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Structures and Formation Mechanisms of Folding Intermediates of Proteins

蛋白质折叠中间体的结构和形成机制

基本信息

批准号：
06304051
负责人：
KATAOKA Mikio
金额：
$ 6.34万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Co-operative Research (A)
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

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06304051/
关键词：
protein folding molten globule X-ray solution scattering protein denturation high pressure NMR stopped-flow CD chaperonine gene engineering 蛋白質変性変性静電的相互作用疎水的相互作用蛋白質非天然構造

项目摘要

Solution structures of molten globules (MG) of various proteins were investigated in detail by solution X-ray scattering. The structure of MG can be classified into two categories : one is close to native structure and the other is composed of a hydrophobic core and flaring tail (s) . Some MG's are stabilized mainly by hydrophobic interaction, the other MG's are stabilized mainly by intramolecular SS bonds. The denatured states cannot be generalized by a term, random coil, because the structural diversity and variety in the denatured states were also indicated.Kinetic studies of beta-lactoglobulin folding demonstrated the accumulation of intermediate with non-native secondary structure, which suggests the importance of non-hierarchical model for folding. Isothermal titration calorimetric measurements on MG formation indicated that the hydrophobic interaction existed in MG is almost 40% of that in native state.It was revealed that denaturant-induced denaturation of alpha-subunit of tryp … More tophane synthase is 3 states, however its thermal denaturation is 2 states. Kinetic studies on proline mutants indicated the existence of two successive intermediates in the folding process of alpha-subunit of tryptophane synthase. Proline residues are contributed to the stability of the late intermediate.The mechanism of target recognition by chaperonine have been investigated using MG of alpha-lactalbumin to reveal the importance of electro-static interaction.Theoretical studies were performed on the global structures of MG and the determinant, especially the contribution of water to the MG formation. Models of MG were proposed to various proteins and the calculated scattering profiles were compared with the observed ones. Consequently the proposed theoretical method was turned out to be quite promising for the folding studies.High pressure NMR method for protein solution was developed and applied to thermal denaturation of RNase A.It was revealed that RNase A undergoes two-state transition even under high pressure. A volume change by denaturation was negative and also a heat capacity at constant pressure was decreased significantly by addition of pressure. Adiabatic compressibility upon denaturation was measured precisely. Less

用溶液X射线散射法研究了不同蛋白质熔融球的溶液结构。MG的结构可以分为两类：一类是接近天然结构，另一类是由疏水核心和张开的尾部组成。一些MG主要通过疏水相互作用稳定，其它MG主要通过分子内SS键稳定。β-乳球蛋白的折叠动力学研究表明，β-乳球蛋白的折叠过程中存在着非天然二级结构的中间产物的积累，这表明了非分级折叠模式的重要性。对MG形成的等温滴定量热分析表明，MG中存在的疏水相互作用约为天然状态的40%，表明变性剂诱导的色氨酸α亚基的变性 ...更多信息 tophane合酶是三态的，而它的热变性是两态的。对脯氨酸突变体的动力学研究表明，在色氨酸合酶α亚基的折叠过程中存在两个连续的中间体。本文以α-乳清蛋白的微球蛋白为研究对象，探讨了分子伴侣作用下的靶点识别机制，揭示了静电相互作用的重要性，并对微球蛋白及其决定簇的整体结构进行了理论研究，特别是水对微球蛋白形成的贡献。模型的MG提出了各种蛋白质和计算的散射剖面与观察到的。建立了蛋白质溶液的高压核磁共振方法，并将其应用于RNase A的热变性研究，发现RNase A在高压下也会发生两态转变。变性引起的体积变化是负的，并且恒压下的热容也通过加压而显著降低。精确测量了变性后的绝热压缩率。少