In vitro Selection of Chemically Modified DNAzymes

Abstract

DNAzymes are in vitro selected DNA oligonucleotides with catalytic activities. RNA cleavage is one of the most extensively studied DNAzyme reactions. To expand the chemical functionality of DNA, various chemical modifications have been made during and after selection. In this review, we summarize examples of RNA‐cleaving DNAzymes and focus on those modifications introduced during in vitro selection. By incorporating various modified nucleotides via polymerase chain reaction (PCR) or primer extension, a few DNAzymes were obtained that can be specifically activated by metal ions such as Zn2+ and Hg2+. In addition, some modifications were introduced to mimic RNase A that can cleave RNA substrates in the absence of divalent metal ions. In addition, single modifications at the fixed regions of DNA libraries, especially at the cleavage junctions, have been tested, and examples of DNAzymes with phosphorothioate and histidine‐glycine modified tertiary amine were successfully obtained specific for Cu2+, Cd2+, Zn2+, and Ni2+. Labeling fluorophore/quencher pair right next to the cleavage junction was also used to obtain signaling DNAzymes for detecting various metal ions and cells. Furthermore, we reviewed work on the cleavage of 2′‐5′ linked RNA and L‐RNA substrates. Finally, applications of these modified DNAzymes as biosensors, RNases, and biochemical probes are briefly described with a few future research opportunities outlined at the end.

N-Methoxy-1,3-oxazinane nucleic acids (MOANAs) – a configurationally flexible backbone modification allows post-synthetic incorporation of base moieties

“…Modified or completely artificial base moieties can imbue oligonucleotides with new properties, such as alternative base pairing patterns, [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16] catalytic activity 17,18 or the ability to bind metal ions, either coordinatively [19][20][21][22][23][24] or through formation of a carbon-metal bond. 25,26 Unnatural base moieties are most commonly introduced into oligonucleotides as phosphoramidite building blocks of the corresponding nucleoside analogues by automated solid-phase synthesis.…”

DNAzymes, Novel Therapeutic Agents in Cancer Therapy: A Review of Concepts to Applications

“…Incorporating inverted thymidine at the 3′ terminus of the DNAzyme leading to a 3′-3′ linkage increases the stability and resistance against 3′-exonuclease in human serum [16]. Phosphorothioate linkages in DNAzymes are characterized by the presence of a sulfur atom in one of the nonbridging phosphate oxygen atoms in the cleavage site [17]. These enhance the stability of DNAzymes by providing more resistance to endogenous nucleases.…”

Construction of DNA Biosensors for Mercury (II) Ion Detection Based on Enzyme-Driven Signal Amplification Strategy

“…Thus, the DNAzymes can at least be used to design a biosensor to detect their cofactors. Because Hg 2+ has a weak interaction with phosphate, and high concentration of Hg 2+ can cause the denaturation of DNA, it is difficult to obtain Hg 2+ -specific DNAzyme [ 80 ]. The Hg 2+ -specific DNAzyme named 10–13 was selected using amino/imidazole modified library by Perrin group, and then this group designed a DNAzyme biosensor for Hg 2+ [ 81 ].…”