱方法的补充，是研究非天然态的有力方法，非天然态本质上是多构象态，但它本身不能确定蛋白质的结构。在本研究中，我们开发了一种方法来弥补SXS方法的上述缺点，并分析了蛋白质在非天然状态下的结构：(1)考虑到极性基团的多水化和肽链内旋角的概率分布，我们设计了一种在计算机中生成各种构象的方法。(2)在此基础上，我们开发了一种生成非天然状态多肽多部（MP）结构模型构象的方法，其中多肽链由具有与N态相同结构的片段和具有完全展开结构的片段组成。(3)我们设计了一种新的SXS模拟算法，在连续介质近似下明确考虑溶剂的贡献来评估x射线散射剖面。(4)将上述方法应用于几种天然蛋白质和未折叠链，证实了其性能和有效性，并表明了准确考虑溶剂影响的重要性。(5)将MP模型应用于三种处于MG状态的蛋白，将我们的预测与文献数据进行了比较，证实了它们的SXS谱，特别是细胞色素c的SXS谱可以很好地解释该模型。然而，在α -乳清蛋白和肌红蛋白中发现了细微的差异，这表明有必要为它们改进模型。少