A DNA structure-mediated fluorescent biosensor for apurinic/apyrimidinic endonuclease 1 activity detection with ultra-high sensitivity and selectivity

Abstract Human apurinic/apyrimidinic endonuclease 1 (APE1) plays a crucial role in DNA repair and gene regulation. The abnormal variations of the concentration of APE1 in the human blood and tissue cells are highly correlated to various diseases. Thus, APE1 can be used as a biomarker to aid clinical diagnosis of diseases. Detection of APE1 activity with high sensitivity and selectivity simultaneously is difficult to achieve. Here we provide a DNA structure-mediated fluorescent biosensor to solve the above problem. Upon the existence of APE1, the apurinic/apyrimidinic site will be cleaved, and the terminal structure of the hairpin DNA substrate will change. Then the terminal deoxynucleotidyl transferase and endonuclease IV could provide dual fluorescent signal amplification. Through the ingenious structure design of the biosensor, we have improved the reactivity of APE1 and minimized the interference of other enzymes. The results indicate that the detection limit for APE1 is as low as 1.7 × 10-6 U/mL. The biosensor has been successfully applied to the detection of APE1 in real biological samples and the screening of APE1 inhibitors.