Enzyme-Mediated Endogenous and Bioorthogonal Control of a DNAzyme Fluorescent Sensor for Imaging Metal Ions in Living Cells
Enzyme-Mediated Endogenous and Bioorthogonal Control of a DNAzyme Fluorescent Sensor for Imaging Metal Ions in Living Cells
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DOI：
10.1002/anie.201910343
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发表时间：
2019-10-15
影响因子：
16.6
通讯作者：
Lu, Yi
中科院分区：
文献类型：
--
作者：
Lin, Yao;Yang, Zhenglin;Lu, Yi
生物正交控制的金属离子传感器的活细胞中的金属离子成像是重要的了解金属离子的分布和波动。本文报道了含有归巢核酸内切酶（I-SceI）的18碱基对识别位点的DNAzyme荧光传感器的内源性和生物正交激活，该位点在7 × 10（10）bp的基因组序列中仅偶然发现一次，因此可以与I-SceI形成用于DNAzyme激活的近生物正交对，对活细胞的影响最小。一旦I-SceI在细胞内表达，它就在识别位点裂解，使DNA酶采用其活性构象。然后，激活的DNAzyme传感器能够在Mg 2+存在下特异性催化底物链的切割，以释放荧光团标记的DNA片段并产生Mg 2+的荧光开启信号。因此，I-SceI生物正交激活10-23 DNA酶，用于HeLa细胞中Mg 2+的成像。
Bioorthogonal control of metal-ion sensors for imaging metal ions in living cells is important for understanding the distribution and fluctuation of metal ions. Reported here is the endogenous and bioorthogonal activation of a DNAzyme fluorescent sensor containing an 18-base pair recognition site of a homing endonuclease (I-SceI), which is found by chance only once in 7x10(10) bp of genomic sequences, and can thus form a near bioorthogonal pair with I-SceI for DNAzyme activation with minimal effect on living cells. Once I-SceI is expressed inside cells, it cleaves at the recognition site, allowing the DNAzyme to adopt its active conformation. The activated DNAzyme sensor is then able to specifically catalyze cleavage of a substrate strand in the presence of Mg2+ to release the fluorophore-labeled DNA fragment and produce a fluorescent turn-on signal for Mg2+. Thus I-SceI bioorthogonally activates the 10-23 DNAzyme for imaging of Mg2+ in HeLa cells.