A fluorescence sensor for protein kinase activity detection based on gold nanoparticles/copper nanoclusters system

Abstract

In this work, we develop a novel biosensor for the detection of protein kinase A (PKA) based on the fluorescence inner-filter effect (IFE) between peptide functionalized gold nanoparticles (p-AuNPs) and polyethyleneimine protected copper nanoclusters (PEI-CuNCs). The fluorescence of PEI-CuNCs could be quenched by p-AuNPs through IFE. In the presence of adenosine 5′-triphosphate (ATP) and PKA, the substrate peptide of p-AuNPs was phosphorylated and then selective aggregation of phosphorylated p-AuNPs was triggered by Zr4+ ion coordination. Compared with the monodispersed p-AuNPs, the aggregated p-AuNPs have less fluorescence quenching effect on PEI-CuNCs. So the activity of PKA could be effectively monitored according to the change of fluorescence signal in the range of 0.1–6.0U/mL, and the detection limit (LOD) is 0.038U/mL. In addition, we explored the inhibitory effect of H-89 on protein kinase activity. The developed method was also applied to the investigation of drug-induced PKA activation in HepG-2 cell, which provides one promising tool for kinase-related biochemical fundamental research and inhibitor screening with excellent performance.