Evaluation of the sequence-dependent relative activity of APE1 for optimal biosensing design.

Apurinic/apyrimidinic endonuclease 1 (APE1) can selectively incise the AP site of DNA, thus is universal for various DNA substrates for flexible endonuclease-assisted signal amplification. However, the substrate preference of APE1 has never been systematically investigated. Therefore in this work, the detailed sequence-dependent relative activity of APE1 was determined. It turned out that the APE1 activity did vary with the change of the adjacent and opposite bases, and over 10-fold relative activity difference was observed for different sequence combinations. Such difference is appreciable enough to induce evident impact on APE1-involved biosensing. With an APE1 probe designed for cycled signal amplification, the sensitivities followed exactly with the above activity order. Compared with Nb.BbvCl, the sensitivity of the APE1 probe varied between higher and lower than the Nb.BbvCl probe (with varied substrates), demonstrating the importance of the sequence-dependent relative activity of APE1 for optimal biosensor development. Moreover, the above APE1 probe design was harvested and engineered for sensitive biosensing of uracil-DNA glycosylase (UDG). Through theoretical analysis of the interaction between APE1 and the substrates, the accuracy of the determined sequence-dependent relative activity of APE1 was partially confirmed.