Small-molecule-mediated rescue of protein function by an inducible proteolytic shunt
Small-molecule-mediated rescue of protein function by an inducible proteolytic shunt
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DOI：
10.1073/pnas.0700816104
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发表时间：
2007-07-03
影响因子：
11.1
通讯作者：
Muir, Tom W.
中科院分区：
文献类型：
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作者：
Pratt, Matthew R.;Schwartz, Edmund C.;Muir, Tom W.
通过翻译后操作控制蛋白质功能已经成为现有遗传系统的一种有吸引力的补充技术。通常，这些方法涉及开发药理学试剂来探测蛋白质功能，而无需为每个蛋白质家族生成独特的化合物。一种常见的策略是使用小分子作为二聚化的化学诱导剂，通过介导两种蛋白质的相互作用。在这里，我们报告使用的二聚化的化学诱导剂的翻译后技术的发展，用于操纵蛋白质的功能。该系统，分裂泛素拯救功能（SURF），将遗传分裂泛素的互补置于雷帕霉素诱导的FK 506结合蛋白和FKBP 12-雷帕霉素结合蛋白二聚化的控制下。在互补作用之前，一个“降解决定子”注定了一个蛋白质会被蛋白酶破坏。加入雷帕霉素通过诱导泛素互补和从降解决定子中去除目的蛋白质，导致蛋白水解分流远离降解。重要的是，天然蛋白质被拯救。我们用萤火虫荧光素酶表征了该系统，并将其应用于三种重要蛋白质的成员：蛋白酶（caspase-3），激酶（v-Src）和转录因子（Smad 3）。这种一般策略应该允许以维持其天然结构和功能的方式诱导拯救各种蛋白质。
Controlling protein function through posttranslational manipulations has emerged as an attractive complementary technology to existing genetic systems. Often these methods involve developing pharmacological agents to probe protein function without the need to generate a unique compound for each protein family. One common strategy uses small molecules that act as chemical inducers of dimerization by mediating the interaction of two proteins. Herein we report the use of a chemical inducer of dimerization for the development of a posttranslational technology for the manipulation of protein function. This system, split ubiquitin for the rescue of function (SURF), places the complementation of genetically split ubiquitin under the control of rapamycin-induced dimerization of FK506-binding protein and FKBP12-rapamycin-binding protein. Before complementation a "degron" dooms a protein of interest for destruction by the proteasorne. Addition of rapamycin results in a proteolytic shunt away from degradation by inducing ubiquitin complementation and deavage of the protein of interest from the degron. importantly, the native protein is rescued. We characterized this system with firefly luciferase and went on to apply it to members of three important dasses of proteins: proteases (caspase-3), kinases (v-Src), and transcription factors (Smad3). This general strategy should allow for inducible rescue of a variety of proteins in such a way that their native structure and function are maintained.