Near-Infrared Fluorescence Probes for Enzymes Based on Binding Affinity Modulation of Squarylium Dye Scaffold
Near-Infrared Fluorescence Probes for Enzymes Based on Binding Affinity Modulation of Squarylium Dye Scaffold
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DOI：
10.1021/ac300061a
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发表时间：
2012-05-15
影响因子：
7.4
通讯作者：
Nagano, Tetsuo
中科院分区：
文献类型：
--
作者：
Oushiki, Daihi;Kojima, Hirotatsu;Nagano, Tetsuo
我们提出了一种新的近红外（NIR）荧光探针的设计策略，利用染料-蛋白质相互作用作为荧光增强的触发器。其设计原理涉及用可裂解的官能团修饰聚甲炔染料，从而降低染料的蛋白质结合亲和力。当这些官能团通过与靶酶的特异性相互作用被去除时，染料的蛋白质亲和力恢复，蛋白质结合发生，并且染料的荧光强烈增强。为了验证这一策略，我们首先设计并合成了一种碱性磷酸酶（ALP）传感器，通过将磷酸盐引入方酸染料支架;该传感器能够在Western印迹分析中检测ALP标记的二抗。其次，我们合成了一种探针β-半乳糖苷酶（广泛用作基因表达的报告）的β-半乳糖取代的方酸盐支架的装置，这种传感器能够可视化β-半乳糖苷酶的活性在体外和体内。我们的策略应适用于获得广泛的目标酶的近红外荧光探针。
We present a novel design strategy for near-infrared (NIR) fluorescence probes utilizing dye-protein interaction as a trigger for fluorescence enhancement. The design principle involves modification of a polymethine dye with cleavable functional groups that reduce the dye's protein-binding affinity. When these functional groups are removed by specific interaction with the target enzymes, the dye's protein affinity is restored, protein binding occurs, and the dye's fluorescence is strongly enhanced. To validate this strategy, we first designed and synthesized an alkaline phosphatase (ALP) sensor by introducing phosphate into the squarylium dye scaffold; this sensor was able to detect ALP-labeled secondary antibodies in Western blotting analysis. Second, we synthesized a probe for beta-galactosidase (widely used as a reporter of gene expression) by means of beta-galactosyl substitution of the squarylium scaffold; this sensor was able to visualize beta-galactosidase activity both in vitro and in vivo. Our strategy should be applicable to obtain NIR fluorescence probes for a wide range of target enzymes.