Zr-mediated hybrid chain reaction and its application for highly sensitive electrochemical detection of protein kinase A.

An electrochemical platform has been developed to detect protein kinase activity through the combined actions of Zr mediated signal transition and hybridization chain reaction (HCR)-stimulated DNAzymes nanowires. First of all, protein kinase A (PKA) phosphorylates substrate peptides immobilized on gold electrode surface. Thereafter, the DNA1 containing 5'-phosphoryl ends is linked to the phosphorylated substrate peptide via the robust phosphate-Zr-phosphate linkages. By the introduction of molecular beacons (MBs), the DNA1 can open the hairpin structures of MBs through toehold mediated strand displacement (TMSDR), leading to an autonomous stem-opening process and subsequent assembly of G-quadruplex-containing DNA chains by HCR. After the addition of hemin, the formed HRP-mimicking DNAzymes can catalyze the hydroquinone-HO system to generate amplified electrochemical signals. As expected, this method can achieve ultrahigh analytical performance with a low detection limit of 0.02U/mL and exhibit high cost-savings potential without the need for antibody, protease and labeling. Therefore, this method can serve as a new tool for the assay of protein kinase A and its inhibitor screening in the future.