Engineering oligonucleotide-based peroxidase mimetics for the colorimetric assay of S1 nuclease

Abstract

Herein we propose a rational design to construct peroxidase mimetics in response to S1 nuclease by employing guanine-rich oligonucleotides and Cu²⁺ ions. The enzymatic activities of DNA–Cu(II) are highly dependent upon the binding stoichiometry between Cu²⁺ and DNA. In the H₂O₂-mediated oxidation of 3,3′,5,5′-tetramethylbenzidine, the catalytic activity of Cu²⁺ can be accelerated approximately 88 times in the presence of AG₃(T₂AG₃)₃, whereas it shows slight enhancement in the presence of a mononucleotide (GMP). The coordination of Cu²⁺ with DNA structures greatly strengthens its ability to generate reactive oxygen species. The most intriguing finding in this study is that DNA–Cu(II) complexes lose their enzymatic activities after the cleavage of DNA scaffolds by S1 nuclease. A colorimetric method was established for selectively evaluating the S1 nuclease activity with a limit of detection of 4.7 × 10⁻³ U mL⁻¹, only by using label-free oligonucleotides and copper ions without complicated synthesis and apparatus. This facile method can also be applicable for quantitative analysis in biological fluids.