An RNA‐Cleaving Catalytic DNA Accelerated by Freezing

Abstract

The EtNa DNAzyme was isolated during the isopropanol precipitation step of an in vitro selection effort. Although inactive with the intended cofactor, its RNA cleavage activity was observed under a few conditions. With Na , EtNa was highly active in ∼50 % ethanol, whereas in water, it was highly active with Ca . In this work, we showed that the EtNa DNAzyme was accelerated by freezing in water in the presence of Na . The apparent K value reached 6.2 mm Na under the frozen condition, over 20 times tighter than that in water at room temperature. With 10 mm Na , EtNa had a cleavage rate of 0.12 h after freezing at -20 °C. This effect was unique to EtNa, as all other tested DNAzymes were inhibited by freezing except for the Na -specific NaA43. Freezing also inhibited EtNa if Ca was used. We attributed this to the concentrations of EtNa and Na in the micropockets between ice crystals, but divalent metals might misfold DNA. Overall, we have systematically studied the effect of freezing on the RNA-cleavage activity of DNAzymes. The DNAzyme sequence and the metal ion species are both crucial to determine the effect of freezing.

Freezing‐Assisted Conjugation of Unmodified Diblock DNA to Hydrogel Nanoparticles and Monoliths for DNA and Hg2+ Sensing

“…[18] During freezing, aportion of the water crystallized to form ice,leaving the solutes in the remaining non-ice regions where the monomers and DNAwere likely to be concentrated (Figure 2C). [19] Such ac oncentration effect can accelerate reactions, [20] which may favor DNAa ttachment and gelation. To study the relationship between freezing and gel yield, we weighed the gels (formed at À20 8 8C).…”

Nucleic Acid Catalysis under Potential Prebiotic Conditions

“…[203] Eutectic freezing can also activate EtNa, while other DNAzymes that depend on divalent or trivalent metals are inhibited under these conditions. [204] This again highlights the interchangeability of freezing, organic dehydration or evaporation to achieve activation of metal-independent nucleic acid catalysts.…”

“…Given that EtNa shows cooperative binding of and activation by Ca 2+ (in contrast to Mg 2+ ) it can also be used as an ultrasensitive biosensor capable of detecting Ca 2+ levels down to 1.4 μ m Ca 2+ . Eutectic freezing can also activate EtNa, while other DNAzymes that depend on divalent or trivalent metals are inhibited under these conditions . This again highlights the interchangeability of freezing, organic dehydration or evaporation to achieve activation of metal‐independent nucleic acid catalysts.…”

DNA Assembly‐Based Stimuli‐Responsive Systems

“…[74] Protein and Small Molecule Sensors: The aptamers of a variety of proteins and small molecules have been used in sensing systems combined with stimuli-responsive DNA nanotechnology. [150,182,295,[321][322][323][324][325][326] For example, a pH-based ATP fluorescence sensing system was realized by regulation of the distance between the two segments of aptamer by the i-motif structure. [326] Xiang's group developed a thrombin detection strategy by integrating proximity binding-induced strand displacement and metal ion-dependent DNAzyme recycling amplification (Figure 9c).…”