A label-free electrochemical biosensor for methyltransferase activity detection and inhibitor screening based on graphene quantum dot and enzyme-catalyzed reaction

Abstract

A label-free electrochemical biosensor for methyltransferase (M.SssI MTase) activity detection and inhibitor screening was developed based on the amplification of graphene quantum dot (GQD) and enzyme-catalyzed reaction. In the construction of the electrochemical biosensor, auxiliary DNA hybridized with capture DNA to form the double-stranded DNA structure which contained a specific recognition sequence (5′-CCGG-3′) for both M.SssI MTase and restriction endonuclease HpaII. The presence of M.SssI MTase caused the methylation of CpG site in the ds-DNA, which could not be digested by HpaII, causing GQD bound to the undigested ds-DNA and acted as new platform for the immobilization of horseradish peroxidase (HRP) through noncovalant assembly. The modified HRP catalyzed the hydrogen peroxide-mediated oxidation of 3, 3′, 5, 5′-tetramethylbenzidine, resulting an electrochemical signal output. The proposed biosensor realized sensitivity detection of M.SssI MTase activity in the range of 1–40UmL−1 with a detection limit of 0.3UmL−1, and this dual-amplified biosensor had been successfully applied in M.SssI MTase activity inhibitor screening of procaine and epicatechin. Furthermore, this proposed strategy can also be extended for other methyltransferase detection, which had a promising application in clinical diagnosis and drug development.