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，是一种相对分子质量为39,630的单体蛋白质， 在pH值在6到8.5之间稳定。在pH 6以下，胃蛋白酶原自我激活 通过蛋白质分解失去它的前44个氨基酸，产生一个 具有酶活性的蛋白质，胃酶。胃酶只有在pH低于以下时才稳定 6.胃酶和胃蛋白酶原通过暴露在高pH、高温度下而展开 或浓度的变性剂，如尿素。然而，展开了 当返回天然状态时，胃蛋白酶原可以重新折叠到其正常结构 条件，而胃酶不能。我对它的机制很感兴趣 这种折叠反应以及序列上的差异如何影响 两种蛋白质的重折叠。我们使用了快速动力学技术， (停流和T跳跃)，以检测部分折叠的中间形式 折叠反应；它们的结构已部分确定，并 将它们与原生生物分开的化学反应的性质 并展开调查表格。紫外线等技术 吸光度、圆二色光谱和荧光光谱 用化学修饰和多肽化学来表征 这些蛋白质和其他蛋白质的天然和未折叠形式的结构， 包括色氨酸合成酶、人核甲状腺激素受体和 硫酸盐转移酶。

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 it's 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 rapid kinetic techniques, (stopped-flow and T-jump), to detect partly folded intermediate 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. Techniques such as ultra-violet absorbance, circular dichroic and fluorescence spectroscopies, together with chemical modification and peptide chemistry, to characterize the structures of native and unfolded forms of these and other proteins, including tryptophan synthase, human nuclear thyroid hormone receptor and sulphotransferase enzymes.