Misfolding of a DNAzyme for ultrahigh sodium selectivity over potassium

Herein, the excellent Na(+) selectivity of a few RNA-cleaving DNAzymes was exploited, where Na(+) can be around 3000-fold more effective than K(+) for promoting catalysis. By using a double mutant based on the Ce13d DNAzyme, and by lowering the temperature, increased 2-aminopurine (2AP) fluorescence was observed with addition of both Na(+) and K(+). The fluorescence increase was similar for these two metals at below 10 mM, after which K(+) took a different pathway. Since 2AP probes its local base stacking environment, K(+) can be considered to induce misfolding. Binding of both Na(+) and K(+) was specific, since single base mutations could fully inhibit 2AP fluorescence for both metals. The binding thermodynamics was measured by temperature-dependent experiments revealing enthalpy-driven binding for both metals and less coordination sites compared to G-quadruplex DNA. Cleavage activity assays indicated a moderate cleavage activity with 10 mM K(+), while further increase of K(+) inhibited the activity, also supporting its misfolding of the DNAzyme. For comparison, a G-quadruplex DNA was also studied using the same system, where Na(+) and K(+) led to the same final state with only around 8-fold difference in K(d). This study provides interesting insights into strategies for discriminating Na(+) and K(+).