Zettl Thomas, Das Rhiju, Harbury Pehr A B, Herschlag Daniel, Lipfert Jan, Mathew Rebecca S, Shi Xuesong
Department of Physics, Nanosystems Initiative Munich, and Center for Nanoscience, LMU Munich, Munich, Germany.
Department of Biochemistry, Stanford University, Stanford, California.
Curr Protoc Nucleic Acid Chem. 2018 Jun;73(1):e54. doi: 10.1002/cpnc.54. Epub 2018 Jun 7.
Most structural techniques provide averaged information or information about a single predominant conformational state. However, biological macromolecules typically function through series of conformations. Therefore, a complete understanding of macromolecular structures requires knowledge of the ensembles that represent probabilities on a conformational free energy landscape. Here we describe an emerging approach, X-ray scattering interferometry (XSI), a method that provides instantaneous distance distributions for molecules in solution. XSI uses gold nanocrystal labels site-specifically attached to a macromolecule and measures the scattering interference from pairs of heavy metal labels. The recorded signal can directly be transformed into a distance distribution between the two probes. We describe the underlying concepts, present a detailed protocol for preparing samples and recording XSI data, and provide a custom-written graphical user interface to facilitate XSI data analysis. © 2018 by John Wiley & Sons, Inc.
大多数结构技术提供的是平均信息或关于单一主要构象状态的信息。然而，生物大分子通常通过一系列构象发挥功能。因此，要全面了解大分子结构，就需要了解在构象自由能景观上表示概率的系综。在此，我们描述一种新兴方法——X射线散射干涉测量法（XSI），这是一种能为溶液中的分子提供瞬时距离分布的方法。XSI使用位点特异性附着于大分子的金纳米晶体标记，并测量重金属标记对之间的散射干涉。记录的信号可直接转换为两个探针之间的距离分布。我们阐述了其基本概念，给出了制备样品和记录XSI数据的详细方案，并提供了一个定制的图形用户界面以方便XSI数据分析。© 2018约翰威立国际出版公司
