Abstract
DNAzymes are catalytically active DNA molecules that are normally isolated through in vitro selection methods, among which RNA-cleaving DNAzymes that catalyze the cleavage of a single RNA linkage embedded within a DNA strand are the most studied group of this DNA enzyme family. Recent advances in DNA nanotechnology and engineering have generated many RNA-cleaving DNAzymes with unique recognition and catalytic properties. Over the past decade, numerous RNA-cleaving, DNAzymes-based functional probes have been introduced into many research areas, such as in vitro diagnostics, intracellular imaging, and in vivo therapeutics. This review focus on the fundamental insight into RNA-Cleaving DNAzymes and technical tricks for their intracellular and in vivo applications, highlighting the recent progress in the clinical trial of RNA-Cleaving DNAzymes with selected examples. The challenges and opportunities for the future translation of RNA-cleaving DNAzymes for biomedicine are also discussed.
Highlights
Catalysis in biology has long been considered to be the function of catalytic RNAs and protein enzymes [1]; many studies have reported the discovery of catalytic DNAs since Breaker and Joyce’s pioneer work on DNAzymes in 1994 [2,3,4].These DNAzymes are normally isolated through in vitro selection methods from a random DNA library consisting of approximately 1015 DNA sequences [5]
Among the abovementioned DNAzymes, RNA-cleaving DNAzymes that catalyze the cleavage of a single RNA linkage embedded within a DNA strand are the most studied group of this DNA
Developed the first RCDs-based probe for metal ions in living cells based on the integration of gold nanoparticles (AuNPs), a uranyl-specific 39E DNAzyme, and a 39S substrate strand labeled with a Cy3 fluorophore and a BHQ-2 quencher, respectively (Figure 4A)
Summary
Catalysis in biology has long been considered to be the function of catalytic RNAs (ribozymes) and protein enzymes [1]; many studies have reported the discovery of catalytic DNAs (called DNAzymes hereafter) since Breaker and Joyce’s pioneer work on DNAzymes in 1994 [2,3,4]. The specific catalytic functionalities of these DNAzymes depend on the sequences of the DNAzymes and in many cases on the presence of additional cofactors, such as metal ions and amino acids [6] These DNAzymes have been identified to catalyze many chemical reactions, including RNA cleavage, oxidative or hydrolytic DNA cleavage, DNA/RNA ligation, and DNA phosphorylation [4,5]. The substrate strand contains an rA linkage that serves as a cleavage site, and in the presence of the cofactor, the enzyme strand forms a defined second structure with the catalytic ability to cleave the rA linkage, resulting in two fragments with a 20 -30 cyclic phosphate and a 50 -OH terminus, respectively (Figure 1B) This catalytic property of DNAzyme is highly specific to the substrate strand, and even a single base mismatch in the antisense arms could significantly decrease the cleavage efficiency.
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