Key Laboratory of Interfacial Reaction and Sensing Analysis in Universities of Shandong, School of Chemistry and Chemical Engineering , University of Jinan , Jinan 250022 , P. R. China.
Center of Bio and Micro/Nano Functional Materials, State Key Laboratory of Crystal Materials , Shandong University , Jinan 250100 , P. R. China.
Anal Chem. 2019 Mar 5;91(5):3704-3709. doi: 10.1021/acs.analchem.8b05822. Epub 2019 Feb 12.
Mitochondria membrane potential (MMP) play significant roles during metabolism, signaling, and other important bioevents. Visualization of MMP levels is essential for many biological researches. However, fluorescent probes for monitoring MMP levels in dual emission colors are still deficient, which greatly limited the development of relative research areas. In this work, a pair of fluorescent probes have been designed and synthesized to monitor the MMP levels in dual emission colors based on Forster resonance energy transfer (FRET) mechanism. The FRET donor (FixD) is constructed by linking a benzyl chloride group to a fluorophore with bright-green emission. The FixD could target mitochondria and be immobilized in mitochondria by linking to the thiol group of mitochondrial proteins. The FRET acceptor (LA) is designed with green absorption and deep-red emission. In live cells with high MMP levels, FixD and LA both target mitochondria, and deep-red (DR) emission could be detected with the excitation of 405 nm. Particularly, the spectral shift of fluorescence upon the decrease of MMP is up to 110 nm, which is greatly favorable for the clear observation of MMP levels. With the decrease of MMP, LA would be released from mitochondria while FixD would still be immobilized in mitochondria, and decreased DR emission and increased green fluorescence could be detected due to the absence of FRET. In this manner, the MMP levels could be monitored in dual emission colors.
线粒体膜电位 (MMP) 在代谢、信号传递和其他重要生物事件中发挥着重要作用。MMP 水平的可视化对于许多生物学研究至关重要。然而，用于监测双发射颜色 MMP 水平的荧光探针仍然缺乏，这极大地限制了相关研究领域的发展。在这项工作中，设计并合成了一对基于Förster 共振能量转移 (FRET) 机制的荧光探针，用于监测双发射颜色的 MMP 水平。FRET 供体 (FixD) 通过将苄基氯基团连接到具有亮绿色发射的荧光团上来构建。FixD 可以通过与线粒体蛋白的巯基连接靶向线粒体并固定在其中。FRET 受体 (LA) 设计为具有绿色吸收和深红色发射。在 MMP 水平较高的活细胞中，FixD 和 LA 均靶向线粒体，并且可以用 405nm 的激发光检测到深红色 (DR) 发射。特别地，MMP 降低时荧光的光谱位移高达 110nm，这非常有利于 MMP 水平的清晰观察。随着 MMP 的降低，LA 将从线粒体中释放出来，而 FixD 将仍然固定在线粒体中，由于没有 FRET，会检测到 DR 发射减少和绿色荧光增加。通过这种方式，可以在双发射颜色中监测 MMP 水平。
