A Fluorogenic RNA-Based Sensor Activated by Metabolite-Induced RNA Dimerization
A Fluorogenic RNA-Based Sensor Activated by Metabolite-Induced RNA Dimerization
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DOI：
10.1016/j.chembiol.2019.09.013
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发表时间：
2019-12-19
影响因子：
8.6
通讯作者：
Jaffrey, Samie R.
中科院分区：
文献类型：
--
作者：
Kim, Hyaeyeong;Jaffrey, Samie R.
玉米是形成高亲和力准对称同源二聚体的荧光RNA适体。玉米二聚体界面结合DFHO，导致高度光稳定的黄色荧光。由于其光稳定性，玉米将用于基于RNA的小分子生物传感器，其中定量准确性将受到光漂白的影响。在这里，我们描述了一种策略，用于将组成性玉米二聚体转化为小分子调节的荧光生物传感器，其在体外和活细胞中检测S-腺苷甲硫氨酸（SAM）。我们将玉米适体融合到螺旋茎中，该螺旋茎通过从SAM-III核糖开关循环置换SAM适体而工程化。在不存在SAM的情况下，该融合RNA的Corn部分不能二聚化。然而，在结合SAM时，RNA二聚化并结合DFHO。这种基于RNA的生物传感器能够检测活哺乳动物细胞中的SAM动力学。总之，这些数据描述了一类基于RNA的生物传感器的基础上小分子调控的玉米二聚化。
Corn is a fluorogenic RNA aptamer that forms a high-affinity quasi-symmetric homodimer. The Corn dimer interface binds DFHO, resulting in highly photostable yellow fluorescence. Because of its photostability, Corn would be useful in RNA-based small-molecule biosensors, where quantitative accuracy would be affected by photobleaching. Here we describe a strategy for converting the constitutive Corn dimer into a small-molecule-regulated fluorescent biosensor that detects S-adenosylmethionine (SAM) in vitro and in living cells. We fused the Corn aptamer into a helical stem that was engineered by circularly permuting the SAM aptamer from the SAM-III riboswitch. In the absence of SAM, the Corn portion of this fusion RNA is unable to dimerize. However, upon binding SAM, the RNA dimerizes and binds DFHO. This RNA-based biosensor enables detection of SAM dynamics in living mammalian cells. Together, these data describe a class of RNA-based biosensor based on small-molecule-regulated dimerization of Corn.