A novel DNA/hemin complex with enzyme-like activity selected from a hairpin DNAs library at zero H2O2 concentration

• A newly designed DNA in a hairpin shape structure called G5H with an enzyme-like activity was identified through a fast colorimetric screening experiment. • G5H showed oxidase/proxidase like activity in a complex form with hemin. • G5H in a complex form with hemin (G5HH) can oxidase TMB and some chemicals without H2O2. • Hemin interacts with distinct forms of DNA hairpins and can be active. • These findings offer clues that there are structures of DNA –simpler than G4 DNAs- which capable of mimicking enzyme activity without the need to form G-quadruplexes. Here, a library of short DNAs possessing stem-loop conformation is examined for enzyme-like activity at zero H 2 O 2 concentration. The stem part of all hairpins was identical and the loop sequences were diversified with runs of different nucleotides (A, T, C, G). According to the results, we found that G-type hairpins with 3, 4 and 5 guanines in the loop part are able to be complex with hemin and, oxidize TMB. Among them, a complex named G5HH showed an apparent oxidase-like activity and, was chosen for further studies. UV-thermal analysis, gel shift assay, mutation and structural tests with molecular docking results confirmed that G-tetrads are not formed in our experimental system. So, in this study we describe the functionality of a distinct type of DNA in the hairpin fold as a new class of DNAzymes. Discovering such property for simple DNA hairpins would hold great promise for wide applications in bio-nanotechnology and other fields of research.