STUDIES ON NON-NATIVE STRUCTURES OF PROTEINS

1.利用小角X射线散射、圆二色谱和核磁共振波谱分析变性蛋白质的结构。这三种技术的结合揭示了枯草杆菌蛋白酶抑制剂（SSI）的冷变性状态、细胞色素c、肌红蛋白和溶菌酶的甲醇诱导变性状态的特征结构。在后者中，既观察到了紧密的变性状态（熔融球），也观察到了膨胀的螺旋状态，这取决于甲醇浓度和pH值。2.高压下蛋白质的结构和热力学稳定性。这使得天然结构的“变形”和压力变性的测量。3.蛋白质解折叠的温度-跳跃NMR研究了核糖核酸酶A的解折叠过程中的微波脉冲温度-跳跃NMR光谱在我们的实验室开发的。该方法使我们能够在蛋白质解折叠后的几百毫秒内检测到一个中间物种。4.二硫键在折叠过程中的作用研究了四个二硫键中的一个、两个、三个或全部被去除的基因工程溶菌酶的折叠。结果阐明了不同二硫键在折叠过程中的作用。

1.Structure of denatured proteins using small angle x ray scattering, circular dichroism, and NMR spectroscopy.The combination of the three techniques revealed the characteristic structure of the cold denatured state of Streptomyces subtilisin inhibitor (SSI), methanol-induced denatured states of cytochrome c, myoglobin, and lysozyme. In the latter, the compact denatured states (molten globule) and the expanded helical states were both observed, depending on the methanol concentration and pH.2.Structure and thermodynamic stability of proteins at high pressure.A versatile high pressure NMR technique was developed for use with a high field NMR spectrometer. This allowed measurement of "deformation" of the native structure and denaturation by pressure.3.Protein unfolding by temperature-jump NMR.The process of unfolding of ribonuclease A was studied with the microwave-pulsed temperature-jump NMR spectroscopy developed in our laboratory. The method allowed us to detect an intermediate species within a few hundred milliseconds upon unfolding of the protein.4.The role of disulfide bridges on the folding process.An genetically engineered lysozymes in which one, two, three or all of the four disulfide bridges were removed were investigated for their folding. The results clarified the roles of different disulfide bridges in the folding process.