Ligase-mediated synthesis of CuII-responsive allosteric DNAzyme with bifacial 5-carboxyuracil nucleobases

Abstract

A Cu^II-responsive allosteric DNAzyme has been developed by introducing bifacial 5-carboxyuracil (caU) nucleobases, which form both hydrogen-bonded caU–A and metal-mediated caU–Cu^II–caU base pairs. The base sequence was logically designed based on a known RNA-cleaving DNAzyme so that the caU-modified DNAzyme (caU-DNAzyme) can form a catalytically inactive structure containing three caU–A base pairs and an active form with three caU–Cu^II–caU pairs. The caU-DNAzyme was synthesized by joining short caU-containing fragments with a standard DNA ligase. The activity of caU-DNAzyme was suppressed without Cu^II, but enhanced 21-fold with the addition of Cu^II. Furthermore, the DNAzyme activity was turned on and off during the reaction by the addition and removal of Cu^II ions. Both ligase-mediated synthesis and Cu^II-dependent allosteric regulation were achieved by the bifacial base pairing properties of caU. This study provides a new strategy for designing stimuli-responsive DNA molecular systems.