A Label-Free Deoxyribozyme Sensor for m⁶A Demethylase Activity Detection

A label-free deoxyribozyme (DNAzyme) biosensor for m(6)A demethylase activity detection is developed. When demethylase FTO (Fat mass and obesity-associated protein), an important m(6)A demethylase of ALKBH demethylase family is present, the m(6)A group is specifically demethylated, and the cleavage activity of DNAzyme is restored. This allows the cleavage product that contains G-quadruplex sequence to bind to thioflavin T (ThT) and generate fluorescence signals. The biosensor shows high specificity and sensitivity, and fast reaction speed. Our study demonstrates a new design of allosteric DNAzyme for sensing. This method represents the first label-free nucleic acid biosensor for FTO assay, providing a feasible route towards inhibitors screening. [GRAPHICS]