Smart Dual Quenching Strategy Enhances the Detection Sensitivity of Intracellular Furin

Abstract

Development of sensitive fluorescence "Turn-On" strategies for imaging enzyme activity in living cells is of disease-diagnostic importance but remains challenging. Herein, by employing a click condensation reaction and rational design of a single quenched probe Cys(StBu)-Lys(Gly-Lys(DABCYL)-Gly-Gly-Arg-Arg-Val-Arg-Gly-FITC)-CBT (1), we developed a "smart" dual quenching strategy and applied it to detect intracellular furin activity with enhanced sensitivity. At physiological conditions, 1 was subjected to reduction-controlled condensation reaction to form 1-NPs and its fluorescence intensity further dropped to 1/2.8 of its original. Upon furin cleavage in vitro, the dual quenched 1-NPs had fluorescence "Turn-On" contrast 11-fold more than that of single quenched control probe FITC-Gly-Arg-Val-Arg-Arg-Gly-Gly-Lys(DABCYL)-Gly-OH (1-P). Live cell imaging results indicated that 1 showed fluorescence "Turn-On" contrast 6.3-fold of that of 1-P for sensing intracellular furin activity. We envision that, by replacing the RVRR substrate with other enzyme-cleavable ones, our versatile "smart" dual quenching strategy could be easily adjusted for the detection (or imaging) of other intracellular enzymes' activity with enhanced sensitivity.

Increasing Photothermal Efficacy by Simultaneous Intra‐ and Intermolecular Fluorescence Quenching

“…However, efficiency of above self‐quenching only relies on the packing density of fluorophores in the aggregated state via weak intermolecular interactions (e.g., π–π stacking, hydrogen bond, Van der Waals' force) which we denote as intermolecular quenching herein. If the self‐quenching of the fluorophores was achieved after a covalent bond formation (or chemical reaction) drawing the fluorophores nearer (herein we denote this type of quenching as intramolecular quenching), we have proven that intramolecular quenching is even more efficient than intermolecular quenching . Thus, we naturally thought, after a small organic PTA entering tumor cells, if its fluorescence was quenched by both intra‐ and intermolecular quenching, PTT efficiency of the PTA should be greatly enhanced.…”

Spatiotemporal‐Controlled Reporter for Cell‐Surface Proteolytic Enzyme Activity Visualization

“…Another significant feature of the PFP is the ability to visualize the proteolysis activities specifically on the cell surface. To this end, an investigation was conducted to confirm the ability of the hydrophobic lipid chain to assist in localizing the fluorescent reporter within the cell membranes –. Generally, MDA‐MB‐468 and LoVo cells were incubated with PFP for 1 h during the process of live‐cell fluorescence imaging.…”

Furin‐Controlled Fe3O4 Nanoparticle Aggregation and 19F Signal “Turn‐On” for Precise MR Imaging of Tumors

“…[22] It is well-documented that Arg-X-Lys/Arg-Arg↓X peptide can be cleaved by furin. [24] But to the best of our knowledge, a furin-controlled simultaneous T 2 1 H MRI enhancement and 19 F MRI "Turn-On" for precise tumor imaging has not been reported. [24] But to the best of our knowledge, a furin-controlled simultaneous T 2 1 H MRI enhancement and 19 F MRI "Turn-On" for precise tumor imaging has not been reported.…”