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Methods of reconstitution to investigate membrane protein function
Methods ( IF 4.8 ) Pub Date : 2018-09-01 , DOI: 10.1016/j.ymeth.2018.02.012
Ruth Skrzypek , Shagufta Iqbal , Richard Callaghan

Membrane proteins are notoriously difficult to investigate in isolation. The focus of this chapter is the key step following extraction and purification of membrane proteins; namely reconstitution. The process of reconstitution re-inserts proteins into a lipid bilayer that partly resembles their native environment. This native environment is vital to the stability of membrane proteins, ensuring that they undergo vital conformational transitions and maintain optimal interaction with their substrates. Reconstitution may take many forms and these have been classified into two broad categories. Symmetric systems enable unfettered access to both sides of a bilayer. Compartment containing systems contain a lumen and are ideally suited to measurement of transport processes. The investigator is encouraged to ascertain what aspects of protein function will be undertaken and to apply the most advantageous reconstitution system or systems. It is important to note that the process of reconstitution is not subject to defined protocols and requires empirical optimisation to specific targets.

中文翻译:

研究膜蛋白功能的重建方法

众所周知,膜蛋白很难单独研究。本章的重点是膜蛋白提取纯化后的关键步骤;即重组。重建过程将蛋白质重新插入脂质双层中,该脂质双层与它们的天然环境部分相似。这种天然环境对膜蛋白的稳定性至关重要,可确保它们经历重要的构象转变并保持与其底物的最佳相互作用。重构可以采取多种形式,这些形式被分为两大类。对称系统可以不受限制地访问双层的两侧。隔间系统包含一个内腔,非常适合测量运输过程。鼓励研究人员确定将进行蛋白质功能的哪些方面并应用最有利的重建系统或系统。需要注意的是,重构过程不受定义的协议的约束,需要针对特定​​目标进行经验优化。
更新日期:2018-09-01
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