Combining Multi-scale Modeling with Multi-pronged Experiments to Unveil Conformational Changes of Macromolecular Complexes

大分子复合物的结构表征及其功能状态是分子生物学中的一项重要任务，因为多组分分子机器完成许多重要的细胞功能。由于这些大的复合物很难通过X射线晶体学研究，替代的低分辨率实验技术，如cryo-EM，SAXS和FRET通常用于表征其构象状态。这些实验方法中的每一种都提供了不同的优势，并遇到了不同的陷阱，工件和限制。因此，如果把所有的实验数据结合起来注释构象状态，可以得到更准确的描述。将开发计算方法，通过整合来自各种低分辨率实验和X射线晶体学的多个数据，获得大分子复合物中发生的构象转变的原子水平描述。将通过使用分子动力学模拟和/或正常模式分析以物理上有意义的方式使已知的X射线结构变形以与cryo-EM、SAXS和/或FRET数据一致来构建结构模型。已经使用多组实验数据进行了广泛表征的核糖体将用作测试系统，以使用真实的实验数据来验证该方法。然后，这些方法将被应用到研究的一个小的植物病毒，番茄丛矮化病毒的肿胀转变，以获得更好的理解，通过这个模型系统，病毒的生活周期。这个项目将涉及本科生和研究生。学生将学习生物物理学，计算方法和结构生物学的理论，通过促进方法的开发，测试和后续应用。新开发的工具将以程序包的形式分发给科学界，可通过互联网免费下载。这些新工具的引人注目的方法是，不仅研究小组可以用这些算法分析他们自己的实验数据，而且他们还可以将其他已发表的结果纳入建模，以获得更准确的构象转变描述。首席研究员还参与了对当地图森高中女生的外联活动。首席研究员将与高中教师合作制定一项推广计划，在夏季为高中女生提供大学研究机会，鼓励她们追求科学事业。

Structural characterization of macromolecular complexes and their functional states is a crucial task in molecular biology, since multi-component molecular machines accomplish many important cellular functions. As these large complexes are difficult to study by X-ray crystallography, alternative low-resolution experimental techniques such as cryo-EM, SAXS and FRET are often used to characterize their conformational states. Each of these experimental approaches offers different advantages and meets with different pitfalls, artifacts and limitations. Therefore more accurate descriptions could be obtained if all pieces of experimental data were taken together to annotate conformational states. Computational methods will be developed to obtain atomic level descriptions of conformational transitions occurring in macromolecular complexes by integrating multiple data from a variety of low-resolution experiments and X-ray crystallography. Structural models will be constructed by deforming, in a physical meaningful way using molecular dynamics simulation and/or normal mode analysis, a known X-ray structure to agree with cryo-EM, SAXS and/or FRET data. The ribosome, which has been extensively characterized using multiple sets of experimental data, will serve as a test system to validate the approach with real experimental data. These methods will then be applied to study the swelling transition of a small plant virus, Tomato Bushy Stunt Virus to get a better understanding, through this model system, of the life cycle of viruses. This project will involve undergraduate and graduate students. Students will learn theories of biophysics, computational approaches and structural biology by contributing to the development, testing and subsequent applications of the methods. The newly developed tools will be distributed to the scientific community in the form of a program package that will be freely available through the Internet for download. The compelling approach with these new tools is that not only research groups could analyze their own experimental data with those algorithms, but also they could incorporate other published results into the modeling to obtain a more accurate description of the conformational transitions. The principal investigator is also involved in outreach activities to local Tucson high-school girls. The principal investigator will develop in concert with high-schools teachers an outreach program to offer research opportunities at the University to high-school girls during the summers to encourage them to pursue scientific careers.