A Selective Na + Aptamer Dissected by Sensitized Tb 3+ Luminescence

A previous study of two RNA‐cleaving DNAzymes, NaA43 and Ce13d, revealed the possibility of a common Na + aptamer motif. Because Na + binding to DNA is a fundamental biochemical problem, the interaction between Ce13d and Na + was studied in detail by using sensitized Tb 3+ luminescence spectroscopy. Na + displaces Tb 3+ from the DNAzyme, and thus quenches the emission from Tb 3+ . The overall requirement for Na + binding includes the hairpin and the highly conserved 16‐nucleotide loop in the enzyme strand, along with a few unpaired nucleotides in the substrate. Mutation studies indicate good correlation between Na + binding and cleavage activity, thus suggesting a critical role of Na + binding for the enzyme activity. Ce13d displayed a K d of ∼20 m m with Na + (other monovalent cations: 40–60 m m ). The K d values for other metal ions are mainly due to non‐specific competition. With a single nucleotide mutation, the specific Na + binding was lost. Another mutant improved K d to 8 m m with Na + . This study has demonstrated a Na + aptamer with important biological implications and analytical applications. It has also defined the structural requirements for Na + binding and produced an improved mutant.