MEASURING COLLAPSE KINETICS OF LAMBDA REPRESSOR

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子项目和研究者（PI）可能从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。所列机构为中心，不一定是研究者所在机构。蛋白质折叠通常会被自由能垒减缓，但蛋白质可以被设计得非常稳定，并且在没有明显垒的情况下快速折叠。这种超稳定蛋白质可以作为引入新功能的“支架”，并制造“设计蛋白质”。“我们正在研究这种不寻常的蛋白质，工程λ阻遏物的结构形成。虽然蛋白质折叠非常迅速，通过将其在粘性溶液中冷却下来，我们可以在阿贡的BioCAT光束线上通过时间分辨X射线散射跟踪紧凑结构的形成，这是水溶性蛋白质散射研究的最亮来源。我们已经进行了初步研究的蛋白质和我们的新的停止低装置，以优化信号，并继续我们的研究，在2005年与三种蛋白质变体，野生型，一个含有D14 A突变，第三个A37 G/A49 G突变。选择这些以涵盖一系列稳定性和聚集倾向。小蛋白质折叠的默认假设是X射线散射，荧光和红外光谱将产生相同的折叠时间（所谓的双态折叠）。我们相信，D14 A突变体将违反这一假设，而其他两个将符合，根据我们的设计。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Protein folding is usually slowed down by a free energy barrier, yet proteins can be designed to be very stable and to fold rapidly without a significant barrier. Such hyperstable proteins could serve as "scaffolds" for introducing new function and make "designer proteins." We are studying the formation of structure in one such unusual protein, engineered lambda repressor. Although the protein folds very rapidly, by cooling it down in viscous solutions, we can follow the formation of compact structure by time-resolved X-ray scattering at the BioCAT beamline at Argonne, the brightest source for aqueous protein scattering studies. We have conducted initial studies of the protein and our new stopped-low apparatus to optimize signals, and are continuing our studies in 2005 with three protein variants, the wildtype, one containing a D14A mutation, the third a A37G/A49G mutation. These were chosen to cover a range of stabilities and aggregation propensities. The default hypothesis for the folding of small proteins is that X-ray scattering, fluorescence and infrared spectroscopies will yield identical folding times (so-called two-state folding). We believe that the D14A mutant will violate this hypothesis, while the other two will conform, based on our design.