In Situ Imaging of Furin Activity with a Highly Stable Probe by Releasing of Precipitating Fluorochrome
| Autor: | Tanggang Deng, Xiao-Bing Zhang, Shuai Xu, Shuangyan Huan, Ke Li, Hong-Wen Liu, Xiaoxiao Hu, Jun-Bin Li |
|---|---|
| Rok vydání: | 2018 |
| Předmět: |
In situ
animal structures Fluorophore viruses Golgi Apparatus 010402 general chemistry 01 natural sciences Furin activity Analytical Chemistry chemistry.chemical_compound Cell Line Tumor Humans Tumor growth Furin Fluorescent Dyes biology 010405 organic chemistry Chemistry Cobalt Fluorescence 0104 chemical sciences Microscopy Fluorescence embryonic structures Biophysics biology.protein Hypoxia-Inducible Factor 1 Proprotein Convertases Peptides Linker |
| Zdroj: | Analytical chemistry. 90(19) |
| ISSN: | 1520-6882 |
| Popis: | Furin, a kind of trans-Golgi proprotein convertases, plays important role in various physiological processes. It is overexpressed in many cancers and relates to tumor growth and migration. In situ detection and imaging of furin is of great significance for obtaining real-time information about its activity. However, the previously reported fluorescent probes for furin usually failed to realize in situ detection and long-term bioimaging, because these probes are based on water-soluble fluorophores, which tend to diffuse away from the reaction sites after converted by furin. Such a problem can be addressed by designing a probe, which releases a precipitating fluorophore upon furin conversion. Herein, we developed a probe HPQF for in situ detection of endogenous furin activity and long-term bioimaging by integrating a strictly insoluble solid-state fluorophore 6-chloro-2-(2-hydroxyphenyl) quinazolin-4(3H)-one (Cl-HPQ) with a furin specific peptide substrate (RVRR) through a self-immolative linker. The HPQF probe shows high selectivity and sensitivity to furin. Upon converted by furin, HPQF releases free Cl-HPQ, which precipitates near the enzyme active site. The precipitates emit bright solid-state fluorescence for in situ imaging. HPQF could truly visualize the location of intracellular furin, which was further confirmed by colocalization and immunofluorescence experiments. Excitingly, the long-term bioimaging was also achieved benefiting from its outstanding signal-stability and antidiffusion ability. HPQF was further utilized to monitor the level change of furin under stabilizing of hypoxia-inducible factor (HIF) regulated by cobalt chloride (CoCl |
| Databáze: | OpenAIRE |
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