Early events of folding of β-structure-rich protein

蛋白质通过一定的途径折叠。折叠研究的目的是寻找其折叠途径。我们研制了以圆二色、x射线散射和荧光为探针的冷冻止流仪。我们研究了各种类型的蛋白质。对α -螺旋蛋白的研究结果表明，α -螺旋形成的折叠速率非常快，我们无法测量α -螺旋形成的速率。相反，以β结构为主的蛋白质，在其形成的最初阶段就采取了富含α -螺旋的结构。我们关注了SH3的折叠，它在其天然状态下采用β构象。SH3在折叠的最初阶段采用富含α -螺旋的中间产物。然后我们研究了其他富含β结构的蛋白质，发现中间产物无一例外地出现。将脉冲振幅与其他参数进行了比较，发现脉冲振幅与螺旋分数有很好的关系，该关系由程序Helix2计算。该程序主要基于螺旋线圈跃迁理论，并结合相邻残基的实验数据。这表明最初的折叠核是由短寿命的α螺旋相互作用形成的。通过SH3 （Alanine 45 to Glycine）的一个残基突变，我们发现在pH3下构象从富含β变为富含α。这表明从β到α的转换比预期发生得更频繁。

Protein folds via definite pathway. The purpose of the folding study is to find its folding pathway. We have developed cryo-stopped-flow apparatus with circular dichrosism, x-ray scattering and fluoresecence as probes. We have investigated various types of proteins. Results with alpha-helical protein show the folding rate of alpha-helix formation is so rapid that we cannot measure the rate of alpha-helix formation. In contrast, proteins, mainly consisting of beta-structure, take alpha-helix-rich intrmeditate at its earliest stage. We have focused the folding of SH3, which takes beta-conformation at its native state.SH3 takes alpha-helix-rich intermediate at its earliest stage of folding. We then investigated other beta-structure-rich proteins, and found the intermediate appeared without exception. We compared the amplitude of the burst with other parameters, and found the burst amplitude has good relationship with the helical fraction calculated by a program, Helix2. The program is mainly based on helix-coil transition theory with experimental data of adjacent residues. This suggest that the initial folding cores are formed by the interaction of short-lived alpha-helices.By the one residue mutation of SH3 (Alanine 45 to Glycine), we found the conformation changed from beta-rich to alpha-rich at pH3. This indicates the conversion of beta to alpha occurs more frequently than expected.