A general strategy to construct small molecule biosensors in eukaryotes
A general strategy to construct small molecule biosensors in eukaryotes
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DOI：
10.7554/elife.10606
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发表时间：
2015-12-29
期刊：
影响因子：
7.7
通讯作者：
Baker, David
中科院分区：
文献类型：
--
作者：
Feng, Justin;Jester, Benjamin W.;Baker, David
用于小分子的生物传感器可用于从代谢工程到转录的正交控制的应用。在这里，我们通过使用原则上可以应用于任何目标分子的方法来生产基于配体结合域（LBD）的生物传感器。LBD与荧光蛋白或转录激活因子融合，并通过突变而不稳定，使得融合物仅在含有靶配体的细胞中积累。我们通过开发地高辛和孕酮的生物传感器来说明这种方法的力量。向表达生物传感器的酵母、哺乳动物或植物细胞中添加配体以高达100倍的动态范围激活转录。我们使用生物传感器来改善酵母中双烯醇酮向孕酮的生物转化，并调节哺乳动物细胞中的CRISPR活性。这项工作提供了一个通用的方法来开发生物传感器的广泛的分子在真核生物。
Biosensors for small molecules can be used in applications that range from metabolic engineering to orthogonal control of transcription. Here, we produce biosensors based on a ligandbinding domain (LBD) by using a method that, in principle, can be applied to any target molecule. The LBD is fused to either a fluorescent protein or a transcriptional activator and is destabilized by mutation such that the fusion accumulates only in cells containing the target ligand. We illustrate the power of this method by developing biosensors for digoxin and progesterone. Addition of ligand to yeast, mammalian, or plant cells expressing a biosensor activates transcription with a dynamic range of up to 100 -fold. We use the biosensors to improve the biotransformation of pregnenolone to progesterone in yeast and to regulate CRISPR activity in mammalian cells. This work provides a general methodology to develop biosensors for a broad range of molecules in eukaryotes.