Studying structure and function of membrane proteins with PELDOR/DEER spectroscopy - The crystallographers' perspective.
Studying structure and function of membrane proteins with PELDOR/DEER spectroscopy - The crystallographers' perspective.
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利用 PELDOR/DEER 光谱研究膜蛋白的结构和功能 - 晶体学家的视角
DOI：
10.1016/j.ymeth.2018.03.002
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发表时间：
2018
期刊：
影响因子：
4.8
通讯作者：
Hagelueken G
中科院分区：
文献类型：
--
作者：
Glaenzer J;Peter MF;Hagelueken G
1985年，第一个膜蛋白的X射线结构被确定。三十多年后的今天，更多的结构已经被解决。然而，研究膜蛋白的结构仍然是一项非常具有挑战性的任务。由于其固有的构象灵活性，拥有单一的 X 射线结构通常只是真正理解这些动态分子功能的第一步。因此，需要额外的方法来提供补充信息，特别是关于构象灵活性的信息。脉冲电子-电子双共振光谱（PELDOR，也称为DEER）就是这样一种方法。它可用于精确测量大分子中固有的或人工引入的自旋中心之间的纳米距离分布，从而探测大分子的构象状态。 PELDOR 可应用于溶液、洗涤剂、脂质双层甚至细胞内。然而，PELDOR 是一种先进的光谱技术，需要专门的设备和培训。本章旨在成为希望更好地了解 PELDOR 光谱及其应用的晶体学家和其他结构生物学家的起点。它可以深入了解实验的规划阶段（即哪些旋转标签是可能的以及将它们放置在哪里）、如何进行 PELDOR 实验以及如何解释结果。为此，使用来自霍乱弧菌TRAP转运蛋白的底物结合蛋白（SBP）作为分步示例。此外，本章还举例说明了过去如何应用 PELDOR 光谱来克服现代综合结构生物学方法中 X 射线晶体学的已知局限性。
In 1985, the first X-ray structure of a membrane protein was determined. Today, more than 30 years later, many more structures have been solved. Nevertheless, studying the structure of membrane proteins remains a very challenging task. Due to their inherent conformational flexibility, having a single X-ray structure is usually only the first step towards truly understanding the function of these dynamic molecules. For this reason, additional methods are needed that can provide complementary information, especially about conformational flexibility. Pulsed electron-electron double resonance spectroscopy (PELDOR, also known as DEER) is such a method. It can be used to precisely measure nanometer distance distributions between intrinsic or artificially introduced spin-centers in macromolecules and thereby to probe the conformational state of the macromolecule. PELDOR can be applied in solution, in detergent, in lipid bilayers and even within cells. However, PELDOR is an advanced spectroscopy technique and requires specialised equipment and training. This chapter aims to be a starting point for crystallographers and other structural biologists who want to get a better understanding of PELDOR spectroscopy and its application. It gives an insight into the planning stages of the experiment (i.e., which spin labels are possible and where to place them), how a PELDOR experiment is conducted and how the results are interpreted. For this purpose, the substrate binding protein (SBP) from aVibrio choleraeTRAP transporter is used as a step-by-step example. Further, the chapter gives examples of how PELDOR spectroscopy has previously been applied to overcome known limitations of X-ray crystallography in modern integrative structural biology approaches.
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影响因子：
1
作者：
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