An integrated G-quadruplex-hemin DNAzyme for detection of RNase H activity and its inhibition

In this work, a novel DNAzyme for rapid and sensitive detection of RNase H activity and its inhibitors was developed based on hemin-modified DNA (G4-hemin complex). G4-hemin exhibited superior peroxidase-like activity. It could be accomplished in one step that neither required K⁺ for G-quadruplex assembly nor relied on externally added hemin. Consequently, the background interference induced by free hemin was substantially diminished. Experimental results showed a linear relationship with RNase H at concentrations ranging from 0 to 0.8 U mL^-1, with the limit of detection (LOD) being 5.7 × 10^-3 U mL^-1. Furthermore, the inhibitory effects of the two inhibitors were also evaluated experimentally. The G4-hemin complex exhibited good selectivity and sensitivity for RNase H, which could be applied to screen inhibitors of RNase H. This method enables the evaluation of RNase H activity through straightforward visual color inspection, providing a powerful tool for point-of-care testing (POCT) and research on RNase H activity and inhibition.