COMBINED SAXS AND THEORETICAL DETERMINATION OF THE STRUCTURAL ENSEMBLE

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 越来越清楚的是，蛋白质功能的调节通常是通过活性和非活性构象群的变化来实现的。X射线晶体分析只给出了溶液中填充的可能配置的快照。利用我们在蛋白质折叠和组装的理论和实验分析方面的专业知识，我们试图使用SAXS和理论相结合的方法来定义存在于多结构域蛋白质功能景观中的结构系综。这种方法超越了目前对SAXS数据的模拟分析，包括了在功能上很重要的局部和全局折叠/展开过渡。我们在开发这一方法学时最初选择的是CSK酶，因为它不仅是一个重要的治疗靶点，而且是一个我们根据最佳酶和折叠条件来表征的系统。越来越多的证据表明，结构集合中的偏见可能在CSK的调控中发挥重要作用。建立的方法学广泛适用于生物大分子，并将提供关于哪些构象系综与实验数据相一致的有用信息，以及生物学中普遍存在的动态可逆折叠/组装过程。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. It is becoming clear that regulation of protein function is often acheived via shifts in the populations of active and inactive conformers. X-ray crystallographic analysis gives only snapshots of possible configurations populated in solution. Taking advantage of our expertises in theoretical and experimental analysis of protein folding and assembly, we seek to use a combined SAXS and theoretical approach to define the structural ensembles present in the functional landscapes of multidomain proteins. This approach extends beyond current simulation analysis of SAXS data to include local and global folding/unfolding transitions that are important in function. Our initial choice in developing this methodology is the enzyme Csk as it is not only an important therapeutic target but also is a system we have characterized in terms of the optimal enzymatic and folding conditions. There is growing evidence that biases in the structural ensemble may play an important role in the regulation of Csk. The methodology developed here is broadly applicable to biological macromolecules and will provide useful information about what ensembles of conformations are consistent with the experimental data as well as the the ubiquitous dynamic reversible folding/assembly processes inherent in biology.