A densely modified M 2+ -independent DNAzyme that cleaves RNA efficiently with multiple catalytic turnover.

Sequence-specific cleavage of RNA targets in the absence of a divalent metal cation (M2+ ) has been a long-standing goal in bioorganic chemistry. Herein, we report the in vitro selection of novel RNA cleaving DNAzymes that are selected using 8-histaminyl-deoxyadenosine (dAim TP), 5-guanidinoallyl-deoxyuridine (dUga TP), and 5-aminoallyl-deoxycytidine (dCaa TP) along with dGTP. These modified dNTPs provide key functionalities reminiscent of the active sites of ribonucleases, notably RNase A. Of several such M2+ -free DNAymes, DNAzyme 7-38-32 cleaves a 19 nt all-RNA substrate with multiple-turnover, under simulated physiological conditions wherein only 0.5 mM Mg2+ was present, attaining values of k cat of 1.06 min-1 and a K M of 1.37 μM corresponding to a catalytic efficiency of ∼106 M-1 min-1 . Therefore, Dz7-38-32 represents a promising candidate towards the development of therapeutically efficient DNAzymes.