Exploration of Catalytic Nucleic Acids on Porphyrin Metalation and Peroxidase Activity by in Vitro Selection of Aptamers for N-Methyl Mesoporphyrin IX

摘要

To develop a novel light-up probe and DNAzyme, we selected aptamers for N-methyl mesoporphyrin IX (NMM), a common fluorogenic analogue of coenzyme hemin, by a modified affinity chromatography-based systematic evolution of ligands by exponential enrichment (SELEX). Two truncated aptamers Nm1 and Nm2 with low micromolar dissociation constants (0.75 and 13.27 mu M) were obtained after 11 rounds of selection and the final minimized 39-mer aptamer Nm2.1 showed 24-fold fluorescence enhancement for NMM at saturated concentration. Study of the interactions between aptamers and other porphyrin compounds by circular dichroism (CD) and absorption spectroscopy showed that Nm1 mainly assembled as a stem-loop structure, which exhibited a catalytic activity for the metal insertion reaction of mesoporphyrin IX with 3.3-fold rate enhancement. In contrast, the G-rich Nm2 and Nm2.1 were likely to form G-quadruplexes in the presence of alkali metal cations (K+ and Na+), which displayed excellent peroxidase activity exhibiting 19-fold higher catalytic efficiency than hemin alone. The selected aptamers could therefore be used as novel light-up fluorescent probes and DNAzymes by pairing with porphyrin compounds that have potential to construct sensors for various applications.