c-Myc derived DNAzyme and its application for the detection of a novel probiotic bacteria L. fermentum using the...

Catalytically active DNA molecules are called DNAzymes. Single-stranded guanine-rich DNA molecules that can fold into G-quadruplex structures, on interaction with hemin, can exhibit peroxidase-like activity. In this study, we report a c-Myc G-quadruplex forming DNA sequence in complex with hemin, which demonstrates DNAzyme properties and aim to develop a rapid, simple, cost-effective, and culture-free method for detecting probiotic bacteria. The interaction between hemin and c-Myc G-quadruplex DNA sequence was analyzed employing UV-visible absorption and Circular Dichroism spectroscopy. Kinetic analysis of the DNAzyme was conducted with UV-visible absorption and Stopped flow experiments. Electron Paramagnetic Resonance (EPR) was used to study the mechanism of DNAzyme. A novel bacterial strain of Limosilactobacllus fermentum was isolated from a curd sample and the physiological screening and 16S rDNA sequencing was done. Probiotic properties were studied spectrophotometrically. The proposed DNAzyme was employed to detect this hydrogen peroxide-producing bacterium using ABTS as a colorimetric reporter molecule. Hemin binds to c-Myc G-quadruplex with high affinity K_a = 3.77 ± 0.3 × 10⁷ M^-1. The DNAzyme exhibited peroxidase-mimicking activity with a K_m value of 5.8 mM and compound 1 like intermediates were observed. The H₂O₂ produced by the bacteria was used to detect the Limosilactobacillus fermentum with the proposed DNAzyme. With colorimetric, peroxidase mimicking activity, this method could be utilized for the quality check of L. fermentum-containing probiotics, and screening of H₂O₂-producing bacteria in different samples.