Characterization of New Detergents and Detergent Mimetics by Scattering Techniques for Membrane Protein Crystallization
Characterization of New Detergents and Detergent Mimetics by Scattering Techniques for Membrane Protein Crystallization
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DOI：
10.1007/978-1-4939-7151-0_9
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发表时间：
2017-01-01
期刊：
影响因子：
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通讯作者：
Loll, Patrick J.
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文献类型：
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作者：
Bonnete, Francoise;Loll, Patrick J.
膜蛋白一旦从其天然膜中除去就难以操纵和稳定。然而，尽管存在这些困难，由于化学和技术的进步，膜蛋白结构测定的成功持续了二十多年。这些进展中的许多都来自于专注于蛋白质工程、高通量表达和洗涤剂筛选的开发的努力，所有这些都是为了增强蛋白质的稳定性以用于生物化学和生物物理研究。相比之下，相当少的工作已经做了破译的基本机制，蛋白质洗涤剂复合物的结构，并描述洗涤剂的结构对稳定和结晶的影响。这些问题可以使用散射技术（采用光、X射线和/或中子）来解决，散射技术适合于描述溶液中大分子的结构和构象，以及评估颗粒之间的弱相互作用，这两者都与结晶密切相关。这些技术可以用于批量模式或耦合到尺寸排阻色谱，并提供了潜在的描述洗涤剂溶解的膜蛋白的构象和定量和模型洗涤剂结合到蛋白质，以优化晶体包装。我们将描述相关的技术和散射实验，它允许一个探索胶束之间和膜蛋白复合物之间的相互作用，并将这些相互作用与膜蛋白结晶的例子。
Membrane proteins are difficult to manipulate and stabilize once they have been removed from their native membranes. However, despite these difficulties, successes in membrane-protein structure determination have continued to accumulate for over two decades, thanks to advances in chemistry and technology. Many of these advances have resulted from efforts focused on protein engineering, high-throughput expression, and development of detergent screens, all with the aim of enhancing protein stability for biochemistry and biophysical studies. In contrast, considerably less work has been done to decipher the basic mechanisms that underlie the structure of protein-detergent complexes and to describe the influence of detergent structure on stabilization and crystallization. These questions can be addressed using scattering techniques (employing light, X-rays, and/or neutrons), which are suitable to describe the structure and conformation of macromolecules in solution, as well as to assess weak interactions between particles, both of which are clearly germane to crystallization. These techniques can be used either in batch modes or coupled to sizeexclusion chromatography, and offer the potential to describe the conformation of a detergent-solubilized membrane protein and to quantify and model detergent bound to the protein in order to optimize crystal packing. We will describe relevant techniques and present examples of scattering experiments, which allow one to explore interactions between micelles and between membrane protein complexes, and relate these interactions to membrane protein crystallization.