A New Na+-Dependent RNA-Cleaving DNAzyme with over 1000-fold Rate Acceleration by Ethanol

Enzymes working in organic solvents are important for analytical chemistry, catalysis, and mechanistic studies. Although a few protein enzymes are highly active in organic solvents, little is known regarding nucleic acid-based enzymes. Herein, we report the first RNA-cleaving DNAzyme, named EtNa, that works optimally in concentrated organic solvents containing only monovalent Na+. The EtNa DNAzyme has a rate of 2.0 h(-1) in 54% ethanol (with 120 mm NaCl and no divalent metal ions), and a Kd of 21 mm Na+. It retains activity even in 72% ethanol as well as in DMSO. With 4 mm Na+, the rate in 54% ethanol is >1000-fold higher than that in water. We also demonstrated the use of EtNa to measuring the ethanol content in alcoholic drinks. In total, this DNAzyme has three unique features: divalent metal independent activity, Na+ selectivity among monovalent metals, and acceleration by organic solvents.