Nucleoside Triphosphates — Building Blocks for the Modification of Nucleic Acids

Abstract

Nucleoside triphosphates are moldable entities that can easily be functionalized at various locations. The enzymatic polymerization of these modified triphosphate analogues represents a versatile platform for the facile and mild generation of (highly) functionalized nucleic acids. Numerous modified triphosphates have been utilized in a broad palette of applications spanning from DNA-tagging and -labeling to the generation of catalytic nucleic acids. This review will focus on the recent progress made in the synthesis of modified nucleoside triphosphates as well as on the understanding of the mechanisms underlying their polymerase acceptance. In addition, the usefulness of chemically altered dNTPs in SELEX and related methods of in vitro selection will be highlighted, with a particular emphasis on the generation of modified DNA enzymes (DNAzymes) and DNA-based aptamers.

Nucleoside Triphosphates - From Synthesis to Biochemical Characterization

“…However, the phosphoramidite building blocks need to be resilient to the rather harsh conditions imposed by this methodology, which in turn imposes a severe restriction onto the nature of the chemical functionality 47 . Instead, the enzyme-mediated polymerization of modified nucleoside triphosphates allows for the introduction of a broader range of functionalities, since the sole restriction is that they act as substrates for polymerases 1,2 . Even though there is a noticeable lack of generally applicable methodology, reliable and robust synthetic and analytical methodologies have been developed for the synthesis of modified dNTPs.…”

“…The additional side-chains that are introduced by the polymerization of the modified dNTPs are thought to increase the chemical space that can be explored during a selection experiment and supplement the rather poor functional arsenal of nucleic acids 37 . However, despite these attractive traits and the recent progress made in the development of both synthetic and analytical methods, no universally applicable and high-yielding procedure exists for the crafting of modified nucleoside triphosphates 2,38 .…”

“…In addition to their role in these fundamental biological transformations, their modified counterparts have advanced as a versatile and mild platform for the introduction of functional groups into oligonucleotides, a methodology that nicely complements the automated solid-phase synthesis that is usually applied 1,2 . Indeed, provided the (d)NTPs can act as substrates for RNA and DNA polymerases 3 , a wealth of functional groups including amino acids [4][5][6][7][8][9][10][11][12][13] , boronic acids 14,15 , nornbornene 16 , diamondoid-like residues 17 , side-chains for organocatalysis 18 , bile acids 19 , and even oligonucleotides 20 can be introduced into oligonucleotides.…”

DNA Three Way Junction Core Decorated with Amino Acids-Like Residues-Synthesis and Characterization

“…With the aim to use in vitro selection methods, the Hollenstein s group developed triphosphate deoxynucleosides bearing the amino acids functionalities grafted on the purine or pyrimidine base at a position that would interfere neither with polymerase nor with duplex formation ability [6]. It turned out that these modified triphosphates were good substrates for various DNA polymerases and were fully compatible for the selection process [7].…”

Preparation of fluorescent nucleic acids generating unique emission by primer extension reaction using pyrene-labeled deoxyuridine triphosphate derivatives

“…[1][2][3] To expand their applications as versatile tools, various kinds of oligodeoxynucleotide modified with functional molecules have been prepared by a general synthetic approach using the corresponding phosphoroamidite units during solid-phase DNA synthesis and the combination with post-synthetic oligonucleotides modification. 4,5 As an alternative approach to the conventional DNA synthesis for the preparation of the functionalized oligonucleotides, an enzymatic method using the polymerase extension (PEX) reaction has attracted much attention because of the facile preparation of longer oligonucleotides from DNA templates and multiple incorporation of functional groups/molecules into DNA based on the sequence of the templates. 6,7 Certain kinds of deoxynucleotide triphosphate (dNTP) derivatives possessing a fluorescent reporter group at an appropriate position on the nucleobase can work as effective substrates for the enzymatic incorporation by polymerase and are used as fluorescent probes for the detection of the hybridization and amplification of DNA.…”