Electrochemical detection of poly(ADP-ribose) polymerase-1 with silver nanoparticles as signal labels by integrating the advantages of homogeneous reaction with surface-tethered detection

Abstract

Poly(ADP-ribose)polymerase-1 (PARP1) could be activated by binding to nucleic acids with specific sequences, thus catalyzing the poly-ADP-ribosylation (PARylation) of target proteins including PARP1 itself. Most of the previously reported electrochemical methods for the determination of PARP1 were relied on the electrostatic interactions, which required the pre-immobilization of DNA on an electrode for the capture of PARP1. Herein, we reported an “immobilization−free” electrochemical strategy for the assays of PARP1 on the basic of avidin-biotin interaction. Once PARP1 was activated by binding with the specific double-stranded DNA (dsDNA) in a homogeneous solution, the biotinylated nicotinamide adenine dinucleotide (biotin-NAD+) was transferred onto PARP1, resulting in the formation of biotinylated PAR polymers. The resulting biotinylated PAR polymers were then captured by a neutravidin (NA)-modified electrode through avidin-biotin interactions. The rich biotin moieties in the PAR polymers allowed for the capture of NA-modified silver nanoparticles (NA-AgNPs) through the avidin-biotin interactions. The surface-tethered AgNPs produced a well-defined electrochemical signal due to the characteristic solid-state Ag/AgCl process. The “immobilization−free”, electrostatic interaction-independent electrochemical biosensor exhibited low background current, high sensitivity, and good stability. It has achieved the determination of PARP1 with a detection limit down to 0.7 mU. The biosensor was further applied to determine the inhibition efficiency of potential inhibitors with a satisfactory result. This method shows promising potential applications in PARP1-related clinical diagnosis and drug discovery.