Imaging dynamics of membrane proteins of live cells in a wash-free and real-time manner has been a challenging task. Herein, we report unprecedented applications of malachite green(MG), an organic dye widely used in p...Imaging dynamics of membrane proteins of live cells in a wash-free and real-time manner has been a challenging task. Herein, we report unprecedented applications of malachite green(MG), an organic dye widely used in pigment industry, as a switchable fluorophore to monitor membrane enzymes or noncatalytic proteins in live cells. Conformationally flexible MG is non-fluorescent in aqueous solution, yet covalent binding with endogenous proteins of cells significantly enhances its fluorescence at 670 nm by restricting flexibility of dye. Integrating a phosphate-caged quinone methide precursor with MG yielded a covalent labeling fluorogenic probe, allowing real-time imaging of membrane alkaline phosphatase(ALP,a model catalytic protein) activity in live cells with over 100-fold enhancement of fluorescence intensity.Moreover, MG is also applicable to image non-catalytic protein by conjugation with protein-specific ligand. A fluorogenic probe consisted of c-RGDf K peptide and MG proved to be compatible with wash-free and real-time visualization of non-catalytic integrin α_(v)β_(3) in live cells with high contrast.展开更多
基金financially supported by the National Natural Science Foundation of China (No. 22077031)the Research Program of State Key Laboratory of Bioreactor Engineeringthe Fundamental Research Funds for the Central Universities。
文摘Imaging dynamics of membrane proteins of live cells in a wash-free and real-time manner has been a challenging task. Herein, we report unprecedented applications of malachite green(MG), an organic dye widely used in pigment industry, as a switchable fluorophore to monitor membrane enzymes or noncatalytic proteins in live cells. Conformationally flexible MG is non-fluorescent in aqueous solution, yet covalent binding with endogenous proteins of cells significantly enhances its fluorescence at 670 nm by restricting flexibility of dye. Integrating a phosphate-caged quinone methide precursor with MG yielded a covalent labeling fluorogenic probe, allowing real-time imaging of membrane alkaline phosphatase(ALP,a model catalytic protein) activity in live cells with over 100-fold enhancement of fluorescence intensity.Moreover, MG is also applicable to image non-catalytic protein by conjugation with protein-specific ligand. A fluorogenic probe consisted of c-RGDf K peptide and MG proved to be compatible with wash-free and real-time visualization of non-catalytic integrin α_(v)β_(3) in live cells with high contrast.
