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[蛋白质互补:用于研究活细胞中蛋白质-蛋白质相互作用的工具]

[Protein complementation as tool for studying protein-protein interactions in living cells].

作者信息

Chumakov S P, Kravchenko Iu E, Chumakov P M

出版信息

Mol Biol (Mosk). 2012 Sep-Oct;46(5):699-711.

Abstract

Association and degradation of protein complexes play essential role in a majority of normal and pathologic processes, which take place in living cell. Studying the underlying mechanisms of those interactions would give deeper understanding of specific causes of disease progression and would allow developing new therapeutic strategies. The majority of technical approaches currently used for detecting protein association include in vitro protein extraction and purification, whereas more relevant results require methods that can be used in vivo. One of a few approaches for in vivo protein association detection is based on reporter protein fragment complementation. Reporter systems based on protein complementation rely on reconstitution of reporter protein fluorescent or enzymatic activity which occurs upon reassociation of protein fragments and could be measured by colorimetry, luminometry or fluorimetry. Protein complementation is widely used to develop reporter systems for analysis of protein interactions, for functional dissection of signal transduction pathways and for performing high-throughput screenings to discover new protein interaction partners. Currently developed approaches that utilize protein fragment complementation have possibilities that extend far beyond simple detection of interaction in a pair of proteins.

摘要

蛋白质复合物的缔合与降解在发生于活细胞中的大多数正常和病理过程中起着至关重要的作用。研究这些相互作用的潜在机制将更深入地理解疾病进展的具体原因,并有助于开发新的治疗策略。目前用于检测蛋白质缔合的大多数技术方法包括体外蛋白质提取和纯化,而更具相关性的结果则需要能够在体内使用的方法。少数几种体内蛋白质缔合检测方法之一是基于报告蛋白片段互补。基于蛋白质互补的报告系统依赖于报告蛋白荧光或酶活性的重建,这种重建发生在蛋白质片段重新缔合时,可通过比色法、发光法或荧光法进行测量。蛋白质互补被广泛用于开发报告系统,以分析蛋白质相互作用、对信号转导途径进行功能剖析以及进行高通量筛选以发现新的蛋白质相互作用伙伴。目前利用蛋白质片段互补开发的方法具有的可能性远远超出了简单检测一对蛋白质之间相互作用的范围。

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