High-contrast and real-time visualization of membrane proteins in live cells with malachite green-based fluorogenic probes

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.

Selective inhibition of resistant bacterial pathogens using a β-lactamase-activatable antimicrobial peptide with significantly reduced cytotoxicity

The expression ofβ-lactamase,particularly metallo-β-lactamase(MBL)in bacteria has caused significant resistance to clinically importantβ-lactam antibiotics,including life-saving carbapenems.Antimicrobial peptides(AMPs)have emerged as promising therapeutic agents to combat antibiotic resistance.However,the cytotoxic AMPs has been one of the major concerns for their applications in clinical practice.Herein,we report a novel cephalosporin-caged AMP,which shows significantly reduced cytotoxicity,hemolytic activity,and antibacterial activity but turns highly active against bacteria upon specific hydrolysis by the antimicrobial resistance-causativeβ-lactamase.Further investigations demonstrate thisβ-lactamaseactivatable AMP selectively inactivates resistant bacterial pathogens over susceptible bacteria.This strategy should be applicable to other AMPs as a potential solution for the treatment of infectious diseases caused byβ-lactamase-expressing pathogenic bacteria.