Ribosome purification approaches for studying interactions of regulatory proteins and RNAs with the ribosome.
Ribosome purification approaches for studying interactions of regulatory proteins and RNAs with the ribosome.
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DOI：
10.1007/978-1-61779-949-5_18
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发表时间：
2012
期刊：
影响因子：
--
通讯作者：
Karzai AW
中科院分区：
文献类型：
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作者：
Mehta P;Woo P;Venkataraman K;Karzai AW
核糖体是RNA和蛋白质的大型复合物，在细胞中执行蛋白质合成的基本任务。核糖体也是几种与免疫相关功能的起始点。为了有效地执行这些任务，核糖体与无数的非核糖体蛋白和RNA相互作用。考虑到这些相互作用中的大多数是瞬时的，纯化与核糖体复合的相互作用因子可能是一项具有挑战性的工作。本文就核糖体的分离和核糖体相关因子的研究方法作一综述。我们还讨论了设计和执行核糖体缔合研究的关键参数。最后，我们提出了一个详细的协议报告为基础的富集测定，用于选择性地分离核糖体翻译特定的信息的兴趣。这些方案可用于研究广泛的核糖体相关功能。
Ribosomes are large complexes of RNA and protein that perform the essential task of protein synthesis in the cell. Ribosomes also serve as the initiation point for several translation-associated functions. To perform these tasks efficiently, ribosomes interact with a myriad of non-ribosomal proteins and RNAs. Given that most of these interactions are transient, purification of the interacting factors in complex with the ribosome can be a challenging undertaking. Here, we review methods commonly used to isolate ribosomes and study ribosome associated factors. We also discuss crucial parameters for designing and executing ribosome association studies. Finally, we present a detailed protocol for reporter based enrichment assays that are employed to selectively isolate ribosomes translating a particular message of interest. These protocols can be used to study a wide range of ribosome-associated functions.