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

通过溶液X射线散射详细研究了各种蛋白质熔融球球（MG）的溶液结构。 MG的结构可以分为两类：一个接近天然结构，另一种是由疏水核心和凸起的尾巴组成的。某些MG主要通过疏水相互作用稳定，另一个MG主要通过分子内SS键稳定。变性状态不能通过一个术语，随机线圈概括，因为还指示了变性状态的结构多样性和多样性。β-乳球蛋白折叠的运动研究证明了具有非本性二级结构中间体的准确性，这表明非层次模型的重要性是非等级模型的折叠模型。等温滴定对Mg形成的cal含量测量表明，Mg中存在的疏水相互作用几乎是天然状态下的40％。它表明，变性剂诱导的Tryp的α-亚基的变性……更多的tophane合成酶是3个状态，但是其热变性为2个状态2个状态。对脯氨酸突变体的动力学研究表明，在色氨酸合酶的α-亚基折叠过程中存在两个成功的中间体。脯氨酸残留物有助于晚期中间体的稳定性。伴侣蛋白的靶标识别机制已使用α-乳乳蛋白的MG进行了研究，以揭示电静电相互作用的重要性。对MG的全球结构进行了理论研究的重要性。提出了对各种蛋白质的MG模型，并将计算出的散射曲线与观察到的散射曲线进行了比较。因此，提出的理论方法被证明是折叠研究的很有希望的。开发了蛋白质溶液的高压NMR方法，并将其应用于RNase A的热变性A. IT显示RNase A即使在高压下也会经历两态过渡。通过变性的体积变化为负，并且通过加压力会显着降低恒定压力下的热容量。精确测量变性时的绝热可压缩性。较少的