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首页> 外文期刊>Current Protocols in Cell Biology >In vitro Assays for Targeting and Insertionof Tail-Anchored Proteins Into the ERMembrane
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In vitro Assays for Targeting and Insertionof Tail-Anchored Proteins Into the ERMembrane

机译：用于靶向和插入尾锚蛋白的体外测定进入Ermembrane

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摘要

Membrane proteins mediate numerous essential cellular functions. Due to theaggregation propensity of hydrophobic transmembrane domains in aqueous environments,the targeting and insertion of membrane proteins pose major challengesto cells. In the Guided Entry of Tail-anchored protein (GET) pathway,an essential class of newly synthesized tail-anchored proteins (TAs) are chaperonedand guided by multiple targeting factors to the endoplasmic reticulum(ER) membrane. Deciphering the molecular mechanism of this cellular processhas benefitted from successful in vitro reconstitution of individual molecularevents in the GET pathway with purified components. Here we describe recentlydeveloped protocols for in vitro reconstitution of functional complexesof TA substrates with their targeting factors, for monitoring the transfer ofTAs between targeting factors, and for the insertion of TA into the microsomalmembrane. These procedures are generally applicable to the interrogation ofother post-translational membrane protein targeting pathways.
机译：膜蛋白介导众多必需的细胞功能。由于疏水性跨膜结构域在含水环境中的术语，膜蛋白姿势姿势主要挑战细胞的靶向和插入。在尾锚式蛋白（Get）途径的引导过程中，新合成的尾锚蛋白（TAS）是伴侣，由多个靶向因子引导到内质网（ER）膜。解入这种细胞过程的分子机制受益于在Get途径中的单个分子中的成功体外重建，具有纯化的组分。在这里，我们描述了最近的开发方案，用于在其靶向因子进行TA底物的体外重建功能复合物的体外重建，用于监测靶向因子之间的转移，以及插入微瘤之间的转移。这些程序通常适用于其他翻译后膜蛋白靶向途径的询问。

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