THERMODYNAMIC AND KINETIC STUDIES OF PROTEIN STRUCTURE AND ENZYMIC MECHANISMS

这个实验室从事蛋白质结构和蛋白质结构的研究。 蛋白质折叠的机制 研究的主要对象是猪 胃蛋白酶原，分子量= 39，630的单体蛋白质， 在pH值6到8.5之间是稳定的 pH低于6时胃蛋白酶原会自行激活 通过蛋白水解损失其前44个氨基酸，以产生 酶活性蛋白，胃蛋白酶。 胃蛋白酶只有在pH值 小于6. 胃蛋白酶和胃蛋白酶原通过暴露于高pH而解折叠， 温度或变性剂如尿素的浓度。 然而，在这方面， 未折叠的胃蛋白酶原可以重新折叠成正常结构， 自然条件下，而胃蛋白酶则不能。 我对 这种重折叠反应的机制以及序列的差异 影响两种蛋白质的重折叠。 我们使用技术 例如紫外、圆二色性和荧光光谱， 与化学修饰和肽化学一起， 表征天然和未折叠物种的结构。 我们有 使用快速动力学技术，如停流和T跳，以检测 在折叠反应中部分折叠的形式;它们的结构已经被 部分确定和化学反应， 将它们与所研究的天然和未折叠形式分开。

This laboratory is engaged in studies on protein structure and the mechanism of protein folding. The main subject of research is swine pepsinogen, a monomeric protein of molecular weight= 39,630, which is stable at pH's between 6 and 8.5. Below pH 6 pepsinogen activates itself by proteolytic loss of its first 44 amino acids, to produce an enzymatically active protein, pepsin. Pepsin is stable only at pH's below 6. Pepsin and pepsinogen are unfolded by exposure to high pH, temperature or concentrations of denaturants, such as urea. However, unfolded pepsinogen can refold to its normal structure, when returned to native conditions, whereas pepsin cannot. I am interested in the mechanism of this refolding reaction and how the difference in sequence influences the refolding of the two proteins. We have used techniques such as ultra-violet, circular dichroic and fluorescence spectroscopies, together with chemical modification and peptide chemistry, to characterize the structures of the native and unfolded species. We have used rapid kinetic techniques, such as stopped-flow and T-jump, to detect partly folded forms in the folding reaction; their structures have been partially determined and the nature of the chemical reactions which separate them from the native and unfolded forms investigated.