A densely modified M²⁺-independent DNAzyme that cleaves RNA efficiently with multiple catalytic turnover

Abstract

Modified dNTPs permit selection of DNAzymes that cleave RNA targets in the absence of a divalent metal cation (M2+) to meet a long-standing goal in bioorganic chemistry.

Evaluating the Catalytic Potential of a General RNA‐Cleaving FANA Enzyme

“…The strong preference of DNAzyme 10–23 for natural RNA substrates over a chimeric DNA substrate is striking when compared with other DNAzymes and likely reflects the strong selective pressure used to isolate and optimize its activity by in vitro selection and directed evolution. By comparison, most DNAzymes that were selected for RNA substrates show only modest decreases in activity when challenged with a chimeric DNA substrate . In fact, this substrate specificity seems to be more consistent with DNAzymes that are unable to cleave RNA substrates because they were selected to recognize and cleave chimeric DNA substrates .…”

“…By comparison, most DNAzymes that were selected for RNA substrates show only modest decreases in activity when challenged with a chimeric DNA substrate. [13] In fact, this substrate specificity seems to be more consistent with DNAzymes that are unable to cleave RNA substrates because they were selected to recognize and cleave chimeric DNA substrates. [2] Conformationally, a chimeric substrate with DNA binding arms that adopt a B-form duplex is more favorable to RNA cleavage through transesterification.…”

Catalytic Nucleic Acids: Biochemistry, Chemical Biology, Biosensors, and Nanotechnology

“…To solve this problem, using modified nucleotides to mimic the chemical functionalities of RNase A has been attempted and metal-free cleavage was achieved in some cases. An example is the DNAzyme 7-38-32 ( Figure 5D), which cleaves an RNA substrate with a catalytic rate of 1.06 min À1 in the presence of only 0.5 mM Mg 2+ (Wang et al, 2018). Other efforts such as the selection of divalent metal-free DNAzymes have also been made, some of which required only Na + for activity (Geyer and Sen, 1997;Torabi et al, 2015;Zhou et al, 2017a).…”

A Trisbenzimidazole Phosphoramidite Building Block Enables High-Yielding Syntheses of RNA-Cleaving Oligonucleotide Conjugates

“…Although in modern life sciences gapmers [2,14,15] and siRNAs [16] are the preferred tools for achieving the site-specific cleavage of RNA strands, there is still interest in synthetic ribonucleases and recent years have seen important progress in the field. Effective catalysts that are based on metal ions [17][18][19][20][21][22][23], guanidines or polyamines [24][25][26][27], peptide conjugates [28][29][30][31][32], and deoxyribozymes [33,34] have been described. Starting from heterocyclic guanidine analogs [35], we have investigated the conjugates of tris(2-aminobenzimidazoles) and different types of oligonucleotides, such as DNA [36], PNA [37,38], or DNA-LNA mixmers [39], which were shown to act as sequence-specific metal-free RNA cleavers.…”