SAXS-DETECTED DYNAMICS OF FAST-FOLDING PROTEINS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 这一进行中的项目是对先前BioCAT项目的后续行动，该项目通过结合小角X射线散射和圆二色谱来研究lambda抑制子的快速折叠动力学(Dumont等人，蛋白质科学15,2596-2604(2006))。我们正在研究lambda抑制子和fyn SH3的聚集，以获得分子间平均作用力的势，从而获得临界聚集核的大小。临界核是衡量蛋白质聚集的重要指标，但很难确定。我们还用停流小角X射线散射法研究了一个Lambda抑制因子的下坡折叠突变体的折叠动力学。我们的目标是验证一种假设，即疏水相互作用的丧失最终会导致折叠率的“逆转”。我们最近有了这个项目的第一次光束时间，下一次计划时间在2007年2月。我们已经完成了对蛋白质的辐射损伤测试，聚集研究所需的所有浓度研究(目前处于为准备发表而对数据进行建模的阶段)，以及下坡lambda阻遏突变体的一些动力学测量。接下来将对这两种蛋白质进行更多的动力学，包括在不同最终变性剂浓度下的测量，以提供旋转半径的温度依赖和时间依赖的完整图。

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. This ongoing project follows up on a previous BioCAT project to study fast folding dynamics of lambda repressor by a combination of small angle X-ray scattering and circular dichroism spectroscopy (Dumont et al., Protein Science 15, 2596-2604 (2006)). We are examining the aggregation of lambda repressor and of fyn SH3 to obtain the potential of mean force between molecules and hence the size of the critical aggregation nucleus. The critical nucleus is an important measure of protein aggregation, but difficult to determine. We are also studying the folding dynamics of a downhill-folding mutant of lambda repressor by stopped-flow small angle X-ray scattering. The goal is to test the hypothesis that loss of the hydrophobic interaction eventually leads to a "turn-around" in the folding rate. We recently had our first beam time for this project, with the next planned time coming up in February 2007. We have completed radiation damage test on the proteins, all the concentration studies needed for the aggregation study (which is now in a phase of modeling the data to prepare for publication), and some of the kinetics measurements for the downhill lambda repressor mutants. More kinetics, including measurements at different final denaturant concentrations will be taken for both proteins next, to provide a complete map of the temperature dependence and time dependence of the radius of gyration.