DNA as a Versatile Chemical Component for Catalysis, Encoding, and Stereocontrol

Silverman, Scott

doi:10.1002/anie.200906345

Cited by 231 publications

(76 citation statements)

References 266 publications

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“…Reactivity in DNA can be modulated using a templating approach that holds the catalytic nucleophile of non-native ribose sugar in an appropriate geometry can be positioned to accelerate hydrolysis [19]. This structural modification can be called templating since it brings two strands into proximity using the molecular recognition of the DNA binding arms to permit catalysis as shown in Figure 4.…”

Section: Dnazyme Templates For Enhanced Nucleophilicitymentioning

confidence: 99%

“…DNA catalysts have been also designed using intercalators that contain metal centers [17,18]. Templated structures that use the advantage of recognition in chemistry have been employed to advantage in the development of novel hydrolysis catalysts using both divalent ions and embedded amino acid functionality [19][20][21]. Addition of chemical functionality that causes large changes in DNA or RNA properties requires consideration of the structural and chemical aspects of nucleic acids in applications ranging from binding interactions to enzymatic catalysis.…”

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Thermodynamics of Nucleic Acid Structural Modifications for Biotechnology Applications

Franzen¹

2011

Biotechnology of Biopolymers

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IntroductionThe repertoire of chemistry available to native nucleotides in DNA and RNA is limited to the purine and pyrimidine functional groups, along with the special role of the 2'-hydroxyl of RNA. The nucleobases have exocyclic amino groups and imines, neither of which is highly reactive or a good candidate for catalytic function. One might argue that the limited range of chemical reactivity is an evolutionary advantage in molecules whose functions are primarily to store (DNA) and process (RNA) genetic information. The most important attribute of both DNA and RNA is the molecular recognition through base pairing. The hydrogen bond pattern combined with the conformation of the ribose sugar gives rise to the specific B-form double helix in DNA and A-form helix with a range of additional standard folds in RNA, loops, bulges and pseudoknots. These structures are well known for their ability to interact with proteins to provide a scaffold for transcription (DNA → RNA) to create messenger RNA (mRNA), and processing of mRNA by enzymes that have active components composed of RNA molecules. The chemical action of RNA on other RNAs by means of the 2'-hydroxyl is an exception to the lack or reactivity of DNA and RNA. Selfsplicing by action of the 2'-hydroxyl as a nucleophile was the process that broke the dogma that RNA is always a passive molecule that simply transmits information [1]. Indeed, RNA i s v e r y a c t i v e i n p r o c e s s i n g o t h e r R N A s u s i n g t h e 2 ' -h y d r o x y l a s a n u c l e o p h i l e f o r hydrolysis of the phosphodiester bond leading to cleavage or to rearrangements of structure such as RNA splicing. Outside of this reactivity, the components of the purine and pyrimidine rings are largely inert. Aromatic amines are poor nucleophiles, and imines are even less reactive. The purine and pyrimidine rings are not particularly electrophilic because of the nitrogen heteroatoms and carbonyls. Using in vitro selection (also known as SELEX) [2][3][4], and appropriate modification, RNA and DNA have been developed for numerous applications that transcend their biological functions. In this chapter we will consider the modifications of the nucleobase as a means to expand upon the native function. The extensive literature on modifications of the phosphodiester backbone and the ribose sugar will not be considered in this chapter due to space limitations. Base modifications may consist of expansions of the purine/pyrimidine ring, appended functional group or chemical modification to increase the stability of the backbone with respect to hydrolysis. Expanded DNA or xDNA has been developed as mimics of native DNA with potential biotechnology applications [5,6]. In these molecules, the purine and pyrimidine rings are fused to phenyl or naphthyl rings to give rise to

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Section: Dnazyme Templates For Enhanced Nucleophilicitymentioning

confidence: 99%

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confidence: 99%

Thermodynamics of Nucleic Acid Structural Modifications for Biotechnology Applications

Franzen¹

2011

Biotechnology of Biopolymers

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“…Nature's biomolecular protein and RNA catalysts are responsible for a wide range of chemical reactions, and protein enzymes in particular can achieve large rate enhancements (1,2). Although DNA catalysts are unknown in nature, in vitro selection [first pioneered for RNA (3)] is readily applied to identify catalytically active artificial DNA sequences (4)(5)(6). Importantly, DNA (and RNA) catalysts can be identified by starting with entirely random sequence pools, whereas directed evolution of proteins typically requires a known, catalytically active starting point (7,8).…”

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confidence: 99%

“…Importantly, DNA (and RNA) catalysts can be identified by starting with entirely random sequence pools, whereas directed evolution of proteins typically requires a known, catalytically active starting point (7,8). A growing range of chemical reactions has been shown to be catalyzed by DNA (4)(5)(6). For DNA phosphodiester hydrolysis, the uncatalyzed (spontaneous) half-life for P-O bond cleavage of ∼30 million y is reduced to as little as 0.5 min by a DNA catalyst (9,10).…”

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Catalytic DNA with phosphatase activity

Chandrasekar

Silverman

2013

Proc. Natl. Acad. Sci. U.S.A.

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Catalytic DNA sequences (deoxyribozymes, DNA enzymes, or DNAzymes) have been identified by in vitro selection for various catalytic activities. Expanding the limits of DNA catalysis is an important fundamental objective and may facilitate practical utility of catalysts that can be obtained from entirely unbiased (random) sequence populations. In this study, we show that DNA can catalyze Zn 2+ -dependent phosphomonoester hydrolysis of tyrosine and serine side chains (i.e., exhibit phosphatase activity). The best deoxyribozyme decreases the half-life for phosphoserine hydrolysis from as high as >10 10 y to <1 h. The phosphatase activity also occurs with nonpeptidic substrates but with reduced efficiency, indicating a preference for phosphopeptides. The newly identified deoxyribozymes can function with multiple turnover using free peptide substrates, have activity in the presence of human cell lysate or BSA, and catalyze dephosphorylation of a larger protein substrate, suggesting broader application of DNA catalysts as artificial phosphatases. D evelopment of catalysts is a major impetus for much of modern chemical research. Nature's biomolecular protein and RNA catalysts are responsible for a wide range of chemical reactions, and protein enzymes in particular can achieve large rate enhancements (1, 2). Although DNA catalysts are unknown in nature, in vitro selection [first pioneered for RNA (3)] is readily applied to identify catalytically active artificial DNA sequences (4-6). Importantly, DNA (and RNA) catalysts can be identified by starting with entirely random sequence pools, whereas directed evolution of proteins typically requires a known, catalytically active starting point (7,8). A growing range of chemical reactions has been shown to be catalyzed by DNA (4-6). For DNA phosphodiester hydrolysis, the uncatalyzed (spontaneous) half-life for P-O bond cleavage of ∼30 million y is reduced to as little as 0.5 min by a DNA catalyst (9, 10). However, in this reaction the DNA catalyst interacts with its DNA substrate by extensive Watson-Crick base pairing, and such an approach cannot be generalized to nonoligonucleotide substrates such as peptides and proteins. With the exception of the ribosome, the natural ribozymes catalyze RNA cleavage and ligation, and they generally have more modest rate enhancements (11-13), limited by the relatively high uncatalyzed half-lives of these reactions.DNA catalysts that covalently modify peptide and protein substrates are fundamentally interesting and likely have practical value, especially for biologically relevant chemical modifications.We have initiated studies into DNA-catalyzed modifications of amino acid side chains of peptide substrates, such as nucleopeptide linkage formation involving tyrosine and serine (14-16). A major challenge in such studies is to achieve catalysis even though the DNA catalyst cannot engage in any preprogrammable WatsonCrick binding interactions with the substrate. In this report, we show that DNA can catalyze Zn 2+ -dependent hydrolysis of tyrosine...

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“…[6][7][8] These molecules can be isolated from a vast random-sequence DNA pool (which contains as many as 10 16 individual sequences) by a process known as "in vitro selection" or "SELEX" (systematic evolution of ligands by exponential enrichment). [9][10][11][12][13][14][15][16] These special DNA molecules have been widely examined in recent years as molecular tools for biosensing applications. [6][7][8] Our laboratory has established in vitro selection procedures for isolating RNA-cleaving fluorescent DNAzymes (RFDs; Fig.…”

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Detection of Bacteria Using Fluorogenic DNAzymes

Aguirre

Ali

Kanda

et al. 2012

JoVE

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Outbreaks linked to food-borne and hospital-acquired pathogens account for millions of deaths and hospitalizations as well as colossal economic losses each and every year. Prevention of such outbreaks and minimization of the impact of an ongoing epidemic place an ever-increasing demand for analytical methods that can accurately identify culprit pathogens at the earliest stage. Although there is a large array of effective methods for pathogen detection, none of them can satisfy all the following five premier requirements embodied for an ideal detection method: high specificity (detecting only the bacterium of interest), high sensitivity (capable of detecting as low as a single live bacterial cell), short time-toresults (minutes to hours), great operational simplicity (no need for lengthy sampling procedures and the use of specialized equipment), and cost effectiveness. For example, classical microbiological methods are highly specific but require a long time (days to weeks) to acquire a definitive result.1 PCR-and antibody-based techniques offer shorter waiting times (hours to days), but they require the use of expensive reagents and/or sophisticated equipment. 2-4 Consequently, there is still a great demand for scientific research towards developing innovative bacterial detection methods that offer improved characteristics in one or more of the aforementioned requirements. Our laboratory is interested in examining the potential of DNAzymes as a novel class of molecular probes for biosensing applications including bacterial detection. 5DNAzymes (also known as deoxyribozymes or DNA enzymes) are man-made single-stranded DNA molecules with the capability of catalyzing chemical reactions. [6][7][8] These molecules can be isolated from a vast random-sequence DNA pool (which contains as many as 10 16 individual sequences) by a process known as "in vitro selection" or "SELEX" (systematic evolution of ligands by exponential enrichment). 9-16 These special DNA molecules have been widely examined in recent years as molecular tools for biosensing applications. 6-8Our laboratory has established in vitro selection procedures for isolating RNA-cleaving fluorescent DNAzymes (RFDs; Fig. 1) and investigated the use of RFDs as analytical tools. [17][18][19][20][21][22][23][24][25][26][27][28][29] RFDs catalyze the cleavage of a DNA-RNA chimeric substrate at a single ribonucleotide junction (R) that is flanked by a fluorophore (F) and a quencher (Q). The close proximity of F and Q renders the uncleaved substrate minimal fluorescence. However, the cleavage event leads to the separation of F and Q, which is accompanied by significant increase of fluorescence intensity.More recently, we developed a method of isolating RFDs for bacterial detection. 5 These special RFDs were isolated to "light up" in the presence of the crude extracellular mixture (CEM) left behind by a specific type of bacteria in their environment or in the media they are cultured (Fig. 1). The use of crude mixture circumvents the tedious process of purifyi...

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DNA as a Versatile Chemical Component for Catalysis, Encoding, and Stereocontrol

Cited by 231 publications

References 266 publications

Thermodynamics of Nucleic Acid Structural Modifications for Biotechnology Applications

Thermodynamics of Nucleic Acid Structural Modifications for Biotechnology Applications

Catalytic DNA with phosphatase activity

Detection of Bacteria Using Fluorogenic DNAzymes

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