Guanine-rich nucleic acid sequences can fold into G-quadruplex (G4) structures of different topologies. Previous studies have shown that antiparallel G4 structures, when in complex with hemin, act as catalysts for the peroxidation of organic compounds. Therefore, such nucleic acid sequences can be referred to as peroxidase-like deoxyribozymes (pDz) – functional analogues of horseradish peroxidase, a protein enzyme widely used in various colorimetric assays. The catalytic properties of pDz have been explored in designing sensors for the analysis of metal ions, small molecules, and nucleic acids.  A number of organic indicators that become colored upon oxidation have been suggested in the literature. Here, we systemically compared a selection of common organic indicators available for colorimetric assays utilizing horseradish peroxidase or pDz. We utilized PW17, a short oligonucleotide that inherently folds into a pDz, as a model to study the peroxidation reaction of 3,3′-diaminobenzidine (DAB), 2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), 3,3′,5,5′-tetramethylbenzidine (TMB), 2,3-benzopyrrole (Indole), and 3-amino-9-ethylcarbazole (AEC). The results of this study detail the comparative strengths and weaknesses of these substrates for use in peroxidase-based colorimetric assays.

Comparison of substrates for colorimetric assays using peroxidase-like deoxyribozyme PW17

255th National Meeting (2018)

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