Characterization of deoxyribozymes with site-specific oxidative cleavage activity against DNA obtained by in vitro selection

Wang, Ming-Qi; Dong, Juan; Zhang, Huafan; Tang, Zhuo

doi:10.1039/c6ob00148c

Cited by 7 publications

(10 citation statements)

References 24 publications

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“…This cleavage process occurs by a superoxide/H 2 O 2 -dependent radical mechanism rather than an ionic mechanism and leads to well-defined reaction products that are identical to those formed by the action of the glycopeptide natural product bleomycin on double-stranded DNA (dsDNA) . Unlike the reported examples of DNA-catalyzed oxidative DNA cleavage, , the reactions described here do not require redox-active metal ion cofactors such as Cu 2+ or Mn 2+ . Instead, only redox-inactive Zn 2+ and Mg 2+ are needed.…”

Section: Introductionmentioning

confidence: 85%

“…In contrast, the Cu 2+ /Mn 2+ -dependent oxidatively cleaving deoxyribozymes do not lead to well-defined products, instead forming a variety of fragments via “oxidative destruction” of one or more nucleosides. 11,12 …”

Section: Discussionmentioning

confidence: 99%

“…11,12 The current mechanistic model for the RadDz3/RadDz6 reaction pathway has unresolved aspects. The homogeneity of the reaction products indicates that the C4′ radical is formed at a specific guanosine nucleoside of the substrate, presumably as directed by the deoxyribozyme, but the details of C4′ radical generation are undefined by the current data.…”

Section: Discussionmentioning

confidence: 99%

“…9 We and others also found deoxy-ribozymes that deglycosylate DNA, with subsequent elimination reactions that lead to strand scission. 10 Other reported DNA-catalyzed cleavage reactions include oxidative DNA cleavage 11,12 and phosphoramidate cleavage. 13 …”

Section: Introductionmentioning

confidence: 99%

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DNA-Catalyzed DNA Cleavage by a Radical Pathway with Well-Defined Products

et al. 2016

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We describe an unprecedented DNA-catalyzed DNA cleavage process in which a radical-based reaction pathway cleanly results in excision of most atoms of a specific guanosine nucleoside. Two new deoxyribozymes (DNA enzymes) were identified by in vitro selection from N40 or N100 random pools initially seeking amide bond hydrolysis, although they both cleave simple single-stranded DNA oligonucleotides. Each deoxyribozyme generates both superoxide (O2-• or HOO•) and hydrogen peroxide (H2O2) and leads to the same set of products (3′-phosphoglycolate, 5′-phosphate, and base propenal) as formed by the natural product bleomycin, with product assignments by mass spectrometry and colorimetric assay. We infer the same mechanistic pathway, involving formation of the C4′ radical of the guanosine nucleoside that is subsequently excised. Consistent with a radical pathway, glutathione fully suppresses catalysis. Conversely, adding either superoxide or H2O2 from the outset strongly enhances catalysis. The mechanism of generation and involvement of superoxide and H2O2 by the deoxyribozymes is not yet defined. The deoxyribozymes do not require redox-active metal ions and function with a combination of Zn2+ and Mg2+, although including Mn2+ increases the activity, and Mn2+ alone also supports catalysis. In contrast to all of these observations, unrelated DNA-catalyzed radical DNA cleavage reactions require redox-active metals and lead to mixtures of products. This study reports an intriguing example of a well-defined, DNA-catalyzed, radical reaction process that cleaves single-stranded DNA and requires only redox-inactive metal ions.

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Section: Introductionmentioning

confidence: 85%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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DNA-Catalyzed DNA Cleavage by a Radical Pathway with Well-Defined Products

et al. 2016

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“…These reactions include Cu 2+ -dependent oxidative cleavage [46–48], Cu 2+ /Mn 2+ -dependent oxidative nucleoside excision [49,50], DNA phosphodiester hydrolysis [51,52], RNA hydrolysis (involving a water molecule, a normally disfavored reaction relative to RNA cleavage by the Figure 1 mechanism) [53], and deglycosylation with subsequent backbone cleavage [54–56]. Oligonucleotide end modification has also been achieved by DNA 5′-phosphorylation (kinase activity) [57] and adenylylation (capping) [58] followed by ligation [59].…”

Section: Reaction Scope Of Catalysis By Dnamentioning

confidence: 99%

Catalytic DNA: Scope, Applications, and Biochemistry of Deoxyribozymes

Silverman

2016

Trends in Biochemical Sciences

309

215

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The discovery of natural RNA enzymes (ribozymes) prompted the pursuit of artificial DNA enzymes (deoxyribozymes) by in vitro selection methods. A key motivation is the conceptual and practical advantages of DNA relative to proteins and RNA. Early studies focused on RNA-cleaving deoxyribozymes, and more recent experiments have expanded the breadth of catalytic DNA to many other reactions. Including modified nucleotides has the potential to widen the scope of DNA enzymes even further. Practical applications of deoxyribozymes include their use as sensors for metal ions and small molecules. Structural studies of deoxyribozymes are just beginning; mechanistic experiments will surely follow. From the first report 21 years ago, the field of deoxyribozymes has promise for both fundamental and applied advances in chemistry, biology, and other disciplines.

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