Harnessing Nature’s Insights: Synthetic Small Molecules with Peroxidase‐Mimicking DNAzyme Properties

Improved method for chemiluminescent determination of peroxidase-mimicking DNAzyme activity

“…As shown in the separate experiment, the value of the dissociation constant (K d ) for the PMDNAzyme was 2.7 Â 10 À7 M (see online supplementary material). Similar K d values for other PMDNAzymes were reported previously [12,44]. Searching for new hemin aptamers with higher affinity may improve the sensitivity of the PMDNAzyme assay.…”

Insights into how nucleotide supplements enhance the peroxidase-mimicking DNAzyme activity of the G-quadruplex/hemin system

“…ATP, H 2 O 2 , ABTS/TMB and hemin concentrations) in order to establish an ATP-boosted DNAzyme standard protocol (as a function of the probe). These evaluations were performed in our ‘classical’ conditions (24,28), i.e. in Caco.KTD buffer (10 mM lithium cacodylate buffer (pH 7.2) plus 10 mM KCl/90 mM LiCl, 0.05% Triton X-100 and 0.1% DMSO) with 22AG (a quadruplex-forming 22-nucleotide sequence d[AG 3 (T 2 AG 3 ) 3 ] that mimics the 3’-overhang of the human telomeres).…”

“…the study of the influence of the concentration of ATP, H 2 O 2 , hemin and chromogenic probes on the efficiency of the oxidation protocol. The catalytic capability of a system initially composed of 22AG (0.5 mM), hemin (1.0 µM) and H 2 O 2 (0.6 mM) to oxidize ABTS (2.0 mM) is monitored via the appearance of the ABTS ·+ UV-Vis signal at 420 nm (24,28). As further detailed in the Supplementary Data, several rows of experiments were conducted to optimize the ABTS oxidation protocol: experiments were performed with variable amounts of ATP (from 0 to 10 mM, see Supplementary Figure 1A), H 2 O 2 (from 0.6 to 6.0 mM, see Supplementary Figure 1B), hemin (from 0.25 to 5.0 µM, see Supplementary Figure 1C) and ABTS (from 0.5 to 15 mM, see Supplementary Figure 1D), and it was concluded, in light of these results, that the standard DNAzyme protocol for 22AG-mediated ABTS oxidation is composed of 10 mM ATP, 6 mM H 2 O 2 , 1 µM hemin and 5 mM ABTS, in Caco.KTD buffer.…”

“…The above-mentioned DOTASQ approach is in this way interesting but suffers, despite its overall success, from two drawbacks: first, DOTASQ is itself a pre-catalyst for DNAzyme process (28), and consequently it is responsible for an elevated background reaction that makes the lowering of the detection limit hardly feasible; second, even if the chemical access of DOTASQ is straightforward (27), it is not commercially available, and consequently it will not be readily used in every common laboratories. Because we are convinced that it is worth finding efficient DNAzyme-enhancing agents, we decided to invest efforts to circumvent these drawbacks: results presented herein, which highlight the very positive role of ATP—and more generally of nucleotides—for boosting DNAzyme processes, are inspired by a recent report from D.-M. Kong and co-workers (29); ATP indeed fulfils the requirements of an ideal boosting agent since it is commercially available (from classical suppliers), cheap (∼10€/g at the highest purity grade) and practically convenient (notably in terms of both chemical stability and water-solubility).…”

Structure–activity relationship study for design of highly active covalent peroxidase-mimicking DNAzyme

“…[10][11][12][13][14] Nevertheless, the widespread application of non-covalent PMDNAzymes is restricted because of their relatively low sensitivity. 15,[17][18][19] However, this drawback could be overcome by a covalent binding of hemin to its aptamer resulting in formation of a so called covalent PMDNAzyme. 16 One of the main reasons of the low sensitivity of non-covalent PMDNAzyme determination is dissociation of the PMDNAzyme molecules to the initial components (DNA and hemin) at nano-and picomolar concentrations because of a poor affinity of hemin to its aptamers (K d was ca.…”